Tephrosia purpurea (Linn.) attenuates progression of acute liver failure by reducing oxidative stress and inflammation via modulating cytoprotective enzyme hemoxygenase-1 activity

Tephrosia purpurea (Linn.) attenuates progression of acute liver failure by reducing oxidative stress and inflammation via modulating cytoprotective enzyme hemoxygenase-1 activity | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Tephrosia purpurea (Linn.) attenuates progression of acute liver failure by reducing oxidative stress and inflammation via modulating cytoprotective enzyme hemoxygenase-1 activity Samrat Rakshit, Nisha Sahu, Satendra Kumar Nirala, Shamli S Gupte, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1596610/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background: Acute liver failure (ALF), often associated with elevated oxidative stress and sustained inflammation, is one of the major clinical manifestations of liver disease with limited therapeutic options. Tephrosia purpurea is mentioned in Ayurvedic literature to be effective against various health problems. Additionally, this plant with various unknown bioactive components has not been fully explored against acute organ injury, especially against D-galactosamine and lipopolysaccharide induced acute hepatitis. Thus, we aimed to characterize and investigate the efficacy of hydroethanolic extract of Tephrosia purpurea to attenuate ALF progression. Methods: Different doses of Tephrosia purpurea extract (50, 100 and 200 mg/kg) were administered orally once a day for 6 days. In vivo injury was induced via intraperitoneal injection of D-galactosamine (300 mg/kg) followed by lipopolysaccharide (50 μg/kg). Presence of bioactive compounds, functional groups, nature and size of particles, thermal properties of the extract were determined. Results: Levels of lipid peroxidation, proinflammatory cytokines were significantly elevated after D-galactosamine and lipopolysaccharide administration that were restored by the administration of Tephrosia purpurea via modulating hemoxygenase-1 enzyme. Tephrosia purpurea alleviated serum transaminase levels, hematological variables and markedly reduced histological and ultrastructural alterations. Tephrosia purpurea reduced iron overload and maintained glutathione homeostasis (P£ 0.05). Not only that, it also eliminated lipid droplets, restored biomacromolecule homeostasis, maintained stability of the membrane, cellular organelles and the DNA. Conclusions: These findings provided novel insights into the anti-inflammatory and antioxidative mechanisms of Tephrosia purpurea and suggested its use as a potential therapeutic candidate against ALF. Acute hepatitis D-Galactosamine Hemoxygenase-1 HPLC Inflammation Lipopolysaccharide Natural antioxidants Oxidative stress Reactive oxygen species Tephrosia purpurea Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Introduction Acute liver failure (ALF) or acute liver injury (ALI) is one of the severe forms of liver disease. Pathological characteristics include extensive hepatic necrosis with severe loss of parenchyma, violent oxidative stress, and sustained inflammation. Despite enormous clinical efforts, prognosis and high mortality rate haven’t improved significantly [ 1 , 2 ]. Administration of specific hepatotoxin D-galactosamine (D-GalN) in combination with endotoxin lipopolysaccharide (LPS) greatly induces acute liver injury and mimics liver disease in human. This model of acute liver injury is extensively used worldwide to study underlying mechanism of ALF and develop effective therapeutic drugs [ 1 , 2 ]. It is indispensable that D-GalN and LPS induce oxidative stress and inflammatory responses, which promotes hepatic necrosis, contributing directly in the occurrence and progression of ALF. Therefore, taking care of increased oxidative stress and inflammation may be the first step for the treatment of ALF [ 3 ]. Cytoprotective enzymes such as heme oxygenase (HO)-1 play major role in the well-being of a cell under immense oxidative stress and inflammation. Thus, modulation of these cytoprotective enzymes related to oxidative stress and inflammation could be an effective therapeutic strategy for D-GalN and LPS induced ALF [ 4 ]. Traditional systems of medicine remain invaluable since ages due to their multiple health outcomes and minimal side effects as compared to modern drugs [ 5 ]. These plants have long been thought to be potential hepatoprotective agents for their various biological activities including antioxidant and anti-inflammatory by regulating multiple signal molecules [ 5 ]. The use of crude plant extracts as therapeutic supplement showed various plausible health benefits including amelioration of metabolic complications, oxidative stress and inflammation induced injury [ 6 ]. Plants possess many advantageous biologically active phytochemicals with minimal to no side effects, when taken as a part of human diet or at effective doses [ 7 ]. Phytochemicals present in plant extract often exert synergistic effect in improving the efficacy of plant extracts than administered alone. Thus, various plants are often screened for beneficial effect in ameliorating health problems [ 8 ]. Tephrosia purpurea (Linn.) pers. (Fabaceae) is a self-generating tropical perennial herb, known as “Sarwawranvishapaha” in Ayurveda for its usefulness in hepatic and splenic disorders, phantom tumor, poisonous bite and almost all type of wounds [ 9 ]. The usefulness of Tephrosia purpurea is also mentioned in an old Ayurvedic literature “Bhavaprakasha Nighantu Guduchyadi Varga, 169” that it can alleviate cough, cure blood related diseases, dyspnea and fever [ 9 , 10 ]. In modern day, hepatoprotective formulations such as “Tephroli” and “Yakrifit” have Tephrosia purpurea as one of their major constituents [ 11 ]. Tephrosia purpurea is reported to possess multiple phenolic compounds including glycosides, flavonoids and flavones, responsible for its various protective properties [ 9 , 12 , 13 ]. Tephrosia purpurea showed a very high therapeutic index as the LD 50 of its ethanolic extract is 2000 mg/kg in rats [ 14 ]. Traditional healers claim that Tephrosia purpurea is useful in treating diabetes, rheumatism, asthma, diarrhea, ulcers [ 15 ], gonorrhea, bronchitis, impotency and other kidney and heart disorders [ 16 ]. These claims about Tephrosia purpurea , in our traditional healing system need scientific validation to prove its effectiveness. To the best of our knowledge, no scientific literature is available that explains biophysical properties of Tephrosia purpurea and its effectiveness, particularly against D-GalN and LPS induced acute liver injury. Thus, present study was focused to investigate biophysical characteristics of Tephrosia purpurea and its therapeutic potential against D-GalN and LPS induced acute liver injury. Materials And Methods Preparation of hydroethanolic extract of Tephrosia purpurea The plant was identified as Tephrosia purpurea (Linn.) pers. and a Voucher no: GGV/BOT/H/FAB/DKS/210 was obtained at Botany Department, Guru Ghasidas University, Chhattisgarh, India. Whole plant of Tephrosia purpurea was collected form Guru Ghasidas University campus and air dried in shade for 3 weeks and pulverized. 15 grams of pulverized sample was extracted by Soxhlet apparatus (worm method) for 30 minutes using 70% (v/v) ethanol in water. The hydroethanolic extract was concentrated in a rotary evaporator and lyophilized. The yield of the extract (2.9g, 19.33%) was obtained preserved for further use. Reverse phase-HPLC instrumentation and chromatographic conditions The isocratic reverse phase HPLC analysis was carried out in a Shimadzu HPLC system (LC-20AD) consisted with double pumps (LC 20-AD), column oven (CTO-20AC), sample injector system (SIL-20ACHT) and a spectrophotometric detector (SPD-20A) with a computer assisted analysis software (Shimadzu LC solution 1.25sp1). The stationary phase used for this analysis was a reverse-phase C18 column (150mm × 4.6 mm, 5 𝜇m) and the mobile phase consisted of aqueous acid solution (0.5 M phosphoric acid) : methanol (50 : 50 v/v) mixture with a stable flow rate of 0.9 mL/min. The mobile phase was filtered through 0.45 μm membrane filter and degassed by ultrasonic bath prior to use. The temperature of the column was maintained at room temperature (25 ± 2ºC). 20 𝜇L of each sample were filtered through a 0.22 μm nylon membrane before injecting into the HPLC system for analysis. The presence of bioactive phytochemicals including tannic acid, ascorbic acid, catechin, gallic acid, caffeic acid, rutin, vanillic acid, morin, malic acid, quercetin and ellagic acid were investigated. Presence of these compounds were confirmed by comparing chromatogram peaks of plant extract with reference standards by comparing their retention time. Stock solutions of 40 ppm for all the standard references and 2000 ppm for plant extract were prepared in mobile phase. The detection wavelength was 280 nm for tannic acid, 265 nm for ascorbic acid, 276 nm for catechin, 220 nm for gallic acid, 324 nm for caffeic acid, 260 nm rutin, 220 nm for vanillic acid, 370 nm for morin, 230 nm for malic acid, 256 nm for quercetin and 368 nm for ellagic acid. The detection wavelength for Tephrosia purpurea extract was 269 nm. The absorption spectra were determined by UV-VIS spectroscopy. UV–Vis spectra analysis Hydroethanolic (70% ethanol in water) extract of Tephrosia purpurea at 1000 ppm concentration was observed by UV–Vis spectroscopy within the ranges between 200 nm and 800 nm to identify phytoconstituents containing σ-bonds, π-bonds and lone pair of electrons, chromophores and aromatic rings present in it. Particle size analysis (PSA) The crude plant extract, having various particle size ranges were determined using particle size analyzer (Shimadzu’s SALD- 2300). Sample was dissolved in 20 ml 70% ethanol followed by sonication for 10 minutes. Particle sizes were measured based on the time dependent scattering of laser light by the particles with a pump speed of 5.0 cm 3 /min. Thermogravimetry analysis (TGA) The changes in mass of Tephrosia purpurea extract over the time with changes in temperature were assessed. Dynamic thermogravimetric curves of extract were obtained by thermogravimetric analysis with Shimadzu’s TGA -50, with 10°C/min heating rate ranging from 25°C to a temperature of 800°C under nitrogen atmosphere with a flow of 100 mL/min. The plant extract was packed in a platinum crucible and subjected to analysis. Calcium oxalate monohydrate was used for calibration of the instrument to characterize weight loss steps. Differential scanning colorimetry (DSC) The DSC curves of Tephrosia purpurea were obtained using a Shimadzu calorimeter, model Shimadzu’s DSC-60 Plus. The instrument was coupled to a photovisual system and an oven with a temperature range of 20-300°C. The whole analysis was done under nitrogen atmosphere with constant flow of 100 ml/min and a heating rate of 5°C/min. The plant extract was packed in alumina crucible for analysis. X-ray diffraction (XRD) The structural analysis of Tephrosia purpurea extract was carried out by using X ray diffraction techniques on Rigaku’s Mini Flex 600 diffractometer. The Tephrosia purpurea extract was subjected to Cu-Kα radiations (λ=0.154056 nm, 2θ= 2-90°) operating at a voltage of 40 kV and a current of 15 mA, at 10°C/min speed at a drive axis of 2θ. The average size of the crystallite (D in nm) was also calculated from the XRD peak using Debye Scherrer’s relation [17, 18]. Where, k = 0.9 is the Scherrer’s constant , λ= 1.540562Å is wavelength of X-rays and β (in radian) is the full width of half maxima (FWHM) of the diffraction peak at an angle θ. Fourier transform infrared spectroscopy (FT-IR) Functional groups are responsible for medicinal properties of plants. The whole plant extract of Tephrosia purpurea was subject to FT-IR analysis on a Perkin Elmer’s Spectrum Two using KBr stressed disks within a spectral range of 4000–450 cm −1 to determine presence of possible functional groups. Experimental animals Experimental animals ( Wistar rats, 150±10g) were purchased from Defense Research and Development Establishment, Gwalior, and kept in departmental animal house for 15 days to get acclimatized. Animals were exposed to a 12 h periodic light and dark cycle with a constant temperature about 25°±2°C. Rats were provided with palleted rat diet and water ad libitum . Animal right Animal care and experimental procedures were carried out by sincerely following the guidelines set by the committee for the purpose of control and supervision of experiments on animal, India. The experimental procedure was approved by institutional animal ethics committee (994/Ere/Go/06/CPCSEA) on 01/08/2016 (Ref:162/IAEC/Pharmacy/2016). Chemicals Pure and analytical grade chemicals were procured from Sisco Research Laboratories Pvt. Ltd (SRL) and Himedia Laboratories Pvt. Ltd., India in this study. LPS (Lot # 114M4009V) was procured from Sigma Aldrich Co Ltd, USA whereas D-GalN (Batch # 3647503) was procured from SRL, India. D-GalN and LPS model of acute multiorgan injury The D-GalN and LPS was prepared in saline (0.9% NaCl in water). Acute organ injury was induced by D-GalN (300 mg/2ml/kg) and LPS (50µg/2ml/kg) administered intraperitonially as per previous scientific literature [19]. Preparation of different doses of Tephrosia purpurea extract Freeze dried plant extract was dissolved in distilled water using 1% (w/v) gum acacia as the suspending agent [19]. Three different doses of Tephrosia purpurea (50 mg/kg, 100 mg/kg and 200 mg/kg) were taken for evaluation where the highest dose (200 mg/kg) corresponds to 1/10 th of the LD 50 . Experimental design Well acclimatized thirty-six animals were divided equally into six groups. Group I: designated as control, received 1% gum acacia as vehicle. Group II: designated as TP per se, received 200 mg/kg dose of Tephrosia purpurea extract. Group III: designated as D-GalN+LPS (experimental control), received 300 mg/kg dose of D-GalN followed by 50 µg/kg dose of LPS. Group IV-VI: designated as TP50, TP100 and TP200 respectively, received 50, 100 and 200 mg/kg dose of Tephrosia purpurea extract respectively along with D-GalN and LPS. Vehicle and different doses of Tephrosia purpurea were given orally for 6 days straight to their respective groups. The D-GalN followed by LPS (only after 1 hour of D-GalN) were given intraperitonially to group III to VI on the 6 th day evening. All the animals were fasted overnight and sacrificed 16 hours after D-GalN and LPS treatment using diethyl ether. To generate an individual data point in each of the independent experiments, samples were used in triplicates. Hematological study Blood was collected by puncturing retro-orbital venous sinus into heparinized tubes and kept at 4°C for hematological studies. Analysis of blood for determining white blood cell count (WBC), red blood cell count (RBC), platelet count (PLT), hemoglobulin (Hb), hematocrit (HCT), lymphocyte and monocyte count was done by semi-automatic blood analyzer (Analytica HEMA 2062+) [20]. Serological study The blood was also collected in heparin free tubes and allowed to clot at room temperature for 30 minutes. The blood was centrifuged to obtain serum for determination of serological parameters such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin, urea, uric acid, creatinine, glucose, triglyceride and cholesterol with the help of respective diagnostic kits following direction of use given on kit manual (The ERBA Chem 5 v 3 Germany). Oxidative stress assessment in hepatic and renal tissue samples For determination of lipid peroxidation (LPO), 10% homogenates of hepatic and renal tissues were prepared in KCl solution. The LPO in liver, kidney and microsomes were determined from quantified TBARS [21]. Antioxidant status assessment in hepatic and renal tissue samples Sucrose solution (1%) was used to prepare homogenates to assess reduced glutathione (GSH) [22]. NaCl (0.9%) was used to prepare tissue homogenates to determine superoxide dismutase [23] and catalase activities [24]. Lipid status in hepato-renal tissue samples To determine triglycerides [25] and cholesterol [26] in tissue, homogenates of liver and kidney were prepared in chilled hypotonic solution. Microsomal CYP2E1 activity , protein and lipid peroxidation Microsomes were prepared by CaCl 2 precipitation method [27]. The CYP2E1 activity was determined in terms of aniline hydroxylase activity using aniline as a substrate [28]. Microsomal protein was assessed using Folin and Ciocalteu phenol reagent [29]. Histopathological preparation of hepatic and renal tissue samples Small pieces of liver and kidney were immediately fixed in Bouin’s fixative. Fixed tissues were further processed and embedded in paraffin wax before cutting 05 mm thick sections. Sections were placed on a glass slide and stained with hematoxylin and eosin (H&E) stain [30]. Approximately 10-15 observations for each slide were taken with light microscope. From the above-mentioned results, it can be clearly observed that, 200 mg/kg dose of Tephrosia purpurea perform better than the other two lower doses. Thus, only 200 mg/kg dose of Tephrosia purpurea treated groups was further considered for determination of antioxidant enzymes of GSH cycle, proinflammatory cytokines, electron microscopy, genotoxicity and vital parameters of RBC degradation cycle. Enzymes of GSH cycle in hepato-renal tissue samples Glutathione reductase [31], glutathione peroxidase [32], glucose-6-phosphate dehydrogenase [33] and glutathione-S-transferase [34] were determined from tissue homogenate freshly prepared in 1.15% KCl solution. Assessment of proinflammatory cytokines in serum Proinflammatory cytokines including tumor necrosis factor- α (TNF-α) and interleukin-6 (IL-6) were assessed in serum using ELISA kits (Ray Biotech, Inc; Norcross, GA 30092 USA) to assess inflammation. RBC degradation cycle in hepatic tissue sample Hepatic hemoglobin [35], heme content [36], ferritin [37], hemosiderin [38], iron [39], bilirubin [40], biliverdin [41], biliverdin reductase [42] and hemoxygenase-1 (HO-1) enzyme [43], were determined in freshly prepared liver homogenate. Processing of hepatic and renal tissue samples for ultra-structural observation Liver and kidney tissue of about 1 mm 3 in size were fixed immediately in Karnovsky’s fixative and then in 1% osmium tetroxide [44]. Fixed tissues were eventually embedded in resin and were cut into 70-90 nm thick sections and placed on a copper grid. The grid along with tissue sections were stained with positive staining (Uranyl acetate and lead citrate) prior to the observation with transmission electron microscope [45, 46]. Genotoxicity study in hepato-renal tissue samples Single cell gel electrophoresis study was performed to observe genotoxicity [47]. Liver tissue homogenate (10%) was taken on a thin film of agarose spread glass slide. Slides were placed in electrophoresis buffer to start electrophoresis. Slides were immediately washed with HCl buffer and stained with ethidium bromide to observe with a microscope. Images were analyzed by ImageJ software. Statistical analysis Results are expressed as mean ± SE (n=6). Data were analyzed for statistical significance using one-way analysis of variance (P≤0.05) followed by Tukey’s post hoc honestly significant difference test ( post hoc HSD test) to draw a comparison among different treatment groups (P≤0.05) [48] using Microsoft Excel worksheet-2019. A p value ≤0.05 was considered as significant. Results HPLC fingerprinting analysis of Tephrosia purpurea extract The HPLC fingerprinting analysis of Tephrosia purpurea extract and other standards are summarized in Fig 1. Tephrosia purpurea extract showed various phytochemicals as evident by presence of peaks at various retention time. Based on the retention time, compounds were identified and presented in table 1. Results revealed the presence of some medicinally active well-known phytoconstituents as well as some unknown compounds. The first peak between retention time 2 to 4 comprises multiple active components including tannic acid (2.806), L-ascorbic acid (2.964), catechin (2.920), gallic acid (3.269) and caffeic acid (3.639). The peak at 4.020 is rutin (4.035), peak at 4.542 is vanillic acid (4.517), peak at 5.442 is morin (5.447) or malic acid (5.489), peak at 6.260 is quercetin (6.379) and the peak at 7.349 is ellagic acid (7.265). There were some unidentified peaks at retention time 9.581, 12.026, 18.342 and 23.398. UV-Vis Spectral analysis of Tephrosia purpurea extract Scanning analysis of the extract showed two peaks at UV range as well as one in visible range. The peaks at 260, 280 and 660 nm showed absorption of 3.994, 3.721, 0.242 respectively (Fig. 2). The first peak appeared at 260 nm was intense than that of 280 nm. A very light peak appeared at 660 nm suggested presence of colored compound in the extract. Particle size analysis of Tephrosia purpurea extract Fig. 3 represents the histograms of the dispersed particles present in Tephrosia purpurea extract as obtained from particle size analyzer. The size of the particles ranges between 0.031 to 0.071 µm. The percentile of particle size of particular diameter was explained in table 2. The most common particle size was 44 nm which comprises of 24.24%. The average particle size of 43 nm was determined from the mean value of the histograms. TGA and DSC analysis of Tephrosia purpurea extract The thermogram from TGA analysis of Tephrosia purpurea extract (Fig. 4A) showed thermal decomposition processes, which occurs between 126.58°C and 266.80°C with a midpoint at 241.33°C. The total weight loss was 17.188 mg (62.001%). The DSC curves of Tephrosia purpurea extract (Fig. 4B) showed thermal reaction processes, which occur between 18.64 and 128.49°C. The event of endothermic reaction was observed at 21.46 (D = 15.56 J g -1 ) and 120.88°C (D = 32.24 J g -1 ). Several other endothermic and exothermic peaks were observed between 128.49 and 232.28°C. The major weight loss was observed between 200-500°C. X-ray powder diffraction analysis of Tephrosia purpurea extract The diffractogram obtained from XRPD analysis of Tephrosia purpurea hydroethanolic extract showed 10 sharp peaks of variable height at various 2θ (theta) values (Fig. 5). XRD pattern showed presence of crystals in the extract. The crystallite size was calculated using the XRD data, suggested the presence of nanoparticles in the size range between 25 nm and 47 nm. The average crystallite size of the plant extract was found to be 34.3 nm. Furrier transform infrared spectroscopy of Tephrosia purpurea extract The FT-IR spectra of Tephrosia purpurea extract (Fig. 6) showed absorbance bands centered at 523, 1044, 1178, 1285, 1403, 1587, 2849, 2917 and 3216 cm −1 . Different band Peak represented different functional group present in the extract (Table 3). Effect of Tephrosia purpurea extract on hematological variables Table 4 represents alterations in different hematological variables after combined administration of D-GalN and LPS. Total number of RBCs, platelets, amount of hemoglobin in blood and hematocrit value were significantly declined, whereas differential cell counting of WBCs, including lymphocytes and monocytes were significantly raised (P£0.05). Treatment of Tephrosia purpurea showed dose dependent protection in most of the hematological variables. The 50 mg/kg dose of Tephrosia purpurea significantly checked monocytes count towards control. The 100 mg/kg dose boosted hemoglobin, RBCs, hematocrit in one hand and on the other hand, controlled WBCs, including monocytes significantly. The 200 mg/kg dose of Tephrosia purpurea significantly protected all the studied variables from alteration. It was observed that the 100 and 200 mg/kg doses of Tephrosia purpurea exhibited considerably therapeutic effects in comparison to 50 mg/kg dose. Effect of Tephrosia purpurea extract on serological variables and hepatic glycogen Table 5 exemplifies protective effect of Tephrosia purpurea pretreatment against toxic demonstration of D-GalN and LPS on serological variables. Exposure to D-GalN and LPS resulted in substantial increase of serum transaminases (AST and ALT), ALP, urea, uric acid and creatinine (P£0.05). All the three doses of Tephrosia purpurea provided protection against toxic outcome of D-GalN and LPS on serological indices. The 100 and 200 mg /kg dose of Tephrosia purpurea offered better protection in comparison to 50 mg/kg dose. Administration of D-GalN and LPS caused considerable upsurge of serum triglyceride, cholesterol and bilirubin, whereas level of albumin, glucose and hepatic glycogen were dwindled abruptly (Table 6). All the three doses of Tephrosia purpurea instigated reduction in triglycerides, cholesterol and bilirubin while increment in albumin, glucose and hepatic glycogen was noted towards their respective control. The 100 mg/kg dose was better than 50 mg/kg in reducing triglycerides and bilirubin level. The 200 mg/kg dose of Tephrosia purpurea appeared as better protective force in all the parameters except triglyceride and cholesterol in serum. It was also revealed that 200 mg/kg dose was better than 100 mg/kg dose in maintaining bilirubin and triglycerides. Effect of Tephrosia purpurea extract on oxidative stress The LPO was found to be enhanced drastically in hepatic and renal tissues (Fig. 7) after D-GalN and LPS intoxication (P£0.05). Pretreatment of Tephrosia purpurea at all the three doses repressed hepatorenal LPO towards control (P£0.05). Tephrosia purpurea at 100 and 200 mg/kg dose was more effective than 50 mg/kg dose in decreasing LPO in both tissues. Dose of 200 mg/kg was more efficient than 100 mg/kg in reducing LPO. Effect of Tephrosia purpurea extract on antioxidant status The GSH was significantly demoted (Fig. 7) in hepatorenal tissues after D-GalN and LPS exposure (P£0.05). Administration of Tephrosia purpurea increased GSH level in all the tissues at 50 mg/kg dose. Tukey’s post hoc analysis disclosed the fact that Tephrosia purpurea at all doses showed similar recovery pattern in uplifting tissue GSH level. The 100 mg/kg dose of Tephrosia purpurea showed no significant disparity with control when hepatic GSH was considered only. The 200 mg/kg dose of Tephrosia purpurea has no such difference with control even the renal GSH was considered too. Enzymatic activity of superoxide dismutase and catalase (Fig. 7) in hepatic and renal tissues was severely declined (P£0.05) after exposure to D-GalN and LPS. In Tephrosia purpurea treated groups, activity of superoxide dismutase and catalase was found to be enhanced in dose dependent manner. The 100 mg/kg dose of Tephrosia purpurea was better than 50 mg/kg dose in recovering enzymatic activity of hepatic superoxide dismutase and catalase. The 200 mg/kg dose had better protective efficacy in enhancing superoxide dismutase and catalase activities towards their respective control. Effect of Tephrosia purpurea extract on tissue lipid profile Effect of D-GalN and LPS administration on triglycerides and cholesterol in hepatic and renal tissues (P£0.05) have been summarized in Fig. 8. A sharp rise in triglycerides and cholesterol in hepatic and renal tissues was observed. The 100 mg/kg dose of Tephrosia purpurea significantly reversed renal triglycerides and hepatic cholesterol as compared to 50 mg/kg dose. The 50 and 200 mg/kg dose of Tephrosia purpurea showed significant differences in recovering hepatorenal triglycerides and cholesterol. The 100 and 200 mg/kg dose showed almost identical recovery pattern. Tephrosia purpurea at 200 mg/kg dose treated group reversed the variables very much near to control. Effect of Tephrosia purpurea extract on microsomal LPO, protein and CYP2E1 activity Acute hepatic injury induction by D-GalN and LPS resulted amplified microsomal LPO and CYP2E1 activity (P£0.05), whereas microsomal protein was severely diminished (Fig. 9). Tephrosia purpurea treatment resettled microsomal LPO, protein and CYP2E1 activity towards control. Tephrosia purpurea at 100 and 200 mg/kg dose were better when compared to 50 mg/kg dose in maintaining LPO and CYP2E1 activity. Tephrosia purpurea showed dose dependent recovery pattern for hepatic CYP2E1 activity. All the altered values were found very near to control in 200 mg/kg dose of Tephrosia purpurea treated group. Histopathological observations Photomicrographs of hepatic and renal histological observations suggested structural component disorganization in D-GalN and LPS exposed group. Prophylactic treatment with Tephrosia purpura at three different doses appreciably protected hepato-renal tissues from these alterations. The hepatic and renal histological observations were explained in legends to figures 10 and 11, respectively. Quantitative representation of variables to assess hepato-renal injury as well as improvement has been presented in table 7 and 8 respectively. It was observed from above-mentioned variables that, 200 mg/kg dose of Tephrosia purpurea extract showed better efficacy than the other two lower doses. Thus, only 200 mg/kg dose of Tephrosia purpurea extract treated groups were further considered for the determination of antioxidant enzymes of GSH cycle, electron microscopy, genotoxicity, proinflammatory cytokines (TNF-α and IL-6) and RBC degradation cycle including HO-1. Effect of Tephrosia purpurea extract on enzymes of GSH cycle Effects of D-GalN and LPS administration on hepatorenal enzymatic antioxidant status (P£0.05) were summarized in Fig. 12. After 16 hours of D-GalN and LPS treatment animals showed significant decrement in glutathione reductase, glutathione peroxidase, glucose-6-phosphate dehydrogenase and glutathione-S-transferase activities was observed in hepatorenal tissue at 5% level of significance. Oral administration of Tephrosia purpurea exhibited significant reversal in enzymatic activities of glutathione reductase, glutathione peroxidase, glucose-6-phosphate dehydrogenase and glutathione-S-transferase ( P £ 0.05 ) in liver and kidney. Tephrosia purpurea showed no substantial difference with control in recovering all these enzymatic antioxidants except for renal glutathione peroxidase. Effect of Tephrosia purpurea extract on proinflammatory cytokines Administration of D-GalN and LPS significantly induced inflammation via producing proinflammatory cytokines including TNF-α and IL-6 in serum (Fig. 13). The 200 mg/kg dose of Tephrosia purpurea extract significantly (P £ 0.05) protected the elevation of these cytokines. Effect of Tephrosia purpurea extract on RBC degradation cycle Table 9 depicted therapeutic effect of Tephrosia on the components of RBC degradation cycle against toxic exposure to LPS and D-GalN. The bilirubin, biliverdin, Hb, heme, free iron (ferritin and hemosiderin) and heme iron along with biliverdin reductase and HO-1 activities in the hepatic tissue were significantly elevated (Table 9) after D-GalN and LPS intoxication (P £ 0.05). Tephrosia purpurea at 200 mg/kg dose showed therapeutic effect on all the variables by reducing their level towards control in liver cells except HO-1 activity. The HO-1 activity was further induced up to 3.4 folds by Tephrosia purpurea at 5% level of significance as compared to control. The therapy of Tephrosia purpurea at 200 mg/kg dose was able to recover most of the variables altered due to inflammation and oxidative stress induced by LPS and D-GalN. Ultra-structural observations Electron microscopic study of liver and kidney have been presented in figure 14. The figure of control liver depicted regular hepatocytes with obvious nuclear membrane (NM) with visible nuclear pores (NP), dense cytoplasm, numerous well-formed mitochondrion (M) and rough endoplasmic reticulum (RER) around nuclear membrane (Fig. 14A). The D-GalN and LPS administered rat (Fig. 14B) showed loss of nuclear membrane integrity, dilated endoplasmic reticulum and mitochondrial degeneration. Loss of genetic material as indicated by the electron lucent area inside the nucleus and steatosis indicated by deposition of lipid droplets (LD) were also observed (Fig. 14B). Prophylactic treatment with Tephrosia purpurea extract at 200 mg/kg dose (Fig. 14C) showed almost regular shaped nucleus with well-maintained nuclear membrane and nucleolus. Homogenous distribution of euchromatin throughout the nuclear area, regular shaped and more dense mitochondria were also observed along with fine network of ER attached to the nucleus. Decreased steatosis and more dense cytoplasm were noticed as compared to the D-GalN and LPS exposed group. Electron micrograph obtained from transmission electron microscopy revealed that control kidney showed its regular architecture with well-defined membranes, mitochondrial abundance and peroxisomes with circular outline and homogenous content (Fig. 14D). The D-GalN and LPS administered rats showed swelling in mitochondria, loss of cytoplasmic and mitochondrial structural integrity, and lipid droplet accumulation in the cytoplasm (Fig. 14E). Prophylactic treatment with 200 mg/kg dose of Tephrosia purpurea showed better mitochondrial structure, reduced lipid droplet accumulation and peroxisome with homogenous content (Fig. 14F). Effect of Tephrosia purpurea extract on genetic material Exposure to D-GalN and LPS caused severe damage to DNA in hepatorenal tissue. DNA damage was studied in terms of comet assay (Fig. 15) and its numeric data is presented in table 10. Exposure to D-GalN and LPS induced fragmentation of DNA in liver and kidney (Fig. 15, A-F). A significant increase in tail length and high movement translated it into comet and was considered as DNA damage. Comparison between control group (Fig. 15, A and D) and D-GalN + LPS (Fig. 15, B and E) administered group suggested that D-GalN and LPS administration causes significant increase in tail length, tail moment and tail DNA % in hepatorenal tissues (Table 10). Therapy of 200 mg/kg dose of Tephrosia purpurea controlled DNA damage significantly as indicated by decreased tail length and tail moment (Fig. 15, C and F). Discussion Bioactive phytoconstituents are well-known for their protective roles against a variety of health problems including acute organ injury. As Tephrosia purpurea extract have major phytoconstituents including rutin, gallic acid, quercetin and many more, they might be responsible for its protective efficacy in this present study [ 19 ]. Furthermore, presence of single or multiple peaks in the UV-Vis spectra is a clear indication of the presence of unsaturated groups and heteroatoms such as Sulphur, Nitrogen and Oxygen. The peak at positions 660 nm indicated presence of organic chromophores [ 49 ]. The loss of volatile products in the first step of thermal decomposition below 200°C is mainly due to free water and ethanol, as this event occurs in fusion tracks and with vaporization of the substance [ 50 ]. The major weight loss between 200°C and 500°C may be due to the degradation of a wide variety of secondary metabolites, principally phenolics [ 51 ]. The FT-IR is the most suitable technique of the non-destructive spectroscopic methods in the analysis of pharmaceutical solids [ 50 ]. Since the extract was hydroethanolic, ethanol and water bands could be present in FTIR spectra. The strong absorption band between 3200–3400 cm − 1 indicated the presence of polymeric hydroxyl derivatives [ 52 ]. Carboxylic acids have an absorbance range of 2500–3300 cm − 1 , thus, bands at 3216, 2917 and 2849 cm − 1 could be due to OH stretching of carboxylic acids [ 53 ]. Carboxylic acid serves as a main pharmaceutical product in curing many health problems, including ulcers, jaundice, stomatitis, fever, wounds, edema and rheumatic joint pains [ 52 ]. The more intense bands occurring at 2917 cm − 1 and 2849 cm − 1 corresponding to C–H stretching vibrations could also be attributed to the presence of carbonyl functional groups [ 52 ] or methylene group of aliphatic compounds [ 54 ]. The absence of any peak between 2220 and 2260 cm − 1 indicated absence of any toxic substances as this region comprises of cyanide groups [ 52 ]. A band at 1587 cm − 1 could be attributed to C = C bonds in the aromatic rings due to presence of flavonoids and amino acids [ 50 , 55 ]. The bands at 1403 cm − 1 would be related to C-H bending vibration of methyl group. The band at 1285 cm − 1 is probably related to C–O stretching of polyols [ 55 ]. The band at 1178 cm − 1 and 1044 cm − 1 could be related to C–O stretching and –OH deformation of primary and tertiary alcohols respectively [ 56 ]. This band could also be related to C–O stretching of ether, esters and carboxylic acids. This indicated the presence of a wide variety of metabolites, such as tannins, flavonoids, carbohydrates (polysaccharides) and anthraquinones among others [ 57 – 59 ]. The band at 523 cm − 1 indicated the presence of iodate or brominate compounds [ 54 ], which plays role of disinfectant [ 52 ]. The XRPD analysis of Tephrosia purpurea extract exhibits several size dependent features leading to an irregular peak position, height and width. The sharp peak reveals that the extract is crystalline in nature and the particles are in the Nano regime [ 60 ]. The remaining peaks may be due to other phytochemicals present in Tephrosia purpurea extract [ 61 ]. Studies have suggested that D-GalN and LPS induced hepatorenal injury involves many mechanisms, including oxidative stress, inflammation, apoptosis and necrosis [ 62 ]. Compromised cellular integrity due to increased LPO, resulted leakage of cellular constituents especially transaminases (AST and ALT), ALP, bilirubin and albumin into serum impairing liver functions [ 63 ]. Various blood components such as WBC, RBC, platelets and hemoglobin gets altered due to sepsis, caused by LPS [ 19 , 63 ]. Recycling of excess purines due to necrosis increased serum urea, uric acid and creatinine, a clear indication of renal injury [ 64 ]. Increase in CYP activity could be attributed to p450 functional failure due to increased requirement of D-GalN and LPS biotransformation with a decrease in protein synthesis and accumulation of triglycerides [ 19 ]. Recent studies have shown that antioxidant and anti-inflammatory agents can ameliorate inflammatory responses of LPS induced injury [ 5 ]. Tephrosia purpurea mainly stabilized the membrane [ 15 ] to prevent cellular leakage [ 65 ] probably due to the presence of flavonoids that influenced hepatic regeneration [ 66 , 67 ]. Multiple beneficial functional groups present in Tephrosia purpurea extract, as evident in FTIR, might play crucial role in regulating hematological variables, liver and kidney functions. The D-GalN and LPS trigger excessive reactive oxygen species (ROS), leading to serious oxidative insult in liver injury via lipid peroxidation [ 62 ]. Antioxidant molecules such as glutathione, SOD and catalase neutralized oxidative stress by quenching free radicals [ 67 ]. The SOD eliminated superoxide anions and produced H 2 O 2 , which was then decomposed by catalase [ 68 ]. Tephrosia purpurea protects the depletion of these antioxidant molecules by acting as an antioxidant and provided free electron [ 9 , 66 ]. GPx harvested electron from glutathione and neutralized free radical by giving free electron, whereas, GR harvested electron from NADPH and reduced oxidized glutathione to make it available again for GPx. Thereafter, G6PDH obtained electron from glucose 6 phosphate, simultaneously converting NADP + into NADPH by giving that electron. Proper regulation of this cycle helped to maintain antioxidant status, but as cellular ROS increased due to D-GalN and LPS, components of this cycle gets depleted in maintaining redox balance [ 69 ]. Glutathione-S-transferase enabled conjugation of glutathione with toxic intermediates to remove them successfully out of the system. Depletion of glutathione-S-transferase, glutathione and CYP2E1 resulted as exposure to D-GalN and LPS produced a number of toxic intermediates upon metabolism [ 70 ]. Tephrosia purpurea act as a modifier of oxidant response which responded against oxidant-mediated generation of oxidative stress [ 71 ]. These enzymes related to GSH cycle were well maintained by Tephrosia purpurea as this plant might have the ability to form conjugates with toxic intermediates and scavenge free radicals [ 72 , 73 ]. The antihyperlipidemic activity of Tephrosia purpurea is might be due to presence of saponins and flavonoids. Tephrosia purpurea prevented elevation of triglyceride and cholesterol in D-GalN and LPS exposed animal by increasing the lipoprotein lipase and inhibition of HMG Co A reductase activities respectively [ 74 ]. When D-GalN and LPS reaches to the liver, massive pro-inflammatory cytokines including interleukin (IL)-6, IL-1β, and tumor necrosis factor-alpha (TNF-α), were released, leading to serious inflammation damage [ 62 ]. The anti-inflammatory activity of Tephrosia purpurea helps to reduced inflammation associated hepatorenal injury and maintaining cellular integrity [ 75 ]. High production of ROS caused DNA fragmentation and denaturation, thereby, morphological and structural appearance of the nucleus had changed [ 76 ]. In the comet assay, the size of the comet's tail signifies DNA damage and the amount of DNA fragments that has migrated from the nucleus determined the tail length [ 77 ]. Tephrosia purpurea might have quenched free radicals from the nucleus and provided stability to the membrane. As the particles present in the extract are nanosized, they might have interacted with DNA to stabilize it [ 78 ]. The rapid degradation of RBCs in the liver is associated with enormous oxidative stress and inflammation, resulted in massive production of heme and iron [ 79 ]. Unregulated heme contributed towards oxidative injury via ROS generation as the iron from the heme porphyrin ring is rapidly lost and contributes in ferrous state [ 80 ]. Furthermore, ferritin and hemosiderin were released as an acute phase protein to store free intracellular iron [ 81 , 82 ]. This increased iron deposition in liver which lead to tissue damage and dysregulation of function resulted in increased morbidity and mortality [ 83 ]. Tephrosia purpurea extract reduced iron overload in liver might be due to having good metal chelating ability [ 84 ]. The HO-1 is not only involved in normal physiology but also has a role in pathophysiological states [ 85 ]. LPS administration induced release of HO-1 in liver from 6–24 hours [ 4 ], but excessive oxidative stress and free heme could also induce HO-1 [ 86 ]. Induced HO-1 decreased macrophage infiltration, pro-oxidant and proinflammatory transcription factors to reduce oxidative stress and inflammation in order to protect organs during sepsis. Thus, upregulation of HO-1 is linked with decreased injury during endotoxemia [ 4 ]. Tephrosia purpurea interfere with RBC degradation and manages to induce HO-1 enzyme up to many folds. HO-1 reduced oxidative stress and inflammation by activating various antioxidant genes to employ cyto-protection [ 62 ]. This induced activity of HO-1 may influence other NADPH and oxygen-consuming pathways to reduce oxidative stress [ 87 ]. Increased conversion of heme into biliverdin and finally into unconjugated bilirubin by biliverdin reductase leads to the depletion of biliverdin reductase and increased biliverdin accumulation in hepatocytes [ 88 ]. Glucuronidation of bilirubin is an essential process to produce conjugated bilirubin for excretion. But, the depletion of uridine-diphosphoglucuronic glucuronosyl transferase (UDPGT) due to D-GalN administration resulted accumulation of unconjugated bilirubin [ 88 , 89 ]. Administration of Tephrosia purpurea might have smoothen the conversion of bilirubin from biliverdin by acting as an alternative of NADPH and glucuronidation of bilirubin by reducing depletion of UDPGT to minimize the accumulation [ 90 , 91 ]. Similar observation was also reported by Sahu et al., [ 75 ]. The phytocompounds present in the hydroalcoholic extract of Tephrosia purpurea might be responsible for protecting tissue from oxidative stress and inflammation induced injury by neutralizing those free radicals while enhancing the body’s defense systems against degenerative diseases [ 92 , 93 ]. Conclusion The pharmaceutical industry disposes of valuable tools used for quality control of products of plant origin and raw materials which require attentive characterization. Characterization of Tephrosia purpurea provides important information about its stability, composition, nature and others. This information is valuable to insure its quality and, therefore, its safety and efficacy. The exposure to D-GalN and LPS resulted increase in the level of oxidative stress and inflammation, which leads to acute organ injury. Treatment with bioactive component rich extract of Tephrosia purpurea demonstrated inhibition of D-GalN and LPS induced inflammatory response and oxidative stress. Preventive effect was supported by regular serological and hematological values, improved histoarchitecture, ultrastructure, comet assay photographs, induced HO-1, improved enzymatic antioxidant status and controlled lipid peroxidation in vital organs. These findings may provide a new insight to understand the molecular mechanism behind the anti-inflammatory and anti-oxidative action of Tephrosia purpurea , and may provide a better understanding of its use as a traditional medicine to treat various inflammatory and oxidative stress disorders. Abbreviations ALF = Acute liver failure; ALI = Acute liver injury; ALP = Alkaline phosphatase; ALT = Alanine amino transferase; ANOVA = Analysis of variance; AST = Aspartate amino transferase; CaCl 2 = Calcium chloride; CAT = Catalase; CYP2E1 = Cytochrome p 450 2E1; D-GalN = D-galactosamine; DSC = Differential scanning colorimetry; FTIR = Fourier transform infrared spectroscopy; G6PDH = Glucose 6 phosphate dehydrogenase; GPx = Glutathione peroxidase; GR = Glutathione reductase; GSH = Reduced glutathione; G-S-T = Glutathione S transferase; H&E = Hematoxylin and eosin; Hb = Hemoglobulin; HCT = Hematocrit; HO-1 = Hemoxygenase-1; HSD = Honestly significant difference; IL-6 = Interleukin 6; KCl = Potassium chloride; LPO = Lipid peroxidation; LPS = Lipopolysaccharide; NaCl = Sodium chloride; PLT = Platelets; PSA = Particle size analysis; RBC = Red blood cell; ROS = Reactive oxygen species; RP-HPLC = Reverse phase high performance liquid chromatography; SE = Standard error; SOD = Superoxide dismutase; TBARS = Thio-barbituric acid reactive substances; TGA = Thermogravimetric analysis; TNF-α = Tumor necrosis factor alpha; TP = Tephrosia purpurea; UV-Vis = Ultra violet-visible spectroscopy; WBC = White blood cell; XRPD = X-ray powder diffraction. Declarations Acknowledgements The first author greatly acknowledged electron microscopy division, AIIMS, New Delhi, India for providing facilities to carry out ultrastructural study. Funding: The first author greatly acknowledged Department of Zoology, Guru Ghasidas Vishwavidyalaya for providing Laboratory facilities and fellowship. Authors are also grateful to UGC, New Delhi [Startup Grant {20-1/2012(BSR)/20-12(3/2012) (BSR)] for partial financial assistance. Conflict of interests: There is no conflict of interest between authors for this manuscript. The manuscript is not communicated in any other journal. Contribution of authors: Samrat Rakshit: Conceptualization, Investigation, Formal analysis, Writing- Original draft preparation. Nisha Sahu: Investigation, Formal analysis. Satendra Kumar Nirala: Conceptualization, Writing- Review and editing, Supervision, Validation. Shamli S Gupte: Formal analysis. Sangeeta Shukla: Supervision, Resources. 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Tables Table 1: Multiple bioactive constituents are present in the hydroethanolic extract of Tephrosia purpurea Peak # Retention time Phytoconstituents Area Hight Area % Hight % 1 2-4 Tannic acid, L-ascorbic acid, catechin, gallic acid and caffeic acid Not measured ٭ Not measured ٭ Not measured ٭ Not measured ٭ 2 4.020 Rutin 50067 2964 1.048 2.765 3 4.542 Vanillic acid 181386 11854 3.796 11.058 4 5.442 Morin and malic acid 14559 903 0.305 0.842 5 6.260 Quercetin 508192 11930 10.635 11.129 6 7.349 Ellagic acid 55092 7091 1.153 6.615 7 9.581 Unknown 809107 23292 16.933 21.728 8 12.026 Unknown 342990 5229 7.178 4.878 9 18.342 Unknown 1864374 31651 39.017 29.525 10 23398 Unknown 952551 12287 19.935 11.462 ٭ There can be many phytoconstituents present between retention time 2 to 4, so, we did not measured the area and height of the peak. Table 2: Particle size distribution statistics analysis of Tephrosia purpurea extract Cum Q3 (%) 100.000 99.834 94.601 76.789 54.075 29.834 20.360 0.039 Diff q3(%) 0.166 5.232 17.813 22.713 24.242 9.474 7.525 0.035 Particle diameter (μm) 0.071 0.063 0.056 0.050 0.044 1.949 1.949 0.031 Median Diameter (μm) 0.043 Modal Diameter (μm) 0.039 Mean Value (μm) 0.043 Standard Deviation 0.079 Refractive Index 1.70-0.20i Median: The size of particle where the cumulative line crosses the 50th percentile; Modal: The size of particle that is most often seen; Mean Value: The average size of the particles; Standard Deviation: The amount of variance, in microns, between each trial; Cum Q3 (%) = Cumulative %: The percent of particles below that micron size; Diff q3 (%) = Differential Distribution: The percent of particles at that micron size. Table 3: Structural analysis of Tephrosia purpurea extract by FTIR spectrum Wave numbers (cm -1 ) Vibration Assignment 3216 cm −1 OH stretching Polymeric hydroxyl derivatives and carboxylic acids 2849 and 2917 cm −1 OH stretching C-H stretching Carboxylic acid Carbonyl functional groups Methylene group of aliphatic compounds 1587 cm −1 C=C stretching Aromatic rings of flavonoids and amino acids 1403 cm −1 C–H bending Methyl group 1285 cm −1 C–O stretching polyols 1178 cm −1 C–O stretching Primary alcohol, ether, ester and carboxylic acid 1044 cm −1 –OH deformation Tertiary alcohol 523 cm −1 C–I/Br stretching Iodide or Bromide Table 4: Protective effect of Tephrosia purpurea on hematological variables against D-GalN and LPS Parameters Groups Control TP per se TP 200 mg/kg D-GalN+LPS D-GalN+LPS+ TP 50 mg/kg D-GalN+LPS+ TP 100 mg/kg D-GalN+LPS+ TP 200 mg/kg Hemoglobin (g/dl) 15.4±0.71 15.5±0.71 11.4±0.52 * 12.6±0.58 * 14.1±0.65 14.9±0.69 Y Hematocrit (%) 44.2±2.03 45.1±2.07 31.5±1.45 * 39.1±1.8 41.1±1.89 Y 43.2±1.98 Y RBC (10 6 /mm 3 ) 7.66±0.35 7.58±0.35 5.75±0.26 * 6.75±0.31 7.18±0.33 Y 7.42±0.34 Y WBC (10 3 /mm 3 ) 11.9±0.55 11.8±0.54 16.8±0.77 * 14.7±0.68 * 13.2±0.61 Y 12.8±0.59 Y Platelets (10 3 /mm 3 ) 710±32.6 725±33.3 452±20.7 * 521±23.9 * 594±27.3 Y 651±29.8 Y j Lymphocyte Count (10 2 /mm 3 ) 28.1±1.29 29.3±1.34 63.7±2.92 * 61.3±2.81 * 56.4±2.59 * 43.1±1.98 Y * j F Monocyte Count (10 2 /mm 3 ) 6.80±0.31 6.70±0.31 16.2±0.74 * 9.7±0.45 Y * 9.20±0.42 Y * 8.70±0.40 Y Data are expressed as mean ± SE, (n=6). * P≤0.05 versus Control. Y P≤0.05 versus D-GalN+LPS. j P≤0.05 versus TP 50 mg/kg. F P≤0.05 versus TP 100 mg/kg. Table 5: Protective effect of Tephrosia purpurea on hepatic and renal functions against D-GalN and LPS Parameters Groups Control TP per se TP 200 mg/kg D-GalN+LPS D-GalN+LPS+ TP 50 mg/kg D-GalN+LPS+ TP 100 mg/kg D-GalN+LPS+ TP 200 mg/kg AST (IU/L) 81.3±3.73 74.1±3.40 471±21.6 * 295±13.5 Y * 201±9.23 Y * j 108±4.96 Y j F ALT (IU/L) 36.3±1.66 35.4±1.62 172±7.92 * 129±5.93 Y * 90.4±4.15 Y * j 51.6±2.37 Y j F ALP (IU/L) 193±8.86 184±8.45 389±17.8 * 297±13.7 * Y 248±11.4 * Y 212±9.75 Y j Urea (mg/dl) 29.1±1.34 28.8±1.32 70.6±3.24 * 58.9±2.70 Y * 44.1±2.02 Y * j 36.0±1.65 Y j Uric Acid (mg/dl) 1.41±0.06 1.31±0.06 5.01±0.23 * 3.97±0.18 Y * 2.87±0.13 Y * j 2.0±0.09 Y j F Creatinine (mg/dl) 0.70±0.03 0.65±0.03 2.03±0.09 * 1.91±0.09 * 1.42±0.07 Y * j 1.03±0.05 * Y j F Data are expressed as mean ± SE, (n=6). * P≤0.05 versus Control. Y P≤0.05 versus D-GalN+LPS. j P≤0.05 versus TP 50 mg/kg. F P≤0.05 versus TP 100 mg/kg. Table 6: Protective effect of Tephrosia purpurea on serum biochemical variables and hepatic glycogen against D-GalN and LPS Parameters Groups Control TP per se TP 200 mg/kg D-GalN+LPS D-GalN+LPS+ TP 50 mg/kg D-GalN+LPS+ TP 100 mg/kg D-GalN+LPS+ TP 200 mg/kg Glucose (mg/dl) 121±5.55 114±5.23 71.3±3.72 * 89.5±4.11 * 102±4.68 Y 110±5.06 Y j Triglyceride (mg/dl) 32.3±1.48 30.1±1.38 120±5.49 * 86.7±3.98 Y * 65.8±3.02 Y * j 51.5±2.36 * Y j F Cholesterol (mg/dl) 21.3±0.97 20.5±0.94 49.7±2.28 * 37.2±1.71 Y * 35.1±1.61 Y * 29.7±1.36 Y * j Total Bilirubin (mg/dl) 0.39±0.02 0.37±0.02 0.89±0.04 * 0.71±0.03 Y * 0.59±0.03 Y * j 0.49±0.02 Y j Direct Bilirubin (mg/dl) 0.1±0.01 0.11±0.01 0.70±0.03 * 0.54±0.02 Y * 0.39±0.01 Y * j 0.19±0.01 Y j F Albumin (g/dl) 4.61±0.21 4.74±0.22 2.41±0.11 * 2.89±0.13 * 3.51±0.16 * Y 4.01±0.18 Y j Glycogen (mg/100g) 3200±146 3185±146 1345±61.5 * 2339±107 Y * 2555±117 Y * 2736±125 Y Data are expressed as mean ± SE, (n=6). * P≤0.05 versus Control. Y P≤0.05 versus D-GalN+LPS. j P≤0.05 versus TP 50 mg/kg. F P≤0.05 versus TP 100 mg/kg. Table 7: Severity of liver injuries in different treatments. [─] no alterations; [+] mild alterations; [++] moderate alterations; [+++] severe alterations. Parameters Groups Control TP per se TP 200 mg/kg D-GalN+LPS D-GalN+LPS+ TP 50 mg/kg D-GalN+LPS+ TP 100 mg/kg D-GalN+LPS+ TP 200 mg/kg Necrosis ─ ─ +++ ++ + ─ Vacuolation ─ ─ +++ ++ + ─ Distorted hepatic cords ─ ─ +++ + ─ ─ Inflammatory cell infiltration ─ ─ +++ ++ + ─ Pyknotic nuclei ─ ─ +++ ++ + ─ Damaged central vain ─ ─ +++ + ─ ─ Table 8: Severity of kidney injuries in different treatments. [─] no alterations; [+] mild alterations; [++] moderate alterations; [+++] severe alterations. Parameters Groups Control TP per se TP 200 mg/kg D-GalN+LPS D-GalN+LPS+ TP 50 mg/kg D-GalN+LPS+ TP 100 mg/kg D-GalN+LPS+ TP 200 mg/kg Tubular dilatation ─ ─ +++ ++ + ─ Tubular degeneration ─ ─ +++ ++ + ─ Tubular vacuolization ─ ─ ++ ++ + ─ Glomerular damage ─ ─ +++ + ─ ─ Loss of glomerular space ─ ─ +++ + ─ ─ Distortion of endothelial lining ─ ─ + ─ ─ ─ Table 9: Protective effect of Tephrosia purpurea on RBC degradation cycle in liver Groups Hb (mg/100g liver) Heme (µg/g liver) Ferritin (µg Fe/g liver) Hemosiderin (µg Fe/g liver) Free iron (µg Fe/g liver) Total Iron (µg Fe/g liver) Bilirubin (µg/100 g liver) Biliverdin (µg/100 g liver) Biliverdin reductase (p mol bilirubin /min/mg protein) HO-1 (p mol bilirubin /h/mg protein) Control 7.39±0.48 8.48±0.55 39±2.53 20±1.29 140±9.08 199±12.9 477±30.9 182±11.8 3.52±0.23 21.1±1.37 LPS+D-GalN 12.8±0.83 * 50.7±3.29 * 162±10.5 * 83±5.39 * 545±35.4 * 790±51.3 * 865±56.1 * 394±25.6 * 7.88±0.51 * 47.3±3.07 * LPS+D-GalN+ TP 200 mg/kg 8.4±0.55 Y 21.3±1.38 Y * 52±3.38 Y 33±2.14 Y * 175±11.4 Y 260±16.9 Y 546±35.4 Y 217±14.1 Y 3.92±0.25 Y 91.8±5.96 Y * Data are expressed as mean ± SE, (n=6). * P≤0.05 versus Control. Y P≤0.05 versus D-GalN+LPS. j P≤0.05 versus TP 50 mg/kg. F P≤0.05 versus TP 100 mg/kg. Table 10: Protective effect of Tephrosia purpurea on hepato-renal comet assay Groups Parameter Hepatic Renal Tail length Tail movement Tail DNA (%) Tail length Tail movement Tail DNA (%) Control 0.1±0.005 0.002±0.0001 10.0±0.50 0.29±0.015 0.001±0.0001 7.00±0.35 LPS+D-GalN 22.0±1.11 * 21.4±1.08 * 97±4.88 * 24.0±1.21 * 23.9±1.20 * 82.0±4.12 * LPS+D-GalN+ TP 20 mg/kg 16.0±0.81 Y * 13.6±0.68 Y * 41±2.06 Y * 14.0±0.70 Y * 13.2±0.66 Y * 47.0±2.36 Y * Data are expressed as mean ± SE, (n=6). * P≤0.05 versus Control. Y P≤0.05 versus D-GalN+LPS. j P≤0.05 versus TP 50 mg/kg. F P≤0.05 versus TP 100 mg/kg. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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condition\u003c/strong\u003e\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig01.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/71e65c8e66c97b0df7d7c3f7.png"},{"id":21565601,"identity":"8588b266-df1a-4c3d-8641-ae17b33ad0ba","added_by":"auto","created_at":"2022-05-17 15:24:58","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":8480,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUV-VIS absorption spectra of hydroethanolic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Fig02.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/f30d82e93f4e7055ed001abd.png"},{"id":21565603,"identity":"be3bb875-1bfe-4ee3-98d0-5aa612ffa01b","added_by":"auto","created_at":"2022-05-17 15:24:58","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":21614,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eParticle size distribution histogram of hydroethanolic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig03.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/273f402c0bc3c42cf23ea4a6.png"},{"id":21565378,"identity":"114dd0d2-2773-4b7f-9b88-421902a7d47a","added_by":"auto","created_at":"2022-05-17 15:19:58","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":28604,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eTGA and DSC analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract \u003c/strong\u003e\u003c/p\u003e\u003cp\u003e(A) \u0026nbsp;Thermo gravimetric scan of \u003cem\u003eTephrosia purpurea\u003c/em\u003e\u003c/p\u003e\u003cp\u003e(B)\u0026nbsp;\u0026nbsp;Differential scanning colorimetry scan of \u003cem\u003eTephrosia purpurea\u003c/em\u003e\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig04.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/c00f29be7dda70633a727d07.png"},{"id":21565373,"identity":"df06ca8c-b927-45da-a229-3ca9526f3a5a","added_by":"auto","created_at":"2022-05-17 15:19:58","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":2989,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eXRD patterns of the crystal structure of \u003cem\u003eTephrosia purpurea\u003c/em\u003e hydroethanolic extract\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig05.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/58bf5046612e41e2cb664c3f.png"},{"id":21566312,"identity":"eb00b39e-3068-43a1-a7b9-973369aafe42","added_by":"auto","created_at":"2022-05-17 15:34:58","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":10092,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eFTIR spectra of hydroethanolic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e \u003c/strong\u003e\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig06.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/6510557c85725fbcc5e877b1.png"},{"id":21565607,"identity":"f31a9145-1afa-4779-943b-9259fe5e8a70","added_by":"auto","created_at":"2022-05-17 15:24:58","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":178338,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective efficacy of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on lipid peroxidation, reduced glutathione, superoxide dismutase and catalase activity on hepatic and renal tissue against D-GalN and LPS induced oxidative stress \u003c/strong\u003e\u003c/p\u003e\u003cp\u003eData are expressed as mean ± SE, (n=6). \u003csup\u003e\u003cstrong\u003e*\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus Control. \u003csup\u003e\u003cstrong\u003eY\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus D-GalN+LPS. \u003csup\u003e\u003cstrong\u003ej\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 50 mg/kg. \u003csup\u003e\u003cstrong\u003eF\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 100 mg/kg. I=Control group. II= TP 200 mg/kg group. III= D-GalN+LPS group. IV= D-GalN+LPS+ TP 50 mg/kg group. V= D-GalN+LPS + TP 100 mg/kg group. VI= D-GalN+LPS + TP 200 mg/kg group.\u003c/p\u003e","description":"","filename":"Fig07.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/5c595354a96280ec7c267cb1.png"},{"id":21565610,"identity":"b30180a1-94aa-4a68-aa60-e6c9569790cc","added_by":"auto","created_at":"2022-05-17 15:24:59","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":75719,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective potential of \u003cem\u003eTephrosia purpurea\u003c/em\u003e against D-GalN and LPS induced changes in hepatorenal triglyceride and cholesterol contents\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eData are expressed as mean ± SE, (n=6). \u003csup\u003e\u003cstrong\u003e*\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus Control. \u003csup\u003e\u003cstrong\u003eY\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus D-GalN+LPS. \u003csup\u003e\u003cstrong\u003ej\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 50 mg/kg. \u003csup\u003e\u003cstrong\u003eF\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 100 mg/kg. I=Control group. II= TP 200 mg/kg group. III= D-GalN+LPS group. IV= D-GalN+LPS+ TP 50 mg/kg group. V= D-GalN+LPS + TP 100 mg/kg group. VI= D-GalN+LPS + TP 200 mg/kg group.\u003c/p\u003e","description":"","filename":"Fig08.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/ea2a344eb6cf4c30796ebd9f.png"},{"id":21567277,"identity":"d0803cb5-9933-4817-b973-8e74f2e5fa48","added_by":"auto","created_at":"2022-05-17 15:39:58","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":95907,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective efficacy of \u003cem\u003eTephrosia purpurea\u003c/em\u003e against D-GalN and LPS induced lipid peroxidation, CYP2E1 activity and protein on hepatic microsomes\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eData are expressed as mean ± SE, (n=6). \u003csup\u003e\u003cstrong\u003e*\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus Control. \u003csup\u003e\u003cstrong\u003eY\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus D-GalN+LPS. \u003csup\u003e\u003cstrong\u003ej\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 50 mg/kg. \u003csup\u003e\u003cstrong\u003eF\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 100 mg/kg. I=Control group. II= TP 200 mg/kg group. III= D-GalN+LPS group. IV= D-GalN+LPS+ TP 50 mg/kg group. V= D-GalN+LPS + TP 100 mg/kg group. VI= D-GalN+LPS + TP 200 mg/kg group.\u003c/p\u003e","description":"","filename":"Fig09.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/44e621c2e32647a6b3b07a2c.png"},{"id":21565381,"identity":"db3e9dbd-4c4d-4a27-8171-0c41776bd682","added_by":"auto","created_at":"2022-05-17 15:19:58","extension":"png","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":628634,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHistological observation of rat liver sections under light microscope\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eA: control (10X); B: control (40X); C: TP \u003cem\u003eper se\u003c/em\u003e (10X); D: D-GalN+ LPS (10X); E and F: D-GalN+ LPS (40X); G: TP 50 (10X); H: TP 50 (40X); I: TP 100 (10X); J: TP 100 (40X); K: TP 200 (10X); L: TP 200 (40X); K: Kupffer cell; N: Nucleus; ND: Nuclear division;\u0026nbsp;Figure A and B shows regular structure of liver with tightly packed pink stained hepatocytes. The sinusoidal space (SS) can be seen as pale stained spaces between the cord of hepatocytes. Micrograph C shows hepatic architecture with central vein (CV), array of hepatocytes (H) and sinusoidal space (SS). Photomicrograph D, E and F shows hepatic centrilobular mononuclear cell infiltration (LI), distortion in shape of central vein and dilation in sinusoidal spaces (SS). Figures G-H, I-J and K-L shows a gradual recovery pattern with increasing dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e. Well-shaped CV, surrounded by array of hepatocytes, proper sinusoidal spaces and lesser number of neutrophils were also observed.\u0026nbsp;\u0026nbsp;\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig10.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/54b3896ce5c09c0ab70b4ae3.png"},{"id":21566313,"identity":"b3b55e30-e367-40f7-91e1-4e85d8d77eeb","added_by":"auto","created_at":"2022-05-17 15:34:58","extension":"png","order_by":11,"title":"Figure 11","display":"","copyAsset":false,"role":"figure","size":697070,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHistological observation of rat kidney sections under light microscope\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eA: control (10X); B: control (40X); C: TP \u003cem\u003eper se\u003c/em\u003e (10X); D: TP \u003cem\u003eper se\u003c/em\u003e (40X); E: D-GalN+ LPS (10X); F: D-GalN+ LPS (40X); G: TP 50 (10X); H: TP 50 (40X); I: TP 100 (10X); J: TP 100 (40X); K: TP 200 (10X); L: TP 200 (40X); BC: Bowman’s capsule; CS: Capsular space; G: Glomerulus. Figure A-D shows peculiar histoarchitecture of kidney with normal glomerulus (G) and Bowman’s capsule (BC). Figure E and F of kidney treated with LPS and D-GalN indicated swelling in glomerulus that occupied maximum capsular space of Bowman’s capsule and disturbed cellular matrix of Bowman’s capsule. Representation of figure G-H, I-J, and K-L of kidney shows gradual recovery pattern increasing dose of \u003cem\u003eTephrosia \u003c/em\u003epurpurea as evident by well-shaped glomerulus with reduced swelling and more capsular spaces (CS). The cellular matrix of Bowman’s capsule of purpurin treated groups is similar to histological appearance of control group.\u003c/p\u003e","description":"","filename":"Fig11.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/53d6694441881595d1fd835b.png"},{"id":21567278,"identity":"de34e419-7891-4104-ba96-8a549f85eece","added_by":"auto","created_at":"2022-05-17 15:39:59","extension":"png","order_by":12,"title":"Figure 12","display":"","copyAsset":false,"role":"figure","size":95459,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective potential of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on D-GalN and LPS induced alteration in hepatorenal GSH cycle\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eData are expressed as mean ± SE, (n=6). \u003csup\u003e\u003cstrong\u003e*\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus Control. \u003csup\u003e\u003cstrong\u003eY\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus D-GalN+LPS. \u003csup\u003e\u003cstrong\u003ej\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 50 mg/kg. \u003csup\u003e\u003cstrong\u003eF\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 100 mg/kg. I=Control group. III= D-GalN+LPS group. VI= D-GalN+LPS + TP 200 mg/kg group.\u003c/p\u003e","description":"","filename":"Fig12.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/15802cc2c13344875e7226f6.png"},{"id":21565999,"identity":"89c86850-7cda-4fb0-857a-be4d554d70b3","added_by":"auto","created_at":"2022-05-17 15:29:58","extension":"png","order_by":13,"title":"Figure 13","display":"","copyAsset":false,"role":"figure","size":17223,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective potential of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on D-GalN and LPS induced alteration in serum TNF-α and IL-6\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eData are expressed as mean ± SE, (n=6). \u003csup\u003e\u003cstrong\u003e*\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus Control. \u003csup\u003e\u003cstrong\u003eY\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus D-GalN+LPS. \u003csup\u003e\u003cstrong\u003ej\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 50 mg/kg. \u003csup\u003e\u003cstrong\u003eF\u003c/strong\u003e\u003c/sup\u003eP≤0.05 versus TP 100 mg/kg. I=Control group. III= D-GalN+LPS group. VI= D-GalN+LPS + TP 200 mg/kg group.\u003c/p\u003e","description":"","filename":"Fig13.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/0d28caf9e8630d7b89627a89.png"},{"id":21566315,"identity":"d99f6cb2-2cbe-4aba-9492-d5a4fed4437b","added_by":"auto","created_at":"2022-05-17 15:34:58","extension":"png","order_by":14,"title":"Figure 14","display":"","copyAsset":false,"role":"figure","size":351790,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUltrastructural observation of rat liver and kidney under transmission electron microscope\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eFigure 14A (2550X) represents electron micrograph of liver of control group, figure 14B (2550X) represents liver of D-GalN and LPS exposed group and figure 14C (2550X) represents liver of prophylactic treatment with \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 200 mg/kg dose followed by D-GalN and LPS.\u003c/p\u003e\u003cp\u003eElectron micrograph of kidney of control group treated with vehicle only is shown in figure 14D (2550X), figure 14B (2550X) represents kidney of D-GalN and LPS exposed group and figure 14C (2550X) represents kidney of prophylactic treatment with \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 200 mg/kg dose followed by D-GalN and LPS.\u003c/p\u003e","description":"","filename":"Fig14.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/d787c8329d2e6ee93489da21.png"},{"id":21565384,"identity":"3e4c9162-bac7-44d1-ab16-df4fd87cd2b9","added_by":"auto","created_at":"2022-05-17 15:19:59","extension":"png","order_by":15,"title":"Figure 15","display":"","copyAsset":false,"role":"figure","size":348050,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProtective efficacy of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on D-GalN and LPS induced genotoxicity \u003c/strong\u003e\u003c/p\u003e\u003cp\u003ePhotographs of liver section obtained from comet assay analysis were presented from A to C, while kidney sections were presented from D to F. A and D: control; B and E: D-GalN + LPS only; C and F: TP 200 with D-GalN + LPS. Photomicrographs of control group shows round shaped nucleus with compact DNA. The D-GalN + LPS treated group shows formation of a comet shaped appearance of nuclear material in liver (B) and kidney (E) indicating DNA damage. Animals treated with \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 200 mg/kg dose showed lesser head and tail (comet) of nuclear material in liver (C) and kidney (F) as a sign of protection of DNA.\u003c/p\u003e","description":"","filename":"Fig15.png","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/42a8de303e493193e80d01ee.png"},{"id":21737046,"identity":"0f3d744b-9f74-402a-be9f-6e04e70d6c70","added_by":"auto","created_at":"2022-05-21 12:41:48","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4006682,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1596610/v1/3c8c0f2d-f0c6-4421-b3e2-f9475f1fdaae.pdf"}],"financialInterests":"","formattedTitle":"\u003cp\u003e\u003cem\u003eTephrosia purpurea\u003c/em\u003e (Linn.) attenuates progression of acute liver failure by reducing oxidative stress and inflammation via modulating cytoprotective enzyme hemoxygenase-1 activity\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAcute liver failure (ALF) or acute liver injury (ALI) is one of the severe forms of liver disease. Pathological characteristics include extensive hepatic necrosis with severe loss of parenchyma, violent oxidative stress, and sustained inflammation. Despite enormous clinical efforts, prognosis and high mortality rate haven\u0026rsquo;t improved significantly [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Administration of specific hepatotoxin D-galactosamine (D-GalN) in combination with endotoxin lipopolysaccharide (LPS) greatly induces acute liver injury and mimics liver disease in human. This model of acute liver injury is extensively used worldwide to study underlying mechanism of ALF and develop effective therapeutic drugs [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. It is indispensable that D-GalN and LPS induce oxidative stress and inflammatory responses, which promotes hepatic necrosis, contributing directly in the occurrence and progression of ALF. Therefore, taking care of increased oxidative stress and inflammation may be the first step for the treatment of ALF [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Cytoprotective enzymes such as heme oxygenase (HO)-1 play major role in the well-being of a cell under immense oxidative stress and inflammation. Thus, modulation of these cytoprotective enzymes related to oxidative stress and inflammation could be an effective therapeutic strategy for D-GalN and LPS induced ALF [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTraditional systems of medicine remain invaluable since ages due to their multiple health outcomes and minimal side effects as compared to modern drugs [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. These plants have long been thought to be potential hepatoprotective agents for their various biological activities including antioxidant and anti-inflammatory by regulating multiple signal molecules [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. The use of crude plant extracts as therapeutic supplement showed various plausible health benefits including amelioration of metabolic complications, oxidative stress and inflammation induced injury [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Plants possess many advantageous biologically active phytochemicals with minimal to no side effects, when taken as a part of human diet or at effective doses [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Phytochemicals present in plant extract often exert synergistic effect in improving the efficacy of plant extracts than administered alone. Thus, various plants are often screened for beneficial effect in ameliorating health problems [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cem\u003eTephrosia purpurea\u003c/em\u003e (Linn.) pers. (Fabaceae) is a self-generating tropical perennial herb, known as \u0026ldquo;Sarwawranvishapaha\u0026rdquo; in \u003cem\u003eAyurveda\u003c/em\u003e for its usefulness in hepatic and splenic disorders, phantom tumor, poisonous bite and almost all type of wounds [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. The usefulness of \u003cem\u003eTephrosia purpurea\u003c/em\u003e is also mentioned in an old \u003cem\u003eAyurvedic\u003c/em\u003e literature \u0026ldquo;Bhavaprakasha Nighantu Guduchyadi Varga, 169\u0026rdquo; that it can alleviate cough, cure blood related diseases, dyspnea and fever [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. In modern day, hepatoprotective formulations such as \u0026ldquo;Tephroli\u0026rdquo; and \u0026ldquo;Yakrifit\u0026rdquo; have \u003cem\u003eTephrosia purpurea\u003c/em\u003e as one of their major constituents [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e is reported to possess multiple phenolic compounds including glycosides, flavonoids and flavones, responsible for its various protective properties [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e showed a very high therapeutic index as the LD\u003csub\u003e50\u003c/sub\u003e of its ethanolic extract is 2000 mg/kg in rats [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTraditional healers claim that \u003cem\u003eTephrosia purpurea\u003c/em\u003e is useful in treating diabetes, rheumatism, asthma, diarrhea, ulcers [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], gonorrhea, bronchitis, impotency and other kidney and heart disorders [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. These claims about \u003cem\u003eTephrosia purpurea\u003c/em\u003e, in our traditional healing system need scientific validation to prove its effectiveness. To the best of our knowledge, no scientific literature is available that explains biophysical properties of \u003cem\u003eTephrosia purpurea\u003c/em\u003e and its effectiveness, particularly against D-GalN and LPS induced acute liver injury. Thus, present study was focused to investigate biophysical characteristics of \u003cem\u003eTephrosia purpurea\u003c/em\u003e and its therapeutic potential against D-GalN and LPS induced acute liver injury.\u003c/p\u003e"},{"header":"Materials And Methods","content":"\u003cp\u003e\u003cstrong\u003ePreparation of hydroethanolic extract of\u003cem\u003e\u0026nbsp;Tephrosia purpurea\u003c/em\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe plant was identified as \u003cem\u003eTephrosia purpurea\u003c/em\u003e (Linn.) pers. and a Voucher no: GGV/BOT/H/FAB/DKS/210 was obtained at Botany Department, Guru Ghasidas University, Chhattisgarh, India. Whole plant of \u003cem\u003eTephrosia purpurea\u003c/em\u003e was collected form Guru Ghasidas University campus and air dried in shade for 3 weeks and pulverized. 15 grams of pulverized sample was extracted by Soxhlet apparatus (worm method) for 30 minutes using 70% (v/v) ethanol in water. The hydroethanolic extract was concentrated in a rotary evaporator and lyophilized. The yield of the extract (2.9g, 19.33%) was obtained preserved for further use.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eReverse phase-HPLC instrumentation and chromatographic conditions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe isocratic reverse phase HPLC analysis was carried out in a Shimadzu HPLC system (LC-20AD) consisted with double pumps (LC 20-AD), column oven (CTO-20AC), sample injector system (SIL-20ACHT) and a spectrophotometric detector (SPD-20A) with a computer assisted analysis software (Shimadzu LC solution 1.25sp1). The stationary phase used for this analysis was a reverse-phase C18 column (150mm \u0026times; 4.6 mm, 5 𝜇m) and the mobile phase consisted of aqueous acid solution (0.5 M phosphoric acid) : methanol (50 : 50 v/v) mixture with a stable flow rate of 0.9 mL/min. The mobile phase was filtered through 0.45 \u0026mu;m membrane filter and degassed by ultrasonic bath prior to use. The temperature of the column was maintained at room temperature (25 \u0026plusmn; 2\u0026ordm;C). 20 𝜇L of each sample were filtered through a 0.22 \u0026mu;m nylon membrane before injecting into the HPLC system for analysis. The presence of bioactive phytochemicals including tannic acid, ascorbic acid, catechin, gallic acid, caffeic acid, rutin, vanillic acid, morin, malic acid, quercetin and ellagic acid were investigated. Presence of these compounds were confirmed by comparing chromatogram peaks of plant extract with reference standards by comparing their retention time. Stock solutions of 40 ppm for all the standard references and 2000 ppm for plant extract were prepared in mobile phase. The detection wavelength was 280 nm for tannic acid, 265 nm for ascorbic acid, 276 nm for catechin, 220 nm for gallic acid, 324 nm for caffeic acid, 260 nm rutin, 220 nm for vanillic acid, 370 nm for morin, 230 nm for malic acid, 256 nm for quercetin and 368 nm for ellagic acid. The detection wavelength for \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract was 269 nm. The absorption spectra were determined by UV-VIS spectroscopy.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eUV\u0026ndash;Vis spectra analysis\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eHydroethanolic (70% ethanol in water) extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 1000 ppm concentration was observed by UV\u0026ndash;Vis spectroscopy within the ranges between 200 nm and 800 nm to identify phytoconstituents containing \u0026sigma;-bonds, \u0026pi;-bonds and lone pair of electrons, chromophores and aromatic rings present in it.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eParticle size analysis (PSA)\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe crude plant extract, having various particle size ranges were determined using particle size analyzer (Shimadzu\u0026rsquo;s SALD- 2300). Sample was dissolved in 20 ml 70% ethanol followed by sonication for 10 minutes. Particle sizes were measured based on the time dependent scattering of laser light by the particles with a pump speed of 5.0 cm\u003csup\u003e3\u003c/sup\u003e/min.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThermogravimetry analysis (TGA)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe changes in mass of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract over the time with changes in temperature were assessed. Dynamic thermogravimetric curves of extract were obtained by thermogravimetric analysis with Shimadzu\u0026rsquo;s TGA -50, with 10\u0026deg;C/min heating rate ranging from 25\u0026deg;C to a temperature of 800\u0026deg;C under nitrogen atmosphere with a flow of 100 mL/min. The plant extract was packed in a platinum crucible and subjected to analysis. Calcium oxalate monohydrate was used for calibration of the instrument to characterize weight loss steps.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDifferential scanning colorimetry (DSC)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe DSC curves of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003ewere obtained using a Shimadzu calorimeter, model\u0026nbsp;Shimadzu\u0026rsquo;s DSC-60 Plus. The instrument was\u0026nbsp;coupled to a photovisual system and an oven with a temperature range of 20-300\u0026deg;C. The whole analysis was done under nitrogen atmosphere with constant flow of 100 ml/min and a heating rate of 5\u0026deg;C/min. The plant extract was packed in alumina\u0026nbsp;crucible\u0026nbsp;for analysis.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eX-ray diffraction (XRD)\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe structural analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract was carried out by using X ray diffraction techniques on Rigaku\u0026rsquo;s Mini Flex 600 diffractometer. The \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract was subjected to Cu-K\u0026alpha; radiations (\u0026lambda;=0.154056 nm, 2\u0026theta;= 2-90\u0026deg;) operating at a voltage of 40 kV and a current of 15 mA, at 10\u0026deg;C/min speed at a drive axis of 2\u0026theta;. The average size of the crystallite (D in nm) was also calculated from the XRD peak using Debye Scherrer\u0026rsquo;s relation [17, 18].\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/69519_bce2c0439cd956a6/69519_custom_files/img1652726192.PNG\"\u003e\u003c/p\u003e\n\u003cp\u003eWhere, \u003cem\u003ek\u003c/em\u003e= 0.9 is the\u0026nbsp;Scherrer\u0026rsquo;s constant\u003cstrong\u003e, \u0026lambda;=\u0026nbsp;\u003c/strong\u003e1.540562\u0026Aring; is wavelength of X-rays and \u0026beta; (in radian) is the full width of half maxima (FWHM) of the diffraction peak at an angle \u0026theta;.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFourier transform infrared spectroscopy (FT-IR)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFunctional groups are responsible for medicinal properties of plants. The whole plant extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e was subject to FT-IR analysis on a Perkin Elmer\u0026rsquo;s Spectrum Two using KBr stressed disks within a spectral range of 4000\u0026ndash;450 cm\u003csup\u003e\u0026minus;1\u0026nbsp;\u003c/sup\u003eto determine presence of possible functional groups.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExperimental animals\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eExperimental animals (\u003cem\u003eWistar\u003c/em\u003e rats, 150\u0026plusmn;10g) were purchased from Defense Research and Development Establishment, Gwalior, and kept in departmental animal house for 15 days to get acclimatized. Animals were exposed to a 12 h periodic light and dark cycle with a constant temperature about 25\u0026deg;\u0026plusmn;2\u0026deg;C. Rats were provided with palleted rat diet and water \u003cem\u003ead libitum\u003c/em\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAnimal right\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAnimal care and experimental procedures were carried out by sincerely following the guidelines set by the committee for the purpose of control and supervision of experiments on animal, India. The experimental procedure was approved by institutional animal ethics committee (994/Ere/Go/06/CPCSEA) on 01/08/2016 (Ref:162/IAEC/Pharmacy/2016).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eChemicals\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePure and analytical grade chemicals were procured from Sisco Research Laboratories Pvt. Ltd (SRL) and Himedia Laboratories Pvt. Ltd., India in this study. LPS (Lot # 114M4009V) was procured from Sigma Aldrich Co Ltd, USA whereas D-GalN (Batch # 3647503) was procured from SRL, India.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eD-GalN and LPS model of acute multiorgan injury\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe D-GalN and LPS was prepared in saline (0.9% NaCl in water). Acute organ injury was induced by D-GalN (300 mg/2ml/kg) and LPS (50\u0026micro;g/2ml/kg) administered intraperitonially as per previous scientific literature [19].\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePreparation of different doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFreeze dried plant extract was dissolved in distilled water using 1% (w/v) gum acacia as the suspending agent [19]. Three different doses of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003e(50 mg/kg, 100 mg/kg and 200 mg/kg) were taken for evaluation where the highest dose (200 mg/kg) corresponds to 1/10\u003csup\u003eth\u0026nbsp;\u003c/sup\u003eof the LD\u003csub\u003e50\u003c/sub\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExperimental design\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWell acclimatized thirty-six animals were divided equally into six groups. Group I: designated as control, received 1% gum acacia as vehicle. Group II: designated as TP\u003cem\u003e\u0026nbsp;per\u0026nbsp;\u003c/em\u003ese, received 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract. Group III: designated as D-GalN+LPS (experimental control), received 300 mg/kg dose of D-GalN followed by 50 \u0026micro;g/kg dose of LPS. Group IV-VI: designated as TP50, TP100 and TP200 respectively, received 50, 100 and 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract respectively along with D-GalN and LPS. Vehicle and different doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e were given orally for 6 days straight to their respective groups. The D-GalN followed by LPS (only after 1 hour of D-GalN) were given intraperitonially to group III to VI on the 6\u003csup\u003eth\u003c/sup\u003e day evening. All the animals were fasted overnight and sacrificed 16 hours after D-GalN and LPS treatment using diethyl ether. To generate an individual data point in each of the independent experiments, samples were used in triplicates.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHematological study\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBlood was collected by puncturing retro-orbital venous sinus into heparinized tubes and kept at 4\u0026deg;C for hematological studies. Analysis of blood for determining\u0026nbsp;white blood cell count (WBC), red blood cell count (RBC), platelet count (PLT), hemoglobulin (Hb), hematocrit (HCT), lymphocyte and monocyte count was done by\u0026nbsp;semi-automatic blood analyzer (Analytica HEMA 2062+)\u0026nbsp;[20].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSerological study\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe blood was also collected in heparin free tubes and allowed to clot at room temperature for 30 minutes. The blood was centrifuged to obtain serum for determination of serological parameters such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin, urea, uric acid, creatinine, glucose, triglyceride and cholesterol with the help of respective diagnostic kits following direction of use given on kit manual (The ERBA Chem 5 v 3 Germany).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eOxidative stress assessment in hepatic and renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFor determination of lipid peroxidation (LPO), 10% homogenates of hepatic and renal tissues were prepared in KCl solution. The LPO in liver, kidney and microsomes were determined from quantified TBARS [21].\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAntioxidant status assessment in hepatic and renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSucrose solution (1%) was used to prepare homogenates to assess reduced glutathione (GSH)\u0026nbsp;[22].\u0026nbsp;NaCl (0.9%)\u0026nbsp;was used to prepare tissue homogenates to determine superoxide dismutase\u0026nbsp;[23]\u0026nbsp;and catalase activities\u0026nbsp;[24].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eLipid status in hepato-renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo determine triglycerides\u0026nbsp;[25]\u0026nbsp;and cholesterol\u0026nbsp;[26]\u0026nbsp;in tissue, homogenates of liver and kidney were prepared in chilled hypotonic solution.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMicrosomal CYP2E1 activity\u003c/strong\u003e\u003cstrong\u003e, protein and lipid peroxidation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMicrosomes were prepared by CaCl\u003csub\u003e2\u003c/sub\u003e precipitation method [27]. The CYP2E1 activity was determined in terms of aniline hydroxylase activity using aniline as a substrate [28]. Microsomal protein was assessed using Folin and Ciocalteu phenol reagent [29].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHistopathological preparation of hepatic and renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSmall pieces of liver and kidney were immediately fixed in Bouin\u0026rsquo;s fixative. Fixed tissues were further processed and embedded in paraffin wax before cutting 05\u0026nbsp;mm thick sections. Sections were placed on a glass slide and stained with hematoxylin and eosin (H\u0026amp;E) stain\u0026nbsp;[30]. Approximately 10-15 observations for each slide were taken with light microscope.\u003c/p\u003e\n\u003cp\u003eFrom the above-mentioned results, it can be clearly observed that, 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e perform better than the other two lower doses. Thus, only 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e treated groups was further considered for determination of antioxidant enzymes of GSH cycle, proinflammatory cytokines, electron microscopy, genotoxicity and vital parameters of RBC degradation cycle.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEnzymes of GSH cycle in hepato-renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eGlutathione reductase\u0026nbsp;[31], glutathione peroxidase\u0026nbsp;[32], glucose-6-phosphate dehydrogenase\u0026nbsp;[33]\u0026nbsp;and glutathione-S-transferase\u0026nbsp;[34]\u0026nbsp;were determined from tissue homogenate freshly prepared in 1.15% KCl solution.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAssessment of proinflammatory cytokines in serum\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eProinflammatory cytokines including tumor necrosis factor- \u0026alpha; (TNF-\u0026alpha;) and interleukin-6 (IL-6) were assessed in serum using ELISA kits (Ray Biotech, Inc; Norcross, GA 30092 USA) to assess inflammation.\u003cstrong\u003e\u003cbr\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eRBC degradation cycle in hepatic tissue sample\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Hepatic hemoglobin [35], heme content [36], ferritin [37], hemosiderin [38], iron [39], bilirubin [40], biliverdin [41], biliverdin reductase [42] and hemoxygenase-1 (HO-1) enzyme [43], were determined in freshly prepared liver homogenate.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eProcessing of hepatic and renal tissue samples for ultra-structural observation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eLiver and kidney tissue of about 1 mm\u003csup\u003e3\u003c/sup\u003e in size were fixed immediately in Karnovsky\u0026rsquo;s fixative and then in 1% osmium tetroxide [44]. Fixed tissues were eventually embedded in resin and were cut into 70-90 nm thick sections and placed on a copper grid. The grid along with tissue sections were stained with positive staining (Uranyl acetate and lead citrate) prior to the observation with transmission electron microscope [45, 46].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGenotoxicity study in hepato-renal tissue samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSingle cell gel electrophoresis study was performed to observe genotoxicity\u0026nbsp;[47]. Liver tissue homogenate (10%) was taken on a thin film of agarose spread glass slide. Slides were placed in electrophoresis buffer to start electrophoresis. Slides were immediately washed with HCl buffer and stained with ethidium bromide to observe with a microscope. Images were analyzed by ImageJ software.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatistical analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eResults are expressed as mean \u0026plusmn; SE (n=6). Data were analyzed for statistical significance using one-way analysis of variance (P\u0026le;0.05) followed by Tukey\u0026rsquo;s \u003cem\u003epost hoc\u003c/em\u003e honestly significant difference test (\u003cem\u003epost hoc\u003c/em\u003e HSD test) to draw a comparison among different treatment groups (P\u0026le;0.05) [48] using Microsoft Excel worksheet-2019. A p value \u0026le;0.05 was considered as significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eHPLC fingerprinting analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe HPLC fingerprinting analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract and other standards are summarized in Fig 1. \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract showed various phytochemicals as evident by presence of peaks at various retention time. Based on the retention time, compounds were identified and presented in table 1. Results revealed the presence of some medicinally active well-known phytoconstituents as well as some unknown compounds. The first peak between retention time 2 to 4 comprises multiple active components including tannic acid (2.806), L-ascorbic acid (2.964), catechin (2.920), gallic acid (3.269) and caffeic acid (3.639). The peak at 4.020 is rutin (4.035), peak at 4.542 is vanillic acid (4.517), peak at 5.442 is morin (5.447) or malic acid (5.489), peak at 6.260 is quercetin (6.379) and the peak at 7.349 is ellagic acid (7.265). There were some unidentified peaks at retention time 9.581, 12.026, 18.342 and 23.398.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eUV-Vis Spectral analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eScanning analysis of the extract showed two peaks at UV range as well as one in visible range. The peaks at 260, 280 and 660 nm showed absorption of 3.994, 3.721, 0.242 respectively (Fig. 2). The first peak appeared at 260 nm was intense than that of 280 nm. A very light peak appeared at 660 nm suggested presence of colored compound in the extract.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eParticle size analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFig. 3 represents the histograms of the dispersed particles present in \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract as obtained from particle size analyzer. The size of the particles ranges between 0.031 to 0.071 \u0026micro;m. The percentile of particle size of particular diameter was explained in table 2. The most common particle size was 44 nm which comprises of 24.24%. The average particle size of 43 nm was determined from the mean value of the histograms.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTGA and DSC\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe thermogram from TGA analysis of\u0026nbsp;\u003cem\u003eTephrosia purpurea\u003c/em\u003e extract (Fig. 4A) showed thermal decomposition processes, which occurs between 126.58\u0026deg;C and 266.80\u0026deg;C with a midpoint at 241.33\u0026deg;C. The total weight loss was 17.188 mg (62.001%).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe DSC curves of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract (Fig. 4B) showed thermal reaction processes, which occur between 18.64 and 128.49\u0026deg;C. The event of endothermic reaction was observed at 21.46 (D\u0026nbsp;= 15.56 J g\u003csup\u003e-1\u003c/sup\u003e) and 120.88\u0026deg;C (D\u0026nbsp;= 32.24 J g\u003csup\u003e-1\u003c/sup\u003e). Several other endothermic and exothermic peaks were observed between 128.49 and 232.28\u0026deg;C. The major weight loss was observed between 200-500\u0026deg;C.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eX-ray powder diffraction analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe diffractogram obtained from XRPD analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e hydroethanolic extract showed 10 sharp peaks of variable height at various 2\u0026theta; (theta) values (Fig. 5). XRD pattern showed presence of crystals in the extract.\u0026nbsp;The crystallite size was calculated using the XRD data, suggested the presence of nanoparticles in the size range between 25 nm and 47 nm.\u0026nbsp;The average crystallite size of the plant extract was found to be 34.3 nm.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFurrier transform infrared spectroscopy of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe FT-IR spectra of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract (Fig. 6) showed absorbance bands centered at 523, 1044, 1178, 1285, 1403, 1587, 2849, 2917 and 3216 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e. Different band Peak represented different functional group present in the extract (Table 3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on hematological variables\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTable 4 represents alterations in different hematological variables after combined administration of D-GalN and LPS. \u0026nbsp;Total number of RBCs, platelets, amount of hemoglobin in blood and hematocrit value were significantly declined, whereas differential cell counting of WBCs, including lymphocytes and monocytes were significantly raised (P\u0026pound;0.05). Treatment of \u003cem\u003eTephrosia purpurea\u003c/em\u003e showed dose dependent protection in most of the hematological variables. The 50 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e significantly checked monocytes count towards control. The 100 mg/kg dose boosted hemoglobin, RBCs, hematocrit in one hand and on the other hand, controlled WBCs, including monocytes significantly. The 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e significantly protected all the studied variables from alteration. It was observed that the 100 and 200 mg/kg doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e exhibited considerably therapeutic effects in comparison to 50 mg/kg dose.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cbr\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on serological variables and hepatic glycogen\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTable 5 exemplifies protective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e pretreatment against toxic demonstration of D-GalN and LPS on serological variables. Exposure to D-GalN and LPS resulted in substantial increase of serum transaminases (AST and ALT), ALP, urea, uric acid and creatinine (P\u0026pound;0.05). All the three doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e provided protection against toxic outcome of D-GalN and LPS on serological indices. The 100 and 200 mg /kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e offered better protection in comparison to 50 mg/kg dose.\u003c/p\u003e\n\u003cp\u003eAdministration of D-GalN and LPS caused considerable upsurge of serum triglyceride, cholesterol and bilirubin, whereas level of albumin, glucose and hepatic glycogen were dwindled abruptly (Table 6). All the three doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e instigated reduction in triglycerides, cholesterol and bilirubin while increment in albumin, glucose and hepatic glycogen was noted towards their respective control. The 100 mg/kg dose was better than 50 mg/kg in reducing triglycerides and bilirubin level. The 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e appeared as better protective force in all the parameters except triglyceride and cholesterol in serum. It was also revealed that 200 mg/kg dose was better than 100 mg/kg dose in maintaining bilirubin and triglycerides.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on oxidative stress\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe LPO was found to be enhanced drastically in hepatic and renal tissues (Fig. 7) after D-GalN and LPS intoxication (P\u0026pound;0.05). Pretreatment of \u003cem\u003eTephrosia purpurea\u003c/em\u003e at all the three doses repressed hepatorenal LPO towards control (P\u0026pound;0.05). \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 100 and 200 mg/kg dose was more effective than 50 mg/kg dose in decreasing LPO in both tissues. Dose of 200 mg/kg was more efficient than 100 mg/kg in reducing LPO.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on antioxidant status\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe GSH was significantly demoted (Fig. 7) in hepatorenal tissues after D-GalN and LPS exposure (P\u0026pound;0.05). Administration of \u003cem\u003eTephrosia purpurea\u003c/em\u003e increased GSH level in all the tissues at 50 mg/kg dose. Tukey\u0026rsquo;s \u003cem\u003epost hoc\u003c/em\u003e analysis disclosed the fact that \u003cem\u003eTephrosia purpurea\u003c/em\u003e at all doses showed similar recovery pattern in uplifting tissue GSH level. The 100 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e showed no significant disparity with control when hepatic GSH was considered only. The 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e has no such difference with control even the renal GSH was considered too.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eEnzymatic activity of superoxide dismutase and catalase (Fig. 7) in hepatic and renal tissues was severely declined (P\u0026pound;0.05) after exposure to D-GalN and LPS. In \u003cem\u003eTephrosia purpurea\u003c/em\u003e treated groups, activity of superoxide dismutase and catalase was found to be enhanced in dose dependent manner. The 100 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e was better than 50 mg/kg dose in recovering enzymatic activity of hepatic superoxide dismutase and catalase. The 200 mg/kg dose had better protective efficacy in enhancing superoxide dismutase and catalase activities towards their respective control.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on tissue lipid profile\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEffect of D-GalN and LPS administration on triglycerides and cholesterol in hepatic and renal tissues (P\u0026pound;0.05) have been summarized in Fig. 8. A sharp rise in triglycerides and cholesterol in hepatic and renal tissues was observed. The 100 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e significantly reversed renal triglycerides and hepatic cholesterol as compared to 50 mg/kg dose. The 50 and 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e showed significant differences in recovering hepatorenal triglycerides and cholesterol. The 100 and 200 mg/kg dose showed almost identical recovery pattern. \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 200 mg/kg dose treated group reversed the variables very much near to control.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia\u003c/em\u003e \u003cem\u003epurpurea\u003c/em\u003e extract on microsomal LPO, protein and CYP2E1 activity\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAcute hepatic injury induction by D-GalN and LPS resulted amplified microsomal LPO and CYP2E1 activity (P\u0026pound;0.05), whereas microsomal protein was severely diminished (Fig. 9). \u003cem\u003eTephrosia purpurea\u003c/em\u003e treatment resettled microsomal LPO, protein and CYP2E1 activity towards control. \u003cem\u003eTephrosia purpurea\u003c/em\u003e at 100 and 200 mg/kg dose were better when compared to 50 mg/kg dose in maintaining LPO and CYP2E1 activity. \u0026nbsp;\u003cem\u003eTephrosia purpurea\u003c/em\u003e showed dose dependent recovery pattern for hepatic CYP2E1 activity. \u0026nbsp;All the altered values were found very near to control in 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003etreated group.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHistopathological observations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePhotomicrographs of hepatic and renal histological observations suggested structural component disorganization\u0026nbsp;in D-GalN and LPS exposed group.\u0026nbsp;Prophylactic treatment with \u003cem\u003eTephrosia purpura\u003c/em\u003e at three different doses appreciably protected hepato-renal tissues from these alterations. The hepatic and renal histological observations were explained in legends to figures 10 and 11, respectively. Quantitative representation of variables to assess hepato-renal injury as well as improvement has been presented in table 7 and 8 respectively.\u003c/p\u003e\n\u003cp\u003eIt was observed from above-mentioned variables that, 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003eextract showed better efficacy than the other two lower doses. Thus, only 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003eextract treated groups were further considered for the determination of antioxidant enzymes of GSH cycle, electron microscopy, genotoxicity, proinflammatory cytokines (TNF-\u0026alpha; and IL-6) and RBC degradation cycle including HO-1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on enzymes of GSH cycle\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEffects of D-GalN and LPS administration on hepatorenal enzymatic antioxidant status (P\u0026pound;0.05) were summarized in Fig. 12.\u0026nbsp;After 16 hours of D-GalN and LPS treatment animals showed significant decrement in glutathione reductase, glutathione peroxidase, glucose-6-phosphate dehydrogenase and glutathione-S-transferase activities was observed in hepatorenal tissue at 5% level of significance. Oral administration of \u003cem\u003eTephrosia\u0026nbsp;\u003c/em\u003e\u003cem\u003epurpurea\u003c/em\u003e exhibited significant reversal in enzymatic activities of glutathione reductase, glutathione peroxidase, glucose-6-phosphate dehydrogenase and glutathione-S-transferase (\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u003cem\u003e\u0026pound;\u003c/em\u003e\u003cem\u003e\u0026nbsp;0.05\u003c/em\u003e) in liver and kidney. \u003cem\u003eTephrosia\u0026nbsp;\u003c/em\u003e\u003cem\u003epurpurea\u003c/em\u003e showed no substantial difference with control in recovering all these enzymatic antioxidants except for renal glutathione peroxidase.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract on proinflammatory cytokines\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAdministration of D-GalN and LPS significantly induced inflammation via producing proinflammatory cytokines including TNF-\u0026alpha; and IL-6 in serum (Fig. 13). The 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract significantly (P\u0026nbsp;\u0026pound;\u0026nbsp;0.05) protected the elevation of these cytokines.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia\u003c/em\u003e \u003cem\u003epurpurea\u003c/em\u003e extract on RBC degradation cycle\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTable 9 depicted therapeutic effect of \u003cem\u003eTephrosia\u003c/em\u003e on the components of RBC degradation cycle against toxic exposure to LPS and D-GalN. The bilirubin, biliverdin, Hb, heme, free iron (ferritin and hemosiderin) and heme iron along with biliverdin reductase and HO-1 activities in the hepatic tissue were significantly elevated (Table 9) after D-GalN and LPS intoxication (P\u0026nbsp;\u0026pound;\u0026nbsp;0.05). \u003cem\u003eTephrosia\u0026nbsp;\u003c/em\u003e\u003cem\u003epurpurea\u003c/em\u003e at 200 mg/kg dose showed therapeutic effect on all the variables by reducing their level towards control in liver cells except HO-1 activity. The HO-1 activity was further induced up to 3.4 folds by \u003cem\u003eTephrosia\u003c/em\u003e \u003cem\u003epurpurea\u003c/em\u003e at 5% level of significance as compared to control. The therapy of \u003cem\u003eTephrosia\u003c/em\u003e \u003cem\u003epurpurea\u003c/em\u003e at 200 mg/kg dose was able to recover most of the variables altered due to inflammation and oxidative stress induced by LPS and D-GalN.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eUltra-structural observations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eElectron microscopic study of\u0026nbsp;liver and kidney have been presented\u0026nbsp;in figure 14. The figure of\u0026nbsp;control liver depicted regular hepatocytes with obvious nuclear membrane (NM)\u0026nbsp;with visible nuclear pores (NP), dense cytoplasm, numerous well-formed mitochondrion (M) and rough endoplasmic reticulum (RER) around nuclear membrane (Fig. 14A). The D-GalN and LPS administered rat\u0026nbsp;(Fig. 14B)\u0026nbsp;showed loss of nuclear membrane integrity,\u0026nbsp;dilated endoplasmic reticulum and mitochondrial degeneration.\u0026nbsp;Loss of genetic material as indicated by the electron lucent area inside the nucleus and steatosis indicated by deposition of lipid droplets (LD) were also observed\u0026nbsp;(Fig. 14B).\u003c/p\u003e\n\u003cp\u003eProphylactic\u0026nbsp;treatment with \u003cem\u003eTephrosia\u0026nbsp;\u003c/em\u003e\u003cem\u003epurpurea\u003c/em\u003e extract at 200 mg/kg dose (Fig. 14C) showed almost regular shaped nucleus with well-maintained nuclear membrane and nucleolus. Homogenous distribution of euchromatin throughout the nuclear area, regular shaped and more dense mitochondria were also observed along with fine network of ER attached to the nucleus. Decreased steatosis and more dense cytoplasm were noticed as compared to the D-GalN and LPS exposed group.\u003c/p\u003e\n\u003cp\u003eElectron micrograph obtained from transmission electron microscopy\u0026nbsp;revealed that control kidney showed its regular architecture with well-defined membranes, mitochondrial abundance and peroxisomes with circular outline and homogenous content (Fig. 14D). The D-GalN and LPS administered rats showed swelling in mitochondria, loss of cytoplasmic and mitochondrial structural integrity, and lipid droplet accumulation in the cytoplasm (Fig. 14E). Prophylactic treatment with 200 mg/kg dose of \u003cem\u003eTephrosia\u0026nbsp;\u003c/em\u003e\u003cem\u003epurpurea\u003c/em\u003eshowed better mitochondrial structure, reduced lipid droplet accumulation and peroxisome with homogenous content (Fig. 14F).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEffect of \u003cem\u003eTephrosia\u003c/em\u003e \u003cem\u003epurpurea\u003c/em\u003e extract on genetic material\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eExposure to D-GalN and LPS caused severe damage to DNA in hepatorenal tissue. DNA damage was studied in terms of comet assay (Fig. 15) and its numeric data is presented in table 10.\u0026nbsp;Exposure to D-GalN and LPS induced fragmentation of DNA in liver and kidney (Fig. 15, A-F). A significant increase in tail length and high movement translated it into comet and was considered as DNA damage. Comparison between control group (Fig. 15, A and D) and D-GalN + LPS (Fig. 15, B and E) administered group suggested that D-GalN and LPS administration causes significant increase in tail length, tail moment and tail DNA % in hepatorenal tissues (Table 10). Therapy of 200 mg/kg dose of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003econtrolled DNA damage significantly as indicated by decreased tail length and tail moment (Fig. 15, C and F).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eBioactive phytoconstituents are well-known for their protective roles against a variety of health problems including acute organ injury. As \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract have major phytoconstituents including rutin, gallic acid, quercetin and many more, they might be responsible for its protective efficacy in this present study [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Furthermore, presence of single or multiple peaks in the UV-Vis spectra is a clear indication of the presence of unsaturated groups and heteroatoms such as Sulphur, Nitrogen and Oxygen. The peak at positions 660 nm indicated presence of organic chromophores [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe loss of volatile products in the first step of thermal decomposition below 200\u0026deg;C is mainly due to free water and ethanol, as this event occurs in fusion tracks and with vaporization of the substance [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. The major weight loss between 200\u0026deg;C and 500\u0026deg;C may be due to the degradation of a wide variety of secondary metabolites, principally phenolics [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe FT-IR is the most suitable technique of the non-destructive spectroscopic methods in the analysis of pharmaceutical solids [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. Since the extract was hydroethanolic, ethanol and water bands could be present in FTIR spectra. The strong absorption band between 3200\u0026ndash;3400 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e indicated the presence of polymeric hydroxyl derivatives [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. Carboxylic acids have an absorbance range of 2500\u0026ndash;3300 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, thus, bands at 3216, 2917 and 2849 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e could be due to OH stretching of carboxylic acids [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e]. Carboxylic acid serves as a main pharmaceutical product in curing many health problems, including ulcers, jaundice, stomatitis, fever, wounds, edema and rheumatic joint pains [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. The more intense bands occurring at 2917 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and 2849 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e corresponding to C\u0026ndash;H stretching vibrations could also be attributed to the presence of carbonyl functional groups [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e] or methylene group of aliphatic compounds [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e]. The absence of any peak between 2220 and 2260 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e indicated absence of any toxic substances as this region comprises of cyanide groups [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. A band at 1587 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e could be attributed to C\u0026thinsp;=\u0026thinsp;C bonds in the aromatic rings due to presence of flavonoids and amino acids [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e, \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e]. The bands at 1403 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e would be related to C-H bending vibration of methyl group. The band at 1285 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is probably related to C\u0026ndash;O stretching of polyols [\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e]. The band at 1178 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and 1044 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e could be related to C\u0026ndash;O stretching and \u0026ndash;OH deformation of primary and tertiary alcohols respectively [\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e]. This band could also be related to C\u0026ndash;O stretching of ether, esters and carboxylic acids. This indicated the presence of a wide variety of metabolites, such as tannins, flavonoids, carbohydrates (polysaccharides) and anthraquinones among others [\u003cspan additionalcitationids=\"CR58\" citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e]. The band at 523 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e indicated the presence of iodate or brominate compounds [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e], which plays role of disinfectant [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe XRPD analysis of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract exhibits several size dependent features leading to an irregular peak position, height and width. The sharp peak reveals that the extract is crystalline in nature and the particles are in the Nano regime [\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e]. The remaining peaks may be due to other phytochemicals present in \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract [\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eStudies have suggested that D-GalN and LPS induced hepatorenal injury involves many mechanisms, including oxidative stress, inflammation, apoptosis and necrosis [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e]. Compromised cellular integrity due to increased LPO, resulted leakage of cellular constituents especially transaminases (AST and ALT), ALP, bilirubin and albumin into serum impairing liver functions [\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e]. Various blood components such as WBC, RBC, platelets and hemoglobin gets altered due to sepsis, caused by LPS [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e]. Recycling of excess purines due to necrosis increased serum urea, uric acid and creatinine, a clear indication of renal injury [\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e]. Increase in CYP activity could be attributed to p450 functional failure due to increased requirement of D-GalN and LPS biotransformation with a decrease in protein synthesis and accumulation of triglycerides [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Recent studies have shown that antioxidant and anti-inflammatory agents can ameliorate inflammatory responses of LPS induced injury [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e mainly stabilized the membrane [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] to prevent cellular leakage [\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e] probably due to the presence of flavonoids that influenced hepatic regeneration [\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e, \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e]. Multiple beneficial functional groups present in \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract, as evident in FTIR, might play crucial role in regulating hematological variables, liver and kidney functions.\u003c/p\u003e \u003cp\u003eThe D-GalN and LPS trigger excessive reactive oxygen species (ROS), leading to serious oxidative insult in liver injury via lipid peroxidation [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e]. Antioxidant molecules such as glutathione, SOD and catalase neutralized oxidative stress by quenching free radicals [\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e]. The SOD eliminated superoxide anions and produced H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e, which was then decomposed by catalase [\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e protects the depletion of these antioxidant molecules by acting as an antioxidant and provided free electron [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e]. GPx harvested electron from glutathione and neutralized free radical by giving free electron, whereas, GR harvested electron from NADPH and reduced oxidized glutathione to make it available again for GPx. Thereafter, G6PDH obtained electron from glucose 6 phosphate, simultaneously converting NADP\u003csup\u003e+\u003c/sup\u003e into NADPH by giving that electron. Proper regulation of this cycle helped to maintain antioxidant status, but as cellular ROS increased due to D-GalN and LPS, components of this cycle gets depleted in maintaining redox balance [\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e]. Glutathione-S-transferase enabled conjugation of glutathione with toxic intermediates to remove them successfully out of the system. Depletion of glutathione-S-transferase, glutathione and CYP2E1 resulted as exposure to D-GalN and LPS produced a number of toxic intermediates upon metabolism [\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e act as a modifier of oxidant response which responded against oxidant-mediated generation of oxidative stress [\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e]. These enzymes related to GSH cycle were well maintained by \u003cem\u003eTephrosia purpurea\u003c/em\u003e as this plant might have the ability to form conjugates with toxic intermediates and scavenge free radicals [\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e, \u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e73\u003c/span\u003e]. The antihyperlipidemic activity of \u003cem\u003eTephrosia purpurea\u003c/em\u003e is might be due to presence of saponins and flavonoids. \u003cem\u003eTephrosia purpurea\u003c/em\u003e prevented elevation of triglyceride and cholesterol in D-GalN and LPS exposed animal by increasing the lipoprotein lipase and inhibition of HMG Co A reductase activities respectively [\u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e74\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eWhen D-GalN and LPS reaches to the liver, massive pro-inflammatory cytokines including interleukin (IL)-6, IL-1β, and tumor necrosis factor-alpha (TNF-α), were released, leading to serious inflammation damage [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e]. The anti-inflammatory activity of \u003cem\u003eTephrosia purpurea\u003c/em\u003e helps to reduced inflammation associated hepatorenal injury and maintaining cellular integrity [\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e]. High production of ROS caused DNA fragmentation and denaturation, thereby, morphological and structural appearance of the nucleus had changed [\u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e76\u003c/span\u003e]. In the comet assay, the size of the comet's tail signifies DNA damage and the amount of DNA fragments that has migrated from the nucleus determined the tail length [\u003cspan citationid=\"CR77\" class=\"CitationRef\"\u003e77\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e might have quenched free radicals from the nucleus and provided stability to the membrane. As the particles present in the extract are nanosized, they might have interacted with DNA to stabilize it [\u003cspan citationid=\"CR78\" class=\"CitationRef\"\u003e78\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe rapid degradation of RBCs in the liver is associated with enormous oxidative stress and inflammation, resulted in massive production of heme and iron [\u003cspan citationid=\"CR79\" class=\"CitationRef\"\u003e79\u003c/span\u003e]. Unregulated heme contributed towards oxidative injury via ROS generation as the iron from the heme porphyrin ring is rapidly lost and contributes in ferrous state [\u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e80\u003c/span\u003e]. Furthermore, ferritin and hemosiderin were released as an acute phase protein to store free intracellular iron [\u003cspan citationid=\"CR81\" class=\"CitationRef\"\u003e81\u003c/span\u003e, \u003cspan citationid=\"CR82\" class=\"CitationRef\"\u003e82\u003c/span\u003e]. This increased iron deposition in liver which lead to tissue damage and dysregulation of function resulted in increased morbidity and mortality [\u003cspan citationid=\"CR83\" class=\"CitationRef\"\u003e83\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract reduced iron overload in liver might be due to having good metal chelating ability [\u003cspan citationid=\"CR84\" class=\"CitationRef\"\u003e84\u003c/span\u003e]. The HO-1 is not only involved in normal physiology but also has a role in pathophysiological states [\u003cspan citationid=\"CR85\" class=\"CitationRef\"\u003e85\u003c/span\u003e]. LPS administration induced release of HO-1 in liver from 6\u0026ndash;24 hours [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e], but excessive oxidative stress and free heme could also induce HO-1 [\u003cspan citationid=\"CR86\" class=\"CitationRef\"\u003e86\u003c/span\u003e]. Induced HO-1 decreased macrophage infiltration, pro-oxidant and proinflammatory transcription factors to reduce oxidative stress and inflammation in order to protect organs during sepsis. Thus, upregulation of HO-1 is linked with decreased injury during endotoxemia [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. \u003cem\u003eTephrosia purpurea\u003c/em\u003e interfere with RBC degradation and manages to induce HO-1 enzyme up to many folds. HO-1 reduced oxidative stress and inflammation by activating various antioxidant genes to employ cyto-protection [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e]. This induced activity of HO-1 may influence other NADPH and oxygen-consuming pathways to reduce oxidative stress [\u003cspan citationid=\"CR87\" class=\"CitationRef\"\u003e87\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIncreased conversion of heme into biliverdin and finally into unconjugated bilirubin by biliverdin reductase leads to the depletion of biliverdin reductase and increased biliverdin accumulation in hepatocytes [\u003cspan citationid=\"CR88\" class=\"CitationRef\"\u003e88\u003c/span\u003e]. Glucuronidation of bilirubin is an essential process to produce conjugated bilirubin for excretion. But, the depletion of uridine-diphosphoglucuronic glucuronosyl transferase (UDPGT) due to D-GalN administration resulted accumulation of unconjugated bilirubin [\u003cspan citationid=\"CR88\" class=\"CitationRef\"\u003e88\u003c/span\u003e, \u003cspan citationid=\"CR89\" class=\"CitationRef\"\u003e89\u003c/span\u003e]. Administration of \u003cem\u003eTephrosia purpurea\u003c/em\u003e might have smoothen the conversion of bilirubin from biliverdin by acting as an alternative of NADPH and glucuronidation of bilirubin by reducing depletion of UDPGT to minimize the accumulation [\u003cspan citationid=\"CR90\" class=\"CitationRef\"\u003e90\u003c/span\u003e, \u003cspan citationid=\"CR91\" class=\"CitationRef\"\u003e91\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eSimilar observation was also reported by Sahu et al., [\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e]. The phytocompounds present in the hydroalcoholic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e might be responsible for protecting tissue from oxidative stress and inflammation induced injury by neutralizing those free radicals while enhancing the body\u0026rsquo;s defense systems against degenerative diseases [\u003cspan citationid=\"CR92\" class=\"CitationRef\"\u003e92\u003c/span\u003e, \u003cspan citationid=\"CR93\" class=\"CitationRef\"\u003e93\u003c/span\u003e].\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe pharmaceutical industry disposes of valuable tools used for quality control of products of plant origin and raw materials which require attentive characterization. Characterization of \u003cem\u003eTephrosia purpurea\u003c/em\u003e provides important information about its stability, composition, nature and others. This information is valuable to insure its quality and, therefore, its safety and efficacy. The exposure to D-GalN and LPS resulted increase in the level of oxidative stress and inflammation, which leads to acute organ injury. Treatment with bioactive component rich extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e demonstrated inhibition of D-GalN and LPS induced inflammatory response and oxidative stress. Preventive effect was supported by regular serological and hematological values, improved histoarchitecture, ultrastructure, comet assay photographs, induced HO-1, improved enzymatic antioxidant status and controlled lipid peroxidation in vital organs. These findings may provide a new insight to understand the molecular mechanism behind the anti-inflammatory and anti-oxidative action of \u003cem\u003eTephrosia purpurea\u003c/em\u003e, and may provide a better understanding of its use as a traditional medicine to treat various inflammatory and oxidative stress disorders.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eALF = Acute liver failure; ALI = Acute liver injury; ALP = Alkaline phosphatase; ALT = Alanine amino transferase; ANOVA = Analysis of variance; AST = Aspartate amino transferase; CaCl\u003csub\u003e2\u003c/sub\u003e = Calcium chloride; CAT = Catalase; CYP2E1 = Cytochrome p 450 2E1; D-GalN = D-galactosamine; DSC = Differential scanning colorimetry; FTIR = Fourier transform infrared spectroscopy; G6PDH = Glucose 6 phosphate dehydrogenase; GPx = Glutathione peroxidase; GR = Glutathione reductase; GSH = Reduced glutathione; G-S-T = Glutathione S transferase; H\u0026amp;E = Hematoxylin and eosin; Hb = Hemoglobulin; HCT = Hematocrit; HO-1 = Hemoxygenase-1; HSD = Honestly significant difference; IL-6 = Interleukin 6; KCl = Potassium chloride; LPO = Lipid peroxidation; LPS = Lipopolysaccharide; NaCl = Sodium chloride; PLT = Platelets; PSA = Particle size analysis; RBC = Red blood cell; ROS = Reactive oxygen species; RP-HPLC = Reverse phase high performance liquid chromatography; SE = Standard error; SOD = Superoxide dismutase; TBARS = Thio-barbituric acid reactive substances; TGA = Thermogravimetric analysis; TNF-\u0026alpha; = Tumor necrosis factor alpha; TP = \u003cem\u003eTephrosia purpurea;\u0026nbsp;\u003c/em\u003eUV-Vis = Ultra violet-visible spectroscopy; WBC = White blood cell; XRPD = X-ray powder diffraction.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe first author greatly acknowledged electron microscopy division, AIIMS, New Delhi, India for providing facilities to carry out ultrastructural study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding:\u0026nbsp;\u003c/strong\u003eThe first author greatly acknowledged Department of Zoology, Guru Ghasidas Vishwavidyalaya for providing Laboratory facilities and fellowship. Authors are also grateful to UGC, New Delhi [Startup Grant {20-1/2012(BSR)/20-12(3/2012) (BSR)] for partial financial assistance.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interests:\u0026nbsp;\u003c/strong\u003eThere is no conflict of interest between authors for this manuscript. The manuscript is not communicated in any other journal.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContribution of authors: Samrat Rakshit:\u003c/strong\u003e Conceptualization, Investigation, Formal analysis, Writing- Original draft preparation. \u003cstrong\u003eNisha Sahu:\u003c/strong\u003e Investigation, Formal analysis. \u003cstrong\u003eSatendra Kumar Nirala:\u003c/strong\u003e Conceptualization, Writing- Review and editing, Supervision, Validation. \u003cstrong\u003eShamli S Gupte:\u003c/strong\u003e Formal analysis. \u003cstrong\u003eSangeeta Shukla:\u003c/strong\u003e Supervision, Resources. \u003cstrong\u003eSadhana Shrivastava:\u003c/strong\u003e Supervision, Validation. \u003cstrong\u003eBhawana Bhadauria:\u003c/strong\u003e Validation, Resources. \u003cstrong\u003eMonika Bhadauria:\u003c/strong\u003e Conceptualization, Writing- Review and editing, Supervision, Validation. All data were generated in-house, and no paper mill was used. All authors agree to be accountable for all aspects of work ensuring integrity and accuracy.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate:\u0026nbsp;\u003c/strong\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication:\u0026nbsp;\u003c/strong\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and material:\u0026nbsp;\u003c/strong\u003eNot applicable.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eLiao CC, Day YJ, Lee HC, Liou JT, Chou AH, Liu FC. ERK Signaling Pathway Plays a Key Role in Baicalin Protection Against Acetaminophen-Induced Liver Injury. 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Int J Pharm Pharma Sci. 2010;2(3):30\u0026ndash;3.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003e\u003cstrong\u003eTable 1: Multiple bioactive constituents are present in the hydroethanolic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003ePeak #\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003eRetention time\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003ePhytoconstituents\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eArea\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eHight\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eArea %\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eHight %\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e2-4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eTannic acid, L-ascorbic acid, catechin, gallic acid and caffeic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eNot measured\u003cspan dir=\"RTL\"\u003e٭\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eNot measured\u003cspan dir=\"RTL\"\u003e٭\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eNot measured\u003cspan dir=\"RTL\"\u003e٭\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003eNot measured\u003cspan dir=\"RTL\"\u003e٭\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e4.020\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eRutin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e50067\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e2964\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e1.048\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e2.765\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e4.542\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eVanillic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e181386\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e11854\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e3.796\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e11.058\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e5.442\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eMorin and malic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e14559\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e903\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e0.305\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e0.842\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e6.260\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eQuercetin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e508192\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e11930\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e10.635\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e11.129\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e7.349\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eEllagic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e55092\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e7091\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e1.153\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e6.615\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e9.581\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eUnknown\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e809107\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e23292\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e16.933\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e21.728\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e12.026\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eUnknown\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e342990\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e5229\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e7.178\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e4.878\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e18.342\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eUnknown\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e1864374\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e31651\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e39.017\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e29.525\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"9.936575052854122%\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"17.653276955602536%\"\u003e\n \u003cp\u003e23398\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.978858350951374%\"\u003e\n \u003cp\u003eUnknown\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e952551\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e12287\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e19.935\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.10782241014799%\"\u003e\n \u003cp\u003e11.462\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cspan dir=\"RTL\"\u003e٭\u003c/span\u003eThere can be many phytoconstituents present between retention time 2 to 4, so, we did not measured the area and height of the peak.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2: Particle size distribution statistics analysis of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003eextract\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.430594900849858%\"\u003e\n \u003cp\u003eCum Q3 (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.48158640226629%\"\u003e\n \u003cp\u003e100.000\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e99.834\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e94.601\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e76.789\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e54.075\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e29.834\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e20.360\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.34844192634561%\"\u003e\n \u003cp\u003e0.039\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.430594900849858%\"\u003e\n \u003cp\u003eDiff q3(%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.48158640226629%\"\u003e\n \u003cp\u003e0.166\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e5.232\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e17.813\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e22.713\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e24.242\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e9.474\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e7.525\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.34844192634561%\"\u003e\n \u003cp\u003e0.035\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.430594900849858%\"\u003e\n \u003cp\u003eParticle diameter (\u0026mu;m)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.48158640226629%\"\u003e\n \u003cp\u003e0.071\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e0.063\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e0.056\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e0.050\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e0.044\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76487252124646%\"\u003e\n \u003cp\u003e1.949\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.339943342776204%\"\u003e\n \u003cp\u003e1.949\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.34844192634561%\"\u003e\n \u003cp\u003e0.031\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.407355021216407%\"\u003e\n \u003cp\u003eMedian Diameter (\u0026mu;m)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"8\" valign=\"top\" width=\"83.5926449787836%\"\u003e\n \u003cp\u003e0.043\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.407355021216407%\"\u003e\n \u003cp\u003eModal Diameter (\u0026mu;m)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"8\" valign=\"top\" width=\"83.5926449787836%\"\u003e\n \u003cp\u003e0.039\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.407355021216407%\"\u003e\n \u003cp\u003eMean Value (\u0026mu;m)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"8\" valign=\"top\" width=\"83.5926449787836%\"\u003e\n \u003cp\u003e0.043\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.407355021216407%\"\u003e\n \u003cp\u003eStandard Deviation\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"8\" valign=\"top\" width=\"83.5926449787836%\"\u003e\n \u003cp\u003e0.079\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"16.407355021216407%\"\u003e\n \u003cp\u003eRefractive Index\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"8\" valign=\"top\" width=\"83.5926449787836%\"\u003e\n \u003cp\u003e1.70-0.20i\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eMedian: The size of particle where the cumulative line crosses the 50th percentile; Modal: The size of particle that is most often seen; Mean Value: The average size of the particles; Standard Deviation: The amount of variance, in microns, between each trial; Cum Q3 (%) = Cumulative %: The percent of particles below that micron size; Diff q3 (%) = Differential Distribution: The percent of particles at that micron size.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3: Structural analysis of \u003cem\u003eTephrosia purpurea\u0026nbsp;\u003c/em\u003eextract by FTIR spectrum\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e\u003cstrong\u003eWave numbers (cm\u003csup\u003e-1\u003c/sup\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003e\u003cstrong\u003eVibration\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003e\u003cstrong\u003eAssignment\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e3216 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eOH stretching\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003ePolymeric hydroxyl derivatives and carboxylic acids\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e2849 and 2917 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eOH stretching\u003c/p\u003e\n \u003cp\u003eC-H stretching\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003eCarboxylic acid\u003c/p\u003e\n \u003cp\u003eCarbonyl functional groups\u003c/p\u003e\n \u003cp\u003eMethylene group of aliphatic compounds\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e1587 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eC=C stretching\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003eAromatic rings of flavonoids and amino acids\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e1403 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eC\u0026ndash;H bending\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003eMethyl group\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e1285 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eC\u0026ndash;O stretching\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003epolyols\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e1178 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eC\u0026ndash;O stretching\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003ePrimary alcohol, ether, ester and carboxylic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e1044 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003e\u0026ndash;OH deformation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003eTertiary alcohol\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"34.22913719943423%\"\u003e\n \u003cp\u003e523 cm\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.66053748231966%\"\u003e\n \u003cp\u003eC\u0026ndash;I/Br stretching\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"46.11032531824611%\"\u003e\n \u003cp\u003eIodide or Bromide\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eTable 4: \u0026nbsp;Protective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on hematological variables against D-GalN and LPS\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" width=\"23.401950162513543%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameters\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"76.59804983748646%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"13.012729844413013%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.256011315417258%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTP \u003cem\u003eper se\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTP 200 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.52899575671853%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.256011315417258%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 50 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.256011315417258%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 100 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.690240452616692%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eHemoglobin (g/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e15.4\u0026plusmn;0.71\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e15.5\u0026plusmn;0.71\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e11.4\u0026plusmn;0.52\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e12.6\u0026plusmn;0.58\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e14.1\u0026plusmn;0.65\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e14.9\u0026plusmn;0.69\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eHematocrit (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e44.2\u0026plusmn;2.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e45.1\u0026plusmn;2.07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e31.5\u0026plusmn;1.45\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e39.1\u0026plusmn;1.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e41.1\u0026plusmn;1.89\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e43.2\u0026plusmn;1.98\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eRBC (10\u003csup\u003e6\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e7.66\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e7.58\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e5.75\u0026plusmn;0.26\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e6.75\u0026plusmn;0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e7.18\u0026plusmn;0.33\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e7.42\u0026plusmn;0.34\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eWBC (10\u003csup\u003e3\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e11.9\u0026plusmn;0.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e11.8\u0026plusmn;0.54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e16.8\u0026plusmn;0.77\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e14.7\u0026plusmn;0.68\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e13.2\u0026plusmn;0.61\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e12.8\u0026plusmn;0.59\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003ePlatelets (10\u003csup\u003e3\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e710\u0026plusmn;32.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e725\u0026plusmn;33.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e452\u0026plusmn;20.7\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e521\u0026plusmn;23.9\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e594\u0026plusmn;27.3\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e651\u0026plusmn;29.8\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eLymphocyte Count (10\u003csup\u003e2\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e28.1\u0026plusmn;1.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e29.3\u0026plusmn;1.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e63.7\u0026plusmn;2.92\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e61.3\u0026plusmn;2.81\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e56.4\u0026plusmn;2.59\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e43.1\u0026plusmn;1.98\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"23.401950162513543%\"\u003e\n \u003cp\u003eMonocyte Count (10\u003csup\u003e2\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.967497291440953%\"\u003e\n \u003cp\u003e6.80\u0026plusmn;0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e6.70\u0026plusmn;0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.192849404117009%\"\u003e\n \u003cp\u003e16.2\u0026plusmn;0.74\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e9.7\u0026plusmn;0.45\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.217768147345613%\"\u003e\n \u003cp\u003e9.20\u0026plusmn;0.42\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.784398699891657%\"\u003e\n \u003cp\u003e8.70\u0026plusmn;0.40\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eData are expressed as mean \u0026plusmn; SE, (n=6). \u003cstrong\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus Control. \u003cstrong\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus D-GalN+LPS. \u003cstrong\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 50 mg/kg. \u003csup\u003eF\u003c/sup\u003eP\u0026le;0.05 versus TP 100 mg/kg.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 5: \u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eProtective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on hepatic and renal functions against D-GalN and LPS\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" width=\"18.574514038876888%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameters\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"81.42548596112312%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"15.0066401062417%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.529880478087648%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTP \u003cem\u003eper se\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTP 200 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.334661354581673%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.334661354581673%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 50 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.529880478087648%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 100 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.264276228419654%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eAST (IU/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e81.3\u0026plusmn;3.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e74.1\u0026plusmn;3.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e471\u0026plusmn;21.6\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e295\u0026plusmn;13.5\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e201\u0026plusmn;9.23\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e108\u0026plusmn;4.96\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eALT (IU/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e36.3\u0026plusmn;1.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e35.4\u0026plusmn;1.62\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e172\u0026plusmn;7.92\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e129\u0026plusmn;5.93\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e90.4\u0026plusmn;4.15\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e51.6\u0026plusmn;2.37\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eALP (IU/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e193\u0026plusmn;8.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e184\u0026plusmn;8.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e389\u0026plusmn;17.8\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e297\u0026plusmn;13.7\u003csup\u003e*\u003c/sup\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e248\u0026plusmn;11.4\u003csup\u003e*\u003c/sup\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e212\u0026plusmn;9.75\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eUrea (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e29.1\u0026plusmn;1.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e28.8\u0026plusmn;1.32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e70.6\u0026plusmn;3.24\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e58.9\u0026plusmn;2.70\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e44.1\u0026plusmn;2.02\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e36.0\u0026plusmn;1.65\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eUric Acid (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e1.41\u0026plusmn;0.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e1.31\u0026plusmn;0.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e5.01\u0026plusmn;0.23\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e3.97\u0026plusmn;0.18\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e2.87\u0026plusmn;0.13\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e2.0\u0026plusmn;0.09\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"18.594594594594593%\"\u003e\n \u003cp\u003eCreatinine (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.216216216216216%\"\u003e\n \u003cp\u003e0.70\u0026plusmn;0.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e0.65\u0026plusmn;0.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e2.03\u0026plusmn;0.09\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.297297297297296%\"\u003e\n \u003cp\u003e1.91\u0026plusmn;0.09\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.27027027027027%\"\u003e\n \u003cp\u003e1.42\u0026plusmn;0.07\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.054054054054054%\"\u003e\n \u003cp\u003e1.03\u0026plusmn;0.05\u003csup\u003e*\u003c/sup\u003e\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eData are expressed as mean \u0026plusmn; SE, (n=6). \u003cstrong\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus Control. \u003cstrong\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus D-GalN+LPS. \u003cstrong\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 50 mg/kg. \u003cstrong\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 100 mg/kg.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eTable 6: Protective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on serum biochemical variables and hepatic glycogen against D-GalN and LPS\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" width=\"19.934994582881906%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameters\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"80.06500541711809%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"13.008130081300813%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.344173441734416%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTP \u003cem\u003eper se\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTP 200 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.311653116531165%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.88617886178862%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 50 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.88617886178862%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 100 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.56368563685637%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eGlucose (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e121\u0026plusmn;5.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e114\u0026plusmn;5.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e71.3\u0026plusmn;3.72\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e89.5\u0026plusmn;4.11\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e102\u0026plusmn;4.68\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e110\u0026plusmn;5.06\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eTriglyceride (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e32.3\u0026plusmn;1.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e30.1\u0026plusmn;1.38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e120\u0026plusmn;5.49\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e86.7\u0026plusmn;3.98\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e65.8\u0026plusmn;3.02\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e51.5\u0026plusmn;2.36\u003csup\u003e*\u003c/sup\u003e\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eCholesterol (mg/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e21.3\u0026plusmn;0.97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e20.5\u0026plusmn;0.94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e49.7\u0026plusmn;2.28\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e37.2\u0026plusmn;1.71\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e35.1\u0026plusmn;1.61\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e29.7\u0026plusmn;1.36\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eTotal Bilirubin (mg/dl) \u0026nbsp; \u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e0.39\u0026plusmn;0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e0.37\u0026plusmn;0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e0.89\u0026plusmn;0.04\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e0.71\u0026plusmn;0.03\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e0.59\u0026plusmn;0.03\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e0.49\u0026plusmn;0.02\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eDirect Bilirubin (mg/dl) \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e0.1\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e0.11\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e0.70\u0026plusmn;0.03\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e0.54\u0026plusmn;0.02\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e0.39\u0026plusmn;0.01\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e0.19\u0026plusmn;0.01\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eAlbumin (g/dl)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e4.61\u0026plusmn;0.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e4.74\u0026plusmn;0.22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e2.41\u0026plusmn;0.11\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e2.89\u0026plusmn;0.13\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e3.51\u0026plusmn;0.16\u003csup\u003e*\u003c/sup\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e4.01\u0026plusmn;0.18\u003csup\u003eY\u003c/sup\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"19.95661605206074%\"\u003e\n \u003cp\u003eGlycogen (mg/100g) \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.412147505422993%\"\u003e\n \u003cp\u003e3200\u0026plusmn;146\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.882863340563992%\"\u003e\n \u003cp\u003e3185\u0026plusmn;146\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.25596529284165%\"\u003e\n \u003cp\u003e1345\u0026plusmn;61.5\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e2339\u0026plusmn;107\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.316702819956616%\"\u003e\n \u003cp\u003e2555\u0026plusmn;117\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.859002169197398%\"\u003e\n \u003cp\u003e2736\u0026plusmn;125\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eData are expressed as mean \u0026plusmn; SE, (n=6). \u003cstrong\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus Control. \u003cstrong\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus D-GalN+LPS. \u003cstrong\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 50 mg/kg. \u003cstrong\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 100 mg/kg.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 7: Severity of liver injuries in different treatments.\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e[─] no alterations; [+] mild alterations; [++] moderate alterations; [+++] severe alterations.\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" width=\"22.282023681377826%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameters\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"77.71797631862218%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"15.905947441217151%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.767634854771785%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTP \u003cem\u003eper se\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTP 200 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.767634854771785%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.597510373443985%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 50 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.25726141078838%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 100 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.704011065006917%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003eNecrosis\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003eVacuolation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003eDistorted hepatic cords\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003eInflammatory cell infiltration\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003ePyknotic nuclei\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"22.258064516129032%\"\u003e\n \u003cp\u003eDamaged central vain\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.365591397849462%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.258064516129032%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.903225806451612%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.193548387096774%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.763440860215054%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003cstrong\u003eTable 8:\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eSeverity of kidney injuries in different treatments.\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e[─] no alterations; [+] mild alterations; [++] moderate alterations; [+++] severe alterations.\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" width=\"25.643776824034333%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameters\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"74.35622317596567%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.987012987012987%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.594516594516595%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTP \u003cem\u003eper se\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTP 200 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.584415584415584%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.181818181818183%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 50 mg/kg\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.181818181818183%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 100 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.47041847041847%\"\u003e\n \u003cp\u003e\u003cstrong\u003eD-GalN+LPS+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eTubular dilatation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eTubular degeneration\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eTubular vacuolization\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eGlomerular damage\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eLoss of glomerular space\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e+++\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"25.643776824034333%\"\u003e\n \u003cp\u003eDistortion of endothelial lining \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.656652360515022%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.339055793991417%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.587982832618026%\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.51931330472103%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.733905579399142%\"\u003e\n \u003cp\u003e─\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eTable 9: Protective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on RBC degradation cycle in liver\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"9.66702470461869%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.378088077336198%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHb (mg/100g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHeme (\u0026micro;g/g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.16326530612245%\"\u003e\n \u003cp\u003e\u003cstrong\u003eFerritin (\u0026micro;g Fe/g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.063372717508056%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHemosiderin (\u0026micro;g Fe/g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e\u003cstrong\u003eFree iron (\u0026micro;g Fe/g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTotal Iron (\u0026micro;g Fe/g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.022556390977444%\"\u003e\n \u003cp\u003e\u003cstrong\u003eBilirubin (\u0026micro;g/100 g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e\u003cstrong\u003eBiliverdin (\u0026micro;g/100 g liver)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e\u003cstrong\u003eBiliverdin reductase\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(p mol bilirubin\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e/min/mg protein)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHO-1\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(p mol bilirubin\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e/h/mg protein)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"9.66702470461869%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.378088077336198%\"\u003e\n \u003cp\u003e7.39\u0026plusmn;0.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e8.48\u0026plusmn;0.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.16326530612245%\"\u003e\n \u003cp\u003e39\u0026plusmn;2.53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.063372717508056%\"\u003e\n \u003cp\u003e20\u0026plusmn;1.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e140\u0026plusmn;9.08\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e199\u0026plusmn;12.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.022556390977444%\"\u003e\n \u003cp\u003e477\u0026plusmn;30.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e182\u0026plusmn;11.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e3.52\u0026plusmn;0.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e21.1\u0026plusmn;1.37\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"9.66702470461869%\"\u003e\n \u003cp\u003e\u003cstrong\u003eLPS+D-GalN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.378088077336198%\"\u003e\n \u003cp\u003e12.8\u0026plusmn;0.83\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e50.7\u0026plusmn;3.29\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.16326530612245%\"\u003e\n \u003cp\u003e162\u0026plusmn;10.5\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.063372717508056%\"\u003e\n \u003cp\u003e83\u0026plusmn;5.39\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e545\u0026plusmn;35.4\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e790\u0026plusmn;51.3\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.022556390977444%\"\u003e\n \u003cp\u003e865\u0026plusmn;56.1\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e394\u0026plusmn;25.6\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e7.88\u0026plusmn;0.51\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e47.3\u0026plusmn;3.07\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"9.66702470461869%\"\u003e\n \u003cp\u003e\u003cstrong\u003eLPS+D-GalN+ TP 200 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.378088077336198%\"\u003e\n \u003cp\u003e8.4\u0026plusmn;0.55\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e21.3\u0026plusmn;1.38\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.16326530612245%\"\u003e\n \u003cp\u003e52\u0026plusmn;3.38\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.063372717508056%\"\u003e\n \u003cp\u003e33\u0026plusmn;2.14\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e175\u0026plusmn;11.4\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.270676691729323%\"\u003e\n \u003cp\u003e260\u0026plusmn;16.9\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.022556390977444%\"\u003e\n \u003cp\u003e546\u0026plusmn;35.4\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e217\u0026plusmn;14.1\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.237379162191193%\"\u003e\n \u003cp\u003e3.92\u0026plusmn;0.25\u003csup\u003eY\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.344790547798066%\"\u003e\n \u003cp\u003e91.8\u0026plusmn;5.96\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eData are expressed as mean \u0026plusmn; SE, (n=6). \u003cstrong\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus Control. \u003cstrong\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus D-GalN+LPS. \u003cstrong\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 50 mg/kg. \u003cstrong\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 100 mg/kg.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 10: Protective effect of \u003cem\u003eTephrosia purpurea\u003c/em\u003e on hepato-renal comet assay \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" width=\"12.875536480686696%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"6\" width=\"87.12446351931331%\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameter\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"3\" width=\"49.26108374384236%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHepatic\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" width=\"50.73891625615764%\"\u003e\n \u003cp\u003e\u003cstrong\u003eRenal\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"15.737704918032787%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail length\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.19672131147541%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail movement\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.245901639344263%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail DNA (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.901639344262295%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail length\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.0327868852459%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail movement\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.885245901639344%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTail DNA (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.857142857142858%\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.714285714285714%\"\u003e\n \u003cp\u003e0.1\u0026plusmn;0.005\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.857142857142858%\"\u003e\n \u003cp\u003e0.002\u0026plusmn;0.0001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.285714285714286%\"\u003e\n \u003cp\u003e10.0\u0026plusmn;0.50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.857142857142858%\"\u003e\n \u003cp\u003e0.29\u0026plusmn;0.015\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.714285714285714%\"\u003e\n \u003cp\u003e0.001\u0026plusmn;0.0001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.714285714285714%\"\u003e\n \u003cp\u003e7.00\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.857142857142858%\"\u003e\n \u003cp\u003e\u003cstrong\u003eLPS+D-GalN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.714285714285714%\"\u003e\n \u003cp\u003e22.0\u0026plusmn;1.11\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.857142857142858%\"\u003e\n \u003cp\u003e21.4\u0026plusmn;1.08\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.285714285714286%\"\u003e\n \u003cp\u003e97\u0026plusmn;4.88\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.857142857142858%\"\u003e\n \u003cp\u003e24.0\u0026plusmn;1.21\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.714285714285714%\"\u003e\n \u003cp\u003e23.9\u0026plusmn;1.20\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.714285714285714%\"\u003e\n \u003cp\u003e82.0\u0026plusmn;4.12\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.857142857142858%\"\u003e\n \u003cp\u003e\u003cstrong\u003eLPS+D-GalN+ TP 20 mg/kg \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.714285714285714%\"\u003e\n \u003cp\u003e16.0\u0026plusmn;0.81\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.857142857142858%\"\u003e\n \u003cp\u003e13.6\u0026plusmn;0.68\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.285714285714286%\"\u003e\n \u003cp\u003e41\u0026plusmn;2.06\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.857142857142858%\"\u003e\n \u003cp\u003e14.0\u0026plusmn;0.70\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.714285714285714%\"\u003e\n \u003cp\u003e13.2\u0026plusmn;0.66\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.714285714285714%\"\u003e\n \u003cp\u003e47.0\u0026plusmn;2.36\u003csup\u003eY\u003c/sup\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eData are expressed as mean \u0026plusmn; SE, (n=6). \u003cstrong\u003e\u003csup\u003e*\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus Control. \u003cstrong\u003e\u003csup\u003eY\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus D-GalN+LPS. \u003cstrong\u003e\u003csup\u003ej\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 50 mg/kg. \u003cstrong\u003e\u003csup\u003eF\u003c/sup\u003e\u003c/strong\u003eP\u0026le;0.05 versus TP 100 mg/kg. \u003cstrong\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Acute hepatitis, D-Galactosamine, Hemoxygenase-1, HPLC, Inflammation, Lipopolysaccharide, Natural antioxidants, Oxidative stress, Reactive oxygen species, Tephrosia purpurea","lastPublishedDoi":"10.21203/rs.3.rs-1596610/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1596610/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground:\u003c/strong\u003e Acute liver failure (ALF), often associated with elevated oxidative stress and sustained inflammation, is one of the major clinical manifestations of liver disease with limited therapeutic options. \u003cem\u003eTephrosia purpurea\u003c/em\u003e is mentioned in \u003cem\u003eAyurvedic\u003c/em\u003e literature to be effective against various health problems. Additionally, this plant with various unknown bioactive components has not been fully explored against acute organ injury, especially against D-galactosamine and lipopolysaccharide induced acute hepatitis. Thus, we aimed to characterize and investigate the efficacy of hydroethanolic extract of \u003cem\u003eTephrosia purpurea\u003c/em\u003e to attenuate ALF progression. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e Different doses of \u003cem\u003eTephrosia purpurea\u003c/em\u003e extract (50, 100 and 200 mg/kg) were administered orally once a day for 6 days. \u003cem\u003eIn vivo\u003c/em\u003e injury was induced via intraperitoneal injection of D-galactosamine (300 mg/kg) followed by lipopolysaccharide (50 μg/kg). Presence of bioactive compounds, functional groups, nature and size of particles, thermal properties of the extract were determined. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e Levels of lipid peroxidation, proinflammatory cytokines were significantly elevated after D-galactosamine and lipopolysaccharide administration that were restored by the administration of \u003cem\u003eTephrosia purpurea\u003c/em\u003e via modulating hemoxygenase-1 enzyme. \u003cem\u003eTephrosia purpurea\u003c/em\u003e alleviated serum transaminase levels, hematological variables and markedly reduced histological and ultrastructural alterations. \u003cem\u003eTephrosia purpurea\u003c/em\u003e reduced iron overload and maintained glutathione homeostasis (P£ 0.05). Not only that, it also eliminated lipid droplets, restored biomacromolecule homeostasis, maintained stability of the membrane, cellular organelles and the DNA.\u003cstrong\u003e \u003c/strong\u003e\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eConclusions: \u003c/strong\u003eThese findings provided novel insights into the anti-inflammatory and antioxidative mechanisms of \u003cem\u003eTephrosia purpurea\u003c/em\u003e and suggested its use as a potential therapeutic candidate against ALF.\u003c/p\u003e","manuscriptTitle":"Tephrosia purpurea (Linn.) attenuates progression of acute liver failure by reducing oxidative stress and inflammation via modulating cytoprotective enzyme hemoxygenase-1 activity","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-05-17 15:19:56","doi":"10.21203/rs.3.rs-1596610/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"87aa5155-3f66-4b5f-8739-a9b85ba84e8a","owner":[],"postedDate":"May 17th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2022-05-21T12:41:38+00:00","versionOfRecord":[],"versionCreatedAt":"2022-05-17 15:19:56","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1596610","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1596610","identity":"rs-1596610","version":["v1"]},"buildId":"J0_U0BvcaRcwD8yVFaRlm","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}