Innovative strategy for extraction of green cardamom via Super critical fluid extractor at different levels of pressure with its application against microorganisms in vitro and in silico

Innovative strategy for extraction of green cardamom via Super critical fluid extractor at different levels of pressure with its application against microorganisms in vitro and in silico | 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 Innovative strategy for extraction of green cardamom via Super critical fluid extractor at different levels of pressure with its application against microorganisms in vitro and in silico Sulaiman A. Alsalamah, Mohammed Ibrahim Alghonaim, Khatib Sayeed Ismail, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6480943/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 08 Dec, 2025 Read the published version in Bioresources and Bioprocessing → Version 1 posted 4 You are reading this latest preprint version Abstract There is broad approach in the search for resource-use strategies that are both economically viable and value-based is the extraction of active compounds from various plants. Supercritical fluid extraction (SFE), has become a popular technique for extraction significant plant-based compounds. Our investigation contrasted the yield, biological functions and phytochemical compositions of green cardamom extracts generated with SFE at 100, 200, and 300 bar of pressures. The maximal obtained weight was 0.279 gm upon applying 300 bar. There is a proportional elevation in the levels of most of phenolic compounds which detected using HPLC upon raising the pressure levels for extraction. Antimicrobial action of extract types towards test food-born microbes where the produced extract at 300 bar had the maximal antimicrobial action towards Bacillus subtilis , and Candida albicans with inhibition zones 23.33 ± 0.58, and 22.17 ± 1.04 mm, subsequently. Antibiofilm and hemolysis inhibition in presence of test microbes of various extract forms reveal the potential impact of raising pressure for extraction to enhance impact of extract to reach maximal level at 300 bar. Killing kinetics assay showed the gradual raise of the green cardamom extract versus test microbes at various time points where the extract at 300 bar had the best outcomes. Transmission electron microscopy visualized alterations in ultrastructure features of B. subtilis and C. albicans using the green cardamom extract produced at 300 bar. The molecular docking performance of the main constituents in green cardamom extracts gallic acid and syringic acid against B. subtilis (PDB ID: 5VX6) and S. aureus (PDB ID: 3V8J) using the Molecular Operating Environment (MOE) software was evaluated. The docking scores (S), RMSD_refine values, and energy terms (E_conf, E_place, E_score1, E_refine, E_score2) were analyzed to assess binding affinities. Key interactions, including hydrogen bonds, were identified, with distances and energies quantified. Syringic acid exhibited better binding (S = -4.27 to -5.04) compared to gallic acid (S = -4.11 to -4.68) across both targets. Interactions with residues like GLU 187 and ARG 172 in 5VX6, and ASP 239 in 3V8J, highlighted critical binding motifs. These results suggest both compounds as potential inhibitors for bacterial targets, warranting further experimental validation. Super critical fluid extractor cardamom antimicrobial antihemolytic Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Introduction The academic sector has long been interested in aromatic and therapeutic plants with antimicrobial capabilities because they provide numerous biologically active substances that can be applied to counteract the spreading of microorganisms that cause serious and potentially deadly infections [ 1 , 2 ]. In Mediterranean areas, spices and herbaceous plants are commonly used in preparing food since they have the ability to fight towards several pathological bacteria [ 3 ]. The frequent consumption of antimicrobial agents among consumers to combat pathogens explains the rise in resistance produced in most populations towards microbial infections which urge the need to use innovative tools to overcome these infections [ 4 , 5 ]. The green cardamom species ( Elettaria cardamomum) is a big evergreen herb rhizomatous monocot species that is a member of the Zingiberaceae and Elettaria families [ 6 ]. It is widely recognized for the flavorful components and bioactive substances found in dry mature fruit, that make them valuable commodities for the food, medicinal products and cosmetic sectors [ 7 ]. In the food industry, cardamom is frequently used to flavor prepared meals, soft drinks, and confections. Furthermore, it is employed in the cosmetics industry for body and hair care items, aromatherapy, and scents [ 8 ]. In the drug sector, cardamom has also been utilized as a potent antioxidant, antimicrobial agent [ 9 ]. The process of separating physiologically active compounds from various plants is an overall framework in the research for resource use strategies that are both ecologically sound and driven by value. Both the pharmaceutical and food sectors have shown a growing fascination in developing and implementing green extraction methods in recent decades in an effort to reduce their negative effects on the environment and advance sustainability [ 10 , 11 ]. In this regard, supercritical extraction methods have become a crucial path, with their long history that reflects the development of extraction techniques. Supercritical CO 2 extraction is thought to be a more ecologically friendly and efficient process than solvent-based extractions as it has a positive values including its non-flammability, non-toxicity, and ease of recycling [ 12 , 13 ]. The bioactive components are being characterized and quantified using a variety of methods involving chromatography following the extraction of phytochemicals. Cardamom phytochemicals have been shown to offer several health benefits in recent years, including antioxidative, antibacterial, and anti-hypertensive effects [ 14 , 15 ]. In rats fed a high-fat diet, cardamom phytochemicals dramatically reduced the levels of triglycerides [ 16 ]. Cardamom phytochemicals were shown to suppress the growth of bacteria and fungi that cause foodborne illnesses [ 17 ]. The reactive nature of free radicals can be inhibited by antioxidant substances [ 18 ]. An inherent means of defense versus reactive oxygen species (ROS) is the production of indigenous or intracellular antioxidants by the human body. It was discovered that plant-based antioxidants were more successful than chemical-based ones in reducing ROS levels [ 19 ]. Secondary plant metabolites involve molecules known as polyphenols, which include phenolic and flavonoid chemicals, are molecules that contain several phenolic rings with several useful roles [ 20 , 21 ]. A useful technique that simplifies the conduct of in vitro investigations by doing away with the need for high-throughput testing of a large number of molecules commonly used in the drug development process is molecular docking investigation [ 22 – 25 ]. This work was designed to test the impact of applying of various pressure levels (150, 200, and 300 Bar) in supercritical fluid extraction (SFE) of green cardamom, illustrating the variations in polyphenols levels in each treatment using HPLC, biological applications of various resulted extracts as well as molecular docking for molecules with high levels in the extract. Materials and Methods Plant & Chemicals Green cardamom powder was purchased from an Egyptian market authorized retailer (Giza, Egypt Code: 03547). The container of plastic was used to hold the powder at the ambient temperature for further experimentation. The used chemicals were purchased from (MOPOCO Co. INC., Egypt). SFE condition for green cardamom In a supercritical fluid extraction device, five grams of green cardamom powder are processed. A high-pressure piston forced the carbon dioxide of the cylindrical vessel onto the chiller and to the extraction tank. The instrument panel of the extraction device was set to the extract's length (15 minutes for statistics, and 45 minutes for dynamic extraction), the temperature at which it operated (60°C), and the pressures of 100 (Code: 1), 200 (Code: 2), and 300 bar (Code: 3). The extraction device unsealed the fitting across the pump and the specimen cartridge, enabling 5.0 mL/min of CO2 to flow across the specimen once the required temperature and pressure were reached. Following each the extraction procedure, the weight was determined and the extract was taken away in a glass container [ 26 ]. HPLC testing for green cardamom extracts A PerkinElmer LC300 was used to perform an HPLC test on 5.0 µL of each sample. Using a KNALJER column of C18 (4.7 mm × 250.0 mm, 6 µm, 42°C), the separation procedure occurred. The mobile phase for the HPLC was 0.06% trifluoroacetic acid diluted in acetonitrile (2nd) at an average flow rate of 0.8 mL/min and H2O (1st). In the subsequent control, the mobile phase was progressively altered in a straight gradient from 0 to 6 minutes (81%), 6 to 9 minutes (60%), 9 to 13 minutes (60%), for minutes 13 to 16 (83% 1st), for minutes 16 to 18 (82%), and for minutes 18 to 23 (83%). Using an HPLC detection device, a signal at 285 nm was detected [ 27 ]. Action of various extracts of green cardamom samples towards test microbes A group of various test food-born microbes including: Bacillus subtilis (ATCC6051), Staphylococcus aureus (ATCC43300), Klebsiella pneumoniae (ATCC2146), Salmonella typhi (ATCC14028), Candida albicans (ATCC90028), Penicillium glabrum (ATCC11080) were applied as test microbes. The fungi were streaked onto the area of the malt extract culture in the petri dishes, and the examined bacteria were injected onto the surface of the nutrient agar. Wells (5.0 mm) were formed in the infected agar layer, subsequently 100 µL of various extracts of green cardamom were placed in each well. The standard antifungal was fluconazole at 1000 µg/mL, while the standard antibiotic was gentamicin at 20 µg/mL. To allow the studied components to diffuse before microbial development, the plates were stored at 5°C for 20 minutes. Following a 24-hour incubation period at 37°C (for bacteria), 5–7 days at 28°C (for fungi) the inhibition zones surrounding the wells were measured [ 28 ]. Evaluation the values of MIC and MBC of green cardamom samples towards test microbes The minimal inhibitory levels (MIC) of investigated samples versus microorganisms were determined using the micro-dilution technique. 100 µl of repetitions of each created level were evaluated in a fluid medium made up for each location in 96 tissue culture plates, while each specimen had been diluted for values ranging from 0.95 to 1000.0 µg/mL. A 1.3X 10 6 CFU/ml was obtained by inoculating microbe cultures (1.0 McFarland standard) with 2.9 µl of sanitized 0.9% sodium chloride for every well. The bacteria were cultivated in the wells for 3 days at 38°C, while the fungal strains were left to grow for 5–8 days at 28°C. While specimens containing 100% inhibitory agent were moved to the holes for appropriate incubation periods of time, the organisms being studied were cultivated on their appropriate media for minimal bactericidal levels (MBC), which was the lowest level of specimens that could not sustain the growth of examined microbes in the examined circumstance [ 24 ]. Antibiofilm action of prepared specimens of green cardamom The effect of the tested materials on the growth of bacterial biofilms was evaluated in 96-well plates. To put it succinctly, 300 µL of freshly introduced trypticase soy yeast broth (TSY) was divided into each microplate well, and it was cultured with previously determined sub-lethal concentrations of MBC (75, 50, and 25%). The ultimate amount of TSY was 10 6 CFU/mL. Wells containing the medium, methanol, and no extracts were used as controls. The plates were then left at 38°C for 2 days. Following the incubation process the supernatant was discarded, and sterile distilled water was used to thoroughly clean the free-floating bacterial cells from every well in the experiment. After 35 minutes of air drying, the plates were dyed with a 0.1% crystal violet fluid solution for 15 minutes at ambient temperature to reveal the biofilm that had formed. After incubation, sterile distilled water was used to remove any remaining stain. Lastly, 250 µL of 95% ethanol was added to each well to dissolve the dye that was bound to the bacterial cells. After 16 minutes of incubation, absorption was determined at an intensity of 580 nm employing an elisa reader [ 29 ]. Hemolysis inhibition test The hemolysin effects of samples of green cardamom in sub-MIC (25.0%, 50.0%, and 75.0% of MIC) were measured using the Rossignol et al. technique. After being adjusted to an OD 595 of 0.4, the tested bacteria underwent spinning at 20,000× g for 23 minutes with 25.0%, 50.0%, and 75.0% of MIC (sub-MIC) or samples that were untreated. 500 µL of supernatants were combined with freshly made erythrocyte solution (2.0%) in 0.80 mL of saline, kept at 36°C for 120 minutes, and then spun at 15,000 × g for ten minutes at 4°C. To generate a positive control, an erythrocyte suspension was combined with 0.1% sodium dodecyl sulfate. For generating a control set of un-hemolyzed erythrocytes, erythrocytes were cultured in LB broth under similar circumstances. Monitoring was carried out in triplicate, and the amount of hemoglobin produced was determined through assessing absorption at 545 nm. Mean ± standard error of the variation in percentage from the hemolysis of the control group with no treated settings was used to represent the hemolysis that occurred with samples that were tested in sub-MIC [ 30 ]. Killing Kinetic times test This investigation was conducted to investigate the time dying of evaluated for test specimens against the test microorganisms. 0.7 McFarland solution was combined with fresh bacterial stains. Containers containing broth media infected with 1.10×10 6 bacterial strains were combined with the MIC values of the examined oil forms. At 0, 30, 60, 90, 120, 150, and 180 minutes, 0.5 ml of samples were taken, cultured on dishes, and stored at 38°C for the night. However, organisms lacking tested samples were present in control sample. Over the course of the tests, a shift in the number of bacterial cells was calculated [ 31 ]. Ultrastructure examination using transmission electron microscope The specimens of B. subtili s and C. albicans , comprising untreated microbes and those subjected to the MIC of green cardamom (extracted at 300 bar), each, had been fixed in an aqueous glutaraldehyde (2.6%) solution for 48 hours at 5°C, post-fixed with a 1% osmium tetroxide liquid for 4.0 hours at 5°C, sectioned via an ultra-microtome (Leica, Germany), and then investigated with a TEM (JEOL 1010) [ 32 ]. Docking analysis Molecular docking was performed using MOE (Chemical Computing Group) with the following parameters: 1-Protein Preparation Structures 5VX6 and 3V8J were retrieved from the PDB, protonated, and energy-minimized then the site finder created the active binding sites, which served as the binding pocket's dummy sites. 2-Ligand Preparation Gallic acid and Syringic acid were optimized with the MMFF94x force field to achieve a low-energy conformation. 3-Docking Procedure The ligands were put at the site using the triangle matcher approach, and the stiff receptor atoms were docked for 100 nanoseconds. The GBVI/WSA dG processes were used for rescoring, with the London dG acting as a scoring function. For each ligand-protein pair, several postures were created, and the top five were chosen for further investigation. 2D and 3D interaction diagrams were created to show how both ligands attach to each protein's active regions. These graphical representations focused on specific interactions. The docked complexes were examined to identify the interactions between the studied ligands and the active site residues of the protein. Statistics for the results The findings of the study were verified three times using the mean standard deviation (± SD). The outcome of experiments was evaluated by a one-way ANOVA applying the Graph Pad Prism V8 program (CA, USA). Result and discussion In this work, SFE-CO 2 of green cardamom seeds powder packed in the Extractor cartilage and exposed to three different pressures 100, 200, and 300 bar. It is clear that this approach was effective in yield of extract. As the pressure raised from 150 to 200 to 300 the yield of extract increased from 0.189 to 0.217 to 0.279 g (Table 1 ). According to earlier study on the SFE fundamental terms, higher pressure raised the SFE's frequency and diffusion, which improved recovery [ 33 ]. Furthermore, according to a different investigation, higher pressure has essential influence on the use of SFE in medicinal plants recovery [ 34 ]. Table 1 SFE-CO 2 of green cardamom at different pressure levels. Condition code Temp. 0 C Pressure Bar Extraction Static time (min) Extraction dynamic time (min) Quantity (gm) Extract quantity (gm) 1 60 100 15 45 5.0 0.189 ± 0.01 2 60 200 15 45 5.0 0.217 ± 0.01 3 60 300 15 45 5.0 0.279 ± 0.02 Based on the injected standards of phenols and flavonoids in HPLC, 13 compounds were appeared in the extract but with various quantities depending on the extraction pressure (Table 2 and Figs. 1 – 3 ). The concentration of eight compounds was increased with the increasing of extraction pressure while the concentration of four compounds was decreased with the increasing of extraction pressure. One compound namely rosmarinic acid was appeared in the extract at extraction condition of 200 and 300 with concentration 24.84 and 57.29 µg/g, respectively but didn’t appear in the extract at extraction condition of 100. For instance, Gallic acid increased from 77.17 to 203.93 to 461.52 µg/g, Caffeic acid increased from 87.87 to 137.81 to 216.83 µg/g, Syringic acid increased from 144.90 to 220.52 to 315.57µg/g, while Chlorogenic acid decreased from 345.20 to 216.64 to 242.36 µg/g, Rutin decreased from 139.98 to 144.98 to 45.00 µg/g, Methyl gallate decreased from 115.83 to 92.51 to 74.64 µg/g when the extraction pressure increased 100 to 200 to 300, respectively. Our findings indicated that pressure was effective for releasing the compounds from the extract. Moreover, the obtained results were matching with other investigation studied the SFE at different pressures. The maceration technique [ 35 ], extraction through supercritical fluid [ 36 ], and the use of Soxhlet extraction [ 37 ] are the techniques most commonly employed for obtaining flavonoid and phenolic components from spices. The main determinants of the quantity of antioxidants obtained are the extraction process and time [ 38 ]. In addition to pressure, other factors that can impact the yields obtained from extraction of molecules, which can be measured as total flavonoid and phenolic contents, include length of time and temperature [ 39 ]. Table 2 Phenols and flavonoid compounds of in the extract of green cardamom at different pressure levels. Detected constituents 100 200 300 RT Area % Conc. (µg/g) RT Area % Conc. (µg/ml) RT Area % Conc. (µg/g) Gallic acid 3.52 4.13 77.17 3.52 10.07 203.93 3.52 17.48 461.52 Chlorogenic acid 4.23 9.71 345.20 4.23 5.62 216.64 4.22 4.82 242.36 Methyl gallate 5.51 8.12 115.83 5.51 5.98 92.51 5.51 3.70 74.64 Caffeic acid 5.79 6.72 87.87 5.80 9.71 137.81 5.80 11.72 216.83 Syringic acid 6.29 9.66 144.90 6.29 13.55 220.52 6.29 14.88 315.57 Rutin 6.94 3.67 139.98 6.95 3.50 144.98 6.94 0.83 45.00 Coumaric acid 8.58 27.76 254.42 8.58 28.06 278.96 8.58 21.73 281.54 Vanillin 8.94 11.18 112.15 8.95 12.40 114.72 8.94 10.85 141.94 Ferulic acid 9.60 2.40 38.56 9.61 2.97 41.27 9.61 2.21 46.24 Naringenin 10.20 1.93 49.22 10.16 2.78 65.45 10.15 2.33 77.62 Rosmarinic acid 11.74 0.00 0.00 11.83 0.92 24.84 11.83 1.64 57.29 Quercetin 17.24 0.67 23.06 17.55 1.22 38.72 17.54 1.42 63.83 Cinnamic acid 19.10 11.04 54.56 19.13 6.40 34.31 19.14 6.37 44.52 Table 3 Antimicrobail action (mm) & MIC and MBC (µg/ml) levels of green cardamom extracts upon using various pressure levels (100, 200, and 300 bar) (outcomes are represnted as means ± SD). Tested microbes Zone of inhibition (mm) MIC/MBC (µg/ml) 100 200 300 standard 100 200 300 B. subtilis 18.17 ± 0.29 20.83 ± 0.29 23.33 ± 0.58 25.17 ± 1.26 125/500 31.25/62.5 15.62/15.62 S. aureus 15.50 ± 0.87 17.83 ± 0.29 18.67 ± 1.53 16.33 ± 0.58 125/250 62.5/62.5 31.25/31.25 K. pneumoniae 8.67 ± 0.58 15.83 ± 1.04 20.33 ± 0.58 18.67 ± 1.53 1000/-- 125/250 15.62/31.25 S. typhi NA 10.75 ± 0.43 16.33 ± 0.58 14.67 ± 1.15 ---/-- 250/500 250/500 C. albicans 14.33 ± 0.58 19.33 ± 2.08 22.17 ± 1.04 21.83 ± 0.29 125/250 62.5/125 31.25/62.5 P. glabrum NA NA NA 28.33 ± 2.52 ---/-- ---/-- ---/-- The recorded data in table (3) and Figure (4) showed that the obtained extract at 100, 200, and 300 possess antimicrobial activity against B. subtilis, S. aureus, K. pneumonia , and C. albicans with various inhibition zones based on the extraction condition and tested microbe. The inhibition zones were 18.17 ± 0.29, 20.83 ± 0.29, and 23.33 ± 0.58 mm for B. subtilis , 8.67 ± 0.58, 15.83 ± 1.04, and 20.33 ± 0.58 mm for K. pneumonia , 14.33 ± 0.58, 19.33 ± 2.08, and 22.17 ± 1.04 mm for C. albicans employing the extract at 100, 200, and 300, respectively. On the other hand the extract at 100 of the extraction condition faild to inhibit S. typhi while exhibited 10.75 ± 0.43 and 16.33 ± 0.58 mm inhibition zones at 200, and 300 of extraction condition. The filamentous fungus P. glabrum didn’t inhibited by the extract at any studied extraction condition. The recorded results were compared to the inhibitory action of standard antibiotic/ antifungal drugs. Furthermore, the measured MIC and MBC confirmed that extract efficacy increased with increasing extraction condition of pressure, where less MIC and MBC quantities at 300 were recorded than that recorded at 100 or 200 against tested microbes. Cardamom seeds add a sweet and tangy flavor. As an antimicrobial substance that treats dental cavities, it is frequently used as a breath freshener to preserve oral health [ 40 ]. Numerous investigations on the extraction of naturally occurring non-antibiotic compounds with possible antibacterial effects—like SFE extracts—for the treatment of multidrug-resistant microbes have been published [ 41 ]. Antibiofilm activity of the extract was estimated against tested bacteria as depicted in (Fig. 5 ). The biofilm formation was inhibited by the extract and increased with the increasing the extract concentration. At the same time the inhibition of biofilm formation was more affected by the extract at the extraction condition at 300 Bar than that by the extract at the extraction condition at 100 and 200. The biofilm of B. subtilis was the most sensitive followed by S. typhi, K. pneumonia , and S. aureus where biofilm inhibition was 95.64 ± 0.98, 94.00 ± 1.22, 92.96 ± 0.66, and 92.32 ± 1.33%, respectively by the extract at extraction condition of 300 bar (Fig. 6 ). A bacterial biofilm is a multidimensional framework made up of mono- or multi-microbial populations attached to an outer layer of cells that gives microorganisms resistance to challenging circumstances, antimicrobial agents, and the human immune system's strategies [ 42 ]. Furthermore, Nassar et al. [ 43 ] observed that microorganisms that cause a variety of human diseases and infections acquired in hospitals have demonstrated a biofilm origin. Accordingly, safe antibacterial substances that can hinder biofilm formation are needed to eradicate long-lasting infections linked to biofilms and lessen their detrimental effect on the health of people. The present investigation revealed that the raise of produced compounds upon elevation of the applied pressure enhance the antibiofilm impact of green cardamom. Investigation on shrub phenolics' antibiofilm properties has shown that, in addition to their harmful effects on bacteria, these compounds also have "softer" effects that minimize biofilms by interfering with bacterial regulating mechanisms like quorum recognition or other universal regulatory frameworks, all without influencing bacterial growth [ 29 ]. The ability of tested extract to inhibite the hemolysis in the presence of tested bacteria was measured (Figs. 7 , 8 ). As the concentration of extract increase, the hemolysis inhibition % increased as well as the extract at extraction condition 300 was effective for hemolysis inhibition % compared to the extract at extraction condition 200 and 100 in all cases of tested bacteria. At 75% MIC of the extract at extraction conditions 100, 200, and 300, the hemolysis inhibition % was 92.53 ± 2.41, 93.53 ± 1.10, and 94.27 ± 0.46 in the presence of S. aureus , 90.60 ± 3.11, 93.67 ± 1.76, and 94.80 ± 0.17 in the presence of S. typhi , 89.37 ± 0.47, 93.53 ± 3.41, and 96.66 ± 2.52 in the presence of B. subtilis , 90.33 ± 1.53, 93.70 ± 1.00, and 97.66 ± 2.08 in the presence of K. pneumonia. Since hemolysin is the primary feature of S. aureus strains' virulence, it has been suggested that antivirulence/antitoxin strategies ought to prioritize it [ 44 ]. Flavonoids have been demonstrated to aggressively counteract the hemolytic properties of S. aureus 's α-hemolysin, disrupting the structure of the toxin [ 45 ]. An other way natural extracts exhibits antihemolytic effect is by strengthening the erythrocyte membrane's resistance to the toxin [ 46 ]. Various types of green cardamom extracts which prepared upon using different pressure values (100, 200, and 300 bar) towards different examined bacteria at different time points could be seen at (Table 4 ). A daramatic reduction ( P ≤ 0.05) in bacterial count could be seen upon using green cardamom extract which prepared at 300 bar relaive to the other two forms of green cardamom extracts. The prepared cardamom extract at 100 bar showed a minimal impact towards B. subtilis at different time points where 324 ± 12 bacterial cells could be detected after 180 minutes. There is no bacterial cells after exposing various bacteria for cardamom extract at 300 bar after 150 minutes revealing its effective impact at earlier examined time points. Time killing kinetics confirmed the bactericidal impact of cardamom extract especially at the prepared one at 300 bar. A few research investigations have been conducted on the time-kill kinetics of cardamom, while numerous studies of natural product extracts have been published [ 47 ]. Killing examination revealed that the level of time-varying inhibition of microbes differed between bacteria and natural product extracts [ 48 ]. Plant-derived compounds and reactions following microbial infection may be taken into consideration when evaluating the antibacterial attributes [ 49 ]. Table 4 Effect of different Killing Kinetic times of green cardamom extracts upon using various pressure levels (100, 200, and 300 bar) towards B. subtilis , S. aureus , K. pneumonia , and S. typhi , outcomes are represnted as means ± SD). Killing Kinetic time (min) CFU B. subtilis S. aureus 100 200 300 100 200 300 0 26×10 5 ±12 26×10 5 ±9 26×10 5 ±2.0 29×10 5 ±11 29×10 5 ±12 29×10 5 ±8 30 6×10 5 ±7 56×10 4 ±6 287×10 3 ±1 232×10 4 ±9 22×10 4 ±4 15×10 4 ±6 60 128×10 4 ± 8 28×10 3 ± 11 18×10 2 ± 2 117×10 3 ±7 79×10 3 ±8 23×10 2 ±4 120 184×10 3 ±11 174×10 ± 4 70×10 ± 3 13×10 2 ±6 210 ± 6 167 ± 3 150 25 ×10 2 ±12 291 ± 3 0 380 21 ± 2 0 180 324 ± 12 0 0 0 0 0 Killing Kinetic time (min) CFU 100 200 300 100 200 300 K. pneumoniae S. typhi 24×10 5 ±8 24×10 5 ±12 24×10 5 ±9 5×10 5 ±6 5×10 5 ±1 5×10 5 ±4 30 239×10 4 ±6 30×10 4 ±14 13×10 4 ±8 221×10 4 ±8 94×10 4 ±2 19×10 4 ±11 60 157×10 3 ±5 17×10 3 ±8 213×10 2 ±11 242×10 3 ±4 142×10 3 ±1 12×10 3 ±14 120 176×10 ± 4 250 ± 7 231 126×10 2 ±6 15×10 2 ±2 200 ± 8 150 193 ± 3 11 ± 4 0 292 294 0 180 0 0 0 0 0 0 The green cardamom extract obtained by SFE extraction at 300 Bar caused numerous ultrastructural changes in B. subtilis and C. albicans cells in the current investigation. TEM images (Fig. 9 A) of the untreated B. subtilis specimen showed that each individual cell was healthy, rod-shaped, and encompassed by the outermost and innermost layers with no any damage to the cell. When B. subtilis cells were treated with green cardamom extract, their size drastically shrank and several lysed cells were visible (Fig. 9 B). TEM pictures (Fig. 9 C) showed that individual C. albicans cells in the reference specimen were well, rounded, and classically shaped; however, when green cardamom was applied, the C. albicans cells were destructed, resulting in deformed cells with a diminished shape (Fig. 9 D). To fight the microbes that cause different infections and illnesses, new antimicrobial substances had to be created [ 50 ]. The TEM technique can provide valuable insight into the antibacterial mechanisms of action of new antibacterial substances to confirm the antimicrobial outcomes through visualization of various alterations upon treatments [ 51 ]. In silico investigation though docking analysis Syringic acid showed stronger binding (S = -4.27 to -4.64 for 3V8J; S = -4.69 to -5.04 for 5VX6) than gallic acid (S = -4.11 to -4.45 for 3V8J; S = -4.52 to -4.68 for 5VX6). Lower RMSD_refine values indicated stable poses. E_conf values reflecting conformational stability. E_place and E_refine values were consistently favorable. Key Interactions for B. subtilis (5VX6) indicated that gallic acid formed H-bonds with GLU 187 (2.73 Å, -5.4 kcal/mol) and ARG 172 (2.94 Å, -4.4 kcal/mol). Syringic acid interacted with GLU 187 (2.82 Å, -7.3 kcal/mol), ARG 172 (3.10 Å, -1.4 kcal/mol), and AR 104 (3.04 Å, -0.8 kcal/mol). In S. aureus (3V8J): Both acids bound to ASP 239 (Gallic acid: 2.85 Å, -5.4 kcal/mol; Syringic acid: 2.86 Å, -4.8 kcal/mol). The important docking metrics, such as docking scores, interaction types, binding energies, and distances, have been reviewed to determine binding affinities and interaction processes, as shown in Tables 5 – 8 . The best-fitted diagrams of 2D and 3D poses select by the syringic acid and gallic acid are described (Fig. 10 ). Regarding the discussion of docking analysis, the results reveal nuanced differences in the binding behavior of gallic acid and syringic acid against Bacillus subtilis (5VX6) and S. aureus (3V8J), offering insights into their potential as antimicrobial agents. The presence of methoxy groups in Syringic acid likely enhances its binding. While methoxy groups are not directly involved in hydrogen bonding, their electron-donating effects may subtly modulate the electronic environment of Syringic acid’s hydroxyl groups, enhancing their ability to form stable bonds with residues like GLU 187. In B. subtilis (5VX6), syringic acid forms a stronger hydrogen bond with GLU 187 (-7.3 kcal/mol) compared to Gallic acid (-5.4 kcal/mol), likely due to optimal alignment of its hydroxyl groups. Additional interactions with ARG 172 and ARG 104 (Table 7 ) may stabilize its pose despite the absence of methoxy group participation. Table 5 Docking scores and energies of gallic acid and syringic acid with structure of B. subtilis (PDB ID: 5VX6). Mol S rmsd_refine E_conf E_place E_score1 E_refine E_score2 Gallic acid -4.67921 1.7332464 -31.8259 -56.9843 -9.70131 -23.0596 -4.67921 Gallic acid -4.59409 0.74464136 -32.8286 -68.1666 -10.2249 -24.6589 -4.59409 Gallic acid -4.58793 1.7696532 -33.2433 -63.2359 -9.91201 -25.4289 -4.58793 Gallic acid -4.56066 2.6160221 -31.7368 -52.0508 -10.3878 -23.347 -4.56066 Gallic acid -4.51797 2.1812108 -33.1922 -47.8645 -9.88048 -21.0348 -4.51797 Syringic acid -5.03796 0.52116263 -14.0667 -78.0995 -10.1364 -28.6602 -5.03796 Syringic acid -4.94237 0.94439775 -13.9608 -70.5491 -10.0975 -25.0605 -4.94237 Syringic acid -4.92053 2.111726 -5.60061 -64.9127 -10.0869 -22.5365 -4.92053 Syringic acid -4.69705 1.4523685 -13.7555 -67.6678 -10.1653 -22.6829 -4.69705 Syringic acid -4.67621 1.2140613 -7.97653 -61.7188 -10.9312 -21.6721 -4.67621 Table 6 Docking scores and energies of gallic acid and syringic acid with structure of S. aureus (PDB ID: 3V8J). Mol S rmsd_refine E_conf E_place E_score1 E_refine E_score2 Gallic acid -4.45374 1.4267793 -32.8569 -50.573 -9.60852 -22.523 -4.45374 Gallic acid -4.28895 1.3745345 -30.6581 -53.7743 -9.88156 -19.5394 -4.28895 Gallic acid -4.19007 2.6599638 -33.9026 -41.8718 -11.3473 -17.8853 -4.19007 Gallic acid -4.18197 1.3183807 -32.5074 -69.8018 -11.9495 -17.2698 -4.18197 Gallic acid -4.11419 1.4595817 -32.7511 -55.7456 -9.92507 -15.8203 -4.11419 Syringic acid -4.64437 0.73497051 -12.4447 -60.9532 -10.6667 -23.4391 -4.64437 Syringic acid -4.52733 0.98655659 -7.59219 -61.9997 -10.1858 -19.1627 -4.52733 Syringic acid -4.31496 1.6934763 -10.5394 -60.394 -10.1252 -19.3342 -4.31496 Syringic acid -4.28199 1.5533125 -10.5155 -61.2405 -9.95271 -17.4618 -4.28199 Syringic acid -4.2695 3.3460429 -10.8598 -58.6044 -10.1436 -17.6905 -4.2695 Table 7 Interaction of gallic acid and syringic acid with structure of B. subtilis (PDB ID: 5VX6). Mol Ligand Receptor Interaction Distance E (kcal/mol) Gallic acid O 13 OE2 GLU 187 (A) H-donor 2.73 -5.4 O 13 NH1 ARG 172 (A) H-acceptor 2.94 -4.4 Syringic acid O 23 OE2 GLU 187 (A) H-donor 2.82 -7.3 O 14 NH1 ARG 172 (A) H-acceptor 3.10 -1.4 O 22 NE ARG 104 (A) H-acceptor 3.04 -0.8 Table 8 Interaction of gallic acid and syringic acid with structure of S. aureus (PDB ID: 3V8J). Mol Ligand Receptor Interaction Distance E (kcal/mol) Gallic acid O 17 OD2 ASP 239 (A) H-donor 2.85 -5.4 Syringic acid O 23 OD2 ASP 239 (A) H-donor 2.86 -4.8 Steric Compatibility Despite its larger size, Syringic acid’s structure appears better accommodated in the binding pockets, as reflected in lower RMSD_refine values (e.g., 0.52 Å in 5VX6), indicating stable pose retention post-refinement. Energy Term Analysis indicated that gallic acid showed more favorable conformational energy (E_conf) values (-31.8 to -33.9 kcal/mol vs. Syringic acid’s -5.6 to -14.1 kcal/mol), suggesting its simpler structure adopts lower-energy conformations. However, syringic acid’s higher (less negative) E_conf may reflect conformational adjustments to maximize interactions, ultimately yielding stronger overall binding (S). Placement and Refinement Energies : Syringic acid’s superior E_place (e.g., -78.10 kcal/mol in 5VX6) indicates efficient initial docking, while its moderate E_refine penalties (e.g., -28.66 kcal/mol) suggest minor adjustments to optimize interactions. Several investigators employed the molecular docking to document the activity of some phenolic and flavonoids against cancer cells, pathogenic bacteria, and yeasts [ 52 – 56 ]. For example, catechin and gallic acid were docked with prostate cancer cells 3 PDB ID: 2Q7L with binding affinities − 5.27521 and − 6.3204 kcal/mol [ 57 ]. Rosmarinic acid was docked with C. tropicalis (PDB ID: 6ZD6), G. candidum (PDB ID: 6ISV), and C. albicans (PDB ID: 1ZAP) with binding affinities − 6.8224 kcal/mol, -6.79996, and − 6.15839 kcal/mol, respectively [ 13 ]. According to Alsalamah et al. [ 58 ], ellagic acid showed − 4.5145 kcal/mol and − 6.18615 kcal/mol, while chlorogenic acid showed − 5.69876 kcal/mol and − 7.84379 kcal/mol as docked with G. candidum (4ZZT) and C. albicans (4ZZT) proteins, respectively. Conclusion This is a novel investigation to extract a substantial quantity of phenolic components from green cardamom employing the SFE approach. The SFE apparatus's pressure has been adjusted to maximize both the yield and the quality of the extracted substances. Additionally, a large number of chemicals in extracted green cardamom have been found to have about 13 bioactive components using the HPLC method. These compounds are recognized to have both medicinal and effective antimicrobial properties. Remarkably, in contrast to the other conditions of treatment, the extracted output rose under high pressure (300 bar). High antimicrobial, antibiofilm, and antihemolytic properties were demonstrated by all green cardamom extracts; however, when contrasted with the other two pressures, the strongest activity was noted at 300 bar. Syringic and gallic acids exhibit promising binding to B. subtilis and S. aureus targets, with syringic acid showing superior affinity. The interactions with key catalytic residues (GLU 187, ASP 239) provide a mechanistic basis for inhibition. Declarations Ethics approval and consent to participate : Not applicable Consent for publication : Not applicable Data Availability Statement: The results from the current investigation are available from the corresponding author upon reasonable appeal. Conflicts of Interest: The authors declare no conflicts of interest Author Contributions: S.A.A. Investigation, writing—review and editing; M.I.A., and K.S.I., A.M., and T.M.A. Conceptualization and methodology. All authors agreed to the published version of the manuscript. ACKNOWLEDGMENT The authors wish to appreciate the Deanship of Scientific Research at Imam Mohammad Ibn Saud Islamic University (IMSIU) for supported and funded the current study (Grant number grant number IMSIU-DDRSP2501) FUNDING This work was supported and funded by the Deanship of Scientific Research at Imam Mohammad Ibn Saud Islamic University (IMSIU) (grant number IMSIU-DDRSP2501). References Abdelghany, T. M., Hassan, M. M., El-Naggar, M. A., & Abd El-Mongy, M. 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Alsalamah","email":"","orcid":"","institution":"Al-Imam University: Imam Muhammad Ibn Saud Islamic University","correspondingAuthor":false,"prefix":"","firstName":"Sulaiman","middleName":"A.","lastName":"Alsalamah","suffix":""},{"id":447969196,"identity":"85c722a6-3464-42cd-a55f-779943d66dde","order_by":1,"name":"Mohammed Ibrahim Alghonaim","email":"","orcid":"","institution":"Al-Imam University: Imam Muhammad Ibn Saud Islamic University","correspondingAuthor":false,"prefix":"","firstName":"Mohammed","middleName":"Ibrahim","lastName":"Alghonaim","suffix":""},{"id":447969197,"identity":"bb9bb695-b6e5-478a-b1ad-94e654fb3549","order_by":2,"name":"Khatib Sayeed Ismail","email":"","orcid":"","institution":"Jazan University","correspondingAuthor":false,"prefix":"","firstName":"Khatib","middleName":"Sayeed","lastName":"Ismail","suffix":""},{"id":447969198,"identity":"ce41ff3e-bd78-4b69-abff-8a45cf7d473e","order_by":3,"name":"Abdullah Mashraqi","email":"","orcid":"","institution":"Jazan University College of Science","correspondingAuthor":false,"prefix":"","firstName":"Abdullah","middleName":"","lastName":"Mashraqi","suffix":""},{"id":447969199,"identity":"5428c36c-4720-493c-bf88-7cea4567dec9","order_by":4,"name":"Tarek Mohamed","email":"data:image/png;base64,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","orcid":"https://orcid.org/0000-0001-6418-5890","institution":"Al-Azhar University","correspondingAuthor":true,"prefix":"","firstName":"Tarek","middleName":"","lastName":"Mohamed","suffix":""}],"badges":[],"createdAt":"2025-04-18 18:45:48","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6480943/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6480943/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s40643-025-00951-z","type":"published","date":"2025-12-08T15:57:03+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":81639532,"identity":"d6175fe0-b675-4c6f-8ec0-e337f92a8c5e","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":44093,"visible":true,"origin":"","legend":"\u003cp\u003eHPLC chromatogram for green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e upon SFE extraction at 100 bar.\u003c/p\u003e","description":"","filename":"1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/ceeb10bd1ffab3e65865c1de.jpg"},{"id":81640882,"identity":"ce4a9bed-5c05-42a0-b659-f31cd60b352c","added_by":"auto","created_at":"2025-04-29 13:26:46","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":44248,"visible":true,"origin":"","legend":"\u003cp\u003eHPLC chromatogram for green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e upon SFE extraction at 200 bar.\u003c/p\u003e","description":"","filename":"2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/9d77340b9479ce0d79028451.jpg"},{"id":81639536,"identity":"1bdf84bb-1b25-4b51-a28c-96a66ab15122","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":41330,"visible":true,"origin":"","legend":"\u003cp\u003eHPLC chromatogram for green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003eupon SFE extraction at 300 bar.\u003c/p\u003e","description":"","filename":"3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/eeffa2ec660411471ba063a4.jpg"},{"id":81639533,"identity":"25b04809-dbef-4976-bf3c-c8078fb3be94","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":93891,"visible":true,"origin":"","legend":"\u003cp\u003eAntimicrobail action of of green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e extracts upon using various pressure levels (100, 200, and 300 bar)(-ve: negative control, SD: standard drug).\u003c/p\u003e","description":"","filename":"4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/0c464cd9c1d5cfcb377e87bd.jpg"},{"id":81641163,"identity":"360aa7bc-ae98-4d09-81ad-321e58f5d954","added_by":"auto","created_at":"2025-04-29 13:34:46","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":113260,"visible":true,"origin":"","legend":"\u003cp\u003eAntibiofilm action of green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e extracts upon using various pressure levels (100, 200, and 300 bar)towards \u003cem\u003e(A) K. pneumonia, \u003c/em\u003e(B) \u003cem\u003eS. typhi,\u003c/em\u003e (C) \u003cem\u003eB. subtilis\u003c/em\u003e and (D)\u003cem\u003eS. aureus\u003c/em\u003e.\u003c/p\u003e","description":"","filename":"5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/c9439a12b8f114a10ee4bfa2.jpg"},{"id":81640883,"identity":"c70472a2-8e9d-4fd5-a5a0-1a3783ade770","added_by":"auto","created_at":"2025-04-29 13:26:46","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":77743,"visible":true,"origin":"","legend":"\u003cp\u003eStatistical analysis for antibiofilm action of green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e extracts upon using various pressure levels (100, 200, and 300 bar) towards (A)\u003cem\u003e K. pneumonia, \u003c/em\u003e(B) \u003cem\u003eS. typhi,\u003c/em\u003e (C) \u003cem\u003eB. subtilis\u003c/em\u003e and (D)\u003cem\u003eS. aureus\u003c/em\u003e outcomes are represnted as means ±SD).\u003c/p\u003e","description":"","filename":"6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/4c94852f83f8b580b324d302.jpg"},{"id":81640018,"identity":"5cf2d63d-64e5-45f9-bbcf-baf2d964b077","added_by":"auto","created_at":"2025-04-29 13:18:46","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":92584,"visible":true,"origin":"","legend":"\u003cp\u003eHemolysis inhibition %. SAA \u003cem\u003eS. aureus\u003c/em\u003e at 100 bar, SAB \u003cem\u003eS. aureus\u003c/em\u003e at 200 bar, SAC \u003cem\u003eS. aureus\u003c/em\u003e at 300 bar. KNA \u003cem\u003eK. pneumonia\u003c/em\u003e at 100 pressure, KNB \u003cem\u003eK. pneumonia\u003c/em\u003e at 200 bar, KNC \u003cem\u003eK. pneumonia\u003c/em\u003e at 300 bar. STA \u003cem\u003eS. typhi at\u003c/em\u003e 100 bar, STB \u003cem\u003eS. typhi \u003c/em\u003eat 200 pressure, STC \u003cem\u003eS. typhi \u003c/em\u003eat 300 bar. BSA \u003cem\u003eB. subtilis \u003c/em\u003eat 100 bar, BS \u003cem\u003eB. subtilis \u003c/em\u003eat 200 pressure, BS \u003cem\u003eB. subtilis \u003c/em\u003eat 300 bar.\u003c/p\u003e","description":"","filename":"7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/22bcd361fbfc95bb433ebc17.jpg"},{"id":81639539,"identity":"246cd7d9-245a-4f75-bdda-c22c3c423e26","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":73197,"visible":true,"origin":"","legend":"\u003cp\u003eStatistical analysis for Hemolysis inhibition % of green \u003ca href=\"https://context.reverso.net/%D8%A7%D9%84%D8%AA%D8%B1%D8%AC%D9%85%D8%A9/%D8%A7%D9%84%D8%A5%D9%86%D8%AC%D9%84%D9%8A%D8%B2%D9%8A%D8%A9-%D8%A7%D9%84%D8%B9%D8%B1%D8%A8%D9%8A%D8%A9/cardamom\"\u003ecardamom\u003c/a\u003e extracts upon using various pressure levels (100, 200, and 300 bar) in presence of \u003cem\u003e(A) K. pneumonia, \u003c/em\u003e(B) \u003cem\u003eS. typhi,\u003c/em\u003e (C) \u003cem\u003eB. subtilis\u003c/em\u003e and (D)\u003cem\u003eS. aureus\u003c/em\u003e outcomes are represnted as means ±SD).\u003c/p\u003e","description":"","filename":"8.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/14a855e81310d35f05fde10d.jpg"},{"id":81639542,"identity":"7b0cf233-89d6-4da1-aa03-1826a74a4065","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":51286,"visible":true,"origin":"","legend":"\u003cp\u003eTEM micrographs for (A) \u003cem\u003eB. subtilis\u003c/em\u003e, (\u003cstrong\u003eB) \u003c/strong\u003e\u003cem\u003eB. subtilis\u003c/em\u003e treated by cardamom extract obtained by SFE extraction at 300 Bar, (C) control \u003cem\u003eC. albicans, \u003c/em\u003eand (D)\u003cem\u003e C. albicans\u003c/em\u003e cells treated by cardamom extract obtained by SFE extraction at 300 Bar (Magnification 10,000 X).\u003c/p\u003e","description":"","filename":"9.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/13c61b9b699944888215ddc4.jpg"},{"id":81639545,"identity":"b5df7f2e-9c00-4cdb-9070-bb7e686ca4cd","added_by":"auto","created_at":"2025-04-29 13:10:46","extension":"jpg","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":166906,"visible":true,"origin":"","legend":"\u003cp\u003e2D and 3D diagrams show the interaction between gallic acid and active sites of \u003cem\u003eB. subtilis\u003c/em\u003e 5VX6 protein (A), 2D and 3D diagrams show the interaction between syringic acid and active sites of \u003cem\u003eB. subtilis\u003c/em\u003e 5VX6 protein (B), 2D and 3D diagrams show the interaction between gallic acid and active sites of \u003cem\u003eS. aureus\u003c/em\u003e 3V8J protein (C), 2D and 3D diagrams show the interaction between syringic acid and active sites of \u003cem\u003eS. aureus\u003c/em\u003e 3V8J protein (D), and the representative key for the types of interaction between ligands and selected protein receptors (E)\u003c/p\u003e","description":"","filename":"10.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/df4d90089ea270337872e4d9.jpg"},{"id":98243967,"identity":"6269a896-62d8-44fb-8eb6-3272fb40bd68","added_by":"auto","created_at":"2025-12-15 16:11:40","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2329439,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/d3da4e43-f3f2-47a9-b600-f75dc9bb6119.pdf"},{"id":81640025,"identity":"82194f7d-e475-410c-abcf-ee1e7f613359","added_by":"auto","created_at":"2025-04-29 13:18:46","extension":"jpg","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":71130,"visible":true,"origin":"","legend":"","description":"","filename":"GRAPHICAL.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6480943/v1/6ec89c36fcb4a56c1ef85631.jpg"}],"financialInterests":"","formattedTitle":"Innovative strategy for extraction of green cardamom via Super critical fluid extractor at different levels of pressure with its application against microorganisms in vitro and in silico","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe academic sector has long been interested in aromatic and therapeutic plants with antimicrobial capabilities because they provide numerous biologically active substances that can be applied to counteract the spreading of microorganisms that cause serious and potentially deadly infections [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. In Mediterranean areas, spices and herbaceous plants are commonly used in preparing food since they have the ability to fight towards several pathological bacteria [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. The frequent consumption of antimicrobial agents among consumers to combat pathogens explains the rise in resistance produced in most populations towards microbial infections which urge the need to use innovative tools to overcome these infections [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe green cardamom species (\u003cem\u003eElettaria cardamomum)\u003c/em\u003e is a big evergreen herb rhizomatous monocot species that is a member of the Zingiberaceae and Elettaria families [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. It is widely recognized for the flavorful components and bioactive substances found in dry mature fruit, that make them valuable commodities for the food, medicinal products and cosmetic sectors [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. In the food industry, cardamom is frequently used to flavor prepared meals, soft drinks, and confections. Furthermore, it is employed in the cosmetics industry for body and hair care items, aromatherapy, and scents [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. In the drug sector, cardamom has also been utilized as a potent antioxidant, antimicrobial agent [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe process of separating physiologically active compounds from various plants is an overall framework in the research for resource use strategies that are both ecologically sound and driven by value. Both the pharmaceutical and food sectors have shown a growing fascination in developing and implementing green extraction methods in recent decades in an effort to reduce their negative effects on the environment and advance sustainability [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. In this regard, supercritical extraction methods have become a crucial path, with their long history that reflects the development of extraction techniques. Supercritical CO\u003csub\u003e2\u003c/sub\u003e extraction is thought to be a more ecologically friendly and efficient process than solvent-based extractions as it has a positive values including its non-flammability, non-toxicity, and ease of recycling [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe bioactive components are being characterized and quantified using a variety of methods involving chromatography following the extraction of phytochemicals. Cardamom phytochemicals have been shown to offer several health benefits in recent years, including antioxidative, antibacterial, and anti-hypertensive effects [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. In rats fed a high-fat diet, cardamom phytochemicals dramatically reduced the levels of triglycerides [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Cardamom phytochemicals were shown to suppress the growth of bacteria and fungi that cause foodborne illnesses [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe reactive nature of free radicals can be inhibited by antioxidant substances [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. An inherent means of defense versus reactive oxygen species (ROS) is the production of indigenous or intracellular antioxidants by the human body. It was discovered that plant-based antioxidants were more successful than chemical-based ones in reducing ROS levels [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Secondary plant metabolites involve molecules known as polyphenols, which include phenolic and flavonoid chemicals, are molecules that contain several phenolic rings with several useful roles [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. A useful technique that simplifies the conduct of \u003cem\u003ein vitro\u003c/em\u003e investigations by doing away with the need for high-throughput testing of a large number of molecules commonly used in the drug development process is molecular docking investigation [\u003cspan additionalcitationids=\"CR23 CR24\" citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. This work was designed to test the impact of applying of various pressure levels (150, 200, and 300 Bar) in supercritical fluid extraction (SFE) of green cardamom, illustrating the variations in polyphenols levels in each treatment using HPLC, biological applications of various resulted extracts as well as molecular docking for molecules with high levels in the extract.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003ePlant \u0026amp; Chemicals\u003c/h2\u003e \u003cp\u003eGreen cardamom powder was purchased from an Egyptian market authorized retailer (Giza, Egypt Code: 03547). The container of plastic was used to hold the powder at the ambient temperature for further experimentation. The used chemicals were purchased from (MOPOCO Co. INC., Egypt).\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eSFE condition for green cardamom\u003c/h3\u003e\n\u003cp\u003eIn a supercritical fluid extraction device, five grams of green cardamom powder are processed. A high-pressure piston forced the carbon dioxide of the cylindrical vessel onto the chiller and to the extraction tank. The instrument panel of the extraction device was set to the extract's length (15 minutes for statistics, and 45 minutes for dynamic extraction), the temperature at which it operated (60°C), and the pressures of 100 (Code: 1), 200 (Code: 2), and 300 bar (Code: 3). The extraction device unsealed the fitting across the pump and the specimen cartridge, enabling 5.0 mL/min of CO2 to flow across the specimen once the required temperature and pressure were reached. Following each the extraction procedure, the weight was determined and the extract was taken away in a glass container [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e\n\u003ch3\u003eHPLC testing for green cardamom extracts\u003c/h3\u003e\n\u003cp\u003eA PerkinElmer LC300 was used to perform an HPLC test on 5.0 µL of each sample. Using a KNALJER column of C18 (4.7 mm × 250.0 mm, 6 µm, 42°C), the separation procedure occurred. The mobile phase for the HPLC was 0.06% trifluoroacetic acid diluted in acetonitrile (2nd) at an average flow rate of 0.8 mL/min and H2O (1st). In the subsequent control, the mobile phase was progressively altered in a straight gradient from 0 to 6 minutes (81%), 6 to 9 minutes (60%), 9 to 13 minutes (60%), for minutes 13 to 16 (83% 1st), for minutes 16 to 18 (82%), and for minutes 18 to 23 (83%). Using an HPLC detection device, a signal at 285 nm was detected [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e].\u003c/p\u003e\n\u003ch3\u003eAction of various extracts of green cardamom samples towards test microbes\u003c/h3\u003e\n\u003cp\u003eA group of various test food-born microbes including: \u003cem\u003eBacillus subtilis\u003c/em\u003e (ATCC6051), \u003cem\u003eStaphylococcus aureus\u003c/em\u003e (ATCC43300), \u003cem\u003eKlebsiella pneumoniae\u003c/em\u003e (ATCC2146), \u003cem\u003eSalmonella typhi\u003c/em\u003e (ATCC14028), \u003cem\u003eCandida albicans\u003c/em\u003e (ATCC90028), \u003cem\u003ePenicillium glabrum\u003c/em\u003e (ATCC11080) were applied as test microbes. The fungi were streaked onto the area of the malt extract culture in the petri dishes, and the examined bacteria were injected onto the surface of the nutrient agar. Wells (5.0 mm) were formed in the infected agar layer, subsequently 100 µL of various extracts of green cardamom were placed in each well. The standard antifungal was fluconazole at 1000 µg/mL, while the standard antibiotic was gentamicin at 20 µg/mL. To allow the studied components to diffuse before microbial development, the plates were stored at 5°C for 20 minutes. Following a 24-hour incubation period at 37°C (for bacteria), 5–7 days at 28°C (for fungi) the inhibition zones surrounding the wells were measured [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e].\u003c/p\u003e\n\u003ch3\u003eEvaluation the values of MIC and MBC of green cardamom samples towards test microbes\u003c/h3\u003e\n\u003cp\u003eThe minimal inhibitory levels (MIC) of investigated samples versus microorganisms were determined using the micro-dilution technique. 100 µl of repetitions of each created level were evaluated in a fluid medium made up for each location in 96 tissue culture plates, while each specimen had been diluted for values ranging from 0.95 to 1000.0 µg/mL. A 1.3X 10\u003csup\u003e6\u003c/sup\u003e CFU/ml was obtained by inoculating microbe cultures (1.0 McFarland standard) with 2.9 µl of sanitized 0.9% sodium chloride for every well. The bacteria were cultivated in the wells for 3 days at 38°C, while the fungal strains were left to grow for 5–8 days at 28°C. While specimens containing 100% inhibitory agent were moved to the holes for appropriate incubation periods of time, the organisms being studied were cultivated on their appropriate media for minimal bactericidal levels (MBC), which was the lowest level of specimens that could not sustain the growth of examined microbes in the examined circumstance [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e].\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eAntibiofilm action of prepared specimens of green cardamom\u003c/h2\u003e \u003cp\u003eThe effect of the tested materials on the growth of bacterial biofilms was evaluated in 96-well plates. To put it succinctly, 300 µL of freshly introduced trypticase soy yeast broth (TSY) was divided into each microplate well, and it was cultured with previously determined sub-lethal concentrations of MBC (75, 50, and 25%). The ultimate amount of TSY was 10\u003csup\u003e6\u003c/sup\u003e CFU/mL. Wells containing the medium, methanol, and no extracts were used as controls. The plates were then left at 38°C for 2 days. Following the incubation process the supernatant was discarded, and sterile distilled water was used to thoroughly clean the free-floating bacterial cells from every well in the experiment. After 35 minutes of air drying, the plates were dyed with a 0.1% crystal violet fluid solution for 15 minutes at ambient temperature to reveal the biofilm that had formed. After incubation, sterile distilled water was used to remove any remaining stain. Lastly, 250 µL of 95% ethanol was added to each well to dissolve the dye that was bound to the bacterial cells. After 16 minutes of incubation, absorption was determined at an intensity of 580 nm employing an elisa reader [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eHemolysis inhibition test\u003c/h3\u003e\n\u003cp\u003eThe hemolysin effects of samples of green cardamom in sub-MIC (25.0%, 50.0%, and 75.0% of MIC) were measured using the Rossignol et al. technique. After being adjusted to an OD\u003csub\u003e595\u003c/sub\u003e of 0.4, the tested bacteria underwent spinning at 20,000× g for 23 minutes with 25.0%, 50.0%, and 75.0% of MIC (sub-MIC) or samples that were untreated. 500 µL of supernatants were combined with freshly made erythrocyte solution (2.0%) in 0.80 mL of saline, kept at 36°C for 120 minutes, and then spun at 15,000 × g for ten minutes at 4°C. To generate a positive control, an erythrocyte suspension was combined with 0.1% sodium dodecyl sulfate. For generating a control set of un-hemolyzed erythrocytes, erythrocytes were cultured in LB broth under similar circumstances. Monitoring was carried out in triplicate, and the amount of hemoglobin produced was determined through assessing absorption at 545 nm. Mean ± standard error of the variation in percentage from the hemolysis of the control group with no treated settings was used to represent the hemolysis that occurred with samples that were tested in sub-MIC [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e].\u003c/p\u003e\n\u003ch3\u003eKilling Kinetic times test\u003c/h3\u003e\n\u003cp\u003eThis investigation was conducted to investigate the time dying of evaluated for test specimens against the test microorganisms. 0.7 McFarland solution was combined with fresh bacterial stains. Containers containing broth media infected with 1.10×10\u003csup\u003e6\u003c/sup\u003e bacterial strains were combined with the MIC values of the examined oil forms. At 0, 30, 60, 90, 120, 150, and 180 minutes, 0.5 ml of samples were taken, cultured on dishes, and stored at 38°C for the night. However, organisms lacking tested samples were present in control sample. Over the course of the tests, a shift in the number of bacterial cells was calculated [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e].\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eUltrastructure examination using transmission electron microscope\u003c/h2\u003e \u003cp\u003eThe specimens of \u003cem\u003eB. subtili\u003c/em\u003es and \u003cem\u003eC. albicans\u003c/em\u003e, comprising untreated microbes and those subjected to the MIC of green cardamom (extracted at 300 bar), each, had been fixed in an aqueous glutaraldehyde (2.6%) solution for 48 hours at 5°C, post-fixed with a 1% osmium tetroxide liquid for 4.0 hours at 5°C, sectioned via an ultra-microtome (Leica, Germany), and then investigated with a TEM (JEOL 1010) [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eDocking analysis\u003c/h2\u003e \u003cp\u003eMolecular docking was performed using MOE (Chemical Computing Group) with the following parameters:\u003c/p\u003e \u003cp\u003e \u003cstrong\u003e1-Protein Preparation\u003c/strong\u003e \u003c/p\u003e\u003cp\u003eStructures 5VX6 and 3V8J were retrieved from the PDB, protonated, and energy-minimized then the site finder created the active binding sites, which served as the binding pocket's dummy sites.\u003c/p\u003e \u003cp\u003e\u003c/p\u003e \u003cp\u003e \u003cstrong\u003e2-Ligand Preparation\u003c/strong\u003e \u003c/p\u003e\u003cp\u003eGallic acid and Syringic acid were optimized with the MMFF94x force field to achieve a low-energy conformation.\u003c/p\u003e \u003cp\u003e\u003c/p\u003e \u003cp\u003e \u003cstrong\u003e3-Docking Procedure\u003c/strong\u003e \u003c/p\u003e\u003cp\u003eThe ligands were put at the site using the triangle matcher approach, and the stiff receptor atoms were docked for 100 nanoseconds. The GBVI/WSA dG processes were used for rescoring, with the London dG acting as a scoring function. For each ligand-protein pair, several postures were created, and the top five were chosen for further investigation. 2D and 3D interaction diagrams were created to show how both ligands attach to each protein's active regions. These graphical representations focused on specific interactions. The docked complexes were examined to identify the interactions between the studied ligands and the active site residues of the protein.\u003c/p\u003e \u003cp\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eStatistics for the results\u003c/h2\u003e \u003cp\u003eThe findings of the study were verified three times using the mean standard deviation (± SD). The outcome of experiments was evaluated by a one-way ANOVA applying the Graph Pad Prism V8 program (CA, USA).\u003c/p\u003e \u003c/div\u003e "},{"header":"Result and discussion","content":"\u003cp\u003eIn this work, SFE-CO\u003csub\u003e2\u003c/sub\u003e of green cardamom seeds powder packed in the Extractor cartilage and exposed to three different pressures 100, 200, and 300 bar. It is clear that this approach was effective in yield of extract. As the pressure raised from 150 to 200 to 300 the yield of extract increased from 0.189 to 0.217 to 0.279 g (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). According to earlier study on the SFE fundamental terms, higher pressure raised the SFE's frequency and diffusion, which improved recovery [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. Furthermore, according to a different investigation, higher pressure has essential influence on the use of SFE in medicinal plants recovery [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e].\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSFE-CO\u003csub\u003e2\u003c/sub\u003e of green cardamom at different pressure levels.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCondition code\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTemp. \u003csup\u003e0\u003c/sup\u003eC\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePressure Bar\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eExtraction Static time (min)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eExtraction dynamic time (min)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eQuantity (gm)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eExtract quantity (gm)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e45\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.189 ± 0.01\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e45\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.217 ± 0.01\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e45\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.279 ± 0.02\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003eBased on the injected standards of phenols and flavonoids in HPLC, 13 compounds were appeared in the extract but with various quantities depending on the extraction pressure (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e and Figs.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e–\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). The concentration of eight compounds was increased with the increasing of extraction pressure while the concentration of four compounds was decreased with the increasing of extraction pressure. One compound namely rosmarinic acid was appeared in the extract at extraction condition of 200 and 300 with concentration 24.84 and 57.29 µg/g, respectively but didn’t appear in the extract at extraction condition of 100. For instance, Gallic acid increased from 77.17 to 203.93 to 461.52 µg/g, Caffeic acid increased from 87.87 to 137.81 to 216.83 µg/g, Syringic acid increased from 144.90 to 220.52 to 315.57µg/g, while Chlorogenic acid decreased from 345.20 to 216.64 to 242.36 µg/g, Rutin decreased from 139.98 to 144.98 to 45.00 µg/g, Methyl gallate decreased from 115.83 to 92.51 to 74.64 µg/g when the extraction pressure increased 100 to 200 to 300, respectively. Our findings indicated that pressure was effective for releasing the compounds from the extract. Moreover, the obtained results were matching with other investigation studied the SFE at different pressures. The maceration technique [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e], extraction through supercritical fluid [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e], and the use of Soxhlet extraction [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e] are the techniques most commonly employed for obtaining flavonoid and phenolic components from spices. The main determinants of the quantity of antioxidants obtained are the extraction process and time [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. In addition to pressure, other factors that can impact the yields obtained from extraction of molecules, which can be measured as total flavonoid and phenolic contents, include length of time and temperature [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e].\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePhenols and flavonoid compounds of in the extract of green cardamom at different pressure levels.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"10\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eDetected constituents\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c7\" namest=\"c5\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c10\" namest=\"c8\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eRT\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eArea\u003c/p\u003e \u003cp\u003e%\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eConc. (µg/g)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eRT\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eArea\u003c/p\u003e \u003cp\u003e%\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eConc. (µg/ml)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eRT\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eArea\u003c/p\u003e \u003cp\u003e%\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eConc. (µg/g)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3.52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4.13\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e77.17\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3.52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e10.07\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e203.93\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e3.52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e17.48\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e461.52\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eChlorogenic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4.23\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.71\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e345.20\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4.23\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.62\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e216.64\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e4.22\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e4.82\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e242.36\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMethyl gallate\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.51\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8.12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e115.83\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5.51\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5.98\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e92.51\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e5.51\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e3.70\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e74.64\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCaffeic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.79\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6.72\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e87.87\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5.80\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e9.71\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e137.81\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e5.80\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e11.72\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e216.83\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.66\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e144.90\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e6.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e13.55\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e220.52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e6.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e14.88\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e315.57\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eRutin\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.94\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.67\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e139.98\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e6.95\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.50\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e144.98\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e6.94\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.83\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e45.00\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCoumaric acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e27.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e254.42\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e8.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e28.06\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e278.96\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e8.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e21.73\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e281.54\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eVanillin\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.94\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.18\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e112.15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e8.95\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e12.40\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e114.72\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e8.94\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e10.85\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e141.94\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFerulic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.40\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e38.56\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e9.61\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.97\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e41.27\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e9.61\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e2.21\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e46.24\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNaringenin\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10.20\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.93\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e49.22\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e10.16\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.78\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e65.45\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e10.15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e2.33\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e77.62\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eRosmarinic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e11.74\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e11.83\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.92\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e24.84\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e11.83\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.64\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e57.29\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eQuercetin\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e17.24\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.67\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e23.06\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e17.55\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.22\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e38.72\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e17.54\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.42\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e63.83\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCinnamic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e19.10\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.04\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e54.56\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e19.13\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e6.40\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e34.31\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e19.14\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e6.37\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e44.52\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntimicrobail action (mm) \u0026amp; MIC and MBC (µg/ml) levels of green cardamom extracts upon using various pressure levels (100, 200, and 300 bar) (outcomes are represnted as means ± SD).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"8\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTested microbes\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003eZone of inhibition (mm)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c8\" namest=\"c6\"\u003e \u003cp\u003eMIC/MBC (µg/ml)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003estandard\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eB. subtilis\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e18.17 ± 0.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e20.83 ± 0.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e23.33 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e25.17 ± 1.26\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e125/500\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e31.25/62.5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e15.62/15.62\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e15.50 ± 0.87\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e17.83 ± 0.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e18.67 ± 1.53\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e16.33 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e125/250\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e62.5/62.5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e31.25/31.25\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eK. pneumoniae\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.67 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e15.83 ± 1.04\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e20.33 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e18.67 ± 1.53\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1000/--\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e125/250\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e15.62/31.25\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eS. typhi\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10.75 ± 0.43\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e16.33 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e14.67 ± 1.15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e---/--\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e250/500\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e250/500\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eC. albicans\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e14.33 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e19.33 ± 2.08\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e22.17 ± 1.04\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e21.83 ± 0.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e125/250\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e62.5/125\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e31.25/62.5\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP. glabrum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNA\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e28.33 ± 2.52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e---/--\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e---/--\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e---/--\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003eThe recorded data in table (3) and Figure (4) showed that the obtained extract at 100, 200, and 300 possess antimicrobial activity against \u003cem\u003eB. subtilis, S. aureus, K. pneumonia\u003c/em\u003e, and \u003cem\u003eC. albicans\u003c/em\u003e with various inhibition zones based on the extraction condition and tested microbe. The inhibition zones were 18.17 ± 0.29, 20.83 ± 0.29, and 23.33 ± 0.58 mm for \u003cem\u003eB. subtilis\u003c/em\u003e, 8.67 ± 0.58, 15.83 ± 1.04, and 20.33 ± 0.58 mm for \u003cem\u003eK. pneumonia\u003c/em\u003e, 14.33 ± 0.58, 19.33 ± 2.08, and 22.17 ± 1.04 mm for \u003cem\u003eC. albicans\u003c/em\u003e employing the extract at 100, 200, and 300, respectively. On the other hand the extract at 100 of the extraction condition faild to inhibit \u003cem\u003eS. typhi\u003c/em\u003e while exhibited 10.75 ± 0.43 and 16.33 ± 0.58 mm inhibition zones at 200, and 300 of extraction condition. The filamentous fungus \u003cem\u003eP. glabrum\u003c/em\u003e didn’t inhibited by the extract at any studied extraction condition. The recorded results were compared to the inhibitory action of standard antibiotic/ antifungal drugs. Furthermore, the measured MIC and MBC confirmed that extract efficacy increased with increasing extraction condition of pressure, where less MIC and MBC quantities at 300 were recorded than that recorded at 100 or 200 against tested microbes. Cardamom seeds add a sweet and tangy flavor. As an antimicrobial substance that treats dental cavities, it is frequently used as a breath freshener to preserve oral health [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. Numerous investigations on the extraction of naturally occurring non-antibiotic compounds with possible antibacterial effects—like SFE extracts—for the treatment of multidrug-resistant microbes have been published [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAntibiofilm activity of the extract was estimated against tested bacteria as depicted in (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). The biofilm formation was inhibited by the extract and increased with the increasing the extract concentration. At the same time the inhibition of biofilm formation was more affected by the extract at the extraction condition at 300 Bar than that by the extract at the extraction condition at 100 and 200. The biofilm of \u003cem\u003eB. subtilis\u003c/em\u003e was the most sensitive followed by \u003cem\u003eS. typhi, K. pneumonia\u003c/em\u003e, and \u003cem\u003eS. aureus\u003c/em\u003e where biofilm inhibition was 95.64 ± 0.98, 94.00 ± 1.22, 92.96 ± 0.66, and 92.32 ± 1.33%, respectively by the extract at extraction condition of 300 bar (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). A bacterial biofilm is a multidimensional framework made up of mono- or multi-microbial populations attached to an outer layer of cells that gives microorganisms resistance to challenging circumstances, antimicrobial agents, and the human immune system's strategies [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. Furthermore, Nassar et al. [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e] observed that microorganisms that cause a variety of human diseases and infections acquired in hospitals have demonstrated a biofilm origin. Accordingly, safe antibacterial substances that can hinder biofilm formation are needed to eradicate long-lasting infections linked to biofilms and lessen their detrimental effect on the health of people. The present investigation revealed that the raise of produced compounds upon elevation of the applied pressure enhance the antibiofilm impact of green cardamom. Investigation on shrub phenolics' antibiofilm properties has shown that, in addition to their harmful effects on bacteria, these compounds also have \"softer\" effects that minimize biofilms by interfering with bacterial regulating mechanisms like quorum recognition or other universal regulatory frameworks, all without influencing bacterial growth [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe ability of tested extract to inhibite the hemolysis in the presence of tested bacteria was measured (Figs.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e, \u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e). As the concentration of extract increase, the hemolysis inhibition % increased as well as the extract at extraction condition 300 was effective for hemolysis inhibition % compared to the extract at extraction condition 200 and 100 in all cases of tested bacteria. At 75% MIC of the extract at extraction conditions 100, 200, and 300, the hemolysis inhibition % was 92.53 ± 2.41, 93.53 ± 1.10, and 94.27 ± 0.46 in the presence of \u003cem\u003eS. aureus\u003c/em\u003e, 90.60 ± 3.11, 93.67 ± 1.76, and 94.80 ± 0.17 in the presence of \u003cem\u003eS. typhi\u003c/em\u003e, 89.37 ± 0.47, 93.53 ± 3.41, and 96.66 ± 2.52 in the presence of \u003cem\u003eB. subtilis\u003c/em\u003e, 90.33 ± 1.53, 93.70 ± 1.00, and 97.66 ± 2.08 in the presence of \u003cem\u003eK. pneumonia.\u003c/em\u003e Since hemolysin is the primary feature of \u003cem\u003eS. aureus\u003c/em\u003e strains' virulence, it has been suggested that antivirulence/antitoxin strategies ought to prioritize it [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e]. Flavonoids have been demonstrated to aggressively counteract the hemolytic properties of \u003cem\u003eS. aureus\u003c/em\u003e's α-hemolysin, disrupting the structure of the toxin [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]. An other way natural extracts exhibits antihemolytic effect is by strengthening the erythrocyte membrane's resistance to the toxin [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eVarious types of green cardamom extracts which prepared upon using different pressure values (100, 200, and 300 bar) towards different examined bacteria at different time points could be seen at (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). A daramatic reduction (\u003cem\u003eP\u003c/em\u003e ≤ 0.05) in bacterial count could be seen upon using green cardamom extract which prepared at 300 bar relaive to the other two forms of green cardamom extracts. The prepared cardamom extract at 100 bar showed a minimal impact towards \u003cem\u003eB. subtilis\u003c/em\u003e at different time points where 324 ± 12 bacterial cells could be detected after 180 minutes. There is no bacterial cells after exposing various bacteria for cardamom extract at 300 bar after 150 minutes revealing its effective impact at earlier examined time points. Time killing kinetics confirmed the bactericidal impact of cardamom extract especially at the prepared one at 300 bar. A few research investigations have been conducted on the time-kill kinetics of cardamom, while numerous studies of natural product extracts have been published [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e]. Killing examination revealed that the level of time-varying inhibition of microbes differed between bacteria and natural product extracts [\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e]. Plant-derived compounds and reactions following microbial infection may be taken into consideration when evaluating the antibacterial attributes [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e].\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eEffect of different Killing Kinetic times of green cardamom extracts upon using various pressure levels (100, 200, and 300 bar) towards \u003cem\u003eB. subtilis\u003c/em\u003e, \u003cem\u003eS. aureus\u003c/em\u003e, \u003cem\u003eK. pneumonia\u003c/em\u003e, and \u003cem\u003eS. typhi\u003c/em\u003e, outcomes are represnted as means ± SD).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eKilling Kinetic time (min)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"6\" nameend=\"c7\" namest=\"c2\"\u003e \u003cp\u003eCFU\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e \u003cp\u003e\u003cem\u003eB. subtilis\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c7\" namest=\"c5\"\u003e \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e26×10\u003csup\u003e5\u003c/sup\u003e ±12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e26×10\u003csup\u003e5\u003c/sup\u003e ±9\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e26×10\u003csup\u003e5\u003c/sup\u003e ±2.0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e29×10\u003csup\u003e5\u003c/sup\u003e±11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e29×10\u003csup\u003e5\u003c/sup\u003e±12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e29×10\u003csup\u003e5\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e30\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6×10\u003csup\u003e5\u003c/sup\u003e±7\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e56×10\u003csup\u003e4\u003c/sup\u003e±6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e287×10\u003csup\u003e3\u003c/sup\u003e±1\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e232×10\u003csup\u003e4\u003c/sup\u003e±9\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e22×10\u003csup\u003e4\u003c/sup\u003e±4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e15×10\u003csup\u003e4\u003c/sup\u003e±6\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e128×10\u003csup\u003e4\u003c/sup\u003e± 8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e28×10\u003csup\u003e3\u003c/sup\u003e± 11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e18×10\u003csup\u003e2\u003c/sup\u003e± 2\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e117×10\u003csup\u003e3\u003c/sup\u003e±7\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e79×10\u003csup\u003e3\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e23×10\u003csup\u003e2\u003c/sup\u003e±4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e120\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e184×10\u003csup\u003e3\u003c/sup\u003e ±11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e174×10 ± 4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e70×10 ± 3\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e13×10\u003csup\u003e2\u003c/sup\u003e ±6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e210 ± 6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e167 ± 3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e150\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25 ×10\u003csup\u003e2\u003c/sup\u003e ±12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e291 ± 3\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e380\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e21 ± 2\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e180\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e324 ± 12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eKilling Kinetic time (min)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colspan=\"6\" nameend=\"c7\" namest=\"c2\"\u003e \u003cp\u003eCFU\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e200\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e300\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e \u003cp\u003e\u003cem\u003eK. pneumoniae\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colspan=\"3\" nameend=\"c7\" namest=\"c5\"\u003e \u003cp\u003e\u003cem\u003eS. typhi\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e24×10\u003csup\u003e5\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e24×10\u003csup\u003e5\u003c/sup\u003e±12\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e24×10\u003csup\u003e5\u003c/sup\u003e±9\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5×10\u003csup\u003e5\u003c/sup\u003e±6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e5×10\u003csup\u003e5\u003c/sup\u003e±1\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e5×10\u003csup\u003e5\u003c/sup\u003e±4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e30\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e239×10\u003csup\u003e4\u003c/sup\u003e±6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e30×10\u003csup\u003e4\u003c/sup\u003e±14\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e13×10\u003csup\u003e4\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e221×10\u003csup\u003e4\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e94×10\u003csup\u003e4\u003c/sup\u003e±2\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e19×10\u003csup\u003e4\u003c/sup\u003e±11\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e157×10\u003csup\u003e3\u003c/sup\u003e±5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e17×10\u003csup\u003e3\u003c/sup\u003e±8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e213×10\u003csup\u003e2\u003c/sup\u003e±11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e242×10\u003csup\u003e3\u003c/sup\u003e±4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e142×10\u003csup\u003e3\u003c/sup\u003e±1\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e12×10\u003csup\u003e3\u003c/sup\u003e±14\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e120\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e176×10 ± 4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e250 ± 7\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e231\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e126×10\u003csup\u003e2\u003c/sup\u003e ±6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e15×10\u003csup\u003e2\u003c/sup\u003e ±2\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e200 ± 8\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e150\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e193 ± 3\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11 ± 4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e292\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e294\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e180\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003eThe green cardamom extract obtained by SFE extraction at 300 Bar caused numerous ultrastructural changes in \u003cem\u003eB. subtilis\u003c/em\u003e and \u003cem\u003eC. albicans\u003c/em\u003e cells in the current investigation. TEM images (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003eA) of the untreated B. subtilis specimen showed that each individual cell was healthy, rod-shaped, and encompassed by the outermost and innermost layers with no any damage to the cell. When B. subtilis cells were treated with green cardamom extract, their size drastically shrank and several lysed cells were visible (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003eB). TEM pictures (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003eC) showed that individual \u003cem\u003eC. albicans\u003c/em\u003e cells in the reference specimen were well, rounded, and classically shaped; however, when green cardamom was applied, the \u003cem\u003eC. albicans\u003c/em\u003e cells were destructed, resulting in deformed cells with a diminished shape (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003eD). To fight the microbes that cause different infections and illnesses, new antimicrobial substances had to be created [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. The TEM technique can provide valuable insight into the antibacterial mechanisms of action of new antibacterial substances to confirm the antimicrobial outcomes through visualization of various alterations upon treatments [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e \u003cb\u003eIn silico\u003c/b\u003e \u003cb\u003einvestigation though docking analysis\u003c/b\u003e\u003c/p\u003e\u003cp\u003eSyringic acid showed stronger binding (S = -4.27 to -4.64 for 3V8J; S = -4.69 to -5.04 for 5VX6) than gallic acid (S = -4.11 to -4.45 for 3V8J; S = -4.52 to -4.68 for 5VX6). Lower RMSD_refine values indicated stable poses. E_conf values reflecting conformational stability. E_place and E_refine values were consistently favorable.\u003c/p\u003e\u003cp\u003eKey Interactions for \u003cem\u003eB. subtilis\u003c/em\u003e (5VX6) indicated that gallic acid formed H-bonds with GLU 187 (2.73 Å, -5.4 kcal/mol) and ARG 172 (2.94 Å, -4.4 kcal/mol). Syringic acid interacted with GLU 187 (2.82 Å, -7.3 kcal/mol), ARG 172 (3.10 Å, -1.4 kcal/mol), and AR 104 (3.04 Å, -0.8 kcal/mol). In S. aureus (3V8J): Both acids bound to ASP 239 (Gallic acid: 2.85 Å, -5.4 kcal/mol; Syringic acid: 2.86 Å, -4.8 kcal/mol). The important docking metrics, such as docking scores, interaction types, binding energies, and distances, have been reviewed to determine binding affinities and interaction processes, as shown in Tables\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e5\u003c/span\u003e–\u003cspan refid=\"Tab8\" class=\"InternalRef\"\u003e8\u003c/span\u003e. The best-fitted diagrams of 2D and 3D poses select by the syringic acid and gallic acid are described (Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e10\u003c/span\u003e). Regarding the discussion of docking analysis, the results reveal nuanced differences in the binding behavior of gallic acid and syringic acid against Bacillus subtilis (5VX6) and \u003cem\u003eS. aureus\u003c/em\u003e (3V8J), offering insights into their potential as antimicrobial agents. The presence of methoxy groups in Syringic acid likely enhances its binding. While methoxy groups are not directly involved in hydrogen bonding, their electron-donating effects may subtly modulate the electronic environment of Syringic acid’s hydroxyl groups, enhancing their ability to form stable bonds with residues like GLU 187. In \u003cem\u003eB. subtilis\u003c/em\u003e (5VX6), syringic acid forms a stronger hydrogen bond with GLU 187 (-7.3 kcal/mol) compared to Gallic acid (-5.4 kcal/mol), likely due to optimal alignment of its hydroxyl groups. Additional interactions with ARG 172 and ARG 104 (Table\u0026nbsp;\u003cspan refid=\"Tab7\" class=\"InternalRef\"\u003e7\u003c/span\u003e) may stabilize its pose despite the absence of methoxy group participation.\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDocking scores and energies of gallic acid and syringic acid with structure of B. subtilis (PDB ID: 5VX6).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"8\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMol\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ermsd_refine\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eE_conf\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eE_place\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eE_score1\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eE_refine\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eE_score2\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.67921\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.7332464\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-31.8259\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-56.9843\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.70131\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-23.0596\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.67921\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.59409\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.74464136\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-32.8286\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-68.1666\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.2249\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-24.6589\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.59409\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.58793\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.7696532\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-33.2433\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-63.2359\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.91201\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-25.4289\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.58793\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.56066\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.6160221\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-31.7368\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-52.0508\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.3878\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-23.347\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.56066\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.51797\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.1812108\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-33.1922\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-47.8645\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.88048\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-21.0348\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.51797\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-5.03796\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.52116263\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-14.0667\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-78.0995\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.1364\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-28.6602\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-5.03796\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.94237\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.94439775\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-13.9608\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-70.5491\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.0975\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-25.0605\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.94237\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.92053\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.111726\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-5.60061\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-64.9127\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.0869\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-22.5365\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.92053\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.69705\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.4523685\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-13.7555\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-67.6678\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.1653\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-22.6829\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.69705\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.67621\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.2140613\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-7.97653\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-61.7188\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.9312\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-21.6721\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.67621\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab6\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 6\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDocking scores and energies of gallic acid and syringic acid with structure of \u003cem\u003eS. aureus\u003c/em\u003e (PDB ID: 3V8J).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"8\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMol\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ermsd_refine\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eE_conf\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eE_place\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eE_score1\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eE_refine\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eE_score2\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.45374\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.4267793\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-32.8569\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-50.573\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.60852\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-22.523\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.45374\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.28895\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.3745345\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-30.6581\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-53.7743\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.88156\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-19.5394\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.28895\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.19007\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.6599638\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-33.9026\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-41.8718\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-11.3473\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-17.8853\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.19007\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.18197\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.3183807\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-32.5074\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-69.8018\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-11.9495\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-17.2698\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.18197\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.11419\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.4595817\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-32.7511\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-55.7456\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.92507\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-15.8203\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.11419\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.64437\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.73497051\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-12.4447\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-60.9532\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.6667\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-23.4391\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.64437\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.52733\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.98655659\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-7.59219\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-61.9997\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.1858\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-19.1627\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.52733\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.31496\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.6934763\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-10.5394\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-60.394\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.1252\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-19.3342\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.31496\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.28199\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.5533125\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-10.5155\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-61.2405\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-9.95271\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-17.4618\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.28199\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-4.2695\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.3460429\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-10.8598\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-58.6044\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-10.1436\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-17.6905\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-4.2695\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab7\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 7\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eInteraction of gallic acid and syringic acid with structure of \u003cem\u003eB. subtilis\u003c/em\u003e (PDB ID: 5VX6).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMol\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLigand\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eReceptor\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eInteraction\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eDistance\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eE (kcal/mol)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 13\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eOE2 GLU 187 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-donor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.73\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-5.4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 13\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNH1 ARG 172 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-acceptor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.94\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-4.4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 23\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eOE2 GLU 187 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-donor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.82\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-7.3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 14\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNH1 ARG 172 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-acceptor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3.10\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-1.4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 22\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNE ARG 104 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-acceptor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3.04\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-0.8\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab8\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 8\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eInteraction of gallic acid and syringic acid with structure of \u003cem\u003eS. aureus\u003c/em\u003e (PDB ID: 3V8J).\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMol\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLigand\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eReceptor\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eInteraction\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eDistance\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eE (kcal/mol)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGallic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 17\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eOD2 ASP 239 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-donor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.85\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-5.4\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSyringic acid\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eO 23\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eOD2 ASP 239 (A)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eH-donor\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.86\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-4.8\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003e \u003cstrong\u003eSteric Compatibility\u003c/strong\u003e \u003c/p\u003e\u003cp\u003eDespite its larger size, Syringic acid’s structure appears better accommodated in the binding pockets, as reflected in lower RMSD_refine values (e.g., 0.52 Å in 5VX6), indicating stable pose retention post-refinement.\u003c/p\u003e\u003cp\u003e \u003cb\u003eEnergy Term Analysis\u003c/b\u003e indicated that gallic acid showed more favorable conformational energy (E_conf) values (-31.8 to -33.9 kcal/mol vs. Syringic acid’s -5.6 to -14.1 kcal/mol), suggesting its simpler structure adopts lower-energy conformations. However, syringic acid’s higher (less negative) E_conf may reflect conformational adjustments to maximize interactions, ultimately yielding stronger overall binding (S).\u003c/p\u003e\u003cp\u003e \u003cb\u003ePlacement and Refinement Energies\u003c/b\u003e: Syringic acid’s superior E_place (e.g., -78.10 kcal/mol in 5VX6) indicates efficient initial docking, while its moderate E_refine penalties (e.g., -28.66 kcal/mol) suggest minor adjustments to optimize interactions. Several investigators employed the molecular docking to document the activity of some phenolic and flavonoids against cancer cells, pathogenic bacteria, and yeasts [\u003cspan additionalcitationids=\"CR53 CR54 CR55\" citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e–\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e]. For example, catechin and gallic acid were docked with prostate cancer cells 3 PDB ID: 2Q7L with binding affinities − 5.27521 and − 6.3204 kcal/mol [\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e]. Rosmarinic acid was docked with \u003cem\u003eC. tropicalis\u003c/em\u003e (PDB ID: 6ZD6), \u003cem\u003eG. candidum\u003c/em\u003e (PDB ID: 6ISV), and \u003cem\u003eC. albicans\u003c/em\u003e (PDB ID: 1ZAP) with binding affinities − 6.8224 kcal/mol, -6.79996, and − 6.15839 kcal/mol, respectively [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. According to Alsalamah et al. [\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e], ellagic acid showed − 4.5145 kcal/mol and − 6.18615 kcal/mol, while chlorogenic acid showed − 5.69876 kcal/mol and − 7.84379 kcal/mol as docked with \u003cem\u003eG. candidum\u003c/em\u003e (4ZZT) and \u003cem\u003eC. albicans\u003c/em\u003e (4ZZT) proteins, respectively.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis is a novel investigation to extract a substantial quantity of phenolic components from green cardamom employing the SFE approach. The SFE apparatus's pressure has been adjusted to maximize both the yield and the quality of the extracted substances. Additionally, a large number of chemicals in extracted green cardamom have been found to have about 13 bioactive components using the HPLC method. These compounds are recognized to have both medicinal and effective antimicrobial properties. Remarkably, in contrast to the other conditions of treatment, the extracted output rose under high pressure (300 bar). High antimicrobial, antibiofilm, and antihemolytic properties were demonstrated by all green cardamom extracts; however, when contrasted with the other two pressures, the strongest activity was noted at 300 bar. Syringic and gallic acids exhibit promising binding to \u003cem\u003eB. subtilis\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e targets, with syringic acid showing superior affinity. The interactions with key catalytic residues (GLU 187, ASP 239) provide a mechanistic basis for inhibition.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003cstrong\u003e:\u003c/strong\u003e Not applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003cstrong\u003e:\u003c/strong\u003e Not applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability Statement:\u003c/strong\u003e The results from the current investigation are available from the corresponding author upon reasonable appeal.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of Interest:\u003c/strong\u003e The authors declare no conflicts of interest\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions:\u003c/strong\u003e S.A.A.\u0026nbsp;Investigation, writing—review and editing; M.I.A., and K.S.I., A.M., and T.M.A. Conceptualization and methodology. All authors agreed to the published version of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eACKNOWLEDGMENT\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors wish to appreciate the Deanship of Scientific Research at Imam Mohammad Ibn Saud Islamic University (IMSIU) for supported and funded the current study (Grant number grant number IMSIU-DDRSP2501)\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFUNDING\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;This work was supported and funded by the Deanship of Scientific Research at Imam Mohammad Ibn Saud Islamic University (IMSIU) (grant number IMSIU-DDRSP2501).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAbdelghany, T. M., Hassan, M. M., El-Naggar, M. A., \u0026amp; Abd El-Mongy, M. (2020). 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Chemical characterization, anticancer, antioxidant and anti-obesity activities with molecular docking studies of Pleurotus ostreatus biomass exposed to moist heat. \u003cem\u003eJournal of Food Measurement and Characterization\u003c/em\u003e\u003cem\u003e \u003c/em\u003e19, 2476\u0026ndash;2495 (2025). https://doi.org/10.1007/s11694-025-03125-9.\u003cspan dir=\"RTL\"\u003e\u0026rlm;\u003c/span\u003e\u003c/li\u003e\n\u003cli\u003eAlsalamah SA, Alghonaim MI, Jusstaniah M, Abdelghany TM, Anti-Yeasts (2023) Antioxidant and healing properties of Henna Pre-Treated by moist heat and molecular Docking of its major constituents, chlorogenic and ellagic acids, with \u003cem\u003eCandida albicans\u003c/em\u003e and \u003cem\u003eGeotrichum candidum\u003c/em\u003e proteins. Life 13(9):1839. https://doi.org/10.3390/life13091839\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bioresources-and-bioprocessing","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"biob","sideBox":"Learn more about [Bioresources and Bioprocessing](http://bioresourcesbioprocessing.springeropen.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/biob/default.aspx","title":"Bioresources and Bioprocessing","twitterHandle":"@SpringerOpen","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Super critical fluid extractor, cardamom, antimicrobial, antihemolytic","lastPublishedDoi":"10.21203/rs.3.rs-6480943/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6480943/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThere is broad approach in the search for resource-use strategies that are both economically viable and value-based is the extraction of active compounds from various plants. Supercritical fluid extraction (SFE), has become a popular technique for extraction significant plant-based compounds. Our investigation contrasted the yield, biological functions and phytochemical compositions of green cardamom extracts generated with SFE at 100, 200, and 300 bar of pressures. The maximal obtained weight was 0.279 gm upon applying 300 bar. There is a proportional elevation in the levels of most of phenolic compounds which detected using HPLC upon raising the pressure levels for extraction. Antimicrobial action of extract types towards test food-born microbes where the produced extract at 300 bar had the maximal antimicrobial action towards \u003cem\u003eBacillus subtilis\u003c/em\u003e, and \u003cem\u003eCandida albicans\u003c/em\u003e with inhibition zones 23.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.58, and 22.17\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04 mm, subsequently. Antibiofilm and hemolysis inhibition in presence of test microbes of various extract forms reveal the potential impact of raising pressure for extraction to enhance impact of extract to reach maximal level at 300 bar. Killing kinetics assay showed the gradual raise of the green cardamom extract versus test microbes at various time points where the extract at 300 bar had the best outcomes. Transmission electron microscopy visualized alterations in ultrastructure features of \u003cem\u003eB. subtilis\u003c/em\u003e and \u003cem\u003eC. albicans\u003c/em\u003e using the green cardamom extract produced at 300 bar. The molecular docking performance of the main constituents in green cardamom extracts gallic acid and syringic acid against \u003cem\u003eB. subtilis\u003c/em\u003e (PDB ID: 5VX6) and \u003cem\u003eS. aureus\u003c/em\u003e (PDB ID: 3V8J) using the Molecular Operating Environment (MOE) software was evaluated. The docking scores (S), RMSD_refine values, and energy terms (E_conf, E_place, E_score1, E_refine, E_score2) were analyzed to assess binding affinities. Key interactions, including hydrogen bonds, were identified, with distances and energies quantified. Syringic acid exhibited better binding (S = -4.27 to -5.04) compared to gallic acid (S = -4.11 to -4.68) across both targets. Interactions with residues like GLU 187 and ARG 172 in 5VX6, and ASP 239 in 3V8J, highlighted critical binding motifs. These results suggest both compounds as potential inhibitors for bacterial targets, warranting further experimental validation.\u003c/p\u003e","manuscriptTitle":"Innovative strategy for extraction of green cardamom via Super critical fluid extractor at different levels of pressure with its application against microorganisms in vitro and in silico","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-29 13:10:41","doi":"10.21203/rs.3.rs-6480943/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewerAgreed","content":"","date":"2025-04-25T12:40:42+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-04-25T11:19:02+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-04-25T04:14:18+00:00","index":"","fulltext":""},{"type":"submitted","content":"Bioresources and Bioprocessing","date":"2025-04-23T07:26:47+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bioresources-and-bioprocessing","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"biob","sideBox":"Learn more about [Bioresources and Bioprocessing](http://bioresourcesbioprocessing.springeropen.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/biob/default.aspx","title":"Bioresources and Bioprocessing","twitterHandle":"@SpringerOpen","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"c2af957a-14c5-4935-af3c-3fbbeb3bf57f","owner":[],"postedDate":"April 29th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-12-15T16:03:56+00:00","versionOfRecord":{"articleIdentity":"rs-6480943","link":"https://doi.org/10.1186/s40643-025-00951-z","journal":{"identity":"bioresources-and-bioprocessing","isVorOnly":false,"title":"Bioresources and Bioprocessing"},"publishedOn":"2025-12-08 15:57:03","publishedOnDateReadable":"December 8th, 2025"},"versionCreatedAt":"2025-04-29 13:10:41","video":"","vorDoi":"10.1186/s40643-025-00951-z","vorDoiUrl":"https://doi.org/10.1186/s40643-025-00951-z","workflowStages":[]},"version":"v1","identity":"rs-6480943","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6480943","identity":"rs-6480943","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}