Phytochemical analysis of the extract from berries of Schisandra chinensis Turcz. (Baill.) and its anti-platelet potential in vitro

Phytochemical analysis of the extract from berries of Schisandra chinensis Turcz. (Baill.) and its anti-platelet potential in vitro | 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 Article Phytochemical analysis of the extract from berries of Schisandra chinensis Turcz. (Baill.) and its anti-platelet potential in vitro Natalia Sławińska, Bogdan Kontek, Jerzy Żuchowski, Barbara Moniuszko-Szajwaj, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4346913/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Schisandra chinensis Turcz. (Baill.) is a dioecious vine, belonging to the Schisandraceae family. Itsberries show several beneficial activities, including cardioprotective, antioxidant, and anti-inflammatory. We examined the chemical content of the extract from S. chinensis berries, as well as its antiplatelet potential in washed human blood platelets and whole blood in vitro . We assessed effect of the extract on several hemostasis parameters, including thrombus formation in full blood, platelet activation and adhesion, and coagulation times. Moreover, we evaluated the cytotoxicity of the extract against blood platelets based on extracellular lactate dehydrogenase (LDH) activity. The most important constituents of the extract were dibenzocyclooctadiene lignans; schisandrin was the dominant compound. The extract inhibited thrombus formation, agonist-stimulated platelet activation and adhesion, and was not cytotoxic. These results suggest that S. chinensis berries can be used as a safe, natural supplement with anti-platelet properties. However, more studies are needed to determine their mechanisms of action and in vivo efficiency. Biological sciences/Biochemistry Biological sciences/Plant sciences blood platelet coagulation hemostasis Schisandra chinensis Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1. Introduction Edible berries are an important component of the human diet, serving as a source of nutrients and bioactive compounds. They contain a variety of phytochemicals that exhibit a wide range of biological properties, including cardioprotective activity (Olas, 2017 , 2018 , 2023 ; Sławińska et al., 2023 ; Sławińska & Olas, 2022 ). Schisandra chinensis Turcz. (Baill.) is a dioecious vine, belonging to the Schisandraceae family. It occurs naturally in the forests of south-eastern Siberia (Primorski, Amurski, Khabarovski regions, Sakhalin), north-eastern China, Korea, and Japan. Recently, Valíčková et al. ( 2023 ) have shown that S. chinensis has the potential to be used for the production of nutrient supplements due to its adaptogenic properties. The primary property of all adaptogens is to eliminate the effects of stress and adapt the body to unfavorable external conditions (Szopa et al., 2016 ). S. chinensis fruit, called "fruit of five flavors" (wǔwèizǐ), which results from the fact that the fruit's peel is sweet, the pulp is sour, the seeds are bitter and astringent, and the grain extract is salty, has been used in Chinese medicine to treat different disorders and diseases. Indigenous peoples of the Far-Eastern regions of Russia used it to reduce hunger, thirst, and exhaustion (Panossian & Wikman, 2008 ; Szopa et al., 2017 ). Importantly, S. chinensis berries also have cardioprotective potential (Chun et al., 2014 ; Olas, 2023 ), which is exerted through various molecular pathways. They can also help control oxidative stress, obesity, and inflammation (Lin et al., 2017 ; Su et al., 2022 ; Wu et al., 2017 ). In traditional Chinese medicine, S. chinensis is often used in the treatment of palpitation (Su et al., 2022 ). The 2020-year edition of Chinese Pharmacopoeia contains 100 prescriptions containing S. chinensis (K. Yang et al., 2022 ). Due to medicinal and health promoting properties of this plant, many food supplements made from S. chinensis fruit are available on the market. Different components of S. chinensis , including lignans (especially schisandrin, also called schizandrin, schisandrol A, or wǔwèizǐ sù A) play a significant role in the prophylaxis and treatment of cardiovascular diseases (CVDs), for example in myocardial infarction and hypertension. However, there are no experiments that focus on the response of various elements of hemostasis (including blood platelets, plasma, and others) to S. chinensis berries. Therefore, the present study examined the anti-platelet activity of the extract from S. chinensis berries in washed human blood platelets and human whole blood in vitro . Using washed platelets, we studied the activity of this extract on platelet adhesion to two adhesive proteins – collagen and fibrinogen. We also determined the anti-platelet activity of the tested extract in whole blood using the Total Thrombus-formation Analysis System (T-TAS) and flow cytometry. Moreover, we evaluated the cytotoxicity of the extract against blood platelets based on extracellular lactate dehydrogenase (LDH) activity. We also investigated the effect of S. chinensis berry extract on the parameters of hemostasis in plasma ( in vitro ), including coagulation times: thrombin time (TT), prothrombin time (PT), and activated partial thromboplastin time (APTT). The action of the extract from S. chinensis berries was compared to the extract from sea buckthorn ( Hippophae rhamnoides L.) berries, and a commercial product – Aronox (A ronia melanocarpa berry extract), which have documented anti-platelet properties (Olas et al., 2008 ; Skalski et al., 2020 , 2021 ). 2. Materials and methods 2.1. Reagents Methanol (HPLC grade) was purchased from Fisher Chemical (UK), heptane (for chromatography), and acetonitrile (LC-MS grade) were obtained from Merck (Darmstadt, Germany). Monoclonal antibodies for flow cytometry (CD61-PerCP, CD62P-PE, and PAC-1-FITC) were purchased from Becton Dickinson (New Jersey, USA). Phosphate buffered saline (PBS), fibrinogen, collagen, bovine serum albumin (BSA), tris(hydroxymethyl)aminomethane (Tris), 4-nitrophenyl phosphate, ADP, nicotinamide adenine dinucleotide (NADH), and thrombin were purchased from Merck (Darmstadt, Germany). NaCl, sodium citrate, and citric acid were purchased from POCH (Avantor performance materials, Gliwice, Poland). Reagents needed for coagulation time measurements were purchased from Kselmed (Grudziądz, Poland). All materials and chemicals for T-TAS were purchased from Bionicum Sp. z o.o. (Warsaw, Poland). Other reagents were purchased from commercial distributors and were of the highest available grade. 2.2. Plant material Crimson-red, ripe S. chinensis berries, growing in spike clusters on vines, were collected in mild-September 2022 in the park and palace settlement “Lower Garden” of the Czartoryski family in Pulawy (51°24’47.698” N21° 57’ 20.626” E). The plants were planted about 40 years ago and identified by the park’s curator, dr. Adam Wołk. After harvesting, the berries were frozen at -18°C, and subsequently lyophilized (Gamma 2–16 LSC, Christ, Osterode am Harz, Germany). The reference sample marked 1/09/2022 is located at the Institute of Soil Science and Plant Cultivation (Pulawy, Poland). 2.3. Preparation of the extract from S. chinensis berries Freeze-dried fruits (100 g) were soaked overnight in 80% methanol (v/v; 1.5 L) at ambient temperature, roughly homogenized using a kitchen blender, and subjected to a 15 min. sonication in an ultrasonic bath. The collected extract was centrifuged (3-16KL, Sigma, Osterode am Harz, Germany), and filtered using a glass filter funnel. The extraction procedure was repeated twice, with new portions of the solvent. The pooled extracts were concentrated in a rotary evaporator and defatted by liquid-liquid extraction with heptane. The defatted extract was rotary evaporated to remove organic solvents and reduce its volume, and freeze-dried. The extract (74.57 g) was subsequently cleaned up by SPE, to remove organic acids, sugars, and other highly polar constituents. It was dissolved in 0.1% formic acid in MilliQ water (v/v) and loaded onto a short C18 column (60 × 50 mm; Cosmosil 140C18-Prep, 140 µm). The column was washed with 0.1% formic acid, and the bound compounds were eluted with methanol. The eluate was rotary-evaporated, and the residue was freeze-dried to yield 7.61 g of the purified extract. 2.4. Phytochemical analysis of the extract from S. chinensis berries The composition of S. chinensis fruits was analyzed by LC-HRMS, using a Thermo Ultimate 3000RS (Thermo Fischer Scientific, Waltham, MS, USA) UHPLC system equipped with a corona-charged aerosol detector (CAD) and coupled with a Bruker Impact II HD quadrupole-time of flight (Q-TOF) mass spectrometer (Bruker Daltonics GmbH, Bremen, Germany). Samples were chromatographed on an ACQUITY UPLC HSS T3 column (2.1 × 150 mm, 1.8 µm), equipped with a pre-column, and maintained at 40˚C. The injection volume was 2 µL. The mobile phase A was 0.1% formic acid in MilliQ water, and the mobile phase B was acetonitrile containing 0.1% of formic acid. The flow rate was 0.400 mL min − 1 . The separation program started from a short isocratic elution with 5% B (0.5 min.), followed by a 31 min. gradient from 5 to 95% of solvent B. The elution was continued a t 95% B for 7 min., then the mobile phase composition returned to the initial conditions and the column was equilibrated with 5% B for 5 minutes. MS's analyses were performed in the negative and positive ion mode. MS's settings for negative ion mode: capillary voltage 3 kV; dry gas flow 6 L min − 1 ; dry gas temperature 200°C; nebulizer pressure 2.5 bar; collision RF 750 Vpp; transfer time 100 µs; prepulse storage time 10 µs. Collision energy was set automatically in the range from 2.5 to 80 eV, depending on the m/z of a fragmented ion. The scanning values ranged from m/z 80 to m/z 2000. The settings for positive ion mode: capillary voltage 4 kV; dry gas flow 6 L min − 1 ; dry gas temperature 200°C; nebulizer pressure 2.5 bar; collision RF 750 Vpp; transfer time 100 µs; prepulse storage time 10 µs. Collision energy was set automatically in the range from 2.5 to 50 eV. The scanning range was m/z 100–2000. Quantitative analyses of phenolic compounds were performed using an ACQUITY UPLC® chromatographic system (Waters Inc., Milford, MA, USA), equipped with a PDA detector, and coupled with a triple quadrupole mass detector (TQD; Waters). The S. chinensis extract was separated on an ACQUITY UPLC BEH C18 (2.1 × 100 mm, 1.7 µm; Waters) column. The column temperature was 50°C, the injection volume was 2.5 µL. Mobile phase was composed of Mill-Q water with 0.1% formic acid in (solvent A) and acetonitrile with 0.1% formic acid (solvent B); the elution program was as follows: 0.0-0.5 min., 7% B; 0.5-7.0 min., linear gradient 7–25% B; 7.0-16.9 min., linear gradient 25–75% B; 17.0–18.0 min., 95% B; 18.1–20.0 min., 7% B. The TQ mass spectrometer was operated in negative and positive ion mode. The MS settings in negative ion mode were as follows: capillary voltage 2.80 kV, cone voltage 45 V, source temperature 150°C, desolvation temperature 450°C, cone gas flow (N 2 ) 100 L h − 1 , desolvation gas (N 2 ) flow 800 L h − 1 . In positive ion mode: capillary voltage 3.10 kV, cone voltage 60 V, the remaining parameters were the same as those described above. The scanning range was m/z 100–1200. The content of anthocyanins (the peak integration at λ = 475 nm; y = 232.90079x + 82.94488, R² = 0.9989), flavonoids (the peak integration at λ = 350 nm; y = 442.56566x − 46.00717, R² = 0.9999), and lignans (the peak integration at λ = 255 nm; y = 607.19720x + 131.25948, R² = 0.9999) was calculated on the basis of calibration curves, and was expressed as cyanidin, rutin and schisandrin equivalents, respectively. 2.5 Preparation of stock solution of the extract from S. chinensis berries, the extract from sea buckthorn berries, and the extract from aronia berries The extract from S. chinensis berries and the extract from sea buckthorn berries were dissolved in 50% DMSO (a universal solvent for many plant substances), however the final concentration of DMSO in the tested human blood platelets, human plasma, and human blood was below 0.05% ( v / v ). The addition of a low concentration of DMSO to blood platelets, plasma, and blood has no effect on the hemostatic properties of blood platelets, plasma, and full blood (data not presented). The extract from sea buckthorn berries contained predominantly various flavonol glycosides, mainly isorhamnetin glycosides and acylated glycosides (Skalski et al., 2021 ). A stock solution of commercial product – Aronox (by Agropharm Ltd., Poland; batch No 020/2007k, A. melanocarpa berry extract) was prepared in H 2 O at a concentration of 5 mg/mL. 2.6. Blood, plasma, and blood platelet samples Whole human blood was collected at “Diagnostyka” blood collection center (Brzechwy 7A, Lodz, Poland). All donors were non-smoking, healthy volunteers, which did not drink alcohol or take medication and anti-platelet supplementation for two weeks before blood collection. Peripheral blood was drawn into tubes with benzylsulfonyl-D-Arg-Pro-4-amidinobenzylamide (BAPA) (for T-TAS) or with citrate/phosphate/dextrose/adenine (CPDA) anticoagulant (for the remaining assays). The analysis of blood samples was performed according to the guidelines of the Helsinki Declaration for Human Research. Informed consent form was signed by each participant one day prior to blood collection. Procedures were conducted with the consent of Bioethics Committee at the University of Łódź (2/KBBN-UŁ/III/2014). Blood for the T-TAS assay was used within 2 hours after collection. For coagulation times measurements, plasma was separated from whole blood by differential centrifugation (2800 x g, 20 min., at room temperature). Blood platelets were isolated from fresh blood by differential centrifugation as described previously (Lis et al., 2019 ). The required number of platelets (2.0 × 10 8 /mL) was confirmed by a spectrophotometric measurement in a UV-Visible Helios-α at 800 nm. In every experiment, blood, plasma, or blood platelets were incubated for 30 minutes at 37°C, either with the extract from S. chinensis berries at final concentrations of 0.5–50 µg/mL, or the extract from sea buckthorn berries or aronia berries at final concentration 50 µg/mL. 2.7. Blood platelet adhesion Blood platelet adhesion was measured based on the activation of exoenzyme acid phosphatase in the blood platelets according to the method described by (Bellavite et al., 1994 ). 96-well plates were coated with 100 µg/mL fibrinogen or 0.04 µg/mL collagen. To achieve this, 100 µL of fibrinogen or collagen was added to the wells, and incubated for 24 h at 4°C, on an orbital shaker. Afterward, the wells were washed three times with TBS (pH 7.5), and 200 µL of 1% BSA was added. The plates were incubated with BSA at 37°C for 2 h. Meanwhile, the extracts were added to washed blood platelets at final concentrations of 0.5–50 µg/mL. A control sample was set up containing only blood platelets with Barber’s buffer (a modified Tyrode’s buffer); this value was assumed to be 100%. The samples were incubated for 30 min. at 37°C. BSA was removed, and the wells were washed three times with TBS (pH 7.5) with 0.1 mM CaCl 2 and 0.1 mM MgCl 2 . The samples were added to the wells in triplicate. Agonists (1 U/mL thrombin or 30 µM ADP) or TBS were added to appropriate wells. Then, the plates were incubated for 1h at 37°C. Afterward, the wells were washed three times with PBS. 150 µL of 0.1 M citrate buffer (pH 5.4) with 5 mM p -nitrophenyl phosphate and 0.1% Triton X-100 was added; the samples were incubated for 1 h at room temperature. Lastly, 100 µL of 2M NaOH was added to the wells. Absorbance was read at 405 nm with SPECTROstar Nano Microplate Reader (BMG LABTECH, Germany 2.8. Flow cytometry analysis Platelet activation was assessed based on platelet-derived microparticles (PMP) formation, and the exposition of P-selectin (CD62P) and the active form of GPIIb/IIIa on the surface of blood platelets. To measure the exposition of P-selectin and the active form of GPIIb/IIIa, full blood was mixed with the extracts at the concentrations of 0.5–50 µg/mL and incubated for 15 min. at 37°C. After 15 minutes, agonists were added to the samples (10 µM ADP, 20 µM ADP, or 10 µg/mL collagen), and the incubation continued for another 15 min. (at 37°C). After the incubation, the samples were diluted 10 times with PBS and incubated with antibodies (CD61-PerCP, CD62P-PE, and PAC-1-FITC, 3 µL of each antibody incubated with 10 µL of diluted samples). The samples were stained for 30 minutes in the dark, at room temperature. Afterward, the samples were fixed with 1% CellFix for 1 h at 37°C. The samples were analyzed with a LSR II Flow Cytometer (Becton Dickinson, San Diego, CA, USA); at least 5000 CD61-PerCP-positive objects were recorded. Blood platelets were gated based on an FSC/SSC (forward scatter/side scatter) plot and positive staining with CD61-PerCP. Then, the percent of CD62-PE and PAC-1-FITC positive platelets was calculated in each of the samples. To measure the formation of platelet-derived microparticles, the samples were incubated with S. chinensis extract for 15 minutes at 37°C. Then, 20 µg/mL collagen was added, and the incubation continued for another 15 minutes. The samples were diluted 10 times with PBS and stained with 2 µL of CD61-PE antibody for 30 minutes in the dark, at room temperature. The samples were fixed with 1% CellFix for 1 hour and analyzed with LSR II Flow Cytometer (Becton Dickinson, San Diego, CA, USA). At least 10000 CD61-PE-positive objects were recorded. PMPs were distinguished from CD61-PE-positive objects based on an FSC/SSC plot on a log/log scale. The analysis was carried out with Floreada software ( https://floreada.io , accessed on 26.02.2024). 2.9. Total Thrombus-formation Analysis System (T-TAS) (PL-chip) The T-TAS system can measure the thrombus formation process in semi-physiological conditions. Here, the PL-chip which measures only primary hemostasis was used. The results were recorded as AUC 10 (Area Under the Curve) - area under the flow pressure curve recorded for 10 min. after the start of the test. The AUC 10 depicts the growth, intensity, and stability of thrombus formation. Data was depicted as % of control. Further information about the assay can be found in (Hosokawa et al., 2011 ). First, the extracts were incubated with human full blood at final concentrations of 0.5–50 µg/mL (37°C, 30 minutes). A control sample with 0.9% NaCl instead of the extract was set up. Then, 320 µL of each sample was added to the PL-chip, and pressure was recorded for 10 minutes or until it reached 60 kPa (Time of Occlusion). The results were depicted as % of the control sample. 2.10. Measurement of prothrombin time, thrombin time, and activated partial thromboplastin time Coagulation times were determined coagulometrically, according to the method described by ( Malinowska et al., 2012 ). The extract was incubated with human plasma at 37°C for 30 minutes, at final concentrations of 0.5–50 µg/mL. The measurements were carried out on an Optic Coagulation Analyser, model K-3002 (Kselmed, Grudziadz, Poland). Each sample was measured in duplicate. 2.11. Activity of LDH The toxic effect of the extract from S. chinensis berries on human blood platelets was analyzed by measuring the activity of lactate dehydrogenase (LDH), an enzyme released from damaged blood platelets (Wróblewski & Ladue, 1955 ). Blood platelets were incubated with the extract at final concentrations of 0.5–50 µg/mL, at 37°C for 30 min. Afterward, the samples were centrifuged (2500 rpm, 15 min., 25°C) and 10 µl of the supernatant was added to the wells of a 96-well plate (in triplicate). Then, 0.1 M phosphate buffer (pH 7.4, 270 µL) and 0.25% nicotinamide adenine dinucleotide (NADH) (10 µl) were added to each well. A blank which contained 280 µL of phosphate buffer and 10 µL of NADH was set up as well. The samples were mixed and incubated for 20 minutes at room temperature. Lastly, 0.25% pyruvate (10 µL) was added to the samples. The samples were immediately mixed, and the absorbance was read at 1 minute intervals for 10 minutes, at 340 nm. 2.12. Statistical analysis Statistical analysis was performed with Statistica 10 (StatSoft 13.3, TIBCO Software Inc. Palo Alto, CA, USA). The distribution of data was checked by the Shapiro-Wilk test, and the homogeneity of variance by Levene’s test. Differences within and between groups were assessed with one-way ANOVA followed by Tukey’s test, or Kruskall-Wallis test. Results were presented as means ± SD. The results were considered significant at p ≤ 0.05. Dixon’s Q-test was used to eliminate uncertain data. 3. Results 3.1. Chemical characteristic of the extract from S. chinensis berries The extract contained many lignans, which were the major phenolic constituents (and more generally, main specialized metabolites) of the S. chinensis fruit extract (Table 1 ; Fig. 1 ). Among them, schisandrin (tentatively identified) was the dominant compound. The red color of the extract can be attributed to the presence of cyanidin- Pen-Hex-dHex. S. chinensis fruits also contained lesser amounts of (epi)catechin, dimeric and trimeric B-type proanthocyanidins, quercetin hexoside-deoxyhexoside, and quercetin hexoside. Apart from phenolics, small amounts of highly oxygenated nortriterpenoids (mainly), bisnortriterpenoids, triterpenoids, and homotriterpenoids were also detected (Table 1 ). Moreover, the extract contained also very high amounts of diverse highly polar constituents (Fig. 1 ). Table 1 Specialized metabolites of the S. chinensis fruit extract. No. tR (min.) parent ion ( m/z ) CID delta (ppm) mσ formula tentative identification Ref. 1–3 0.85–1.80 mixtures of polar compounds 4 5.77 203.0823 − 186.0552 (5), 159.0921 (7), 142.0641 (7), 116.0595 (17) 1.3 10.2 C 11 H 12 N 2 O 2 tryptophan 5 7.45 577.1352 − 407.0773 (90), 289.0720 (100), 245.0821 (23) -0.2 8.0 C 30 H 26 O 12 (epi)C-(epi)C 6 7.83 289.0719 − 289.0720 (100), 245.0819 (73), 203.03715 (31) -0.3 1.7 C 15 H 14 O 6 (epi)catechin 7 7.90 865.1982 − 695.1400 (4), 525.0822 (15), 407.0785 (83), 289.0729 (100), 243.0309 (39) 0.4 15.5 C 45 H 38 O 18 (epi)C-(epi)C-(epi)C 8 9.30 725.1927 − 339.0514 (14), 284.0329 (100) 1.0 7.2 C 32 H 39 O 19 cyanidin-Pen-Hex-dHex 1 9 13.23 441.1769 − 441.1769 (78), 397.1854 (28), 330.1319 (100), 217.1227 (30) -0.6 9.3 C 21 H 30 O 10 10 13.45 609.1460 − 300.0279 (100) 0.2 15.4 C 27 H 30 O 16 rutin 11 13.87 463.0882 − 300.0276 (100) 0.1 13.2 C 21 H 20 O 12 Q-3- O -Hex 12 16.40 685.2927 − * 639.2851 (12), 477.2348 (24), 383.1193 (100), 323.0979 (10), 263.0774 (15), 221.0664 (31), 179.0557 (28) -0.3 8.4 C 28 H 48 O 16 unidentified bisnorterpenoid 13 17.09 605.2232 − * 559.2182 (100), 497.2176 (65), 439.1759 (45), 351.1965 (31), 317.1391 (22) 1.3 12.2 C 29 H 36 O 11 unidentified nortriterpenoid 14 17.52 575.2131 − 575.2138 (100), 557.2020 (20), 531.2235 (3), 455.1728 (10) 0.5 8.9 C 29 H 36 O 12 unidentified nortriterpenoid 15 17.74 559.2179 − 559.2209 (100), 541.2087 (65), 523.2031 (50), 465.1557 (47), 447.1440 (55), 439.1785 (70), 421.1656(81) 1.1 8.0 C 29 H 36 O 11 unidentified nortriterpenoid 16 18.22 605.2239 − * 559.2181 (100), 497.2173 (88), 479.2080 (43), 439.1762 (41), 351.1961 (42), 317.1391 (34) 0.1 7.3 C 29 H 36 O 11 unidentified nortriterpenoid 17 18.45 559.2189 − 559.2196 (100), 541.2082 (19), 465.1531 (5), 445.1495 (6), 439.1760 (7) -0.7 3.2 C 29 H 36 O 11 unidentified nortriterpenoid 18 18.85 621.2194 − * 575.2142 (100), 557.2041 (28), 511.2317 (8), 455.1724 (9), 417.1558 (12) -0.9 14.6 C 29 H 36 O 12 unidentified nortriterpenoid 19 19.69 605.2241 − * 559.2188 (100), 541.2083 (14), 445.1504 (6), 347.1493 (3) -0.3 9.1 C 29 H 36 O 11 unidentified nortriterpenoid 20 19.99 589.2296 − * 543.2244 (47), 525.2134 (60), 507.2025(100), 481.2219 (60), 463.2128 (89), 419.2235 (94), 383.1863 (43) -0.9 15.9 C 29 H 36 O 10 unidentified nortriterpenoid 21 20.24 605.2242 − * 559.2184 (100), 541.2082 (40), 515.2278 (32), 497.2186 (49) -0.4 1.6 C 29 H 36 O 11 unidentified nortriterpenoid 22 20.62 591.2453 − * 545.2415 (2), 447.2161 (20), 411.1817 (25), 367.1917 (100)349.1815 (22) -1.0 11 C 29 H 38 O 10 unidentified nortriterpenoid 23 20.7 605.2240 − * 559.2186 (100), 497.1289 (14), 457.1862 (19) 0.0 30.1 C 29 H 36 O 11 unidentified nortriterpenoid 24 21.38 589.2295 − * 543.2245 (100), 525.2143 (46), 499.2354 (38), 481.2238 (77), 407.1863 (26) -0.7 4.3 C 29 H 36 O 10 unidentified nortriterpenoid 25 21.46 593.2603 − * 547.2556 (6), 529.2449 (25), 511.2336 (14), 485.2538 (16), 467.2442 (100), 427.2138 (14) 0.1 12.2 C 29 H 40 O 10 unidentified nortriterpenoid 26 22.83 577.2292 − * 531.2238 (100), 513.2118 (11), 451.2120 (16), 297.1485 (9) -0.3 12.8 C 28 H 36 O 10 wuweizidilactone H / isomer 2 27 23.87 603.208 − * 557.2037 (100)481.1507 (17), 455.1716 (55), 437.1604 (13), 365.1393 (7) 0.5 10.4 C 29 H 34 O 11 unidentified nortriterpenoid 28 24.17 579.2447 − * 533.2387 (100), 497.2195 (29), 475.2005 (35), 453.2270 (28), 435.2166 (24) 0.0 13.1 C 28 H 38 O 10 unidentified bisnorterpenoid 29 24.6 619.2757 − * 573.2733 (1), 531.2596 (100), 513.2490 (16), 495.2399 (8), 211.0975 (9) 0.5 10.5 C 31 H 42 O 10 unidentified homotriterpenoid 30 24.79 633.2554 − * 587.2515 (1), 509.2182 (29), 491.2076 (13), 465.2284 (19), 447.2180 (100), 429.2077 (21) -0.2 6.6 C 31 H 40 O 11 unidentified homotriterpenoid 31 24.79 603.2088 − * 557.2032 (100), 499.1618 (10), 481.1509 (18), 455.1707 (56), 437.1600 (13) 0.0 17.6 C 29 H 34 O 11 unidentified nortriterpenoid 32 24.89 635.2710 − * 589.2654 (4), 571.2546 (21), 509.2544 (100), 491.2440 (14), 467.2435 (54), 449.2329 (34) -0.1 11.9 C 31 H 42 O 11 unidentified homotriterpenoid 33 25.36 575.2476 + 557.2372 (11), 497.2162 (27), 479.2057 (69)461.1951 (33), 437.1953 (51), 419.1846 (100) 1.9 8.9 C 30 H 38 O 11 unidentified triterpenoid 34 25.47 587.2126 − * 541.2071 (100), 483.1655 (16), 465.1552 (20), 439.1759 (34), 365.1394 (12), 215.0715 (4) 1.4 3.9 C 29 H 34 O 10 unidentified nortriterpenoid 35 26.75 577.2638 + 559.2533 (46), 541.2434 (29), 499.2316 (49), 481.2218 (100), 463.2107 (53), 439.2112 (66), 421.2004 (95) 0.9 9.2 C 30 H 40 O 11 unidentified triterpenoid 36 27.02 619.2764 − * 573.2706 (71), 531.2591 (100) -0.6 8.8 C 31 H 42 O 10 unidentified homotriterpenoid 37 28.12 433.2218 + 415.2112 (100) 0.6 10.7 C 24 H 32 O 7 schisandrin 3 , 4 , 5 38 28.93 701.2813 − * 531.2215 (66), 513.2110 (100), 495.2043 (55), 477.1932 (32), 451.2151 (41), 433.2003 (64), 415.1912 (66) 0.2 10.5 C 35 H 44 O 12 unidentified 39 29.13 389.1960 + 389.1963 (100), 357.1699 (7), 319.1180 (3), 287.0920 (3) -0.4 1.7 C 22 H 28 O 6 gomisin J / isomer 3 , 5 40 29.40 417.1911 + 399.1804 (100), 369.1694 (13) -0.8 7.3 C 23 H 28 O 7 schisandrol B / isomer 3 , 4 , 5 41 30.03 391.2116 + 391.2115 (100), 237.1485 (16) -0.2 2.1 C 22 H 30 O 6 pregomisin / isomer 5 42 30.03 501.2482 + 483.2379 (65), 401.1961 (100) 0.1 11.4 C 28 H 36 O 8 angeloylgomisin H / tigloylgomisin H / isomer 3 , 4 , 5 43 30.38 501.2483 + 483.2377 (26), 401.1960 (100) 0.1 5.5 C 28 H 36 O 8 angeloylgomisin H / tigloylgomisin H/ isomer 3 , 4 , 5 44 30.46 523.2321 + 505.2221 (100), 401.1960 (49) 1.0 2.4 C 30 H 34 O 8 benzoylgomisin H / isomer 5 45 30.67 548.2850 +# 431.2063 (100) 0.7 7.9 C 29 H 38 O 9 angeloylgomisin Q / tigloylgomisin Q / isomer 4 , 5 46 30.70 554.2387 +# 415.1754 (100) -0.5 12.2 C 30 H 32 O 9 schisantherin A / gomisin G 3 , 4 , 5 47 30.99 532.2542 +# 415.1758 (100) -0.1 18.1 C 28 H 34 O 9 schisantherin B / isomer 3 , 4 , 5 48 31.12 532.2542 +# 415.1755 (100), 371.1491 (7) -0.2 3.0 C 28 H 34 O 9 gomisin F / angeloylgomisin P / tigloylgomisin P 3 , 5 49 31.32 403.2118 + 403.2120 (100), 371.1857 (6) -0.8 8.1 C 23 H 30 O 6 schisanhenol / isomer 3 , 5 50 32.08 417.2275 + -0.8 4.7 C 24 H 32 O 6 schisandrin A (deoxyschisandrin) / isomer 3 , 4 , 5 51 32.30 401.1962 + 401.1964 (100), 331.1183 (5) -0.9 2.1 C 23 H 28 O 6 gomisin N / isomer 3 , 5 52 32.40 401.1962 + 401.1965 (100), 331.1183 (5) -0.9 5.7 C 23 H 28 O 6 schisandrin B (γ-schisandrin) / isomer 3 , 4 , 5 53 32.49 385.1649 + 385.1648 (100), 355.1544 (5), 315.0871 (5), 285.0765 (3) -0.9 5.5 C 22 H 24 O 6 schisandrin C / isomer 3 , 5 54 32.52 485.2536 + 485.2538 (100), 403.2116 (32) -0.4 7.3 C 28 H 36 O 7 unidentified 55 33.31 279.2326 − 279.2326 (100) 1.1 14.3 C 18 H 32 O 2 octadecadienoic acid − - negative ion mode; + - positive ion mode; * - formic acid adduct; # - NH 4 + adduct; C – catechin, Q – quercetin, dHex – deoxyhexose; Hex – hexose; Pen – pentose 1 . Ma et al., 2012 ; 2 . Li et al., 2017 ; 3 . He et al., 1997 ; 4 . Yang et al., 2011 ; 5 . Liu et al., 2013 Results of the quantitative analysis of phenolic compounds are shown in Table 2 . As mentioned above, lignans were dominant phenolic compounds of the extract from S. chinensis fruit. The total lignan content was 29.61 ± 0.18 mg g -1 of the extract (expressed as schisandrin equivalent). The total flavonoid content was 1.64 ± 0.03 mg g -1 (expressed as rutin equivalent), and the content of anthocyanidins was 17.08 ± 0.35 mg g -1 (expressed as cyanidin equivalent). Table 2 Content of major phenolic compounds in the extract from the fruit of S. chinensis . tR (min.) m/z * identification content (mg g − 1 ) 2.21 727 cyanidin-Pen-Hex-dHex 17.08 ± 0.35 $ 4.17 611 rutin 1.12 ± 0.02 4.32 465 quercetin-Hex 0.52 ± 0.01 # 11.39 433 schisandrin 23.07 ± 0.31 11.86 389 gomisin J traces 11.99 417 schisandrol B 1.16 ± 0.01 ^ 12.97 501 angeloylgomisin H / tigloylgomisin H 1.61 ± 0.02 ^ 15.81 401 schisandrin B 2.12 ± 0.11 ^ 16.15 385 schisandrin C 0.84 ± 0.06 ^ * - positive ion mode; $ - cyanidin equivalent; # - rutin equivalent; ^ - schisandrin equivalent; Hex - hexose; dHex - deoxyhexose; Pen - pentose; 3.2. Effect of the extract from S. chinensis berries on the adhesion of washed blood platelets to collagen and fibrinogen The anti-adhesive activity of the extract was studied using human washed blood platelets. The results were presented as percent of adhesion of the control samples. As shown in Fig. 2 A and B the level of adhesion of resting blood platelets and adhesion of thrombin-activated platelets was significantly reduced in the presence of all used concentrations of the extract from S. chinensis berries (0.5–50 µg/mL). For example, at the highest concentration (50 µg/mL), the extract inhibited the adhesion of thrombin-activated blood platelets by about 50% in comparison with control (Fig. 2 B). On the other hand, none of the tested concentrations of the extract (0.5–50 µg/mL) demonstrated anti-adhesive properties, when adhesion of thrombin-activated platelets to fibrinogen was measured (Fig. 2 C). For the ADP-activated blood platelets, three tested concentrations of the extract from S. chinensis berries (0.5, 1, and 5 µg/mL) did not have anti-adhesive activity (Fig. 2 D). Reduced adhesion was only observed for the two highest concentrations (10 and 50 µg/mL) (Fig. 2 D). 3.3. Effect of the extract from S. chinensis berries on parameters of blood platelet activation measured with flow cytometry in whole blood Figures 3 and 4 demonstrate the parameters of activation of platelets stimulated with 10 and 20 µM ADP, and 10 µg/mL collagen (measured with flow cytometry). Changes in platelet activation were noted in whole blood treated with the extract from S. chinensis berries at all tested concentrations (0.5–50 µg/mL), but these changes were not always statistically significant (Figs. 3 and 4 ). For example, the extract from S. chinensis berries (at all used concentrations: 0.5–50 µg/mL) inhibited the exposition of the active form of GPIIb/IIIa on the surface of blood platelets stimulated with 10 µM ADP (Fig. 3 A). In blood platelets stimulated with 20 µM ADP, statistically significant reduction of the exposition of the active form of GPIIb/IIIa was observed only for the highest concentration of the extract (50 µg/mL) (Fig. 3 B). On the other hand, none of the concentrations of the extract (0.5–50 µg/mL) significantly impacted the exposition of GPIIb/IIIa on platelets stimulated by 10 µg/mL collagen, and the exposition of P-selectin on the surface of blood platelets stimulated by 10 and 20 µM ADP (Fig. 3 C, 4 A and B). However, in platelets activated by collagen, significant inhibition of the exposition of P-selectin was observed for three concentrations of the extract (5, 10, and 50 µg/mL) (Fig. 4 C). Moreover, none of the used concentrations (0.5–50 µg/mL) of the extract from S. chinensis berries significantly impacted platelet-derived microparticle formation and the exposition of GPIIb/IIIa and P-selectin on resting platelets (data not presented). 3.4. Effect of the extract from S. chinensis berries on thrombus formation with T-TAS (PL-chip) in whole blood Only the highest concentration of the extract from S. chinensis berries (50 µg/mL) significantly decreased the AUC 10 value measured by T-TAS in whole blood (Fig. 5 ). 3.5. Effect of the extract from S. chinensis berries on coagulation parameters (PT, TT, and APTT) in plasma The analysis of the effect of the extract from S. chinensis berries on the coagulation times of human plasma showed that none of the used concentrations of the extract (0.5–50 µg/mL) changed APTT, PT, and TT (data not demonstrated). 3.6. Effect of the extract from S. chinensis berries on a marker of blood platelet damage To determine the toxic effect of the extract on blood platelets, the level of LDH activity was measured. The results demonstrate no significant difference in platelet viability after exposure to S. chinensis berry extract at all used concentrations (0.5–50 µg/mL) (data not presented). Table 3 compares the effects of the extract from S. chinensis berries, the extract from sea buckthorn berries, and the extract from aronia berries (as a positive control) (50 µg/mL) on selected markers of blood platelet activation. The strongest anti-platelet activity was demonstrated by the extract from S. chinensis berries and the extract from aronia berries. The anti-platelet potential was observed for six markers. Table 3 A comparison of the anti-platelet activity of the extract from S. chinensis berries (50 µg/mL), the extract from sea buckthorn berries (50 µg/mL), and the extract from aronia berries (50 µg/mL) in washed blood platelets (measured by adhesion to adhesive proteins (collagen and fibrinogen) and in whole blood (measured by T-TAS and flow cytometry). S. chinensis berries Sea buckthorn berries Aronia berries Inhibition of adhesion of thrombin-activated platelet to collagen Anti-platelet activity No effect Anti-platelet activity Inhibition of adhesion of thrombin-activated platelet to fibrinogen No effect No effect No effect Inhibition of adhesion of ADP-activated platelet to fibrinogen Anti-platelet activity Anti-platelet activity No effect Inhibition of thrombus formation (measured by T-TAS) Anti-platelet activity Anti-platelet activity No effect Inhibition of GPIIb/IIIa exposition in 10 µM ADP-activated platelets Anti-platelet activity No effect Anti-platelet activity Inhibition of GPIIb/IIIa exposition in 20 µM ADP-activated platelets Anti-platelet activity No effect Anti-platelet activity Inhibition of GPIIb/IIIa exposition in collagen-activated platelets No effect No effect Anti-platelet activity Inhibition of P-selectin exposition in 10 µM ADP-activated platelets No effect Anti-platelet activity No effect Inhibition of P-selectin exposition in 20 µM ADP-activated platelets No effect Anti-platelet activity Anti-platelet activity Inhibition of P-selectin exposition in collagen-activated platelets Anti-platelet activity No effect Anti-platelet activity 4. Discussion Our LC-HRMS analysis showed that S. chinensis berry extract contained diverse lignans, which were its most important phenolic constituents. Most of these tentatively identified compounds were dibenzocyclooctadiene lignans, except for pregomisin, which belongs to the dibenzylbutane type. The presence of dibenzocyclooctadiene lignans is a characteristic feature of Schisandraceae family; they are regarded as the main bioactive constituents of S. chinensis (S. Yang & Yuan, 2021 ). Schisandrin was the dominant compound; putative schisandrol B, angelogomisin H, deoxyschisandrin, and schisandrin B were other major lignans. Our results are mostly in line with other studies (He et al., 1997 ; Liu et al., 2013 ; Sheng et al., 2022 ; Yang et al., 2011 ). The red color of S. chinensis berries is caused by an anthocyanin, cyanidin- Pen-Hex-dHex. According to the literature data, this compound is most probably cyanidin 3- O -xylosylrutinoside (Liao et al., 2016 ; Ma et al., 2012 ). Flavonoids, represented rutin and quercetin hexoside, occurred in small quantities. Our results are supported by the work of (Mocan et al., 2014 ), who found small amounts of rutin, quercetin 3- O -glucoside, and quercetin 3- O -galactoside in S. chinensis berries. Apart from phenolics, the extract also contained numerous terpenoid compounds, mainly highly oxygenated nortriterpenoids, as well as bisnortriterpenoids, triterpenoids and homotriterpenoids, all present in small amounts. Substances with identical or similar molecular formulas were previously isolated from the aerial parts or fruits of S. chinensis (Huang et al., 2008 ; Li et al., 2017 ; Shi et al., 2014 ; S. Yang & Yuan, 2021 ). One of the detected bisnortriterpenoids was tentatively identified (on the basis of its determined formula) as wuweizidilactone H; this compound was earlier purified from the berry of S. chinensis by other research groups (Li et al., 2017 ; Xue et al., 2010 ). Hemostasis is a complex process depending on many interconnecting factors, among which blood platelets play a key role (Kannan et al., 2019 ; Khodadi, 2020 ). Platelets are small (approximately 2–4 µm), discoid, anucleate blood elements. They can activate due to contact with various agonists, like thrombin, ADP, thromboxane A 2 , or collagen (Gremmel et al., 2016 ; Rubenstein & Yin, 2018 ; Tomaiuolo et al., 2017 ). Upon activation, platelets change their shape and expose various proteins that aid in the coagulation process. One of those proteins is Pselectin, which in resting platelets is located in α granules. After activation, α granules fuse with the cell membrane, which exposes P-selectin on the surface of platelets, allowing for their adhesion to leukocytes and/or endothelial cells. Because P-selectin is exposed only on the surface of activated platelets, it is often used as an activation marker (Kannan et al., 2019 ; Rubenstein & Yin, 2018 ). GPIIb/IIIa (integrin α IIb β 3 ) is another protein that plays a vital role in hemostasis. In resting platelets, GPIIb/IIIa is exposed on the cell membrane in its low affinity state, and transforms into a high affinity (active) form upon platelet activation. GPIIb/IIIa plays a key role in aggregation, as it binds to fibrinogen, which in turn binds to GPIIb/IIIa on other platelets, facilitating the formation of platelet aggregates (Gremmel et al., 2016 ; Kannan et al., 2019 ; Rubenstein & Yin, 2018 ; Tomaiuolo et al., 2017 ). Currently, there are two GPIIb/IIIa inhibitors used as anti-platelet drugs - tirofiban and eptifibatide (Tummala & Rai, 2024 ). Platelet adhesion is mediated mainly by GPIb-IX-V (which binds to von Willebrand factor (vWF), which in turn binds to collagen) and GPIV, which binds to collagen directly (Gremmel et al., 2016 ; Kannan et al., 2019 ; Rubenstein & Yin, 2018 ; Tomaiuolo et al., 2017 ). Another change that occurs in platelets during activation is increased shedding of platelet-derived microparticles (PMPs), also known as microvesicles. PMPs are 1 µm or less but are bigger than exosomes (30–100 nm). Their plasma membrane exposes negatively charged phospholipids and a number of platelet receptors, including P-selectin, GPIIb/IIIa, and vWF (Nignpense et al., 2019; Kannan et al., 2019 ). Cardiovascular diseases are a leading cause of death worldwide. An imbalance in hemostasis which leads to excessive clotting is a major issue in many CVDs and can lead to severe conditions, like myocardial infarction or stroke (Kannan et al., 2019 ; Khodadi, 2020 ). Increased platelet activation plays a major role in the pathology of many CVDs and is associated with adverse prognosis. To prevent this, many cardiovascular patients must take anti-platelet medication, among which acetylsalicylic acid (aspirin) is used most often. Aspirin inhibits cyclooxygenase (COX) activity, decreasing the production of thromboxane A 2 (Kannan et al., 2019 ). Unfortunately, aspirin and many other anti-platelet drugs can cause adverse effects, including bleeding from the gastrointestinal tract. For this reason, researchers are searching for new anti-platelet compounds with a lower risk of side effects (Gremmel et al., 2016 ; Olas, 2020 ). The most important aspect of our findings is the confirmation of the anti-platelet activity of the extract from S. chinensis berries in various in vitro models. We used flow cytometry and TTAS to study platelet activation in whole blood, which is a more natural environment than media in which blood platelets are suspended after isolation. For the first time, we noted that S. chinensis extract (at the highest tested concentration – 50 µg/mL) significantly prolonged the time of occlusion, showing anti-platelet activity in vitro . We also observed that all the tested concentrations (0.5–50 µg/mL) of the extract inhibit the exposition of the active form of GPIIb/IIIa in 10 µM ADP-stimulated platelets. In addition, we observed the anti-adhesive potential of the extract from S. chinensis berries using an in vitro model based on human washed blood platelets. Results of (Chang et al., 2005 ) also demonstrated that the extract from S. chinensis inhibits arachidonic acid-induced blood platelet aggregation. Moreover, they suggested that the inhibition of cyclooxygenase is its primary mechanism of action. On the other hand, our results demonstrate that S. chinensis berry extract did not influence plasma coagulation times. Various studies showed that lignans (including schisandrin, schisandrin A, B, and C) isolated from S. chinensis are its main active constituents, and possess a variety of nutritional properties and biological activities (Jia et al., 2023 ; Wang et al., 2024 ; Zhou et al., 2021 ). We propose that the most active component of S. chinensis berry extract may be schisandrin, which could be the major determinant of its anti-platelet activity in vitro . Other in vitro and in vivo models have also demonstrated that schisandrin has cardioprotective properties (Gong et al., 2021 ; Zhang et al., 2019 ). For example, (Gong et al., 2021 ) observed its cardioprotective activity in a myocardial ischemia/reperfusion injury murine model ( in vivo ) and H9c2 cardiomyocyte cell line subjected to hypoxia/reoxygenation injury ( in vitro ). (Zhang et al., 2019 ) also found that schisandrin promotes the recovery of myocardial tissues by enhancing cell viability and migration. Recently, more information about the protective effect of schisandrin on the cardiovascular system has been described in a review paper by (Wang et al., 2024 ). However, it did not cover the effect of this compound on blood platelets. The bioavailability and toxicity of various plant preparations (including extracts and chemical compounds) that could be used as drugs or supplements are a crucial element in the evaluation of their biological properties. In our study, none of the used concentrations of the extract from S. chinensis berries caused damage to human blood platelets (which was determined by a measurement of LDH that leaked out from the damaged cells into the extracellular medium). These results indicate, that this extract should be safe for use as a natural supplement with anti-platelet activity, but it still lacks validation in clinical settings. However, it has been found that extract from S. chinensis berries has little to no toxicity toward various animals, including mice, rats, pigs, and rabbits (Panossian & Wikman, 2008 ; K. Yang et al., 2022 ). Moreover, the results of (Chen et al., 2018 ) demonstrated that it did not have toxic effects in an atherosclerosis rat model. Researchers have incorporated schisandrin as a key ingredient in various formulations, including tablets, capsules, liquids, and injections, to investigate its efficacy. Interestingly, schisandrin exhibited a good level of absorption. Hydroxylation and demethylation pathways are its main metabolic modifications (Wang et al., 2024 ). Another active components that we identified in S. chinensis berries are triterpenoids, which display a wide range of biological activities, including antitumor, antiviral, hepatoprotective, neuroprotective, and anti-inflammatory (Jia et al., 2023 ). Moreover, other studies indicate that triterpenoids and their derivatives present in fractions and extracts from various organs of sea buckthorn have anti-platelet activity (Skalski et al., 2021 , 2023 ). Other compounds that can significantly affect the circulatory system, including blood platelets, are phenolic compounds. They occur in large quantities in many plant species, including sea buckthorn and aronia berries (Olas, 2017 , 2018 , 2023 ; Olas et al., 2008 ; Skalski et al., 2020 , 2021 ). Some mechanisms of their anti-platelet activity are the inhibition of cyclooxygenase and blocking the binding of surface receptors to adhesion proteins. An additional advantage of supplementation with phenolic compounds is the lack of side effects (Luo et al., 2017 ; Olas, 2017 ). To conclude, our results indicate that S. chinensis berries could be used to produce natural nutrient supplements with cardioprotective potential, including anti-platelet activity. Future investigations into this extract and its chemical components, including schisandrin, should prioritize the following areas: (1) further understanding of the molecular mechanisms underlying its anti-platelet action, and the identification of specific targets; (2) bridging the gaps in knowledge about its in vivo efficiency by using animal models and performing clinical studies on healthy people and patients with different CVDs. Abbreviations ADI - Acceptable Daily Intake; APTT - activated partial thromboplastin time; BSA - bovine serum albumin; CPDA - citrate/phosphate/dextrose/adenine; CVDs – cardiovascular diseases; FDA - Food and Drug Administration; LDH - lactate dehydrogenase; PBS - Phosphate buffered saline; PT - prothrombin time; T-TAS - Total Thrombus-formation Analysis System; TT - thrombin time; WHO - The World Health Organization Declarations Acknowledgements : The Authors would like to thank Mariusz Kowalczyk, PhD for performing UHPLC-HRMS analyses. Conflict of interest: the authors declare no conflict of interest. The data availability: The datasets used and/or analysed during the current study available from the corresponding author on reasonable request. Author Contribution N.S. and J.Z. - methodology, formal analysis, investigation, writing-original draft, prepared figures; B.L., J.B., and K. Z. - methodology, formal analysis, investigtion; B.M.Sz. , P.B., and A.S. - methodology; B.O. - conceptualization, writing-rewiew and editing.All authors reviewed the manuscript. References Bellavite, P., Andrioli, G., Guzzo, P., Arigliano, P., Chirumbolo, S., Manzato, F., & Santonastaso, C. (1994). A Colorimetric Method for the Measurement of Platelet Adhesion in Microtiter Plates. Analytical Biochemistry , 216 (2), 444–450. https://doi.org/10.1006/abio.1994.1066 Chang, G.-T., Kang, S.-K., Kim, J.-H., Chung, K.-H., Chang, Y.-C., & Kim, C.-H. (2005). Inhibitory effect of the Korean herbal medicine, Dae-Jo-Whan, on platelet-activating factor-induced platelet aggregation. Journal of Ethnopharmacology , 102 (3), 430–439. https://doi.org/10.1016/j.jep.2005.07.003 Chen, X., Cao, J., Sun, Y., Dai, Y., Zhu, J., Zhang, X., Zhao, X., Wang, L., Zhao, T., Li, Y., Liu, Y., Wei, G., Zhang, T., & Yan, Z. (2018). Ethanol extract of Schisandrae chinensis fructus ameliorates the extent of experimentally induced atherosclerosis in rats by increasing antioxidant capacity and improving endothelial dysfunction. Pharmaceutical Biology , 56 (1), 612–619. https://doi.org/10.1080/13880209.2018.1523933 Chun, J. N., Cho, M., So, I., & Jeon, J.-H. (2014). The protective effects of Schisandra chinensis fruit extract and its lignans against cardiovascular disease: A review of the molecular mechanisms. Fitoterapia , 97 , 224–233. https://doi.org/10.1016/j.fitote.2014.06.014 Gong, S., Liu, J., Wan, S., Yang, W., Zhang, Y., Yu, B., Li, F., & Kou, J. (2021). Schisandrol A Attenuates Myocardial Ischemia/Reperfusion-Induced Myocardial Apoptosis through Upregulation of 14-3-3θ. Oxidative Medicine and Cellular Longevity , 2021 , 1–15. https://doi.org/10.1155/2021/5541753 Gremmel, T., Frelinger, A., & Michelson, A. (2016). Platelet Physiology. Seminars in Thrombosis and Hemostasis , 42 (03), 191–204. https://doi.org/10.1055/s-0035-1564835 He, X.-G., Lian, L.-Z., & Lin, L.-Z. (1997). Analysis of lignan constituents from Schisandra chinensis by liquid chromatography-electrospray mass spectrometry. Journal of Chromatography A , 757 , 81–87. Hosokawa, K., Ohnishi, T., Kondo, T., Fukasawa, M., Koide, T., Maruyama, I., & Tanaka, K. A. (2011). A novel automated microchip flow-chamber system to quantitatively evaluate thrombus formation and antithrombotic agents under blood flow conditions. Journal of Thrombosis and Haemostasis , 9 (10), 2029–2037. https://doi.org/10.1111/J.1538-7836.2011.04464.X Huang, S.-X., Han, Q.-B., Lei, C., Pu, J.-X., Xiao, W.-L., Yu, J.-L., Yang, L.-M., Xu, H.-X., Zheng, Y.-T., & Sun, H.-D. (2008). Isolation and characterization of miscellaneous terpenoids of Schisandra chinensis. Tetrahedron , 64 (19), 4260–4267. https://doi.org/10.1016/j.tet.2008.02.085 Jia, M., Zhou, L., Lou, Y., Yang, X., Zhao, H., Ouyang, X., & Huang, Y. (2023). An analysis of the nutritional effects of Schisandra chinensis components based on mass spectrometry technology. Frontiers in Nutrition , 10 . https://doi.org/10.3389/fnut.2023.1227027 Kannan, M., Ahmad, F., & Saxena, R. (2019). Platelet activation markers in evaluation of thrombotic risk factors in various clinical settings. Blood Reviews , 37 , 100583. https://doi.org/10.1016/j.blre.2019.05.007 Khodadi, E. (2020). Platelet Function in Cardiovascular Disease: Activation of Molecules and Activation by Molecules. Cardiovascular Toxicology 20 (1), 1-10 https://doi.org/10.1007/s12012-019-09555-4 Li, F., Zhang, T., Sun, H., Gu, H., Wang, H., Su, X., Li, C., Li, B., Chen, R., & Kang, J. (2017). A New Nortriterpenoid, a Sesquiterpene and Hepatoprotective Lignans Isolated from the Fruit of Schisandra chinensis. Molecules , 22 (11), 1931. https://doi.org/10.3390/molecules22111931 Liao, J., Zang, J., Yuan, F., Liu, S., Zhang, Y., Li, H., Piao, Z., & Li, H. (2016). Identification and analysis of anthocyanin components in fruit color variation in Schisandra chinensis . Journal of the Science of Food and Agriculture , 96 (9), 3213–3219. https://doi.org/10.1002/jsfa.7503 Lin, Q., Qin, X., Shi, M., Qin, Z., Meng, Y., Qin, Z., & Guo, S. (2017). Schisandrin B inhibits LPS-induced inflammatory response in human umbilical vein endothelial cells by activating Nrf2. International Immunopharmacology , 49 , 142–147. https://doi.org/10.1016/j.intimp.2017.05.032 Lis, B., Rolnik, A., Jedrejek, D., Soluch, A., Stochmal, A., & Olas, B. (2019). Dandelion ( Taraxacum officinale L.) root components exhibit anti-oxidative and antiplatelet action in an in vitro study. Journal of Functional Foods , 59 , 16–24. https://doi.org/10.1016/j.jff.2019.05.019 Liu, H., Lai, H., Jia, X., Liu, J., Zhang, Z., Qi, Y., Zhang, J., Song, J., Wu, C., Zhang, B., & Xiao, P. (2013). Comprehensive chemical analysis of Schisandra chinensis by HPLC–DAD–MS combined with chemometrics. Phytomedicine , 20 (12), 1135–1143. https://doi.org/10.1016/j.phymed.2013.05.001 Luo, X., Du, C., Cheng, H., Chen, J., & Lin, C. (2017). Study on the Anticoagulant or Procoagulant Activities of Type II Phenolic Acid Derivatives. Molecules , 22 (12), 2047. https://doi.org/10.3390/molecules22122047 Ma, C., Yang, L., Yang, F., Wang, W., Zhao, C., & Zu, Y. (2012). Content and Color Stability of Anthocyanins Isolated from Schisandra chinensis Fruit. International Journal of Molecular Sciences , 13 (12), 14294–14310. https://doi.org/10.3390/ijms131114294 Malinowska, J., Kołodziejczyk-Czepas, J., Moniuszko-Szajwaj, B., Kowalska, I., Oleszek, W., Stochmal, A., & Olas, B. (2012). Phenolic fractions from Trifolium pallidum and Trifolium scabrum aerial parts in human plasma protect against changes induced by hyperhomocysteinemia in vitro. Food and Chemical Toxicology: An International Journal Published for the British Industrial Biological Research Association , 50 (11), 4023–4027. https://doi.org/10.1016/J.FCT.2012.07.065 Mocan, A., Crișan, G., Vlase, L., Crișan, O., Vodnar, D., Raita, O., Gheldiu, A.-M., Toiu, A., Oprean, R., & Tilea, I. (2014). Comparative Studies on Polyphenolic Composition, Antioxidant and Antimicrobial Activities of Schisandra chinensis Leaves and Fruits. Molecules , 19 (9), 15162–15179. https://doi.org/10.3390/molecules190915162 Ed Nignpense, B., Chinkwo, K. A., Blanchard, C. L., & Santhakumar, A. B. (2019). Polyphenols: Modulators of Platelet Function and Platelet Microparticle Generation? International Journal of Molecular Sciences , 21 (1), 146. https://doi.org/10.3390/ijms21010146 Olas, B. (2017). The multifunctionality of berries toward blood platelets and the role of berry phenolics in cardiovascular disorders. Platelets , 28 (6), 540–549. https://doi.org/10.1080/09537104.2016.1235689 Olas, B. (2018). Berry Phenolic Antioxidants – Implications for Human Health? Frontiers in Pharmacology , 9 (MAR), 78. https://doi.org/10.3389/fphar.2018.00078 Olas, B. (2020). Biochemistry of blood platelet activation and the beneficial role of plant oils in cardiovascular diseases. In Advances in Clinical Chemistry (Vol. 95, pp. 219–243). Academic Press Inc. https://doi.org/10.1016/bs.acc.2019.08.006 Olas, B. (2023). Cardioprotective Potential of Berries of Schisandra chinensis Turcz. (Baill.), Their Components and Food Products. Nutrients , 15 (3). https://doi.org/10.3390/nu15030592 Olas, B., Wachowicz, B., Tomczak, A., Erler, J., Stochmal, A., & Oleszek, W. (2008). Comparative anti-platelet and antioxidant properties of polyphenol-rich extracts from: berries of Aronia melanocarpa , seeds of grape and bark of Yucca schidigera in vitro . Platelets , 19 (1), 70–77. https://doi.org/10.1080/09537100701708506 Panossian, A., & Wikman, G. (2008). Pharmacology of Schisandra chinensis Bail.: An overview of Russian research and uses in medicine. Journal of Ethnopharmacology , 118 (2), 183–212. https://doi.org/10.1016/j.jep.2008.04.020 Rubenstein, D. A., & Yin, W. (2018). Platelet‐Activation Mechanisms and Vascular Remodeling. In Comprehensive Physiology (Vol. 8, Issue 3, pp. 1117–1156). Wiley. https://doi.org/10.1002/cphy.c170049 Sheng, Y., Wang, R., Wang, Y., Li, M., Liu, F., Huang, X., & Chen, C. (2022). Evaluation of Multicomponent Changes of Schisandra chinensis Fruits with Different Drying Process by UPLC-QQQ-MS-Based Targeted Metabolomics Analysis. Journal of Chemistry , 2022 , 1–10. https://doi.org/10.1155/2022/2616122 Shi, Y.-M., Wang, L.-Y., Zou, X.-S., Li, X.-N., Shang, S.-Z., Gao, Z.-H., Liang, C.-Q., Luo, H.-R., Li, H.-L., Xiao, W.-L., & Sun, H.-D. (2014). Nortriterpenoids from Schisandra chinensis and their absolute configurational assignments by electronic circular dichroism study. Tetrahedron , 70 (4), 859–868. https://doi.org/10.1016/j.tet.2013.12.023 Skalski, B., Rywaniak, J., Szustka, A., Żuchowski, J., Stochmal, A., & Olas, B. (2021). Anti-Platelet Properties of Phenolic and Nonpolar Fractions Isolated from Various Organs of Elaeagnus rhamnoides (L.) A. Nelson in Whole Blood. International Journal of Molecular Sciences , 22 (6), 3282. https://doi.org/10.3390/ijms22063282 Skalski, B., Rywaniak, J., Żuchowski, J., Stochmal, A., & Olas, B. (2023). The changes of blood platelet reactivity in the presence of Elaeagnus rhamno ides (L.) A. Nelson leaves and twig extract in whole blood. Biomedicine & Pharmacotherapy , 162 , 114594. https://doi.org/10.1016/j.biopha.2023.114594 Skalski, B., Stochmal, A., Żuchowski, J., Grabarczyk, Ł., & Olas, B. (2020). Response of blood platelets to phenolic fraction and non-polar fraction from the leaves and twigs of Elaeagnus rhamnoides (L.) A. Nelson in vitro. Biomedicine & Pharmacotherapy , 124 , 109897. https://doi.org/10.1016/j.biopha.2020.109897 Sławińska, N., & Olas, B. (2022). Selected Seeds as Sources of Bioactive Compounds with Diverse Biological Activities. Nutrients , 15 (1), 187. https://doi.org/10.3390/nu15010187 Sławińska, N., Żuchowski, J., Stochmal, A., & Olas, B. (2023). Extract from Sea Buckthorn Seeds—A Phytochemical, Antioxidant, and Hemostasis Study; Effect of Thermal Processing on Its Chemical Content and Biological Activity In Vitro . Nutrients , 15 (3), 686. https://doi.org/10.3390/nu15030686 Su, D., Luo, J., Ge, J., Liu, Y., Jin, C., Xu, P., Zhang, R., Zhu, G., Yang, M., Ai, Z., & Song, Y. (2022). Raw and Wine Processed Schisandra chinensis Regulate NREM-Sleep and Alleviate Cardiovascular Dysfunction Associated with Insomnia by Modulating HPA Axis. Planta Medica , 88 (14), 1311–1324. https://doi.org/10.1055/a-1721-4971 Szopa, A., Ekiert, R., & Ekiert, H. (2017). Current knowledge of Schisandra chinensis (Turcz.) Baill. (Chinese magnolia vine) as a medicinal plant species: a review on the bioactive components, pharmacological properties, analytical and biotechnological studies. Phytochemistry Reviews , 16 (2), 195–218. https://doi.org/10.1007/s11101-016-9470-4 Szopa, A., Klimek, M., & Ekiert, H. (2016). Chinese magnolia vine ( Schisandra chinensis ) - therapeutic and cosmetic importance. Polish Journal of Cosmetology , 19 (4), 274–284. Tomaiuolo, M., Brass, L. F., & Stalker, T. J. (2017). Regulation of Platelet Activation and Coagulation and Its Role in Vascular Injury and Arterial Thrombosis. Interventional Cardiology Clinics , 6 (1), 1–12. https://doi.org/10.1016/j.iccl.2016.08.001 Tummala, R., & Rai, M. P. (2024). Glycoprotein IIb/IIIa Inhibitors . Valíčková, J., Zezulka, Š., Maršálková, E., Kotlík, J., Maršálek, B., & Opatřilová, R. (2023). Bioactive compounds from Schisandra chinensis - Risk for aquatic plants? Aquatic Toxicology (Amsterdam, Netherlands) , 254 , 106365. https://doi.org/10.1016/j.aquatox.2022.106365 Wang, X., Wang, X., Yao, H., Shen, C., Geng, K., & Xie, H. (2024). A comprehensive review on Schisandrin and its pharmacological features. Naunyn-Schmiedeberg’s Archives of Pharmacology , 397 (2), 783–794. https://doi.org/10.1007/s00210-023-02687-z Wróblewski, F., & Ladue, J. S. (1955). Lactic dehydrogenase activity in blood. Proc. Soc. Exp. Biol. Med , 90 , 210–221. Wu, J., Jia, J., Liu, L., Yang, F., Fan, Y., Zhang, S., Yan, D., Bu, R., Li, G., Gao, Y., & Chen, Y. (2017). Schisandrin B displays a protective role against primary pulmonary hypertension by targeting transforming growth factor β1. Journal of the American Society of Hypertension , 11 (3), 148-157.e1. https://doi.org/10.1016/j.jash.2016.12.007 Xue, Y.-B., Zhang, Y.-L., Yang, J.-H., Du, X., Pu, J.-X., Zhao, W., Li, X.-N., Xiao, W.-L., & Sun, H.-D. (2010). Nortriterpenoids and Lignans from the Fruit of Schisandra chinensis. Chemical and Pharmaceutical Bulletin , 58 (12), 1606–1611. https://doi.org/10.1248/cpb.58.1606 Yang, J., IP, S. P., J. Yeung, H., & Che, C. (2011). HPLC-MS analysis of Schisandra lignans and their metabolites in Caco-2 cell monolayer and rat everted gut sac models and in rat plasma. Acta Pharmaceutica Sinica B , 1 (1), 46–55. https://doi.org/10.1016/j.apsb.2011.04.007 Yang, K., Qiu, J., Huang, Z., Yu, Z., Wang, W., Hu, H., & You, Y. (2022). A comprehensive review of ethnopharmacology, phytochemistry, pharmacology, and pharmacokinetics of Schisandra chinensis (Turcz.) Baill. and Schisandra sphenanthera Rehd. et Wils. Journal of Ethnopharmacology , 284 , 114759. https://doi.org/10.1016/j.jep.2021.114759 Yang, S., & Yuan, C. (2021). Schisandra chinensis : A comprehensive review on its phytochemicals and biological activities. Arabian Journal of Chemistry , 14 (9), 103310. https://doi.org/10.1016/j.arabjc.2021.103310 Zhang, X., Zhao, Y., Bai, D., Yuan, X., & Cong, S. (2019). Schizandrin protects H9c2 cells against lipopolysaccharide‐induced injury by downregulating Smad3. Journal of Biochemical and Molecular Toxicology , 33 (5). https://doi.org/10.1002/jbt.22301 Zhou, Y., Men, L., Sun, Y., Wei, M., & Fan, X. (2021). Pharmacodynamic effects and molecular mechanisms of lignans from Schisandra chinensis Turcz. (Baill.), a current review. European Journal of Pharmacology , 892 , 173796. https://doi.org/10.1016/j.ejphar.2020.173796 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4346913","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":301476306,"identity":"731541c8-2fd2-4f76-bb92-a2a5551785d5","order_by":0,"name":"Natalia Sławińska","email":"","orcid":"","institution":"University of Lodz","correspondingAuthor":false,"prefix":"","firstName":"Natalia","middleName":"","lastName":"Sławińska","suffix":""},{"id":301476307,"identity":"5d93978c-9330-40a6-a80a-d17c2f00d3ac","order_by":1,"name":"Bogdan Kontek","email":"","orcid":"","institution":"University of Lodz","correspondingAuthor":false,"prefix":"","firstName":"Bogdan","middleName":"","lastName":"Kontek","suffix":""},{"id":301476308,"identity":"c872e98f-39de-42c5-976d-1ac569760ced","order_by":2,"name":"Jerzy Żuchowski","email":"","orcid":"","institution":"Institute of Soil Science and Plant Cultivation – State Research Institute","correspondingAuthor":false,"prefix":"","firstName":"Jerzy","middleName":"","lastName":"Żuchowski","suffix":""},{"id":301476309,"identity":"b82382ff-40b7-45d7-8026-ff7504f096ac","order_by":3,"name":"Barbara Moniuszko-Szajwaj","email":"","orcid":"","institution":"Institute of Soil Science and Plant Cultivation – State Research Institute","correspondingAuthor":false,"prefix":"","firstName":"Barbara","middleName":"","lastName":"Moniuszko-Szajwaj","suffix":""},{"id":301476310,"identity":"1e15bebc-bc6b-427a-8dad-c6c0603c74a3","order_by":4,"name":"Jacek Białecki","email":"","orcid":"","institution":"University of Lodz","correspondingAuthor":false,"prefix":"","firstName":"Jacek","middleName":"","lastName":"Białecki","suffix":""},{"id":301476311,"identity":"aebfef08-0595-4cff-8c31-ae3d5c8ef1d7","order_by":5,"name":"Kamil Zakrzewski","email":"","orcid":"","institution":"University of Lodz","correspondingAuthor":false,"prefix":"","firstName":"Kamil","middleName":"","lastName":"Zakrzewski","suffix":""},{"id":301476312,"identity":"c9523259-b42f-47f4-ae7a-a7d1c54e913f","order_by":6,"name":"Paulina Bogusz","email":"","orcid":"","institution":"Lukasiewicz Research Network – New Chemical Synthesis Institute","correspondingAuthor":false,"prefix":"","firstName":"Paulina","middleName":"","lastName":"Bogusz","suffix":""},{"id":301476313,"identity":"1b33563f-1aff-4f92-8a71-082c8d80c429","order_by":7,"name":"Anna Stochmal","email":"","orcid":"","institution":"Institute of Soil Science and Plant Cultivation – State Research Institute","correspondingAuthor":false,"prefix":"","firstName":"Anna","middleName":"","lastName":"Stochmal","suffix":""},{"id":301476314,"identity":"928f3042-ceda-4b03-a4a5-e5c836302109","order_by":8,"name":"Beata Olas","email":"data:image/png;base64,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","orcid":"","institution":"University of Lodz","correspondingAuthor":true,"prefix":"","firstName":"Beata","middleName":"","lastName":"Olas","suffix":""}],"badges":[],"createdAt":"2024-04-30 07:07:26","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4346913/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4346913/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":56403469,"identity":"81bc0a42-e7bd-4123-b0d1-86e584c11657","added_by":"auto","created_at":"2024-05-13 17:32:52","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":55269,"visible":true,"origin":"","legend":"\u003cp\u003eUHPLC-CAD (\u003cstrong\u003eA\u003c/strong\u003e), and UV (λ = 255 nm (\u003cstrong\u003eB\u003c/strong\u003e)) chromatograms of the extract from the fruit of \u003cem\u003eS. chinensis\u003c/em\u003e. Numbers above the chromatic peaks correspond to those from Table 1.\u003c/p\u003e","description":"","filename":"1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/1f3b1e33bc062515904ff8e6.jpg"},{"id":56402882,"identity":"e4d7379a-81d3-46da-a3ae-80c56de071f3","added_by":"auto","created_at":"2024-05-13 17:24:56","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":86941,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (at concentrations 0.5, 1, 5, 10, and 50 μg/mL) on the adhesion of resting platelets to collagen (A), thrombin-activated platelets to collagen (B), thrombin-activated platelets to fibrinogen (C), and ADP-activated platelets to fibrinogen (D) (n=9). In the graphs, platelet adhesion is expressed as a percentage of the control sample (blood platelets without the tested extract). The data are expressed as means ± SD. The results were considered significant at p\u0026lt;0.05 (* p\u0026lt;0.05).\u003c/p\u003e","description":"","filename":"2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/e85d8d30285e072e1b8c85e3.jpg"},{"id":56402886,"identity":"e561f2c9-e207-445a-8724-9ccccc3d60d5","added_by":"auto","created_at":"2024-05-13 17:24:56","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":70081,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (at concentrations 0.5, 1, 5, 10, and 50 μg/mL) on the exposition of the active form of GPIIb/IIIa on 10 µM ADP-stimulated blood platelets (A), 20 µM ADP-stimulated blood platelets (B), and 10 µg/mL collagen-stimulated blood platelets (C) in whole blood samples. Blood platelets were distinguished based on the exposition of CD61. For each sample, 5000 CD61-positive objects were acquired. For the assessment of GPIIb/IIIa exposition, samples were labeled with fluorescently conjugated monoclonal antibody PAC-1/FITC. Results are shown as the percentage of platelets binding PAC-1/FITC. Data represent the means ± SD. The blood samples were drawn from 6 healthy volunteers. The activity of the tested extract was compared to the control sample. The results were considered as significant at p\u0026gt;0.05 (*p\u0026lt;0.05).\u003c/p\u003e","description":"","filename":"3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/6d27f528282439b4695ebb97.jpg"},{"id":56402881,"identity":"1c91582d-5d57-4bec-9fb3-5ca252c40064","added_by":"auto","created_at":"2024-05-13 17:24:56","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":76617,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (at concentrations 0.5, 1, 5, 10, and 50 μg/mL) on the exposition of P-selectin on 10 µM ADP-stimulated blood platelets (A), 20 µM ADP-stimulated blood platelets (B), and 10 µg/mL collagen-stimulated blood platelets (C) in whole blood samples. Blood platelets were distinguished based on the exposition of CD61. For each sample, 5000 CD61-positive objects were acquired. For the assessment of P-selectin exposition, samples were labeled with fluorescently conjugated monoclonal antibody CD62P/PE. Results are shown as the percentage of platelets expressing CD62P. Data represent the means ± SD. The blood samples were drawn from 6 healthy volunteers. The activity of the tested extract was compared to the control sample. The results were considered as significant at p\u0026gt;0.05 (*p\u0026lt;0.05).\u003c/p\u003e","description":"","filename":"4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/bd78d7e7204e31d1ec072ca4.jpg"},{"id":56402884,"identity":"70b0ed58-d7d1-4302-81a2-af2119040e04","added_by":"auto","created_at":"2024-05-13 17:24:56","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":50079,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (at concentrations 0.5, 1, 5, 10, and 50 μg/mL) on thrombus formation in whole blood (n=7). The samples were analyzed with T-TAS PL-chip, at the shear stress rates of 1500/s. The results are calculated as AUC\u003csub\u003e10\u003c/sub\u003e (area under the curve). In the graphs, AUC\u003csub\u003e10\u003c/sub\u003e is expressed as a percentage of the control sample (blood without the tested extract). The data are expressed as means ± SD. The results were considered significant at p\u0026lt;0.05 (*\u0026nbsp;p\u0026lt;0.05).\u003c/p\u003e","description":"","filename":"5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/3f704c025ecd725ac96df899.jpg"},{"id":59912308,"identity":"127c3c37-7dde-431f-8d75-3431ac9848d0","added_by":"auto","created_at":"2024-07-09 08:18:24","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1709486,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4346913/v1/311fb38e-4c1a-4ab8-94a8-b4ff6594a59d.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Phytochemical analysis of the extract from berries of Schisandra chinensis Turcz. (Baill.) and its anti-platelet potential in vitro","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eEdible berries are an important component of the human diet, serving as a source of nutrients and bioactive compounds. They contain a variety of phytochemicals that exhibit a wide range of biological properties, including cardioprotective activity (Olas, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2017\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Sławińska et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Sławińska \u0026amp; Olas, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cem\u003eSchisandra chinensis\u003c/em\u003e Turcz. (Baill.) is a dioecious vine, belonging to the Schisandraceae family. It occurs naturally in the forests of south-eastern Siberia (Primorski, Amurski, Khabarovski regions, Sakhalin), north-eastern China, Korea, and Japan. Recently, Val\u0026iacute;čkov\u0026aacute; et al. (\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2023\u003c/span\u003e) have shown that \u003cem\u003eS. chinensis\u003c/em\u003e has the potential to be used for the production of nutrient supplements due to its adaptogenic properties. The primary property of all adaptogens is to eliminate the effects of stress and adapt the body to unfavorable external conditions (Szopa et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2016\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cem\u003eS. chinensis\u003c/em\u003e fruit, called \"fruit of five flavors\" (wǔw\u0026egrave;izǐ), which results from the fact that the fruit's peel is sweet, the pulp is sour, the seeds are bitter and astringent, and the grain extract is salty, has been used in Chinese medicine to treat different disorders and diseases. Indigenous peoples of the Far-Eastern regions of Russia used it to reduce hunger, thirst, and exhaustion (Panossian \u0026amp; Wikman, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Szopa et al., \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Importantly, \u003cem\u003eS. chinensis\u003c/em\u003e berries also have cardioprotective potential (Chun et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Olas, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), which is exerted through various molecular pathways. They can also help control oxidative stress, obesity, and inflammation (Lin et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Su et al., \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Wu et al., \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). In traditional Chinese medicine, \u003cem\u003eS. chinensis\u003c/em\u003e is often used in the treatment of palpitation (Su et al., \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). The 2020-year edition of Chinese Pharmacopoeia contains 100 prescriptions containing \u003cem\u003eS. chinensis\u003c/em\u003e (K. Yang et al., \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Due to medicinal and health promoting properties of this plant, many food supplements made from \u003cem\u003eS. chinensis\u003c/em\u003e fruit are available on the market.\u003c/p\u003e \u003cp\u003eDifferent components of \u003cem\u003eS. chinensis\u003c/em\u003e, including lignans (especially schisandrin, also called schizandrin, schisandrol A, or wǔw\u0026egrave;izǐ s\u0026ugrave; A) play a significant role in the prophylaxis and treatment of cardiovascular diseases (CVDs), for example in myocardial infarction and hypertension. However, there are no experiments that focus on the response of various elements of hemostasis (including blood platelets, plasma, and others) to \u003cem\u003eS. chinensis\u003c/em\u003e berries. Therefore, the present study examined the anti-platelet activity of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries in washed human blood platelets and human whole blood \u003cem\u003ein vitro\u003c/em\u003e. Using washed platelets, we studied the activity of this extract on platelet adhesion to two adhesive proteins \u0026ndash; collagen and fibrinogen. We also determined the anti-platelet activity of the tested extract in whole blood using the Total Thrombus-formation Analysis System (T-TAS) and flow cytometry. Moreover, we evaluated the cytotoxicity of the extract against blood platelets based on extracellular lactate dehydrogenase (LDH) activity. We also investigated the effect of \u003cem\u003eS. chinensis\u003c/em\u003e berry extract on the parameters of hemostasis in plasma (\u003cem\u003ein vitro\u003c/em\u003e), including coagulation times: thrombin time (TT), prothrombin time (PT), and activated partial thromboplastin time (APTT). The action of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries was compared to the extract from sea buckthorn (\u003cem\u003eHippophae rhamnoides\u003c/em\u003e L.) berries, and a commercial product \u0026ndash; Aronox (A\u003cem\u003eronia melanocarpa\u003c/em\u003e berry extract), which have documented anti-platelet properties (Olas et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Skalski et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\n \u003ch2\u003e2.1. Reagents\u003c/h2\u003e\n \u003cp\u003eMethanol (HPLC grade) was purchased from Fisher Chemical (UK), heptane (for chromatography), and acetonitrile (LC-MS grade) were obtained from Merck (Darmstadt, Germany). Monoclonal antibodies for flow cytometry (CD61-PerCP, CD62P-PE, and PAC-1-FITC) were purchased from Becton Dickinson (New Jersey, USA). Phosphate buffered saline (PBS), fibrinogen, collagen, bovine serum albumin (BSA), tris(hydroxymethyl)aminomethane (Tris), 4-nitrophenyl phosphate, ADP, nicotinamide adenine dinucleotide (NADH), and thrombin were purchased from Merck (Darmstadt, Germany). NaCl, sodium citrate, and citric acid were purchased from POCH (Avantor performance materials, Gliwice, Poland). Reagents needed for coagulation time measurements were purchased from Kselmed (Grudziądz, Poland). All materials and chemicals for T-TAS were purchased from Bionicum Sp. z o.o. (Warsaw, Poland). Other reagents were purchased from commercial distributors and were of the highest available grade.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\n \u003ch2\u003e2.2. Plant material\u003c/h2\u003e\n \u003cp\u003eCrimson-red, ripe \u003cem\u003eS. chinensis\u003c/em\u003e berries, growing in spike clusters on vines, were collected in mild-September 2022 in the park and palace settlement \u0026ldquo;Lower Garden\u0026rdquo; of the Czartoryski family in Pulawy (51\u0026deg;24\u0026rsquo;47.698\u0026rdquo; N21\u0026deg; 57\u0026rsquo; 20.626\u0026rdquo; E). The plants were planted about 40 years ago and identified by the park\u0026rsquo;s curator, dr. Adam Wołk. After harvesting, the berries were frozen at -18\u0026deg;C, and subsequently lyophilized (Gamma 2\u0026ndash;16 LSC, Christ, Osterode am Harz, Germany). The reference sample marked 1/09/2022 is located at the Institute of Soil Science and Plant Cultivation (Pulawy, Poland).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\n \u003ch2\u003e2.3. Preparation of the extract from S. chinensis berries\u003c/h2\u003e\n \u003cp\u003eFreeze-dried fruits (100 g) were soaked overnight in 80% methanol (v/v; 1.5 L) at ambient temperature, roughly homogenized using a kitchen blender, and subjected to a 15 min. sonication in an ultrasonic bath. The collected extract was centrifuged (3-16KL, Sigma, Osterode am Harz, Germany), and filtered using a glass filter funnel. The extraction procedure was repeated twice, with new portions of the solvent. The pooled extracts were concentrated in a rotary evaporator and defatted by liquid-liquid extraction with heptane. The defatted extract was rotary evaporated to remove organic solvents and reduce its volume, and freeze-dried. The extract (74.57 g) was subsequently cleaned up by SPE, to remove organic acids, sugars, and other highly polar constituents. It was dissolved in 0.1% formic acid in MilliQ water (v/v) and loaded onto a short C18 column (60 \u0026times; 50 mm; Cosmosil 140C18-Prep, 140 \u0026micro;m). The column was washed with 0.1% formic acid, and the bound compounds were eluted with methanol. The eluate was rotary-evaporated, and the residue was freeze-dried to yield 7.61 g of the purified extract.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e\n \u003ch2\u003e2.4. Phytochemical analysis of the extract from S. chinensis berries\u003c/h2\u003e\n \u003cp\u003eThe composition of \u003cem\u003eS. chinensis\u003c/em\u003e fruits was analyzed by LC-HRMS, using a Thermo Ultimate 3000RS (Thermo Fischer Scientific, Waltham, MS, USA) UHPLC system equipped with a corona-charged aerosol detector (CAD) and coupled with a Bruker Impact II HD quadrupole-time of flight (Q-TOF) mass spectrometer (Bruker Daltonics GmbH, Bremen, Germany). Samples were chromatographed on an ACQUITY UPLC HSS T3 column (2.1 \u0026times; 150 mm, 1.8 \u0026micro;m), equipped with a pre-column, and maintained at 40˚C. The injection volume was 2 \u0026micro;L. The mobile phase A was 0.1% formic acid in MilliQ water, and the mobile phase B was acetonitrile containing 0.1% of formic acid. The flow rate was 0.400 mL min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. The separation program started from a short isocratic elution with 5% B (0.5 min.), followed by a 31 min. gradient from 5 to 95% of solvent B. The elution was continued a t 95% B for 7 min., then the mobile phase composition returned to the initial conditions and the column was equilibrated with 5% B for 5 minutes. MS\u0026apos;s analyses were performed in the negative and positive ion mode. MS\u0026apos;s settings for negative ion mode: capillary voltage 3 kV; dry gas flow 6 L min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e; dry gas temperature 200\u0026deg;C; nebulizer pressure 2.5 bar; collision RF 750 Vpp; transfer time 100 \u0026micro;s; prepulse storage time 10 \u0026micro;s. Collision energy was set automatically in the range from 2.5 to 80 eV, depending on the \u003cem\u003em/z\u003c/em\u003e of a fragmented ion. The scanning values ranged from \u003cem\u003em/z\u003c/em\u003e 80 to m/z 2000. The settings for positive ion mode: capillary voltage 4 kV; dry gas flow 6 L min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e; dry gas temperature 200\u0026deg;C; nebulizer pressure 2.5 bar; collision RF 750 Vpp; transfer time 100 \u0026micro;s; prepulse storage time 10 \u0026micro;s. Collision energy was set automatically in the range from 2.5 to 50 eV. The scanning range was \u003cem\u003em/z\u003c/em\u003e 100\u0026ndash;2000.\u003c/p\u003e\n \u003cp\u003eQuantitative analyses of phenolic compounds were performed using an ACQUITY UPLC\u0026reg; chromatographic system (Waters Inc., Milford, MA, USA), equipped with a PDA detector, and coupled with a triple quadrupole mass detector (TQD; Waters). The \u003cem\u003eS. chinensis\u003c/em\u003e extract was separated on an ACQUITY UPLC BEH C18 (2.1 \u0026times; 100 mm, 1.7 \u0026micro;m; Waters) column. The column temperature was 50\u0026deg;C, the injection volume was 2.5 \u0026micro;L. Mobile phase was composed of Mill-Q water with 0.1% formic acid in (solvent A) and acetonitrile with 0.1% formic acid (solvent B); the elution program was as follows: 0.0-0.5 min., 7% B; 0.5-7.0 min., linear gradient 7\u0026ndash;25% B; 7.0-16.9 min., linear gradient 25\u0026ndash;75% B; 17.0\u0026ndash;18.0 min., 95% B; 18.1\u0026ndash;20.0 min., 7% B. The TQ mass spectrometer was operated in negative and positive ion mode. The MS settings in negative ion mode were as follows: capillary voltage 2.80 kV, cone voltage 45 V, source temperature 150\u0026deg;C, desolvation temperature 450\u0026deg;C, cone gas flow (N\u003csub\u003e2\u003c/sub\u003e) 100 L h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, desolvation gas (N\u003csub\u003e2\u003c/sub\u003e) flow 800 L h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. In positive ion mode: capillary voltage 3.10 kV, cone voltage 60 V, the remaining parameters were the same as those described above. The scanning range was \u003cem\u003em/z\u003c/em\u003e 100\u0026ndash;1200. The content of anthocyanins (the peak integration at \u0026lambda;\u0026thinsp;=\u0026thinsp;475 nm; y\u0026thinsp;=\u0026thinsp;232.90079x\u0026thinsp;+\u0026thinsp;82.94488, R\u0026sup2; = 0.9989), flavonoids (the peak integration at \u0026lambda;\u0026thinsp;=\u0026thinsp;350 nm; y\u0026thinsp;=\u0026thinsp;442.56566x \u0026minus;\u0026thinsp;46.00717, R\u0026sup2; = 0.9999), and lignans (the peak integration at \u0026lambda;\u0026thinsp;=\u0026thinsp;255 nm; y\u0026thinsp;=\u0026thinsp;607.19720x\u0026thinsp;+\u0026thinsp;131.25948, R\u0026sup2; = 0.9999) was calculated on the basis of calibration curves, and was expressed as cyanidin, rutin and schisandrin equivalents, respectively.\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cstrong\u003e2.5 Preparation of stock solution of the extract from S. chinensis berries, the extract from sea buckthorn berries, and the extract from aronia berries\u003c/strong\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eThe extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries and the extract from sea buckthorn berries were dissolved in 50% DMSO (a universal solvent for many plant substances), however the final concentration of DMSO in the tested human blood platelets, human plasma, and human blood was below 0.05% (\u003cem\u003ev\u003c/em\u003e/\u003cem\u003ev\u003c/em\u003e). The addition of a low concentration of DMSO to blood platelets, plasma, and blood has no effect on the hemostatic properties of blood platelets, plasma, and full blood (data not presented).\u003c/p\u003e\n \u003cp\u003eThe extract from sea buckthorn berries contained predominantly various flavonol glycosides, mainly isorhamnetin glycosides and acylated glycosides (Skalski et al., \u003cspan class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eA stock solution of commercial product \u0026ndash; Aronox (by Agropharm Ltd., Poland; batch No 020/2007k, \u003cem\u003eA. melanocarpa\u003c/em\u003e berry extract) was prepared in H\u003csub\u003e2\u003c/sub\u003eO at a concentration of 5 mg/mL.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\n \u003ch2\u003e2.6. Blood, plasma, and blood platelet samples\u003c/h2\u003e\n \u003cp\u003eWhole human blood was collected at \u0026ldquo;Diagnostyka\u0026rdquo; blood collection center (Brzechwy 7A, Lodz, Poland). All donors were non-smoking, healthy volunteers, which did not drink alcohol or take medication and anti-platelet supplementation for two weeks before blood collection. Peripheral blood was drawn into tubes with benzylsulfonyl-D-Arg-Pro-4-amidinobenzylamide (BAPA) (for T-TAS) or with citrate/phosphate/dextrose/adenine (CPDA) anticoagulant (for the remaining assays).\u003c/p\u003e\n \u003cp\u003eThe analysis of blood samples was performed according to the guidelines of the Helsinki Declaration for Human Research. Informed consent form was signed by each participant one day prior to blood collection. Procedures were conducted with the consent of Bioethics Committee at the University of Ł\u0026oacute;dź (2/KBBN-UŁ/III/2014).\u003c/p\u003e\n \u003cp\u003eBlood for the T-TAS assay was used within 2 hours after collection.\u003c/p\u003e\n \u003cp\u003eFor coagulation times measurements, plasma was separated from whole blood by differential centrifugation (2800 x g, 20 min., at room temperature).\u003c/p\u003e\n \u003cp\u003eBlood platelets were isolated from fresh blood by differential centrifugation as described previously (Lis et al., \u003cspan class=\"CitationRef\"\u003e2019\u003c/span\u003e). The required number of platelets (2.0 \u0026times; 10\u003csup\u003e8\u003c/sup\u003e/mL) was confirmed by a spectrophotometric measurement in a UV-Visible Helios-\u0026alpha; at 800 nm.\u003c/p\u003e\n \u003cp\u003eIn every experiment, blood, plasma, or blood platelets were incubated for 30 minutes at 37\u0026deg;C, either with the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries at final concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL, or the extract from sea buckthorn berries or aronia berries at final concentration 50 \u0026micro;g/mL.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\n \u003ch2\u003e2.7. Blood platelet adhesion\u003c/h2\u003e\n \u003cp\u003eBlood platelet adhesion was measured based on the activation of exoenzyme acid phosphatase in the blood platelets according to the method described by (Bellavite et al., \u003cspan class=\"CitationRef\"\u003e1994\u003c/span\u003e). 96-well plates were coated with 100 \u0026micro;g/mL fibrinogen or 0.04 \u0026micro;g/mL collagen. To achieve this, 100 \u0026micro;L of fibrinogen or collagen was added to the wells, and incubated for 24 h at 4\u0026deg;C, on an orbital shaker. Afterward, the wells were washed three times with TBS (pH 7.5), and 200 \u0026micro;L of 1% BSA was added. The plates were incubated with BSA at 37\u0026deg;C for 2 h. Meanwhile, the extracts were added to washed blood platelets at final concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL. A control sample was set up containing only blood platelets with Barber\u0026rsquo;s buffer (a modified Tyrode\u0026rsquo;s buffer); this value was assumed to be 100%. The samples were incubated for 30 min. at 37\u0026deg;C. BSA was removed, and the wells were washed three times with TBS (pH 7.5) with 0.1 mM CaCl\u003csub\u003e2\u003c/sub\u003e and 0.1 mM MgCl\u003csub\u003e2\u003c/sub\u003e. The samples were added to the wells in triplicate. Agonists (1 U/mL thrombin or 30 \u0026micro;M ADP) or TBS were added to appropriate wells. Then, the plates were incubated for 1h at 37\u0026deg;C. Afterward, the wells were washed three times with PBS. 150 \u0026micro;L of 0.1 M citrate buffer (pH 5.4) with 5 mM \u003cem\u003ep\u003c/em\u003e-nitrophenyl phosphate and 0.1% Triton X-100 was added; the samples were incubated for 1 h at room temperature. Lastly, 100 \u0026micro;L of 2M NaOH was added to the wells. Absorbance was read at 405 nm with SPECTROstar Nano Microplate Reader (BMG LABTECH, Germany\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\n \u003ch2\u003e2.8. Flow cytometry analysis\u003c/h2\u003e\n \u003cp\u003ePlatelet activation was assessed based on platelet-derived microparticles (PMP) formation, and the exposition of P-selectin (CD62P) and the active form of GPIIb/IIIa on the surface of blood platelets. To measure the exposition of P-selectin and the active form of GPIIb/IIIa, full blood was mixed with the extracts at the concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL and incubated for 15 min. at 37\u0026deg;C. After 15 minutes, agonists were added to the samples (10 \u0026micro;M ADP, 20 \u0026micro;M ADP, or 10 \u0026micro;g/mL collagen), and the incubation continued for another 15 min. (at 37\u0026deg;C). After the incubation, the samples were diluted 10 times with PBS and incubated with antibodies (CD61-PerCP, CD62P-PE, and PAC-1-FITC, 3 \u0026micro;L of each antibody incubated with 10 \u0026micro;L of diluted samples). The samples were stained for 30 minutes in the dark, at room temperature. Afterward, the samples were fixed with 1% CellFix for 1 h at 37\u0026deg;C. The samples were analyzed with a LSR II Flow Cytometer (Becton Dickinson, San Diego, CA, USA); at least 5000 CD61-PerCP-positive objects were recorded. Blood platelets were gated based on an FSC/SSC (forward scatter/side scatter) plot and positive staining with CD61-PerCP. Then, the percent of CD62-PE and PAC-1-FITC positive platelets was calculated in each of the samples.\u003c/p\u003e\n \u003cdiv class=\"BlockQuote\"\u003e\n \u003cp\u003eTo measure the formation of platelet-derived microparticles, the samples were incubated with \u003cem\u003eS. chinensis\u003c/em\u003e extract for 15 minutes at 37\u0026deg;C. Then, 20 \u0026micro;g/mL collagen was added, and the incubation continued for another 15 minutes. The samples were diluted 10 times with PBS and stained with 2 \u0026micro;L of CD61-PE antibody for 30 minutes in the dark, at room temperature. The samples were fixed with 1% CellFix for 1 hour and analyzed with LSR II Flow Cytometer (Becton Dickinson, San Diego, CA, USA). At least 10000 CD61-PE-positive objects were recorded. PMPs were distinguished from CD61-PE-positive objects based on an FSC/SSC plot on a log/log scale. The analysis was carried out with Floreada software (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://floreada.io\u003c/span\u003e\u003c/span\u003e, accessed on 26.02.2024).\u003c/p\u003e\n \u003c/div\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec10\" class=\"Section2\"\u003e\n \u003ch2\u003e2.9. Total Thrombus-formation Analysis System (T-TAS) (PL-chip)\u003c/h2\u003e\n \u003cp\u003eThe T-TAS system can measure the thrombus formation process in semi-physiological conditions. Here, the PL-chip which measures only primary hemostasis was used. The results were recorded as AUC\u003csub\u003e10\u003c/sub\u003e (Area Under the Curve) - area under the flow pressure curve recorded for 10 min. after the start of the test. The AUC\u003csub\u003e10\u003c/sub\u003e depicts the growth, intensity, and stability of thrombus formation. Data was depicted as % of control. Further information about the assay can be found in (Hosokawa et al., \u003cspan class=\"CitationRef\"\u003e2011\u003c/span\u003e). First, the extracts were incubated with human full blood at final concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL (37\u0026deg;C, 30 minutes). A control sample with 0.9% NaCl instead of the extract was set up. Then, 320 \u0026micro;L of each sample was added to the PL-chip, and pressure was recorded for 10 minutes or until it reached 60 kPa (Time of Occlusion). The results were depicted as % of the control sample.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\n \u003ch2\u003e2.10. Measurement of prothrombin time, thrombin time, and activated partial thromboplastin time\u003c/h2\u003e\n \u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eCoagulation times were determined coagulometrically, according to the method described by (\u003c/span\u003eMalinowska et al., \u003cspan class=\"CitationRef\"\u003e2012\u003c/span\u003e). \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eThe extract was incubated with human plasma at 37\u0026deg;C for 30 minutes, at final concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL. The measurements were carried out on an Optic Coagulation Analyser, model K-3002 (Kselmed, Grudziadz, Poland). Each sample was measured in duplicate.\u003c/span\u003e\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\n \u003ch2\u003e2.11. Activity of LDH\u003c/h2\u003e\n \u003cp\u003eThe toxic effect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries on human blood platelets was analyzed by measuring the activity of lactate dehydrogenase (LDH), an enzyme released from damaged blood platelets (Wr\u0026oacute;blewski \u0026amp; Ladue, \u003cspan class=\"CitationRef\"\u003e1955\u003c/span\u003e). Blood platelets were incubated with the extract at final concentrations of 0.5\u0026ndash;50 \u0026micro;g/mL, at 37\u0026deg;C for 30 min. Afterward, the samples were centrifuged (2500 rpm, 15 min., 25\u0026deg;C) and 10 \u0026micro;l of the supernatant was added to the wells of a 96-well plate (in triplicate). Then, 0.1 M phosphate buffer (pH 7.4, 270 \u0026micro;L) and 0.25% nicotinamide adenine dinucleotide (NADH) (10 \u0026micro;l) were added to each well. A blank which contained 280 \u0026micro;L of phosphate buffer and 10 \u0026micro;L of NADH was set up as well. The samples were mixed and incubated for 20 minutes at room temperature. Lastly, 0.25% pyruvate (10 \u0026micro;L) was added to the samples. The samples were immediately mixed, and the absorbance was read at 1 minute intervals for 10 minutes, at 340 nm.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\n \u003ch2\u003e2.12. Statistical analysis\u003c/h2\u003e\n \u003cp\u003eStatistical analysis was performed with Statistica 10 (StatSoft 13.3, TIBCO Software Inc. Palo Alto, CA, USA). The distribution of data was checked by the Shapiro-Wilk test, and the homogeneity of variance by Levene\u0026rsquo;s test. Differences within and between groups were assessed with one-way ANOVA followed by Tukey\u0026rsquo;s test, or Kruskall-Wallis test. Results were presented as means\u0026thinsp;\u0026plusmn;\u0026thinsp;SD. The results were considered significant at p\u0026thinsp;\u0026le;\u0026thinsp;0.05. Dixon\u0026rsquo;s Q-test was used to eliminate uncertain data.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\n \u003ch2\u003e3.1. Chemical characteristic of the extract from S. chinensis berries\u003c/h2\u003e\n \u003cdiv class=\"BlockQuote\"\u003e\n \u003cp\u003eThe extract contained many lignans, which were the major phenolic constituents (and more generally, main specialized metabolites) of the \u003cem\u003eS. chinensis\u003c/em\u003e fruit extract (Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). Among them, schisandrin (tentatively identified) was the dominant compound. The red color of the extract can be attributed to the presence of cyanidin- Pen-Hex-dHex. \u003cem\u003eS. chinensis\u003c/em\u003e fruits also contained lesser amounts of (epi)catechin, dimeric and trimeric B-type proanthocyanidins, quercetin hexoside-deoxyhexoside, and quercetin hexoside. Apart from phenolics, small amounts of highly oxygenated nortriterpenoids (mainly), bisnortriterpenoids, triterpenoids, and homotriterpenoids were also detected (Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). Moreover, the extract contained also very high amounts of diverse highly polar constituents (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u0026nbsp;\u003ctable id=\"Tab1\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eSpecialized metabolites of the \u003cem\u003eS. chinensis\u003c/em\u003e fruit extract.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eNo.\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003etR\u003c/p\u003e\n \u003cp\u003e(min.)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eparent ion\u003c/p\u003e\n \u003cp\u003e(\u003cem\u003em/z\u003c/em\u003e)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCID\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003edelta (ppm)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003em\u0026sigma;\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eformula\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003etentative identification\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eRef.\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1\u0026ndash;3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.85\u0026ndash;1.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003emixtures of polar compounds\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e203.0823\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e186.0552 (5), 159.0921 (7), 142.0641 (7), 116.0595 (17)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e11\u003c/sub\u003eH\u003csub\u003e12\u003c/sub\u003eN\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003etryptophan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e577.1352\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e407.0773 (90), 289.0720 (100), 245.0821 (23)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e30\u003c/sub\u003eH\u003csub\u003e26\u003c/sub\u003eO\u003csub\u003e12\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(epi)C-(epi)C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e289.0719\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e289.0720 (100), 245.0819 (73), 203.03715 (31)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e15\u003c/sub\u003eH\u003csub\u003e14\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(epi)catechin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e865.1982\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e695.1400 (4), 525.0822 (15), 407.0785 (83), 289.0729 (100), 243.0309 (39)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e45\u003c/sub\u003eH\u003csub\u003e38\u003c/sub\u003eO\u003csub\u003e18\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(epi)C-(epi)C-(epi)C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e725.1927\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e339.0514 (14), 284.0329 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e32\u003c/sub\u003eH\u003csub\u003e39\u003c/sub\u003eO\u003csub\u003e19\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ecyanidin-Pen-Hex-dHex\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e441.1769\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e441.1769 (78), 397.1854 (28), 330.1319 (100), 217.1227 (30)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e21\u003c/sub\u003eH\u003csub\u003e30\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e609.1460\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e300.0279 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e27\u003c/sub\u003eH\u003csub\u003e30\u003c/sub\u003eO\u003csub\u003e16\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003erutin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.87\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e463.0882\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e300.0276 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e21\u003c/sub\u003eH\u003csub\u003e20\u003c/sub\u003eO\u003csub\u003e12\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eQ-3-\u003cem\u003eO\u003c/em\u003e-Hex\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e685.2927\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e639.2851 (12), 477.2348 (24), 383.1193 (100), 323.0979 (10), 263.0774 (15), 221.0664 (31), 179.0557 (28)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e48\u003c/sub\u003eO\u003csub\u003e16\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified bisnorterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e605.2232\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2182 (100), 497.2176 (65), 439.1759 (45), 351.1965 (31), 317.1391 (22)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e575.2131\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e575.2138 (100), 557.2020 (20), 531.2235 (3), 455.1728 (10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e12\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.74\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2179\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2209 (100), 541.2087 (65), 523.2031 (50), 465.1557 (47), 447.1440 (55), 439.1785 (70), 421.1656(81)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18.22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e605.2239\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2181 (100), 497.2173 (88), 479.2080 (43), 439.1762 (41), 351.1961 (42), 317.1391 (34)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2189\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2196 (100), 541.2082 (19), 465.1531 (5), 445.1495 (6), 439.1760 (7)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18.85\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e621.2194\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e575.2142 (100), 557.2041 (28), 511.2317 (8), 455.1724 (9), 417.1558 (12)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e12\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e605.2241\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2188 (100), 541.2083 (14), 445.1504 (6), 347.1493 (3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e589.2296\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e543.2244 (47), 525.2134 (60), 507.2025(100), 481.2219 (60), 463.2128 (89), 419.2235 (94), 383.1863 (43)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20.24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e605.2242\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2184 (100), 541.2082 (40), 515.2278 (32), 497.2186 (49)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20.62\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e591.2453\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e545.2415 (2), 447.2161 (20), 411.1817 (25), 367.1917 (100)349.1815 (22)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e38\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e605.2240\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2186 (100), 497.1289 (14), 457.1862 (19)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e589.2295\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e543.2245 (100), 525.2143 (46), 499.2354 (38), 481.2238 (77), 407.1863 (26)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e593.2603\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e547.2556 (6), 529.2449 (25), 511.2336 (14), 485.2538 (16), 467.2442 (100), 427.2138 (14)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e40\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e577.2292\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e531.2238 (100), 513.2118 (11), 451.2120 (16), 297.1485 (9)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ewuweizidilactone H / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e2\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.87\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e603.208\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e557.2037 (100)481.1507 (17), 455.1716 (55), 437.1604 (13), 365.1393 (7)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e579.2447\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e533.2387 (100), 497.2195 (29), 475.2005 (35), 453.2270 (28), 435.2166 (24)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e38\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified bisnorterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e619.2757\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e573.2733 (1), 531.2596 (100), 513.2490 (16), 495.2399 (8), 211.0975 (9)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e31\u003c/sub\u003eH\u003csub\u003e42\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified homotriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.79\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e633.2554\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e587.2515 (1), 509.2182 (29), 491.2076 (13), 465.2284 (19), 447.2180 (100), 429.2077 (21)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e31\u003c/sub\u003eH\u003csub\u003e40\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified homotriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.79\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e603.2088\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e557.2032 (100), 499.1618 (10), 481.1509 (18), 455.1707 (56), 437.1600 (13)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e635.2710\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e589.2654 (4), 571.2546 (21), 509.2544 (100), 491.2440 (14), 467.2435 (54), 449.2329 (34)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e31\u003c/sub\u003eH\u003csub\u003e42\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified homotriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e25.36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e575.2476\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e557.2372 (11), 497.2162 (27), 479.2057 (69)461.1951 (33), 437.1953 (51), 419.1846 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e30\u003c/sub\u003eH\u003csub\u003e38\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified triterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e25.47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e587.2126\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e541.2071 (100), 483.1655 (16), 465.1552 (20), 439.1759 (34), 365.1394 (12), 215.0715 (4)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified nortriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e26.75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e577.2638\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e559.2533 (46), 541.2434 (29), 499.2316 (49), 481.2218 (100), 463.2107 (53), 439.2112 (66), 421.2004 (95)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e30\u003c/sub\u003eH\u003csub\u003e40\u003c/sub\u003eO\u003csub\u003e11\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified triterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.02\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e619.2764\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e573.2706 (71), 531.2591 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e31\u003c/sub\u003eH\u003csub\u003e42\u003c/sub\u003eO\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified homotriterpenoid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e28.12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e433.2218\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e415.2112 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e24\u003c/sub\u003eH\u003csub\u003e32\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e28.93\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e701.2813\u003csup\u003e\u0026minus;\u003c/sup\u003e*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e531.2215 (66), 513.2110 (100), 495.2043 (55), 477.1932 (32), 451.2151 (41), 433.2003 (64), 415.1912 (66)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e35\u003c/sub\u003eH\u003csub\u003e44\u003c/sub\u003eO\u003csub\u003e12\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e29.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e389.1960\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e389.1963 (100), 357.1699 (7), 319.1180 (3), 287.0920 (3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e22\u003c/sub\u003eH\u003csub\u003e28\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003egomisin J / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e29.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e417.1911\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e399.1804 (100), 369.1694 (13)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e23\u003c/sub\u003eH\u003csub\u003e28\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrol B / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e41\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e391.2116\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e391.2115 (100), 237.1485 (16)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e22\u003c/sub\u003eH\u003csub\u003e30\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003epregomisin / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e501.2482\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e483.2379 (65), 401.1961 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eangeloylgomisin H / tigloylgomisin H / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e43\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e501.2483\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e483.2377 (26), 401.1960 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eangeloylgomisin H / tigloylgomisin H/ isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e523.2321\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e505.2221 (100), 401.1960 (49)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e30\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ebenzoylgomisin H / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e548.2850\u003csup\u003e+#\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e431.2063 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e29\u003c/sub\u003eH\u003csub\u003e38\u003c/sub\u003eO\u003csub\u003e9\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eangeloylgomisin Q / tigloylgomisin Q / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e554.2387\u003csup\u003e+#\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e415.1754 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e30\u003c/sub\u003eH\u003csub\u003e32\u003c/sub\u003eO\u003csub\u003e9\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisantherin A / gomisin G\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e30.99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e532.2542\u003csup\u003e+#\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e415.1758 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e9\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisantherin B / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e31.12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e532.2542\u003csup\u003e+#\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e415.1755 (100), 371.1491 (7)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e34\u003c/sub\u003eO\u003csub\u003e9\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003egomisin F / angeloylgomisin P / tigloylgomisin P\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e31.32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e403.2118\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e403.2120 (100), 371.1857 (6)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e23\u003c/sub\u003eH\u003csub\u003e30\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisanhenol / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32.08\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e417.2275\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e24\u003c/sub\u003eH\u003csub\u003e32\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin A (deoxyschisandrin) / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e401.1962\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e401.1964 (100), 331.1183 (5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e23\u003c/sub\u003eH\u003csub\u003e28\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003egomisin N / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e401.1962\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e401.1965 (100), 331.1183 (5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e23\u003c/sub\u003eH\u003csub\u003e28\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin B (\u0026gamma;-schisandrin) / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e4\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32.49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e385.1649\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e385.1648 (100), 355.1544 (5), 315.0871 (5), 285.0765 (3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e22\u003c/sub\u003eH\u003csub\u003e24\u003c/sub\u003eO\u003csub\u003e6\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin C / isomer\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003e3\u003c/strong\u003e, \u003cstrong\u003e5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e32.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e485.2536\u003csup\u003e+\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e485.2538 (100), 403.2116 (32)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-0.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e28\u003c/sub\u003eH\u003csub\u003e36\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eunidentified\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e33.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e279.2326\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e279.2326 (100)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eC\u003csub\u003e18\u003c/sub\u003eH\u003csub\u003e32\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eoctadecadienoic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"9\"\u003e\n \u003cp\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e - negative ion mode; + - positive ion mode; * - formic acid adduct; \u003csup\u003e#\u003c/sup\u003e - NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e adduct; C \u0026ndash; catechin, Q \u0026ndash; quercetin, dHex \u0026ndash; deoxyhexose; Hex \u0026ndash; hexose; Pen \u0026ndash; pentose\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"9\"\u003e\n \u003cp\u003e\u003cstrong\u003e1\u003c/strong\u003e. Ma et al., \u003cspan class=\"CitationRef\"\u003e2012\u003c/span\u003e; \u003cstrong\u003e2\u003c/strong\u003e. Li et al., \u003cspan class=\"CitationRef\"\u003e2017\u003c/span\u003e; \u003cstrong\u003e3\u003c/strong\u003e. He et al., \u003cspan class=\"CitationRef\"\u003e1997\u003c/span\u003e; \u003cstrong\u003e4\u003c/strong\u003e. Yang et al., \u003cspan class=\"CitationRef\"\u003e2011\u003c/span\u003e; \u003cstrong\u003e5\u003c/strong\u003e. Liu et al., \u003cspan class=\"CitationRef\"\u003e2013\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cp\u003eResults of the quantitative analysis of phenolic compounds are shown in Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e. As mentioned above, lignans were dominant phenolic compounds of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e fruit. The total lignan content was 29.61\u0026thinsp;\u0026plusmn;\u0026thinsp;0.18 mg g\u003csup\u003e-1\u003c/sup\u003e of the extract (expressed as schisandrin equivalent). The total flavonoid content was 1.64\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03 mg g\u003csup\u003e-1\u003c/sup\u003e (expressed as rutin equivalent), and the content of anthocyanidins was 17.08\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35 mg g\u003csup\u003e-1\u003c/sup\u003e (expressed as cyanidin equivalent).\u003c/p\u003e\n \u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u0026nbsp;\u003ctable id=\"Tab2\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eContent of major phenolic compounds in the extract from the fruit of \u003cem\u003eS. chinensis\u003c/em\u003e.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003etR (min.)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003em/z\u003c/em\u003e\u003csup\u003e\u003cem\u003e*\u003c/em\u003e\u003c/sup\u003e\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eidentification\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003econtent (mg g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ecyanidin-Pen-Hex-dHex\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.08\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003csup\u003e$\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e611\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003erutin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.12\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4.32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e465\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003equercetin-Hex\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.52\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003e#\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11.39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e433\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23.07\u0026thinsp;\u0026plusmn;\u0026thinsp;0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e389\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003egomisin J\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003etraces\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11.99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e417\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrol B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.16\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003csup\u003e^\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e501\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eangeloylgomisin H / tigloylgomisin H\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.61\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003csup\u003e^\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.81\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e401\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.12\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11\u003csup\u003e^\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e385\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eschisandrin C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003csup\u003e^\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"4\"\u003e\n \u003cp\u003e* - positive ion mode; \u003cspan\u003e$\u003c/span\u003e - cyanidin equivalent; # - rutin equivalent; ^ - schisandrin equivalent; Hex - hexose; dHex - deoxyhexose; Pen - pentose;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003c/div\u003e\u003cspan\u003e\n \u003cp\u003e\u003cem\u003e3.2. Effect of the extract from S. chinensis berries on the adhesion of washed blood platelets to collagen and fibrinogen\u003c/em\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eThe anti-adhesive activity of the extract was studied using human washed blood platelets. The results were presented as percent of adhesion of the control samples. As shown in Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003eA and B the level of adhesion of resting blood platelets and adhesion of thrombin-activated platelets was significantly reduced in the presence of all used concentrations of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (0.5\u0026ndash;50 \u0026micro;g/mL). For example, at the highest concentration (50 \u0026micro;g/mL), the extract inhibited the adhesion of thrombin-activated blood platelets by about 50% in comparison with control (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003eB). On the other hand, none of the tested concentrations of the extract (0.5\u0026ndash;50 \u0026micro;g/mL) demonstrated anti-adhesive properties, when adhesion of thrombin-activated platelets to fibrinogen was measured (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003eC).\u003c/p\u003e\n \u003cp\u003eFor the ADP-activated blood platelets, three tested concentrations of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (0.5, 1, and 5 \u0026micro;g/mL) did not have anti-adhesive activity (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003eD). Reduced adhesion was only observed for the two highest concentrations (10 and 50 \u0026micro;g/mL) (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003eD).\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cem\u003e3.3. Effect of the extract from S. chinensis berries on parameters of blood platelet activation measured with flow cytometry in whole blood\u003c/em\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eFigures \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e and \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e demonstrate the parameters of activation of platelets stimulated with 10 and 20 \u0026micro;M ADP, and 10 \u0026micro;g/mL collagen (measured with flow cytometry). Changes in platelet activation were noted in whole blood treated with the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries at all tested concentrations (0.5\u0026ndash;50 \u0026micro;g/mL), but these changes were not always statistically significant (Figs. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e and \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e). For example, the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (at all used concentrations: 0.5\u0026ndash;50 \u0026micro;g/mL) inhibited the exposition of the active form of GPIIb/IIIa on the surface of blood platelets stimulated with 10 \u0026micro;M ADP (Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003eA). In blood platelets stimulated with 20 \u0026micro;M ADP, statistically significant reduction of the exposition of the active form of GPIIb/IIIa was observed only for the highest concentration of the extract (50 \u0026micro;g/mL) (Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003eB). On the other hand, none of the concentrations of the extract (0.5\u0026ndash;50 \u0026micro;g/mL) significantly impacted the exposition of GPIIb/IIIa on platelets stimulated by 10 \u0026micro;g/mL collagen, and the exposition of P-selectin on the surface of blood platelets stimulated by 10 and 20 \u0026micro;M ADP (Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003eC, \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003eA and B). However, in platelets activated by collagen, significant inhibition of the exposition of P-selectin was observed for three concentrations of the extract (5, 10, and 50 \u0026micro;g/mL) (Fig. \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003eC).\u003c/p\u003e\n \u003cp\u003eMoreover, none of the used concentrations (0.5\u0026ndash;50 \u0026micro;g/mL) of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries significantly impacted platelet-derived microparticle formation and the exposition of GPIIb/IIIa and P-selectin on resting platelets (data not presented).\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cem\u003e3.4. Effect of the extract from S. chinensis berries on thrombus formation with T-TAS (PL-chip) in whole blood\u003c/em\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eOnly the highest concentration of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (50 \u0026micro;g/mL) significantly decreased the AUC\u003csub\u003e10\u003c/sub\u003e value measured by T-TAS in whole blood (Fig. \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cem\u003e3.5. Effect of the extract from S. chinensis berries on coagulation parameters (PT, TT, and APTT) in plasma\u003c/em\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eThe analysis of the effect of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries on the coagulation times of human plasma showed that none of the used concentrations of the extract (0.5\u0026ndash;50 \u0026micro;g/mL) changed APTT, PT, and TT (data not demonstrated).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\n \u003ch2\u003e3.6. Effect of the extract from S. chinensis berries on a marker of blood platelet damage\u003c/h2\u003e\n \u003cp\u003eTo determine the toxic effect of the extract on blood platelets, the level of LDH activity was measured. The results demonstrate no significant difference in platelet viability after exposure to \u003cem\u003eS. chinensis\u003c/em\u003e berry extract at all used concentrations (0.5\u0026ndash;50 \u0026micro;g/mL) (data not presented).\u003c/p\u003e\n \u003cp\u003eTable \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e compares the effects of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries, the extract from sea buckthorn berries, and the extract from aronia berries (as a positive control) (50 \u0026micro;g/mL) on selected markers of blood platelet activation. The strongest anti-platelet activity was demonstrated by the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries and the extract from aronia berries. The anti-platelet potential was observed for six markers.\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n \u003ctable id=\"Tab3\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eA comparison of the anti-platelet activity of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries (50 \u0026micro;g/mL), the extract from sea buckthorn berries (50 \u0026micro;g/mL), and the extract from aronia berries (50 \u0026micro;g/mL) in washed blood platelets (measured by adhesion to adhesive proteins (collagen and fibrinogen) and in whole blood (measured by T-TAS and flow cytometry).\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eS. chinensis\u003c/em\u003e berries\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eSea buckthorn berries\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eAronia berries\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of adhesion of thrombin-activated platelet to collagen\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of adhesion of thrombin-activated platelet to fibrinogen\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of adhesion of ADP-activated platelet to fibrinogen\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of thrombus formation (measured by T-TAS)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of GPIIb/IIIa exposition in 10 \u0026micro;M ADP-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of GPIIb/IIIa exposition in 20 \u0026micro;M ADP-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of GPIIb/IIIa exposition in collagen-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of P-selectin exposition in 10 \u0026micro;M ADP-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of P-selectin exposition in 20 \u0026micro;M ADP-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eInhibition of P-selectin exposition in collagen-activated platelets\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNo effect\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnti-platelet activity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eOur LC-HRMS analysis showed that \u003cem\u003eS. chinensis\u003c/em\u003e berry extract contained diverse lignans, which were its most important phenolic constituents. Most of these tentatively identified compounds were dibenzocyclooctadiene lignans, except for pregomisin, which belongs to the dibenzylbutane type. The presence of dibenzocyclooctadiene lignans is a characteristic feature of Schisandraceae family; they are regarded as the main bioactive constituents of \u003cem\u003eS. chinensis\u003c/em\u003e (S. Yang \u0026amp; Yuan, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Schisandrin was the dominant compound; putative schisandrol B, angelogomisin H, deoxyschisandrin, and schisandrin B were other major lignans. Our results are mostly in line with other studies (He et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; Liu et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Sheng et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Yang et al., \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). The red color of \u003cem\u003eS. chinensis\u003c/em\u003e berries is caused by an anthocyanin, cyanidin- Pen-Hex-dHex. According to the literature data, this compound is most probably cyanidin 3-\u003cem\u003eO\u003c/em\u003e-xylosylrutinoside (Liao et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Ma et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). Flavonoids, represented rutin and quercetin hexoside, occurred in small quantities. Our results are supported by the work of (Mocan et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), who found small amounts of rutin, quercetin 3-\u003cem\u003eO\u003c/em\u003e-glucoside, and quercetin 3-\u003cem\u003eO\u003c/em\u003e-galactoside in \u003cem\u003eS. chinensis\u003c/em\u003e berries. Apart from phenolics, the extract also contained numerous terpenoid compounds, mainly highly oxygenated nortriterpenoids, as well as bisnortriterpenoids, triterpenoids and homotriterpenoids, all present in small amounts. Substances with identical or similar molecular formulas were previously isolated from the aerial parts or fruits of \u003cem\u003eS. chinensis\u003c/em\u003e (Huang et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Li et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Shi et al., \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; S. Yang \u0026amp; Yuan, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). One of the detected bisnortriterpenoids was tentatively identified (on the basis of its determined formula) as wuweizidilactone H; this compound was earlier purified from the berry of \u003cem\u003eS. chinensis\u003c/em\u003e by other research groups (Li et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Xue et al., \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2010\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eHemostasis is a complex process depending on many interconnecting factors, among which blood platelets play a key role (Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Khodadi, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Platelets are small (approximately 2\u0026ndash;4 \u0026micro;m), discoid, anucleate blood elements. They can activate due to contact with various agonists, like thrombin, ADP, thromboxane A\u003csub\u003e2\u003c/sub\u003e, or collagen (Gremmel et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Rubenstein \u0026amp; Yin, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Tomaiuolo et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Upon activation, platelets change their shape and expose various proteins that aid in the coagulation process. One of those proteins is Pselectin, which in resting platelets is located in α granules. After activation, α granules fuse with the cell membrane, which exposes P-selectin on the surface of platelets, allowing for their adhesion to leukocytes and/or endothelial cells. Because P-selectin is exposed only on the surface of activated platelets, it is often used as an activation marker (Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Rubenstein \u0026amp; Yin, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). GPIIb/IIIa (integrin α\u003csub\u003eIIb\u003c/sub\u003eβ\u003csub\u003e3\u003c/sub\u003e) is another protein that plays a vital role in hemostasis. In resting platelets, GPIIb/IIIa is exposed on the cell membrane in its low affinity state, and transforms into a high affinity (active) form upon platelet activation. GPIIb/IIIa plays a key role in aggregation, as it binds to fibrinogen, which in turn binds to GPIIb/IIIa on other platelets, facilitating the formation of platelet aggregates (Gremmel et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Rubenstein \u0026amp; Yin, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Tomaiuolo et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Currently, there are two GPIIb/IIIa inhibitors used as anti-platelet drugs - tirofiban and eptifibatide (Tummala \u0026amp; Rai, \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Platelet adhesion is mediated mainly by GPIb-IX-V (which binds to von Willebrand factor (vWF), which in turn binds to collagen) and GPIV, which binds to collagen directly (Gremmel et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Rubenstein \u0026amp; Yin, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Tomaiuolo et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Another change that occurs in platelets during activation is increased shedding of platelet-derived microparticles (PMPs), also known as microvesicles. PMPs are 1 \u0026micro;m or less but are bigger than exosomes (30\u0026ndash;100 nm). Their plasma membrane exposes negatively charged phospholipids and a number of platelet receptors, including P-selectin, GPIIb/IIIa, and vWF (Nignpense et al., 2019; Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eCardiovascular diseases are a leading cause of death worldwide. An imbalance in hemostasis which leads to excessive clotting is a major issue in many CVDs and can lead to severe conditions, like myocardial infarction or stroke (Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Khodadi, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Increased platelet activation plays a major role in the pathology of many CVDs and is associated with adverse prognosis. To prevent this, many cardiovascular patients must take anti-platelet medication, among which acetylsalicylic acid (aspirin) is used most often. Aspirin inhibits cyclooxygenase (COX) activity, decreasing the production of thromboxane A\u003csub\u003e2\u003c/sub\u003e (Kannan et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Unfortunately, aspirin and many other anti-platelet drugs can cause adverse effects, including bleeding from the gastrointestinal tract. For this reason, researchers are searching for new anti-platelet compounds with a lower risk of side effects (Gremmel et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Olas, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe most important aspect of our findings is the confirmation of the anti-platelet activity of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries in various \u003cem\u003ein vitro\u003c/em\u003e models. We used flow cytometry and TTAS to study platelet activation in whole blood, which is a more natural environment than media in which blood platelets are suspended after isolation. For the first time, we noted that \u003cem\u003eS. chinensis\u003c/em\u003e extract (at the highest tested concentration \u0026ndash; 50 \u0026micro;g/mL) significantly prolonged the time of occlusion, showing anti-platelet activity \u003cem\u003ein vitro\u003c/em\u003e. We also observed that all the tested\u003c/p\u003e \u003cp\u003econcentrations (0.5\u0026ndash;50 \u0026micro;g/mL) of the extract inhibit the exposition of the active form of GPIIb/IIIa in 10 \u0026micro;M ADP-stimulated platelets. In addition, we observed the anti-adhesive potential of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries using an \u003cem\u003ein vitro\u003c/em\u003e model based on human washed blood platelets. Results of (Chang et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2005\u003c/span\u003e) also demonstrated that the extract from \u003cem\u003eS. chinensis\u003c/em\u003e inhibits arachidonic acid-induced blood platelet aggregation. Moreover, they suggested that the inhibition of cyclooxygenase is its primary mechanism of action. On the other hand, our results demonstrate that S. \u003cem\u003echinensis\u003c/em\u003e berry extract did not influence plasma coagulation times.\u003c/p\u003e \u003cp\u003eVarious studies showed that lignans (including schisandrin, schisandrin A, B, and C) isolated from \u003cem\u003eS. chinensis\u003c/em\u003e are its main active constituents, and possess a variety of nutritional properties and biological activities (Jia et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Wang et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Zhou et al., \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). We propose that the most active component of \u003cem\u003eS. chinensis\u003c/em\u003e berry extract may be schisandrin, which could be the major determinant of its anti-platelet activity \u003cem\u003ein vitro\u003c/em\u003e. Other \u003cem\u003ein vitro\u003c/em\u003e and \u003cem\u003ein vivo\u003c/em\u003e models have also demonstrated that schisandrin has cardioprotective properties (Gong et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Zhang et al., \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). For example, (Gong et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) observed its cardioprotective activity in a myocardial ischemia/reperfusion injury murine model (\u003cem\u003ein vivo\u003c/em\u003e) and H9c2 cardiomyocyte cell line subjected to hypoxia/reoxygenation injury (\u003cem\u003ein vitro\u003c/em\u003e). (Zhang et al., \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) also found that schisandrin promotes the recovery of myocardial tissues by enhancing cell viability and migration. Recently, more information about the protective effect of schisandrin on the cardiovascular system has been described in a review paper by (Wang et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). However, it did not cover the effect of this compound on blood platelets.\u003c/p\u003e \u003cp\u003eThe bioavailability and toxicity of various plant preparations (including extracts and chemical compounds) that could be used as drugs or supplements are a crucial element in the evaluation of their biological properties. In our study, none of the used concentrations of the extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries caused damage to human blood platelets (which was determined by a measurement of LDH that leaked out from the damaged cells into the extracellular medium). These results indicate, that this extract should be safe for use as a natural supplement with anti-platelet activity, but it still lacks validation in clinical settings. However, it has been found that extract from \u003cem\u003eS. chinensis\u003c/em\u003e berries has little to no toxicity toward various animals, including mice, rats, pigs, and rabbits (Panossian \u0026amp; Wikman, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; K. Yang et al., \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Moreover, the results of (Chen et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2018\u003c/span\u003e) demonstrated that it did not have toxic effects in an atherosclerosis rat model.\u003c/p\u003e \u003cp\u003eResearchers have incorporated schisandrin as a key ingredient in various formulations, including tablets, capsules, liquids, and injections, to investigate its efficacy. Interestingly, schisandrin exhibited a good level of absorption. Hydroxylation and demethylation pathways are its main metabolic modifications (Wang et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eAnother active components that we identified in \u003cem\u003eS. chinensis\u003c/em\u003e berries are triterpenoids, which display a wide range of biological activities, including antitumor, antiviral, hepatoprotective, neuroprotective, and anti-inflammatory (Jia et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Moreover, other studies indicate that triterpenoids and their derivatives present in fractions and extracts from various organs of sea buckthorn have anti-platelet activity (Skalski et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2021\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2023\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOther compounds that can significantly affect the circulatory system, including blood platelets, are phenolic compounds. They occur in large quantities in many plant species, including sea buckthorn and aronia berries (Olas, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2017\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Olas et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Skalski et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Some mechanisms of their anti-platelet activity are the inhibition of cyclooxygenase and blocking the binding of surface receptors to adhesion proteins. An additional advantage of supplementation with phenolic compounds is the lack of side effects (Luo et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Olas, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTo conclude, our results indicate that \u003cem\u003eS. chinensis\u003c/em\u003e berries could be used to produce natural nutrient supplements with cardioprotective potential, including anti-platelet activity. Future investigations into this extract and its chemical components, including schisandrin, should prioritize the following areas: (1) further understanding of the molecular mechanisms underlying its anti-platelet action, and the identification of specific targets; (2) bridging the gaps in knowledge about its \u003cem\u003ein vivo\u003c/em\u003e efficiency by using animal models and performing clinical studies on healthy people and patients with different CVDs.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eADI - Acceptable Daily Intake;\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eAPTT - activated partial thromboplastin time; BSA -\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003ebovine serum albumin; CPDA - citrate/phosphate/dextrose/adenine;\u0026nbsp;CVDs \u0026ndash; cardiovascular diseases;\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eFDA - Food and Drug Administration;\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eLDH\u003cstrong\u003e\u0026nbsp;-\u0026nbsp;\u003c/strong\u003elactate dehydrogenase; PBS - Phosphate buffered saline; PT - prothrombin time; T-TAS - Total Thrombus-formation Analysis System; TT - thrombin time; WHO -\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eThe World Health Organization\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e: The Authors would like to thank Mariusz Kowalczyk, PhD for performing UHPLC-HRMS analyses.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest:\u003c/strong\u003e the authors declare no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe data availability:\u003c/strong\u003e \u003cstrong\u003eThe datasets used and/or analysed during the current study available from the corresponding author on reasonable request.\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eN.S. and J.Z. - methodology, formal analysis, investigation, writing-original draft, prepared figures; B.L., J.B., and K. Z. - methodology, formal analysis, investigtion; B.M.Sz. , P.B., and A.S. - methodology; B.O. - conceptualization, writing-rewiew and editing.All authors reviewed the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBellavite, P., Andrioli, G., Guzzo, P., Arigliano, P., Chirumbolo, S., Manzato, F., \u0026amp; Santonastaso, C. (1994). A Colorimetric Method for the Measurement of Platelet Adhesion in Microtiter Plates. \u003cem\u003eAnalytical Biochemistry\u003c/em\u003e, \u003cem\u003e216\u003c/em\u003e(2), 444\u0026ndash;450. https://doi.org/10.1006/abio.1994.1066\u003c/li\u003e\n\u003cli\u003eChang, G.-T., Kang, S.-K., Kim, J.-H., Chung, K.-H., Chang, Y.-C., \u0026amp; Kim, C.-H. (2005). Inhibitory effect of the Korean herbal medicine, Dae-Jo-Whan, on platelet-activating factor-induced platelet aggregation. \u003cem\u003eJournal of Ethnopharmacology\u003c/em\u003e, \u003cem\u003e102\u003c/em\u003e(3), 430\u0026ndash;439. https://doi.org/10.1016/j.jep.2005.07.003\u003c/li\u003e\n\u003cli\u003eChen, X., Cao, J., Sun, Y., Dai, Y., Zhu, J., Zhang, X., Zhao, X., Wang, L., Zhao, T., Li, Y., Liu, Y., Wei, G., Zhang, T., \u0026amp; Yan, Z. (2018). Ethanol extract of \u003cem\u003eSchisandrae chinensis fructus\u003c/em\u003e ameliorates the extent of experimentally induced atherosclerosis in rats by increasing antioxidant capacity and improving endothelial dysfunction. \u003cem\u003ePharmaceutical Biology\u003c/em\u003e, \u003cem\u003e56\u003c/em\u003e(1), 612\u0026ndash;619. https://doi.org/10.1080/13880209.2018.1523933\u003c/li\u003e\n\u003cli\u003eChun, J. N., Cho, M., So, I., \u0026amp; Jeon, J.-H. (2014). The protective effects of Schisandra chinensis fruit extract and its lignans against cardiovascular disease: A review of the molecular mechanisms. \u003cem\u003eFitoterapia\u003c/em\u003e, \u003cem\u003e97\u003c/em\u003e, 224\u0026ndash;233. https://doi.org/10.1016/j.fitote.2014.06.014\u003c/li\u003e\n\u003cli\u003eGong, S., Liu, J., Wan, S., Yang, W., Zhang, Y., Yu, B., Li, F., \u0026amp; Kou, J. (2021). Schisandrol A Attenuates Myocardial Ischemia/Reperfusion-Induced Myocardial Apoptosis through Upregulation of 14-3-3\u0026theta;. \u003cem\u003eOxidative Medicine and Cellular Longevity\u003c/em\u003e, \u003cem\u003e2021\u003c/em\u003e, 1\u0026ndash;15. https://doi.org/10.1155/2021/5541753\u003c/li\u003e\n\u003cli\u003eGremmel, T., Frelinger, A., \u0026amp; Michelson, A. (2016). Platelet Physiology. \u003cem\u003eSeminars in Thrombosis and Hemostasis\u003c/em\u003e, \u003cem\u003e42\u003c/em\u003e(03), 191\u0026ndash;204. https://doi.org/10.1055/s-0035-1564835\u003c/li\u003e\n\u003cli\u003eHe, X.-G., Lian, L.-Z., \u0026amp; Lin, L.-Z. (1997). Analysis of lignan constituents from Schisandra chinensis by liquid chromatography-electrospray mass spectrometry. \u003cem\u003eJournal of Chromatography A\u003c/em\u003e, \u003cem\u003e757\u003c/em\u003e, 81\u0026ndash;87.\u003c/li\u003e\n\u003cli\u003eHosokawa, K., Ohnishi, T., Kondo, T., Fukasawa, M., Koide, T., Maruyama, I., \u0026amp; Tanaka, K. A. (2011). A novel automated microchip flow-chamber system to quantitatively evaluate thrombus formation and antithrombotic agents under blood flow conditions. \u003cem\u003eJournal of Thrombosis and Haemostasis\u003c/em\u003e, \u003cem\u003e9\u003c/em\u003e(10), 2029\u0026ndash;2037. https://doi.org/10.1111/J.1538-7836.2011.04464.X\u003c/li\u003e\n\u003cli\u003eHuang, S.-X., Han, Q.-B., Lei, C., Pu, J.-X., Xiao, W.-L., Yu, J.-L., Yang, L.-M., Xu, H.-X., Zheng, Y.-T., \u0026amp; Sun, H.-D. (2008). Isolation and characterization of miscellaneous terpenoids of Schisandra chinensis. \u003cem\u003eTetrahedron\u003c/em\u003e, \u003cem\u003e64\u003c/em\u003e(19), 4260\u0026ndash;4267. https://doi.org/10.1016/j.tet.2008.02.085\u003c/li\u003e\n\u003cli\u003eJia, M., Zhou, L., Lou, Y., Yang, X., Zhao, H., Ouyang, X., \u0026amp; Huang, Y. (2023). An analysis of the nutritional effects of Schisandra chinensis components based on mass spectrometry technology. \u003cem\u003eFrontiers in Nutrition\u003c/em\u003e, \u003cem\u003e10\u003c/em\u003e. https://doi.org/10.3389/fnut.2023.1227027\u003c/li\u003e\n\u003cli\u003eKannan, M., Ahmad, F., \u0026amp; Saxena, R. (2019). Platelet activation markers in evaluation of thrombotic risk factors in various clinical settings. \u003cem\u003eBlood Reviews\u003c/em\u003e, \u003cem\u003e37\u003c/em\u003e, 100583. https://doi.org/10.1016/j.blre.2019.05.007\u003c/li\u003e\n\u003cli\u003eKhodadi, E. (2020). Platelet Function in Cardiovascular Disease: Activation of Molecules and Activation by Molecules. \u003cem\u003eCardiovascular Toxicology\u003c/em\u003e \u003cem\u003e20\u003c/em\u003e(1), 1-10 https://doi.org/10.1007/s12012-019-09555-4\u003c/li\u003e\n\u003cli\u003eLi, F., Zhang, T., Sun, H., Gu, H., Wang, H., Su, X., Li, C., Li, B., Chen, R., \u0026amp; Kang, J. (2017). A New Nortriterpenoid, a Sesquiterpene and Hepatoprotective Lignans Isolated from the Fruit of Schisandra chinensis. \u003cem\u003eMolecules\u003c/em\u003e, \u003cem\u003e22\u003c/em\u003e(11), 1931. https://doi.org/10.3390/molecules22111931\u003c/li\u003e\n\u003cli\u003eLiao, J., Zang, J., Yuan, F., Liu, S., Zhang, Y., Li, H., Piao, Z., \u0026amp; Li, H. (2016). Identification and analysis of anthocyanin components in fruit color variation in \u003cem\u003eSchisandra chinensis\u003c/em\u003e. \u003cem\u003eJournal of the Science of Food and Agriculture\u003c/em\u003e, \u003cem\u003e96\u003c/em\u003e(9), 3213\u0026ndash;3219. https://doi.org/10.1002/jsfa.7503\u003c/li\u003e\n\u003cli\u003eLin, Q., Qin, X., Shi, M., Qin, Z., Meng, Y., Qin, Z., \u0026amp; Guo, S. (2017). Schisandrin B inhibits LPS-induced inflammatory response in human umbilical vein endothelial cells by activating Nrf2. \u003cem\u003eInternational Immunopharmacology\u003c/em\u003e, \u003cem\u003e49\u003c/em\u003e, 142\u0026ndash;147. https://doi.org/10.1016/j.intimp.2017.05.032\u003c/li\u003e\n\u003cli\u003eLis, B., Rolnik, A., Jedrejek, D., Soluch, A., Stochmal, A., \u0026amp; Olas, B. (2019). Dandelion (\u003cem\u003eTaraxacum officinale\u003c/em\u003e L.) root components exhibit anti-oxidative and antiplatelet action in an in vitro study. \u003cem\u003eJournal of Functional Foods\u003c/em\u003e, \u003cem\u003e59\u003c/em\u003e, 16\u0026ndash;24. https://doi.org/10.1016/j.jff.2019.05.019\u003c/li\u003e\n\u003cli\u003eLiu, H., Lai, H., Jia, X., Liu, J., Zhang, Z., Qi, Y., Zhang, J., Song, J., Wu, C., Zhang, B., \u0026amp; Xiao, P. (2013). Comprehensive chemical analysis of \u003cem\u003eSchisandra chinensis\u003c/em\u003e by HPLC\u0026ndash;DAD\u0026ndash;MS combined with chemometrics. \u003cem\u003ePhytomedicine\u003c/em\u003e, \u003cem\u003e20\u003c/em\u003e(12), 1135\u0026ndash;1143. https://doi.org/10.1016/j.phymed.2013.05.001\u003c/li\u003e\n\u003cli\u003eLuo, X., Du, C., Cheng, H., Chen, J., \u0026amp; Lin, C. (2017). Study on the Anticoagulant or Procoagulant Activities of Type II Phenolic Acid Derivatives. \u003cem\u003eMolecules\u003c/em\u003e, \u003cem\u003e22\u003c/em\u003e(12), 2047. https://doi.org/10.3390/molecules22122047\u003c/li\u003e\n\u003cli\u003eMa, C., Yang, L., Yang, F., Wang, W., Zhao, C., \u0026amp; Zu, Y. (2012). Content and Color Stability of Anthocyanins Isolated from Schisandra chinensis Fruit. \u003cem\u003eInternational Journal of Molecular Sciences\u003c/em\u003e, \u003cem\u003e13\u003c/em\u003e(12), 14294\u0026ndash;14310. https://doi.org/10.3390/ijms131114294\u003c/li\u003e\n\u003cli\u003eMalinowska, J., Kołodziejczyk-Czepas, J., Moniuszko-Szajwaj, B., Kowalska, I., Oleszek, W., Stochmal, A., \u0026amp; Olas, B. (2012). Phenolic fractions from Trifolium pallidum and Trifolium scabrum aerial parts in human plasma protect against changes induced by hyperhomocysteinemia in vitro. \u003cem\u003eFood and Chemical Toxicology: An International Journal Published for the British Industrial Biological Research Association\u003c/em\u003e, \u003cem\u003e50\u003c/em\u003e(11), 4023\u0026ndash;4027. https://doi.org/10.1016/J.FCT.2012.07.065\u003c/li\u003e\n\u003cli\u003eMocan, A., Crișan, G., Vlase, L., Crișan, O., Vodnar, D., Raita, O., Gheldiu, A.-M., Toiu, A., Oprean, R., \u0026amp; Tilea, I. (2014). Comparative Studies on Polyphenolic Composition, Antioxidant and Antimicrobial Activities of \u003cem\u003eSchisandra chinensis\u003c/em\u003e Leaves and Fruits. \u003cem\u003eMolecules\u003c/em\u003e, \u003cem\u003e19\u003c/em\u003e(9), 15162\u0026ndash;15179. https://doi.org/10.3390/molecules190915162\u003c/li\u003e\n\u003cli\u003eEd Nignpense, B., Chinkwo, K. A., Blanchard, C. L., \u0026amp; Santhakumar, A. B. (2019). Polyphenols: Modulators of Platelet Function and Platelet Microparticle Generation? \u003cem\u003eInternational Journal of Molecular Sciences\u003c/em\u003e, \u003cem\u003e21\u003c/em\u003e(1), 146. https://doi.org/10.3390/ijms21010146\u003c/li\u003e\n\u003cli\u003eOlas, B. (2017). The multifunctionality of berries toward blood platelets and the role of berry phenolics in cardiovascular disorders. \u003cem\u003ePlatelets\u003c/em\u003e, \u003cem\u003e28\u003c/em\u003e(6), 540\u0026ndash;549. https://doi.org/10.1080/09537104.2016.1235689\u003c/li\u003e\n\u003cli\u003eOlas, B. (2018). Berry Phenolic Antioxidants \u0026ndash; Implications for Human Health? \u003cem\u003eFrontiers in Pharmacology\u003c/em\u003e, \u003cem\u003e9\u003c/em\u003e(MAR), 78. https://doi.org/10.3389/fphar.2018.00078\u003c/li\u003e\n\u003cli\u003eOlas, B. (2020). Biochemistry of blood platelet activation and the beneficial role of plant oils in cardiovascular diseases. In \u003cem\u003eAdvances in Clinical Chemistry\u003c/em\u003e (Vol. 95, pp. 219\u0026ndash;243). Academic Press Inc. https://doi.org/10.1016/bs.acc.2019.08.006\u003c/li\u003e\n\u003cli\u003eOlas, B. (2023). Cardioprotective Potential of Berries of \u003cem\u003eSchisandra chinensis\u003c/em\u003e Turcz. (Baill.), Their Components and Food Products. \u003cem\u003eNutrients\u003c/em\u003e, \u003cem\u003e15\u003c/em\u003e(3). https://doi.org/10.3390/nu15030592\u003c/li\u003e\n\u003cli\u003eOlas, B., Wachowicz, B., Tomczak, A., Erler, J., Stochmal, A., \u0026amp; Oleszek, W. (2008). Comparative anti-platelet and antioxidant properties of polyphenol-rich extracts from: berries of \u003cem\u003eAronia melanocarpa\u003c/em\u003e, seeds of grape and bark of \u003cem\u003eYucca schidigera in vitro\u003c/em\u003e. \u003cem\u003ePlatelets\u003c/em\u003e, \u003cem\u003e19\u003c/em\u003e(1), 70\u0026ndash;77. https://doi.org/10.1080/09537100701708506\u003c/li\u003e\n\u003cli\u003ePanossian, A., \u0026amp; Wikman, G. (2008). Pharmacology of Schisandra chinensis Bail.: An overview of Russian research and uses in medicine. \u003cem\u003eJournal of Ethnopharmacology\u003c/em\u003e, \u003cem\u003e118\u003c/em\u003e(2), 183\u0026ndash;212. https://doi.org/10.1016/j.jep.2008.04.020\u003c/li\u003e\n\u003cli\u003eRubenstein, D. A., \u0026amp; Yin, W. (2018). Platelet‐Activation Mechanisms and Vascular Remodeling. In \u003cem\u003eComprehensive Physiology\u003c/em\u003e (Vol. 8, Issue 3, pp. 1117\u0026ndash;1156). Wiley. https://doi.org/10.1002/cphy.c170049\u003c/li\u003e\n\u003cli\u003eSheng, Y., Wang, R., Wang, Y., Li, M., Liu, F., Huang, X., \u0026amp; Chen, C. (2022). Evaluation of Multicomponent Changes of \u003cem\u003eSchisandra chinensis\u003c/em\u003e Fruits with Different Drying Process by UPLC-QQQ-MS-Based Targeted Metabolomics Analysis. \u003cem\u003eJournal of Chemistry\u003c/em\u003e, \u003cem\u003e2022\u003c/em\u003e, 1\u0026ndash;10. https://doi.org/10.1155/2022/2616122\u003c/li\u003e\n\u003cli\u003eShi, Y.-M., Wang, L.-Y., Zou, X.-S., Li, X.-N., Shang, S.-Z., Gao, Z.-H., Liang, C.-Q., Luo, H.-R., Li, H.-L., Xiao, W.-L., \u0026amp; Sun, H.-D. (2014). Nortriterpenoids from Schisandra chinensis and their absolute configurational assignments by electronic circular dichroism study. \u003cem\u003eTetrahedron\u003c/em\u003e, \u003cem\u003e70\u003c/em\u003e(4), 859\u0026ndash;868. https://doi.org/10.1016/j.tet.2013.12.023\u003c/li\u003e\n\u003cli\u003eSkalski, B., Rywaniak, J., Szustka, A., Żuchowski, J., Stochmal, A., \u0026amp; Olas, B. (2021). Anti-Platelet Properties of Phenolic and Nonpolar Fractions Isolated from Various Organs of \u003cem\u003eElaeagnus rhamnoides\u003c/em\u003e (L.) A. Nelson in Whole Blood. \u003cem\u003eInternational Journal of Molecular Sciences\u003c/em\u003e, \u003cem\u003e22\u003c/em\u003e(6), 3282. https://doi.org/10.3390/ijms22063282\u003c/li\u003e\n\u003cli\u003eSkalski, B., Rywaniak, J., Żuchowski, J., Stochmal, A., \u0026amp; Olas, B. (2023). The changes of blood platelet reactivity in the presence of \u003cem\u003eElaeagnus rhamno\u003c/em\u003eides (L.) A. Nelson leaves and twig extract in whole blood. \u003cem\u003eBiomedicine \u0026amp; Pharmacotherapy\u003c/em\u003e, \u003cem\u003e162\u003c/em\u003e, 114594. https://doi.org/10.1016/j.biopha.2023.114594\u003c/li\u003e\n\u003cli\u003eSkalski, B., Stochmal, A., Żuchowski, J., Grabarczyk, Ł., \u0026amp; Olas, B. (2020). Response of blood platelets to phenolic fraction and non-polar fraction from the leaves and twigs of \u003cem\u003eElaeagnus rhamnoides \u003c/em\u003e(L.) A. Nelson in vitro. \u003cem\u003eBiomedicine \u0026amp; Pharmacotherapy\u003c/em\u003e, \u003cem\u003e124\u003c/em\u003e, 109897. https://doi.org/10.1016/j.biopha.2020.109897\u003c/li\u003e\n\u003cli\u003eSławińska, N., \u0026amp; Olas, B. (2022). Selected Seeds as Sources of Bioactive Compounds with Diverse Biological Activities. \u003cem\u003eNutrients\u003c/em\u003e, \u003cem\u003e15\u003c/em\u003e(1), 187. https://doi.org/10.3390/nu15010187\u003c/li\u003e\n\u003cli\u003eSławińska, N., Żuchowski, J., Stochmal, A., \u0026amp; Olas, B. (2023). Extract from Sea Buckthorn Seeds\u0026mdash;A Phytochemical, Antioxidant, and Hemostasis Study; Effect of Thermal Processing on Its Chemical Content and Biological Activity \u003cem\u003eIn Vitro\u003c/em\u003e. \u003cem\u003eNutrients\u003c/em\u003e, \u003cem\u003e15\u003c/em\u003e(3), 686. https://doi.org/10.3390/nu15030686\u003c/li\u003e\n\u003cli\u003eSu, D., Luo, J., Ge, J., Liu, Y., Jin, C., Xu, P., Zhang, R., Zhu, G., Yang, M., Ai, Z., \u0026amp; Song, Y. (2022). Raw and Wine Processed Schisandra chinensis Regulate NREM-Sleep and Alleviate Cardiovascular Dysfunction Associated with Insomnia by Modulating HPA Axis. \u003cem\u003ePlanta Medica\u003c/em\u003e, \u003cem\u003e88\u003c/em\u003e(14), 1311\u0026ndash;1324. https://doi.org/10.1055/a-1721-4971\u003c/li\u003e\n\u003cli\u003eSzopa, A., Ekiert, R., \u0026amp; Ekiert, H. (2017). Current knowledge of \u003cem\u003eSchisandra chinensis\u003c/em\u003e (Turcz.) Baill. (Chinese magnolia vine) as a medicinal plant species: a review on the bioactive components, pharmacological properties, analytical and biotechnological studies. \u003cem\u003ePhytochemistry Reviews\u003c/em\u003e, \u003cem\u003e16\u003c/em\u003e(2), 195\u0026ndash;218. https://doi.org/10.1007/s11101-016-9470-4\u003c/li\u003e\n\u003cli\u003eSzopa, A., Klimek, M., \u0026amp; Ekiert, H. (2016). Chinese magnolia vine (\u003cem\u003eSchisandra chinensis\u003c/em\u003e) - therapeutic and cosmetic importance. \u003cem\u003ePolish Journal of Cosmetology\u003c/em\u003e, \u003cem\u003e19\u003c/em\u003e(4), 274\u0026ndash;284.\u003c/li\u003e\n\u003cli\u003eTomaiuolo, M., Brass, L. F., \u0026amp; Stalker, T. J. (2017). Regulation of Platelet Activation and Coagulation and Its Role in Vascular Injury and Arterial Thrombosis. \u003cem\u003eInterventional Cardiology Clinics\u003c/em\u003e, \u003cem\u003e6\u003c/em\u003e(1), 1\u0026ndash;12. https://doi.org/10.1016/j.iccl.2016.08.001\u003c/li\u003e\n\u003cli\u003eTummala, R., \u0026amp; Rai, M. P. (2024). \u003cem\u003eGlycoprotein IIb/IIIa Inhibitors\u003c/em\u003e.\u003c/li\u003e\n\u003cli\u003eVal\u0026iacute;čkov\u0026aacute;, J., Zezulka, \u0026Scaron;., Mar\u0026scaron;\u0026aacute;lkov\u0026aacute;, E., Kotl\u0026iacute;k, J., Mar\u0026scaron;\u0026aacute;lek, B., \u0026amp; Opatřilov\u0026aacute;, R. (2023). Bioactive compounds from \u003cem\u003eSchisandra chinensis\u003c/em\u003e - Risk for aquatic plants? \u003cem\u003eAquatic Toxicology (Amsterdam, Netherlands)\u003c/em\u003e, \u003cem\u003e254\u003c/em\u003e, 106365. https://doi.org/10.1016/j.aquatox.2022.106365\u003c/li\u003e\n\u003cli\u003eWang, X., Wang, X., Yao, H., Shen, C., Geng, K., \u0026amp; Xie, H. (2024). A comprehensive review on Schisandrin and its pharmacological features. \u003cem\u003eNaunyn-Schmiedeberg\u0026rsquo;s Archives of Pharmacology\u003c/em\u003e, \u003cem\u003e397\u003c/em\u003e(2), 783\u0026ndash;794. https://doi.org/10.1007/s00210-023-02687-z\u003c/li\u003e\n\u003cli\u003eWr\u0026oacute;blewski, F., \u0026amp; Ladue, J. S. (1955). Lactic dehydrogenase activity in blood. \u003cem\u003eProc. Soc. Exp. Biol. Med\u003c/em\u003e, \u003cem\u003e90\u003c/em\u003e, 210\u0026ndash;221.\u003c/li\u003e\n\u003cli\u003eWu, J., Jia, J., Liu, L., Yang, F., Fan, Y., Zhang, S., Yan, D., Bu, R., Li, G., Gao, Y., \u0026amp; Chen, Y. (2017). Schisandrin B displays a protective role against primary pulmonary hypertension by targeting transforming growth factor \u0026beta;1. \u003cem\u003eJournal of the American Society of Hypertension\u003c/em\u003e, \u003cem\u003e11\u003c/em\u003e(3), 148-157.e1. https://doi.org/10.1016/j.jash.2016.12.007\u003c/li\u003e\n\u003cli\u003eXue, Y.-B., Zhang, Y.-L., Yang, J.-H., Du, X., Pu, J.-X., Zhao, W., Li, X.-N., Xiao, W.-L., \u0026amp; Sun, H.-D. (2010). Nortriterpenoids and Lignans from the Fruit of Schisandra chinensis. \u003cem\u003eChemical and Pharmaceutical Bulletin\u003c/em\u003e, \u003cem\u003e58\u003c/em\u003e(12), 1606\u0026ndash;1611. https://doi.org/10.1248/cpb.58.1606\u003c/li\u003e\n\u003cli\u003eYang, J., IP, S. P., J. Yeung, H., \u0026amp; Che, C. (2011). HPLC-MS analysis of Schisandra lignans and their metabolites in Caco-2 cell monolayer and rat everted gut sac models and in rat plasma. \u003cem\u003eActa Pharmaceutica Sinica B\u003c/em\u003e, \u003cem\u003e1\u003c/em\u003e(1), 46\u0026ndash;55. https://doi.org/10.1016/j.apsb.2011.04.007\u003c/li\u003e\n\u003cli\u003eYang, K., Qiu, J., Huang, Z., Yu, Z., Wang, W., Hu, H., \u0026amp; You, Y. (2022). A comprehensive review of ethnopharmacology, phytochemistry, pharmacology, and pharmacokinetics of \u003cem\u003eSchisandra chinensis\u003c/em\u003e (Turcz.) Baill. and Schisandra sphenanthera Rehd. et Wils. \u003cem\u003eJournal of Ethnopharmacology\u003c/em\u003e, \u003cem\u003e284\u003c/em\u003e, 114759. https://doi.org/10.1016/j.jep.2021.114759\u003c/li\u003e\n\u003cli\u003eYang, S., \u0026amp; Yuan, C. (2021). \u003cem\u003eSchisandra chinensis\u003c/em\u003e: A comprehensive review on its phytochemicals and biological activities. \u003cem\u003eArabian Journal of Chemistry\u003c/em\u003e, \u003cem\u003e14\u003c/em\u003e(9), 103310. https://doi.org/10.1016/j.arabjc.2021.103310\u003c/li\u003e\n\u003cli\u003eZhang, X., Zhao, Y., Bai, D., Yuan, X., \u0026amp; Cong, S. (2019). Schizandrin protects H9c2 cells against lipopolysaccharide‐induced injury by downregulating Smad3. \u003cem\u003eJournal of Biochemical and Molecular Toxicology\u003c/em\u003e, \u003cem\u003e33\u003c/em\u003e(5). https://doi.org/10.1002/jbt.22301\u003c/li\u003e\n\u003cli\u003eZhou, Y., Men, L., Sun, Y., Wei, M., \u0026amp; Fan, X. (2021). Pharmacodynamic effects and molecular mechanisms of lignans from \u003cem\u003eSchisandra chinensis\u003c/em\u003e Turcz. (Baill.), a current review. \u003cem\u003eEuropean Journal of Pharmacology\u003c/em\u003e, \u003cem\u003e892\u003c/em\u003e, 173796. https://doi.org/10.1016/j.ejphar.2020.173796\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"blood platelet, coagulation, hemostasis, Schisandra chinensis","lastPublishedDoi":"10.21203/rs.3.rs-4346913/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4346913/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cem\u003eSchisandra chinensis\u003c/em\u003e Turcz. (Baill.) is a dioecious vine, belonging to the Schisandraceae family. Itsberries show several beneficial activities, including cardioprotective, antioxidant, and anti-inflammatory. We examined the chemical content of the extract from \u003cem\u003eS. chinensis \u003c/em\u003eberries, as well as its antiplatelet potential in washed human blood platelets and whole blood \u003cem\u003ein vitro\u003c/em\u003e. We assessed effect of the extract on several hemostasis parameters, including thrombus formation in full blood, platelet activation and adhesion, and coagulation times. Moreover, we evaluated the cytotoxicity of the extract against blood platelets based on extracellular lactate dehydrogenase (LDH) activity. The most important constituents of the extract were dibenzocyclooctadiene lignans; schisandrin was the dominant compound. The extract inhibited thrombus formation, agonist-stimulated platelet activation and adhesion, and was not cytotoxic. These results suggest that \u003cem\u003eS. chinensis\u003c/em\u003eberries can be used as a safe, natural supplement with anti-platelet properties. However, more studies are needed to determine their mechanisms of action and \u003cem\u003ein vivo\u003c/em\u003e efficiency.\u003c/p\u003e","manuscriptTitle":"Phytochemical analysis of the extract from berries of Schisandra chinensis Turcz. (Baill.) and its anti-platelet potential in vitro","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-05-13 17:24:01","doi":"10.21203/rs.3.rs-4346913/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"a066cecf-c42b-4a4b-b811-04a948f29d9a","owner":[],"postedDate":"May 13th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":31809432,"name":"Biological sciences/Biochemistry"},{"id":31809433,"name":"Biological sciences/Plant sciences"}],"tags":[],"updatedAt":"2024-07-09T08:10:16+00:00","versionOfRecord":[],"versionCreatedAt":"2024-05-13 17:24:01","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4346913","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4346913","identity":"rs-4346913","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}