水飞蓟果实、果皮及其提取物质量评价法的研究

目的建立水飞蓟果实、果皮及其提取物综合质量评价方法。方法用UV法在 2 87nm下测定水飞蓟素的含量 ;并以HPLC法测定指标成分水飞蓟宾的含量 :采用HypersilC1 8(2 5 0mm×4 6mm ,5 μm)色谱柱 ;以甲醇 乙腈 磷酸二氢铵缓冲液 (2 4∶2 4∶5 0 ,pH =5 )为流动相 ;流速为1 0mL min ;柱温为 4 0℃ ;检测波长为 2 87nm。结果UV法和HPLC法线性分别在 8 0 4～4 0 2 μmol L和 0 2 76～ 1 3 8μg;加样回收率 :UV法 98 5 % ,HPLC法 97 3 % ;水飞蓟果实、果皮中水飞蓟素和水飞蓟宾含量分别为 3 88%和 1 3 2 %、8 0 1 %和 2 77% ;3种水飞蓟提取物中水飞蓟素含量均在 80 %以上 ,但水飞蓟宾含量在 2 8 7%～ 3 2 8% ,有明显差异。结论 2法合用可用于水飞蓟原料药材及提取物的质量综合评价和控制     

Evidence for differences in regioselective and stereoselective glucuronidation of silybin diastereomers from milk thistle (Silybum marianum) by human UDP-glucuronosyltransferases

The flavonolignan silybin, the main component of silymarin, extract from the seeds of Silybum marianum, is used mostly as a hepatoprotectant. Silybin is almost 1:1 mixture of two diastereomers A and B. The individual UDP-glucuronosyltransferases (UGTs) contributing to the metabolism of silybin diastereomers have not been identified yet. In this study, the contribution of UGTs to silybin metabolism was examined. The potential silybin metabolites were formed in vitro by incubating silybin (i) with the human liver microsomal fraction, (ii) with human hepatocytes and finally (iii) with 12 recombinant UGTs (UGT1A1, 1A3, 1A4, 1A6, 1A7, 1A8, 1A9, 1A10, 2B4, 2B7, 2B15 and 2B17). High-performance liquid chromatographic (HPLC) techniques with UV detection and additionally MS detection were used for metabolite identification. Hepatocytes and microsomes formed silybin A-7-O-β-D-glucuronides, B-7-O-β-D-glucuronides, A-20-O-β-D-glucuronides and B-20-O-β-D-glucuronides. With recombinant UGTs, the major role of the UGT1A1, 1A3, 1A8 and 1A10 enzymes but also of the UGT1A6, 1A7, 1A9, 2B7 and 2B15 in the stereoselective reactions leading to the respective silybin glucuronides was confirmed. UGT1A4, UGT2B4 and UGT2B17 did not participate in silybin glucuronidation. The predominant formation of 7-O-β-D-glucuronides and the preferential glucuronidation of silybin B diastereomer in vitro by human UGTs were confirmed.     

Evidence for differences in regioselective and stereoselective glucuronidation of silybin diastereomers from milk thistle (Silybum marianum) by human UDP-glucuronosyltransferases

1. The flavonolignan silybin, the main component of silymarin, extract from the seeds of Silybum marianum, is used mostly as a hepatoprotectant. Silybin is almost 1:1 mixture of two diastereomers A and B. The individual UDP-glucuronosyltransferases (UGTs) contributing to the metabolism of silybin diastereomers have not been identified yet. In this study, the contribution of UGTs to silybin metabolism was examined.

2. The potential silybin metabolites were formed in vitro by incubating silybin (i) with the human liver microsomal fraction, (ii) with human hepatocytes and finally (iii) with 12 recombinant UGTs (UGT1A1, 1A3, 1A4, 1A6, 1A7, 1A8, 1A9, 1A10, 2B4, 2B7, 2B15 and 2B17). High-performance liquid chromatographic (HPLC) techniques with UV detection and additionally MS detection were used for metabolite identification.

3. Hepatocytes and microsomes formed silybin A-7-O-β-d-glucuronides, B-7-O-β-d-glucuronides, A-20-O-β-d-glucuronides and B-20-O-β-d-glucuronides. With recombinant UGTs, the major role of the UGT1A1, 1A3, 1A8 and 1A10 enzymes but also of the UGT1A6, 1A7, 1A9, 2B7 and 2B15 in the stereoselective reactions leading to the respective silybin glucuronides was confirmed. UGT1A4, UGT2B4 and UGT2B17 did not participate in silybin glucuronidation.

4. The predominant formation of 7-O-β-d-glucuronides and the preferential glucuronidation of silybin B diastereomer in vitro by human UGTs were confirmed.

     

Determination of T-2 and HT-2 Toxins in Seed of Milk Thistle [Silybum marianum (L.) Gaertn.] Using Immunoaffinity Column by UPLC-MS/MS

Milk thistle [Silybum marianum (L.) Gaertn.] achieved a significant increase in interest over the past few years from local and foreign pharmaceutical corporations. The silymarin complex of constituents extracted from milk thistle achenes provides compelling health benefits primarily thanks to antioxidant activities and hepatoprotective effects. However, consuming mycotoxin-contaminated plant material can cause immunosuppression and hepatotoxic problems. The aim of this study was to develop and validate a method for the determination of mycotoxin content in milk thistle. Fusarium toxins as T-2 and HT-2 toxins in grown milk thistle harvested from a breeding station in the Czech Republic during 2020–2021 were studied. The analysis of T-2 and HT-2 toxins was performed by UPLC-MS/MS after immunoaffinity columns EASI-EXTRACT^® T-2 & HT-2 clean up. All analysed samples of milk thistle were contaminated with T-2 toxin and HT-2 toxin. The content of T-2 toxin in the samples from 2020 was in the range of 122.7–290.2 µg/kg and HT-2 toxin 157.0–319.0 µg/kg. In 2021, the content of T-2 toxin was in the range of 28.8–69.9 µg/kg and HT-2 toxin was 24.2–75.4 µg/kg. The results show that the climatic conditions of the year of harvesting have a highly statistically significant effect on the content of T-2 and HT-2 toxins in milk thistle.     

Effect of the flavanolignans of Silybum marianum L. on lipid peroxidation in rat liver microsomes and freshly isolated hepatocytes.

The effect of several flavanolignans (silicristin, silidianin, silybin and isosilybin) present in silymarin, the extract of Silybum marianum fruits, was tested on lipid peroxidation in rat liver microsomes and freshly isolated hepatocytes. In microsomes lipid peroxidation was generated by ADP/Fe2+ and NADPH. All flavanolignans inhibited peroxidation in a concentration dependent manner. In hepatocytes lipid peroxidation was induced by ADP/Fe3+ complex and cell damage was evaluated as LDH activity released in the medium. The inhibition of the peroxidative process by flavanolignans was also evident in this model, even if with a potency order different from that found in microsomes. In contrast, the effect on LDH release was significant only for silybin and isosilybin, the other compounds being inactive on this parameter.     

The effect of silymarin (Silybum marianum) on human skin fibroblasts in an in vitro wound healing model

Context: Silymarin, a flavonolignan from Silybum marianum (L.) Gaertn. (Asteraceae), has been reported to have antioxidant and anti-inflammatory properties. Therefore, it may be worthwhile to study the effect of silymarin on wound healing.

Objective: To evaluate the effect of silymarin on human fibroblast cells in an in vitro model of wound healing.

Materials and methods: Human fibroblast cells were treated with different concentrations (4.5, 9, 18, 36 µg/mL) of silymarin. The effects of silymarin on cell viability, proliferation, collagen synthesis, and expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthetase (iNOS) were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, 5-bromo-2′-deoxy-uridine, hydroxyproline analysis and real-time PCR, respectively. The effect of silymarin on cellular antioxidant status was determined by protection against hydrogen peroxide (H₂O₂)-induced cell injury and free radical scavenging activity (ABTS assay) of the cells.

Results: Results of the present study indicate that pretreatment of fibroblast cells with silymarin significantly protected cells against H₂O₂-induced injury (p < 0.05). After an 18?h treatment of cells with 36 µg/mL silymarin, total antioxidant capacity of cells significantly increased (p < 0.05). Furthermore, pretreatment of human fibroblast cells with silymarin significantly inhibited lipopolysaccharide (LPS)-induced COX-2 mRNA expression (p < 0.001). There was no significant difference in fibroblast proliferation and collagen synthesis between treatment and control groups (p > 0.05).

Discussion and conclusion: Silymarin may be useful as a therapeutic agent for the treatment of cutaneous wounds through its antioxidation and anti-inflammation effects.     

Cancer Preventive Agents. 7. Antitumor-Promoting Effects of Seven Active Flavonolignans from Milk Thistle (Silybum marianum.) on Epstein-Barr Virus Activation

Abstract

Seven pure flavonolignans were isolated from an extract of milk thistle [Silybum marianum.(L.) Gaertn (Asteraceae)], by semipreparative reverse-phase HPLC, and identified based on spectroscopic and LC-MS/IT-TOF data. All seven compounds were screened as potential antitumor-promoting agents by using the in vitro. short-term 12-O.-tetradecanoylphorbol-13-acetate (TPA)-induced Epstein-Barr virus early antigen (EBV-EA) activation assay in Raji cells. They showed good inhibitory activity (87.7–94.9%) at 1000 mol ratio/TPA. Silychristin A (1) and silychristin B (2) were slightly more potent than the well-known antitumor promoter β.-carotene. Silychristins A (1) and B (2) and isosilybins A (6) and B (7) were more active than the clinically proven cancer prevention components silybins A (4) and B (5).     

苦荞麦种子的化学成分

目的对苦荞麦〔Fagopyrum tataricum(L.)Gaertn.〕的种子中的化学成分进行研究。方法采用硅胶柱色谱等方法进行分离纯化,根据理化性质和光谱数据进行结构鉴定。结果分离得到11个化合物,分别鉴定为乌苏酸(ursolic acid,1)、山柰酚(kaempferol,2)、槲皮素(quercetin,3)、大黄素(emodin,4)、山柰酚-3-O-芸香糖苷(kaempferol-3-O-rutinoside,5)、芦丁(rutin,6)、3′,5′-二甲氧基-4′-O-β-D-吡喃葡萄糖基桂皮酸(3′,5′-dimethoxy-4′-O-β-D-glucopyranosyl-cinnamic-acid,7)、β-谷甾醇(β-sitosterol,8)、胡萝卜苷(daucosterol,9)、蔗糖(sucrose,10)、果糖(fructose,11)。结论化合物7是从荞麦属中首次分离得到,化合物1是从该植物中首次分离得到。     

苦荞麦的化学成分

目的研究苦荞麦的化学成分。方法利用硅胶S、ephadex LH-20等柱色谱方法分离化学成分,根据理化性质和波谱数据分析鉴定化合物的结构。结果从苦荞麦种子中初步分离得到5个化合物,分别鉴定为乌苏酸(ursolic acid,1)、7-羟基香豆素(7-hydroxycoumarin,2)、大黄素(emodin,3)、尿嘧啶(uracil,4)、原儿茶酸(protocatechuic acid,5)。结论化合物2为首次从荞麦属植物中分离得到,化合物1、5为首次从苦荞麦中分离得到。     

A new sulphated nor-sesquiterpene from mangrove Laguncularia racemosa (L.) Gaertn. F

A new sulphated nor-sesquiterpene, 3alpha-hydroxysulphonyloxy-5alpha,6alpha-epoxy-megastigmen-9-one (1), and a known sulphated lipid, 5'-(hydroxysulphonyloxy) jasmonic acid (2), have been isolated from the twigs and leaves of mangrove Laguncularia racemosa (L.) Gaertn. F. The structure of the new compound was elucidated on the basis of detailed analysis of spectroscopic data and chemical reaction.     

The effect of silybin dihemisuccinate on cholesterol biosynthesis in rat liver homogenates (author's transl)

The effect of silybin dihemisuccinate on the hepatic biosynthesis of cholesterol was studied by measuring the incorporation of [1-14C]-acetate and [2-14C]-mevalonate in the postmitochondrial supernatant of liver homogenates. 2. Under absolute in vitro conditions, silybin dihemisuccinate inhibits the biosynthesis of cholesterol in dependence on its concentration. With [1-14C]-acetate or [2-14C]-mevalonate the concentrations of silybin for 50% inhibition are 0.37 mM or 0.40 mM, resp. 3. 30 min after i.v. injection of 150.6 mg silybin dihemisuccinate/kg with both precursors the biosynthesis of cholesterol studied in vitro is diminished in comparison to controls. 60 min or 24 h after i.v. injection of silybin no effects on in vitro biosynthesis of cholesterol can be observed.     

The clinical utility of milk thistle (Silybum marianum) in cirrhosis of the liver

Milk thistle (Silybum marianum) is a flowering herb utilized for its potentially protective effects on the liver. Although the mechanism of action is not fully understood, one explanation may be that it concentrates in the hepatocytes and competes with toxins for hepatocyte binding and penetration. Preliminary clinical evaluations of milk thistle for cirrhosis of the liver indicate potential benefits in healthier patients with alcoholic cirrhosis. However, major flaws in many of the studies make it difficult to draw solid conclusions. Milk thistle appears to be relatively safe, even with long-term use.     

磷脂酰胆碱对小鼠过氧化损伤的保护作用(英文)

目的：研究磷脂酰胆碱对小鼠过氧化损伤的保护作用。方法：以小鼠为试验动物,检测心、脑组织超氧化物歧化酶（SOD）活性,过氧化脂质（LPO）和脂褐素含量指标。结果：磷脂酰胆碱能有效提高心、脑组织超氧化物歧化酶（SOD）活性,降低过氧化脂质（LPO）和脂褐素含量。结论：磷脂酰胆碱具有明显的拮抗过氧化损伤、抗衰老作用。     

The influence of phosphatidyl serine on the release of histamine from isolated rat mast cells induced by different agents.

The influence of phosphatidyl serine (PS) on histamine release from isolated rat mast cells induced by antigen, compound 48/80, adenosine-5'-triphosphate (ATP), the ionophore A23187, and decylamine was studied. PS enhanced antigen-induced release but inhibited the release caused by compound 48/80, A23187, and decylamine. PS did not influence the release induced by ATP. The different effects of PS on the action of the various histamine releasing agents do not conform to a unifying model for the action of PS on the release process. Possible interactions between PS and the agents in the incubation medium as well as at specific reactive sites on the plasma membrane might explain some of the effects of PS. Consequently, the results cannot be used as evidence for the existence of basic differences in the release process induced by various calcium- and energy-dependent releasing agents.     

Effect of milk thistle (Silybum marianum) and black cohosh (Cimicifuga racemosa) supplementation on digoxin pharmacokinetics in humans.

Phytochemical-mediated modulation of P-glycoprotein (P-gp) and other drug transporters may underlie many herb-drug interactions. Serial serum concentration-time profiles of the P-gp substrate, digoxin, were used to determine whether supplementation with milk thistle or black cohosh modified P-gp activity in vivo. Sixteen healthy volunteers were randomly assigned to receive a standardized milk thistle (900 mg daily) or black cohosh (40 mg daily) supplement for 14 days, followed by a 30-day washout period. Subjects were also randomized to receive rifampin (600 mg daily, 7 days) and clarithromycin (1000 mg daily, 7 days) as positive controls for P-gp induction and inhibition, respectively. Digoxin (Lanoxicaps, 0.4 mg) was administered orally before and at the end of each supplementation and control period. Serial digoxin serum concentrations were obtained over 24 h and analyzed by chemiluminescent immunoassay. Comparisons of area under the serum concentration time curves from 0 to 3 h (AUC(0-3)), AUC(0-24), Cmax, apparent oral clearance of digoxin (CL/F), and elimination half-life were used to assess the effects of milk thistle, black cohosh, rifampin, and clarithromycin on digoxin pharmacokinetics. Rifampin produced significant reductions (p < 0.01) in AUC(0-3), AUC(0-24), and Cmax, whereas clarithromycin increased these parameters significantly (p < 0.01). Significant changes in digoxin half-life and CL/F were also observed with clarithromycin. No statistically significant effects on digoxin pharmacokinetics were observed following supplementation with either milk thistle or black cohosh, although digoxin AUC(0-3) and AUC(0-24) approached significance (p = 0.06) following milk thistle administration. When compared with rifampin and clarithromycin, supplementation with these specific formulations of milk thistle or black cohosh did not appear to affect digoxin pharmacokinetics, suggesting that these supplements are not potent modulators of P-gp in vivo.     

Effects of antiarrhythmic drugs on rat erythrocytes, isolated hepatocytes and dipalmitoyl phosphatidyl choline (DPPC)-liposomes

The effects of antiarrhythmic drugs, aprindine, mexiletine and lidocaine, on rat erythrocytes, isolated rat hepatocytes and DPPC-liposomes were studied at various concentrations. Maximal inhibition of aprindine on the hypotonic hemolysis was observed at a concentration of 2 X 10(-4) M. In isolated rat hepatocytes, aprindine caused an increase in GOT leakage above 4 X 10(-4) M. Mexiletine and lidocaine caused a slight decrease in GOT. Only aprindine caused an increase in LDH leakage above 2 X 10(-4) M. In the relationship between the surface tension and pH conditions (pH 5.7, 7.4 and 8.0), aprindine and mexiletine indicated a depression of surface tension at a dose of 10(-4) M to 10(-3) M under all pH conditions. Lidocaine indicated a depression of surface tension at a dose of 10(-4) M at pH 8.0 only. Aprindine and mexiletine depressed the phase transition temperature (Tc) of DPPC-liposomes. The depression of Tc by aprindine was greater than that by mexiletine. The rank by order of surface activity was the same as that of enzyme leakage from hepatocytes, hemolysis of erythrocytes and depression of Tc in DPPC-liposomes in vitro. These results suggest that differences in membrane damage produced by antiarrhythmic drugs may by related to surface activity, which in turn may determine the extent of adsorption onto cell membranes.