儿茶药材化学成分分析——反相高效液相色谱法测定儿茶中儿茶素和 …

目的：测定儿茶中主要成分儿茶素及表儿茶素的含量。方法：采用反相高效液相色谱法,以儿茶素及表儿茶素为对照品。结果：按选定的色谱条件,儿茶素及表儿茶素分别在０．２５６－１．２８,０．７２８－３．６４１μｇ范围内线性良好。２种成分的平均加样回收率分别为１００．１％和９６．０％,ＲＳＤ分别为２．４％和２．７％结论：本方法操作简便、快捷、重现性好。     

RP-HPLC测定家兔血浆中的儿茶素和表儿茶素

目的 采用RP-HPLC法测定家兔血浆中的儿茶素和表儿茶素.方法 色谱柱为Hypersil C18柱(250 mm×4.6 mm,10 μm),测定儿茶素和表儿茶的流动相分别为乙腈-水-三乙胺(8:92:0.3、11:89:0.3),流速均为1.0 ml·min-1,检测波长分别为206、204 nm,采用甲醇直接沉淀蛋白,取上清液进样.结果 家兔血浆中内源性成分测定无干扰;儿茶素0.68～6.50μg·ml-1呈良好的线性关系(r=0.9994),平均方法回收率为94.7%,日内和日间RSD分别为2.72%和2.84%;表儿茶素0.25～9.92 μg·ml-1呈良好的线性关系(r=0.9991),平均方法回收率为99.5%,日内和日间RSD分别为2.52%和5.92%.结论 所建方法简便,灵敏度较高,结果准确,可为儿茶药材及含儿茶中药制剂的体内研究提供参考.     

薄层色谱法鉴别儿茶中儿茶素和表儿茶素

采用薄层色谱法,对儿茶中儿茶素及表儿茶素进行了鉴别。     

HPLC法测定复方儿茶软膏中儿茶素和表儿茶素含量

目的:采用高效液相色谱法测定复方儿茶软膏中儿茶素和表儿茶素含量。方法:采用 DiamonsiL~(TM)C_(18)柱(4.6mm×250mm,5μm),以甲醇-0.04mol-L~(-1)柠檬酸溶液—N,N-二甲基甲酰胺(13:45:8)为流动相,流速1 mL·min~(-1),紫外检测器于280nm 测定。结果:HPLC 法测定,阴性对照品与样品分离良好,儿茶素和表儿茶素线性范围分别为10.5～105μg·mL~(-1)(r=0.9994)和6.6～66μg·mL~(-1)(r=0.9996),平均回收率分别为99.5%和99.6%,样品溶液在12h 内稳定。结论:本法简便,稳定,能有效控制该制剂的质量。     

HPLC法测定制痂酊中儿茶素和表儿茶素的含量

目的：建立高效液相色谱法同时测定制痂酊中儿茶素和表儿茶素的含量。方法：色谱柱：Hypersil C18柱（250mm×4．6mm,5μm）,流动相：2．2％冰醋酸一乙腈（90：10）,流速：0．8ml·min^-1,检测波长：280nm,柱温：35℃。结果：儿茶素和表儿茶素分别在0．10～0．80ml·min^-1、0．01—0．20mg·ml。之间呈良好的线性关系,平均回收率分别为101．1％（RSD=0．95％,n=6）和100．2％（RSD=1．90％,n=6）。结论：该方法快速、定量准确,可用于制痂酊的质量控制。     

HPLC法测定儿茶配方颗粒中儿茶素和表儿茶素的含量

目的:建立同时测定儿茶配方颗粒中儿茶素和表儿茶素含量的方法。方法:采用高效液相色谱法。色谱柱为Shim-pack CLC-ODS-C18,流动相为0.04 mol/L枸橼酸溶液-N,N-二甲基甲酰胺-四氢呋喃(45∶8∶2,V/V/V),流速为1.0 ml/min,柱温为35℃,检测波长为280 nm。结果:儿茶素和表儿茶素的进样量分别在0.301 6~1.508 0、0.202 0~1.010 0μg范围内与各自峰面积呈良好的线性关系(r=0.999 9、0.999 8);精密度、稳定性、重复性试验的RSD均<2%;平均加样回收率分别为96.60%(RSD=1.46%,n=9)、96.36%(RSD=1.30%,n=9)。结论:该方法简便、准确、重复性好,可用于测定儿茶配方颗粒中儿茶素和表儿茶素的含量。     

超声波法提取儿茶中总儿茶素工艺研究

目的 研究超声波法提取儿茶中儿茶素和表儿茶素的最佳工艺。方法 以儿茶素和表儿茶素的含量总和占儿茶重量的百分比为提取率作为考察指标,采用正交试验,考察乙醇浓度、料液比、提取温度和超声时间4个因素的影响。结果 超声波提取优化工艺条件为：乙醇浓度为50%,料液比为1∶12,超声时间为35 min,提取温度为60 ℃。结论 超声波提取儿茶中儿茶素和表儿茶素的工艺具有提取率高、时间短、溶剂用量少、能量消耗小等优点,适用于儿茶素和表儿茶素的提取。     

高效液相色谱法测定茶黄酊中儿茶素、表儿茶素和黄芩苷的含量

目的：建立高效液相色谱法测定茶黄酊中儿茶素、表儿茶素和黄芩苷含量。方法：采用Agilent ZORBAX Eclipse XDB—C18（150mm&#215;4．6mm，5μm）色谱柱；流动相A为0．5％三乙胺溶液（以磷酸调pH至3．0），流动相B为乙腈，梯度洗脱（0～12min，流动相B10％；12．1～30min，流动相B22％）；流速：1mL&#183;min^-1，检测波长278nm。结果：儿茶素、表儿茶素和黄芩苷线性范围分别为26．75—214．0μg&#183;mL^-1（r=0．9998），5．568～44．54μg&#183;mL^-1（r=0．9998），3．520～28．16μg&#183;mL^-1（r=0．9999）。回收率（n=6）分别为99．8％，98．9％，102．6％；RSD分别为0．61％，1．2％，0．92％。结论：本方法简便、准确、稳定，重复性好，可用于茶黄酊中儿茶素、表儿茶素和黄芩苷的含量测定。     

HPLC法同时测定复方儿茶酊中儿茶素与原儿茶素的含量

目的：建立同时测定复方儿茶酊中儿茶素与原儿茶素含量的方法。方法：采用高效液相色谱法。色谱柱为YMC ODS（250mm×4.6mm,5μm）,流动相为0.04mol·L-1枸橼酸溶液-N,N-二甲基甲酰胺-四氢呋喃（45∶8∶2,V/V/V）,流速为1.0mL·min-1,检测波长为280nm,柱温为35℃。结果：儿茶素与原儿茶素的检测浓度分别在0.0518～0.2590、0.0101～0.2030mg·mL-1（r均为0.9999）范围内与各自峰面积积分值呈良好的线性关系;平均加样回收率分别为101.63%和100.49%,RSD分别为1.27%和1.12%（n均为9）。结论：本方法简便、准确、稳定,能有效控制复方儿茶酊的质量。     

反相高效液相色谱法测定楤木属植物中黄酮类和香豆精的含量

目的：对木属植物中草本和木本两个自然群进行化学分类学的研究。方法：应用反相高效液相色谱法测定木属植物中４种化学成分［ｎｅｏｌｉｇｕｉｒｉｔｉｎ（Ｉ）,５,８ｄｉｍｅｔｈｏｘｙｃｏｕｍａｒｉｎ（ＩＩ）,ｉｓｏｌｉｑｕｒｉｔｏｓｉｄｅ（ＩＩ）,８［（２″,３″）ｐｒｅｎｙｌ］４′ｍｅｔｈｏｘｙｆｌａｖｏｎｅ７ＯβＤｇｌｕｃｏｐｙｒａｎｏｓｙｌ（２→１）αＬｒｈａｍｎｏｐｙｒａｎｏｓｉｄｅ（ＩＶ）］,对本属１１种植物中该４种成分进行了定量分析。色谱柱固定相为ＳｕｐｅｌｃｏＳＩＬＬＣ１８;流动相为４０％甲醇—甲醇梯度洗脱,根据４种成分紫外吸收波长的不同而改变检测波长;流速１０ｍｌ·ｍｉｎ－１;４种成分的线性范围分别为（０５６００～００１７５）μｇ,（０３７６０～００１１８）μｇ,（０２８７５～０００９０）μｇ和（０３０９０～０００９７）μｇ,回收率分别为９８９２％,９８２７％,９４７８％和９７４１％。结果：木本自然群中香豆精成分含量较高。结论：香豆精为木本自然群区别于草本自然群的特征成分之一     

Screening and Toxicity Analysis of Catechin Isomers Against FemA Protein

Fem proteins are the essential structural proteins of various gram-positive bacteria. These are of three different types namely FemX (FmhB), FemA and FemB. Only two Fem protein crystallographic structures are available till date, one for FemA in Staphylococcus aureus and another for FemX in Weissella viridescensis. In this study, computational methods are used to evaluate interaction of FemA protein with catechin and epicatechin analogues. The interaction of FemA protein with catechin and epicatechin analogues are confirmed by binding energy and scores given by Autodock Vina and UCSF Dock docking softwares, which is followed by Lipinski filters and toxicity studies using online Lipinski server of SCFBIO and OSIRIS. Catechin gallate has been found as the best ligand for FemA protein in all aspects and it has outperformed all catechin and epicatechin isomers.     

Development and validation of a LC-method for determination of catechin and epicatechin in aqueous extractives from leaves of Maytenus ilicifolia

A reverse phase-LC method was developed and validated for separation and quantification of catechin and epicatechin in aqueous extractives from leaves of Maytenus ilicifolia. The analysis was performed using a C(18) column with acetic acid-acetonitrile gradient elution. The detection was carried out by UV at 280 nm and the peak identification was based on the retention times and by co-chromatography with reference substances. High coefficients of determination were achieved for both catechin and epicatechin peaks from the standard solutions (0.9996 and 0.9999), as well as from extractives (0.9981 and 0.9982, respectively). The method showed good repeatability (R.S.D. <1.5%), reproducibility (R.S.D. <5%) and good accuracy for both catechin and epicatechin peaks (101.4%, R.S.D. = 1.18% and 100.6%, R.S.D. = 2.07%, respectively).     

Effects of (+)catechin and (−)epicatechin on heterocyclic amines‐induced oxidative DNA damage

The aim of the present study was to evaluate the protective effect of (+)‐catechin and (?)‐epicatechin against 2‐amino‐3,8‐ dimethylimidazo[4,5‐f]quinoxaline (8‐MeIQx), 2‐amino‐3,4,8‐trimethylimidazo[4,5‐f]‐quinoxaline (4,8‐diMeIQx) and 2‐amino‐1‐methyl‐6‐phenyl‐imidazo[4,5‐b]pyridine (PhIP)‐induced DNA damage in human hepatoma cells (HepG2). DNA damage (strand breaks and oxidized purines/pyrimidines) was evaluated by the alkaline single‐cell gel electrophoresis or comet assay. Increasing concentrations of 8‐MeIQx, 4,8‐diMeIQx and PhIP induced a significant increase in DNA strand breaks and oxidized purines and pyrimidines in a dose‐dependent manner. Among those, PhIP (300 µm ) exerted the highest genotoxicity. (+)‐Catechin exerted protection against oxidized purines induced by 8‐MeIQx, 4,8‐diMeIQx and PhIP. Oxidized pyrimidines and DNA strand breaks induced by PhIP were also prevented by (+)‐catechin. Otherwise, (?)‐epicatechin protected against the oxidized pyrimidines induced by PhIP and the oxidized purines induced by 8‐MeIQx and 4,8‐diMeIQx. One feasible mechanism by which (+)‐catechin and (?)‐epicatechin exert their protective effect towards heterocyclic amines‐induced oxidative DNA damage may be by modulation of phase I and II enzyme activities. The ethoxyresorufin O‐deethylation (CYP1A1) activity was moderately inhibited by (+)‐catechin, while little effect was observed by (?)‐epicatechin. However, (+)‐catechin showed the greatest increase in UDP‐glucuronyltransferase activity. In conclusion, our results clearly indicate that (+)‐catechin was more efficient than (?)‐epicatechin in preventing DNA damage (strand breaks and oxidized purines/pyrimidines) induced by PhIP than that induced by 8‐MeIQx and 4,8‐diMeIQx. Copyright © 2010 John Wiley & Sons, Ltd.     

Penicillium expansum Impact and Patulin Accumulation on Conventional and Traditional Apple Cultivars

Penicillium expansum is a necrotrophic plant pathogen among the most ubiquitous fungi disseminated worldwide. It causes blue mould rot in apples during storage, transport and sale, threatening human health by secreting patulin, a toxic secondary metabolite that contaminates apples and apple-derived products. Nevertheless, there is still a lack of sufficient data regarding the resistance of different apple cultivars to P. expansum, especially ancient ones, which showed to possess certain resistance to plant diseases. In this work, we investigated the polyphenol profile of 12 traditional and 8 conventional apple cultivar and their resistance to P. expansum CBS 325.48. Eight polyphenolic compounds were detected; the most prominent were catechin, epicatechin and gallic acid. The highest content of catechin was detected in ‘Apistar’—91.26 mg/100 g of fresh weight (FW), epicatechin in ‘Bobovac’—67.00 mg/100 g of FW, and gallic acid in ‘Bobovac’ and ‘Kraljevčica’—8.35 and 7.40 mg/100 g of FW, respectively. The highest content of patulin was detected in ‘Kraljevčica’ followed by ‘Apistar’—1687 and 1435 µg/kg, respectively. In apple cultivars ‘Brčko’, ‘Adamčica’ and ‘Idared’, patulin was not detected. Furthermore, the patulin content was positively correlated with gallic acid (r = 0.4226; p = 0.002), catechin (r = 0.3717; p = 0.008) and epicatechin (r = 0.3305; p = 0.019). This fact indicates that higher contents of gallic acid, catechin and epicatechin negatively affected and boost patulin concentration in examined apple cultivars. This can be related to the prooxidant activity of polyphenolic compounds and sensitivity of P. expansum to the disturbance of oxidative status.     

Flavonoids protect U-937 macrophages against tert-butylhydroperoxide induced oxidative injury.

The study was carried out to determine the relative efficacies of polyphenolic flavonoids, quercetin, catechin and epicatechin against tert-BOOH induced oxidative stress in human macrophage, U-937 cell line. Exposure of the cells to tert-BOOH oxidative stress resulted in a significant increase in cytotoxicity and reactive oxygen species (ROS) generation. Further, a significant decrease in mitochondrial membrane potential and increase in lipid peroxidation and DNA damage was observed in cells exposed to tert-BOOH. Pretreatment of cells with quercetin, catechin and epicatechin significantly inhibited tert-BOOH induced cytotoxicity by inhibiting ROS generation. The flavonoids inhibited DNA damage induced by tert-BOOH and preserved the mitochondrial transmembrane potential significantly. Epicatechin and catechin were found to be more efficient than quercetin in inhibiting tert-BOOH induced cellular damage.     

Effect of resveratrol and some other natural compounds on tyrosine kinase activity and on cytolysis.

Resveratrol is a phytoalexin with several biological and pharmacological activities including the "French paradox". We investigated the effect of resveratrol on cytolytic activity by oxygen reactive species and on soluble and particulate tyrosine kinases from human placenta and human prostatic adenoma. These effects were compared with those of piceatannol, quercetin, catechin and epicatechin. Fifty percent of erythrocyte lysis due to H2O2-lactoperoxidase-KI incubation, in which I3-, OI- and oxygen singlet are produced, was obtained after 22 +/- 7 (SD) min in the absence of the tested compounds. The 50% lysis was obtained after 66 +/- 15, 129 +/- 35, 196 +/- 21, 240 +/- 63 and 420 +/- 80 min with 40 microM piceatannol, quercetin, resveratrol, epicatechin and catechin respectively. Protection was concentration dependent. The assay of tyrosine kinase activity was performed using two different substrates as follows: substrate A corresponded to the sequence 1-17 of gastrin, and substrate B to sequence 6-20 of cell division kinase p34cdc2. In all experiments, initial velocity was measured. When assayed with both substrates, tyrosine kinase activities from particulate and cytosolic fractions of placenta were more inhibited by piceatannol and quercetin. Resveratrol significantly inhibited the particulate fraction and the cytosolic fraction respectively when substrates A and B were employed: Catechin acted as an inhibitor with substrate A and particulate fraction while in the other experimental conditions it acted as an activator. Resveratrol inhibited the tyrosine kinase of particulate and cytosolic fractions of prostatic adenoma assayed with substrate A and B.     

Polymeric proanthocyanidins from the bark of Hamamelis virginiana

Polymeric proanthocyanidins were isolated from the bark of Hamamelis virginiana L. in yields of about 5 %. Fractionation yielded fractions with similar structures but different molecular weights with DP between 17-29 (thiolysis) and 11-20 (GPC). Polymers were composed predominantly of epicatechin and epigallocatechin as chain extension units at ratio of about 1.3:1. Terminal chain units were catechin (approximately 95 %) and gallocatechin (approximately 5 %). All chain extension units were completely galloylated at position O-3, while chain terminating units were not galloylated. Predominant interflavan linkages were 4 --> 8-bonds.