五爪龙外敷液中芹菜素与槲皮素的HPLC测定

本实验建立芹菜素与槲皮素的HPLC检测法,同时测定草药五爪龙外敷药液中芹菜素与槲皮素的含量。采用反相shim-pack VP-ODS C18色谱柱(4.6 mm×150 mm,5μm),优化条件为:柱温30℃,流动相:甲醇-0.4%磷酸(50∶50,v∶v);流速:1.0 mL/min;检测波长:360nm;进样体积10μL。对草药五爪龙外敷药液中芹菜素与槲皮素的含量进行测定。结果表明:芹菜素与槲皮素均在60~300μg/mL的范围内线性关系良好,r分别为0.9998与0.9996,草药五爪龙乙醇浸液较水煎煮液中芹菜素与槲皮素含量均高。本法方便、快速、准确,可检测草药五爪龙外敷液中的芹菜素与槲皮素的含量。     

反相高效液相色谱法测定不同品种花椒中芦丁和槲皮素的含量

目的:建立RP-HPLC测定花椒中芦丁与槲皮素含量的方法,并对不同种花椒的中芦丁与槲皮素含量进行测定与比较。方法:Zorbax Eclipse C18色谱柱(150 mm×4.6 mm,5μm),流动相∶甲醇-0.4%磷酸(50∶50);流速1 mL/min;检测波长:360 nm;柱温25℃。结果:芦丁在0.25~5.0μg,r=0.999 9峰面积与质量浓度呈良好的线性关系;平均回收率为99.1%,RSD为4.3%(n=3)。槲皮素在0.25~0.5μg,r=0.999 9峰面积与质量浓度呈良好的线性关系;平均回收率为111.2%,RSD为5.1%(n=3)。结论:该方法可用于花椒中芦丁和槲皮素的测定。测定结果表明,韩城红花椒中芦丁含量最高,茂汉红花椒次之,四川青花椒较少,云南青花椒最低。槲皮素在韩城红花椒中含量较高,在其他三种花椒中差别不大。     

反相高效液相色谱法测定石榴皮多酚类物质方法的建立及分析

建立了同时测定石榴皮多酚中绿原酸、表儿茶素、鞣花酸和槲皮素含量的反相高效液相色谱方法。采用色谱柱Hypersil ODS2(250 mm×4.6 mm,5μm),其流动相为乙腈-0.4%磷酸溶液(体积比17∶83),流速为1.0m L/min,检测波长310 nm,柱温为30℃。建立安石榴甙测量方法,色谱柱为Hypersil ODS2(250 mm×4.6 mm,5μm);流动相为甲醇-2%冰醋酸(体积比7∶93);体积流量为1.0 m L/min;检测波长为232 nm;柱温25℃。结果表明:绿原酸、表儿茶素、鞣花酸、槲皮素和安石榴甙在一定浓度范围内与峰面积呈良好的线性关系,其平均回收率分别为100.66%(RSD=2.06%)、100.05%(RSD=0.58%)、100.05%(RSD=0.51%)、99.51%(RSD=1.22%)和99.79%(RSD=0.52%)。此结果说明反相高效液相色谱可用于测定石榴皮多酚类物质。     

RP-HPLC同时测定喙果绞股蓝中芦丁和槲皮素的含量

目的:建立RP-HPLC同时测定喙果绞股蓝中芦丁和槲皮素含量的方法,并揭示其含量动态变化规律。方法:采用大连依利特SinoChromODS-APC18色谱柱(250 mm×4.6 mm,5μm),流动相:甲醇-0.4%磷酸水溶液梯度洗脱,流速:1 mL/min,检测波长:364 nm,柱温:30℃。结果:芦丁在0.1525～3.8120μg范围内线性关系良好,r=0.9998,平均回收率为98.2%(RSD=2.0%);槲皮素在0.0589～1.4720μg范围内线性关系良好,r=0.9999,平均回收率为97.7%(RSD=2.3%)。含量测定结果表明芦丁和槲皮素的含量具季节性动态变化,芦丁8月份含量高,平均质量分数达6.31 mg/g,槲皮素9月份含量最高,平均质量分数达0.86 mg/g。结论:该方法简单,准确度高,为喙果绞股蓝的质量控制提供实验依据,芦丁和槲皮素含量动态变化规律为其开发利用提供参考。     

RP-HPLC法测定不同产地红枣中甜菜碱的含量

目的:通过对影响甜菜碱提取和测定的符种因素进行了研究,建立了一种准确、快速的测定不同产地红枣中甜菜碱的方法.方法:样品采用超声波甲醇提取的方法,得到的样品采用反相C18柱进行分离,色谱条件为:流动相为50mmol/L pH 4.5的KH2PO4溶液;192nm波长测定,流速0.7ml/min.结果:总分析时间:4.5min.甜菜碱平均保留时间2.698min.甜菜碱在5.0～25.0mg/ml线性关系良好(r2=0.9911),平均回收率为96.87%.结论:该法测定红枣中甜菜碱含量简便、快速、准确,重现性好.     

绿肉山楂叶中五种黄酮类成分含量动态变化

为充分利用山楂叶用资源,采用反相高效液相色谱法,以ZORBAX Extend-C18色谱柱（250×4.6 mm,5 μm）,乙腈-0.5%磷酸水溶液梯度洗脱,流速1.0 mL·min^-1,检测波长345 nm,柱温25℃,对不同生长时期的绿肉山楂叶中牡荆苷、牡荆素2″-O-鼠李糖苷、金丝桃苷、芦丁和槲皮素含量进行测定。结果表明,在此分析检测条件下牡荆苷、牡荆素 2″-O-鼠李糖苷、金丝桃苷、芦丁和槲皮素的精密度、重复性、稳定性及平均加样回收率试验的RSD值均小于3%,方法准确可靠。绿肉山楂叶中含有的黄酮类成分主要以牡荆苷和金丝桃苷为主,且二者含量变化规律与其总黄酮含量变化相似,均在10月初含量达到最高;此时采摘,其质量完全符合《中国药典》的有关规定,故绿肉山楂可作为一种新的药用资源来进一步开发利用。     

梯度洗脱高效液相色谱法测定桑叶中3种活性成分的含量

建立同时测定桑叶中绿原酸、芦丁和槲皮素含量的分析方法。色谱柱为NUCLEODUR C18 RP(250 mm×4.6 mm,5μm),光电二极管检测器,流动相为甲醇-质量分数0.5%磷酸水溶液,梯度洗脱程序为0 min(30:70)-15min(30:70)-25min(50:50)-35min(85:15)-40min(30:70);流速0.8 ml.min-1,检测波长为350 nm。绿原酸、芦丁和槲皮素的线性范围分别为0.1144~1.0296μg(r=0.9996)、0.0436~0.3924μg(r=0.9999)和0.0452~0.4068μg(r=0.9997),平均回收率分别为97.7%(RSD=1.7%)、98.4%(RSD=2.2%)和100.6%(RSD=1.5%)。方法快速简便,专属性强,重复性好,可作为桑叶中绿原酸、芦丁和槲皮素的定量分析方法。     

反相高效液相色谱法测定平邑甜茶植株内源ABA、IAA含量

用高效液相色谱法 (HPLC)分析平邑甜茶内源ABA、IAA ,选用SpherisorbC18反相柱 ,用UV检测器进行检测 ,使ABA、IAA得到很好的分离 ,测定方法迅速简便 ,流动相为甲醇、乙酸和水 ,系统研究了pH值、纯化方法对测定结果的影响 ,确定适合平邑甜茶ABA、IAA提取的分离条件。方法的精密度为 :变异系数分别为 2 83和 2 79,最低检出限 (信噪比 =2 )分别为 3 2 7和 3 2 6nmol/L ,线性范围为 50nmol/L -10 0 0nmol/L ,平均回收率为 91 9%和 92 2 %。     

RP-HPLC法测定垫状卷柏中穗花杉双黄酮的含量

本文用反相高效液相色谱法测定垫状卷柏药材中的穗花杉双黄酮.选用Puraspher star RP-C18色谱柱(250 mm×4.6 mm,5μm),以甲醇-0.1%磷酸水溶液(65:35)为流动相,流速为0.8 mL/min,检测波长337 nm,柱温25℃进行检测.结果显示,穗花杉双黄酮的含量在0.038～0.342 μg范围内时进样量与峰面积积分值呈良好的线性关系,r=0.9999,平均回收率为101.2%,RSD为2.3%(n=6).该方法测定垫状卷柏药材中的穗花杉双黄酮含量灵敏、准确、重现性好.     

HPLC法测定芦笋茎秆中芦丁、槲皮素及白藜芦醇含量

为了建立高效液相色谱法对芦笋茎秆中芦丁、槲皮素和白藜芦醇含量的测定方法。采用不同方法提取芦笋茎秆中的芦丁、槲皮素和白藜芦醇,得出甲醇法提取芦丁、甲醇-HCl法提取槲皮素以及乙醇法提取白藜芦醇得率最高;确定了高效液相色谱法对3种物质的最佳色谱条件为流动相:芦丁为0.2 mol/L乙酸钠-甲醇(Me-OH∶H2O=35∶65)溶液(用磷酸调p H 2.80)、槲皮素为无水甲醇-0.4%磷酸溶液(55∶45,V/V),等梯度洗脱,白藜芦醇为水和甲醇,二元线性梯度洗脱;检测波长分别为:254 nm、360 nm和306 nm;流速:0.8 m L/min、1.0 m L/min、0.8 m L/min。通过该方法测得芦丁、槲皮素及白藜芦醇的平均回收率分别为:98%、97%、98%。     

Quercetin both partially attenuates hydrogen peroxide-induced toxicity and decreases viability of rat glial cells

Quercetin is one of the most ubiquitous flavonoids in foods of plant origin. Although quercetin is generally considered to provide protection against oxidative injury, recent studies have shown to be cytotoxic to many cell types. We intended here to determine whether quercetin protects against H2O2-induced toxicity and/or affects viability of rat mixed glial cells. The cells were obtained from 1-3 day olds rat brains and incubated in a humidified atmosphere of 5% CO2, at 37 °C in flasks. In the quercetin groups, different quercetin concentrations (1, 10, 50, 75 or 100 μM) were applied alone for 24 h. For H2O2 cytotoxicity group, the glial cells were treated for 3 h with 100 μM H2O2 which induced 75% cell death. In another group, the cells were treated with 100 μM H2O2 plus respective quercetin concentrations simultaneously for 3 h, the medium was removed and refed for 24 h. MTT test was then applied and statistical significance was ascertained by one way analysis of variance, followed by Tukey's multiple comparison test. Quercetin starting from 50 μM decreased the glia survival significantly. In H2O2 plus quercetin co-treated groups, both 75 and 100 μM quercetin attenuated toxic effect of H2O2 by 15%. In conclusion, quercetin both partially protects H2O2-induced gliotoxicity and decreases rat glial cell viability in primary culture.     

High-performance liquid chromatography with coulometric electrode array detector for the determination of quercetin levels in cells of the immune system

There is mounting evidence emphasizing the importance of intracellular antioxidant levels for maintenance of the immune function. The flavonoid quercetin, a natural antioxidant, has been shown to modulate enzymes involved in the regulation of the inflammatory response. However, up to now, there have been no studies describing quercetin levels in cells of the immune system. A gradient reversed-phase HPLC technique to identify and quantify intracellular levels of quercetin and its application in mice splenocytes are described. Mobile phases were a 0.01 M sodium phosphate monobasic solution adjusted to pH 2.8 with 85% orthophosphoric acid (buffer, Solvent A) and methanol (Solvent B) with a flow rate of 1 ml/min. An eight-channel coulometric electrode array detector was used. In vitro supplementation with increasing concentration of quercetin (25, 50, and 100 microM) raises intracellular quercetin levels in a dose-dependent manner. The method has the required features of specificity and sensitivity for monitoring quercetin uptake in cells of the immune system.     

Bioavailability and metabolism of the flavonol quercetin in the pig

During the last years, much data pointing to putative health-promoting effects of dietary plant-derived flavonoids (stemming mainly from epidemiological and in vitro studies) have been published. Our knowledge, however, concerning the systemic availability of these substances after ingestion with food is only sketchy. In the present study, we have investigated the bioavailability of the flavonol quercetin after intravenous and oral application in pigs equipped with a permanent jugular catheter. Each animal received a single intravenous dose of quercetin (0.4 mg/kg body weight) and one week later an oral dose of 50 mg/kg. A single animal additionally received an oral dose of 500 mg/kg one week after the lower oral dose. Blood samples were drawn at defined intervals over a total period of three days following the application of quercetin. Analysis of quercetin and some of its metabolites (isorhamnetin, tamarixetin, kaempferol) in plasma samples were performed by HPLC. The calculated apparent bioavailability of free, unchanged quercetin after intake of 50 mg quercetin/kg body weight was 0.54+/-0.19%. Bioavailability was, however, considerably increased to 8.6+/-3.8% after additionally taking into account conjugated quercetin and further increased to 17.0+/-7.1% by including quercetin's metabolites. Our results further indicate, that the conjugation of orally administered quercetin with glucuronic and sulfuric acid appears to preferentially occur in the intestinal wall.     

Inhibition of cytochrome P-450 activity in rat liver microsomes by the naturally occurring flavonoid, quercetin

The kinetic characteristics and mechanism of flavonoid inhibition of cytochrome P-450-mediated reactions were examined in rat liver microsomes, using the naturally occurring flavonoid, quercetin (3,3',4',5,7-pentahydroxyflavone). Quercetin inhibited the O-deethylation of ethoxyresorufin in beta-naphthoflavone-induced microsomes by 15-80% at concentrations of 10-250 nM. The pattern of inhibition was dependent on quercetin concentration. Quercetin also inhibited p-nitroanisole demethylation and benzo(a)-pyrene hydroxylation, but did not change the proportions of the individual benzo(a)pyrene metabolites in comparison to controls. Specific steps in the P-450 reaction pathway were tested for sensitivity to quercetin inhibition. The Km values of the P-450 substrates tested were increased in the presence of quercetin; competition for and/or alteration of the substrate binding site contributes to the mechanism of inhibition. In experiments under anaerobic, carbon monoxide-saturated conditions, quercetin did not inhibit cytochrome P-450 reduction by NADPH-cytochrome P-450 reductase. The cumene hydroperoxide-supported O-deethylation of ethoxyresorufin was inhibited by quercetin (15-60% inhibition at concentrations of 50-300 nM), suggesting that quercetin may interfere with the formation or breakdown of the oxygenated heme complex. Stoichiometry experiments established that quercetin is a potent uncoupler of P-450 reactions, elevating the rates of H2O2 formation almost twofold. Structure/activity studies indicated that certain other naturally occurring flavonoids were at least as potent inhibitors of ethoxyresorufin deethylation as quercetin. These findings are of interest in light of the significant dietary exposure of the human population to the flavonoids.     

Liquid chromatography-tandem mass spectroscopy assay for quercetin and conjugated quercetin metabolites in human plasma and urine

A sensitive and specific method was developed and validated for the quantitation of quercetin in human plasma and urine. The application of liquid chromatography-tandem mass spectrometry (LC/MS/MS) with a TurboIonspray (TIS) interface in negative mode under multiple reactions monitoring was investigated. Chromatographic separation was achieved on a C12 column using a mobile phase of acetonitrile/water with 0.2% formic acid (pH 2.4) (40/60, v/v). The detection limit was 100 pg/ml and the lower limit of quantification was 500 pg/ml for plasma samples; the detection limit was 500 pg/ml and the lower limit of quantification was 1 ng/ml for urine samples. The calibration curve was linear from 1 to 800 ng/ml for plasma samples and was linear from 1 to 200 and 50 to 2000 ng/ml for urine samples. All the intra- and inter-day coefficients of variation were less than 11% and intra- and inter-day accuracies were within +/-15% of the known concentrations. This represents a LC/MS/MS assay with the sensitivity and specificity necessary to determine quercetin in human plasma and urine. This assay was used to determine both parent quercetin and the quercetin after enzymatic hydrolysis with beta-glucuronidase/sulfatase in human plasma and urine samples following the ingestion of quercetin 500 mg capsules.     

羽裂蟹甲草对家蝇和白纹伊蚊的杀虫活性和活性成分(英文)

为探明羽裂蟹甲草Cacalia tangutica的杀虫活性及其活性成分, 通过拌糖饲喂法和浸渍法测定了羽裂蟹甲草甲醇提取物对家蝇Musca domestica成虫和白纹伊蚊Aedes albopictus 4龄幼虫的毒杀活性, 并采用色谱分离技术和现代波谱技术对羽裂蟹甲草中的化学成分进行了分离鉴定。结果表明： 该植物不同部位甲醇提取物对家蝇成虫和白纹伊蚊4龄幼虫均具有较高的生物活性。从该植物叶和花甲醇提取物中分离鉴定了豆甾醇、 无羁萜、 7 羟基 8 甲氧基香豆素、 7 羟基香豆素、 7, 8 二羟基香豆素、 瑞香素-8-O-β-D-葡萄糖苷、 槲皮素、 山奈酚和β 胡萝卜苷9个化合物。无羁萜、 槲皮素和山奈酚为首次从该植物中分离得到, 500 μg/g拌糖处理48 h后, 其对家蝇成虫的校正死亡率分别为88.30%, 69.90%和77.04%; 50 μg/mL药液处理72 h后, 其对白纹伊蚊4龄幼虫的校正死亡率分别为88.49%, 72.22%和71.06%; 均与鱼藤酮差异不显著（P<0.05）, 表现出良好的杀虫活性。本研究说明羽裂蟹甲草对家蝇和白纹伊蚊具有明显的毒杀活性, 是一种潜在的卫生害虫控制剂。     

槐米中天然产物槲皮素抗人鼻咽癌CNE1细胞活性研究

本研究以槐米为原材料提纯天然产物槲皮素,以人鼻咽癌细胞系CNE1为试验对象,探索天然产物槲皮素对人鼻咽癌细胞的增殖抑制效应及凋亡诱导效应。研究方法采用超声醇提法从槐米中提取芸香苷,再经酸水解和重结晶制备槲皮素精制品;采用槲皮素标准曲线比色法检测槲皮素精制品中槲皮素的纯度;采用MTT比色法进行细胞毒性试验,检测槲皮素精制品对人鼻咽癌细胞CNE1增殖的抑制效应及半数抑制浓度(IC50);采用流式细胞术结合AnnexinV、PI双染色法进行细胞凋亡试验,检测槲皮素精制品对CNE1细胞凋亡的诱导效应。结果表明,从槐米中提纯的天然产物槲皮素精制品,能呈剂量依赖性地抑制人鼻咽癌细胞CNE1增殖并诱导其凋亡。综上所述,应用超声醇提法,从槐米中制备的天然产物槲皮素精制品,不仅纯度高,且有着较优的抗癌活性,进一步明确了槐米等中药材开发天然产物有效成分,服务于人类健康的潜在应用价值和社会与经济效益。     

Interaction of flavonoids with rabbit muscle phosphorylase kinase

We have examined the effect of several flavonoids on the activity of phosphorylase kinase from rabbit skeletal muscle. From 14 flavonoids tested, the flavones quercetin and fisetin were found to be efficient inhibitors of nonactivated phosphorylase kinase when assayed at pH 8.2, causing 50% inhibition at a concentration of about 50 microM, while the flavanone hesperetin stimulated phosphorylase kinase activity about 2-fold when tested at 250 microM. The efficiency of quercetin in inhibiting the kinase is higher when the enzyme is stimulated either by ethanol or by alkaline pH. Both casein and troponin phosphorylation by phosphorylase kinase and the autophosphorylation of the kinase were inhibited by quercetin. In addition, quercetin was found to be a competitive inhibitor of ATP for the phosphorylation of phosphorylase b at pH 8.2. These observations suggest that the inhibitory effect of the flavone is directly on the phosphorylase kinase molecule. Trypsin-activated phosphorylase kinase was inhibited by quercetin and stimulated by hesperetin, as for the native enzyme.