Characterisation of the phenolic profile of Boerhaavia diffusa L. by HPLC-PAD-MS/MS as a tool for quality control

Phenolic acids and flavonols of nine leaf and three root samples of Boerhaavia diffusa L., collected at different locations and subjected to several drying procedures, were characterised by reversed-phase HPLC-PAD-ESI/MS for the first time. Ten phenolic compounds were identified: 3,4-dihydroxy-5-methoxycinnamoyl-rhamnoside, quercetin 3-O-rhamnosyl(1-->6)galactoside (quercetin 3-O-robinobioside), quercetin 3-O-(2"-rhamnosyl)-robinobioside, kaempferol 3-O-(2"-rhamnosyl)-robinobioside, 3,5,4'-trihydroxy-6,7-dimethoxyflavone 3-O-galactosyl(1-->2)glucoside [eupalitin 3-O-galactosyl(1-->2)glucoside], caffeoyltartaric acid, kaempferol 3-O-robinobioside, eupalitin 3-O-galactoside, quercetin and kaempferol. Quantification was achieved by HPLC-PAD and two phenolic patterns were found for the leaves, in which quercetin 3-O-robinobioside or quercetin 3-O-(2"-rhamnosyl)-robinobioside was the major compound. Caffeoyltartaric acid was only present in the root material where it represented the main phenolic constituent. The results obtained demonstrated that the geographical origin (particularly the nature of the soil), but not the drying process, influences the phenolic composition.     

Flavonoids of Ochradenus baccatus.

From the aerial parts of Ochradenus baccatus, the new flavonoids, quercetin 3-O-beta-glucosyl(1----2)-alpha-rhamnoside-7-O-alpha-rhamnoside and quercetin 3-O-p-coumaryl(1----6)-beta-glucosyl(1----6)-beta-glucoside-7-O-alpha rhamnoside were isolated. The known quercetin glycosides, quercetin 3-gentiobioside, isoquercitrin, quercitrin, together with the known kaempferol glycosides, astragalin and afzelin, were also characterized. The structures were established by conventional methods of analysis and confirmed by spectral analysis.     

湖南猕猴桃属植物地理分布及其区系特点研究

湖南猕猴桃属植物有２１种１３变种,主要分布于湘南和湘西两地区,经分析研究,湖南猕猴桃属植物区系有如下特点：１、种类比较丰富,是构成湖南森林植物区系林间植物的主要成分之一;２、地理成分比较复杂,种的分析显示出以中国特有分布为主的特点;３、种间、种内分化强烈;４、特有种较多;５、与邻近地区猕猴桃属植物区系的关系密切,最后,作者对湖南猕猴桃属植物的起源问题进行了分析和推测。     

Quercetin 3-O-beta-glucoside is better absorbed than other quercetin forms and is not present in rat plasma

The effect of the nature of the sugar moiety on quercetin absorption has been investigated in rats. Four groups of rats received an experimental meal containing 20 mg of quercetin equivalents, supplied as quercetin, quercetin 3-O-β-glucoside, quercetin 3-O-β-rhamnoside or rutin. Four hours after the meal, the metabolites identified in hydrolysed plasma were identical in all groups (3'- and 4'-methylquercetin). However, the total concentration of metabolites was markedly different: 11.2±1.8, 2.5±2.0 and 33.2±3.5 μM for the quercetin, rutin, and quercetin 3-glucoside meals respectively. After quercetin 3-rhamnoside consumption, we failed to detect any metabolites in the plasma. These data suggest that the 3-O-glucosylation improves the absorption of quercetin in the small intestine, whereas the binding of a rhamnose to the aglycone markedly depresses it. Additional experiments have shown that the higher plasma levels measured after quercetin 3-glucoside meal compared to the quercetin meal were maintained throughout the 24-hour period following the meal. Using a multi-electrode coulometric detection, together with suitable chromatographic conditions, we were able to distinguish between the conjugated and the glycosylated forms. Thus, we clearly showed the absence of quercetin 3-O-β-glucoside in the plasma from rats fed a diet containing this glucoside. This result suggests that quercetin 3-O-β-glucoside is hydrolysed before or during its intestinal absorption.     

Coumaroyl flavonol glycosides from the leaves of Ginkgo biloba.

Two coumaroyl flavonol glycosides, isorhamnetin 3-O-alpha-L-[6"'-p-coumaroyl-(beta-D)-glucopyranosyl-(1,2)-rhamnopyranoside], and kaempferol 3-O-alpha-L-[6"'-p-coumaroyl-(beta-D)-glucopyranosyl-(1,2)-rhamnopyranoside]-7-O-beta-D-glucopyranoside, were isolated from the n-BuOH extract of Ginkgo biloba leaves. These two, together with six other flavonol glycosides, kaempferol 3-O-alpha-L-[6"'-p-coumaroyl-(beta-D)-glucopyranosyl-(1,2)-rhamnopyranoside], quercetin 3-O-alpha-L-[6"'-p-coumaroyl-(beta-D)-glucopyranosyl-(1,2)-rhamnopyranoside], quercetin 3-O-alpha-L-[6"'-p-coumaroyl-(beta-D)-glucopyranosyl-(1,2)-rhamnopyranoside]-7-O-beta-D-glucopyranoside, quercetin 3-O-beta-D-glucopyranosyl-(1-2)-alpha-L-rhamnopyranoside, quercetin 3-O-beta-rutinoside, and quercetin 3-O-beta-D-glucopyranoside, showed profound antioxidant activities in DPPH and cytochrome-c reduction assays using the HL-60 cell culture system.     

微生物糖基转移酶催化合成槲皮素糖苷及其抗炎活性评价

利用微生物来源的糖基转移酶GT-1、GT-2和BTGT-1对槲皮素进行糖基化修饰。研究发现这3种酶均能够催化以槲皮素和UDP-葡萄糖为底物生成槲皮素-3-O-β-D-葡萄糖（异槲皮苷）,GT-1、 GT-2和BTGT-1催化反应的产物生成率分别为5.33%、15.18%和63.82%。通过对槲皮素、异槲皮苷和槲皮素-N-乙酰-D-氨基葡萄糖进行水溶性以及体外细胞抗炎活性检测,发现槲皮素经糖基化后其水溶性得到较大提高,异槲皮苷和槲皮素-N-乙酰-D-氨基葡萄糖水溶性分别是槲皮素的13.8倍和15.4倍。同等浓度下槲皮素糖苷对RAW264.7 细胞的毒性作用低于槲皮素,且3种化合物在一定浓度范围内对LPS诱导RAW264.7细胞释放NO、 IL-1β、IL-6 都有显著的抑制作用,且呈剂量依赖性,研究表明3种化合物都具有一定的抗炎作用。     

Identification of quercetin 3-O-beta-D-glucuronide as an antioxidative metabolite in rat plasma after oral administration of quercetin

The potential beneficial effect of dietary quercetin (3,3',4',5,7-pentahydroxyflavone) has attracted much attention in relation to the prevention of cardiovascular disease. It is generally recognized that dietary quercetin is subject to metabolic conversion resulting in conjugated forms during absorption and circulation. However, no quercetin conjugates have yet been identified from biological fluids or tissues. In the present study, we isolated and characterized two quercetin conjugates from the plasma of quercetin-administered rats. The blood plasma was collected from 26 rats 30 min after oral administration of quercetin (250 mg/kg body weight), concentrated, dissolved in 2% acetic acid aqueous solution (pH 2.65), and extracted with ethyl acetate. Two compounds (P2, P3) were obtained from the extract by repeated reversed-phase HPLC. On the other hand, two quercetin glucuronides were synthesized chemically and identified as quercetin 3-O-beta-D-glucuronide (Q3GA) and quercetin 4'-O-beta-D-glucuronide (Q4'GA), as determined from FABMS, 1H- and 13C-NMR, and HMBC data. The retention times of P2 and P3 in the HPLC chromatogram corresponded to those of Q3GA and Q4'GA, respectively. FABMS data demonstrated that P2 and P3 are quercetin monoglucuronides. 1H-NMR data for P2 were completely in agreement with those for Q3GA. P2 was therefore identified as Q3GA. This is, to our knowledge, the first evidence that Q3GA accumulates in vivo after oral administration of quercetin. Q3GA is likely to act as an effective antioxidant in blood plasma low-density lipoprotein, because this conjugated metabolite was found to possess a substantial antioxidant effect on copper ion-induced oxidation of human plasma low-density lipoprotein as well as 1,1-diphenyl-2-picrylhydrazyl radical-scavenging activity.     

Role of quercetin and its in vivo metabolites in protecting H9c2 cells against oxidative stress

The aim of this study was to investigate the potential of quercetin and two of its "in vivo" metabolites, 3'-O-methyl quercetin and 4'-O-methyl quercetin, to protect H9c2 cardiomyoblasts against H(2)O(2)-induced oxidative stress. As limited data are available regarding the potential uptake and cellular effects of quercetin and its metabolites in cardiac cells, we have evaluated the cellular association/uptake of the three compounds and their involvement in the modulation of two pro-survival signalling pathways: ERK1/2 signalling cascade and PI3K/Akt pathway. The three flavonols associated with cells to differing extents. Quercetin and its two O-methylated metabolites were able to reduce intracellular ROS production but only quercetin was able to counteract H(2)O(2) cell damage, as measured by MTT reduction assay, caspase-3 activity and DNA fragmentation assays. Furthermore, only quercetin was observed to modulate pro-survival signalling through ERK1/2 and PI3K/Akt pathway. In conclusion we have demonstrated that quercetin, but not its O-methylated metabolites, exerts protective effects against H(2)O(2) cardiotoxicity and that the mechanism of its action involves the modulation of PI3K/Akt and ERK1/2 signalling pathways.     

Regulation of cyclic AMP level and lactic acid production in Ehrlich ascites tumor cells.

1. Quercetin (3.3',4',5,7-pentahydroxy flavone) at the concentration of 10(-4) M, as well as 2-10(-2) M theophylline and 1.5 - 10(-4) M prostaglandin E2 caused maximal rise of cyclic AMP in Ehrlich ascites tumor cells. 2. No additional increase of cyclic AMP level in these cells was found when both quercetin (10(-4) M) and theophylline (2-10(-2) M) were present in the incubation medium, while combination of quercetin (10(-4) M) and prostaglandin E2 (1.5 - 10(-4) M) has a synergistic effect on the level of cyclic AMP. 3. Degradation of cyclic AMP by homogenate of Ehrlich ascites tumor cells was inhibited by both quercetin and theophylline. 4. Quercetin, and to a smaller but significant extent theophylline, inhibited the lactic acid production in Ehrlich ascites tumor cells while prostaglandin E2 did not change the glycolytic rate in these cells. No synergistic inhibitory effect on lactic acid production was found when combinations of quercetin and prostaglandin E2, quercetin and theophylline or prostaglandin E2 and theophylline were tested. 5. Treatment of Ehrlich ascites tumor cells with dextran sulfate abolished the inhibitory effect of quercetin on lactic acid production, while the effect of the bioflavonoid on cyclic AMP levels was not altered.     

猕猴桃种子发芽试验报告

报道在桂林的条件下猕猴桃属几个主要种类的不同的播种期、贮藏方法、药剂处理等发芽试验的结果:1.中华猕猴桃以三月中旬至四月巾旬为适宜的播种期.此时播种的发芽率为21.75—54.67％;2.沙贮猕猴桃种子可保持较高的发芽率,比袋藏的高7.5—54.75％;3.猕猴桃种子播种前以激动素处理有较高的发芽率,比对照高38.06—48.02％。     

Multiple Regiospecific Methylations of a Flavonoid by Plant O-Methyltransferases Expressed in E. coli

Quercetin was methylated with two O-methyltransferases (OMTs) expressed in E. coli. A construct (RSOMT) was designed to express two OMTs: ROMT-9, which methylates specifically at the 3'-hydroxyl group of quercetin and SOMT-2, which methylates at the 4'-hydroxyl group. Both OMT genes were driven by T7 promoters and had ribosome binding sites. Both ROMT-9 and SOMT-2 were successfully expressed in E. coli transformant harboring RSOMT. Reaction products of quercetin with E. coli transformant containing RSOMT showed two methylation products that corresponded to the 3'-methylated and the 3',4'-dimethylated quercetin, which were confirmed by NMR. More than 90% of quercetin was converted into the 3',4'-dimethylated quercetin after 24 h incubation with E. coli transformant harboring RSOMT.     

Invvestigations on Early Embryogenesis of Actinidia chinensis Planch var. chinensis

This paper deals with early embryogenesis of Actinidia chinensis var. chinensis. 1. Ovary superior consists of 34—45 carpels. Each carpel contains 11–45 ovules. The ovule is uni-integument and tenuinucellar. The ovule is anatropous. The archesporium is formed by a single cell, and directly develops into megaspore mother cell. Sometimes the archesporium consists of 2–3 cells, but only one of them develops into megaspore mother cell and the others are degenerated. 2. The mature pollen grain is two-celled and the embryo sac belongs to olygonum type. In most embryo sacs two polar nuclei are fused before fertilization. One of the synergids was destroyed as the pollen tube penetrated into embryo sac the other one disappeared after fertilization. In most cases the antipodal cells became degenerated in fertilization process, only some remained until the first division of primary endosperm nucleus. 3. In Beijing area the double fertilization of Actinidia chinensis occurred 30–72 hours after pollination. In the fertilization one sperm fused with egg nucleus and the other sperm fused with the secondary nucleus as usual. The fusion of the secondary nucleus with sperm was in advance of the fusion of the egg nudeus. 4. The endosperm is cellular type.     

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.     

Acetylated flavonol diglucosides from Meconopsis quintuplinervia

Four acetylated flavonol diglucosides, quercetin 3-O-[2'-O-acetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], quercetin 3-O-[2',6'-O-diacetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], isorhamnetin 3-O-[2'-O-acetyl-beta-d-glucopyranosyl-(1-->6)-beta-d-glucopyranoside], and quercetin 3-O-[2'-O-acetyl-alpha-l-arabinopyranosyl-(1-->6)-beta-d-glucopyranoside], together with five known flavonol glycosides quercetin 3-O-beta-d-glucopyranoside, kaempferol 3-O-beta-d-glucopyranoside, quercetin 3-O-[beta-d-galactopyranosyl-(1-->6)-glucopyranoside], isorhamnetin 3-O-[beta-d-galactopyranosyl-(1-->6)-beta-d-glucopyranoside], and kaempferol 3-O-[beta-d-glucopyranosyl-(1-->2)-beta-d-glucopyranoside] have been isolated from Meconopsis quintuplinervia. Their structures were determined using chemical and spectroscopic methods including HRFABMS, (1)H-(1)H COSY, HSQC and HMBC experiments.     

Kaempferol and quercetin isolated from Euonymus alatus improve glucose uptake of 3T3-L1 cells without adipogenesis activity

Euonymus alatus as a folk medicine in China has been clinically used to treat type 2 diabetes for many years, and also exerts beneficial effects on hyperglycemia of diabetic animals. Our previous studies have isolated kaempferol and quercetin from the extract of E. alatus. In the present study, we investigated the possible mechanism of antidiabetic activity of these compounds. Kaempferol and quercetin could significantly improve insulin-stimulated glucose uptake in mature 3T3-L1 adipocytes. In addition, further experiments showed that kaempferol and quercetin served as weak partial agonists in the peroxisome proliferator-agonist receptor gamma (PPARgamma) reporter gene assay. Kaempferol and quercetin could not induce differentiation of 3T3-L1 preadipocytes as traditional PPARgamma agonist. When added together with the PPARgamma agonist rosiglitazone to 3T3-L1 preadipocytes, they could inhibit 3T3-L1 differentiation in a dose-dependent manner. Competitive ligand-binding assay confirmed that kaempferol and quercetin could compete with rosiglitazone at the same binding pocket site as PPARgamma. Kaempferol and quercetin showed significant inhibitory effects on NO production in response to lipopolysaccharide treatment in macrophage cells in which the PPARgamma was overexpressed; rosiglitazone was less potent than kaempferol and quercetin. These observations suggest that kaempferol and quercetin potentially act at multiple targets to ameliorate hyperglycemia, including by acting as partial agonists of PPARgamma.     

Flavonol triglycosides containing galactose in tea.

The isolation and structural elucidation of new quercetin and kaempferol triglycosides from Camellia sinensis is described. Their structures were determined as quercetin and kaempferol 3-glucosyl(1----3) rhamnosyl(1----6)galactosides. The content of quercetin glucosylrhamnosylgalactoside ranged between 0 and 87 mg per 100 g, and that of the kaempferol homologue between 0 and 119 mg per 100 g dry wt.     

Flavonol glycosides of Warburgia ugandensis leaves

Four new flavonol gycosides: kaempferide 3-O-beta-xylosyl (1-->2)-beta-glucoside, kaempferol 3-O-alpha-rhamnoside-7,4'-di-O-beta-galactoside, kaempferol 3,7,4'-tri-O-beta-glucoside and quercetin 3-O-[alpha-rhamnosyl (1-->6)] [beta-glucosyl (1-->2)]-beta-glucoside-7-O-alpha-rhamnoside, were characterized from a methanolic leaf extract of Warburgia ugandensis. The known flavonols: kaempferol, kaempferol 3-rhamnoside, kaempferol 3-rutinoside, myricetin, quercetin 3-rhamnoside, kaempferol 3-arabinoside, quercetin 3-glucoside, quercetin, kaempferol 3-rhamnoside-4'-galactoside, myricetin 3-galactoside and kaempferol 3-glucoside were also isolated. Structures were established by spectroscopic and chemical methods and by comparison with authentic samples.     

Regioselective synthesis of flavonoid bisglycosides using Escherichia coli harboring two glycosyltransferases

Regioselective glycosylation of flavonoids cannot be easily achieved due to the presence of several hydroxyl groups in flavonoids. This hurdle could be overcome by employing uridine diphosphate-dependent glycosyltransferases (UGTs), which use nucleotide sugars as sugar donors and diverse compounds including flavonoids as sugar acceptors. Quercetin rhamnosides contain antiviral activity. Two quercetin diglycosides, quercetin 3-O-glucoside-7-O-rhamnoside and quercetin 3,7-O-bisrhamnoside, were synthesized using Escherichia coli expressing two UGTs. For the synthesis of quercetin 3-O-glucoside-7-O-rhamnoside, AtUGT78D2, which transfers glucose from UDP-glucose to the 3-hydroxyl group of quercetin, and AtUGT89C1, which transfers rhamnose from UDP-rhamnose to the 7-hydroxyl group of quercetin 3-O-glucoside, were transformed into E. coli. Using this approach, 67 mg/L of quercetin 3-O-glucoside-7-O-rhamnoside was synthesized. For the synthesis of quercetin 3,7-O-bisrhamnoside, AtUGT78D1, which transfers rhamnose to the 3-hydroxy group of quercetin, and AtUGT89C1 were used. The RHM2 gene from Arabidopsis thaliana was coexpressed to supply the sugar donor, UDP-rhamnose. E. coli expressing AtUGT78D1, AtUGT89C1, and RHM2 was used to obtain 67.4 mg/L of quercetin 3,7-O-bisrhamnoside.     

金叶子中抑制膜结合型PGE2合酶-1表达活性物质的分离与鉴定

在从天然产物筛选抗炎物质的过程中,应用有机溶媒萃取、浓缩及层析分离纯化后,从金叶子中得到三个化合物:槲皮素(1)、番石榴苷(2)、栎素(3).化合物2为首次从此植物中分离鉴定,化合物1显示了强烈的抑制mPGES-1基因表达的活性,其IC50值为35.40μg/mL.