基于Ce(IV)分光光度法的天然产物抗氧化活性的评价

基于在H2SO4,Na2SO4介质中稳定存在的Ce(IV)的还原,在320 nm时,采用紫外可见分光光度计测定Ce(IV)吸光度,建立还原率与样品浓度的剂量关系,以线性关系的斜率为主要评价指标,评价样品的抗氧化活性。结果表明:针对5种结构相近的黄酮化合物,抗氧化活性顺序为:槲皮素>山奈酚>木犀草素>芹菜素>黄芩素;针对中药植物湖北旋覆花,抱石莲和吉祥草的提取物,正丁醇、乙酸乙酯部位是抗氧化活性成分的主体部位,但在旋覆花和抱石莲中,正丁醇部位抗氧化活性更强,而在吉祥草中,乙酸乙酯部位明显强于正丁醇部位;此外,不同提取方法得到的提取物抗氧化活性不同,低温回流和超声提取均可提高效率,但超声提取的时间应该控制在30 min以内。本评价方法操作简单,结果稳定可靠,检测限低,适合于天然产物抗氧化活性植物及活性部位的快速筛选。     

HPLC法测定不同施肥模式栽培的金线莲中黄酮类成分

HPLC法测定不同施肥模式栽培的金线莲中3种黄酮类成分:槲皮素、山奈酚和异鼠李素.超声-微波协同提取法提取4种不同施肥模式下金线莲中的黄酮类成分.HPLC-电喷雾离子化法/质谱(HPLC-ESI-MS)对黄酮类化合物进行定性分析.HPLC分析得槲皮素在0.25~20.0μg/m L(r=0.999 2)、山奈酚在0.25~20.0μg/m L(r=0.999 5)、异鼠李素在1.0~80.0μg/m L(r=0.999 0)范围内峰面积与浓度呈良好的线性关系,不同栽培方法的金线莲药材中槲皮素、山奈酚、异鼠李素的质量分数分别在151.8~240.0、47.2~88.2、16.8~29.8μg/g之间.方法简单、精确,快速,可为科学评价不同培养模式金线莲质量提供依据.栽培过程施加氮、磷、钾(NPK)复合肥及添加腐植素和微量元素,能提高金线莲中3种黄酮类成分的含量.栽培过程施加硫、磷(SP)复合肥,槲皮素、山奈酚和异鼠李素的质量分数分别下降了2.3%、17.2%和18.8%.结果表明采用不同施肥模式直接影响到药材中的药效物质的含量.     

几种天然植物提取物对酪氨酸酶活性的抑制作用

选择合适的溶剂体系,用维生素C(Vc)作为阳性对照物,采用分光光度法测定了不同浓度的茶多酚、丹皮酚及金银花叶子总黄酮提取物(乙酸乙酯相)对酪氨酸酶活性的抑制作用.结果表明:采用1%丙三醇和1%吐温-80(体积比1∶1)混合液作溶剂可显著增加丹皮酚和金银花叶子总黄酮提取物在水中的溶解度;茶多酚、丹皮酚、金银花叶子总黄酮提取物对酪氨酸酶的活性都有一定的抑制作用,其抑制作用从强到弱依次为Vc、Vc∶丹皮酚=1∶2(质量比)、Vc∶丹皮酚=1∶1(质量比)、茶多酚、茶多酚∶丹皮酚=2∶1(质量比)、丹皮酚、金银花叶子总黄酮提取物.     

Determination of antioxidant activity of natural products based on the spectroscopic absorbance of Ce(Ⅳ)

基于在H2SO4,Na2SO4介质中稳定存在的Ce(Ⅳ)的还原,在320 nm时,采用紫外可见分光光度计测定Ce(Ⅳ)吸光度,建立还原率与样品浓度的剂量关系,以线性关系的斜率为主要评价指标,评价样品的抗氧化活性.结果表明:针对5种结构相近的黄酮化合物,抗氧化活性顺序为:槲皮素＞山奈酚＞木犀草素＞芹菜素＞黄芩素;针对中药植物湖北旋覆花,抱石莲和吉祥草的提取物,正丁醇、乙酸乙酯部位是抗氧化活性成分的主体部位,但在旋覆花和抱石莲中,正丁醇部位抗氧化活性更强,而在吉祥草中,乙酸乙酯部位明显强于正丁醇部位;此外,不同提取方法得到的提取物抗氧化活性不同,低温回流和超声提取均可提高效率,但超声提取的时间应该控制在30 min以内.本评价方法操作简单,结果稳定可靠,检测限低,适合于天然产物抗氧化活性植物及活性部位的快速筛选.     

粉笔分离菠菜色素实验条件优化*

采用家用超声波眼镜清洗机辅助提取菠菜色素,考察了菠菜自身水分、不同提取部位、不同提取溶剂以及不同石油醚乙酸乙酯体积比对提取物粉笔分离效果的影响。实验结果表明:干燥菠菜叶、丙酮提取、V_石油醚∶V_乙酸乙酯=17∶2展开时,粉笔分离后呈现出5条清晰的分离色带。改进后实验效果更好,植物成分提取和色谱分离的教学功能更有利于实现。     

镰翅羊耳蒜中的新黄酮苷类化合物

利用多种分离方法对镰翅羊耳蒜的化学成分进行研究,从其全草的乙醇提取物中分离得到两个化合物,经过HR-ESIMS,1D和2D NMR等波谱技术鉴定了结构,分别为山奈酚3-O-α-L-鼠李糖-7-O-[6-(4-羟基-3-异戊二烯基苯甲酸酯)-β-D-葡萄糖]-(1→2)-α-L-鼠李糖(1)和山奈酚-3,7-O-α-L-二鼠李糖(2).化合物1为新的黄酮苷类化合物.     

固相萃取-气相色谱质谱测定动物组织中氯霉素的残留量

对鸡肉、猪肉、猪肝、鸭肝等动物组织样品中的氯霉素残留的检测方法进行了研究。用乙酸乙酯和磷酸盐缓冲溶液提取动物组织中的氯霉素残留,用硅胶和C18固相萃取柱对乙酸乙酯提取物进行净化和富集。净化后的提取物用BSTFA衍生化和气相色谱-质谱-负化学电离源选择离子定量检测(选择离子为376,378,466,468,470)。通过对提取、净化富集条件的优化,建立了动物组织中氯霉素残留量的检测方法。该方法对不同基体样品的加标回收率为80％～100％之间;相对标准偏差小于20％;方法的检出限为0．1μg／kg。     

高效液相色谱法测定野葱中黄酮类化合物

建立了测定野葱中芦丁、黄酮醇类化合物的高效液相色谱方法。80%乙醇超声提取,高效液相色谱分析野葱中芦丁和黄酮醇类化合物的含量。芦丁检测条件:V(甲醇)∶V(0.2%磷酸水)=45∶55,检测波长:360nm;槲皮素、山奈酚、异鼠李素检测条件:V(甲醇)∶V(0.2%磷酸水)=40∶60,检测波长:360 nm。结果表明,野葱中芦丁含量0.22%,槲皮素、山奈酚、异鼠李素含量依次为0.42%、0、0.23%,总黄酮醇类化合物含量为1.63%。     

芜菁籽不同溶剂提取物体外抗氧化活性研究

通过体外试验初步研究芜菁籽提取物的抗氧化活性。用95%乙醇为溶剂提取芜菁籽,采用有机溶剂萃取法将提取物分为石油醚相,乙酸乙酯相,正丁醇相和水相等4个不同极性部位、同时测定了各相提取物对羟自由基,超氧阴离子自由基,DPPH自由基的清除能力,并与合成抗氧化剂BHT进行对照。结果表明,芜菁籽提取物的不同极性部位均有抗氧化活性,其抗氧化效果随着浓度的增加而增强,乙酸乙酯相和正丁醇相的还原能力及清除O-2.,DPPH和.OH的能力均强于水相和石油醚相,乙酸乙酯相和正丁醇相的抗氧化作用强,是天然抗氧化剂的良好来源,应对其进一步分离纯化。     

山奈酚的电喷雾质谱裂解途径

采用电喷雾质谱技术对山奈酚的质谱过程进行了表征,并用量子化学方法对山奈酚的质谱裂解途径进行了理论研究。 依据密度泛函理论,在B3LYP/6-31G(d)水平,对山奈酚的各质谱碎片离子进行了几何结构优化,确定了各碎片的稳定结构;然后,在ROB3LYP/ 6-311++G(2d,2p)水平计算了形成各碎片离子所需的键断裂能,进而推导出了山奈酚的质谱碎裂途径。 计算结果显示,山奈酚主要通过C环碎裂而发生裂解,出现碎片m/z 284.7、256.7、228.7、210.7、184.8、168.7和150.7,其中m/z 210.7的碎片离子键断裂能最小,m/z 150.7的碎片离子键断裂能最大,说明前者很容易由母离子碎片形成,后者较难由母离子碎片形成。     

The uptake behaviors of kaempferol and quercetin through rat primary cultured cortical neurons

The objective of this study was to investigate whether kaempferol and quercetin could be transported into primary cultured cerebral neurons, to establish a practical HPLC method with UV detection for the two flavonols in the neurons, and to study the uptake and transport behaviors of them through the neurons. The present results showed that the level of kaempferol in the neurons increased linearly and then reached a plateau with incubation time at the high concentration of 10 microg/mL, but not at the other two concentrations of 1 and 0.1 microg/mL. However, the levels of quercetin in the neurons were not detected at the three incubating concentrations, and there was a new peak detected in the cell whose retention time was shorter (3.42 min) than that of quercetin (4.65 min). This phenomenon suggested that quercetin might be transported into the neurons and then metabolized quickly to some derivative. Kaempferol could be transported into the neurons in a concentration- and time-dependent manner when the neurons were incubated with the culture medium containing kaempferol at the high dose. There was an apparent correlation between the concentrations of kaempferol in the medium and in the cell, indicating that the uptake of kaempferol in the cell increased along with its dose (10 microg/mL). However, there was a negative correlation between the concentrations of quercetin in the medium and in the cell. The results suggested that kaempferol and quercetin were disposed by the neurons at different way, and this might be an important factor for their different effects on primary cultured cortical cells.     

Microbial metabolism of biologically active secondary metabolites from Nerium oleander L

Ursolic acid (1) and kaempferol (3) are two major constituents of the Mediterranean plant Nerium oleander L. Microbial metabolism of (1) with Aspergillus flavus (ATCC 9170) resulted in the formation of 3-oxo-ursolic acid derivative, ursonic acid (2). On the other hand, Cunninghamella blakesleeana (ATCC 8688A) was able to convert (3) into kaempferol 3-O-beta-D-glucopyranoside (4) as well as the new natural product kaempferol 4'-sulfate (5). Incubation of kaempferol with Mucor ramannianus (ATCC 9628) led to the isolation of one metabolite identified as kaempferol 4'-O-alpha-L-rhamnopyranoside (6). Transformation of kaempferol to the new compound kaempferol 7-O-beta-D-4-O-methylglucopyranoside (7) and herbacetin 8-O-beta-D-glucopyranoside (8) was observed after fermentation with Beauveria bassiana (ATCC 13144). Cytotoxic as well as antioxidant activities of the isolated metabolites were determined.     

Kaempferol glycosides in the flowers of carnation and their contribution to the creamy white flower color

Three flavonol glycosides were isolated from the flowers of carnation cultivars 'White Wink' and 'Honey Moon'. They were identified from their UV, MS, 1H and 13C NMR spectra as kaempferol 3-O-neohesperidoside, kaempferol 3-O-sophoroside and kaempferol 3-O-glucosyl-(1 --> 2)-[rhamnosyl-(1 --> 6)-glucoside]. Referring to previous reports, flavonols occurring in carnation flowers are characterized as kaempferol 3-O-glucosides with additional sugars binding at the 2 and/or 6-positions of the glucose. The kaempferol glycoside contents of a nearly pure white flower and some creamy white flower lines were compared. Although the major glycoside was different in each line, the total kaempferol contents of the creamy white lines were from 5.9 to 20.9 times higher than the pure white line. Thus, in carnations, kaempferol glycosides surely contribute to the creamy tone of white flowers.     

Antiradical and cytotoxic activity of different Helichrysum plicatum flower extracts

Flowers of Helichrysum plicatum were extracted under different experimental conditions, and their antioxidant activity was determined by DPPH radical scavenging assay. Extracts obtained with higher concentration of ethyl acetate (90% or 100%) were found to contain the greatest amount of total phenolics (> 250 mg gallic acid equivalents/g of dried extract), and high correlation between total phenolic content and antiradical activity was observed (r = -0.79). Based on the total phenolic content and antiradical activity, some extracts were selected for investigation of cytotoxic activity toward PC3, HeLa and K562 human cancer cell lines in vitro. All tested extracts exhibited moderate activity against HeLa cells (41.9-42.1 microg/mL), whereas the extract obtained with 100% ethyl acetate was the most active against K562 and PC3 cell lines (25.9 and 39.2 microg/mL, respectively). Statistical analysis revealed significant correlation between total phenolic content and cytotoxic activity against PC3 and K562 cells. HPLC identification of phenolic compounds from the extracts indicated the presence of apigenin, naringenin and kaempferol as free aglycones, and glycosides of apigenin, naringenin, quercetin and kaempferol. Among aglycones, kaempferol displayed moderate cytostatic activity against all cell lines (24.8-64.7 microM).     

The electron transfer reactivity of kaempferol and its interaction with amino acid residues

In this work, the electron transfer reactivity of kaempferol was studied and the interaction in vivo between kaempferol and protein was simulated. Dimethylsulfoxide (DMSO) as an aprotic solvent was employed to simulate the specific environment. Various residues of amino acids were used to study the effect of the amino acids in the active site of protein on the electron transfer reactivity of kaempferol. Experimental results revealed that the redox activity of kaempferol was different in aprotic medium DMSO from that in water, and a new redox process was further found. Of all the residues tested, nitrogenous nucleophile, for example, imidazole, was observed to be able to facilitate the electron transfer of kaempferol, and the mechanism was also proposed. This work might provide a simple model to study the electron transfer reactivity of some small active organic molecules, especially medicines, in specific environment, which might approach a more accurate understanding of the activity of some medicines in vivo.     

Study on the phenolic constituents of the flowers and leaves of Trifolium repens L

The flowers and leaves of Trifolium repens L. (Fabaceae) were subjected to phytochemical investigation in order to identify their major chemical constituents and to evaluate in?vitro antioxidant activity of the isolated compounds against DPPH˙. A total of 12 flavonoids, pterocarpan and methyl caffeate were isolated, then characterised by UV, MS, NMR spectroscopy and identified as quercetin and kaempferol 3-O-(6″-α-rhamnopyranosyl-2″-β-xylopyranosyl)-β-galactopyranosides (1, 2), kaempferol 3-O-(2″,6″-α-dirhamnopyranosyl)-β-galactopyranoside, mauritianin (3), quercetin and kaempferol 3-O-(2″-β-xylopyranosyl)-β-galactopyranosides (4, 5), kaempferol and quercetin 3-O-β-(6″-O-acetyl)-galactopyranosides (6, 7), trifolin (8), hyperoside (9), myricetin 3-O-β-galactopyranoside (10), quercetin (11), ononin (12), medicarpin 3-O-β-glucopyranoside (13) and methyl caffeate (14). Mauritianin, ononin, pterocarpan and methyl caffeate have been reported in this plant for the first time. The compounds 4, 7, 9, 10, and 11 were tested for their antioxidant effect against DPPH˙. All studied compounds were found to have potent activity, but the most effective in the test were compounds 9, 10 and 11 (EC(50) values in the range 7.51-9.52?μM).     

Kaempferol tri- and tetraglycosides from the flowers of Clematis cultivar

A new kaempferol glycoside, kaempferol 3-O-alpha-rhamnopyranosyl-(1 --> 2)-[alpha-rhamnopyranosyl-(1 --> 6)-beta-glucopyranoside]-7-O-beta-glucopyranoside (2) was isolated from the flowers of Clematis cultivar "Jackmanii Superba", together with a known kaempferol 3-O-alpha-rhamnopyranosyl-(1 --> 6)-beta-glucopyranoside-7-O-beta-glucopyranoside (1). The chemical structures of the isolated glycosides were established by UV, LC-MS, characterization of acid hydrolysates, and 1H and 13C NMR spectroscopy.     

Study on the interactions of kaempferol and quercetin with intravenous immunoglobulin by fluorescence quenching, fourier transformation infrared spectroscopy and circular dichroism spectroscopy

The interactions of kaempferol and quercetin with intravenous immunoglobulin (IVIG) were studied in vitro by spectroscopic methods including fluorescence spectra, Fourier transformation infrared (FT-IR) spectra and circular dichroism (CD) spectra. The binding parameters for the reactions calculated according to the Sips equation suggested that the bindings of IVIG to kaempferol and quercetin were characterized by two binding sites with the average affinity constants K(o) at 1.032 x 10(4) M(-1) and 1.849 x 10(4) M(-1), respectively. The binding of IVIG with quercetin is stronger than that of IVIG with kaempferol. They were of non-specific and weak drug-protein interactions. Docking was used to calculate the interaction modes between kaempferol and quercetin with IVIG. The secondary structural compositions of free IVIG and its kaempferol, quercetin complexes were calculated by the FT-IR difference spectra, self-deconvolution, second derivative resolution enhancement and the curve-fitting procedures of amide I band respectively, which are in good agreement with the analyses of CD spectra. The effect of 3'-OH substituent in quercetin is distinct between the interactions of IVIG with kaempferol and quercetin for the secondary structure of the protein. The observed spectral changes indicate a partial unfolding of the protein structure, but the typical beta structural conformation of IVIG is still retentive in the presence of both drugs in aqueous solution. The average binding distances between the chromophores of IVIG with kaempferol (4.30 nm) and quercetin (4.35 nm) were obtained on the basis of the theory of F?rster energy transfer. IVIG can serve as transport protein (carrier) for kaempferol and quercetin.     

液相色谱法同时测定维药蜀葵花中芦丁、槲皮素和山柰酚

维药是祖国医药学不可分割的组成部分。维药现代化,即利用现代技术研究维药的有效成分,是维药科学化、标准化、规范化、商品化和产业化的必经之路。本文建立了维药蜀葵花中有效成分芦丁、槲皮素和山柰酚的选择性提取方法,优化了高效液相色谱法(HPLC)同时测定这3种有效成分的分析条件。采用HC-C₁₈色谱柱(250 mm×4.6 mm, 5 μm)和甲醇-0.4%磷酸(50:50, v/v)流动相,在柱温30 ℃和流速1.00 mL/min的条件下实现了3种物质之间以及和干扰物之间的基线分离。维药蜀葵花中芦丁、槲皮素及山柰酚的线性范围分别为12.5~150 μg/mL (r=0.9998), 12.5~125 μg/mL (r=0.9999)及12.5~125 μg/mL (r=0.9988),加标回收率(n=5)分别为100.3%(RSD=1.1%)、97.60%(RSD=0.47%)、97.75%(RSD=0.71%)。该方法实现了同时测定维药蜀葵花中芦丁、槲皮素及山柰酚,为其他黄酮类物质的开发应用提供了科学依据,同时也可为其他维药分析提供借鉴。