紫甘薯花色苷色素的抑菌作用研究

本文研究了紫甘薯花色苷色素抑制大肠杆菌、金黄色葡萄球菌、啤酒酵母和黑曲霉的作用及机理.结果表明:紫甘薯花色苷色素对大肠杆菌及金黄色葡萄球菌均有抑制作用,并与其浓度呈正相关,而对啤酒酵母和黑曲霉无抑制作用.透射电镜观察和大肠杆菌生长曲线表明,该色素的抑菌作用可能是通过增强细胞膜的通透性,使细胞异常生长,抑制对数生长期的细胞分裂,使细胞质稀薄、细胞解体.SDS-PAGE分析表明,紫甘薯花色苷对大肠杆菌蛋白表达影响不明显,未见特征性条带的消失,仅对部分蛋白质合成量有影响.     

紫甘薯花色苷色素的抑菌作用研究

本文研究了紫甘薯花色苷色素抑制大肠杆菌、金黄色葡萄球菌、啤酒酵母和黑曲霉的作用及机理。结果表明：紫甘薯花色苷色素对大肠杆菌及金黄色葡萄球菌均有抑制作用, 并与其浓度呈正相关, 而对啤酒酵母和黑曲霉无抑制作用。透射电镜观察和大肠杆菌生长曲线表明, 该色素的抑菌作用可能是通过增强细胞膜的通透性, 使细胞异常生长, 抑制对数生长期的细胞分裂, 使细胞质稀薄、细胞解体。SDS-PAGE 分析表明, 紫甘薯花色苷对大肠杆菌蛋白表达影响不明显, 未见特征性条带的消失, 仅对部分蛋白质合成量有影响。     

响应面法优化紫甘薯花色苷水解条件

使用响应面法优化了紫甘薯花色苷的水解条件.使用高效液相色谱-多波长检测器测定花青素含量.采用3因素3水平Box-Behnken中心组合设计,优化了水解时间、盐酸浓度和水解温度对矢车菊素和芍药素峰面积之和的综合影响.结果显示,紫甘薯花色苷的最佳水浴水解条件为:盐酸浓度4.54 mol/L,水浴温度100℃,水解时间45.8 min.在最优条件下进行验证试验,响应值为理论值的96.5％,说明该响应面法优化方法可行.     

耐盐紫甘薯花色苷组分分析和抗氧化性研究

利用高效液相色谱与二极管阵列检测器/电喷雾质谱联用技术研究了耐盐紫甘薯Z103中的花色苷类化合物,确定该品种含有15种花色苷,主要为被咖啡酸、阿魏酸、对羟基苯甲酸等芳香酸酰化的矢车菊苷和芍药苷,其中矢车菊素3-O-对羟基苯甲酰-槐糖苷-5-O-葡糖苷、芍药素3-O-对羟基苯甲酰-槐糖苷-5-O-葡糖苷、芍药素3-O-阿魏酰-槐糖苷-5-O-葡糖苷为首次报道;同时进一步考察了不同体系中,紫甘薯花色苷抑制脂质过氧化能力和对DPPH·、O-·2和HO·的清除作用,结果表明紫甘薯花色苷具有较强的抗氧化能力,且均具有量效关系。紫甘薯花色苷(0.4 mg/mL)对脂质体氧化的抑制率为83.24%,对DPPH·(0.20 mg/mL)、O-·2(4 mg/mL)和HO·(30μg/mL)的清除率分别为94.06%、96.62%和96.12%。     

紫甘薯花青素对肝癌的影响及其机制的研究

为研究紫甘薯花青素对体外培养的人肝癌细胞SNU-387的影响及其可能的机制。在不同终浓度紫甘薯花青素处理下,利用MTT法和Hoechst 33258染色法,测定人肝癌细胞SNU-387增殖活力,通过测定细胞TNF-α含量来考察死亡受体TNFR1介导的外源性凋亡通路的作用,通过测定细胞内Caspase-8表达量变化来考察NF-κB和MAPK信号通路的作用。结果表明在细胞凋亡实验中,不同浓度紫甘薯花青素作用下,人肝癌细胞SNU-387增殖活力降低,出现核固缩等凋亡特征,细胞内TNF-α含量和Caspase-8表达量提高;在阻断剂PDTC作用下,Caspase-8表达量无差异;在阻断剂SB203580作用下,Caspase-8表达量下降。综上,紫甘薯花青素可以通过死亡受体TNFR1介导的外源性凋亡通路和MAPK信号通路参与介导肝癌细胞SNU-387的凋亡过程。     

黑果腺肋花楸花色苷对人皮肤成纤维细胞紫外辐射损伤的保护作用

为了探讨黑果腺肋花楸花色苷对紫外辐射所致人皮肤成纤维细胞氧化损伤的保护作用,将体外培养的人皮肤成纤维细胞分为对照组、辐射组和辐射给药组。采用MTT法检测不同浓度花色苷添加量对细胞增殖的保护作用,以选择最优添加浓度。采用化学荧光法检测细胞活性氧(ROS)含量,ELISA法检测细胞MMP-1分泌水平。结果表明:UVA辐照剂量为10 J/cm2条件下,与辐射组相比,MTT法显示花色苷添加组浓度为125μg/m L对损伤细胞增殖保护作用有极显著的提高(P0.01),同时,125μg/m L花色苷添加组能够显著降低辐射损伤后细胞ROS含量以及MMP-1分泌水平(P0.01)。由此可见,适宜浓度的黑果腺肋花楸花色苷能够降低细胞ROS含量及MMP-1分泌水平,从而对辐射损的细胞起到一定的保护作用。     

三七绿紫过渡地上茎的花色苷和皂苷组织定位及其含量分析

为探究三七绿紫过渡地上茎的花色苷和皂苷组织定位与含量的相关性,采用显微组织化学法研究云南文山三七的一年生植株绿紫过渡地上茎各茎段花色苷和皂苷的组织定位,用分光光度法检测茎段的总花色苷(TAC)和总皂苷含量(TSC),用高效液相色谱检测了茎段的皂苷单体含量。结果表明:(1)在三七绿紫过渡地上茎的中部横截面上,花色苷主要定位在皮层薄壁组织外侧的2层或2~3层细胞中,而皂苷主要定位在维管束中;各茎段的皂苷单体均主要为人参皂苷Rb1。(2)从茎顶向茎基,茎段中TAC、TSC和Rb1的含量总体上分别表现为一条"单峰"、"V形"和"降-升-降三段式"曲线;其中,花色苷主要积累在茎的中、上部,总皂苷在茎的下、基部,Rb1则在茎的上半段,而且TAC最高以及TSC和Rb1含量最低的茎段均恰好定位在中上部的黄金分割点处。(3)不同茎段间的TAC含量差异显著,Rb1含量差异极显著,但TSC含量的差异不显著;不同茎段间的TAC与TSC、Rb1含量呈不同的相关性,整个地上茎的TAC与TSC含量间呈极显著负相关关系、TAC与Rb1含量间呈不显著正相关。研究认为,在三七绿紫过渡地上茎中,花色苷和皂苷的横向组织定位不同,二者含量在纵向上总体呈负相关。     

MTT法检测石刁柏悬浮细胞技术的研究

本研究确定了MTT法测定悬浮培养的石刁柏细胞活力所需的最适条件为:比色波长560nm,最适缓冲液为柠檬酸钠-Vc缓冲液,渗透促进剂为二甲基亚砜,洗脱剂为酸化异丙醇,0.133%浓度的MTT溶液适于对未知培养时期的细胞计数,0.267‰浓度的MTT溶液适于鉴定细胞活力;同时,本研究通过MTT对培养细胞的生长过程进行了测定,确定细胞活性最强时期为指数期;并研究了细胞数量与吸光值之间的关系以及培养时间对MTT染色的影响.     

与内生真菌共培养对葡萄果肉细胞生理生化影响的差异研究

根据与内生真菌共培养过程中对葡萄细胞生理生化的影响差异将内生真菌菌株归类,筛选出具有不同利用价值的内生真菌资源。以分离于玫瑰蜜、赤霞珠和夏黑葡萄叶片的18个属47株内生真菌和佳美葡萄(Vitis vinifera L. cultivar:Gamay)果肉愈伤组织为试验材料,构建内生真菌与葡萄细胞共培养体系,并探索这些内生真菌菌株对葡萄果肉细胞生长、花色苷含量、苯丙氨酸解氨酶(PAL)活性等的生理生化影响。结果表明,与不同内生真菌共培养对葡萄细胞生长、花色苷总量和PAL活性都有不同程度的影响。其中共筛选出20株对葡萄细胞伤害较小的内生真菌;6株显著促进葡萄细胞生长的内生真菌;11株显著促进葡萄细胞PAL活性的内生真菌;5株促进葡萄细胞花色苷总量的内生真菌。此次研究为发掘和利用内生真菌资源调控葡萄生化品质特征提供了技术参考和一定的物质基础。     

结核分枝杆菌融合蛋白Ag85B-Hsp16.3、Ag85B-ESAT6 和分泌蛋白 Hsp16.3 体外对人肝癌细胞HepG-2 的作用*

目的:探讨结核分枝杆菌融合蛋白Ag85B-Hsp16.3、Ag85B-ESAT6及分泌蛋白Hsp16.3对人肝癌细胞HepG-2的作用。方法:将已构建的含3种目的基因的表达载体pProEXHTa-Ag85B-Hsp16.3、pProEXHTa-Ag85B-ESAT6和pProEXHTb-Hsp16.3,分别转入宿主菌E.coliDH5α中,诱导表达后分别获得Ag85B-Hsp16.3、Ag85B-ESAT6和Hsp16.3三种蛋白,采用Ni2+亲和层析柱进行纯化,并用透析方法进行目的蛋白的复性。复性的蛋白按照不同浓度和作用时间分别与肝癌细胞HepG-2反应,用MTT法检测细胞生长情况。结果:三种蛋白被成功纯化并复性。MTT数据统计分析显示,终浓度10μg/ml的三种蛋白对HepG-2细胞生长没有明显作用,当三种蛋白的终浓度分别为20、40、80μg/ml时均能够抑制HepG-2细胞的生长,并且抑制作用随着蛋白终浓度的增大以及作用时间的延长而增强。不同类别的蛋白抑制作用没有明显差别。结论:结核分枝杆菌的部分分泌蛋白能够抑制肝癌细胞HepG-2的生长。     

Protective effect of supplemental anthocyanins on Arabidopsis leaves under high light

Ten anthocyanin components have been detected in roots of purple sweet potato (Ipomoea batatas Lam.) by high‐performance liquid chromatography coupled to diode array detection and electrospray ionization tandem mass spectrometry. All the anthocyanins were exclusively cyanidins or peonidin 3‐sophoroside‐5‐glucosides and their acylated derivatives. The total anthocyanin content in purple sweet potato powder obtained by solid‐phase extraction was 66 mg g^?1. A strong capacity of purple sweet potato anthocyanins (PSPA) to scavenge reactive oxygen species (superoxide, hydroxyl radical) and the stable 1,1‐diphenyl‐2‐picrylhydrazyl organic free radical was found in vitro using the electron spin resonance technique. To determine the functional roles of anthocyanins in leaves in vivo, for the first time, supplemental anthocyanins were infiltrated into leaves of Arabidopsis thaliana double mutant of the ecotype Landsberg erecta (tt3tt4) deficient in anthocyanin biosynthesis. Chlorophyll fluorescence imaging showed that anthocyanins significantly ameliorated the inactivation of photosystems II during prolonged high‐light (1300 µmol m^?2 s^?1) exposure. Comet assay of DNA revealed an obvious role of supplemental PSPA in alleviating DNA damage by high light in leaves. Our results suggest that anthocyanins could function in vitro and in vivo to alleviate the direct or indirect oxidative damage of the photosynthetic apparatus and DNA in plants caused by high‐light stress.     

Antioxidative activity of anthocyanins from purple sweet potato, Ipomoera batatas cultivar Ayamurasaki

We evaluated the antioxidative activity of anthocyanins from an extract of the tuber of purple sweet potato (PSP) (Ipomoea batatas cultivar Ayamurasaki). Anthocyanins from PSP showed stronger 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity than anthocyanins from red cabbage, grape skin, elderberry, or purple corn, and eight major components of the anthocyanins from PSP showed higher levels of activity than ascorbic acid. In PSP anthocyanin-injected rats and PSP beverage-administered volunteers, DPPH radical-scavenging activity in the urine increased. The elevation of plasma transaminase activities induced by carbon tetrachloride was depressed in rats administered PSP anthocyanin solution. Two components, cyanidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D-glucopyranocyl)-beta-D-glucopyranoide)-5-O-beta-D-glucopyranoside and peonidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D-glucopyranocyl)-beta-D-glucopyranoide)-5-O-beta-D-glucopyranoside, which were detected in the plasma, protected low density lipoprotein from oxidation at a physiological concentration. These results indicate that PSP anthocyanins have antioxidative activity in vivo as well as in vitro.     

Evaluation of viability assays for anthocyanins in cultured cells

The MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay is a widely accepted cytotoxicity assay which can produce inaccurate results due to possible interference with the antioxidant property of anthocyanins. Alternative methods to the MTT assay, such as BrdU (DNA-based) and CellTiter-Glo (ATP-based) assays were evaluated to assess anthocyanin cytotoxicity, derived from blackberry in LNCaP, MCF-7 and MDA-MB-453 cell lines. The standard cell counting method was the reference assay. Greater correlation of cell viability values following anthocyanin exposure was obtained from multiple cell lines with the alternative assays when compared with cell counting. MTT and cell counting results were not always correlated, albeit this was a function of cell type. In particular, poor correlations between cell counting and MTT procedures used to assess cytotoxicity of anthocyanins were observed in the MDA-MB-453 cell lines. Comparison of cytotoxicity derived from alternative assays and the MTT assays with the cell counting method was dependent on the assay procedure and the cell type. The LC(50) of blackberry crude extract ranged from 0.4 to 9.4 mg/mL between assays and across all cell lines, whereas a semi-purified anthocyanin extract was not cytotoxic. Cytotoxicity evaluation of polyphenolic-rich extracts using BrdU and CellTiter-Glo assays as alternatives to the MTT method is recommended.     

Absorption of acylated anthocyanins in rats and humans after ingesting an extract of Ipomoea batatas purple sweet potato tuber

We evaluated the absorbability of anthocyanins in humans and rats administered with a beverage prepared from an extract of the tuber of purple sweet potato (Ipomoea batatas Cultivar Ayamurasaki), or with an anthocyanin concentrate. Two major anthocyanin components, cyanidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D-glucopyranosyl)-beta-D-glucopyranoside)-5-O-beta-D-glucopyranoside) and peonidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D-glucopyranosyl)-beta-D-glucopyranoside)-5-O-beta-D-glucopyranoside), were detected in the plasma and urine of both rats and humans by HPLC or liquid chromatography/mass spectrometry (LC/MS). The plasma concentration of anthocyanins in humans reached a maximum 90 minutes after ingestion, and the recovery of anthocyanins in the urine was estimated as 0.01-0.03%. These results indicate that acylated anthocyanins could be selectively absorbed after ingesting food.     

菊花不同花色品种中花青素苷代谢分析

应用高效液相色谱和多级质谱联用技术（HPLC-ESI-MSn）,分析菊花（Chrysanthemum×morifolium）白色、粉色、红色、紫色、红紫色和墨色6个色系共计82个品种中花青素苷合成过程的中间产物和最终产物,发现从白色、粉色、红色、紫色、红紫色到墨色花青素苷含量快速增加,分别为4.68、111.60、366.89、543.56、1220.36和2674.95μg·g-1,不同色系间花青素苷的含量差异显著（P〈0.01）,花青素苷含量越高花色越深;墨色菊花品种中总类黄酮含量显著高于其它花色品种（P〈0.01）,其它不同色系间总类黄酮含量差异不显著（P〉0.05）;随着菊花花色变深,从柚皮素分支到圣草酚的代谢流,以及从圣草酚分支到矢车菊素苷的代谢流比例增加。花青素苷成分分析发现：菊花中只含有矢车菊素苷类化合物。根据花青素苷代谢成分分析结果绘制了菊花中花青素苷代谢路径图,即在菊花类黄酮代谢途径中只存在矢车菊素苷代谢分支途径;菊花不同色系在柚皮素和圣草酚2个关键代谢分支点上向不同方向代谢流的分配比例不同,造成花青素苷产物含量不同,导致不同花色。以上研究结果为菊花花色改良的分子育种提供了理论依据。     

菊花不同花色品种中花青素苷代谢分析

应用高效液相色谱和多级质谱联用技术(HPLC-ESI-MSⁿ), 分析菊花(Chrysanthemum × morifolium)白色、粉色、红色、紫色、红紫色和墨色6个色系共计82个品种中花青素苷合成过程的中间产物和最终产物, 发现从白色、粉色、红色、紫色、红紫色到墨色花青素苷含量快速增加, 分别为4.68、111.60、366.89、543.56、1 220.36和2 674.95 μg·g^–1, 不同色系间花青素苷的含量差异显著(P<0.01), 花青素苷含量越高花色越深; 墨色菊花品种中总类黄酮含量显著高于其它花色品种(P<0.01), 其它不同色系间总类黄酮含量差异不显著(P>0.05); 随着菊花花色变深, 从柚皮素分支到圣草酚的代谢流, 以及从圣草酚分支到矢车菊素苷的代谢流比例增加。花青素苷成分分析发现: 菊花中只含有矢车菊素苷类化合物。根据花青素苷代谢成分分析结果绘制了菊花中花青素苷代谢路径图, 即在菊花类黄酮代谢途径中只存在矢车菊素苷代谢分支途径;菊花不同色系在柚皮素和圣草酚2个关键代谢分支点上向不同方向代谢流的分配比例不同, 造成花青素苷产物含量不同,导致不同花色。以上研究结果为菊花花色改良的分子育种提供了理论依据。     

Expression of a vacuolar protein (VP24) in anthocyanin-producing cells of sweet potato in suspension culture.

VP24, an abundant protein of 24 kD, was found to accumulate in the anthocyanin-containing vacuoles of cells of sweet potato (Ipomoea batatas) in suspension culture. Light-induced expression of VP24 was analyzed by immunoblotting in three different cell lines that produced anthocyanins at different rates. The expression of VP24 was closely correlated with the accumulation of anthocyanin in these cell lines. Immunocytochemical detection of VP24 with specific antibodies on thin sections showed that VP24 was localized in the intravacuolar pigmented globules (cyanoplasts) in the anthocyanin-containing vacuoles and not in the tonoplast. No VP24 immunogold labeling was detected in the vacuoles of the cell line that does not produce anthocyanin. We suggest that VP24 may be involved in the formation of the cyanoplast via an interaction with anthocyanin, and that it may play an important role in the trapping in vacuoles of large amounts of anthocyanins that have been transported into these vacuoles.     

Study and characterization of a novel functional food: purple popcorn

Many phytonutrients seem to be able to combat the effects of oxidants which may lead to chronic diseases. Among them, anthocyanins have been studied for a long time, and different types of functional foods rich in these pigments are already available on the market. In particular, wine, berries and various cereals have already aroused consumers’ awareness, and in this context we propose a new and attractive healthy food: purple popcorn. Popcorn is the most popular American snack, now well known all over the world. A corn rich in anthocyanins, suitable to be transformed into a snack, could help to introduce healthy antioxidant compounds into the diet of many people, contributing to the prevention of chronic diseases. In this work we developed a coloured popcorn variety rich in anthocyanins (about 66 mg/100 g, mainly cyanidin) by a recurrent selection scheme, with the aim of obtaining a healthier snack. The selection was based on some quality characteristics such as anthocyanin content, popping ability and the popping expansion volume. The purple popcorn obtained was further analyzed by high pressure liquid chromatography and 2,2-diphenyl-1-picrylhydrazyl radical scavenging ability, before and after microwave treatment. The results obtained showed that, even though the microwave treatment reduced the anthocyanin content to about 46 %, the remaining anthocyanins exhibited a marked antioxidant capacity compared to the colourless control. Finally taste perception was also checked between coloured and uncoloured popcorn, and no difference was perceived.     

Linkage of the <Emphasis Type="Italic">A</Emphasis> locus for the presence of anthocyanin and <Emphasis Type="Italic">fs10.1</Emphasis>, a major fruit-shape QTL in pepper

The purple color of the foliage, flower and immature fruit of pepper ( Capsicum spp.) is a result of the accumulation of anthocyanin pigments in these tissues. The expression of anthocyanins is controlled by the incompletely dominant gene A. We have mapped A to pepper chromosome 10 in a Capsicum annuum (5226) x Capsicum chinense (PI 159234) F(2) population to a genomic region that also controls anthocyanin expression in two other Solanaceous species, tomato and potato, suggesting that variation for tissue-specific expression of anthocyanin pigments in these plants is controlled by an orthologous gene(s). We mapped an additional locus, Fc, for the purple anther filament in an F(2) population from a cross of IL 579, a C. chinense introgression line and its recurrent parent 100/63, to the same position as A, suggesting that the two loci are allelic. The two anthocyanin loci were linked to a major quantitative trait locus, fs10.1, for fruit-shape index (ratio of fruit length to fruit width), that also segregated in the F(2) populations. This finding verified the observation of Peterson in 1959 of linkage between fruit color and fruit-shape genes in a cross between round and elongated-fruited parents. The linkage relationship in pepper resembles similar linkage in potato, in which anthocyanin and tuber-shape genes were found linked to each other in a cross of round and elongated-tuber parents. It is therefore possible that the shape pattern of distinct organs such as fruit and tuber in pepper and potato is controlled by a similar gene(s).