老黑谷米中多酚化合物的提取及体外抗氧化活性研究

以老黑谷米面粉为原料,提取分离其中的可溶性多酚、游离多酚、酯化多酚、醚化多酚、键合多酚,利用比色法进行多酚和类黄酮含量测定,并采用DPPH自由基清除能力、铁离子还原/抗氧化能力测定（FRAP）、总抗氧化能力测定（TEAC）、还原能力测定（RP）和H2O2清除活性等5种不同的体外抗氧化测定体系进行抗氧化活性测定。结果表明：5种多酚中,可溶性多酚含量最高,酯化多酚含量次之,显著高于游离多酚、醚化多酚、键合多酚。多酚中类黄酮含量最高为键和多酚和可溶性多酚,显著高于其他3种多酚。5种多酚均有较高的体外抗氧化能力,键合多酚的DPPH自由基清除能力最高,可溶性多酚的H2O2清除能力、FRAP还原能力、RP还原能力和TEAC总抗氧化能力最高。本研究为老黑谷米的开发利用提供科学依据。     

诺丽发酵果汁中抗氧化物质的研究

本实验针对茜草科植物诺丽的发酵果汁中抗氧化物质进行了定性和定量研究。采用系统预实验法对其不同极性的洗脱部位可能含有的化学成分进行预实验,探索了各个部位的化学成分。采用DPPH·自由基清除、ABTS+·自由基清除和FRAP三种实验方法对各部位洗脱物的抗氧化活性进行了测定,结果显示50%~90%乙醇洗脱部位抗氧化能力最强,此部位主要含有多酚类、黄酮类、香豆素内酯类和蒽醌类成分,并且研究表明抗氧化能力与多酚成分的含量成极显著相关。本研究结果表明多酚类物质是诺丽发酵果汁发挥抗氧化作用的主要功能成分。     

凤仙花不同部位乙醇提取物多酚、黄酮含量测定及抗氧化活性研究

采用55%的乙醇分别回流提取凤仙花的茎、叶、花中的活性物质,采用比色法测定各部位的多酚及总黄酮含量,测定各提取物对DPPH、羟自由基、超氧阴离子自由基的清除能力和还原力等抗氧化活性指标。结果表明,凤仙花各部位的多酚(以没食子酸计)、总黄酮(以芦丁计)含量由高到低依次为凤仙花的叶、花、茎,多酚含量分别为72.81、66.07、33.07 mg/m L,总黄酮含量分别为112.13、108.06、30.62 mg/m L。各种提取物均有不同程度的抗氧化活性,且抗氧化活性与提取物浓度存在量效关系,其中叶提取物清除DPPH自由基、羟自由基、超氧阴离子自由基活性以及还原力均最高,本研究为凤仙花作为低毒高效的天然抗氧化剂应用提供了依据。     

荸荠皮提取物对DPPH自由基清除活性

采用70%丙酮溶液对荸荠皮中抗氧化物质进行提取,得到红棕色浸膏。通过定性及定量方法分析了荸荠皮提取物中可能存在的具有抗氧化活性的物质;采用DPPH自由基法测定了荸荠皮提取物对DPPH自由基的清除能力。结果显示,荸荠皮提取物中含有多酚类和黄酮类等化合物,其多酚含量为3.31%(w/w,以干物质计)。DPPH自由基法显示,荸荠皮提取物具有一定的清除DPPH自由基能力,其清除能力与提取物浓度之间显示出良好的剂量-效应关系。该提取物(IC50值为130.37 ppm)对DPPH自由基清除能力略低于BHT(IC50值为94.16 ppm)。     

黔产接骨草中总多酚的含量测定及抗氧化活性研究

以黔产接骨草药材为研究对象,建立了福林酚比色法检测接骨草中总多酚含量方法;并采用单因素结合响应面优化设计法优化接骨草中总多酚提取条件;在此基础上,同时测定接骨草不同产地、不同药用部位总多酚的含量,并以清除1,1-二苯基-2-三硝基苯肼(DPPH)自由基和ABTS自由基的能力为评价指标,对接骨草不同药用部位抗氧化活性进行比较研究。结果表明,不同产地、不同药用部位接骨草中总多酚含量差异较大;同批次接骨草中总多酚含量依次为叶全草茎根;接骨草中叶、全草、茎和根清除DPPH的IC50值分别为1.14、1.70、2.25、2.65 mg/m L,清除ABTS的IC50值分别为1.16、1.56、2.17、3.15 mg/m L,且活性大小与总多酚质量浓度呈现明显量效关系。该方法简便、快捷、重复性好,为接骨草基于总多酚含量的质量评价和质量控制提供参考,以IC50作为判定抗氧化活性强弱的指标,4个不同药用部位中以接骨草叶提取物抗氧化能力相对较强。     

紫薇花的抗氧化活性研究

采用DPPH法、ABTS法和FRAP三种测定法对银薇和红花紫薇体外抗氧化活性进行综合评价,并与阳性对照二丁基羟基甲苯(BHT)比较。研究结果发现紫薇花具有较好的抗氧化活性。银薇乙酸乙酯部位清除DPPH自由基的能力(IC50=7.4μg/m L)、清除ABTS自由基的能力(IC50=1.8μg/m L)和还原Fe3+的能力(TEAC=2664.7μmol/g)均强于阳性对照BHT(DPPH方法:IC50=23μg/m L;ABTS方法:IC50=2.3μg/m L;FRAP方法:TEAC=1532.7μmol/g),银薇乙酸乙酯部位抗氧化能力最强。     

冷蒿酚酸及其抗氧化防御酶活性对机械损伤的响应

为了探讨放牧过程中冷蒿(Artemisia frigida)酚酸及其抗氧化酶活性对牲畜采食和践踏损伤的响应,该研究对盆栽冷蒿幼苗枝叶采用不同程度(轻度、中度和重度)机械损伤的方式模拟放牧强度,利用高效液相色谱技术测定了机械损伤处理6 h后冷蒿枝叶和根系9种酚酸含量的变化;同时对多酚氧化酶(PPO)、苯丙氨酸氨裂合酶(PAL)和抗氧化酶活性进行了测定,分析了酚酸含量与PPO和PAL活性之间的相关性。结果表明:轻度机械损伤可以诱导冷蒿枝叶和根系中PPO、PAL和抗氧化酶迅速做出响应,活性显著升高;随着机械损伤强度增加,冷蒿枝叶游离态咖啡酸、丁香酸、阿魏酸和肉桂酸含量显著增加,与对照相比中度损伤处理分别增加了150.4%、93.5%、154.4%和121.7%,与PAL活性呈正相关关系;游离态绿原酸和邻苯二酚含量减少,与对照相比重度损伤处理后降低了91.1%和69.3%,与PPO活性呈负相关关系;与对照相比重度损伤处理下没食子酸和原儿茶酸分别增加了280.6%和215.7%。随损伤强度增加,根系中9种游离态酚酸含量均显著增加,其含量均小于叶片。机械损伤导致冷蒿枝叶中游离态和结合态酚酸总量增加,根系中结合态酚酸减少。上述结果表明机械损伤首先诱导冷蒿体内应激防御系统中抗氧化酶和酚酸合成关键酶活性升高,然后促进抗氧化物质酚酸的积累,调节冷蒿植株内木质素合成以及醌类和单宁形成,增强冷蒿抗机械损伤能力和提高冷蒿的耐牧性。     

丹参生品及炮制品的抗氧化活性研究

采用清除二苯代苦味酰基(DPPH)自由基、清除[2,2'-连氨-(3-乙基苯并噻唑啉-6-磺酸)二铵盐](ABTS)自由基及铁离子还原/抗氧化能力(FRAP)测定法,以二丁基羟基甲苯(BHT)为阳性对照,对丹参生品及炮制品进行抗氧化活性评价。实验结果表明,丹参生品及其炮制品均有一定的抗氧化活性。其中,丹参炭乙酸乙酯部位清除DPPH自由基的能力最强,IC50值为13.9μg/mL;炒丹参乙酸乙酯部位、酒丹参乙酸乙酯部位和丹参炭正丁醇部位清除ABTS自由基能力最强,IC50值均为2.1μg/mL;米丹参乙酸乙酯部位的FRAP值最高为1517.81μmol/g。不同炮制方法对丹参抗氧化活性的能力有所不同,其中,丹参炭的整体抗氧化活性相对较好。     

药用真菌桑黄液体培养过程中的抗氧化活性研究

桑黄Sanghuangporus sanghuang是一种珍稀的药用真菌,具有较大的应用潜力。本研究主要以菌丝体生物量、多酚含量、黄酮含量、抗坏血酸(Ascorbic acid,AA)含量、总抗氧化能力(Total antioxidant capacity,T-AOC)、抑制羟自由基能力(Restraining ability to hydroxyl free radicals,RAHFR)、超氧阴离子清除能力、1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基清除能力、2,2’-连氮-双(3-乙基苯并噻唑-6-磺酸)[2,2’-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid),ABTS]自由基清除能力、铁离子还原能力(Ferric reducing antioxidant power,FRAP)和亚铁离子螯合能力为测定指标,对桑黄液体培养过程中的抗氧化活性进行了评价。结果显示,菌丝体生物量在2–10d内增长迅速,第10天达到最大值,在此过程中AA含量和T-AOC也出现了峰值,且多酚含量、黄酮含量和FRAP均呈上升趋势,说明菌株的抗氧化活性与其自身的生长状况、次级代谢产物分泌及还原能力等密切相关。此外,该菌株对羟自由基、超氧阴离子、DPPH自由基和ABTS自由基的清除能力也较强,于第12天、第10天、第2天和第14天分别达到73.06U/m L、46.78%、86.47%和94.16%,显示了较强的抗氧化活性。较高的亚铁离子螯合能力也说明桑黄在氧化胁迫下可启动自身的抗氧化系统以阻断自由基链反应。研究结果为更好地研究、开发和利用药用真菌桑黄提供了基础资料。     

苦碟子注射液抗氧化成分的分离与活性评价研究

苦碟子注射液经盐酸酸化,HPD大孔树脂柱分离,得到总酚酸部分,以抗坏血酸(Vc)做为阳性对照,利用清除二苯代苦味酰基(DPPH)自由基法和铁离子还原/抗氧化能力(FRAP)法对其进行抗氧化活性评价,研究结果表明,总酚酸部分具有较好的抗氧化能力。为此,我们利用大孔树脂柱色谱、MDS-5反相制备色谱对总酚酸进一步分离,共获得6个酚酸类成分。采用上述活性评价的方法对所获得的6个成分进行了抗氧化活性评价,发现除芹菜素-7-O-β-D葡萄糖醛酸苷外,其它5个酚酸类成分均具有很强的抗氧化活性,其清除DPPH自由基能力及总抗氧化能力好于或与阳性对照Vc相当。     

Optimization of theaflavin biosynthesis from tea polyphenols using an immobilized enzyme system and response surface methodology

Theaflavins were synthesized from tea polyphenols extracted from green tea using an immobilized polyphenol oxidase system. To optimize the production of theaflavins, response surface methodology was applied to determine the effects of five critical variables and their mutual interactions on theaflavin biosynthesis at five levels. A total of 52 individual experiments were performed and a statistical model predicted that the highest theaflavin concentration was 0.766mgml^–1 at optimized conditions. Using these optimal parameters under experimental conditions in three independent replicates, the average value of the biosynthesized theaflavin concentration reached 0.75±0.017mgml^–1 and matched the value predicted by the modelRevisions requested 03 November 2004; Revisions received 7 December 2004     

Separate monophenolase and o-diphenolase enzymes in Triticum aestivum

Monophenolase and o-diphenolase activities of polyphenol oxidase are usually thought to be a part of the same enzyme complex. It has now been demonstrated that the two catalytic activities of the polyphenol oxidase of wheat grains are separable and reside in different enzymes. The electrophoretically separated monophenolase enzyme showed specificity only for monophenol (l-tyrosine) after its elution from acrylamide gels. Further, this enzyme is confined to the endosperm tissue and is undetectable in the embryonic region of the seedling.     

Transthyretin Binding Heterogeneity and Anti-amyloidogenic Activity of Natural Polyphenols and Their Metabolites

Transthyretin (TTR) is an amyloidogenic protein, the amyloidogenic potential of which is enhanced by a number of specific point mutations. The ability to inhibit TTR fibrillogenesis is known for several classes of compounds, including natural polyphenols, which protect the native state of TTR by specifically interacting with its thyroxine binding sites. Comparative analyses of the interaction and of the ability to protect the TTR native state for polyphenols, both stilbenoids and flavonoids, and some of their main metabolites have been carried out. A main finding of this investigation was the highly preferential binding of resveratrol and thyroxine, both characterized by negative binding cooperativity, to distinct sites in TTR, consistent with the data of x-ray analysis of TTR in complex with both ligands. Although revealing the ability of the two thyroxine binding sites of TTR to discriminate between different ligands, this feature has allowed us to evaluate the interactions of polyphenols with both resveratrol and thyroxine preferential binding sites, by using resveratrol and radiolabeled T4 as probes. Among flavonoids, genistein and apigenin were able to effectively displace resveratrol from its preferential binding site, whereas genistein also showed the ability to interact, albeit weakly, with the preferential thyroxine binding site. Several glucuronidated polyphenol metabolites did not exhibit significant competition for resveratrol and thyroxine preferential binding sites and lacked the ability to stabilize TTR. However, resveratrol-3-O-sulfate was able to significantly protect the protein native state. A rationale for the in vitro properties found for polyphenol metabolites was provided by x-ray analysis of their complexes with TTR.     

Characterisation of phenol oxidase and peroxidase from maize silk

Silk of some maize genotypes contains a high level of phenolics that undergo enzymatic oxidation to form quinones, which condense among themselves or with proteins to form brown pigments. Two phenolic oxidizing enzymes, peroxidase (POD; EC 1.11.1.7) and polyphenol oxidase (PPO; EC 1.10.3.1), from maize (Zea mays L.) silk were characterised with respect to their preferred substrate, different isoforms and specific effectors. One browning silk sample with high, and two non‐browning samples with low phenolic content were investigated. Although POD oxidizes a wide range of phenolic substrates in vitro, its activity rate was independent of silk phenolic content. PPO activity, detected with o‐diphenolic substrates, was abundant only in browning silk, and low or absent in non‐browning silk. Pollination increased POD but not PPO activity. Isoelectric‐focusing (IEF) and specific staining for POD and PPO showed a high degree of polymorphism that varied with silk origin. The IEF pattern of POD revealed a number of anionic and several cationic isoenzymes, with the most pronounced having neutral pI 7 and a basic isoform with pI 10. Detected isoforms of PPO were anionic, except for one neutral form found only in browning silk, and occupied positions different from those of POD. Different inhibitory effects of NaN₃, EDTA, KCN, and L‐cysteine, as well as different impacts of a variety of cations on the oxidation of chlorogenic acid, mediated by PPO or POD, were detected. The findings are discussed in terms of a possible roles of these enzymes in defence and pollination.     

葡萄多酚物质提取方法的研究

对葡萄多酚物质提取分离影响因子进行正交试验,以确定各提取工艺路线的最佳条件,并对各种方法进行了比较。在实验室条件下,有机溶剂提取是进行葡萄多酚物质相关研究的适宜方法,其最佳因子组合为C3D2A2B1,即在70C,50％乙醇浓度,5倍体积比提取60min。同时对提取、萃取的次数和提取时材料的干燥程度进行了研究。     

逆相蒸发法制备茶多酚脂质体及质量评价

采用逆相蒸发法制备茶多酚脂质体并进行质量评价。通过二次回归旋转组合设计优化茶多酚脂质体制备工艺及配方,对其形态、结构、粒径分布等性质进行考察。研究结果表明,最佳配方为m（大豆卵磷脂）：m（胆固醇）=3：1、茶多酚质量浓度为7mg／mL、V（有机相）：V（水相）=4：1、磷酸盐缓冲液浓度15mmoL／L,此条件下包封率为50．37％;所制备的茶多酚脂质体形态呈圆球形或椭球形,为大单室脂质体,有效粒径为165．3nm,Zeta电位为-69．3mV。逆相蒸发法制备茶多酚脂质体方法简单可行,所制备的脂质体具有一定缓释性。     

Green tea polyphenol (−)‐epigallocatechin gallate suppressed the differentiation of murine osteoblastic MC3T3‐E1 cells

Recently, various physiological effects of the tea polyphenol catechin for alleviating diseases such as cancer, arteriosclerosis, hyperlipidaemia and osteoporosis have been reported. However, the physiological effect of catechin on bone metabolism remains unclear. We examined the physiological effect of EGCG [(?)‐epigallocatechin‐3‐gallate], which is the main component of green tea catechin, on osteoblast development using the precursor cell line of osteoblasts, MC3T3‐E1, and co‐culture of the osteoblasts from mouse newborn calvaria and mouse bone marrow cells. Although EGCG did not affect the viability and proliferation of MC3T3‐E1 cells, EGCG inhibited the osteoblast differentiation. Furthermore, EGCG did not affect the mineralization of differentiated MC3T3‐E1 cells, and reduced osteoclast formation in co‐culture. These results suggest that EGCG can effectively suppress bone resorption, and can be used as an effective medicine in the treatment of the symptoms of osteoporosis.     

Folin-Ciocalteu比色法测定花椒中多酚含量的研究

研究了以没食子酸为标准品,采用Folin-Ciocalteu比色法,测定花椒多酚含量的适宜条件。结果表明:当Folin-Ciocalteu试剂与10%Na2CO3溶液之比为1∶2(V∶V),Folin-Ciocalteu试剂体积为4 mL,反应温度为25℃,反应时间为120 min时,多酚含量在1~6μg/mL范围内与吸光值呈良好的线性关系。测定了4种花椒中的多酚含量,四川青花椒多酚含量最高。     

彩绒革盖菌多酚氧化酶活性研究

本文测定了彩绒革盖菌在ＰＤＹ液体培养基中的多酚氧化酶活性。在３０℃,１１０ｒ／ｍｉｎ,恒温振荡培养条件下,多酚氧化酶第１４天达产酶高峰,最高酶活５４９．０ｕ;酶作用的最适酸碱度为ｐＨ５．０;最适作用温度３０℃;Ｍｎ２＋、Ｂａ２＋、Ｍｇ２＋等离子对多酚氧化酶有激活作用,Ａｇ＋、Ｆｅ３＋等离子对酶活则有明显的抑制作用     

苹果多酚氧化酶的纯化与表征

运用丙酮浸漬干燥、磷酸盐缓冲液提取、低温离心、硫酸铵沉淀、DEAE-Sephadex(A-50)、Sephadex(G-75) 和DEAE-celluse(DE-52)层析等方法从苹果中分离获得一种新的含铜酶蛋白,该酶被命名为多酚氧化酶Ⅱ(polyphenol oxidase Ⅱ, PPOⅡ),纯化倍数是215,纯化收率是23%.PAGE、SDS-PAGE和MALDI-TOF 等技术用于测定所获的酶的纯度和分子量.在PAGE和SDS-PAGE 均显示一条带,表明PPOⅡ只由一个亚基组成,且已达到单一组分(MALDI-TOF的结果更证实了这一点).SDS-PAGE 和 MALDI-TOF 的结果都表明PPO的分子量为 38204 Da.pH值对酶活性和稳定性研究的结果显示,从pH值4.0～7.0随着pH值的增加,酶活性也不断增加;从pH值 7.0～11.0, 酶活性不断降低.PPOⅡ的最适pH值为6.6最适温度为30℃.