光谱法研究甲醇和盐酸对短梗霉黑色素的分离效果

应用紫外和红外光谱对甲醇和盐酸分离的短梗霉黑色素进行了分析。分析结果表明:甲醇和盐酸法分离的黑色素在紫外图谱215 nm处都有最大吸收峰,而甲醇法分离的黑色素其紫外图谱在260、280 nm处无吸收峰表明此法分离的黑色素不含核酸及蛋白质;红外图谱中,在3340 cm-1、1637 cm-1处有很强的吸收峰表现为黑色素的典型特征;同时在对照中发现盐酸多次处理后的黑色素显示较少的结构信息,说明盐酸沉降法有可能破坏黑色素的结构。由此选用甲醇作为沉淀剂有利于黑色素的纯化及确保其结构信息的完整,为进一步分析短梗霉黑色素的结构表征奠定了基础。     

香灰菌黑色素对真菌生长及多酚氧化酶活性的影响

以PDB培养的香灰菌丝体为材料,提取、纯化香灰菌黑色素,并测定香灰菌黑色素对六种真菌生长及对其胞外多酚氧化酶活性的影响.结果表明:香灰菌黑色素对六种真菌的生长没有抑制作用,而对其多酚氧化酶活力有明显的诱导作用.在不同的香灰菌黑色素浓度范围内,六种真菌胞外多酚氧化酶最大酶活的出现时间不同.     

高原链带藻抗氧化蛋白分离纯化与结构鉴定

【背景】微藻Desmodesmus sp. QL96从我国西藏地区分离得到,经形态鉴定隶属于链带藻属。前期研究发现,这种链带藻在4℃和25℃下均可生长,在25℃生长时,干细胞中蛋白质含量可高达71.68%(质量分数),而且蛋白粗提物具有一定的抗氧化活力。【目的】分离纯化Desmodesmus sp. QL96细胞中具有抗氧化活力的蛋白质,并对其结构进行鉴定。【方法】应用柱层析的方法分离纯化Desmodesmus sp. QL96细胞中具有抗氧化活力的蛋白质,通过化学发光法和细胞学实验对该蛋白的抗氧化活性进行检测,并通过质谱技术对其一级结构进行检测。【结果】Desmodesmus sp. QL96细胞中抗氧化蛋白的含量占微藻细胞干重的11.40%(质量分数);纯化的Desmodesmus sp. QL96抗氧化蛋白在一定浓度范围内对OH-、DPPH、ABTS自由基和H2O2具有较好的清除率(超过60%),细胞学实验显示其对H2O2诱导的HepG2细胞氧化损伤具有抑制作用,验证了其抗氧化功能;通过质谱技术检测了Desmodesmus sp. QL96抗氧化蛋白的氨基酸序列,并进行了生物信息学分析,结果显示,这种蛋白质的理论分子量为44.8 kD、pI 5.79,与NCBI中目前已知的其他物种蛋白质的相似性不超过59%。【结论】Desmodesmus sp. QL96可能生产一种具有抗氧化活性的新蛋白质,后续将对其转录本进行分析,验证其遗传信息的同源性,并分析其规模化生产和应用前景。     

基于Py-GC/MS分析黑木耳新品种‘农黑2号’木耳黑色素的热裂解产物

利用热裂解-气相色谱质谱联用(pyrolysis gas chromatography mass spectrometry, Py-GC/MS)分析技术测定黑木耳新品种‘农黑2号’黑色素裂解产物组分的种类及含量,为解析黑色素的结构奠定理论基础。本研究通过碱溶酸沉法提取黑色素,并用紫外可见吸收光谱、傅立叶红外光谱、有机元素分析鉴定黑色素,采用Py-GC/MS技术分析其热裂解产物。结果表明,‘农黑2号’黑色素的最大吸收峰值在波长220 nm处,傅立叶红外光谱符合典型的黑色素特征;经有机元素分析,C、H、N、O元素的含量分别为43.47%、5.24%、11.71%和39.58%,不含硫元素,为真黑色素;检出的热裂解产物主要有苯类、苯酚类、吲哚类、苯基腈类、烃类、酸类等物质,其中苯、苯酚、吲哚等化合物为真黑色素的裂解产物。综上所述,‘农黑2号’黑色素为真黑色素,其基本骨架为苯环和吲哚,本研究为精准解析黑木耳黑色素结构提供科学依据。     

抑制黑色素合成的乳酸菌胞外多糖的筛选和性质研究

【目的】筛选可抑制黑色素合成的乳酸菌胞外多糖。【方法】通过观察凝乳拉丝外观筛选产胞外多糖的乳酸菌菌株,测量胞外多糖对B16黑色素瘤细胞黑色素合成和细胞活力的影响。对胞外多糖进行纯化,并通过PMP衍生-HPLC、红外光谱、抑制酪氨酸酶活性、抗氧化能力对其单糖组成和结构、作用机制进行研究。【结果】筛选到一株乳酸菌Lactobacillus rhamnosus HLAB122,发酵产生的胞外多糖在5 g/L浓度下可使B16细胞黑色素产量下降至空白对照的32.7%,且在96 h内对细胞活力无影响。纯化后的多糖由鼠李糖、葡萄糖、半乳糖构成,各单糖摩尔比为1?5.44?5.37。该胞外多糖不抑制酪氨酸酶活力且抗氧化性微弱。【结论】L.rhamnosus HLAB122产生的胞外多糖在个人护理产品中有潜在应用价值。     

哺乳动物毛色形成机制与影响因素

哺乳动物拥有丰富多彩的毛色,对于多种多样的动物毛色,影响其形成的可能原因较多,遗传物质与环境的相互作用不可忽视。本文针对调控黑色素细胞形成及褐黑色素、真黑色素合成的相关基因,分析了哺乳动物毛色形成的可能机制,并对研究不同毛色动物的同种动物遗传差异的方法进行了讨论。     

短梗霉细胞内黑色素的分离提取

采用单因素试验和正交试验对短梗霉细胞内的黑色素提取条件进行了优化,结果表明:在pH为2～3、温度为70℃、NaOH浓度为1.5mol/L、发酵液与浸提液之比1:1(V)的条件下黑色素的提取量可达2.2g/L。对提取的黑色素进行了纯化,研究了其紫外和红外光谱学性质,紫外光谱图显示随波长的减少其吸收值增大,在215nm处有特征吸收峰;红外光谱图在3和6μm处有吸收峰,证实短梗霉黑色提取物是一种以芳香环为结构主体的异聚体黑色素。     

乌鳖黑色素理化性质及其抗氧化活性研究

本试验以乌鳖肌肉为原料,采用"脱脂-酶解-酸解"三步法制备乌鳖黑色素,研究其结构特性、理化性质和抗氧化活性.采用紫外分光光度计、傅里叶红外光谱仪和元素分析仪对乌鳖肌肉黑色素结构特性进行分析,并进行了颜色检测、溶解性试验、稳定性试验、总抗氧化能力试验、羟基自由基清除能力试验、DPPH自由基清除能力试验、超氧阴离子自由基清...     

萝卜色素的分离、纯化及其性质研究

“心里美”萝卜酒精提取物,经薄层层析、高压液相层析纯化后可得到天然色素为粉红色针状结晶。该色素性质稳定、耐温、耐光,颜色随pH值的不同而变化。经红外光谱分析和质谱鉴定,初步推导其结构可能为报春花色素。     

霍乱弧菌分型噬菌体VP3蛋白的双向电泳分析

目的:噬菌体-生物分型方案具有区分霍乱弧菌潜在致病力的作用,VP3是5个霍乱弧菌分型噬菌体之一。对VP3成熟颗粒的蛋白组成进行测定和分析,以补充基因组注释信息。方法:在全基因组测序及生物信息学分析的基础上,利用双向电泳技术及质谱鉴定,对纯化的成熟VP3噬菌体的结构蛋白进行分离及鉴定。结果:双向电泳分离得到近20个蛋白点,质谱鉴定出了其中的10个,对应于4个VP3蛋白和4个霍乱弧菌蛋白。结论:VP3结构蛋白的组成和T7具有很高的相似性。与噬菌体颗粒一起被纯化分离的宿主蛋白可能在VP3的转染过程中起作用。     

香灰菌黑色素的分离及抗氧化活性研究

从香灰菌中分离得到黑色素,其产量为33.01%。经紫外/可见扫描和红外光谱分析,显示香灰菌黑色素同合成黑色素具有基本一致的化学结构。通过检测抗氧化剂抑制5-硫代-(2-硝基苯甲酸)(TNB)被氧化程度,比较香灰菌黑色素同合成黑色素及Vc的抗氧化活性,结果表明,香灰菌黑色素抗氧化能力强于Vc,略低于合成黑色素。     

Down-regulation of melanin synthesis by a biphenyl derivative and its mechanism

Down-regulation of melanin synthesis is required for recovery of pigmentary disorders and it is known that direct inhibitors of tyrosinase, the key enzyme in melanin synthesis, such as hydroquinone with a phenol structure, suppress melanin synthesis. We screened several phenolic derivatives using B16 melanoma cells and found that a biphenyl derivative, 2,2'-dihydroxy-5,5'-dipropyl-biphenyl (DDB), down-regulated melanin synthesis effectively. Although DDB has a phenol structure, it did not inhibit tyrosinase in vitro, thus we examined its mechanism in detail. Western blotting revealed that the amount of tyrosinase was decreased by DDB, and pulse-chase labeling and immunoprecipitation analysis showed a decrease of mature tyrosinase and acceleration of tyrosinase degradation in its presence. These results suggest that DDB down-regulates melanin synthesis by inhibiting the maturation of tyrosinase, leading to acceleration of tyrosinase degradation.     

Down‐Regulation of Melanin Synthesis by a Biphenyl Derivative and Its Mechanism

Down‐regulation of melanin synthesis is required for recovery of pigmentary disorders and it is known that direct inhibitors of tyrosinase, the key enzyme in melanin synthesis, such as hydroquinone with a phenol structure, suppress melanin synthesis. We screened several phenolic derivatives using B16 melanoma cells and found that a biphenyl derivative, 2,2′‐dihydroxy‐5,5′‐dipropyl‐biphenyl (DDB), down‐regulated melanin synthesis effectively. Although DDB has a phenol structure, it did not inhibit tyrosinase in vitro, thus we examined its mechanism in detail. Western blotting revealed that the amount of tyrosinase was decreased by DDB, and pulse‐chase labeling and immunoprecipitation analysis showed a decrease of mature tyrosinase and acceleration of tyrosinase degradation in its presence. These results suggest that DDB down‐regulates melanin synthesis by inhibiting the maturation of tyrosinase, leading to acceleration of tyrosinase degradation.     

Molecular docking studies of a phlorotannin, dieckol isolated from Ecklonia cava with tyrosinase inhibitory activity

In this study, the phlorotannin dieckol, which was isolated from the brown alga Ecklonia cava, was examined for its inhibitory effects on melanin synthesis. Tyrosinase inhibitors are important agents for cosmetic products. We therefore examined the inhibitory effects of dieckol on mushroom tyrosinase and melanin synthesis, and analyzed its binding modes using the crystal structure of Bacillus megaterium tyrosinase (PDB ID: 3NM8). Dieckol inhibited mushroom tyrosinase with an IC(50) of 20μM and was more effective as a cellular tyrosinase having melanin reducing activities than the commercial inhibitor, arbutin, in B16F10 melanoma cells, and without apparent cytotoxicity. It was found that dieckol behaved as a non-competitive inhibitor with l-tyrosine substrates. For further insight, we predicted the 3D structure of tyrosinase and used a docking algorithm to simulate binding between tyrosinase and dieckol. These molecular modeling studies were successful (calculated binding energy value: -126.12kcal/mol), and indicated that dieckol interacts with His208, Met215, and Gly46. These results suggest that dieckol has great potential to be further developed as a pharmaceutical or cosmetic agent for use in dermatological disorders associated with melanin.     

Role of estradiol and 2-hydroxyestradiol in melanin formation in vitro

Melanin formation from 3,4-dihydroxyphenylalanine (dopa) was studied in the presence of estradiol and 2-hydroxyestradiol by use of a tyrosinase isolated from B16-F10 melanoma cells grown in C57 black female mice. Both steroids were found incorporated into melanin, but the 2-hydroxy compound was incorporated to a higher extent. The melanin was also able to bind substantial amounts of the two steroids, and the more highly oxidized compound showed higher binding. Melanin isolated from incubates of dopa with mushroom tyrosinase has the ability to bind the steroids and to incorporate small amounts into its structure. It is suggested that melanin in mammalian tissues may function as a depository for estrogens, particularly for those which are more highly oxidized.     

大鲵皮肤色素提取工艺及抗氧化研究

为优化大鲵皮肤黑色素的提取工艺条件,探讨大鲵皮肤黑色素组成成分及体外抗氧化活性,采用酶法和碱溶酸沉法提取大鲵皮肤黑色素,以氢氧化钠浓度、液料比、提取温度为影响色素提取率因素,优化黑色素提取工艺条件,用紫外-可见光谱仪、红外光谱仪和超高效液相质谱仪测定黑色素的光谱特性,测定其抗氧化性。结果表明:大鲵皮肤黑色素最佳提取工艺条件为氢氧化钠浓度1.5 mol/L、液料比1∶15、提取温度45℃,黑色素提取率达0.65%。大鲵皮肤黑色素的紫外最大吸收波长为214 nm,由真黑色素和脱黑色素两种色素组成,其对超氧阴离子自由基的清除率为30.51%,对羟基自由基的清除率为54.17%。大鲵皮肤黑色素具有一定的体外抗氧化能力。     

银耳菌糠黑色素的理化性质及抗氧化和抑菌活性

银耳菌糠中存在一种银耳的伴生菌——炭团菌(俗称香灰菌),其菌丝生长到一定阶段后会产生大量的黑色素,具有广泛的应用价值。本研究从银耳菌糠中提取黑色素,研究其理化性质、抗氧化活性及抑菌作用。通过紫外-可见光谱、傅里叶红外光谱对提取的黑色素进行鉴定,表明银耳菌糠黑色素具有黑色素的典型特征。通过对银耳菌糠黑色素理化性质的研究,表明银耳菌糠黑色素是一种趋于黑色并略带红色和黄色的粉末;该黑色素易溶于碱性溶液;具有较好的热稳定性和光稳定性,其稳定性受氧化剂和还原剂的影响较小,受Ca²⁺、Cu²⁺的影响较明显。通过总抗氧化能力(FRAP法)、自由基清除能力检测银耳菌糠黑色素的抗氧化活性,研究表明黑色素具有较高的抗氧化活性,羟自由基、ABTS自由基清除的EC₅₀值分别为0.429 mg/mL和0.016 mg/mL。本研究还检测了黑色素对革兰氏阳性、革兰氏阴性细菌的抑菌能力,结果表明该黑色素在浓度为3.2 mg/mL时对供试菌株的抑菌率超过90%,且对革兰氏阳性菌会较敏感。本研究为银耳菌糠的有效利用及其黑色素产品的开发提供了理论基础,具有较高的经济价值。     

Roles of reactive oxygen species in UVA‐induced oxidation of 5,6‐dihydroxyindole‐2‐carboxylic acid‐melanin as studied by differential spectrophotometric method

Eumelanin photoprotects pigmented tissues from ultraviolet (UV) damage. However, UVA‐induced tanning seems to result from the photooxidation of preexisting melanin and does not contribute to photoprotection. We investigated the mechanism of UVA‐induced degradation of 5,6‐dihydroxyindole‐2‐carboxylic acid (DHICA)‐melanin taking advantage of its solubility in a neutral buffer and using a differential spectrophotometric method to detect subtle changes in its structure. Our methodology is suitable for examining the effects of various agents that interact with reactive oxygen species (ROS) to determine how ROS is involved in the UVA‐induced oxidative modifications. The results show that UVA radiation induces the oxidation of DHICA to indole‐5,6‐quinone‐2‐carboxylic acid in eumelanin, which is then cleaved to form a photodegraded, pyrrolic moiety and finally to form free pyrrole‐2,3,5‐tricarboxylic acid. The possible involvement of superoxide radical and singlet oxygen in the oxidation was suggested. The generation and quenching of singlet oxygen by DHICA‐melanin was confirmed by direct measurements of singlet oxygen phosphorescence.     

Solution structure of copper ion-induced molecular aggregates of tyrosine melanin.

Melanin, the ubiquitous biological pigment, provides photoprotection by efficient filtration of light and also by its antioxidant behavior. In solutions of synthetic melanin, both optical and antioxidant behavior are affected by the aggregation states of melanin. We have utilized small-angle x-ray and neutron scattering to determine the molecular dimensions of synthetic tyrosine melanin in its unaggregated state in D(2)O and H(2)O to study the structure of melanin aggregates formed in the presence of copper ions at various copper-to-melanin molar ratios. In the absence of copper ions, or at low copper ion concentrations, tyrosine melanin is present in solution as a sheet-like particle with a mean thickness of 12.5 A and a lateral extent of approximately 54 A. At a copper-to-melanin molar ratio of 0.6, melanin aggregates to form long, rod-like structures with a radius of 32 A. At a higher copper ion concentration, with a copper-to-melanin ratio of 1.0, these rod-like structures further aggregate, forming sheet-like structures with a mean thickness of 51 A. A change in the charge of the ionizable groups induced by the addition of copper ions is proposed to account for part of the aggregation. The data also support a model for the copper-induced aggregation of melanin driven by pi stacking assisted by peripheral Cu(2+) complexation. The relationship between our results and a previous hypothesis for reduced cellular damage from bound-to-melanin redox metal ions is also discussed.