黑曲霉利用五倍子生料固体发酵产单宁酶优化发酵条件研究

单宁酶全称是单宁酯酰水解酶(Tannase,EC3.1.1.20),它可以水解没食子酸单宁中的酯键和缩酚羧键,生成没食子酸和葡萄糖.主要来源于微生物,存在于细胞内和细胞外,目前研究得较多较为深入的是青霉和黑曲霉.本实验对五倍子生料固体发酵产单宁酶进行了研究,发酵条件的单因素优化后得到:初始水分含量80%,接种量1%,20%五倍子,温度30℃为最佳产酶条件.在此基础上,利用正交实验对显著影响产单宁酶的培养基及培养条件因素进行优化,得到固体发酵黑曲霉诱变菌株最佳产单宁酶条件为:湿度80%、接种量1%、发酵温度33℃、五倍子含量15%.其中温度对黑曲霉产单宁酶酶活性的影响是显著的,并测得在此最佳发酵条件下产单宁酶活性达57.32U/gds.     

单宁酶降解五倍子制备没食子酸的工艺优化

以五倍子为原料,单宁酶酶制剂为酶源,研究酶解温度、酶浓度、酶促反应时间以及酶促反应体系pH对单宁酶降解五倍子制备没食子酸得率的影响,再通过响应曲面法优化单宁酶降解五倍子制备没食子酸的工艺参数。结果表明,对没食子酸得率影响较大的因素依次为酶浓度、酶促反应时间、酶促反应体系pH以及酶解温度,最优工艺技术参数为酶浓度16.26 U/g,酶促反应时间5.88h,酶促反应体系pH 5.78,酶解温度40.70℃。在该参数条件下,没食子酸的得率为59.32%。     

抗氧化剂没食子酸的酶法转化生产研究

没食子酸及其衍生物是一种重要的抗氧化剂,可以用作食品添加剂,也可作为化工原料,用于医药工业、饲料工业、化妆品等工业.没食子酸的传统生产方法是酸水解法和碱水解法,而近年来,微生物发酵法和酶转化法研究正在兴起.黑曲霉B0201直接生料固体发酵五倍子原料,未能检测到没食子酸产物.如果先采用生料固体发酵生产单宁酶,然后再用酶法制备没食子酸却是可行的.研究结果表明,分步制备没食子酸反应4h,0.5L反应液中积累了3.83g的没食子酸.没食子酸的制备过程为:生料发酵生产单宁酶--单宁酶的提取--酶法制备没食子酸,初步构建了一种微生物酶分步制备没食子酸的工艺.     

一株产单宁酶酵母菌株的筛选

单宁酶全称单宁酯酰水解酶(Tannase,EC 3.11.20),能水解水解性单宁酸和没食子酸酯类中的酯键和缩酚羧键,生成没食子酸和葡萄糖。本实验从青柿、未完全成熟的香蕉、茶叶、茶园土壤等富含单宁酸的环境中取样,采用富集培养技术分离筛选出69株产单宁酶的菌株,并对其菌株的外在特性和产酶活力进行了研究和测定,最终获得一株产单宁酶活力较高的酵母菌株FC6-1。在未进行发酵条件优化前,该菌株所产单宁酶酶活达到2.86U/mL。采用薄层色谱法和SBA-40D生物传感分析仪等方法对该酶酶解产物分析,结果表明该菌株产生的单宁酶能够有效的水解单宁酸生成没食子酸和葡萄糖。     

响应面法优化黑曲霉产单宁酶的固体发酵条件

通过固体发酵培养基单因素研究,确定了三个影响单宁酶产率的关键因素,对氮源用量、五倍子用量、培养温度采用响应面法的中心旋转实验设计原理,进行三因素三水平的响应面分析,以获得最佳产单宁酶的培养基及培养条件组成。结果表明,固体发酵黑曲霉最佳产酶条件为：五倍子含量为9％;氮源添加量为2．3％;温度为32℃。在此条件下进行发酵产酶重复实验,酶活力为219．4U／mL。     

五倍子单宁酸同步提取-酶解工艺的研究

单宁酸是一种较强的蛋白质沉淀剂。在酶法制备没食子酸的过程中,高浓度单宁酸会严重抑制单宁酶活性,因此很难直接利用单宁酸进行没食子酸的工业生产。为了解除单宁酸对单宁酶的抑制作用,研究建立了五倍子单宁酸同步提取-酶解工艺,反应90 min,同步提取-酶解工艺没食子酸浓度达到13.9 g/L,没食子酸生产效率明显高于同步提取-酸解工艺和五倍子单宁酸先提取后酶解工艺。对五倍子单宁酸同步提取-酶解工艺进行优化,结果表明在料液比1:50、加酶量32 U/mL、pH6～7、温度50 ℃和提取时间75 min时,没食子酸产量和得率最大,分别达到15.4 g/L和76.8%。在最优条件下进行多次投料试验,没食子酸最终产量可提高至34.0 g/L。     

利用微生物酶两步法生产没食子酸的初步研究

没食子酸及其衍生物是食品、医药等工业的一类重要原料。以黑曲霉B0201为菌种,以五倍子为原料,直接生料固体发酵法不能生成没食子酸,但是,先用生料固体发酵生产单宁酶,再用酶法制备没食子酸是可行的。研究表明,分两步制备没食子酸反应4h时,100mL单宁酶酶液中积累了0.77g没食子酸,建立了一种黑曲霉单宁酶两步法生产没食子酸的制备工艺。     

固态发酵黑曲霉产单宁酶发酵条件研究

经紫外线诱变黑曲霉菌株筛选出1株高产单宁酶的黑曲霉B0201,利用五倍子为诱导物,固态发酵该菌株得到的单宁酶活力有较大提高。单因素优化试验表明,五倍子用量为8%,初始水分含量为50%,初始pH6.0,温度30℃为优化产酶条件。在优化的条件下,培养96h后产单宁酶酶活力达到了58.2 U/g(干基)。因此,黑曲霉固态发酵产单宁酶具有很大的研究意义及广阔的应用前景。     

微生物源单宁酶的研究进展

单宁酶可将单宁水解成没食子酸和葡萄糖，是一种重要的工业用酶。植物、动物和微生物中都含单宁酶，其中微生物是单宁酶的主要来源。基于单宁酶的国内外研究成果，该文归纳总结了不同微生物来源的单宁酶、酶学性质、作用机制，阐述了单宁酶在食品、饲料、制革、精细化工等实际生产中的应用，并对单宁酶今后的研究方向和应用前景进行了展望。     

单宁酶发酵生产的研究进展

单宁酶可水解没食子酸单宁中的酯键和缩酚羧键,生成没食子酸和葡萄糖。单宁酶广泛应用于制革、酿酒、医药、饮料等领域。文中综述了国内外有关单宁酶发酵生产的研究进展,包括产单宁酶菌种的选育、发酵生产方法、底物选择和培养基中添加剂对产酶的影响。     

Synthesis of propyl gallate by transesterification of tannic acid in aqueous media catalysed by immobilised derivatives of tannase from Lactobacillus plantarum

Immobilised derivatives of tannase from Lactobacillus plantarum were able to catalyse the transesterification of tannic acid by using moderate concentrations of 1-propanol in aqueous media. Transesterification of tannic acid was very similar to transesterification of methyl gallate. The synthetic yield depended on the pH and concentration of 1-propanol, although it did not vary much when using 30% or 50% 1-propanol. Synthetic yields of 45% were obtained with 30% of 1-propanol at pH 5.0. The product was chromatographically pure, and the reaction by-product was 55% pure gallic acid. On the other hand, immobilised tannase was fairly stable under optimal reaction conditions.     

单宁酶制备研究进展

单宁酶主要由曲霉属等微生物发酵制备。它能够水解单宁中的酯键和缩酚羧键,生成重要的工业原料没食子酸等化合物。单宁酶已广泛应用于食品饮料、化妆品工业和饲料工业等领域。文章着重综述国内外在单宁酶菌种选育、培养基发酵体系、发酵生产、提取与分离纯化、固定化技术等制备方法方面研究的新进展,分析单宁酶制备研究的发展趋势。     

酸解法制备五倍子抗腐败希瓦氏菌活性物质的工艺

研究酸解法制备五倍子抗腐败希瓦氏菌活性物质的工艺。采用单因素试验、Box-Behnken试验设计和响应面法对酸解法制备五倍子抗腐败希瓦氏菌活性物质的工艺进行优化,优化的酸解工艺为:盐酸浓度4 mol/L、液料比45 mL/g、温度100℃时提取5 h。在最佳酸解工艺下进行验证试验,五倍子提取液的实际抗菌活性为(23.68±0.68)mm,与预测值(22.63 mm)没有显著差异。研究结果表明响应面法应用于优化酸解法制备五倍子抗腐败希瓦氏菌活性物质的工艺是可行的,为进一步鉴定和开发五倍子抗菌活性物质提供了依据。     

玉米芯固定化单宁酶的工艺条件优化

探讨改性玉米芯固定化单宁酶的工艺条件。采用Box-Behnkens试验设计和响应面分析方法,对改性玉米芯固定化单宁酶的条件进行优化,得出其最佳工艺条件为：单宁酶酶液与玉米芯载体比26:1(V/m)、pH 6.8、时间8.0h、温度36℃,在此条件下固定化酶活力达到(16085±5)U/g,酶活力回收率为44.68%。对最佳条件下制备的固定化单宁酶的酶学特性进行研究,发现该固定化酶的最适反应pH值为4.5～5,最适反应温度为60℃。     

Changes in polyphenol and polysaccharide content of grape seed extract and grape pomace after enzymatic treatment

Grape seed extract and grape pomace are rich sources of polyphenols. The aim of this study was to evaluate the release of polyphenols, the solubilisation of carbohydrate, and the antioxidant capacity of these grape by-products after enzymatic reaction with carbohydrases (cellulolytic and pectinolytic activities) and tannase for 24 h. The use of tannase in these by-products, and pectinase in grape pomace changed the galloylated form of catechin to its free form, releasing gallic acid and increasing the antioxidant activity. In grape pomace, cellulase treatment was not efficient for phenolic release and antioxidant activity improvement. The addition of carbohydrases to grape pomace, either alone or in combination, degraded the cell wall polysaccharides, increasing the content of monosaccharides. These results provide relevant data about the potential of pectinase, tannase and combinations of enzymes on the release of polyphenols and monosaccharides from grape by-products, improving the antioxidant capacity and the nutritional value.     

Changes in the content of bioactive polyphenolic compounds of lentils by the action of exogenous enzymes. Effect on their antioxidant activity

The study of the effect of the enzymes tannase, α-galactosidase, phytase and viscozyme on the phenolic composition of lentil flours, in a semi pilot scale stirred fermentor, shows that important modifications occur. Among them, hydroxycinnamic compounds and proanthocyanidins are significantly decreased after the enzymatic treatments. However, quercetin 3-O rutinoside and luteolin increase and reach the highest concentration with tannase. The formation of trans-resveratrol was observed by the action of tannase and phytase, and gallic acid by the action of phytase, α-galactosidase and tannase. The antioxidant capacity of the methanolic extracts was determined by their free radical scavenging activity, using the DPPH test, to study the differences in the behaviour of polyphenolics compounds as antioxidants after the different enzymatic treatments. The treatments with viscozyme, α-galactosidase or tannase produce an increase in the antioxidant activity when compared to raw lentils. The results of the analysis of principal components to examine the relationship among antioxidant activity (EC₅₀) (DPPH test) and the concentrations of polyphenolics in all lentils samples, show that the quercetin 3-O rutinoside appears to be the compound with the greatest influence on the EC₅₀ values.     

Hydrolysis of epigallocatechin gallate using a tannase from Paecilomyces variotii

Epigallocatechin (EGC) and gallic acid (GA) were prepared by the degalloylation of an epigallocatechin gallate (EGCG) extract from green tea. EGCG was entirely hydrolyzed using a tannase (from Paecilomyces variotii) at pH 6.0, incubating at 40 °C for 30 min. The antiradical properties and the reducing power of these samples were assessed using the DPPH and FRAP assays, respectively. This work established a relationship between the antioxidant effects of epigallocatechin gallate and the enzymatic reaction products (epigalocatechin and gallic acid). The enzymatic reaction products showed higher scavenging activity and antioxidant capacity when compared to epigallocatechin gallate.