一种小麦多酚氧化酶活性测定方法的改进研究

参照美国谷物化学家协会标准方法(AACC 22-85),以L-DOPA为底物,测定了22份小麦样品其籽粒的PPO活性,发现测定结果的重复性较差.在籽粒PPO活性测定原理的基础之上,优化了测定参数,改进、完善了一种评价小麦全粉PPO活性的新方法,其操作简便、快速,实验结果的重复性好、稳定性高,与籽粒PPO活性的测定结果极显著相关.     

Inhibitory effect of rice bran extract on polyphenol oxidase of potato and banana

Rice bran was extracted with water and its effects on potato and banana polyphenol oxidase (PPO) were investigated. Rice bran extract (RBE), conc. 0.3 g mL^?1, exhibited PPO inhibition in potato and banana PPO with % inhibition of 69.31% and 47.63%, respectively (P ≤ 0.05). RBE showed a concentration‐dependent inhibition on potato and banana PPO. RBE (conc. 0.3 g mL^?1) inhibited potato PPO higher than ascorbic acid, citric acid, NaCl and EDTA (final conc. 20 mg L^?1); and it also inhibited banana PPO higher than citric acid, NaCl and EDTA (final conc. 20 mg L^?1), respectively. The combination of RBE with citric acid or ascorbic acid appeared to be additive inhibitory effect on banana and potato PPO. Kinetic study of the inhibition on potato and banana PPO by RBE showed that RBE was a mixed‐type inhibitor; however, RBE appeared to be able to act directly on enzyme structure rather than substrate structure.     

在线粉流中小麦多酚氧化酶活性分析

通过对在线粉流中小麦多酚氧化酶(PPO)活性的测定分析,得到PPO活性与小麦粉主要理化指标的关系:PPO活性与小麦粉白度成极显著负相关性,与灰分含量成极显著正相关性;前路粉流PPO活性比后路粉流低;物料含皮较多的在线粉流PPO活性高;物料来源于小麦胚乳外层的粉流PPO活性高。     

小麦粉多酚氧化酶活性测定方法的改进

选取3个小麦粉作为试验材料,对国家标准征求意见稿《粮油检验小麦粉多酚氧化酶测定》的方法进行检验,发现该法因缺少PPO粗酶提取过程,参加反应的酶量较少,不能反映小麦粉真实的PPO活性;其次PPO与底物的反应温度不是最适宜的,导致酶反应不充分;第三酶反应后再离心,使得一部分酶反应产物被沉淀截留,导致检测数据小且不准确。文章在该基础上对其进行改进,考察浸提时间、最适反应温度、底物邻苯二酚添加时间对PPO活性检测的影响,优化该检验方法。结果表明:优化后的检验方法测得的结果较为理想,采用浸提4h、离心后添加邻苯二酚、反应温度70℃,测得的PPO活性数值大大提高,重复性好,稳定可靠。并采用改进后的方法检测了50种小麦粉,得出不同等级的小麦粉PPO的范围。     

响应面法优化黑粒小麦麸皮多酚提取工艺

以黑粒小麦麸皮为原料,选用乙醇为溶剂,考察乙醇浓度、提取温度、料液比、超声波功率、提取时间、提取次数对多酚得率的影响,在单因素实验基础上,通过响应面法优化黑粒小麦麸皮多酚的最佳提取条件。实验结果表明:乙醇浓度58%,料液比1∶22(g/m L),超声功率60 W,50℃条件下超声波作用37 min,提取2次。在此条件下,麸皮中多酚得率为3.545 mg/g,优化后的提取工艺对黑粒小麦麸皮多酚的提取有一定的指导意义。      

影响春小麦面粉中多酚氧化酶活性因素的研究

以邻苯二酚溶液为底物,测定春小麦面粉中的多酚氧化酶,并从不同pH值、不同底物浓度、酶液浓度等方面研究春小麦面粉中多酚氧化酶活性的影响因素,结果表明,以上测定指标对酶活性的影响非常明显。     

Properties of polyphenol oxidase in mango (Mangifera indica) kernel

Polyphenol oxidase (PPO) activity of filtered extract of ground mango kernel suspension (400 g litre⁻¹) was studied spectrophotometrically at 420 nm using catechol as substrate. The enzyme was most active at pH 6·0 and 25°C. Activity was reduced by 50% at pH values of 5·0 and 7·1, and also at temperatures of 14°C and 30°C. The calculated activation energy and the Michaelis constant (K_m) were 21·4 kcal mol⁻¹ °C⁻¹ and 24·6 mM , respectively. The V_max value was 2·14 units g⁻¹ mango kernel. The time to heat inactivate PPO decreased rapidly to < 10 min with increasing temperature of ⩾ 70°C at 50% activity. © 1998 SCI.     

复合风味蛋白酶酶解麦麸蛋白制备麦麸抗氧化肽的研究

以麦麸蛋白为原料,复合风味蛋白酶为水解酶,探讨麦麸蛋白水解度(DH)与麦麸肽抗氧化能力的关系,并以超氧阴离子自由基(O2-·)清除率为指标,采用正交试验设计优化制备麦麸抗氧化肽的工艺条件.试验结果表明,制备麦麸抗氧化肽的最佳酶解条件为pH 5.0,加酶量为6000U/g麦麸蛋白,底物浓度5%,温度55℃,酶解7.5h,在该条件下制得的麦麸抗氧化肽清除超氧阴离子自由基的清除率可达53.70%.     

多酚氧化酶的研究进展

随着研究的不断深入,多酚氧化酶的应用越来越广泛,对此综述了多酚氧化酶的测定方法、分离提纯技术和催化机制,并对其应用及研究的发展趋势进行了展望.     

Properties of polyphenol oxidase from Anamur banana (Musa cavendishii)

Polyphenol oxidase (PPO) was extracted from Anamur banana, grown in Turkey, and its characteristics were studied. The optimum temperature for banana PPO activity was found to be 30 °C. The pH-activity optimum was 7.0. From the thermal inactivation studies, in the range 60–75 °C, the half-life values of the enzyme ranged from 7.3 to 85.6 min. The activation energy (E_a) and Z values were calculated to be 155 kJ mol⁻¹ and 14.2 °C, respectively. K_m and V_max values were 8.5 mM and 0.754 OD₄₁₀ min⁻¹, respectively. Of the inhibitors tested, ascorbic acid and sodium metabisulphite were the most effective.     

Fractionation of wheat bran carbohydrates

A procedure for the fractionation of wheat bran carbohydrates with an emphasis on non-starchy polysaccharides is described. After a complete extraction using a chloroform-methanol (2:1) solvent, bran was destarched using amyloglucosidase and extracted using ammonium oxalate. Then delignification by sodium chlorite followed by hot water extractions was applied. Small quantities of water-soluble hemicellulosic polymers containing glucose, xylose, arabinose with lesser amounts of galactose and mannose were removed by these three steps. The remaining holocellulose (of a low protein content) was extracted using 1M sodium hydroxide, and hemicelluloses A1 and B1 were separated from the extract. Hemicellulose A1 (5.4% of native bran) was determined to be a xylan containing smaller proportions of arabinose and glucose, whereas hemicellulose B1 (12.6% of native bran) was a complex mixture of several types of polysaccharides, the major one being a highly branched arabinoxylan. Hemicelluloses A4 and B4, obtained by 4M sodium hydroxide extraction of the residue after 1M sodium hydroxide treatment, had lesser amounts of arabinose compared with the respective fractions from the preceding step. For hexoses, higher proportions of mannose and glucose were found. α-Cellulose residue retains 19.8% of pentosans.     

杏子多酚氧化酶动力学特性研究

杏子多酚氧化酶是造成杏子果肉褐变的主要因素,研究其动力学特性对杏子深加工中防止果肉褐变有着重要理论意义。采用分光光度计法研究pH、温度、抑制剂和高压二氧化碳处理对杏子多酚氧化酶性质的影响,结果表明:杏子(品种为色买提)的最适底物为邻苯二酚,最适pH5.0,最适温度40℃。多酚氧化酶催化的酶促褐变反应动力学符合米氏方程,该酶促反应的动力学常数Km为0.014mol/L,最大速率为287U/min。控制pH3.0以下,或者采用L-抗坏血酸4mmol/L与柠檬酸8mmol/L复配,或采用高压二氧化碳40MPa处理30min都可以将相对酶活性控制在20%以下;温度90℃处理6min以上或100℃处理4min可以完全抑制酶活性。在实际应用中,可根据产品特性不同选用适宜的处理方法或组合抑制褐变。     

蓝莓中多酚氧化酶的酶学特性

为分析蓝莓中多酚氧化酶的酶学特性,本文对影响蓝莓多酚氧化酶（PPO）的酶活力的主要因素:p H、温度、金属离子、抑制剂及热稳定性进行了研究。结果表明:蓝莓中PPO最适p H为6.5,最适温度为30℃,Ca²⁺和Mn²⁺对蓝莓PPO有抑制作用,Cu²⁺、Mg²⁺、Zn²⁺、Fe³⁺对蓝莓PPO有激活作用。柠檬酸、抗坏血酸、L-半胱氨酸及亚硫酸氢钠能抑制PPO酶活,但柠檬酸抑制效果较差。热稳定性实验结果表明:蓝莓果PPO耐热性较差,高温短时可显著抑制PPO酶活性。      

蕨菜多酚氧化酶的性质

以冷冻丙酮制备蕨菜多酚氧化酶丙酮粉,用分光光度法研究pH、温度、底物浓度,抑制剂对酶活性的影响,结果表明,以邻苯二酚为底物,该酶的最适pH值为7.4,最适温度为25℃;Km为39.86 mmol/L,Vm为0.686 OD/min,90℃热处理50 s可钝化PPO的活性;抗坏血酸、亚硫酸钠为强烈抑制剂,175 mg/L的亚硫酸钠、225 mg/L的抗坏血酸能有效抑制PPO的活性.     

Biochemical studies of cocoa bean polyphenol oxidase

Polyphenol oxidase (EC 1.14.18.1) was isolated and partially purified from cocoa beans. The properties of the enzyme were studied. The Michaelis constant K_m for catechol was 1 × 10^?2 M . The pH optimum of polyphenol oxidase activity assayed with catechol as substrate occurred at pH 6.8 and was characterised by a relatively high thermal stability, 50% of its activity was lost after heating for 40, 25 and 5 min at 60, 69 and 80°C respectively. The optimum temperature for the enzyme activity with catechol as substrate was around 45°C. The enzyme was reactive towards 3-(3,4-dihydroxy phenyl)-DL -alanine, 3-hydroxytyramine hydrochloride and 4-methyl catechol but showed no activity towards tyrosine, p-cresol, and 4-hydroxy-phenol. A rapid deactivation of the enzyme was observed when catechol of concentration > 40 mM was used as substrate. The enzyme activity was inhibited by ascorbic acid, L -cysteine, sodium bisulphite and thiourea.     

Characterization of polyphenol oxidase from Napoleon grape

Polyphenol oxidase (PPO) from Napoleon grape was isolated using a two-phase partitioning approach with Triton X-114. The enzyme was purified in a latent form and could be optimally activated by the presence of 0.2% of sodium dodecyl sulphate (SDS) at pH 6.0. In the absence of SDS, the enzyme showed maximum activity at acid pH (3.0). The enzyme was kinetically characterized at pH 3.0 and pH 6.0 in the presence of 0.2% of SDS, using 4-tert-butylcatechol (TBC) as a substrate. The V_m/K_M ratio showed that Napoleon grape PPO presents greater affinity for TBC at acid pH (0.1 min⁻¹) that at pH 6.0 in the presence of SDS (0.02 min⁻¹). The enzyme was highly heat stable, 80% of activity remaining at 70 °C. Selected inhibitors were also studied, tropolone being the most active with a K_i value of 27 μM at acid pH and pH 6.0 in the presence of 0.2% SDS.     

Some properties of polyphenol oxidase from lily

A study of crude polyphenol oxidase (PPO) from lily bulbs was carried out to provide information useful for guiding food processing operations. Optimum pH for the enzyme activity in the presence of catechol, were 4.0 and 7.0 at room temperature(approximately 20 °C) and the enzyme was stable in the pH range from 5.0 to 6.5 at 4 °C for 10 h. Its optimum temperature was 40 °C and the heat inactivation of the enzyme followed first‐order kinetics. Lily PPO possessed a diphenolase activity toward catechol, catechin and gallic acid; catechin was the best substrate for the enzyme considering the V_max/K_m ratio. The most effective enzyme inhibitor was sodium sulphite, although ascorbic acid, l ‐cysteine and thiourea were also effective inhibitors at high concentration. But NaCl and citric acid were poor inhibitors of the enzyme. Data generated by this study might help to better prevent lily bulbs browning.     

百合中多酚氧化酶的部分性质

通过丙酮酚法从百合中提取出多酚氧化酶(PPO)的粗酶液。以儿茶酚为底物时它有两个最适pH,分别为4.0、7.0,在pH5.0-6.5之间酶活力可以在4℃保持至少10h。PPO的最适温度为40℃,从40℃开始酶出现热失活,热失活速度符合一级反应动力学。PPO除对L-酪氨酸没有活力外,对儿茶酚、儿茶素、没食子酸均有活力,其中对儿茶素具有最好的底物特异性。亚硫酸钠对PPO的抑制作用最强,高浓度的抗坏血酸、半胱氨酸、硫脲也有很好的抑制作用,氯化钠、氯化钙、柠檬酸的抑制作用较差。     

苹果中多酚氧化酶的性质

以冷冻丙酮制备苹果多酚氧化酮粉,用分光光度法研究pH、温度、抑制剂对酶活性的影响。结果表明：以邻苯二酚为底物,PPO的最适pH为6．6,最适温度为28℃;酶热失活遵循一级动力学,80℃半衰期为1．1min亚硫酸钠,半胱氨酸,抗坏血酸为强烈抑制剂。