Characterization of polyphenol oxidase and peroxidase from peach mesocarp (Prunus persica L. cv. Babygold)

The two enzymes involved in enzymatic browning reactions—polyphenol oxidase (PPO) and peroxidase (POD)—were extracted from peach fruit mesocarp. PPO was mainly located in the membrane fraction and was in its latent state. However, POD activity was found in the soluble fraction. The kinetic characterization of PPO and POD was carried out with a natural substrate (chlorogenic acid) and with a non‐physiological substrate. Under native isoelectric focusing (IEF), several PPO isoenzymes were present in the pH range 5.4–5.8. A partially denaturing sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS‐PAGE) showed the presence of two active bands with apparent molecular masses of 49 and 50 kDa. A totally denaturing SDS‐PAGE indicated the presence of a single polypeptide with a molecular mass of 60 kDa, as revealed by western blot. POD was also analyzed by IEF, showing the presence of two strongly basic isoenzymes, which were resolved by cationic native PAGE into two different bands. Copyright © 2007 Society of Chemical Industry     

香蕉皮多酚氧化酶和过氧化物酶特性的研究

分别以邻苯二酚、愈创木酚为底物,采用分光光度计测定氧化产物的方法对香蕉皮多酚氧化酶(PPO)和过氧化物酶(POD)的性质进行研究。结果表明,PPO最适温度为30℃、最适pH5.5、Km=22.42mmol/L、Vmax=0.027OD420/min,75℃水浴处理5min基本完全失活,100℃水浴处理20s可钝化PPO活性,并且香蕉皮PPO有同工酶;香蕉皮过氧化物酶(POD)的最适pH6.0、最适温度30℃,其在80℃下处理5min后基本失活,在沸水浴25s后酶被钝化。     

Browning in processed yams: peroxidase or polyphenol oxidase?

Polyphenol oxidase and peroxidase were purified from white yam (Dioscorea rotundata) using DEAE‐cellulose ionexchange chromatography. Thermoinactivation curves for polyphenol oxidase showed monophasic kinetics, while those for peroxidase were biphasic. Urea partially stabilised peroxidase against irreversible thermoinactivation, but did not do so in the case of polyphenol oxidase. Only peroxidase was capable of regenerating activity after thermoinactivation. The results showed that thermoinactivation of peroxidase was mainly due to conformational changes, while that of polyphenol oxidase was probably due to covalent damage. Peroxidase reactivation might play an important role in the browning of processed yam. © 2002 Society of Chemical Industry     

护色处理对冻藏香蕉片多酚氧化酶活性的影响

通过设计单一护色剂、不同浓度L-半胱氨酸、不同热处理时间及多因子护色处理四个实验研究其对冻藏香蕉片多酚氧化酶(PPO)活性的影响,结果表明,5种护色剂均能有效地抑制冻藏香蕉片PPO活性,但L-半胱氨酸对其抑制效果更佳;随着护色液中Cys浓度的增加,残余PPO活力呈下降趋势,表现为CK>0.05%Cys>0.1%Cys>0.2%Cys;热处理能有效地抑制冻藏香蕉片PPO活性,PPO活性随着热处理时间的增加而降低,但热处理却大大降低了冻藏香蕉片的品质;0.1%L-半胱氨酸、0.05%异抗坏血酸、0.1%蔗糖和0.1%氯化钙构成的护色剂组合能有效地抑制PPO活性,并起到良好的保质增脆效果。     

多酚氧化酶的提取及分离纯化研究进展

多酚氧化酶(PPO)是自然界中分布极广的一种金属蛋白酶,能催化邻-苯二酚氧化成邻-苯二醌,也是许多果蔬等农产品酶促褐变的主要原因。本文综述了国内外多酚氧化酶的提取及分离纯化技术的研究进展,包括传统提取方法(如缓冲液匀浆法、丙酮提取法、丙酮粉提取法等)和新型提取方法(超声波辅助提取法和超高压提取法等),传统经典分离纯化方法(如溶剂法、沉淀法)和新型分离纯化方法(如凝胶色谱分离法和离子交换色谱分离法等),旨在为多酚氧化酶的研究和应用,特别是为抑制农产品酶促褐变提供参考。     

Peroxidase and Polyphenoloxidase Activities in Papaya During Postharvest Ripening and After Freezing/Thawing

Changes in peroxidase (POD) and polyphenoloxidase (PPO) activities of papaya (Caricu papava), cv Sunrise, during ripening, freezing and short frozen storage were studied. Fruits were stored at 14°C and 85 90 % RH until maturity for processing was reached (about 21 days). Fruit were frozen cryogenically and frozen slices were stored at — 18°C. POD activity increased in pulp tissue up to the ripe stage, showing a maximum value after 7 days cold storage. Similarly, PPO activity showed an important increase (4 × initial value) on the same date. The quantity of extractable proteins was at a maximum after 15 days storage at 14°C. Freezing and frozen storage (-18°C) produced an increase of POD activity while EPO activity was only slightly affected.     

虾体多酚氧化酶特性及其抑制技术研究进展

多酚氧化酶是虾体中普遍存在的一种含铜金属蛋白,它是虾体发生黑变的主要原因,也是虾保鲜品质控制中亟待解决的技术关键。研究多酚氧化酶的酶学特性,抑制酶促褐变,对于提高虾的食用价值和商品价值至关重要。本文综合介绍了多酚氧化酶的虾体分布、活性特征、分离纯化、分析测定、黑变机理及其抑制方法。     

化学抑制剂对果蔬食品多酚氧化酶性质影响的研究进展

多酚氧化酶(polyphenol oxidase,PPO)引起的酶促褐变会导致果蔬食品色泽劣变,营养成分降低,甚至使其丧失商品价值.化学抑制剂对PPO具有较好的抑制效果,并且使用方便,在果蔬食品中广泛使用,因此对它的研究具有理论和实际意义.该文论述了羧酸、抗坏血酸及其衍生物、含硫氨基酸、酚酸及其他抑制剂对PPO的抑制作...     

橄榄多酚氧化酶和过氧化物酶的抑制剂筛选及其热失活动力学

本项目研究了七种抑制剂(分别为亚硫酸钠、明矾、O-异抗坏血酸、EDTA-2Na、Ca Cl2、多聚磷酸钠、柠檬酸亚锡二钠)对橄榄多酚氧化酶和过氧化物酶活力的影响,并采用两段模型分析了高温处理对橄榄果实多酚氧化酶(PPO)和过氧化物酶(POD)的钝化动力学。结果表明,橄榄中酚类化合物氧化分解与POD和PPO活性有关,在特定的环境条件下,O-异抗坏血酸和柠檬酸亚锡二钠抑制橄榄PPO酶和POD酶活性效果最好,譬如,其浓度为0.20 g/L时,可完全抑制橄榄PPO酶和POD酶活力;热烫时间和温度对橄榄果实PPO和POD活性影响较大,并随着热烫时间和热烫温度的增加,橄榄果实PPO和POD活性呈现先下降后处于平稳的趋势,故高温处理对橄榄PPO和POD的钝化过程符合两段模型。     

Variations of colour,polyphenol oxidase and peroxidase activities during the production of low temperature dried pasta in various durum wheat genotypes

This research was conducted to determine the effect of milling and pasta making on yellow pigment content, soluble brown pigment content, polyphenol oxidase (PPO) activity and peroxidase (POD) activity in semolina and pasta derived from ten durum genotypes. Results showed that pasta colour was impacted more by the milling than by pasta making. The loss of yellow pigment content, POD activity, PPO activity and soluble brown pigment content due to milling was 1.3×, 4.4×, 6.2× and 17.5×, respectively, greater than losses due to pasta making. Dry pasta contained 62%, 31%, 16% and 25% of the original yellow pigment content, brown pigment content and PPO activity and POD activity in the grain, respectively. Semolina yellow pigment content had a positive effect on pasta colour while semolina protein content, ash content and speck count had negative effects. These results indicate the importance of selecting genotypes that have high yellow pigment content and excellent milling qualities.     

Oxidation of polyphenols in unfermented and partly fermented cocoa beans by cocoa polyphenol oxidase and tyrosinase

Unfermented and partly fermented dried cocoa beans have an excessively astringent and bitter flavour owing to their high polyphenol content. Studies on the remaining polyphenol oxidase activity and the oxidation of polyphenols in these beans have been conducted in relation to two factors, ie incubation time (0, 1, 2, 4, 8 and 16 h) and pH of incubation (3.5, 4.5, 5.5 and 6.5). Owing to unsuccessful polyphenol oxidation during incubation of partly fermented beans, incubation–enrichment treatments were carried out by combining incubation time (0, 8, 16, 24 and 32 h) and addition of crude cocoa polyphenol oxidase and purified tyrosinase from mushroom at 88 and 8800 units g^?1 and pH 5.5. Results showed that the remaining polyphenol oxidase activities of unfermented and partly fermented dried beans were 1 and 0.08% with specific activities of 9 and 1% respectively. The polyphenols of unfermented beans were effectively oxidised by incubation at 45 °C and pH 3.5–6.5 without enzyme enrichment; while those of partly fermented beans required enzyme enrichment. Both crude cocoa polyphenol oxidase and tyrosinase could be used for enzyme enrichment, but tyrosinase seemed to be more effective. © 2002 Society of Chemical Industry     

Properties of peroxidase and polyphenol oxidase in natural complexes from walnuts (Juglans regia) and in active DL-DOPA copolymers

Black polymers were extracted from walnut husk homogenates (Juglans regia L) in a phosphate buffer (pH 8.0) after air-bubbling for 24 h. After several purification stages, ultrafiltration and gel chromatography (Sephadex and Ultrogel), three extracts (two soluble and one insoluble) were isolated. At the same time, a synthetic DL DOPA/POD/PPO copolymer was prepared. Two walnut extracts exhibited POD and PPO activity. Both DL DOPA/POD/PPO copolymers (soluble and insoluble) only presented a POD activity. The insoluble and more complex extracts (either natural or synthetic) presented the lowest oxidase activities. The soluble walnut extract exhibited low though not negligible PPO and POD activities. It was observed using electrophoresis and gel chromatography that both enzymes were present in phenolic complexes. The high level of adsorption observed on polyacrylamide gel, as well as the elementary composition of these complexes enabled a comparison with phytomelanins. PPO and POD in the soluble copolymers (from both natural and synthetic sources) were governed by Michaelis-Menten kinetics. Apparent constants were calculated using the Lineweaver-Burk method. The use of limited proteolysis and an inhibitor showed good resistance of POD activity in walnut and synthetic copolymers and exceptional resistance of PPO activity in walnut extracts (particularly insoluble extract).     

Role of anthocyanins,polyphenol oxidase and phenols in lychee pericarp browning

Postharvest browning of lychee fruits is investigated in relation to anthocyanins, polyphenol oxidase (PPO) and phenols, using a purified PPO, anthocyanin extract and phenolic extract. Lychee PPO cannot oxidise the degradation of anthocyanins, but the presence of phenolic extract stimulates pigment degradation by PPO, leading to the development of brown pigments. The absorbance at 410 nm of the reaction solution is correlated well with the peel browning index. Pyrogallol, catechol and 4‐methylcatechol are good stimulators, but no action with chlorogenic acid, p‐cresol, resorcinol or tyrosine is observed. A decrease in the ascorbic acid content of lychee pericarp is associated with an increase in the peel browning index, and the addition of ascorbic acid to the reaction mixture exerts a preventive action on the anthocyanin degradation. Data here suggest that anthocyanins, PPO and phenols contribute to lychee pericarp browning involved in anthocyanin–PPO–phenol reactions. © 2000 Society of Chemical Industry     

Peroxidase and polyphenol oxidase thermal inactivation by microwaves in green coconut water simulated solutions

Enzymes from coconut water such as peroxidase (POD) and polyphenol oxidase (PPO) when in contact with oxygen begin reactions causing nutritional and color losses. Solutions simulating the chemical constituents of coconut water were submitted to a batch process in a microwave oven. PPO and POD inactivation data could be characterized by: PPO/water D_93 °C=16.5 s (z=35.5 °C); PPO/sugars D_91 °C=18 s (z=33°C); POD/water D_91.5 °C=44 s (z=24 °C) and POD/sugars D_92 °C=20.5 s (z=19.5 °C). The contact between salts and enzymes promoted a drastic reduction of the initial activity. After the incidence of microwave energy at temperatures above 90 °C, enzymes activity was not detected. These results can indicate an adequate choice of temperature conditions to inactivate coconut water enzymes. The knowledge of how green coconut water constituents influence POD and PPO activity will supply useful information about microwave processing of coconut water.     

超高压处理对香蕉果肉多酚氧化酶和过氧化物酶活性抑制的研究

为优化超高压处理对香蕉果肉多酚氧化酶(PPO)和过氧化物酶(POD)的酶活残存率,首先研究压力、温度和保压时间对香蕉果肉PPO 和POD 的酶活残存率的影响,然后采用二次回归正交旋转组合设计试验对工艺进行优化。结果表明,超高压处理香蕉果肉PPO 和POD 的酶活残存率影响因素的主次顺序分别为压力＞温度＞保压时间和温度＞压力＞保压时间;超高压处理香蕉果肉PPO 和POD 的酶活残存率最佳工艺参数为压力480MPa、温度55℃、保压时间10min,在此条件下,PPO 和POD 的酶活残存率分别为0.90% 和3.26%。     

Lipolysis in red clover with different polyphenol oxidase activities in the presence and absence of rumen fluid

This experiment aims to determine whether polyphenol oxidase (PPO) can reduce the extent of lipolysis and the consequent polyunsaturated fatty acid loss through microbial biohydrogenation in red clover when incubated in the presence of rumen fluid. PPO is involved in the browning reaction of red clover leaves when cut or crushed and exposed to air. It starts the browning process by oxidizing endogenous phenols to quinones, which contain electrophilic sites. These sites react with nucleophilic sites of other compounds such as proteins and have been shown to reduce proteolysis and lipolysis in silo. Two lines of red clover (cv. Milvus), a genotypic mutant with reduced PPO activity (L) and the wild type (H) with a high level of PPO activity, were cut 3 cm above soil level, crushed and cut into 1 cm strips before being loaded into incubation bottles. These were then incubated in anaerobic buffer at 39 °C in either the absence (?) or the presence (+) of rumen microorganisms. The incubations were then compared over a 24 h time course in terms of lipolytic activity. Characterization of the tissues showed PPO activities of 25.3 and 5.13 U g^?1 fresh weight for H and L, respectively. Lipolysis, measured as the proportional decline in the membrane lipid, was reduced (P < 0.001) with increasing PPO activity in both the presence (+) and absence (?) of rumen microorganisms. However, values were significantly higher (P < 0.001) in the presence of rumen microorganisms, with values after the 24 h incubation of 0.28, 0.42, 0.72 and 0.82 for H?, L?, H+ and L+, respectively. Biohydrogenation of C18:2 and C18:3 polyunsaturated fatty acids were significantly lower in the H+ treatment than the L+ treatment, with mean values after 24 h incubation of 53% and 57% (P < 0.05) for C18:2 and 65% and 74% (P < 0.01) for C18:3, respectively. Changes that occurred in the lipid fractions (membrane lipid, diacylglycerol, triacylglycerol and free fatty acids) during the incubations are also reported and discussed. These results support the selection of forages high in PPO activity to reduce polyunsaturated fatty acid loses in the rumen. Copyright © 2007 Society of Chemical Industry