Effects of Terpinen-4-ol on Four Metabolic Enzymes and Polyphenol Oxidase (PPO) in Mythimna separta Walker

To study insecticidal mechanism of terpinen-4-ol, a main insecticidal composition in the essential oil of Sabina vulgaris, the 5th instar larvae of Mythimna separta, were investigated with terpinen-4-ol by topical application. The activities of phosphatase, glutathione S-transferase （GSTs）, cytochrome P450 （P450）, and polyphenol oxidase （PPO） of tested insects were determined in all poisoning stages, including exciting stage, convulsing stage, paralysis stage, and recover stage. The result showed that the activities of both acid phosphatase （ACP） and alkaline phosphatase （AKP） in treated insects were induced by terpinen-4-ol, but ACP was inhibited in paralysis stage. The activities of GSTs were inhibited in exciting stage, convulsing stage, and paralysis stage, but gradually recovered in recover stage. O-demethylase activity of cytochrome P450 was inhibited by terpinen-4-ol, and the inhibition rate in all poisoning stages were 26.27, 46.03, 80.24, and 90.22%, respectively. PPO activities were strongly inhibited by terpinen-4-ol both in vitro and in vivo. In conclusion, the activities of P450, GSTs, and PPO could have relation with toxicity of terpinen-4-ol against larvae of the Mythimna separta, but recover stage of the poisoning insects might be related to GSTs induced. As a new insecticide or synergist, terpinen- 4-ol has a potential value in field of insecticide resistance management.

Potential of plant polyphenol oxidases in the decolorization and removal of textile and non-textile dyes

In this study an effort has been made to use plant polyphenol oxidases; potato （Solanum tuberosum） and brinjal （Solanum melongena）, for the treatment of various important dyes used in textile and other industries. The ammonium sulphate fractionated enzyme preparations were used to treat a number of dyes under various experimental conditions. Majority of the treated dyes were maximally decolorized at pH 3.0. Some of the dyes were quickly decolorized whereas others were marginally decolorized. The initial first hour was sufficient for the maximum decolorization of dyes. The rate of decolorization was quite slow on long treatment of dyes. Enhancement in the dye decolorization was noticed on increasing the concentration of enzymes. The complex mixtures of dyes were treated with both preparations of polyphenol oxidases in the buffers of varying pH values. Potato polyphenol oxidase was significantly more effective in decolorizing the dyes to higher extent as compared to the enzyme obtained from brinjal polyphenol oxidase. Decolorization of dyes and their mixtures, followed by the formation of an insoluble precipitate, which could be easily removed simply by centrifugation.

Polyphenol Oxidase, Peroxidase and PhenylalanineAmmonium Lyase Induced in Postharvest Peach Fruitsby Inoculation with Pichia membranefaciensor Rhizopus stolonifer

Rhizopus rot of peach fruits could be significantly suppressed by Pichia membranefaciens. Polyphenol oxidase (PPO), peroxidase (POD) and phenylalanine ammonium-lyase (PAL) activities induced by inoculation with P. membrane faciens or R. stolonifer were studied in postharvest peach fruits. The activities of PPO and PAL in peaches increased significantly after being inoculated with P. membrane faciens + R. stolonifer by 24 h, the activities maintained at a high level throughout the experiment. Under the condition of infected with R. stolonifer alone, activity of PPO and PAL could also increased, but the levels were lower than those treated with P. membrane faciens+ R. stolonifer. However, fruits inoculaed with P. membrane-faciens + R. stolonifer or R. stolonifer alone did not stimulated POD activity. The results suggest that the activation of these defense enzymes is involved in the action of P. membrane faciens against R. stolonifer.

Inhibitory effect of tartary buckwheat seedling extracts and associated flavonoid compounds on the polyphenol oxidase activity in potatoes(Solanum tuberosum L.)

To improve the processing quality of potatoes,phosphate buffer extract(PBE),50%ethanol(E50),and aqueous extract(AE)of tartary buckwheat seedlings were evaluated for their ability to inhibit the enzymatic browning of potatoes.The results suggest that all extracts of tartary buckwheat seedlings exert significant inhibitory effects on the polyphenol oxidase(PPO)activity in potatoes.The relative concentrations required for a 50%reduction in the PPO activity(IC50)were 0.21,0.28 and 0.41 mg mL^-1,for PBE,E50 and AE,respectively.The strongest inhibitory activity was observed for PBE,followed by E50 and AE.Four flavone compounds in the PBE of tartary buckwheat seedlings(i.e.,rutin,kaempferol-3-O-rutinoside,quercetin,and kaempferol)were identified by high-performance liquid chromatography.These compounds were subsequently evaluated for their roles in the inhibition of PPO from potatoes using a model system.The results indicated that rutin exhibited the highest inhibition rate on the PPO of potato.A synergistic inhibitory effect was observed by mixing rutin,kaempferol-3-Orutinoside,quercetin,and proteins.The inhibitory patterns of rutin,kaempferol-3-O-rutinoside,and quercetin on the enzyme were noncompetitive and reversible,with inhibitory constants of 0.12,0.31,and 0.40 mg mL^-1,respectively.Flavonoids from tartary buckwheat seedlings may exhibit a common mechanism with phenolic compounds,involving the blockage of the reaction of oxygen with PPO leading to the inhibition of the enzymes involved in browning.Based on these results,extracts of tartary buckwheat seedlings can be used as potent natural inhibitors.

Study on Polyphenol Oxidase Activity and Total Phenol Content of Docynia indica

[Objectives] To study the polyphenol oxidase (PPO) activity and total phenol content of Docynia indica .[Methods] The tender branches and petioles of the medicinal and edible plant D. indica were used as experimental materials, and the annual changes of total phenol content and polyphenol oxidase (PPO) activity were analyzed.[Results] pH 7.0 was the optimum value of polyphenol oxidase activity in D. indica plants;the best substrate was catechol with the concentration of 0.15 mol/L;the optimum temperature was 25 ℃.[Conclusions] The total phenol content of D. indica plants was the lowest in April, May and September of each year. The polyphenol oxidase activity of D. indica plants at flowering and fruiting stage was significantly higher than that at pre-flowering stage.

Study on Enzymatic Kinetics of Nephelium lappaceum Polyphenol Oxidase

[Objective] This study aimed to investigate the enzymatic kinetics of polyphenol oxidase in peel of Nephelium lappaceum to explore the environmental factors affecting its catalytic activity. [ Method ] With N. lappaceum peel as the experimental material and catechol as substrate, effects of temperature, pH, four inhibitors （EDTA, Vc, sodium bisulfite and citric acid） and substrate concentration on the activity of polyphenol oxidase were investigated. [ Result] The optimal temperature for polyphenol oxidase in N. lappaceum peel was 40 %, and the optimal pH was 6.8. EDTA, Vc, sodium bisulfite and citric acid all showed ideal in- hibitory effects on the activity of polyphenol oxidase; specifically, EDTA had the strongest inhibitory effect. [ Conclusion] At low temperature, EDTA, Vc, sodi- um bisulflte and citric acid with certain concentrations can inhibit the catalytic activity of polyphenol oxidase in peel of N. lappaceum, extend the storage and trans- portation time of N. lappaceum fruit, and inhibit the browning. This study provides theoretical basis for the development and utilization of polyphenol oxidase in peel of N. lappaceum.

Purification and Partial Characterization of Polyphenol Oxidase from Sapodilla Plum (<i>Achras sapota</i>)

The browning of fruits can be considered as an enzymatic oxidation which is believed to be one of the main causes of quality loss during post-harvest handling. The enzymes responsible for this are the oxidoreductases;the polyphenol oxidase (PPO) (monophenol, o-diphenol, oxygen oxidoreductase;EC 1.14.18.1) belongs to this group. This enzyme, which is found in the sapodilla plum (Achras sapota), was purified using a phenylsepharose and a SephacrylS-200 columns. The molecular weight of the purified enzyme was estimated to be about 66 kDa by gel filtration and 29 kDa by SDS-PAGE. A single protein band was found using the latter system (SDS-PAGE), which shows that the PPO of the pulp of the sapodilla plum may be composed of two protein subunits with similar molecular weight. The optimum pH was 7.0 and the optimum temperature 60℃. The most effective inhibitors tested were ascorbic acid, sodium metabisulfite and acetic acid.

Polyphenol Oxidase Inactivation by Microwave Oven and Its Effect on Phenolic Profile of Loquat (<i>Eriobotrya japonica</i>) Fruit

The objective of this research was investigated the effect of polyphenol oxidase microwave treatment on phenolic composition, antioxidant activity and microstructure of loquat fruit. Phenolic profile of methanolic extracts prepared from fresh, and microwave-treated samples were analyzed. Antioxidant activity was also evaluated by 2,2’-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS?+) and 1,1-diphenyl-2-picrylhydrazyl (DPPH+) methods. In addition, polyphenol oxidase inactivation was carried out using a response surface methodology to establish the optimal conditions of treatment. The phenolic content of fresh mesocarp was 311 ± 0.60 mg gallic acid equivalents (GAE)/100g dry weight (DW) and that of microwave-treated mesocarp was 1230 ± 0.36 mg GAE/100g DW. Total phenolic content of water/ methanol extract significantly increases after microwave treatment rather than methanolic extract of fresh loquat. Five glycoside phenolics were identified by HPLC-DAD-MS as 3-caffeoylquinic acid, 3-p-coumaroylquinic acid, 5-caffeoylquinic acid and quercetin-3-O-sambubioside. Methanolic extract of microwave-treated mesocarp showed higher antioxidant activity than that of fresh mesocarp. Thus, polyphenol oxidase inactivation by microwave energy preserved the integrity of phenolic compounds as well as antioxidant activity in mesocarp extracts prepared from loquat fruit. It was also noted that phenolics were more abundant in the microwaved samples than in the fresh samples.

Polyphenol Oxidase Activity and Inhibition in White Yam （Dioscorea Rotundata. Var. Laasirin） Chips as African Fries for Human Consumption

Polyphenol oxidase, a bi-functional enzyme, has been implicated in enzymatic browning of yam and other tubers in a negative way. The objective of this present study was to examine the activity of polyphenol oxidase in Dioscorea rotundata. Var. laasirin and the efficiency of heat and chemical treatments in inhibiting this enzyme. Crude Polyphenol Oxidase （PPO） of Dioscorea rotundata.Var. Laasirin was isolated and the kinetics studied using the lineweaver-burk plot. The activity of the enzyme was evaluated using spectrophotomeric method. Yam PPO catalyzes oxidation of various substrates with catechol being the most readily oxidized substrate. The Michaelis-Menten constant （Km） and maximum reaction velocity （Vmax） for yam PPO were 0.00037 and 0.3125 respectively. Inhibition data showed that the enzyme had least activity （71.70） when blanched at 95℃ for 7 mins with chemical treatment involving a combination of 0.5% Sodium metabisulphite （Food grade） and 0.5% Ascorbic acid （Food grade）. The activity was highest （83.02） when it was blanched at 95 ℃ for 7 rains. This study has shown that it is possible to inhibit polyphenol oxidase activity in white yam using the chemical pretreatments and processing conditions described in this study for possible adoption in the production of packaged frozen yam chips by food industries.

Polyphenol Oxidase Activity and Colour Changes of ‘Starking’ Apple Cubes Coated with Alginate and Dehydrated with Air

The objective was to study the effect of alginate coating on polyphenol oxidase (PPO) activity and colour of ‘Starking’ apple cubes during dehydration with hot air. Apple cubes were dehydrated at 20oC, 35oC, or 40oC, with a parallel airflow. Analysis of PPO activity, colour (L*, a*, b*) and dry matter were performed along the dehydration process at each temperature. All samples presented a peak in relative PPO activity in the beginning of the drying. Exponential models fitted well the experimental data after the peak. Cubes without coating presented lower PPO activity, suggesting lower browning, than coated samples throughout the dehydration process, for all temperatures. Better results for coated samples were obtained with a perpendicular airflow drying at 40oC, after dipping the whole apple in water at 60oC for 10 min. In order to prevent coated samples from browning, drying by perpendicular airflow preceded by a thermal treatment of the whole apple is required.

Immobilization and Properties of Polyphenol Oxidase

[Objectives]This study was conducted to investigate the effects of embedding conditions on activity and catalytic properties of immobilized polyphenol oxidase.[Methods]Polyphenol oxidase was immobilized in a polymer material by the embedding method,and the optimal immobilization conditions were obtained by single factor tests:CaCl_(2) concentration 2.0%,sodium alginate concentration 2.0%,immobilization time 2 h and mass ratio of enzyme to carrier 10 mg/100 g,under which the immobilized enzyme activity was 93.33 U/g.Under the above conditions,the properties of polyphenol oxidase immobilized by sodium alginate(A-PPO)and free polyphenol oxidase were studied.[Results]The thermostability of A-PPO was better than that of the free enzyme,but the pH stability of A-PPO was inhibited.The Michaelis constant K_(m) values of free polyphenol oxidase and A-PPO were 0.37 and 0.48 mmol/L,respectively,and the maximum reaction rate V_(max) values were 0.38 and 0.51 mmol/(L·g),respectively.[Conclusions]This study provides a theoretical basis for the study of the properties of polyphenol oxidase.

铁棍山药PPO的最适底物、热稳定性及褐变控制研究

研究了铁棍山药多酚氧化酶(PPO)的最适底物、热稳定性及不同抑制剂对其活性的影响。结果表明:在试验范围内,铁棍山药PPO最适底物为邻苯二酚;在较低温度(20～40℃)下其PPO能保持较稳定的活性,较高温度(40～80℃)下其活性快速下降,80℃下保温10 min,PPO活性几乎为零;抑制剂对PPO活性的抑制效果依次为:L-抗坏血酸>L-半胱氨酸盐酸盐>植酸>柠檬酸。

不同品种辣椒POD和PPO活性与辣椒素含量的关系

对3个不同辣椒品种胎座和果肉中的辣椒素与其降解相关酶之间的关系进行了研究,结果表明:不同品种辣椒,不同发育时期辣椒素含量及积累特点不同,其中牛角椒胎座中辣椒素含量最高,沈椒4号次之,农大40最少;沈椒4号果肉中辣椒素含量略高于牛角椒中含量,农大40果肉中最少,仅在花后36d检测到极少量的辣椒素,并且胎座中辣椒素的含量均高于果肉中。在整个辣椒果实发育时期,过氧化物酶、多酚氧化酶活性均和辣椒素含量呈一定的负相关,因此,这两种酶有可能参与了辣椒素的氧化降解,且在该过程中有互补的作用。

新疆小麦品种中多酚氧化酶(PPO)活性基因等位变异的分布

为了给新疆选育低PPO活性的小麦品种以及进行面制品色泽的改良提供依据,利用PPO基因的功能标记PPO18、PPO16和PPO29,检测了252份新疆小麦品种(系)(包括农家品种、国内外引进品种和20世纪60年代以来的育成品种)中2A和2D染色体上PPO等位基因Ppo-A1a(高PPO活性)、Ppo-A1b(低PPO活性)、Ppo-D1a(低PPO活性)和Ppo-D1b(高PPO活性)的分布。结果表明,含有Ppo-A1a、Ppo-A1b、Ppo-D1b和Ppo-D1a基因型的频率分别为69.0%、31.0%、13.1%和86.9%,以Ppo-A1a和Ppo-D1a基因型为主;两个PPO基因的等位基因组合Ppo-A1a/Ppo-D1b(高PPO活性)、Ppo-A1a/Ppo-D1a(中等偏高PPO活性)、Ppo-A1b/Ppo-D1b(中等偏高PPO活性)和Ppo-A1b/Ppo-D1a(低PPO活性)的分布频率依次为9.9%、59.1%、3.2%和27.8%。总体来看,新疆小麦品种PPO活性中等偏高的材料最多。农家品种、引进品种和育成品种的PPO活性基因分布频率存在明显差异,其中具有Ppo-A1b/Ppo-D1a等位基因组合类型(低PPO活性)的分布频率依次为21.3%、33.3%和27.1%。共检测出了70个具有Ppo-A1b/Ppo-D1a等位基因组合类型(低PPO活性)的品种(系),并且新疆育成的冬小麦品种(系)携有Ppo-A1b/Ppo-D1a等位基因组合类型(低PPO活性)的数量比春小麦要多,可在培育低PPO活性品种时作为杂交亲本利用。

小麦多酚氧化酶(PPO)活性的基因型、环境及其互作效应的分析

选用 16个有代表性的小麦品种 (系 ) ,分别在山东 8个地区进行小麦多酚氧化酶 (PPO)活性的基因型与环境效应分析。结果表明 ,在随机效应中 ,小麦籽粒PPO活性的品种与试点互作效应最为显著。在对面团白度性状进行选择时 ,既要选择多酚氧化酶 (PPO)活性低的品种 ,又要注意选择在PPO积累低的地区种植 。

小麦多酚氧化酶(PPO)活性的SSR标记筛选与验证

小麦多酚氧化酶(PPO)活性是引起面条和面团储存期间颜色褐变的主要因素。发掘可应用于籽粒PPO活性辅助选择的分子标记,将有助于小麦面粉颜色性状的遗传改良。本研究选用来自全国不同麦区的203份冬小麦品种,验证SSR(simplesequencerepeat)引物Xgwm312的PCR扩增片段大小与籽粒PPO活性的关系。结果表明,198bp扩增片段的有无同籽粒PPO活性大小密切相关,该片段的出现通常意味着籽粒PPO活性较高。Xgwm312可应用于籽粒PPO活性的分子标记辅助育种。

密集烘烤工艺对烟叶多酚类物质含量及PPO活性的影响

为密集烘烤有针对性地调控多酚含量和优化完善烘烤工艺,采用电热式温湿自控密集烤烟箱,研究了3个密集烘烤工艺烘烤过程中烟叶多酚类物质含量及PPO活性的变化。各密集烘烤工艺处理烟叶的总酚、绿原酸、新绿原酸、隐绿原酸、芸香苷含量随着烘烤进程总体呈升高趋势。烤后烟叶的总酚、绿原酸、新绿原酸和隐绿原酸含量以K2工艺处理较高,芸香苷和莨菪亭含量以K3工艺处理较高。棕色化反应的敏感时期(45℃-47℃)的PPO活性以K2工艺处理最低,K3工艺处理最高。研究表明K2工艺处理能很好地抑制酶促棕色化反应的发生,调控密集烘烤过程中多酚类物质的变化,烤后烟叶多酚类物质含量高。

不同品种南瓜POD及PPO同工酶的比较研究

采用垂直平板聚丙烯酰胺凝胶电泳技术 ,对美洲南瓜 (Cucurbita pepo L.)和中国南瓜 (C.moschata Duch.)共 10个不同栽培品种的过氧化物酶 (POD)及多酚氧化酶 (PPO)同工酶进行了分析研究 ,并采用聚类分析法对其亲缘关系进行了比较和分类。结果表明 ,10个不同品种的南瓜根 POD共分离出 10条酶带 ,其中 e1、e4、e5、e6和 e9为 2个栽培种所共有 ;e4、e5、e6分离清晰 ,可作为 2个栽培种的特征酶带 ;PPO同工酶分离出 6条酶带 ,其中 e′1和e′4分离清晰 ,为 2个栽培种所共有。

普通小麦籽粒LOX、PPO活性和类胡萝卜素含量变异及对全麦粉色泽的影响

【目的】研究新鲜全麦粉及储存3个月后全麦粉中脂肪氧化酶(LOX)和多酚氧化酶(PPO)的活性及类胡萝卜素含量的变化情况,分析其色泽性状与LOX、PPO活性和类胡萝卜素含量之间的关系,了解储存时间对全麦粉色泽的影响及其品种间的差异,为全麦粉或面粉色泽的改良和相关的育种工作提供参考依据。【方法】选用63个普通小麦品种(系)为试验材料,用分光光度计法测定其LOX、PPO活性和类胡萝卜素含量,用色差计法测定全麦粉色泽性状。【结果】品种间LOX、PPO活性、类胡萝卜素含量及其它色泽性状差异显著或极显著;新鲜全麦粉和存储3个月的全麦粉的LOX、PPO活性变化较大,储存期间LOX、PPO活性丧失较快,类胡萝卜素含量相对稳定;储存3个月后的白度和亮度等色泽性状存在一定程度的变化,但变化不大。新鲜全麦粉的白度和亮度均与PPO活性呈极显著负相关,黄度与类胡萝卜素含量呈极显著正相关,LOX活性与类胡萝卜素含量呈极显著负相关,而LOX活性与白度、亮度和黄度间相关均不显著;储存3个月的全麦粉的白度和亮度仅与PPO活性呈极显著负相关。【结论】全麦粉色泽改良中,PPO活性和类胡萝卜素的遗传控制是关键;通过储存全麦粉或面粉的方式并不能大幅度的提高其亮度、白度和降低黄度。所以,遗传育种方法是改良小麦粉白度性状和类胡萝卜素含量等品质性状的重要途径。

苹果汁的前褐变及苹果PPO的部分特性研究

苹果汁贮藏过程中色泽加深、色值和透光率不稳定是苹果汁生产中普遍存在的问题,文中尝试采用苹果自身的PPO进行前褐变反应,以减少果汁中的酚类化合物,防止后期褐变。研究表明,果汁经30℃前褐变反应4 0min后,总酚质量含量可下降约30 % ,可溶性固形物质量分数提高了2 8% ,常温下贮藏3个月后,色值和透光率基本保持不变,明显地改善了苹果汁的质量指标。同时,文中还探讨了PPO的最适pH值、最适反应温度及其pH稳定性。