Phenolic compounds and related enzymes are not rate-limiting in browning development of fresh-cut potatoes

The effect of minimal processing on polyphenol oxidase (PPO), peroxidase (POD), phenylalanine ammonia-lyase (PAL), and phenolic compounds was studied in five potato cultivars (Agria, Cara, Liseta, Monalisa, and Spunta). Minimal processing caused an overall increase in PPO, POD, and PAL activities. The isoform pattern of PPO was the same for all of the cultivars before and after processing. No latent PPO was detected. The isoperoxidase pattern was approximately the same among cultivars. An increase in POD activity was related to the specific induction of an acidic isoperoxidase. PAL showed an induction pattern characterized by the presence of a maximum peak of activity after 4 days of processing for all of the cultivars. The sequence of browning susceptibility of potato cultivars was as follows: Monalisa > Spunta > Liseta > Cara > Agria. Browning development was only partially correlated to PAL activity (only during the first 4 days after wounding). However, this correlation could not explain the above sequence of browning susceptibility. Minimal processing caused an increase of chlorogenic acid, whereas tyrosine content remained unchanged. In summary, no significant correlation was found between either rate or degree of browning and any other biochemical and physiological attribute investigated (PPO, POD, hydrogen peroxide, ascorbic acid content, and initial phenolics content as well as total and individual phenolics accumulation).     

Effect of wash water temperature and chlorination on phenolic metabolism and browning of stored iceberg lettuce photosynthetic and vascular tissues

Cut tissues from distinct anatomical locations in iceberg lettuce (Lactuca sativa L.) were subjected to washing in cold (4 degrees C) and warm (47 degrees C) water with or without chlorine to assess their propensity to discoloration during storage. Total protein (Bradford method) and phenolic (TPH; Folin-Ciocalteu method) contents and polyphenol oxidase (PPO; spectrophotometric method using catechol as a substrate), peroxidase (POD; guaiacol substrate), and phenylalanine ammonia-lyase (PAL; phenylalanine substrate) activities were determined in photosynthetic and vascular tissue from outer and inner leaves. Unprocessed photosynthetic and inner leaf tissues had significantly higher (P < 0.05) levels of protein and TPH and PPO and POD activities than vascular and outer leaf tissues. PAL activities (on a fresh weight basis) were similar in all tissues. Changes in browning (light reflectance measurement) and phenolic metabolism in all four tissue types were observed during aerobic storage at 5 degrees C over 10 days. PAL activity increased in all tissues after 1-2 days of storage and then gradually decreased. POD activity also increased steadily for the storage duration. Protein content and PPO activity remained constant. Edge browning (measured with a Minolta Chroma Meter) and TPH increased in all tissues, especially in outer vascular tissue. Cut photosynthetic and vascular tissues washed at 4 and 47 degrees C with and without 100 microg mL(-1) chlorine for 3 min were analyzed during 7 days in storage at 5 degrees C. Enzyme activities and accumulation of phenolics in all tissues washed at 47 degrees C were significantly (P < 0.05) lower compared to controls or tissues washed at 4 degrees C. Chlorine had no additional effect at 47 degrees C but significantly (P < 0.05) reduced browning and accumulation of phenolics in lettuce washed at 4 degrees C. These results showed that inherent differences between tissues affect phenolic metabolism and browning in stored, fresh-cut lettuce.     

石榴果实发育期果皮褐变及相关酶活性变化

为探明石榴果实发育期果皮褐变及其相关酶活性、总酚和抗氧化能力的变化规律及相互关系,以泰山红和泰山三白甜石榴为试材,研究各发育期果皮褐变度、多酚氧化酶(PPO)活性、过氧化物酶(POD)活性、苯丙氨酸解氨酶(PAL)活性、总酚含量和DPPH自由基清除率的变化。结果表明,2个品种果皮褐变度、PPO活性、POD活性、总酚含量和DPPH自由基清除率随着发育天数的增加逐渐降低,均在7月15日达到最高。2个品种果实发育期果皮PAL活性呈先降低后升高再降低的变化趋势,分别在7月15日和9月13日出现峰值。泰山红抗褐变能力高于泰山三白甜,二者分别在9月23日和9月13日采收能减轻褐变的发生。相关性分析表明,PAL、PPO、POD、总酚和DPPH自由基清除率均与石榴果实酶促褐变密切相关,石榴果实酚类物质含量高低与其抗氧化能力极显著正相关。本研究结果为深入揭示石榴果实褐变机理提供了理论依据。     

Biochemical study of leaf browning in minimally processed leaves of lettuce (Lactuca sativa L. var. acephala)

A series of biochemical parameters, including the concentration of total ascorbic acid (ASA(tot)) and the activities of phenylalanine ammonia lyase (PAL), polyphenol oxidase (PPO), and peroxidases (PODs), was investigated during cold storage (72 h at 4 degrees C in the dark) in fresh-cut (minimally processed) leaves of two lettuce (Lactuca sativa L. var. acephala) cultivars differing in the susceptibility to tissue browning: Green Salade Bowl (GSB), susceptible, and Red Salade Bowl (RSB), resistant. The two cultivars showed differences also at the biochemical level. The content in ASA(tot) increased in RSB, as a consequence of increased DHA concentration; conversely, ASA(tot) diminished in GSB, in which ASA was not detectable after 72 h of storage, thus suggesting a disappearance of ascorbate (both ASA and DHA) into nonactive forms. The antioxidant capacity (as determined by using FRAP analysis) decreased significantly during storage in RSB, while a strong increase was observed in GSB. PAL activity increased soon after processing reaching a maximum by 3 h, then it declined to a relatively constant value in RSB, while in GSB it showed a tendency to decrease in the first few hours from harvest and processing. POD activity, at least for chlorogenic acid, increased significantly during storage only in GSB.     

Characterization and role of polyphenol oxidase and peroxidase in browning of fresh-cut melon

Polyphenol oxidase (PPO) and peroxidase (POD) were extracted from two different varieties of melon ( Cucumis melo L. cantalupensis cv. Charentais and C. melo L. inodorus cv. Amarillo) and characterized using reliable spectrophotometric methods. In both cases the enzymes followed Michaelis-Menten kinetics, showing different values of kinetics parameters between the two cultivars: K m = 7.18 +/- 0.70 mM ('Charentais') and 6.66 +/- 0.20 mM ('Amarillo') mM; V max = 7.93 +/- 0.35 units/min ('Charentais') and 13.82 +/- 0.37 units/min ('Amarillo'), relative to PPO; K m = 24.0 +/- 2.10 mM ('Charentais') and 5.05 +/- 0.19 mM ('Amarillo') mM; V max = 344.83 +/- 10.32 units/min ('Charentais') and 80.64 +/- 2.01 units/min ('Amarillo'), relative to POD. Optimum pH for PPO was 7.0 for 'Charentais' and 7.5 for 'Amarillo, whereas it was 4.5 for both cultivars relative to POD. Melon PPO had maximum activity at 60 degrees C in both 'Charentais' and 'Amarillo' cultivars, whereas POD maximum activity was found at 45 degrees C in 'Charentais' and at 25 degrees C in 'Amarillo'. POD from both cultivars showed higher thermolability compared with PPO, losing >90% of relative activity after only 5 min of incubation at 70 degrees C. POD's activation energy was much higher than that of PPO (Delta E (#) = 86.3 and 160.6 kJ mol (-1) for 'Charentais' and 'Amarillo', respectively). PPO and POD activities from both cultivars showed a decreasing pattern as sugar concentration in the assay medium increased, except in POD extract from 'Charentais', which maintained its activity in the presence of high d-glucose concentration (up to 5 M). Changes in L*, a*, b*, chroma, and hue angle values were chosen to describe the browning development in the samples during storage at 5 degrees C. A slight decrease in L* value and a more marked reduction of a* value were noted in both cultivars above all at the end of storage period. POD activity during storage at 5 degrees C was highly correlated with changes of parameters a*, b*, chroma, and hue angle ( r (2) from 0.82 to 0.97) for cultivar 'Charentais'. According to these results, only POD activity seemed to be involved in browning of minimally processed melon.     

Effect of different washing procedures on phenolic metabolism of shredded, packaged iceberg lettuce during storage

Different washing treatments applying chlorinated, ozonated, and tap water were examined for their effect on the phenolic metabolism of minimally processed iceberg lettuce (Lactuca sativa L.) during storage in consumer-sized bags at 4 degrees C for up to 9 days. To eliminate problems associated with raw material inhomogeneity, processing was conducted on a pilot-plant scale under operating conditions of industrial practice. Inherent product heterogeneity caused by diverse lettuce leaf tissues was compensated for by pooling large-sized samples, and frequent sampling ensured significant data about the activities of phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO), and peroxidase (POD), as well as the contents of caffeic acid derivatives over storage time. In the homogeneous lettuce samples, specific responses caused by different washing procedures were detectable. PAL activity in the samples increased for up to 5-8 days of storage. Compared to tap and ozonated water, the use of chlorinated water (100-200 mg/L free chlorine) for washing trimmed heads or shredded lettuce significantly reduced PAL activity and the concomitant rise of 3,5-di-O-caffeoylquinic acid (isochlorogenic acid isomer) concentrations. The phenolic acids O-caffeoyltartaric (caftaric acid), di-O-caffeoyltartaric (chicoric acid), 5-O-caffeoylquinic (chlorogenic acid isomer), and O-caffeoylmalic were less influenced by different washing treatments. Individual contents either were constant or decreased during storage. Additionally, the novel finding of a further caffeic acid isomer, tentatively identified as meso-di-O-caffeoyltartaric acid, is reported. PPO and POD activities were less affected by different washing treatments and thus were less suitable physiological indicators of stress reactions triggered by alternative processing.     

Cinnamaldehyde inhibits enzymatic browning of cut lettuce by repressing the induction of phenylalanine ammonia-lyase without promotion of microbial growth

Cinnamaldehyde treatment inhibited the browning of cut lettuce during cold storage. In this study, to clarify the mechanism of inhibitory action of cinnamaldehyde against the browning and to show its microbiological merit, its effect on the browning of cut lettuce was compared to that of mild heat treatment. Both cinnamaldehyde and mild heat treatments inhibited the induction of phenylalanine ammonia-lyase (PAL) activity because of cutting. As a result, the biosynthesis of polyphenols, which are substrates of polyphenol oxidase, was inhibited. This reduction of polyphenol synthesis caused the inhibition of the browning. Cinnamaldehyde treatment repressed the induction of PAL mRNA, while mild heat treatment did not repress its induction. The increase in microbes in cut lettuce treated with cinnamaldehyde was less than that treated with mild heat after 12 days.     

甘薯抗薯瘟病的苯丙烷类代谢研究

甘薯瘟病是甘薯的重要病害, 严重影响甘薯的品质和产量.本试验在甘薯苗期, 通过剪叶法接菌研究甘薯抗感病品种苯丙烷类代谢的变化规律.结果表明: 甘薯抗感病品种接菌后PAL、POD活性均有所提高, 且提高速度与品种抗性呈正相关; 抗病品种接菌后木质素和绿原酸含量显著增加, 感病品种接菌后绿原酸、总酚和类黄酮含量波动较大, 木质素含量降低.相关性分析表明, 接菌后木质素和绿原酸的积累与甘薯品种抗病性密切相关.接菌后PAL、POD活性迅速提高并积累木质素和绿原酸等抗病物质是甘薯抗薯瘟病的苯丙烷类代谢基础, 在抗病品种鉴定方面有一定的参考价值.     

Effect of natamycin in combination with pure oxygen treatment on postharvest quality and selected enzyme activities of button mushroom (Agaricus bisporus)

The combined effects of natamycin (NA) and pure oxygen (PO) treatment on microbial and physicochemical characteristics of button mushroom ( Agaricus bisporus ) stored at 4 ± 1 °C for 16 days was investigated. Mushroom respiration rate, weight loss, firmness, color, percent open caps, total soluble solids, microbial and activities of polyphenol oxidase (PPO), phenylalanine ammonia lyase (PAL), and peroxidase (POD) were measured. The results indicate that treatment with natamycin + pure oxygen (NAPO) maintained tissue firmness, inhibited increase of respiration rate, delayed browning and cap opening, and reduced microorganism counts of yeasts and molds compared to control treatment. The efficiency was better than that of NA or PO treatment. Furthermore, NAPO inhibited the activities of PPO, PAL, and POD throughout the storage period. Our study suggests that NAPO treatment has the potential to improve the quality of button mushroom and extend the shelf life.     

短波紫外线处理对采后香菇褐变的影响

为了控制采后香菇在贮藏过程中的褐变并提高其贮藏品质,本试验采用不同剂量(2、4、6和8 kJ·m^-2) 短波紫外线(UV-C)对新鲜香菇进行预处理,研究不同剂量UV-C处理对香菇色泽、总酚含量、多酚氧化酶(PPO)活性、过氧化物酶(POD)活性、超氧化物歧化酶(SOD)活性、过氧化氢酶(CAT)活性、苯丙氨酸解氨酶(PAL)活性、可溶性蛋白含量、还原糖含量、游离氨基酸含量以及香菇表面微生物的影响。结果表明,UV-C处理延缓了香菇色泽的下降,保持了较高的总酚含量,并且有利于POD、SOD、CAT和PAL活性维持在较高水平,抑制了PPO活性,同时可以保持较高的还原糖和可溶性蛋白含量,在一定程度上延缓了蛋白质的降解,而且可以有效抑制微生物的生长,其中4 kJ·m^-2 UV-C处理效果最佳。综上所述,UV-C处理可以有效地抑制香菇褐变,延长其贮藏期。本研究结果为香菇的贮藏保鲜提供了理论依据。     

减压贮藏对去壳黄甜竹笋的保鲜效果及其生理和分子机制

为了探讨减压贮藏延缓采后去壳黄甜竹笋木质化和褐变的效果及其作用机制,本试验以黄甜竹笋为试验材料,笋剥壳后于温度6 ±1℃、相对湿度80%~85%、55 kPa减压环境下贮藏10 d,测定其品质指标、竹笋木质化和褐变关键酶的活性和相关基因的相对表达量。结果表明,减压贮藏(中后期)显著抑制了黄甜竹笋基部切面的褐变(P<0.05),显著延缓了黄甜竹笋的呼吸速率、电导率、硬度,以及丙二醛、纤维素和木质素含量的上升(P<0.05),同时也显著抑制了苯丙氨酸解氨酶(PAL)、过氧化物酶(POD)、肉桂醇脱氢酶(CAD)、多酚氧化酶(PPO)的活性及其编码基因的相对表达量(P<0.05)。本研究结果为减压贮藏在黄甜竹笋采后保鲜的应用提供了理论依据。     

Polyphenol oxidase and peroxidase expression in four pineapple varieties (Ananas comosus L.) after a chilling injury

Pineapple internal browning (IB) is a chilling injury that produces enzymatic browning associated with flesh translucency. Pineapple biodiversity allowed the investigation of how polyphenol oxidase (PPO) and peroxidase (POD) activities with their different isoforms are involved in the IB mechanism. Fruits of four varieties that expressed IB symptoms differently, Smooth Cayenne (SCay) and the hybrids MD2, Flhoran 41 (Flh 41), and Flhoran 53 (Flh 53), were stressed by cold. The susceptible varieties showed classical brown spots but different patterns of IB, whereas MD2 and controls showed no IB. Enzymatic activities were measured on fruit protein extracts and PPO and POD isoforms separated on mini-gels (PhastSystem). Only PPO activity was significantly enhanced in the presence of IB. Up to six PPO isoforms were identified in the susceptible varieties. PPO was barely detectable in the nonsusceptible variety MD2 and in controls. The number of PPO isoforms and the total PPO activity after chilling are varietal characteristics.     

莲藕膨大过程中褐变酶活性的变化

本实验以3个熟性不同的莲藕品种为材料,研究了莲藕膨大过程中褐变酶活性的变化。实验表明:随着膨大节数的增加,过氧化物酶(POD)、多酚氧化酶(PPO)的活性增加;第4节藕体随着采收时间的推迟,褐变酶活性增加,且熟性不同品种之间有明显差异。并探讨了品种熟性、藕体大小和环境条件对莲藕褐变酶的影响。     

Characterization of polyphenol oxidase and peroxidase and influence on browning of cold stored strawberry fruit

Polyphenol oxidase and peroxidase were extracted from two different varieties of strawberry fruit (Fragaria x ananassa D, cv. 'Elsanta' and Fragaria vesca L, cv. 'Madame Moutot') and characterized using reliable spectrophotometric methods. In all cases, the enzymes followed Michaelis-Menten kinetics, showing different values of peroxidase kinetics parameters between the two cultivars: Km = 50.68 +/- 2.42 mM ('Elsanta') and 18.18 +/- 8.79 mM ('Madame Moutot') mM and Vmax = 0.14 +/- 0.03 U/g ('Elsanta') and 0.05 +/- 0.01 U/g ('Madame Moutot'). The physiological pH of fruit at the red ripe stage negatively affected the expression of both oxidases, except polyphenol oxidase from 'Madame Moutot' that showed the highest residual activity (68% of the maximum). Peroxidase from both cultivars was much more thermolable as compared with PPO, losing over 60% of relative activity already after 60 min of incubation at 40 degrees C. The POD activation energy was much lower than the PPO activation energy (DeltaE = 97.5 and 57.8 kJ mol-1 for 'Elsanta' and 'Madame Moutot', respectively). Results obtained from d-glucose and d-fructose inhibition tests evidenced a decreasing course of PPO and POD activities from both cultivars as the sugar concentration in the assay medium increased. Changes in CIE L*, a*, b*, chroma, and hue angle values were taken as a browning index of the samples during storage at 4 degrees C. A decrease in L* was evident in both cultivars but more marked in 'Elsanta'. PPO and POD activities from cv. 'Elsanta' were very well-correlated with the parameter L* (r2=0.86 and 0.89, respectively) and hue angle (r2=0.85 and 0.93, respectively). According to these results, the browning of the fruit seemed to be in relation to both oxidase activities.     

减压贮藏对冷藏枇杷果实品质和木质化败坏的影响

为阐明低温减压贮藏对枇杷果实品质和木质化败坏的影响,该研究应用40～50 kPa压力于(0±0.5)℃下贮藏果实49 d,并每隔7 d测定果实的营养品质、出汁率、硬度、褐变指数、呼吸速率、乙烯产生速率、过氧化物酶(POD)和苯丙氨酸解氨酶(PAL)等木质化相关酶活性及木质素含量的变化.结果表明,减压处理可明显抑制冷藏枇杷果实的呼吸强度、乙烯产生和果实褐变,并保持较高的果实可溶性固形物、可滴定酸度和维生素C含量,抑制贮藏21 d后果实硬度的增加,减轻出汁率的下降,降低POD和PAL酶活性的上升趋势,抑制木质素含量的增加.另外,减压贮藏减轻冷藏枇杷果实的木质化败坏,可能与抑制POD和PAL酶活性有关.低温减压贮藏可有效保持果实食用品质,延长贮藏期,是适合枇杷果实贮藏的有效方法.     

黄瓜枯萎菌粗毒素对不同抗性黄瓜种子萌发及幼苗胁迫作用研究

利用黄瓜枯萎菌粗毒素处理不同抗性品种黄瓜,研究粗毒素对黄瓜种子萌发及幼苗生长的影响,结果表明:粗毒素对不同抗性的黄瓜胚根生长和侧根分化均有抑制作用,且随处理浓度增加,抑制作用愈加明显,但粗毒素对感病品种的抑制作用强于抗病品种。粗毒素胁迫使黄瓜胚根的细胞膜透性、MDA含量、可溶性糖含量增加,其中感病品种的细胞膜透性和MDA含量高于抗病品种,但可溶性糖含量低于抗病品种。在粗毒素处理36 h内,抗感品种黄瓜幼苗的PAL、PPO和POD酶活性均呈现先升高后降低的趋势,其中PAL和PPO的峰值出现在粗毒素处理12 h,而POD的峰值出现在粗毒素处理24 h,抗病品种的3种酶活性峰值高于感病品种。表明抗病品种对黄瓜枯萎菌粗毒素胁迫的抵抗能力强于感病品种。     

Relationships between salicylic acid content,phenylalanine ammonia-lyase (PAL) activity,and resistance of barley to aphid infestation

It has been suggested that salicylic acid (SA) is a signal in acquired resistance to pathogens in several plants. Also, it has been suggested that infestation of plants causes an increase in the activity of phenylalanine ammonia-lyase (PAL), a key phenolic biosynthesis enzyme. The purpose of this work was to investigate whether the induction of SA and PAL activity is related to the susceptibility of barley to aphid infestation. The induction of free and conjugated SA in two barley cultivars that differ in susceptibility to aphids was analyzed. Analyses of several physiological parameters showed that cv. UNA-80 was more susceptible to the aphid Schizaphis graminum than cv. LM-109. Salicylic acid was not detected in noninfested plants. Levels of free and conjugated SA in cv. LM-109 and of conjugated SA in cv. UNA-80 increased with aphid infestation, whereas the levels of free SA in cv. UNA-80 remained high under all infestation degrees. Maximum values reached in both cultivars were not significantly different. With respect to PAL activity, cv. LM-109 showed a significantly higher specific activity than cv. UNA-80, the more susceptible cultivar. The relationship between the susceptibility of a plant to aphid and SA induction and PAL activity is discussed.     

Stimulation of phenolic metabolism by silicon contributes to rice resistance to sheath blight

Sheath blight caused by Rhizoctonia solani is a major disease of rice worldwide. Silicon (Si) can enhance rice resistance to sheath blight, but the relation with phenolic metabolism is poorly known. Two rice cultivars with different levels of resistance to R. solani (resistant Teqing and susceptible Ningjing 1) were studied to determine the effects of Si on disease intensity (rated from 0 to 9) and the involvement of phenolic compounds in disease resistance. Variation in the concentrations of phenolics (including total soluble phenolics, flavonoids, and lignin) and in the activities of defense‐related enzymes polyphenoloxidase (PPO) and phenylalanine ammonia‐lyase (PAL) in rice leaf sheaths was investigated. The results show that Si application reduced sheath‐blight disease ratings of Ningjing 1 and Teqing by 2.96 and 0.65, respectively. In uninoculated plants, Si application alone had no significant effects on the concentrations of phenolic compounds or on the activities of PPO and PAL. In inoculated plants, Si application increased phenolics concentrations and PPO and PAL activities only in the susceptible cultivar Ningjing 1. We conclude that Si‐induced enhancement of phenolic metabolism contributed to the improved resistance of rice to sheath blight in the sensitive cultivar.     

植物防御酶与桉树对焦枯病抗性的关系

为筛选鉴定桉树对焦枯病(CylindrocladiumqulnqueSeptatumMorgan)抗性的生化指标,对福建省11个桉树主栽种系接种焦枯病菌前后关键防御酶系中的过氧化物酶(POD)和超氧岐化酶(SOD)活性及其同工酶变化进行了研究.结果表明:无论是桉树健叶还是接种后的病叶,体内POD、SOD活性均表现为抗病种系>中抗种系>中感种系>感病种系,呈规律性变化;接种前后POD同工酶谱带数与桉树对焦枯病的抗性间不呈规律性变化,而SOD谱带数与抗病性成正比.从抗病育种角度出发可将接种前后桉树POD、SOD活性和接种后SOD同工酶谱带作为桉树对焦枯病抗性早期鉴定的生化指标.     

Low-density lipoprotein antioxidant activity of phenolic compounds and polyphenol oxidase activity in selected clingstone peach cultivars

The antioxidant potential of eight clingstone peach cultivars was investigated by determining phenolic compounds and inhibition of low-density lipoprotein (LDL) oxidation. Cultivars low in polyphenol oxidase (PPO) were also selected to minimize enzymatic browning. Inhibition of LDL oxidation varied from 17.0 to 37.1% in peach flesh extract, from 15.2 to 49.8% in whole peach extract, and from 18.2 to 48.1% in peel extract. Total phenols were 432.8-768.1 mg/kg in flesh extract, 483.3-803.0 mg/kg in whole extract, and 910.9-1922.9 mg/kg in peel extract. The correlation coefficient between relative LDL antioxidant activity and concentration of total phenols was 0.76. Peel PPO activity was higher than flesh activity in most cultivars. The lowest PPO and specific activities were found in the Walgant cultivar, followed by Kakamas and 18-8-23. These three cultivars combine the desirable characteristics of strong antioxidant activity, low PPO activity, and lower susceptibility to browning reactions.