食品科学 ›› 2019, Vol. 40 ›› Issue (19): 149-155.doi: 10.7506/spkx1002-6630-20180824-263

• 食品工程 • 上一篇    下一篇

超高压激活鲜榨梨汁多酚氧化酶的酶学特性

尚海涛,宣晓婷,崔燕,林旭东,俞静芬,凌建刚   

  1. (宁波市农业科学研究院农产品加工研究所,宁波市农产品保鲜工程重点实验室,浙江 宁波 315040)
  • 出版日期:2019-10-15 发布日期:2019-10-25
  • 基金资助:
    “十三五”国家重点研发计划重点专项(2017YFD0400700);宁波市科技富民项目(2017C10030)

Characterization of Polyphenol Oxidase from Fresh Pear Juice after High Pressure Processing Activation

SHANG Haitao, XUAN Xiaoting, CUI Yan, LIN Xudong, YU Jingfen, LING Jiangang   

  1. (Ningbo Key Laboratory for Agricultural Products Preservation Engineering, Institute of Agricultural Products Processing, Ningbo Academy of Agricultural Sciences, Ningbo 315040, China)
  • Online:2019-10-15 Published:2019-10-25

摘要: 为探究超高压加速梨汁褐变的机制,以鲜榨翠冠梨汁为原料,采用超高压(500 MPa、5 min)处理,以未处理的梨汁为对照,测定超高压处理对梨汁货架期色泽的影响,检测处理前后梨汁中溶解氧质量浓度和总酚浓度,并研究多酚氧化酶(polyphenol oxidase,PPO)的最适pH值、最适反应温度、热稳定性、米氏常数(Km)。结果表明,超高压组梨汁货架期6 h褐变(色差值(ΔE)=1.61),而未处理组货架期24 h仍未见明显褐变(ΔE=0.56)。鲜榨梨汁PPO存在PPO1(pH 5.0)和PPO2(pH 7.0)两种形式,超高压加速褐变与PPO1和PPO2被激活有关:PPO1活力从6.5 U/mL上升到14.1 U/mL,上升了1.17 倍;PPO2活力从2.6 U/mL上升到25.5 U/mL,上升了8.8 倍。激活效应的内在原因是PPO存在潜在酶活性形式,其本身活力较弱甚至无活力,超高压是激活PPO潜在酶活性的外在原因之一,十二烷基硫酸钠处理、热处理等也可激活PPO潜在酶活性。超高压降低PPO的Km值,提高了其与底物之间的亲和力,从而提高酶促反应速率;PPO1与底物的亲和力1/Km从2.353 L/mol上升到46.909 L/mol,上升了18.9 倍;PPO2与底物的亲和力从2.852 L/mol上升到62.857 L/mol,上升了21.0 倍。超高压同时会降低最大酶促反应速率(Vmax),Vmax对酶促反应速率的影响如超过Km的影响,将会表现出钝化效应。综上,超高压通过激活PPO潜在酶活力、提高PPO与底物的亲和力和酶促反应速率,加速鲜榨梨汁褐变。

关键词: 鲜榨梨汁, 超高压处理, 褐变, 多酚氧化酶, 潜在酶活性, 激活效应

Abstract: This paper aimed to explore the mechanism by which high pressure processing (HPP) accelerates browning of pear juice. The effect of HPP, applied at 500 MPa treatment for 5 min on the color of fresh Cuiguan pear juice during its shelf life was investigated. Untreated samples were used as control. Dissolved oxygen concentration and total phenol concentration in the pear juice were measured before and after HPP treatment, together with enzymatic characteristics of polyphenol oxidase (PPO) such?as?optimal pH, optimal temperature, thermal stability, and Michaelis constant (Km). The results showed that obvious browning in the HPP-treated juice occurred at 6 h (ΔE = 1.61), while no browning was observed in the untreated juice at 24 h (ΔE = 0.56). In the fresh juice there were two forms of PPO: PPO1 (pH 5.0) and PPO2 (pH 7.0), and the accelerated browning of HPP-treated juice was associated with the activation of the two PPOs; the activity of PPO1 was increased from 6.5 to 14.1 U/mL, by 2.17 folds, and the activity of PPO2 from 2.6 to 25.5 U/mL, by 9.8 folds. The potential PPO activities themselves were very weak or even nearly zero, which could be activated by external factors such as HPP, sodium dodecyl sulfate (SDS) and heat treatment. The Km was decreased, and the substrate affinity of PPO was increased by HPP, thereby enhancing the rate of enzymatic reaction. The substrate affinity (1/Km) of PPO1 was increased from 2.353 to 46.909 L/mol, by 19.9 folds, and the substrate affinity of PPO2 from 2.852 to 62.857 L/mol, by 22.0 folds. The maximum reaction rate Vmax was decreased after HPP treatment. Inactivation will occur if the Vmax has a greater effect on the rate of enzymatic reaction than does the Km. To sum up, HPP treatment accelerates browning of fresh pear juice by activating potential PPO, and increasing the substrate affinity of PPO and consequently the enzymatic reaction rate.

Key words: fresh pear juice, high pressure processing, browning, polyphenol oxidase, potential enzyme activity, activating effect

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