摘要
研究了豌豆发酵液(PFB)对莲藕多酚氧化酶(PPO)活性的影响及其机理。采用硫酸铵分级沉淀,DEAE-52和Sephadex G-75柱层析纯化莲藕PPO,测定PFB对PPO的抑制动力学和圆二色谱。结果表明,所得PPO纯化酶纯化倍数、比酶活和蛋白质回收率分别为99.71倍,2 627.36 U/mg和1.85%;PFB(4.0 mg/m L)对PPO的抑制率高达98.65%,其半抑制质量浓度IC50为1.81 mg/m L,对PPO的抑制为竞争性可逆抑制;莲藕PPO蛋白3级结构为全α型,2级结构主要由α-螺旋和无规卷曲构成,PFB作用后α-螺旋含量显著减少,β-折叠含量显著增加。豌豆发酵液是天然有效的多酚氧化酶抑制剂,通过改变酶的2级结构发挥抑制作用。本研究结果为莲藕保鲜提供了新的途径。
The effect and mechanism of pea fermentation broth（PFB） on polyphenol oxidase（PPO） of lotus root was investigated. Extraction and purification of the PPO by ammonium sulphate fractionation, DEAE-52 and Sephadex G-75 column chromatography were performed. Inhibition kinetics and Circular dichroism（CD） spectroscopy was employed to study the inhibition mechanism. Results showed that purification fold, specific activity and yield of the final purified PPO were 99.71, 2627.36 U/mg and 1.85%, respectively. The inhibition of PFB（conc. 4.0 mg/m L） to the PPO was 98.65%and the value of IC50 was 1.81 mg/m L. Kinetic analyses showed that PFB was reversible and competitive inhibitor to the enzyme. CD spectroscopy analysis showed that lotus root PPO was all-α protein and its secondary structure was mainly composed of α-helix and random coil structure. After interacting with PFB, a significant decrease in α-helix and increase in β-sheet was detected. It is concluded that the PFB is a natural and effective inhibitor to the PPO by changing the secondary structure of PPO protein, providing a new approach for fresh-keeping of lotus root.
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2016年第5期76-81,共6页
Journal of Chinese Institute Of Food Science and Technology
基金
国家自然科学基金项目(31071619)
关键词
豌豆发酵液
莲藕
多酚氧化酶
抑制作用
圆二色谱
pea fermentation broth
lotus root
polyphenol oxidase
inhibition
circular dichroism spectroscopy
