超声结合苹果酸处理对双孢蘑菇多酚氧化酶活性及失活动力学的影响

Effect of combing ultrasound with malic acid on activity and inactivation kinetics of polyphenol oxidase from mushroom (Agaricus bisporus)

以双孢蘑菇多酚氧化酶(PPO)为原料,首先考察不同浓度苹果酸处理对PPO酶活性的影响,发现随着苹果酸浓度的增大,PPO酶活性逐渐降低,当苹果酸浓度为60mmol/L时,其活性几乎被完全抑制。然后进一步研究了单独超声处理、中等浓度的苹果酸处理以及超声结合中等浓度苹果酸处理对PPO相对活性的影响,并且对超声结合中等浓度的苹果酸处理PPO的失活动力学进行研究。结果表明:单独超声处理并不能迅速降低PPO的活性,经过60 min处理后PPO活性仍高达81.8%。单独中等浓度的苹果酸处理,在15,20mmol/L苹果酸处理不能显著降低PPO的活性,即使在较高浓度25 mmol/L处理60 min后残留活性仍为63.9%。超声结合中等浓度酸处理能够显著降低PPO的活性,分别经单独超声处理、20mmol/L苹果酸处理和超声结合20 mmol/L苹果酸处理60 min后PPO活性分别为81.8%,79.7%,12.6%。超声结合中等浓度苹果酸处理PPO的失活动力学均符合一级动力学方程,且失活速率常数(k)随着酸浓度的增加而增大,当苹果酸浓度分别为0,25mmol/L时,其失活速率常数(k)为0.33×10-2,4.36×10-2 min-1。同时D值、半衰期(t1/2)随着苹果酸浓度增大而减小并且明显低于单独超声处理的PPO,当苹果酸浓度分别为0,25mmol/L时,其D和t1/2分别为690.9,208.0min和52.6,15.9min。

The effect of malic acid was studied at different concentrations on the activity of polyphenol oxidase （PPO） from mushroom （Agaricu.; bisporus）. It was observed that the activity of PPO decreased gradually with increasing concentration of malie acid, andPPO was almost completely inhibited by 60 mmol/L malie acid. Then effect of ultrasound, malic acid at medium concentration and combing ultrasound with malic acid at medium concentration treatment on PPO was further investigated. It was found that the activity of PPO could not be inhibited by malic acid at medium concentration, and only the ultrasound treatment could not obviously reduce the activity of PPO. However, combing ultrasound with malic acid at medium concentration could significantly reduce the activity of PPO. When PPO was treated by combining ultrasound with 20 mmol/L malic acid, only 12.6% activity was retained. The inhibition kinetics of PPO was treated by combining ultrasound with medium malic acid conforming to first order kinetics equation. The inactivation rate constant （k） increased with increasing concentration of malic acid, and both of the decimal reduction time （D） and half-life （t1/2） reduced significantly with increasing concentration of malic acid. When PPO was treated by ultrasound and combing ultrasound with malic acid at 25 mmol/L, the decimal reduction time （D） and half-life （t 1/2 ） were 690.9, 208. 0 min and 52.6, 15.9 min,respectively.