摘要：
The utilization of the dead livestock and poultry resources is an important way to solve the pollution of livestock and poultry mortality. To study enzymatic hydrolysis conditions of dead pork, in this paper, the pork as experimental raw material, the trypsin as experiment enzyme, the effects of several operating parameters, namely enzyme concentration, substrate concentration, initial hydrolysis pH value and hydrolysis temperature, were studied taking the degree of hydrolysis as criterion by single-factor method. The conditions for trypsin hydrolysis of pork were preliminarily determined by 4 main factors of pork enzymatic hydrolysis tests. The influence law of the 4 main factors on the enzymatic hydrolysis was analyzed. Subsequently, response surface methodology (RSM) based on a three-level, four-variable Box-Behnken experimental design was employed to evaluate the interactive effects. A mathematical model was established, in which the degree of hydrolysis was as a function of 4 variables. The best enzymatic hydrolysis conditions were determined finally. By analysis on the surface plot of response surface model, the interactions of various factors were analyzed intuitively. On this basis, the effect of pork ultrasonic pretreatment on the enzymatic hydrolysis was studied, and the ultrasonic pretreatment conditions were as follows: ultrasonic frequency 20 kHz; ultrasonic power 500 W; ultrasonic time 20 min. The effect of ultrasonic pretreatment on the degree of hydrolysis was analyzed by scanning electron microscopy (SEM), and the microstructure change could be observed. The final test results showed that the order of the effect of 4 factors on the degree of hydrolysis was as follows: Enzyme dosage>hydrolysis temperature>initial hydrolysis pH value>substrate concentration. The optimum conditions for trypsin hydrolysis of pork in the selected experimental ranges were determined as follows: enzyme dosage of 1.15% (by mass), substrate concentration of 80.5 g/L, initial hydrolysis pH value of 7.96, and hydrolysis temperature of 40.6°C. The predicted degree of hydrolysis under these conditions was 16.74%, and the actual degree of hydrolysis was 16.77%, which showed that the experimental results were in agreement with the results of the software analysis. The optimal hydrolysis time was 6 h, and the degree of hydrolysis reached 28.91% at this point. After ultrasonic pretreatment, the optimal hydrolysis time was 4 h, and the degree of hydrolysis could reach up to 32.86% at this time. It could shorten the hydrolysis test cycle by 2 h, and improve the degree of hydrolysis by 4 percentage points under the condition of ultrasonic pretreatment. The reason that ultrasonic pretreatment enhanced the hydrolysis reaction was that protein chemical bond of pork was broken apart and the cell membrane was damaged by ultrasonic cavitation and mechanical action, then the contact probability between the enzyme and substrate was proved, and the enzymatic reaction was promoted finally. This study demonstrates that ultrasonic pretreatment can increase the hydrolysis efficiency of trypsin hydrolysis of pork and shorten hydrolysis time.%病死畜禽资源化利用是解决病死畜禽污染的一条重要途径.为探索病死猪肉酶解工艺条件,该文以猪肉为原料,以胰蛋白酶为试验用酶,以水解度为指标,选取加酶量、底物浓度、pH值、温度作为试验因素,通过单因素试验初步确定了胰蛋白酶的水解条件,并分析了各因素对酶解反应的影响规律.应用Box-Behnken中心组合设计建立数学模型,以水解度为响应值,进行了四因素三水平的响应面优化试验,确定了最佳酶解工艺条件,并通过响应面模型的曲面图直观地分析了各影响因素之间的交互作用.在此基础上,探索频率为20 kHz,功率为500 W的超声波预处理猪肉20 min对酶解效果的影响,并应用扫描电子显微镜在微观结构上对其原因进行了分析.结果表明,各因素对酶解反应的影响大小依次为:加酶量>温度>pH值>底物浓度,在试验范围内得到的酶解最佳工艺条件为:加酶量为1.15%(质量分数)、底物质量浓度为80.5 g/L、pH值为7.96、温度为40.6°C,酶解1 h的预测水解度可达16.74%,验证试验水解度为16.77%,表明试验结果与软件分析结果相符,最佳水解时间为6 h,此时的水解度为28.91%.超声波预处理后,最佳水解时间为4 h,水解度达到32.86%,因此超声波预处理能缩短水解周期2 h,提高水解度4个百分点.由此可见,应用超声波预处理可以提高酶解效率,缩短工作时间.
