双酶分步水解牛骨蛋白工艺的优化

Optimization of Stepwise Dual-enzyme Hydrolysis of Bovine Bone Protein

以新鲜牛骨为原料,探讨牛骨粉的制备工艺。选用枯草杆菌中性蛋白酶与胰蛋白酶分步水解牛骨蛋白,以水解度为特征性指标,采用L9(34)正交试验设计优化双酶水解牛骨粉的工艺条件。结果表明:牛骨粉蛋白质含量高达30.72%,约为鲜骨蛋白质含量的3倍;采用胰蛋白酶先水解,中性蛋白酶后水解的方式分步水解牛骨蛋白,其效果优于两种酶同时水解及中性蛋白酶先水解,胰蛋白酶后水解;中性蛋白酶与胰蛋白酶分步水解牛骨蛋白的最佳工艺条件:固液比1:2,起始pH8.0,胰蛋白酶2000U/g蛋白质于45℃、150r/min振荡水解3h,然后用中性蛋白酶2000U/g蛋白质于45℃以150r/min振荡水解3h,水解度达18.34%;粒径小的活性炭对酶解液脱苦、脱色及除腥效果优于粒径大的活性炭。

Fresh bovine bones were used as raw materials to prepare bone powder. Neutral protease from Bacillus subtilis and trypsin were chosen to hydrolyze bovine protein in a two-step mode. The optimal dual-enzyme hydrolysis conditions of bone proteins were explored for maximlzing degree of hydrolysis using L9（3^4） orthogonal array design. Results indicated that the content of proteins in bone powder was 30.72%, which was 3 times as high as that in fresh bovine bones. The sequential order for enzymatic hydrolysis of bone proteins exhibited an obvious effect on degree of hydrolysis of bone proteins. Sequential hydrolysis of bone proteins by trypsin first and then neutral protease exhibited a higher efficiency when compared with simultaneous dual-enzyme hydrolysis or sequential hydrolysis with neutral protease first. The optimal hydrolysis procedure of bone proteins was based on trypsin hydrolysis for 3 h on a 150 r/min shaking table with a material/liquid ratio of 1：2 and an enzyme dose of 2000 U/g protein at 45 ℃ and pH 8.0 followed by neutral protease hydrolysis for 3 h on a 150 r/min shaking table with an enzyme dose of 2000 U/g protein at 45 ℃. Under such conditions, the overall degree of hydrolysis reached 18.34%. Moreover, a better effect on debittering, decolorization and deodorization was obtained using activated carbon with smaller size.