淀粉液化芽孢杆菌BS5582产蛋白酶的发酵条件优化、酶学性质分析及其在牛皮脱毛中的应用

Fermentation optimization and enzymatic property analysis of proteases produced by Bacillus amyloliquefaciens BS5582 and its application in cattle skin dehairing

该研究旨在优化1株高产蛋白酶淀粉液化芽孢杆菌BS5582的产酶条件,分析其酶学性质,并考察其在牛皮脱毛工艺中的应用效果。结果表明,BS5582菌株的较优产酶培养基组成为(g/L):玉米粉50.0,豆饼粉40.0,Na_(2)HPO_(4)·12H_(2)O 6.0,KH_(2)PO_(4)3.0,MgSO_(4)1.8,CaCl_(2)0.75;培养基初始pH 6;较优发酵条件为:发酵温度35℃,接种量10%,装液量15 mL。在较优条件下,该菌株产蛋白酶活力达到(5270.59±19.61)U/mL,是优化前的21倍。BS5582菌株所产蛋白酶的最适温度和最适pH值分别为40℃和8.5,Mn^(2+)对酶有明显激活作用,而Fe^(2+)、Fe^(3+)和苯甲基磺酰氟对酶活力有显著抑制作用。将该酶应用于牛皮脱毛,发现其具有良好的牛皮脱毛能力且胶原损伤小。相比于传统灰碱法脱毛,该蛋白酶脱毛后牛皮毛孔中无发根残留,毛发完整脱落,脱毛废水的污染指标总固体悬浮物、化学需氧量和五日生化需氧量值分别比化学脱毛法降低约87%、39%和45%,这说明BS5582菌株产蛋白酶具有潜在的清洁脱毛应用价值。

Proteases are considered as clean alternatives to replace chemicals to use in the leather dehairing process to avoid serious pollution.This study aimed to optimize the condition for protease production by Bacillus amyloliquefaciens BS5582 strain,to characterize the enzymatic properties and to evaluate its application in cattle hide dehairing.Results showed that the optimal enzyme production media by BS5582 strain were corn powder 50.0 g/L,soybean powder 40.0 g/L,Na_(2)HPO_(4)·12H_(2)O 6.0 g/L,KKH_(2)PO_(4)3.0 g/L,MgSO_(4)1.8 g/L,and CaCl_(2)0.75 g/L,with the initial pH 6.The optimal fermentation condition was as follows:The fermentation temperature was 35℃,the inoculation volume was 10%,and the loading volume was 15 mL.Under this condition,the crude enzyme activity of the enzyme solution reached(5270.59±19.61)U/mL,which was 21 folds higher than that before optimization.The optimum temperature and pH of the proteases produced by the BS5582 strain were 40℃and 8.5,respectively.Mn^(2+)had a significant activation effect on protease activity while Fe^(2+),Fe^(3+),and PMSF significantly inhibited the enzyme activity.The protease showed good cattle skin hair removal ability with little collagen damage.Compared with traditional chemical dehairing,the application of proteases had no hair root remaining in the pores of cattle skin with the falling of intact hairs.The pollution indexes of the dehairing wastewater,such as total suspended solids,chemical oxygen demand,and five-day biochemical oxygen demand,were reduced by 87%,39%,and 45%,respectively.This indicated that the proteases produced by B.amyloliquefaciens BS5582 were of great potential for application in the leather industry.