响应面法优化含活性肽CC34毕赤酵母重组菌冻干营养保护剂的研究

doi:10.16431/j.cnki.1671-7236.2022.10.011

摘要/Abstract

摘要： 【目的】利用响应面分析法优化含活性肽CC34毕赤酵母重组菌的冻干营养保护剂组合配方。【方法】通过单因素试验,筛选脱脂乳粉、海藻糖、山梨醇和谷氨酸钠提高毕赤酵母重组菌存活率的最佳范围;利用Box-Behnken法建立毕赤酵母重组菌存活率回归方程,分析3种保护剂组合对菌体存活率的效应关系。【结果】脱脂乳粉浓度为10%时毕赤酵母重组菌冻干存活率最高,达到51.23%;海藻糖浓度为15%时冻干存活率最高,达到67.50%;山梨醇浓度为20%时冻干存活率最高,达到52.19%;谷氨酸钠浓度为3%时冻干存活率最高,达到36.92%。通过响应面法获得含活性肽CC34毕赤酵母重组菌冻干保护剂组合中脱脂乳粉、海藻糖、山梨醇的浓度分别为10.75%、15.70%、20.19%,通过最优模型预测毕赤酵母重组菌的存活率为77.00%,应用此最优保护剂组合进行冻干试验,菌体冻干存活率为78.98%。【结论】本试验通过响应面法优化了毕赤酵母重组菌复合冻干保护剂配方,在脱脂乳粉浓度为10.75%、海藻糖浓度为15.70%、山梨醇浓度为20.19%条件下,可获得较高的菌体冻干存活率(78.98%)。该复合冻干保护剂可用于含活性肽CC34毕赤酵母重组菌的保存和后续试验研究。

关键词: 响应面; 冷冻干燥; 冻干保护剂; 毕赤酵母重组菌

Abstract: 【Objective】 Response surface method was used to optimize the formulation of lyophilized nutritional protectants for Pichia pastoris recombinant bacteria containing active peptide CC34.【Method】 The optimum range of skimmed milk powder,trehalose,sorbitol and monosodium glutamate to improve the survival rate of recombinant Pichia pastoris was screened by single factor test. The regression equation of the survival rate of recombinant Pichia pastoris was established by Box-Behnken method,and the effect relationship of three protectant combinations on the survival rate was analyzed.【Result】 When the concentration of skimmed milk powder was 10%,the freeze-drying survival rate of recombinant Pichia pastoris was the highest,reaching 51.23%. When the concentration of trehalose was 15%,the freeze-drying survival rate was the highest,reaching 67.50%. When the concentration of sorbitol was 20%,the freeze-drying survival rate was the highest,reaching 52.19%.And when the concentration of sodium glutamate was 3%,the freeze-drying survival rate was the highest,reaching 36.92%.The concentrations of skimmed milk powder,trehalose and sorbitol in the freeze-drying protectant combination of Pichia pastoris recombinant strain obtained by response surface were 10.75%,15.70% and 20.19%,respectively. The survival rate of recombinant Pichia pastoris was predicted to be 77.00% by the optimal model.The freeze-drying test with the optimal combination of protectants showed that the survival rate of the cell was 78.98%.【Conclusion】 In this experiment, the formulation of compound freeze-drying protectant for recombinant Pichia pastoris was optimized by response surface methodology. Under the conditions of skim milk powder concentration of 10.75%, trehalose concentration of 15.70% and sorbitol concentration of 20.19%, a higher survival rate (78.98%) of recombinant Pichia pastoris could be obtained. The compound freeze-drying protectant could be used for the preservation of recombinant Pichia pastoris containing active peptide CC34 and subsequent experimental research.

Key words: response surface; freeze drying; lyoprotectant; recombinant Pichia pastoris

中图分类号:

S816.3

李爽, 杜家华, 白志辉, 曲可, 苏苗, 姜宁, 张爱忠. 响应面法优化含活性肽CC34毕赤酵母重组菌冻干营养保护剂的研究[J]. 中国畜牧兽医, 2022, 49(10): 3791-3799.

LI Shuang, DU Jiahua, BAI Zhihui, QU Ke, SU Miao, JIANG Ning, ZHANG Aizhong. Optimization of Lyophilized Combination for Pichia pastoris Containing Active Peptide CC34 Recombinant with Response Surface Methodology[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(10): 3791-3799.

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