假交替单胞菌Pseudoalteromonas sp. AJ5 产k-卡拉胶酶的培养条件优化

Optimization of κ-carrageenase production by Pseudoalteromonas sp. AJ5

摘要:

【目的】本研究的目的是优化Pseudoalteromonas sp. AJ5菌株的培养条件使之产生高活性的胞外k-卡拉胶酶。【方法】通过富集培养技术从刺参肠道分离出一株卡拉胶降解菌AJ5，该菌株能利用卡拉胶作为惟一碳源和能源。依据形态学和生理学特征及16S rRNA基因序列分析，将该菌株鉴定为假交替单胞菌属（Pseudoalteromonas）。通过单因素试验和正交试验对Pseudoalteromonas sp. AJ5菌株产胞外k-卡拉胶酶的培养条件进行了优化。【结果】单因素试验结果表明，Pseudoalteromonas sp. AJ5菌株的最佳培养条件为250 mL三角瓶装入75 mL发酵培养基、摇床转速150 r/min、接种量7%、pH8.0。单因素试验和正交试验结果显示该菌株的最佳培养基组成为k-卡拉胶 1 g/L、牛肉膏2 g/L、 NaCl 20 g/L、K2HPO4·3H2O 1 g/L、 MgSO4·7H2O 0.5 g/L、 MnCl2· 4H2O 0.2 g/L、 FePO4 · 4H2O 0.01 g/L； 培养温度为28℃，培养时间为28 h。【结论】Pseudoalteromonas sp. AJ5菌株分泌胞外k-卡拉胶酶，在最佳培养条件下，该菌株的k-卡拉胶酶活力比优化前提高了4倍。

Abstract:

[Objectives] To optimize the culture conditions of Pseudoalteromonas sp. AJ5 for a higher production of extracellular κ-carrageenase. [Methods] A k-carrageenan-degrading bacterium AJ5, capable of utilizing k- carra-geenan as sole source of carbon and energy, was isolated from the intestine of holothurian Apostichopus japonicus by enrichment culture technique. The strain was identified as the genus Pseudoalteromonas sp. according to its morphological and physiological characterization and 16S rRNA gene analysis. Culture conditions for the bacte-rium were standardized for the maximal productivity of the extracellular k-carrageenase by the single factor and orthogonal tests. [Results] According to the single factor test, the optimal culture conditions were: 75 mL medium in 250 mL Erlenmeyer flask, shaking speed of 150 r/min, inoculum’s volume 7%, and pH 8.0. Based on the single factor and orthogonal tests the optimal medium components were: k-carrageenan (1 g/L), beef extract (2 g/L ), NaCl (20 g/L), K2HPO4·3H2O (1 g/L), MgSO4·7H2O (0.5 g/L), MnCl2·4H2O (0.2 g/L), FePO4·4H2O (0.01 g/L), with the incubation temperature and time of 28°C and 28 h. [Conclusions] Pseudoalteromonas sp. AJ5 secreted an extracellular k-carrageenase. Under the optimal culture conditions, four-fold increase in k-carrageenase activity was achieved as compared to the control.