保加利亚乳杆菌LJJ自溶的影响因素分析

Analysis of factors influencing the autolysis of Lactobacillus delbrueckii subsp. bulgaricus LJJ

考察了不同生长阶段的保加利亚乳杆菌LJJ在不同温度及酸度环境中的自溶度变化，不同浓度SDS和Triton X-100对菌体自溶度和溶菌酶活性的影响，并采用扫描电镜观察菌体自溶中的形态变化，以确定LJJ自溶的最适条件并初步探讨溶菌酶在自溶中的作用。结果表明，处于对数生长期的LJJ菌体最易发生自溶。LJJ自溶度随环境温度（4—50℃）的升高，温育时间的延长而相应增大。LJJ自溶的最适温度为50℃，最适pH值范围为6．0—8．0。0．50g／LSDS对LJJ自溶和溶茵酶的抑制率分别为99．6％和97．4％。1．00g／LTritonX-100分别将LJJ的自溶度和溶茵酶活性提升了20．7％和39．5％。扫描电镜观察表明，自溶细胞与正常细胞相比，细胞壁出现孔洞，细胞完整性遭到破坏而0．50g／LSDS可抑制LJJ细胞的破坏。因此，环境pH、温度、温育时间、SDS浓度、TritonX．100浓度及菌体生长阶段可通过溶菌酶活性的作用显著影响UJ的自溶，为调控乳酸菌自溶提供了初步理论基础。

The optimal conditions for Lactobacillus delbrueckii subsp, bulgaricus LJJ autolysis and the effect of lysozyme on autolysis were determined through analyzing the influence of various factors such as temperature, pH of TE buffer, incubation time, cell growth phase, SDS and Triton X-100 concentration. Autolysis was performed on Lactobacillus delbrueckii subsp, bulgaricus LJJ from different growth stages and detecting the autolysis rate and lysozyme activity variation under various levels of SDS and Triton X-100 concentration. The morphological differences of the normal LJJ cells, the autolysis cells in TE buffer after 24h incubation at 37℃ and in TE buffer with 0. 50 g/L SDS were observed by Scanning Electron Microscope （SEM）. The results showed that LJJ had the optimum autolysis temperature of 50℃ and the optimum autolysis pH value in the range of 6.0 -8. 0. Furthermore, with the extension of incubation time, autolysis rate of LJJ was increasing gradually and the rate increase slowed down significantly after 24h incubation. LJJ ceils of the exponential phase showed the maximum autolysis rate, while LJJ cells of the steady growth period gave the minimum autolysis rate. The presence of SDS significantly inhibited the autolysis of LJJ and lysozyme activity. Autolysis of LJJ and lysozyme activity was inhibited by 99.6% and 97.4% with adding 0. 50 g/L SDS respectively. In contrast, the use of 1.00 g/L Triton X-100 improved the activityof lysozyme by 20. 7% and 39.5% respectively. The results of SEM observation revealed that the autolysis cells in TE buffer after 24h incubation at 37℃ compared to the normal cells had holes appearing in the cell wall, cell integrity was destroyed, and intracellular content dissolved out, while the autolysis ceils in TE buffer with 0.50 g/L SDS just became atrophic and the cell surface was crude, but cell wall still kept its integrity. Environmental temperature, pH value, incubation time, cell growth phase, SDS concentration and Triton X-100 concentration can affect the autolysis of LJJ significantly and be used for the regulation of the autolysis of LJJ through controlling the influence factors of autolysins under certain conditions.