深色有隔内生真菌不同培养时间对胞外聚合物合成的影响

Effects of different incubation time periods on synthesis of extracellular polymeric substances by dark septate endophytes

摘要:

【目的】探索不同培养时间下深色有隔内生真菌(CGMCC 17463)合成胞外聚合物(extracellular polymeric substance, EPS)的产量及组成变化。【方法】通过摇瓶培养深色有隔内生真菌(dark septate endophytes, DSE)，分析不同培养时间合成的EPS产量及应用的差异。【结果】摇瓶发酵试验表明，第4-12天，DSE生长趋于对数期和稳定期，EPS合成速率较快，在第12天产量达到1.41 g/L，之后EPS的合成速率逐渐降低。EPS组分分析结果表明，在相同质量的EPS中，第12天的胞外多糖含量较高，随着DSE生长进入稳定期至衰亡期，菌丝裂解，EPS中蛋白含量显著提高。官能团分析结果表明，不同培养时间的EPS中官能团种类并未发生变化，仅存在官能团含量上的改变。扫描电子显微镜和粒径分析结果表明，随着培养时间的增加，EPS的结构逐渐发生改变，EPS中分子粒径小于5 μm所占比例逐渐提高，分子粒径大于100 μm所占比例逐渐降低。另外，EPS的生物活性测定结果表明，不同培养时间合成的EPS均具有清除氧自由基和保水的能力，并且这种能力受EPS组分相对丰度变化的影响。【结论】第12天为提取胞外多糖含量相对较高的EPS最佳时机，第24天为提取蛋白含量相对较高的EPS最佳时机，这为EPS在环境复杂的矿山生态中应用奠定了基础。

Abstract:

[Objective] To explore the changes in the yield and composition of extracellular polymeric substances (EPS) of Alternaria sp. CGMCC 17463, a strain of dark septate endophyte (DSE), cultured for different time periods. [Methods] We conducted the shake flask experiment to compare the yield, structure, composition, and activity of EPS synthesized by a DSE strain cultured for different time periods. [Results] From day 4 to 12, the growth of the DSE strain entered the logarithmic and stationary phases. During this period, the EPS synthesis rate was high, with the yield reaching 1.41 g/L on day 12. Afterwards, the EPS synthesis rate gradually decreased. The component analysis revealed that the extracellular polysaccharide content was the highest on day 12 in the EPS samples of equal mass. As the growth of DSE continued and entered the decline phase, mycelial lysis occurred, significantly increasing the protein content in the EPS. Functional group analysis showed that as the incubation time was extended, the functional groups in the EPS presented changes only in the content but not species. The results of scanning electron microscopy and particle size analysis showed that the EPS composition gradually changed with the increase in incubation time. Specifically, the EPS components with the particle size smaller than 5 μm presented increased volume percentage, while those with the particle size larger than 100 μm showed gradually decreased volume percentage. Furthermore, the EPS possessed the ability to scavenge oxygen free radicals and retain water, which were significantly influenced by the changes in EPS composition. [Conclusion] The day 12 marks the optimal time point for the production of EPS with high polysaccharide content, while the day 24 marks the optimal time point for the production of EPS with high protein content. This result establishes a foundation for the application of EPS in the complex eco-environment of mines.