高产EPS乳酸片球菌的航天育种及其EPS性能研究

张孟雨; 李尧; 彭嘉屹; 陈禹豪; 曾凤婷; 钟青萍

doi:10.13386/j.issn1002-0306.2022110108

高产EPS乳酸片球菌的航天育种及其EPS性能研究

Space Breeding of Pediococcus acidilactici with High Yield of Exopolysaccharide and the Study on Its EPS Performance

摘要

摘要: 本文旨在对乳酸片球菌（Pediococcus acidilactici）进行航天诱变，筛选高产胞外多糖（Exopolysaccharide，EPS）的诱变菌株，并探究其产EPS的功能特性。通过测定EPS产量和遗传稳定性，筛选出一株性能稳定并高产EPS的L21-49菌株，比较了原始菌株及突变菌株的自聚性和疏水性，以及对酸、胆盐及人工模拟胃、肠液的耐受性，并分析其产EPS的抗生物被膜、抗氧化活性和对α-淀粉酶、α-葡萄糖苷酶的抑制效果。结果表明，菌株经航天诱变后，EPS产量为196.23 mg/L，比原始菌株L21产量提高了26.54%；该菌株的自聚性和疏水性良好；体外耐受性较好，在pH为2、3的条件下其活菌数可维持在10⁷ CFU/mL；在胆盐浓度为4.00 g/L时，其活菌数可保持在10⁷ CFU/mL；经过人工模拟胃肠液处理，该菌株活菌数可维持在10⁶ CFU/mL，其所产EPS具有良好的体外抗氧化能力：浓度为8.0 mg/mL时，对DPPH自由基、OH自由基、ABTS⁺自由基和\rmO_2^- 自由基的清除率分别为91.75%、38.44%、54.71%、58.84%；对金黄色葡萄球菌（Staphylococcus aureus）、单增李斯特菌（Listeria monocytogenes）、大肠埃希氏菌（Escherichia coli）和铜绿假单胞菌（Pseudomonas aeruginosa）生物被膜均有一定抑制作用，抑制率分别为40.04%、60.04%、50.59%和33.68%；能够抑制α-淀粉酶和α-葡萄糖苷酶活力，抑制率分别为23.52%、51.15%。本研究探究航天诱变对乳酸菌益生特性及产胞外多糖能力的影响，为乳酸菌及其胞外多糖的研究和开发提供参考依据。

Abstract: Pediococcus acidilactici was mutated by space mutagenesis technology, and the strains with high production of exopolysaccharide (EPS) were screened. The functional characteristics of the EPS were also investigated. EPS production and genetic stability of the strains were measured, and a stable mutant strain L21-49 with high production of EPS was selected. Meanwhile, the self-aggregation and hydrophobicity of the mutant strain and the original strain were investigated, and the tolerances of the strains to acid, bile salts, and artificial simulated gastrointestinal fluid were compared. The anti-biofilm, anti-oxidant, and the inhibitory activities on α-amylase and α-glucosidase of EPS were also analyzed. The results showed that the EPS yield of the mutant strain was 196.23 mg/L, which was 26.54% higher than that of the original strain L21. The strain showed good self-aggregation, hydrophobicity and tolerance in vitro. The viable cell number could be maintained at 10⁷ CFU/mL after being cultured in pH2.0 and 3.0 for 3 h. When treated with 4.00 g/L bile salt for 24 h, the number of viable cells were 10⁷ CFU/mL. The viable count could be maintained at 10⁶ CFU/mL after the treatment of the artificial simulated gastrointestinal fluid. The EPS presented good antioxidant capacity in vitro. When the concentration of EPS was 8.0 mg/mL, the clearance rates on DPPH, OH, ABTS⁺ and \rmO_2^- were 91.75%, 38.44%, 54.71% and 58.84%, respectively. The inhibition rates of L21-49 EPS on the biofilms of Staphylococcus aureus, Listeria monocytogenes, Escherichia coli and Pseudomonas aeruginosa were 40.04%, 60.04%, 50.59% and 33.68%, respectively. It inhibited the activities of α-amylase and α-glucosidase by 23.52% and 51.15%, respectively. In this study, the effects of space mutagenesis on the probiotic characteristics and EPS production of lactic acid bacteria were investigated, so as to provide reference for the research and development of lactic acid bacteria and the EPS.