Abstract: L-lactic acid is the most useful organic acid. In this study，89 strains producing lactic acid were isolated from soil，silage，hot spring water and surrounding soil. The concentration and enantiomeric purity of lactic acid produced by these strains were determined by HPLC. Based on the ability of the strains to produce L-lactic acid，three effective strains S-18，S-32 and S-44 which grew at 50℃ and 55℃ were selected. After 72 h of fermentation，75.0 g/L，81.9 g/L and 86.5 g/L of L-lactic acid were produced from 100 g/L glucose by the three strains，respectively. The enantiomeric purity of the products was 96.3%，96.6% and 97.7%，respectively. Strain S-44 was mutated by the N+ ion beam implantation，and more efficient mutants were obtained. By using the mutant strain Y-4，178.7 g/L of L-lactic acid was produced with the molar yields of 178.7% to the substrate glucose.