There is a close relationship between amino acid metabolism and aminoglycoside antibiotic biosynthesis.Nine kinds of amino acids were added during the fermentation of Streptomyces hygroscopicus 5008 to investigate effects of amino acids on biosynthesis of validamycin A(Val-A).Among these nine kinds of amino acids,seven kinds of them could promote Val-A production.The addition of isoleucine was found to promote Val-A production most to 16.76 g/L which increased by 49% compared to the control.Besides,the addition of methionine showed inhibition on Val-A biosynthesis.Compared to the control,Val-A productivity in methionine addition fermentation decreased by 25% to 8.47 g/L.To further understand the impact of amino acids on Val-A biosynthesis,a series of analysis on protein accumulation,sugar utilization and activities of key enzymes in carbon metabolism which are related to Val-A precursor synthesis,were done.The results revealed addition of amino acids could affect activities of key enzymes in carbon metabolism and redirect their flux which benefits to accumulation of Val-A precursor and stimulates biosynthesis of Val-A.