Abstract: In this paper,the content of vitamin B6 in rice flour samples was determined by microbiological method,and the uncertainty of measurement was analyzed from several aspects.The results showed that,the relative synthetic standard uncertainty was 0.01393,the expanded uncertainty was 0.1653,and the result was (5.932 0.1653)μg/100 g.The main uncertainty component affecting the determination results of vitamin B6 is the capacity difference within micro pipette tips.