Abstract: A new method for the simultaneous determination of riboflavin andpyridoxine by synchronous spectrofluorimetry is described。The measurement was carriedout in a buffer solution of pH 7 with Δλ= 58 nm for synchronous scanning。Under theexperimental conditions two peaks appeared at 526 nm and 389 nm in the obtainedsynchronous fluorescence spectrum ,corresponding to riboflavin and pyridoxine ,respective-ly. This method was proved to be simple and specific. The fluorescence relative intensityis linearly related with concentration over the range of 0 ～1μg/ml and 0 ～1.5 μg/ml for riboflavin and pyridoxine, respectively. The detection limit was 0.5 ng/ml for riboflavinand 1 ng/ml for pyridoxine.The relative errors of the measurements were less than 5% forartifical samples. The proposed method has been applied to the analyses of riboflavin and pyridoxine inthree kinds of commercial multivitamin preparations.The obtained synchronous spectra ofmultivitamin preparations coincided with those of the mixed standard solution ofriboflavin and pyridoxine. Recoveries of 93.5～105.7% were obtained.