Abstract: The toothpaste sample was extracted with methanol by vortex oscillation and ultrasonic extraction. After centrifugation,the supernatant was taken and filtered on 0.45 μm organic microporous membrane. HPLC was applied to the determination of 3 homologues of benzalkonium chloride in the filtrate. GL Sciences CN-3 chromatographic column was used as separation column, and the mixture of acetonitrile and 0.1 mol·L^-1 ammonium acetate solution with volume ratio of 1:1 was used as mobile phase. Detection at 260 nm was adopted in the determination. Linear relationships between values of peak area and mass concentration of 3 homologues of benzalkonium chloride were kept in the same range of 5-500 mg·L^-1, with detection limits (3S/N) in the range of 0.66-1.38 mg·L^-1 and lower limits of determination (10S/N) in the range of 2.20-4.60 mg·L^-1. Tests for recovery were made by standard addition method at the concentration levels of 100, 1 000, 2 000 mg·kg^-1, giving values of recovery and RSDs (n=6) in the ranges of 97.4%-107% and 0.21%-5.2% respectively.

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