建立了免疫亲和柱净化-高效液相色谱法测定乳粉中生物素的方法。该方法样品处理为样品用磷酸盐缓冲液溶解后，样液通过生物素免疫亲和柱净化、甲醇洗脱液洗脱、浓缩富集后直接测定。采用紫外检测器，波长为215 nm；C18色谱柱，柱温为35 ℃，流动相为50 mmol/L磷酸缓冲盐，PH=3.5:甲醇(85:15,v/v)，流速为1.0 mL／min。标准曲线回归方程y=343.2+20.797x，相关系数r=0.9958；回收率为90.7%；RSD%为2.62。本方法检出限为1.0 μg/kg。该法较国标方法具有样品处理简单方便，灵敏度高，重现性好，分析时间短等优点，可以满足调制乳粉及婴幼儿配方乳粉中生物素含量的测定。

The Immunoaffinity column purification - high performance liquid chromatographic method was developed in this paper for the determination of biotin in milk powder. Sample was dissolved with phosphate buffer and purified through biotin immunoaffinity column with ethanol, and concentrated to a direct determination after enrichment. UV detector wavelength was of 215 nm and C18 column was used at 35 ° C. Mobile phase is 50 mmol/L phosphate buffer salt (pH 3.5)-methanol (85:15, v/v) at a flow rate of 1.0mL/min. Standard curve regression equation was y=343.2+220.797x with a correlation coefficient (r) being of 0.9958. The recovery was 90.7% and RSD was of 2.62%. The method detection limit was 1.0 μg/kg. Compared to the national standard method, the method had the advantages of easy sample handling, high sensitivity, good reproducibility, short analysis time to meet modulation milk powder and infant formula milk powder biotin content determination.