HPLC-DAD/FLD测定婴幼儿配方奶粉中B族维生素及不确定度分析

Determination and uncertainty analysis of B vitamins in infant formula by HPLC-DAD/FLD

建立了高效液相色谱法同时测定婴幼儿配方奶粉中维生素B_(2)、烟酸、烟酰胺、吡哆醇、吡哆醛的方法。样品经1%甲醇的0.1%磷酸水溶液提取并沉降蛋白后,采用Thermo Hypersil GOLD^(TM)aQ(5μm,4.6 mm×250 mm)分离,以二极管阵列检测器(DAD)和荧光检测器(FLD)串联模式检测。同时构建了不确定度评定模型,考察了不确定度的来源。结果表明,维生素B_(2)、吡哆醇、吡哆醛在0.025~4.000μg/mL,烟酸、烟酰胺在0.125~20.00μg/mL范围内,标准曲线呈现良好的线性(R^(2)>0.999),加标回收率在93.20%~108.00%(n=6),相对标准偏差(RSD)<8%。当婴幼儿配方奶粉中烟酸、烟酰胺含量为4 mg/100 g时,其扩展不确定度分别为0.300 mg/100 g(k=2)、0.182 mg/100 g(k=2);当吡哆醇、吡哆醛、VB_(2)含量为800μg/100 g时,其扩展不确定度为100μg/100 g(k=2)、71.5μg/100 g(k=2)、41.9μg/100 g(k=2),其不确定度主要来源于标准溶液配制及标准曲线拟合。该方法简便、快速、灵敏、准确,可满足婴幼儿配方奶粉中B族维生素的检测要求。

A method was developed for the simultaneous determination of vitamin B_(2),niacin,nicotinamide,pyridoxine and pyridoxal in infant formula by HPLC.The sample was extracted by 0.1%phosphoric acid aqueous solution of 1%methanol and precipitated protein,separated by Thermo Hypersil GOLD^(TM)aQ(5μm,4.6 mm×250 mm)and detected using diode array detector(DAD)and fluorescence detector(FLD)in series mode.At the same time,an uncertainty evaluation model was constructed to examine the sources of uncertainty.The results showed that calibration of five vitamins were linear in the range of 0.025-4.000μg/ml for vitamin B_(2),pyridoxine,pyridoxal and 0.125-20.00μg/ml for niacin acid,nicotinamide.The standard curve presented a good linearity(R^(2)>0.999),the adding standard recovery was between 93.20%and 108.00%(n=6),and the relative standard deviation(RSD)was less than 8%.When the content of niacin and niacinamide in infant formula was 4 mg/100 g,the expanded uncertainty was 0.300 mg/100 g(k=2)and 0.182 mg/100 g(k=2);When the content of pyridoxol,pyridoxal,and VB_(2)was 800μg/100 g,the expanded uncertainty was 100μg/100 g(k=2),71.5μg/100 g(k=2)and 41.9μg/100 g(k=2).The main source of uncertain was the preparation of standard solutions and the fitting of standard curves.The method was simple,rapid,sensitive and accurate,and could meet the determination requirements of B vitamins in infant formula.