摘要
提出用直接提取法将含有游离态泛酸的食品(如奶粉、米粉等)中将其分离。称取样品5g,加入淀粉酶0.2g和50mL水摇匀后,在(55±5)℃的水浴中振摇酶解30min。从此溶液中定量分取相当于0.5~5.0g的样品,加入泛酸钙[13 C6、15 N]同位素内标溶液100μL,加水至20mL,加入300g·L^-1乙酸锌溶液和150g·L^-1亚铁氰化钾溶液各0.4mL作为沉淀剂,加水定容至25.0mL,混匀,静置10min后离心2min。取其上清液并用0.22μm滤膜过滤,取其滤液按仪器工作条件进行液相色谱-串联质谱法(LC-MS/MS)分析。用酶解法处理含有结合态泛酸的食品样品(谷、薯、肉、蛋、豆类及其制品1~5g,新鲜果蔬5~10g),于样品中加入pH 8.1±0.1Tris缓冲溶液10mL和水40mL,并在121℃和加压条件下水解15min,冷却,加水定容至100.0mL,过滤,取滤液10.0mL加入泛酸钙同位素内标溶液100μL,及三(羟甲基)氨基甲烷-盐酸(Tris-HCl)缓冲溶液5mL,将溶液置于冰浴中,相继加入0.08mol·L^-1碳酸钠溶液0.1mL,0.02g·mL^-1碱性磷酸酶溶液0.4mL,0.5g·L^-1鸽子肝脏提取液0.2mL和水0.4mL,混合均匀,加入甲苯1滴,在37℃反应8h以上使泛酸游离。反应完成后,加水至20mL,以下操作同直接提取法从加入沉淀剂起。在LC分离中,采用Waters BEH C18色谱柱和以不同比例的(A)1%(体积分数)甲酸溶液和(B)乙腈组成的混合液作为流动相进行梯度洗脱,按质谱条件测得泛酸的峰面积。泛酸标准曲线的线性范围为10~1500μgL^-1,其检出限(3S/N)为3.0μg·kg^-1。按标准加入法进行回收试验,测得回收率为91.0%~105%,测定值的相对标准偏差(n=6)为0.46%~3.0%。应用本方法分析了奶粉及其他食品共53种样品,所测得泛酸含量的结果与国标法(微生物法)的测定结果的相对标准偏差均小于10%,并根据t检验的结果,表明本方法的测定值与国标法测定值之间无显著差异。
Direct extraction method was proposed to separate pantothenic acid from those food samples,such as milk powder,rice flour and etc.which contain pantothenic acid in the free state.In this case,5g of the sample were taken and 0.2g of amylase to gether with 50mL of water were added,and the mixture was enzymolyzed by shaking in a water bath heated at(55±5)℃for 10min.An aliquot equivalent to 0.5-5.0g of the sample was taken,and 100μL of the isotopic calcium pantothenate internal standard solution were added,and the solution was diluted to 20mL.0.4mL each of the 2precipitants,300g·L^-1 zinc acetate solution and 150g·L^-1 potassium ferrocyanide solution,was added and extra-pure water was added to make its volume to 25.0mL.After mixing,standing for 30min and centrifuging for 2min,the supernatant was taken and filtered through 0.22μm filtering membrane.The filtrate was used for LC-MS/MS analysis under the instrumental working conditions.For food samples containing pantothenic acid in its combined states of acetyl coenzyme and acyl carriex protein,such as cereals,potato,meat,eggs and beans(taking 1-5g of sample),fresh fruits and vegetables(taking 5-10g of sample),the sample should be first hydrolyzed for 15min in 10mL of Tris-HCl buffer solution(pH 8.1±0.1)and 40mL of water at 121℃under pressure.After cooling,the solution was diluted to 100.0mL with extrapure water and filtered on a dry-filter paper.An aliquot of 10.0mL was taken,and after adding 100μL of the isotopic internal standard solution and 5mL of the Tris-HCl buffer solution,the mixture was placed into an ice-bath,and after adding 0.1mL of 0.08mol·L^-1 sodium carbonate solution,0.4mL of 0.02g·mL^-1 alkaline phosphatese solution,0.2mL of 0.5g·L^-1 pigeon liver extract and 0.4mL of extrapure water were well mixed,where one drop of toluene was added,the reaction mixture was enzymolyzed for at least 8hat 37℃to liberate pantothenic acid from its combined state to free state.After the reaction was completed,the solution was diluted to 20 mL,and the produ
作者
郑国建
解楠
徐琼
杨保刚
彭亚锋
雷涛
宁啸骏
ZHENG Guojian;XIE Nan;XU Qiong;YANG Baogang;PENG Yafeng;LEI Tao;NING Xiaojun(Shanghai Institute of Quality Inspection and Technical Research,Shanghai 200233,China)
出处
《理化检验（化学分册）》
CAS
CSCD
北大核心
2020年第3期326-331,共6页
Physical Testing and Chemical Analysis(Part B:Chemical Analysis)
基金
上海市科学技术委员会科研计划项目(18DZ2201400)。
