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食品研究与开发:2019,40(6):161-165
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肝素亲和柱在乳铁蛋白检测中的应用优化
(1.上海市农业科学院庄行综合试验站,上海201415;2.沈阳市动物疫病预防控制中心,辽宁沈阳110034;3.上海德诺产品检测有限公司,上海200436)
Application of Heparin Affinity Column in the Detection of Lactoferrin
(1.Shanghai Academy of Agricultural Sciences Zhuanghang Comprehensive Experiment Station,Shanghai 201415,China;2.Shenyang Animal Disease Prevention and Control Center,Shenyang 110034,Liaoning,China;3.Denuo Product Testing Service CO.,Ltd.,Shanghai 200436,China)
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投稿时间:2018-08-02    
中文摘要: 乳铁蛋白是一种极具营养价值的活性糖蛋白,广泛应用到婴幼儿乳粉中,目前国内外未建立乳铁蛋白测定相关的标准。将肝素亲和柱(HiTrapTMHeparin HP柱)或相当者应用到乳铁蛋白检测前处理的净化过程中,建立一种高效、便捷的食品中乳铁蛋白测定的分析方法。试样中的乳铁蛋白经磷酸氢二钠溶液(0.20 mol/L,pH=8.00±0.5)提取,提取液经肝素亲和柱净化后,用磷酸氢二钠溶液淋洗,磷酸氢二钠-氯化钠溶液洗脱,并定容至3.0 mL。采用反相高效液相色谱柱分离,以紫外或者二极管阵列检测器于280 nm处检测,外标法定量。该方法系首次提出将肝素亲和柱应用到乳铁蛋白检测前处理过程中。该检测方法适合检测机构或乳品企业日常样品中乳铁蛋白的检测。
Abstract:Lactoferrin is a kind of active glycoprotein with great nutritional value,which is widely used in infant milk powder.At present,there is no relevant standard for the determination of lactoferrin at home and abroad.Heparin affinity column(HiTrapTMHeparin HP column)or equivalent were applied to the purification process of lactoferrin before pretreatment.An efficient and convenient method for the determination of lactoferrin in foods was established.The lactoferrin in the sample was extracted by Na2HPO4buffer solution(0.20 mol/L,pH=8.00±0.5).The extract was purified by heparin affinity column,washed with Na2HPO4buffer solution,and Na2HPO4-NaCl buffer solution was eluted and the final volume to 3.0 mL.High performance liquid chromatography(HPLC)was used with ultraviolet absorption detector(UV)or diode array detector(DAD)at 280 nm,and the external standard method was quantified.It was the first time to put forward the idea that heparin affinity column was applied to the purification of lactoferrin before detection.The improved method was suitable for detection of lactoferrin in detection mechanism or dairy enterprises.
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