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| HPLC柱前衍生法测定棉籽蛋白中17种氨基酸含量 |
| Determination of 17 amino acids in cottonseed protein by pre-column derivatization coupled with high performance liquid chromatography |
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| DOI：10.12166/j.zgyz.1003-7969/2020.12.004 |
| 中文关键词: 棉籽蛋白 氨基酸 高效液相色谱 2,4-二硝基氯苯 衍生 |
| 英文关键词:cottonseed protein amino acid high performance liquid chromatography 2,4-dinitrochlorobenzene derivatization |
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| 中文摘要: |
| 建立了一种高效液相色谱检测棉籽蛋白中17种氨基酸含量的方法。采用2,4-二硝基氯苯（2, 4-dinitrochlorobenzene,CDNB）衍生，利用C18色谱柱分离，紫外检测器进行检测，检测波长360 nm，并对酸解方式、衍生方式和反应时间进行优化，针对样品前处理步骤长，影响检测精密度因素多的现状，采用倒推的方式对每个步骤进行了精密度验证，同时采用内标法定量，消除了检测过程中不确定因素对检测结果的影响，提高检测精密度。结果表明：17种氨基酸在0.004 6~0.045 7 mg/mL范围内呈良好线性关系，相关系数（R2）为0.983 2～0.999 8；精密度除蛋氨酸和胱氨酸较差外，其余氨基酸在1.71%～8.43%之间;除蛋氨酸和胱氨酸外，其余氨基酸加标回收率为9012%～109.67%。 |
| 英文摘要: |
| A method for the determination of 17 amino acids in cottonseed protein by high performance liquid chromatography (HPLC) was developed. The sample was derived with 2,4-dinitrochlorobenzene (CDNB), separated by C18 column and detected by ultraviolet detector with detection wavelength 360 nm. The acid hydrolysis method, derivatization mode and reaction time were optimized. In view of the long sample pretreatment steps and many factors affecting the detection precision, the precision of each step was validated by backward method. At the same time, the internal standard method was used to quantify, which eliminated the influence of uncertain factors on the detection results in the detection process, and improved the detection precision. The results showed that 17 kinds of amino acids showed good linear relationship in the range of 0.004 6-0.045 7 mg/mL, the correlation coefficient was 0.983 2-0.999 8; the precision and the recovery rate were 1.71%-8.43% and 90.12%-109.67% respectiveyly except methionine and cystine. |
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