Albalá-Hurtado S, Bover-Cid S, Izquierdo-Pulido M, Veciana-Nogués M T, Vidal-Carou M C
Unitat de Nutruió i Bromatologia, Facultat de Farmàcia, Universitat de Barcelona, Spain.
J Chromatogr A. 1997 Aug 22;778(1-2):235-41. doi: 10.1016/s0021-9673(97)00299-9.
A high-performance liquid chromatography (HPLC) method to determine available lysine is proposed. Available lysine was measured by an optimised fluorodinitrobenzene method on the basis of the reactivity of the free epsilon-amino group of the lysine. The classical acid hydrolysis has been improved and shortened from the usual time of 12 h to 2 h 30 min using an oil bath. Optimal resolution and quantitation of Nn-dinitrophenyllysine was obtained with a Nova-Pak C18 column using an isocratic elution with 35% methanol and 65% 0.01 M sodium acetate buffer (pH 4.5) and a flow-rate of 1 ml/min. Satisfactory results were obtained for the reliability of the method in terms of linearity from 0.1 to 5.0 mg/l of lysine-free base, precision (R.S.D. values between 4.3% and 7.8%), recovery (91.5%) and sensitivity (detection limit of 0.02 mg/l). The proposed method has also been checked for lack of interferences from other dinitrophenyl-amino acids.
提出了一种测定有效赖氨酸的高效液相色谱（HPLC）方法。基于赖氨酸游离ε-氨基的反应性，采用优化的氟二硝基苯法测定有效赖氨酸。经典的酸水解方法得到了改进，使用油浴将通常的12小时缩短至2小时30分钟。使用Nova-Pak C18柱，以35%甲醇和65% 0.01M醋酸钠缓冲液（pH 4.5）等度洗脱，流速为1ml/min，可获得Nn-二硝基苯基赖氨酸的最佳分离度和定量结果。该方法在0.1至5.0mg/l游离赖氨酸碱的线性度、精密度（相对标准偏差值在4.3%至7.8%之间）、回收率（91.5%）和灵敏度（检测限为0.02mg/l）方面的可靠性得到了满意的结果。还对所提出的方法进行了检查，以确定是否存在来自其他二硝基苯基氨基酸的干扰。
