苦碟子注射液中4 种氨基酸含量的测定

Determination of four amino acids in Kudiezi injection

关键词:  苦碟子注射液  柱前衍生化  2,4-二硝基氟苯  氨基酸  高效液相色谱法

Key words:  Kudiezi injection  pre-column  2, 4-dinitrofluorobenzene  amino acid  HPLC

摘要:

[目的] 建立同时测定苦碟子注射液中4 种氨基酸类成分(缬氨酸、异亮氨酸、亮氨酸、苯丙氨酸)含量的2,4-二硝基氟苯(DNFB)柱前衍生化法。[方法] 采用Diamonsil C₁₈(2)色谱柱(200 mm×4.6 mm,5 um);流动相A 为超纯水,B 为pH=6.40的30 mmol/L乙酸钠(含1%N,N-二甲基甲酰胺),C为乙腈,梯度洗脱。流速为1.0 mL/min,柱温25 ℃,检测波长为350 nm。[结果] 对10 批苦碟子注射液中4 种氨基酸含量进行了测定,缬氨酸、异亮氨酸、亮氨酸、苯丙氨酸分别在0.138 8~4.440、0.124 4~3.980、0.135 0~4.320、0.131 9~4.220 mg/L范围内线性关系良好(r≥0.999 4)。平均回收率分别为98.49%、100.60%、98.50%、98.56%,相对标准偏差(RSD)臆2.0%(n=6)。[结论] 该方法简便、准确,可靠,为苦碟子注射液质量控制提供参考。

Abstract:

[Objective] To establish a pre-column deriving determination method with 2, 4-dinitrofluorobenzene (DNFB) for simultaneous quantification of four amino acids (valine, isoleucine, leucine and phenylalanine) in Kudiezi injection. [Methods] The determination was performed on a Diamonsil C₁₈ (2) column (200 mm×4.6 mm, 5 um). The mobile phase consisted of ultra pure water, 30 mmol/L NaAc (containing 1% N, N-dimethylformamide, pH=6.40) and acetonitrile with gradient elution. The flow rate was 1.0 mL/min. The column temperature was set at 25 ℃and the detection wavelength was 350 nm. [Results] Four amino acids of 10 batches Kudiezi injection were quantified and showed good linear relationship in 0.138 8~4.440, 0.124 4~3.980, 0.135 0~4.320 and 0.131 9~4.220 mg/L, respectively. The mean recoveries of four essential amino acids were 98.49%, 100.60%, 98.50% and 98.56%, RSD ≤2.0% (n=6). [Conclusion] The method is simple, accurate, and reliable for quality control of the Kudiezi injection.