HPLC柱前衍生化法测定发酵虫草制剂中总氨基酸的含量

目的：建立发酵虫草制剂中总氨基酸的含量测定方法，同时评价各发酵虫草制剂的质量。方法：采用Zorbax C₁₈色谱柱（250 mm×4.6 mm，5 μm），以乙酸钠缓冲液（A）-80%乙腈水溶液（B）为流动相进行梯度洗脱（0～20 min，100% A→97% A；20～24 min，97% A→88% A；24～30 min，88% A→78% A；30～50 min，78% A→68% A；50～50.5 min，68% A→0% A；50.5～60 min，0% A），流速为1.0 mL·min^-1，柱温为40℃，检测波长254 nm，运用外标法测定发酵虫草制剂中总氨基酸的含量。结果：各发酵虫草制剂的质量评价，对于独家生产的品种（如金水宝胶囊、百令胶囊和心肝宝胶囊），心肝宝胶囊中总氨基酸含量为199.3～224.9 mg·g^-1；金水宝胶囊中总氨基酸含量为224.0～281.7 mg·g^-1；百令胶囊中总氨基酸含量为294.0～346.7 mg·g^-1，样品各批间一致性较好，可以控制总氨基酸含量结合羟脯氨酸（指标成分）占总氨基酸的含量来控制制剂质量。而对于多厂家生产的发酵虫草制剂（如宁心宝胶囊和至灵胶囊），由于生产工艺的不同和原料来源不同，总氨基酸含量差异较大，宁心宝胶囊中总氨基酸含量为234.9～337.1 mg·g^-1，至灵胶囊总氨基酸含量为241.1～356.1 mg·g^-1。结论：经方法学验证，本文方法可用于测定发酵虫草制剂的氨基酸含量，也可用于评价各发酵虫草制剂的质量。

Objective: To establish a method for the determination of amino acid in fermented Cor-dyceps preparations and quality assessment of fermented Cordyceps preparations by HPLC with precolumn derivatization. Methods: The HPLC analysis was conducted on a Zorbax C₁₈ column(250 mm×4.6 mm,5 μm) with sodium acetate buffer (A) -80% acetonitrile(B) as mobile phase at a flow rate of 1.0 mL·min^-1 by gradient elution(0-20 min,100%A→97%A;20-24 min,97%A→88%A;24-30 min,88%A→78%A;30-50 min,78%A→68%A;50-50.5 min,68%A→0%A;50.5-60 min,0%A). The column temperature was maintained at 40℃,and the detection wavelength was 254 nm. External standard method was adopted for determination of amino acid in fermented Cordyceps preparations. Results: The quality of fermented Cordyceps preparation was comprehensively assessed. The total content of amino acid were 199.3-224.9 mg·g^-1 in Xinganbao capsules,224.0-281.7 mg·g^-1 in Jinshuibao capsules and 294.0-346.7 mg·g^-1 in Bailing capsules,respectively. It is suitable to control the quality of Jinshuibao capsules,Bailing capsules and Xinganbao capsules by controlling the total contents of amino acid combined with the contents of hydroxyproline in the total amino acid. The total content of amino acid was different due to the different production process and different sources of fermented powder for Ningxinbao capsules and Zhiling capsules. The total contents of amino acid were 234.9-337.1 mg·g^-1 in Ningxinbao capsules and 241.1-356.1 mg·g^-1 in Zhiling capsules. Conclusion: By methodology verification,the established method can be used for the determination of the amino acid content in fermented Cordyceps preparations,and can be used to assess fermented Cordyceps preparations.