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目的建立1种同时测定骨瓜提取物注射剂中15种氨基酸含量的RP-HPLC法。方法以异硫氰酸苯酯(PITC)为柱前衍生试剂对样品中15种氨基酸进行衍生化,采用Ocean C18(250 mm×4.6 mm,5μm)色谱柱,流动相A为水-乙腈(体积比1∶4),流动相B为浓度0.1 mol·L-1乙酸钠-乙腈(体积比97∶3),梯度洗脱,流速为1.0 m L·min-1,柱温为25℃,检测波长为254 nm,进样量为10μL。用内标法计算15种氨基酸的含量。结果 15种氨基酸质量浓度在1.24~160.72 mg·L-1内与峰面积呈良好的线性关系(r=0.997 7~1.000 0),15种氨基酸的平均加样回收率在97.7~102.4%内,加样回收率的RSD值在1.7%~3.3%内。结论该方法准确、可靠、灵敏,可以用于同时测定骨瓜提取物注射剂中15种氨基酸的含量。
OBJECTIVE To establish a RP-HPLC method for the simultaneous determination of 15 amino acids in injection of bone pumpkin extract. Methods Fifteen kinds of amino acids in samples were derivatized with phenylisothiocyanate (PITC) as pre-column derivatization reagent. The chromatographic column was Ocean C18 (250 mm × 4.6 mm, 5 μm). The mobile phase A was water-acetonitrile The mobile phase B was eluted with a gradient of 0.1 mol·L-1 sodium acetate-acetonitrile (97: 3 by volume) at a flow rate of 1.0 m L · min-1 with a column temperature of 25 ° C The wavelength is 254 nm and the injection volume is 10 μL. Calculate the content of 15 amino acids by internal standard method. Results The 15 amino acids showed a good linear relationship with the peak area in the range of 1.24-160.72 mg · L-1 (r = 0.997 7-1.000 0). The average recoveries of 15 amino acids were in the range of 97.7- 102.4% The RSD of recovery was within 1.7% ~ 3.3%. Conclusion The method is accurate, reliable and sensitive and can be used for the simultaneous determination of 15 kinds of amino acids in the injection of bone pumpkin extract.