HPLC法测定人凝血酶原复合物中氨基酸含量

目的: 建立用高效液相色谱法测定人凝血酶原复合物中氨基酸含量。 方法: 采用6-氨基喹啉-N-羟基琥珀酰亚氨基氨基甲酸酯(AQC)为衍生剂,与氨基酸柱前衍生后,用Agilent 1200高效液相色谱仪,AccQ·Tag C₁₈柱(waters 150 mm×3.9 mm,4 μm),以水Eluent(醋酸盐-磷酸盐缓冲液)稀释液和乙腈进行梯度洗脱,检测波长为248 nm,柱温37℃,进样量10 μL。 结果: 各氨基酸在32 min内测定完毕,回收率为99.3%~100.6%。RSD均小于1.5%。 结论: 本法分离度好,快速、简便,可作为产品的质量控制方法。

Objective: To establish an HPLC method for determination of amino acids in human prothrombin complex. Method: Each kind of amino acids reacted with 6-aminoquinolyl-N-hydroxysuccinimdyl-carbamate(AQC) respectively, and their corresponding derivatives were analyzed and quantified by the Agilent 1200 HPLC using AccQ-Tag C₁₈ column (waters 150 mm× 3.9 mm, 4 μm) for gradient elution, with the mobile phase of water, eluent diluent (acetic acid salt-phosphate buffer, and acetonitrile at a wavelength of 248 nm by UV-detector, and the column temperature was set at 37℃, the sample volume was 10 μL. Results: The amino acids were completely determined in 32 minutes, and the recoveries were 99.3%-100.6%, RSD less than 1.5%. Conclusion: The method is simple, accurate and suitable for the determination of amino acids in human prothrombin complex.

1 ChP(中国药典).2010.VoL Ⅲ(三部):Appendix(附录) 41
2 NI Wei-fang(倪维芳),JIN Ou(金瓯),ZHANG Jing-song(张劲松). HPLC determination of amino acids in compound amino acid capsules (HPLC法测定复合氨基酸胶囊中氨基酸的含量). Chin J Pharm Anal(药物分析杂志),2004,24(4):402
3 QU Qi-shu(瞿其曙),TANG Xiao-qing(汤晓庆),HU Xiao-ya(胡效亚),et al.Applications of pre-column derivatization for amino acids analysis(柱前衍生化法在氨基酸分析测定中的应用).Prog Chem(化学进展),2006,18(6):689
4 YU Hong(于泓),MOU Shi-fen(牟世芬).Method development for amino acid analysis(氨基酸分析方法的研究进展).Anal Chem(分析化学),2005,33(3):398
5 SONG Zhi-feng(宋志峰),WANG Li(王丽),JI Feng(纪锋),et al.Rapid determination of amino acids in feedstufs by high performance liquid chromatography using internal standard method with online pre-column derivatization(柱前在线衍生-反相高效液相色谱内标法快速测定饲料中氨基酸).Anal Chem(分析化学),2007,35(1):25
6 CHEN Zhi-wei(陈志维),LI Yu-cui(黎玉翠),BAO Mei-jia(暴梅佳)),et al. Determination of glycine and taurine in Pulvis Fellis Suis conjugated cholic acid by pre-column derivation on HPLC(柱前衍生化高效液相色谱法测定猪胆粉结合胆酸中甘氨酸和牛磺酸的含量).Chin J Pharm Anal(药物分析杂志),2010,30(2):288
7 ZHENG Lu-xia(郑璐侠),SHAO Hong(邵泓),CHEN Gang(陈钢),et al.Simultaneous determination of 13 kinds of amino acids and taurine by two internal standard HPLC with pre-column derivatization (柱前衍生RP-HPLC内标法同时检测13种氨基酸和牛磺酸的研究). Chin J Pharm (中国医药工业杂志),2008,39(8):610
8 WANG Xiu-zhong(王秀中),WANG Qing-qing (王清清),SONG Hai-feng (宋海峰),et al. Application progress of derivatization technology in amino acids analysis(衍生化技术在氨基酸分析中的应用进展).Chin J Pharm Anal(药物分析杂志),2010,30(6):1162
9 ZHOU Yuan-hua(周远华),XIN Ni(辛妮),HU Yu-zhu(胡育筑).HPLC with precolumn derivatization determination of amino acid composition of oxytocin(柱前衍生高效液相色谱法测定缩宫素的氨基酸组成).Chin J Pharm Anal(药物分析杂志),2011,31(8):1541
10 Xu ZR,Lan Y,Fan XF,et al.Automated sampling system for the analysis of amino acids using microfluidic capillary electrophoresis.Talanta,2009,78(2):448
11 Carratù B,Boniglia C,Giammarioli S,et al.Free amino acids in botanicals and botanical preparations.J Food Sci,2008,73(5):C323