金振口服液氨基酸特征指纹图谱及29种氨基酸同时定量的质量控制方法研究  

Quality control of Jinzhen Oral Liquid based on amino acids fingerprint and simultaneous determination of 29 amino acids

作　　者：李海波[1] 杨群 区伟珍[3] 王团结[1] 曹亮[1] 王振中[1] 姚新生[2] 于洋[2] 肖伟[1] LI Hai-bo;YANG Qun;QU Wei-zhen;WANG Tuan-jie;CAO Liang;WANG Zhen-zhong;YAO Xinsheng;YU Yang;XIAO Wei(State Key Laboratory of New-tech for Chinese Medicine Pharmaceutical Process,Kanion Pharmaceutical Co.,Ltd.,Lianyungang 222001,China;Institute of Traditional Chinese Medicine and Natural Products,College of Pharmacy,Jinan University,Guangzhou 510632,China;South China Agricultural University,Guangzhou 510642,China;Shenyang Pharmaceutical University,Shenyang 110016,China)

机构地区：[1]江苏康缘药业股份有限公司,中药制药过程新技术国家重点实验室,江苏连云港222001 [2]暨南大学中药及天然药物研究所,广东广州51063 [3]华南农业大学,广东广州510642 [4]沈阳药科大学,辽宁沈阳110016

出　　处：《中草药》2020年第23期5972-5979,共8页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金重点项目(81630097);现代中药创新集群与数字制药技术平台“重大新药创制”科技重大专项(2013ZX09402203)。

摘　　要：目的氨基酸自动分析仪建立金振口服液(JOL)氨基酸特征指纹图谱及29种氨基酸(磷酸丝氨酸、牛磺酸、天冬氨酸、苏氨酸、丝氨酸、天冬酰胺、谷氨酸、谷氨酰胺、肌氨酸、甘氨酸、丙氨酸、瓜氨酸、α-氨基丁酸、缬氨酸、胱氨酸、异亮氨酸、亮氨酸、酪氨酸、苯丙氨酸、β-丙氨酸、β-氨基丁酸、γ-氨基丁酸、鸟氨酸、赖氨酸、组氨酸、3-甲基组氨酸、精氨酸、羟脯氨酸、脯氨酸)同时定量的分析方法。方法采用日立855-4507型离子交换色谱柱(60mm×4.6mm,3μm);检测波长分别为570、440 nm;流动洗脱泵体积流量0.35 mL/min;衍生泵体积流量0.30 mL/min;反应柱温135℃;以柠檬酸缓冲溶液为流动相进行梯度洗脱,建立JOL氨基酸特征指纹图谱,对29个主要特征峰进行化学成分指认,运用中药指纹图谱相似度软件对16批市售制剂进行相似度评价,同时对29种氨基酸进行含量测定。结果建立了JOL氨基酸特征指纹图谱结合29种氨基酸同时定量的方法;16批市售制剂的指纹图谱与对照指纹图谱的相似度值在0.989~1.000;29种定量成分线性关系良好(R2=1.000),且平均回收率为95.71%~101.87%;16批市售制剂29种氨基酸的质量浓度分别为磷酸丝氨酸15.4572~29.3624μg/mL、牛磺酸64.4234~114.2386μg/mL、天冬氨酸19.0564~32.5490μg/mL、苏氨酸6.704 8~7.841 8μg/mL、丝氨酸22.609 4~39.382 8μg/mL、天冬酰胺61.134 0~115.456 0μg/mL、谷氨酸32.254 6~63.127 2μg/mL、谷氨酰胺5.2238~8.9532μg/mL、肌氨酸4.0810~44.0076μg/mL、甘氨酸12.4030~23.5164μg/mL、丙氨酸33.876 8~44.257 4μg/mL、瓜氨酸3.514 2~9.881 0μg/mL、α-氨基丁酸1.126 4~2.287 8μg/mL、缬氨酸11.584 6~15.469 0μg/mL、胱氨酸1.660 2~4.041 8μg/mL、异亮氨酸4.087 8~5.469 2μg/mL、亮氨酸8.295 2~11.724 0μg/mL、酪氨酸7.492 6~10.761 2μg/mL、苯丙氨酸4.856 4~9.248 0μg/mL、β-丙氨酸2.309 4~6.782 6μg/mL、β-氨基丁酸0.175 0~14.790 6μg/mL、γ-氨基�Objective To establish the amino acids fingerprint of Jinzhen Oral Liquid(JOL) and simultaneous determination of 29 amino acids(P-Ser, Tau, Asp, Thr, Ser, Asn, Glu, Gln, Sar, Gly, Ala, Cit, α-ABA, Val, Cys, Ile, Leu, Tyr, Phe, β-Ala, β-AiBA, γ-ABA, Orn, Lys, His, 3-Mehis, Arg, Hypro, Pro) by automatic amino acid analyzer. Methods The separation was developed on HITACHI HPLC PACKED column(60 mm × 4.6 mm, 3 μm) by gradient elution with citric acid buffer solution at 570 and 440 nm, reaction column temperature was maintained at 115 ℃ and the carrier gas flow rate was 0.35 mL/min and flow rate of derivative pump was 0.30 mL/min. An amino acid fingerprint of JOL was set up, and 16 batches of JOL were evaluated by similarity assay. Furthermore, the contents of the main 29 amino acids were determined. Results The amino acid fingerprint of JOL was established with good separation, and 29 amino acids were determined simultaneously. The similarity of 16 batches of JOL was about 0.989 to 1.000. Moreover, good linear relationships were found(R^2 = 1.000), and the average recovery rates were 95.71%—101.87%, the content range of quantitative components in 16 batches of JOL was as following: P-Ser 15.457 2—29.362 4 μg/mL, Tau 64.423 4—114.238 6 μg/mL, Asp 19.056 4—32.549 0 μg/mL, Thr 6.704 8—7.841 8 μg/mL, Ser 22.609 4—39.382 8 μg/mL, Asn 61.134 0—115.456 0 μg/mL, Glu 32.254 6—63.127 2 μg/mL, Gln 5.223 8—8.953 2 μg/mL, Sar 4.081 0—44.007 6 μg/mL, Gly 12.403 0—23.516 4 μg/mL, Ala 33.876 8—44.257 4 μg/mL, Cit 3.514 2—9.881 0 μg/mL, α-ABA 1.126 4—2.287 8 μg/mL, Val 11.584 6—15.469 0 μg/mL, Cys 1.660 2—4.041 8 μg/mL, Ile 4.087 8—5.469 2 μg/mL, Leu 8.295 2—11.724 0 μg/mL, Tyr 7.492 6—10.761 2 μg/mL, Phe 4.856 4—9.248 0 μg/mL, β-Ala 2.309 4—6.782 6 μg/mL, β-AiBA 0.175 0—14.790 6 μg/mL, γ-ABA 17.654 4—26.621 8 μg/mL, Orn 42.338 4—58.172 6 μg/mL, Lys 12.994 9—28.498 0 μg/mL, His 1.802 6—4.067 4 μg/mL, 3-Mehis 2.608 4—4.384 0 μg/mL, Arg 107.141 6

关 键 词：金振口服液 氨基酸自动分析仪 氨基酸特征指纹图谱 磷酸丝氨酸 牛磺酸 天冬氨酸 苏氨酸 丝氨酸 天冬酰胺 谷氨酸 谷氨酰胺 肌氨酸 甘氨酸 丙氨酸 瓜氨酸 α-氨基丁酸 缬氨酸 胱氨酸 异亮氨酸 亮氨酸 酪氨酸 苯丙氨酸 Β-丙氨酸 Β-氨基丁酸 γ-氨基丁酸 鸟氨酸 赖氨酸 组氨酸 3-甲基组氨酸 精氨酸 羟脯氨酸 脯氨酸 

分 类 号：R286.02[医药卫生—中药学]