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对儿茶酚胺甲基转移酶代谢多巴酚丁胺的研究

燕茂河, Anita Pettigrew, Carolyn Myers, Jeffrey L. Blumer   

  1. 1. 山西省儿童医院, 太原 030013;
    2. 彩虹儿童医院, 克里夫兰, 俄亥俄, 美国
  • 收稿日期:1997-08-06 修回日期:1997-11-10 出版日期:1998-03-15 发布日期:1998-03-15

Study on Metabolism of Dobutamine by Human Mononu- clear Cell Catechol-O-Methyltransferase

Mao-He Yan, Anita Pettigrew, Carolyn Myers, Jeffrey L. Blumer   

  1. 1. Shanxi Provincial Children's Hospital, Taiyuan 030013;
    2. Divion of Pediatric Pharmacology and Critical Care, Rainbow Babies and Children's Hospital, Cleveland, Ohio 44106 USA
  • Received:1997-08-06 Revised:1997-11-10 Online:1998-03-15 Published:1998-03-15

摘要: 本文介绍了对多巴酚丁胺代谢的研究。代谢酶选用人类单核细胞中儿茶酚胺甲基转移酶(COMT)。方法选用高效液相色谱。结果阐明了COMT对多巴酚丁胺代谢的最优条件。COMT的反应在pH7.4磷酸盐液中进行。腺苷基蛋氨酸和多巴酚丁胺的Michaelis常数(Km)分别为0.05 μmol·L-150 μmol·L-1。至少在1小时内COMT活性随孵化时间的增加呈线性增加, 蛋白浓度选择在1.7~6 mg·ml-1, 镁离子的最佳浓度(12~18 μmol·L-1), 比以前报道的高8~10倍。方法的日间和日内相对标准偏差分别为9.7%8.9%。此法很适合对多巴酚丁胺的进一步的临床和基础研究。

关键词: 多巴酚丁胺, 儿茶酚胺甲基转移酶, 高效液相色谱

Abstract: The optimal conditions for a HPLC-EC method for measuring human mononu-clear cell catechol-O-methyltransferase (COMT) activity with dobutamine as substrate are reported. The major metabolite of dobutamine by COMT is 3-O-methyldobutamine (3-O-MD). COMT reaction should be conducted in pH 7.4 phosphate buffer. In order to saturate COMT, the concentrations of S-adenosyl-L-methionine (SAM; a methyldonor) and dobutamine should be more than 200 μmol·L-1 and 0.5 mmol·L-1 respectively. The apparent Michaelis constants for S-adenosyl-L-methionine and dobutamine are 0.05 μmol·L-1 and 50 μmol·L-1 respectively. The COMT activity increases linearly with increase of incubation time up to 1 h and with increase of protein concentrations ranged from 1.7 to 6 mg·ml-1 respectively. The optimal concentration of magnesium chloride is ten times higher than that previously reported. The betweenday relative standard deviation for 3-O-methyl dobutamine is 9.7% and that of within-day is 8.9%. This method is ideally adapted to clinical studies.

Key words: Dobutamine, COMT, HPLC

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