Acta Chimica Sinica ›› 2001, Vol. 59 ›› Issue (5): 788-792. Previous Articles     Next Articles

Original Articles

哺乳动物细胞是所得多肽复合酶CAD催化反应产物的 HPLC分析

王 来来;胡霄雪;辛嘉英;崔俊儒;杨树本;朱利民   

  1. 中国科学院兰州化学物理研究所.兰州(730000);中国科学院羰基合成与选择国 家重点实验室;英国北伦敦大学生物和应用科学系
  • 出版日期:2001-05-15 发布日期:2001-05-15

HPLCseparation of the reaction mixture in mammalian cell catalyzed by the multienzyme

Wang lailai;Hu Xiaoxue;Xin Jiaying;Cui Junru;Yang Shuben;Zhu Limin   

  1. Lanzhou Inst Chem Phys, CAS.Lanzhou(730000)
  • Online:2001-05-15 Published:2001-05-15

The product of the first three steps of de novo pathway of pyremidine biosynthesis in mammalian cell catalyzed by carbamoyl phosphate synthetase- asparate transcarbamoylase - dihyroorotase are adenosine diphosphate (ADP) and dihydroorotate (DHO. ). And one of reactant is adenosine tiphosphate(ATP). A HPLC method for separation of ADP, ATP and DHO was developed. A Zarbox column (150mm×4.6mmI.D.) was used. The mobile phase were 0.01mol·L^-1 potassium dihydrogen phosphate buffer(pH4.7) and 0.5mol·L^-1potassium dihydrogen phosphate (pH4.6). The gradient was programmed at 0~5min, (φ=1, volume ratio)0.01mol ·L^-1KH~2PO~4 then 5~50 min, (φ=1,volume ratio) 0.01molL^- 1KH~2PO~4 to (φ=1,volume ratio) 0.5molL^-1KH~2PO~4. The flow rate was 0.7 mL·min^-1. The optimum deteation wavelenght and the sequence of ATP, ADP and DHO peaks were detected. The working cursor was plotted and then the corresponding concentration of ATP, ADP and DHO in the catalytic reaction were calculated.


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