抄録
We studied the optimum conditions for measurement of creatine kinase (EC 2.7.3.2, CK) activity in serum at 30°. Imidazole acetate buffer containing EDTA was superior for both CK and glucose-6-phosphate dehydrogenase (G6PD) activities to Bis-Tris acetate with and without EDTA.
The optimum pH of partially purified CK isoenzymes CK1, CK2, CK3 and serum CK (abundant of CK3) were 6.2-6.6, 6.2-6.6, 6.0-6.4, and 6.4-6.8, respectively.
The affinities for creatine phosphate were significantly different not only among the purified CK isoenzymes but also between the purified CK3 and serum CK (abundant of CK3).
Since approximately the same activities were obtained by several thiol compounds at the concentrations of 10 to 30 mmol/I, N-acetylcysteine was selected because of its easy handling.
The combined additions of AMP (4-5 mmol/l) and diadenosine pentaphosphate (10μmol/l) resulted in remaining of less than 5% of adenylate kinase (AK) activities and 1 to 4% suppression of CK activities.
As a coupling enzyme, G6PD from leuconostoc mescenteroides was recommended because of the low concomitant activity of glucose dehydrogenase and the stability in the CK assay reagents except for its large Km-value for glucose-6-phosphate.
