Ohsuzu F, Bessho M, Yanagida S, Sakata N, Takayama E, Nakamura H
Department of Medicine I, National Defence Medical College, Saitama, Japan.
J Mol Cell Cardiol. 1994 Feb;26(2):203-10. doi: 10.1006/jmcc.1994.1023.
Changes in ATP and creatine phosphate levels during early (up to 150 s) global ischaemia were determined in isolated rat hearts by two chemical extraction methods (a conventional direct perchloric acid extraction and a stepwise extraction using alcohol and perchloric acid solutions) and by qualitative 31P-NMR. No difference in the ATP level hardly changed up to 40 s of ischaemia when measured by the three methods, and slightly decreased at 150 s of ischaemia. In contrast to ATP, creatine phosphate content in the normally perfused tissue was observed to be higher by the stepwise extraction (68-73 nmol/mg protein) than by direct perchloric acid extraction (55 nmol/mg protein). The creatine phosphate rapidly decreased to about 50% of normal value at 40 s of ischaemia, and the difference in the normal creatine phosphate content using the two chemical methods disappeared with the progression of ischaemia. Thus, the creatine phosphate more rapidly decreased when observed by the stepwise method than by the other two methods in this ischaemic condition. These results suggest that (1) creatine phosphate exists in an undetermined chemical state (perhaps neither in simple soluble form nor in so-called "bound" form) in rat cardiac myocytes, and (2) the stepwise extraction method is useful to measure the content of energy metabolites and to examine the intracellular chemical state in cardiac tissues.
采用两种化学提取方法（传统的直接高氯酸提取法和使用酒精及高氯酸溶液的分步提取法）以及定性³¹P-NMR，对离体大鼠心脏在早期（长达150秒）全心缺血期间ATP和磷酸肌酸水平的变化进行了测定。在缺血40秒内，用这三种方法测得的ATP水平几乎没有差异，而在缺血150秒时略有下降。与ATP不同，分步提取法测得的正常灌注组织中的磷酸肌酸含量（68 - 73 nmol/mg蛋白质）高于直接高氯酸提取法（55 nmol/mg蛋白质）。缺血40秒时，磷酸肌酸迅速降至正常值的约50%，随着缺血的进展，两种化学方法测得的正常磷酸肌酸含量差异消失。因此，在这种缺血状态下，分步提取法观察到的磷酸肌酸下降速度比其他两种方法更快。这些结果表明：（1）大鼠心肌细胞中的磷酸肌酸以一种未确定的化学状态存在（可能既不是简单的可溶形式，也不是所谓的“结合”形式）；（2）分步提取法对于测量心脏组织中的能量代谢物含量以及研究细胞内化学状态是有用的。
