Extraction and Analysis of Adenosine Phosphate in Cells of Microcystis Aeruginosa

Abstract

Four methods, perchloric acid extraction, organic solvent extraction, boiling bitter salt solution extraction, and boiling bitter salt solution with ultrasonic extraction, were used for the extraction of adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine momophosphate (AMP) from the cells of Microcystis aeruginosa. The method of boiling bitter salt solution with ultrasonic extraction was chosen as the optimum method. The optimum volume of extraction solution was 2 ml for the method of boiling bitter salt solution with ultrasonic. When ATP, ADP and AMP were placed in boiling water bath, the concentration of ATP and AMP significantly changed, whereas the content of ADP slightly changed. The results demonstrate that the optimum procedure for extracting ATP, ADP and AMP from the cells was that the cells were put into boiling water bath for 10 min and then were staved with ultrasonic for 10 min in bitter salt solution as extraction solution. This method was simple and nonpoisonous. A reversed phase high-performance liquid chromatography combined with UV detection is established to determine of ATP, ADP and AMP. They were distinctly separated within 10 min. Moreover, this method has the advantages of high sensitivity, rapidity and accuracy. The recoveries of ATP, ADP and AMP were 88%–97%, 103%–107%, and 109%–115%, respectively. The standard deviation and relative standard deviation were less than 10%.