Improvements of luciferin-luciferase methodology for determination of adenylate energy charge ratio of marine samples

Abstract

Application of adenylate energy charge (EC) measurements in marine ecological studies requires high accuracy and precision. We have examined the luciferin-luciferase method for EC determination and suggest the following methodological improvements. Replacement of the commonly used potassium phosphate buffer with potassium chloride in the assay medium prevents problems due to precipitation of magnesium phosphates. Interference from ADP and AMP in crude firefly extract can be eliminated by their enzymatic decomposition with apyrase and AMP-deaminase. Impure nucleotide standards are perhaps more common than currently realized, and can lead to inaccurate estimates of EC. The adenylate energy charge ratio (EC = [ATP + %