Quantitation of adenine nucleotides in equine colonic mucosal tissue usinghigh performance liquid chromatography

The objectives were to use high performance liquid chromatography (HPLC) to validate an established method for adenine nucleotide separation in equine colonic mucosal tissue, to determine the inherent variability in the tissu e and extraction method, and to determine the stability of ATP, ADP, and AM P in the tissue with time. Equine colonic mucosal tissue obtained from a si ngle horse was immediately submersed in liquid nitrogen, and stored at -70 degreesC. Samples were lyophilized, extracted, and separated by HPLC. The l imit of quantitation was 0.05 mug/mL. The coefficient of variation for the instrument was less than 10% for all nucleotides measured. When the tissue was not homogenized prior to sampling, there were significant differences i n adenine nucleotide content between samples. However, when the tissue was homogenized prior to analysis, these differences were no longer significant . There was no significant decrease in ATP, ADP, or AMP content over a 54-d ay analysis period.