HPLC-based ATP Quantification in LV

Left ventricular apices were homogenized on ice with 0.4 mmol/L perchloric acid and precipitated with KOH. The samples were centrifuged for 10 min (3000× g) and the supernatants were injected twice onto a pre-equilibrated RP18 column (LiChroCART, Merck, Darmstadt, Germany) as previously published [31 (link)]. For ATP detection a HPLC-apparatus from Knauer (Berlin, Germany) and an UV-detector (PDA Detector 2800, Knauer, Berlin, Germany) were used. Peaks were measured at 259 nm. Standard curves were generated with 4 concentrations of ATP, (25-12.5-6.25-3.125 µg/mL). External standards and ventricular samples were measured together in one HPLC run.