Development and experimental application of an HPLC procedure for the determination of capsaicin and dihydrocapsaicin in serum samples from human subjects

Total capsaicins were extracted from 2 mL aliquots of serum or plasma using methyl-isobutyl ketone, evaporation of the extract to dryness and reconstitution with 200 µL of acetonitrile. The HPLC mobile phase was 40:60 water:acetonitrile pumped at 0.5ml/min through an Phenomenex Luna C18s column. The absorbance of the eluent was monitored at 205 nm. Standardisation used a known mixture of pure capsaicin and dihydrocapsaicin. Accuracies were 98.9% and 100.6% for capsaicin and dihydrocapsaicin respectively. Inter batch reproducibility for both was 15%. The limits of detection were 2.6 and 3.8 ng/mL for capsaicin and dihydrocapsaicin respectively. Analyses of sera obtained previously from human subjects who had eaten chilli containing meals showed that in those that absorbed capsaicins (N=30) then the median, mean and SD of their serum capsaicin were : 13.4 ng/mL, 18.9 ng/mL and 16.3 ng/mL. The corresponding data for those sera (N=13) that had measurable levels of dihydrocapsaicin were : 6.9 ng/mL, 7.5 ng/mL and 3.6 ng/mL. This procedure was deemed suitable for use in prospective studies of the metabolism of orally ingested chilli.