The methylxanthines caffeine and its metabolites theophylline, paraxanthine and theobromine, are stimulants and toxic to dogs. Consequently, residue limits, rather than screening limits are required for sensitivity control. This study aimed to characterise the pharmacokinetics of caffeine and its metabolites in greyhounds and propose scientifically justified residue limits.
Six greyhounds were administered a single 100 mg oral dose of caffeine (No-Doz®). Plasma and urine samples were collected over five days.
Plasma (0.5 mL) or urine (1 mL) was subject to liquid-liquid extraction (LLE) then analysed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) with caffeine-d3 and theophylline-d6 as internal standards for quantification. The limit of quantification was estimated to be 5 ng/mL (caffeine), 1 ng/mL (theophylline), 2.5 ng/mL (paraxanthine) and 25 ng/mL (theobromine) in plasma and urine from validation that also assessed specificity, accuracy, precision, recovery and matrix effects.
A non-compartmental analysis was used for the determination of plasma pharmacokinetic parameters for caffeine. The caffeine plasma concentrations rapidly reach peak concentrations of ~4,000 ng/mL within 1–2 hours and declined with a mean terminal half-life of 6.9 hours followed by a single-phase decline. Urine concentrations peaked at ~7,000 ng/mL by 6 hours. Metabolites theobromine, theophylline, and paraxanthine were also quantified in plasma and urine.
Clifford (1987) identified a physiological response threshold of 2,000 ng/mL plasma in greyhounds. Therefore, this value could be used as the minimum plasma concentration to affect performance and an irrelevant plasma concentration (IPC) of 200 ng/mL in a population of greyhounds could be postulated by applying the safety factor of 10 for the variability in the pharmacokinetics and pharmacodynamics. However, regional limits may need to reflect local factors, such as dietary inputs, to facilitate harmonisation in the long term.