Combinatorial approach of LC–MS/MS and LC–TOF-MS for uncovering in vivo kinetics and biotransformation of ochratoxin A in rat

A combinatorial platform of liquid chromatography–tandem mass spectrometry (LC–MS/MS) and liquid chromatography coupled with time of flight mass spectrometry (LC–TOF-MS) has been developed to investigate the in vivo kinetics and biotransformation of ochratoxin A (OTA) in rats. The stable isotope dilution LC–MS/MS method was first validated by determining the linearity (R2 ≥ 0.9990), sensitivity (lower limit of quantitation of 0.05 ng mL−1), accuracy (83.3–108.3), precision (RSD ≤ 15.6%) and stability (≥75.0%), and was approved for the determination OTA in plasma, heart, liver, spleen, lung, kidney and brain with a run time of 7.0 min. Simultaneously, an LC–TOF-MS method could unambiguously identify the metabolites of OTA in a total run time of 14 min. The subsequent studies on kinetics and distribution after oral administration of 0.2 mg/kg b.w. OTA in rat indicated that OTA could reach a maximum value of 1932.4 ± 124.9 ng mL−1 within 5 h due to its fast absorption, and then was slowly eliminated in plasma with a half-life time (t1/2) of 75.6 ± 29.0 h. Results of tissue accumulation after a daily oral administration of 0.1 mg/kg b.w. OTA during 20 days showed that the highest concentration of OTA was observed in lung (95.9 ± 13.7 ng g−1), followed by liver (76.0 ± 9.7 ng g−1), heart (62.0 ± 4.2 ng g−1) and kidney (55.7 ± 4.7 ng g−1). Furthermore, three less toxic metabolites of OTA were clearly identified: Ochratoxin β (OTβ) and ochratoxin B (OTB) methyl ester were found in kidney and spleen, respectively, while phenylalanine was detected in heart and kidney. Thus, a possible metabolic pathway of OTA was proposed. The above achieved results justified that the application of combinatorial LC–MS/MS and LC–TOF-MS methods are valuable tools to uncover the kinetics and metabolism of OTA for the interpretation of toxicological findings in animals and extrapolation of the resulting data as reference to humans.