A validated LC–MS/MS determination method for the illegal food additive rhodamine B: Applications of a pharmacokinetic study in rats.

Rhodamine B is an illegal and potentially carcinogenic food dye. The aim of this study was to develop a convenient, rapid, and sensitive UHPLC-MS/MS method for pharmacokinetic studies in rats. Rat plasma samples were deproteinized with acetonitrile and separated by UHPLC on a reverse-phase C18e column (100 mm × 2.1 mm, 2 μm) using a mobile phase consisting of methanol—5 mM ammonium acetate (90:10, v/v). Detection was performed using a triple quadrupole tandem mass spectrometer in the selected reaction monitoring mode at [M] + ion m / z 443.39 → 399.28 for rhodamine B and [M+H] + ion m / z 253.17 → 238.02 for 5-methoxyflavone as the internal standard. This method was specific and produced linear results over a concentration range of 0.5–100 ng/mL, with a lower limit of quantitation of 0.5 ng/mL. All validation parameters, including the inter-day, intra-day, matrix effect, recovery, and stability in rat plasma, were acceptable according to the biological method validation guidelines developed by the FDA (2001). This method was successfully applied to a pharmacokinetic study in rats; oral administration of 1 mg/kg of rhodamine B yielded a time to maximum concentration ( T max ) of 1.3 ± 0.4 h and an elimination half-life of 8.8 ± 1.4 h, with a clearance of 229.7 ± 19.4 mL/h/kg. These pharmacokinetic results provide a constructive contribution to our understanding of the absorption mechanism of rhodamine B and support additional food safety evaluations. [ABSTRACT FROM AUTHOR]