Sensitive LC‐MS‐MS method for the determination of tiopronin in human plasma

Abstract

A highly selective and sensitive LC-MS-MS method was developed and validated to quantify tiopronin in human plasma, using fudosteine as the internal standard (IS). L-Cysteine and 1,4-dithiothreitol (DTT) were used as the reducer and the stabilizer to release and stabilify tiopronin from a dimmer and mix forms with endogenous thiols in the treatment of plasma samples. After a simple liquid-liquid extraction with ethyl acetate in acidic condition, the post-treatment samples were analyzed on a C(18) column interfaced with a triple-quadruple tandem mass spectrometer using negative electrospray ionization. Methanol and water (40:60, v/v) were used as the isocratic mobile phase, with 0.2% formic acid and 1.0 mM tris (hydroxymethyl) aminomethane (Tris) in water. The method was validated to demonstrate the specificity, lower limit of quantification, accuracy and precision of measurements. The assay was linear over the concentration range 0.078-10 microg/mL. The correlation coefficients for the calibration curves ranged from 0.9980 to 0.9990. The intra- and inter-day precisions, calculated from quality control samples, were not more than 10.49%. The method was employed in a pharmacokinetic study after oral administration of 200 mg tiopronin tablets to 24 healthy volunteers.

Quantitative determination of tiopronin in human plasma by LC‐MS/MS without derivatization

“…The LLOQ was confirmed to be 40.0 ng/mL for TP, at which the accuracy was in the range 96.1–103.9% and the precision was below 20% (Table ). The result suggested that our method is more sensitive than some previous methods in which no derivatization was performed (Ma et al ., ; Jiang et al ., ).…”

“…Some different methods have been developed to analyze TP in biological fluid, including gas chromatography–mass spectrometry (GC‐MS; Matsuura and Takashina, ) and high‐performance liquid chromatography (HPLC) coupled with fluorescence detection (HPLC‐FLD; Kågedal et al ., ; Leroy et al ., ; Penugonda et al ., ), ultraviolet detection (HPLC‐UV; Huang et al ., ; Kusmierek and Bald, ) and mass spectrometry (LC‐MS; Matsuura et al ., ; Liu et al ., ; Ma et al ., ; Jiang et al ., ). Owing to the absence of chromophore in the structure of TP, TP has to be derivatized with a special reagent that contains chromogenesis groups in all the reported HPLC‐FLD and HPLC‐UV methods (Kågedal et al ., ; Leroy et al ., ; Penugonda et al ., ; Huang et al ., ; Kusmierek and Bald, ).…”

“…() and Jiang et al . () reported LC‐ESI‐MS methods without derivatizition to detect TP in human plasma; however, the processes of sample preparation were laborious and the mobile phase was complex.…”

Spectrophotometric Determination of Tiopronin in Pharmaceutical Samples by Phosphorus Molybdenum Blue

“…Tiopronin i.e., N-(2-mercaptopropionyl)-glycine (TP), a synthetic thiol compound, has been used as a hepatoprotective agent, an antidote to heavy metal poisoning and a radioprotective agent. It has also been successfully applied to prevent kidney injury and to treat cystinuria and rheumatoid arthritis 1,2 . However, tiopronin has a relatively high frequency of side effects, such as the loss of taste or stomach upset.…”

“…Thus, quantitative analysis of the tiopronin can be almost impossible, unless the thiol group can be stabilized in pharmaceutical sample. Up to now, several methods have been reported for the determination of tiopronin such as gas chromatographymass spectrometry (GC-MS) method 3 , chemiluminescent method 4 , LC-MS-MS methods 2 , HPLC-ESI-MS methods 1,5 , fluorimetric methods 6,7 and electrochemical oxidation by amperometric flow injection analysis 8 .…”