Pharmacokinetic, bioavailability and tissue distribution study of MP3950, a new gastroprokinetic candidate compound, in rat using UPLC-MS/MS

Highlights

• For the first time a UPLC–MS/MS method was developed to determine racemic MP3950 in rat plasma and tissues.

• The whole analysis time was 3.0 min per sample.

• MP3950 distributed rapidly and extensively after oral administration and showed high concentration in digestive system.

• Tissue distribution of MP3950 has gender differences.

Abstract

MP3950 is being developed as a gastroprokinetic candidate compound. To illustrate the pharmacokinetic profiles, absolute bioavailability after intravenous administration and oral administration with MP3950 as well as tissue distribution in vivo, an UPLC-MS/MS approach which was rapid and selective was developed to determine MP3950 in plasma and tissue of rat. Sample pre-treatment of plasma sample was one-step protein precipitation. 0.1% formic acid containing 5 mmol/L ammonium acetate-methanol(55/45,v/v) was used for isocratic elution on a Waters ACQUITY UPLC® BEH C18 (50 mm × 2.1 mm, 1.7 μm) to achieve the separation. The analysis was performed in MRM mode via positive ESI mode. LLOQ of the method was 10 ng/mL, and the linearity up to 10,000 ng/mL. The intra-day precision (relative standard deviation, RSD) was 4.0–9.0% and the inter-day precision was 4.2–10.6%. The accuracy (relative error, RE) was −1.2–2.4%. Tissue samples were collected from the brain, heart, liver, spleen, lung, kidney, stomach, duodenum, small intestine, large intestine, appendix and skeletal muscle. The same liquid chromatographic and mass spectrometric conditions were used, and it's proven that this method was feasible to analyze the MP3950 in tissues with good precision and accuracy over the range from 10 to 5000 ng·mL⁻¹. It was found that the concentration of MP3950 is higher in digestive system. The tissue distribution, pharmacokinetic and bioavailability of MP3950 in rats were carried out by the method for the first time, which can provide enough information for the further development and investigation of MP3950.

Introduction

With the accelerating rhythm of life, people with mental pressure have the rising trend of gastrointestinal disorder incidence. Gastrointestinal motility disorders are common digestive tract diseases, including gastroesophageal reflux, functional dyspepsia, gastroparesis, and so on [[1], [2], [3], [4]]. It is seriously affecting the quality of patients' life, and has been one of the hot fields in medical research today for researching and developing a safe and effective drug to promote gastrointestinal function.

As a selective 5-HT4 receptor agonist, mosapride is one of the gastroprokinetic agents widely used in clinic [[5], [6], [7]]. Recent studies indicated that (R,S)-4-amino-5-chloro-2-ethoxy-N-[3-(4-fluorobenzylamino)-2-hydroxypropyl] benzamide (MP3950), an active metabolite of mosapride, may enhance the effect on the contraction of guinea pig ileum in vitro and display a better 5-HT₄ agonizing effect [8, 9]. The pharmacological and toxicological profiles and the in vivo effectiveness of MP3950 indicated that this compound could become an attractive and promising gastroprokinetic agent [10, 11]. As we all know, the preclinical pharmacokinetics has great important influence on the development and investigation of potential candidates with better advice for the further drug design [9]. Pharmacokinetic studies can both provide toxicological and clinical information and direct optimization of drug candidates, as a result, they play necessary parts in drug discovery and development [12]. The pharmacological effects of drugs (drug speed, intensity, duration) mainly depend on the concentration of drug in target organs, so the concentration and pharmacological effect in the organization is directly related to the study. The study of the tissue distribution of drugs is very important to improve drug efficacy and reduce toxicity and guide the formulation design. However, the method of determination of MP3950 in biological matrices hasn't been developed so far. A fast and sensitive bioanalytical method should be warranted to provide information for preclinical and clinical pharmacokinetic study of MP3950. And this study shows the possibility of administration of MP3950 in the form of racemates.

Study on metabolites of mosapride has been conducted since 1993. Four metabolites (des-p-fluorobenzyl (M-1), 5′-oxo-des-p-fluorobenzyl mosapride (M-2), 3-hydroxy des-p-fluorobenzyl (M-3) and 3-hydroxy 5′-oxo-des-p-fluorobenzyl mosapride (M-4)) were isolated from rat urine after oral administration of mosapride by thin layer chromatography (TLC) [13]. With the development of analytical techniques, pharmacokinetics, pharmacological effects, enantiomeric determination and tissue distribution of some metabolites of mosapride were carried out over the next few years [[14], [15], [16], [17]]. The identification and structure elucidation of metabolites of mosapride was investigated in rats by UPLC-MS/MS in 2009. 18 metabolites of mosapride were detected and at least 15 metabolites were reported in rat for the first time. MP3950 is one of the discovered metabolites in the study [9]. After that, some researches on MP3950 have been done by our group in the past few years. Pharmacodynamic studies showed that the racemate and the (S)-enantiomer of MP3950 possessed high efficacy of promoting gastrointestinal motility [18]. We have developed a method to separate the enantiomers of MP3950 in rat plasma by HPLC in 2016. Pharmacokinetic parameters of (R) -MP3950 and (S) -MP3950 were got individually [19]. And pharmacokinetics studies of (S)-MP3950 in normal rats and pathological status rats were studied by UPLC-MS/MS in 2017 [11].

UPLC-MS/MS were used to determine mosapride and some metabolites of it by our group in the previous studies [20, 21]. And an UPLC-MS/MS approach which could analyze MP3950 in rat plasma and tissue was developed and validated for the first time in the present study [22, 23]; And a pharmacokinetic study, bioavailability study after intravenous administration and oral administration with MP3950 and a tissue distribution study of MP3950 in rats were reached by the method.