INTEGRATED BIOANALYTICAL METHOD VALIDATION, PHARMACOKINETICS, AND METABOLITE CHARACTERIZATION OF 19‑MONOAMINOTHIAZOLE (19MAT): A CORRELATIVE IN‑SILICO AND IN‑VIVO STUDY USING LC–MS/MS
DOI:
https://doi.org/10.22159/ijap.2026v18i3.58083Keywords:
LC-MS/MS, Pharmacokinetics, 19MAT, In silico, Biotransformer 3.0, SMARTCyp 3.0, Metabolite identification and Absolute bioavailabilityAbstract
Objective: This study is mainly focused on the development and validation of a robust liquid chromatography–tandem mass spectrometry (LC–MS/MS) method to accurately evaluate the pharmacokinetics (PK) and metabolite profile of 19‑Monoaminothiazole (19MAT), a newly synthesized compound showing promising antiviral activity against dengue virus (DENV). The work additionally integrated in‑silico metabolite prediction tools with experimental in‑vivo metabolite identification data to comprehensively understand the metabolic fate of 19MAT.
Methods: Triple‑quadrupole LC–MS/MS method was developed and validated as per ICH M10 (International council for harmonization of technical requirements for pharmaceuticals for human use) guidelines for its selectivity, linearity (1.27–1270 ng/mL), accuracy/precision, recovery, matrix effect, stability in rat plasma to quantify 19MAT to assess the PK after IV (Intravenous) and PO (Per oral) administration. Waters Xterra RP® C18 (150 mm × 4.6 mm, 5 μm) column was used in reverse phase separation of analyte and internal standard. Pharmacokinetics was assessed in Sprague Dawley rats (n=3 per dose group). BioTransformer 3.0 and SMARTCyp3.0 were used to predict metabolites, and a targeted LC–MS/MS strategy was used to identify the metabolites in plasma, urine, and feces.
Results: The assay showed linear response with r² > 0.99, recovery 48–63% with acceptable matrix factors, precision ≤~11% RSD (Relative standard deviation), and stability up to 45 days (−20/−70 °C). 19MAT exhibited rapid oral absorption (Tmax ≈ 0.42 h), moderate half‑life (PO: 4.20 h), high Vd (IV: ~1977 mL/kg), and bioavailability of 23.82%. Seven metabolites were identified via plasma, urine, and feces involving hydroxylation/dihydroxylation, O-dealkylation, and glucuronidation.
Conclusion: The integrated workflow establishes validated quantitation approach, appropriately identified pharmacokinetic properties, and maps metabolic pathways for 19MAT, supporting preclinical optimization.
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