Bioanalytical Method Validation of Metformin Hydrochloride in Human Plasma by HPLC-UV for Preliminary Population-Based Pharmacokinetic Modeling Study

Dimas Adhi Pradana; Erna Kristin; Akhmad Kharis Nugroho; Dwi Aris Agung Nugrahaningsih; Mustofa Mustofa; Ari Wibowo

doi:10.22146/ijc.77903

Bioanalytical Method Validation of Metformin Hydrochloride in Human Plasma by HPLC-UV for Preliminary Population-Based Pharmacokinetic Modeling Study

https://doi.org/10.22146/ijc.77903

Dimas Adhi Pradana^(1*), Erna Kristin⁽²⁾, Akhmad Kharis Nugroho⁽³⁾, Dwi Aris Agung Nugrahaningsih⁽⁴⁾, Mustofa Mustofa⁽⁵⁾, Ari Wibowo⁽⁶⁾

(1) Department of Pharmacy, Universitas Islam Indonesia, Jl. Kaliurang km. 14, Yogyakarta 55584, Indonesia; Department of Pharmacology and Therapy, Faculty of Medicine, Public Health, and Nursing, Universitas Gadjah Mada, Jl. Farmako, Sekip Utara, Yogyakarta 55281, Indonesia
(2) Department of Pharmacology and Therapy, Faculty of Medicine, Public Health, and Nursing, Universitas Gadjah Mada, Jl. Farmako, Sekip Utara, Yogyakarta 55281, Indonesia
(3) Faculty of Pharmacy, Universitas Gadjah Mada, Sekip Utara, Yogyakarta 55281, Indonesia
(4) Department of Pharmacology and Therapy, Faculty of Medicine, Public Health, and Nursing, Universitas Gadjah Mada, Jl. Farmako, Sekip Utara, Yogyakarta 55281, Indonesia
(5) Department of Pharmacology and Therapy, Faculty of Medicine, Public Health, and Nursing, Universitas Gadjah Mada, Jl. Farmako, Sekip Utara, Yogyakarta 55281, Indonesia
(6) Department of Pharmacy, Universitas Islam Indonesia, Jl. Kaliurang km. 14, Yogyakarta 55584, Indonesia
(*) Corresponding Author

Abstract

This study aims to validate the method for measuring metformin hydrochloride plasma concentrations using High-Performance Liquid Chromatography (HPLC). This research performed chromatography on a 250 mm 4.6 mm 5 µm purosphere® Star RP-18 column at ambient temperature with a UV detector system at 233 nm. The mobile phase components were 70% phosphate buffer (KH₂PO₄) (10 mM), sodium dodecyl sulfate (0.3 mM), and 30% acetonitrile. It was pumped at an isocratic flow rate of 1.2 mL/min. Metformin HCl and ranitidine HCl (internal standard) were extracted using acetonitrile. The calibration curve was linear (R² = 0.9998) in the 0.18–6 µg/mL concentration range. The lower limit of quantification (LLOQ) was 0.18 µg/mL. For intraday accuracy and precision, the percent difference and the coefficient of variation were less than 4 and 7%, and for inter-day were lower than 8 and 6%. The recovery average was 100.96%. The short-term plasma stability test was stable at 24 h at ambient temperature, and the long-term stability test was steady for 30 d at −20 °C. It was also stable after three freeze-thaw cycles. The method meets selectivity, sensitivity, linearity, accuracy, precision, recovery, carryover, and stability requirements and can be applied to population-based pharmacokinetic modeling.

Keywords

human plasma; HPLC-UV; metformin HCl; pharmacokinetic; validation

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DOI: https://doi.org/10.22146/ijc.77903