Simultaneous Determination of Paracetamol and Methocarbamol in Human Plasma By HPLC Using UV Detection with Time Programming: Application to Pharmacokinetic Study

Abstract

Background:

A combination of methocarbamol (MET) and paracetamol (PAR) is a widely used treatment approach. It provides complementary modes of action for treatment of pain associated with muscle spasm. The aim of this work was to develop and validate a new sensitive and reproducible isocratic reversed phase HPLC-UV detection method for simultaneous determination of MET and PAR in human plasma for the routine use in a therapeutic drug monitoring and pharmacokinetic laboratories.

Methods:

A simple HPLC assay was developed and validated for the simultaneous determination of the above-mentioned drugs in small samples of human plasma (0.25 mL). After protein precipitation with methanol, satisfactory separation was achieved on a Hypersil^® BDS C18 column (250 mm×4.6 mm, 5 μm) using a mobile phase comprising 20 mM sodium dihydrogen phosphate buffer (pH=3) and methanol at a ratio of 80:20, v/v; the elution was isocratic at ambient temperature with a flow rate of 1.2 ml/min. The UV detector was programmed at 254 nm for 7.0 min to measure PAR and IS and at 272 nm for the subsequent 3 min to measure MET.

Results:

Linearity was demonstrated over the concentration range from 0.02 to 20 µg/ml (mean R ² =0.9998, n=10). The observed within- and between-day assay precision ranged from 1.11 to 9.4 and 2.46 to 10.0% for PAR and MET, respectively; whereas, accuracy varied between 95.2–101% and 93.9–102.2% for PAR and MET, respectively. Mean drug recovery was 99.8 for PAR and 99.0% for MET. PAR and MET were stable in frozen plasma over a period of 3 months at −80°C.

Conclusions:

The validated method was applied successfully to a bioequivalence study of PAR/MET (500/400 mg) fixed dose combination tablet in healthy volunteers (n=24).

Key words

paracetamol - methocarbamol - HPLC-UV with time ­programming - pharmacokinetics - bioequivalence

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