Different spectrophotometric and TLC-densitometric methods for determination of pyrazinamide in presence of its impurity

Abstract

In this search, three selective spectrophotometric and TLC-densitometric methods have been developed and validated for quantitative determination of Pyrazinamide (PYN) and its impurity, Pyrazine-2-carboxylic acid (PYA). The proposed methods are third derivative spectrophotometric (Method I), first derivative of ratio spectra spectrophotometric (Method II), mean centering of ratio spectra spectrophotometric (MCR) (Method III) and TLC-densitometric (Method IV). In method (I); ³D amplitudes at 276.2 and 274.6 nm were measured and used for determination of PYN and PYA, respectively. In method (II); PYN was determined by measuring the ¹DD peak amplitude at 225.8 nm using 30 µg mL⁻¹ of PYA as a divisor; on the other hand PYA was determined by measuring this amplitude at 245.2 nm using 18 µg mL⁻¹ of PYN as a divisor. For method (III); the amplitudes of the mean centered ratio spectra at 268.4 and 268.8 nm were used for PYN and PYA, respectively. On the other hand, the forth method is TLC-densitometric method at which the chromatographic separation was achieved using silica gel 60 F₂₅₄ TLC plates and mixture of methylene chloride: methanol: ammonia solution (7:3:0.1, by volume) as a developing system followed by UV-scanning at 275 nm.

The proposed methods were successfully applied for determination of the PYN in its pharmaceutical formulation. Also they were statistically compared with the reported method using student’s-t and F-tests and there was no significant difference between them regarding both accuracy and precision.