Optimization of a Stability-Indicating HPLC Method for the Simultaneous Determination of Rifampicin, Isoniazid, and Pyrazinamide in a Fixed-Dose Combination using Artificial Neural Networks

Abstract

The aim of this study is to develop and optimize a simple and reliable high-performance liquid chromatography (HPLC) method for the simultaneous determination of rifampicin (RIF), isoniazid (INH), and pyrazinamide (PZA) in a fixed-dose combination. The method is developed and optimized using an artificial neural network (ANN) for data modeling. Retention times under different experimental conditions (solvent, buffer type, and pH) and using four different column types (referred to as the input and testing data) are used to train, validate, and test the ANN model. The developed model is then used to maximize HPLC performance by optimizing separation. The sensitivity of the separation (retention time) to the changes in column type, concentration, and type of solvent and buffer in the mobile phase are investigated. Acetonitrile (ACN) as a solvent and tetrabutylammonium hydroxide (tBAH), used to adjust pH, have the greatest influence on the chromatographic separation of PZA and INH and are used for the final optimization. The best separation and reasonably short retention times are produced on the micro-bondapak C18, 4.6 x 250-mm column, 10 microm/125 A using ACN-tBAH (42.5:57.5, v/v) (0.0002M) as the mobile phase, and optimized at a final pH of 3.10.

Simultaneous determination of isoniazid, rifampicin, levofloxacin in mouse tissues and plasma by high performance liquid chromatography–tandem mass spectrometry

“…Due to the instability of INH and RFP in aqueous solutions, sample handling of the reported methods usually requires an addition of antioxidant to prevent oxidation during the following extraction, concentration and reconstitution steps [10,11], which was rather complex, labor-and time-consuming. Moreover, in our preliminary experiments it was found that the antioxidant (ascorbic acid) added to samples would result in MS/MS signal suppression of the analytes.…”

“…Over the years, many quantification methods for the individual determination of the three anti-tuberculosis drugs in biological matrices have been reported, but the very different polarities of the three drugs lead to a great difficulty in analyzing them by a single-run chromatographic separation. However, there have been a few reported assay methods for simultaneous quantification of two of the three drugs in different biological matrices recently [8][9][10][11][12]. A C18 column (250 mm × 4.6 mm or 50 mm × 2.0 mm) was used to separate compounds in most of these methods.…”

Electrochemical sensor for Isoniazid based on the glassy carbon electrode modified with reduced graphene oxide–Au nanomaterials

“…A safe, reliable and economical method has to be made to detect the concentration of these drugs simultaneously either in pure form, in pharmaceutical preparations or in biological fluids. At present, various methods have been developed for the determination of INZ, such as high-performance liquid chromatography [2][3][4][5], gas chromatography [6], capillary electrophoresis [7], chemiluminescence [8,9], localized surface plasmon resonance [10], ultraviolet-visible spectrophotometry [11,12], fluorimetry [13] and electrochemical methods [14][15][16][17][18][19][20]. Among these methods, electrochemical methods offer advantages including high sensitivity, simplicity and reproducibility [15,[21][22][23][24][25][26][27].…”

Nano Formulation Analysis: Analytical Method Development of Isoniazid and Simultaneous Estimation of Anti-Tubercular Drugs Isoniazid and Rifampicin by Rp-HPLC

“…Various other methods also have been described for determination and quantification of INH, such as H-point standard addition method [7], selective adsorption using a piezoelectric sensor [8], voltammetric method [9], amperometric method [10], chromatographic methods [11][12][13][14] (HPLC, GC and HPTLC), titrimetric methods [15], and chemiluminescence [16]. In addition, several (HPLC) methods for the simultaneous determination of rifampicin (RIF), INH, and pyrazinamide (PZA) in a fixed-dose combination [17] simultaneous analysis of RIF, INH and PZA in 0.1 M hydrochloric acid dissolution medium and in simulated gastric fluid [18], quantitative determination of INH in plasma, brain, liver and kidney samples and in solid lipid nanoparticles [19]. RIF and INH in combined dosage form estimation by UV spectrophotometric method [20,21] stabilityindicating high performance liquid chromatographic simultaneous determination of RIF, INH and PZA in human plasma [22] Most of the methods are time-consuming (long retention times [RT]) along with gradient methods.…”