Simultaneous Determination of Isoniazid, Pyrazinamide, Rifampicin and Acetylisoniazid in Human Plasma by High-Performance Liquid Chromatography

Abstract

A sensitive and accurate high-performance liquid-chromatography method was developed and validated for the simultaneous determination of pyrazinamide (PZA), isoniazid (INH), rifampicin (RFP) and acetylisoniazid (AcINH) in human plasma. Separation was performed on a Max-RP C12 column using gradient elution and a flow-rate program. The mobile phase was methanol-acetonitrile-buffer (20 mM of heptanesulfonic acid sodium, pH 2.5) with a ratio of 10:8:82 (v/v/v) at the initial phase. All calibration curves had good linearity (r 2 > 0.99) between the test ranges. The intra-and inter-day precision was less than 8.8% in good accuracy (<15%). The limit of detection with a signal-to-noise (S/N) of 3 was 0.014, 0.009, 0.023 and 0.054 µg mL -1 for PZA, AcINH, INH and RFP, respectively. The method was selective, sensitive and reliable, and is a good alternative for routine therapeutic drug monitoring of the four compounds during the treatment of tuberculosis patients.

Assessing Drug-Excipient Interactions in the Formulation of Isoniazid Tablets

“…These agents were selected considering literature studies on separation of tuberculostatic compounds. 10,11 The mobile phase pH was adjusted to 2.5 for the anionic pairing with HS. A slight increase was observed in the resolution and retention time for INH and INAAM when compared to C18 column without the ion pairing mechanism.…”

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

“…Organic and aqueous phases are the two components which are used as mobile phase [3]. As per "World Health Organization" reservation, INH is one the front line agent for the treatment of tuberculosis and INH pertains to the category of nucleoside reverse transcriptase inhibitor [4], with aqueous solubility of 140 mg/ml [5]. To determine polar compounds various analytical methods exists specifically for INH in literature [6].…”

A Sensitive HPLC Method for Determination of Isoniazid in Rat Plasma, Brain, Liver and Kidney

“…Poor retention of polar compounds has been reported earlier on non polar reverse phase HPLC columns earlier [15], however, numerous methods for determination of polar compounds using reverse phase columns specific to isoniazid exists in the literature [16][17][18][19]. Establishment of a reproducible analytical method is always prerogative step before the development of a pharmaceutical product.…”

“…Gradient methods to detect INH in urine upto 1400 ng/mL was reported in 2002 [18]. In addition, methods to detect INH in the presence of its metabolite (acetyl isoniazid), pyrazinamide, rifampicin and ethambutol are also reported in literature [17]. A gradient method using Methanol-Acetonitrile-Buffer (20 mM of heptanesulfonic acid sodium, pH 2.5) as the mobile phase and C-12/18 column chemistry was able to quantify INH at as low as 100 ng/mL concentration [17].…”

“…In addition, methods to detect INH in the presence of its metabolite (acetyl isoniazid), pyrazinamide, rifampicin and ethambutol are also reported in literature [17]. A gradient method using Methanol-Acetonitrile-Buffer (20 mM of heptanesulfonic acid sodium, pH 2.5) as the mobile phase and C-12/18 column chemistry was able to quantify INH at as low as 100 ng/mL concentration [17]. A USP isocratic method with sodium phosphate buffer (pH 6.8)/acetonitrile as the mobile phase has reportedly been used for determination of INH in presence of pyrazinamide, ethambutol and rifampicin in plasma, brain, liver and kidney with LOQ of 200 ng/mL [21], the method validation was however, not been reported in the literature except the LOD and LOQ.…”