Stability studies on flucloxacillin and the design of an HPLC method for the assay of flucloxacillin in capsule dosage forms

Flucloxacillin is extensively used in the treatment of various infections caused by susceptible organisms. It breaks down easily in the presence of moisture and the breakdown products are responsible for the hypersensitivity reactions in susceptible individuals. Stability studies on flucloxacillin sodium for capsule formulations were carried out. Fixed amounts of flucloxacillin sodium (250mg) were mixed with varying amounts of dried starch and dried plain carboxymethyl cellulose (plain cmc). The mixtures were put in well closed containers and kept in a room for two and half months. Iodimetry was used to monitor the amounts of flucloxacillin in the mixtures over the stated period. It was noticed after the period that the mixtures with the dried plain cmc were more stable than mixtures with dried starch. The percentage breakdown for the dried starch mixtures were 28.47, 24.67, 27.32 and 25.20% respectively for 75, 125, 150 and 250mg of dried starch respectively. The percentage breakdown of the dried plain cmc mixtures were 27.40, 22.63, 23.07 and 20.57% respectively for 75, 125, 150 and 250mg of the dried plain cmc. The breakdown process followed pseudo-first order kinetics. The rate constants for the dried starch mixtures were 0.0336, 0.0305, 0.0345 and 0.0297week-1 for 75, 125, 150 and 250mg of dried starch respectively. The rate constants for the dried plain cmc mixtures were 0.0317, 0.0291, 0.0268 and 0.0231week-1 for 75, 125, 150 and 250mg of dried plain cmc respectively. A reversed phase high performance liquid chromatography (RP- HPLC) was also designed to assay a mixture of flucloxacillin sodium, amoxicillin trihydrate, ampicillin trihydrate, benzylpenicillin sodium and phenoxymethylpenicillin sodium. The mobile phase used was 40%v/v redistilled methanol and 60%v/v0.02M potassium dihydrogen phosphate, KH2PO4 (pH = 3.70) on a Phenomenex ® LICHROSORB 10 RP-1 C18 column (250×4.60mm I.D., 5μm particle size). The flow rate was 1.00ml/min with UV detection at 225nm. Mean retention times of 3.14 ± 0.01, 4.36 ± 0.02, 8.50 ± 0.04, 13.57 ± 0.06 and 21.18 ± 0.08minutes (n=18) were recorded for amoxicillin trihydrate (AXT), ampicillin trihydrate (APT), benzylpenicillin sodium (BPS), phenoxymethylpenicillin sodium (PMS) and flucloxacillin sodium (FXS) respectively. The calibration plots for linearity gave straight lines with coefficient of correlation, r2 values of 0.9989, 0.9987, 0.9988, 0.9984 and 0.9995 for AXT, APT, BPS, PMS and FXS respectively in the concentration range of0.005% to 0.10%w/w for BPS, FXS and PMS and 0.003% to 0.06%w/w for AXT and APT. The limit of Detection (LOD) of the method were 0.002578, 0.002830, 0.004397, 0.005184 and 0.003029%w/v for AXT, APT, BPS, PMS and FXS respectively. The Limit of Quantitation (LOQ) of the method were 0.007811, 0.008576, 0.013325, 0.015708 and 0.009178%w/v for AXT, APT, BPS, PMS and FXS respectively. The method was precise with relative standard deviations (RSD) obtained for the intra-day precision/repeatability being 1.75, 1.64, 1.62, 4.03 and 1.75% and the inter-day/intermediate precision being 1.22, 2.23 1.90, 3.97 and 2.25% for AXT, APT, BPS, PMS and FXS respectively in the concentration range stated above. The method was also found to be robust since deliberate alteration of flow rate and wavelength of detection in the range of 1.00±0.20ml/min and 225±2nm respectively did not affect the precision of the method. The accuracy of the method was confirmed with mean percentage recoveries (n=3) of 99.88, 100.66, 100.30, 100.48 and 101.33%w/w at 80% concentration level, 100.57, 100.01, 100.66, 100.48 and 100.51%w/w at 100% concentration level and 99.10, 99.48, 100.79, 100.41 and 100.40% w/w at 120% concentration level obtained for AXT, APT, BPS, PMS and FXS respectively. The method can assay any of these penicillins individually either in single or combined formulations. It assays all five penicillins simultaneously and also detects breakdown products of flucloxacillin. Key words: Amoxicillin trihydrate (AXT), ampicillin trihydrate (APT), benzylpenicillin sodium (BPS), phenoxymethylpenicillin sodium (PMS) flucloxacillin sodium (FXS), dried plain cmc, dried starch, iodimetry and RP-HPLC.

Source: KNUST