Surrogate reference standards in quantitative high performance liquid chromatography; a case study of the analysis of Piroxicam Capsules and Metronidazole Tablets

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2012
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A simple, rapid, accurate and sensitive High Performance Liquid Chromatography (HPLC) method was developed for quantitative analysis of Piroxicam and Metronidazole in Piroxicam capsules and Metronidazole tablets using surrogate reference standards with UV detection. Metronidazole, Metformin and Ascorbic acid pure samples were used as surrogate reference standards for the analysis of Piroxicam. The chromatography parameters were stainless steel ODS C18 Phenomenex 250 x 4.6mm column particle size. The isocratic mobile phase was methanol:phosphate buffer pH 6.2 (50:50; v/v) at a flow rate of at 1.0 mLmin-1. The determinations were performed using UV-Vis detector set at 254 nm. Mean retention times (minutes) of 6.83±0.065, 4.07±0.058, 3.42±0.049 and 2.56±0.055 were recorded for Piroxicam, Metronidazole, Metformin and Ascorbic acid respectively. A constant, K, was determined for Piroxicam using each of the surrogate reference standards. The respective K values obtained when Metronidazole, Metformin and Ascorbic acid were used as surrogate reference standards were 1.9411± 0.004, 1.6200 ± 0.006 and 0.4569 ± 0.006. Ascorbic acid and Para aminophenol pure samples were used as surrogate reference standards for the analysis of Metronidazole. The chromatography parameters were stainless steel ODS C18 Phenomenex 250 x 4.6mm column particle size. The isocratic mobile phase was methanol:phosphate buffer pH 6.2 (50:50; v/v) at a flow rate of at 1.5 mLmin-1. The determinations were performed using UV-Vis detector set at 254 nm. Mean retention times (minutes) of 4.24±0.014, 1.80±0.012 and 2.82±0.015 were recorded for Metronidazole, Ascorbic acid and Para aminophenol respectively. A constant, K, was determined for Metronidazole using each of the surrogate reference standards. The respective K values obtained when Ascorbic acid and Para aminophenol were used as surrogate reference standards were 0.6763 ± 0.004 and 1.1229 ± 0.016. The results obtained when the v methods were used for the analysis of the Piroxicam capsules and Metronidazole tablets were comparable to standard methods of analysis stated in the BP and USP respectively. The method showed adequate precision, with a relative standard deviation (RSD) less than 2% in each case. The HPLC methods developed in this study showed specificity and selectivity with linearity in the working range and good precision and accuracy, making them very suitable for quantification of Piroxicam capsules and Metronidazole tablets.
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A thesis submitted to the Board of Postgraduate Studies, Kwame Nkrumah University of Science and Technology, Kumasi, in partial fulfilment of the requirements for the award of the Degree of Master of Science in Analysis and Quality Control
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