Development and validation of a liquid Chromatographic method for the simultaneous Determination of caffeine, chlorzoxazone, Codeine, diclofenac, doxylamine, ibuprofen and Paracetamol in analgesic preparations

Abstract

Pain is a common menace afflicting many people worldwide. There are a myriad of medications that are available commercially to combat all forms of pain. The quality of medicines is an important aspect in healthcare provision. However, more focus is put on the quality of drugs for the major diseases such as tuberculosis, HIV/AIDS and malaria than for analgesics. There is little quality surveillance of the pain medications that are available commercially in Kenya. There are no official methods for the simultaneous analysis of most of the drug compounds present in the commonly available pain medications. Only the methods for the analysis of paracetamol and caffeine preparations and paracetamol and codeine phosphate preparations are available. This makes the analysis of samples expensive and time consuming as each component would be analyzed individually. In the present study, a simple, rapid, precise, sensitive and robust isocratic elution reversed-phase liquid chromatographic method was developed for the simultaneous determination of caffeine, chlorzoxazone, codeine, diclofenac, doxylamine, ibuprofen and paracetamol. These seven compounds are present in some commonly available formulations used for pain management in Kenya. A mixture of these seven compounds was separated using a liquid chromatographic system with a mobile phase consisting of methanol-0.25 M sodium octanesulfonate-0.2 M ammonium acetate pH 6.5-water (50:2:10:38, % v/v/v/v). This was delivered at a flow rate of 1 mUmin through a column with the dimensions 250 mm in length and 4.6 mm internal diameter packed with an octyldecylsilane stationary reverse phase (Phenomenex Gemini® 511, CIS) maintained at a temperature of 40°C using a column oven and a UV detection wavelength of 220 nm. Validation of the method demonstrated that the limit of detection for caffeine, chlorzoxazone, codeine, diclofenac, doxylamine, ibuprofen and paracetamol were 11.99 ng, 20.72 ng, 25.08 ng, 12.50 ng, 32.93 ng, 12.02 ng, 22.36 ng, and the limit of quantitation were 79.92 ng, 103.60 ng, 104.50 ng, 62.49 ng, 164.64 ng, 48.08 ng, 111.80 Xlll ng respectively. The method was accurate with recovery rates of 100.3% (caffeine), 101.6% (chlorzoxazone), 98.3% (codeine), 98.1% (diclofenac), 102.7% (doxylamine), 99.2% (ibuprofen) and 98.1% (paracetamol). The method was linear over a concentration range of 75% to 125% for all seven compounds with the respective coefficient of determination (R2) values being 0.9995 (caffeine), 0.9986 (chlorzoxazone), 0.9992 (codeine), 0.9993 (diclofenac), 0.9975 (doxylamine), 0.9998 (ibuprofen) and 0.9960 (paracetamol). The method also demonstrated adequate intra-day and intermediate precision with intra-day precision coefficients of variation ranging from 0.15-0.37% and intermediate precision coefficients of variation ranging from 0.91-1. 96% for the seven compounds. The method developed was used for the analysis of four randomly selected commercially available pain medications containing varying combinations of the seven compounds. Three batches of each of the drug samples were analyzed and the results obtained demonstrated that there was minimal batch variation. The assay values for caffeine, chlorzoxazone, diclofenac and ibuprofen ranged from 95.3-102%, 92.0-96.6%, 95.5- 98.6% and 96.7-99.3% respectively. The assay values of paracetamol ranged from 99.6- 117%. One product was found to have values of paracetamol consistently higher than the adopted limits with assay values ranging from 115-117%. The levels of codeine and doxylamine were found to be consistently below the adopted specifications with assay values ranging from 51.3-53.2% and 65.2- 67.2% respectively. The method can be used in the analysis of pain medications containing any combination of the seven compounds and it can therefore be adopted by quality control laboratories for routine analysis and in the post market surveillance of pain medications.