Estimation of Nicotine in Tobacco Extract Using C8 Column in High Performance Liquid Chromatography

Background: Nicotine is the most used toxic substance in spite of mass media awareness. Column used in the estimation of nicotine is found to be the inhibiting factor for its estimation.Objective: To develop a method for nicotine estimation on C8 column in HPLC.Methods: The method involved a model Waters 1515 binary HPLC Pump system interfaced with a 2489 Waters UV/VIS detector, a 4.6 X 250 mm, 5μm beads Symmetry C8 column at 37°C, and an isocratic mobile phase containing 60%: 40% v/ v mixture of 10mM sodium acetate (pH 5.5) and methanol at a flow rate of 1.0ml/min at 256 nm. The method was validated for specificity, linearity, precision, accuracy, the Limit of Detection (LOD), the Limit of Quantitation (LOQ) and system suitability.Results: The HPLC Nicotine peak with average retention time of 3.415 minutes was observed on chromatogram with RSD <0.5%; linearity was greater than 0.99 with acceptable precision (within USP limits of <2.0% RSD) and USP tailing less than 2. LOD was found to be 0.200 μg/ml and Limit of Quantitation (LOQ) was found to be 0.609 μg/ml of nicotine. The average yield of the nicotine extracted by acid base extraction method from tobacco was 1.68% (range: 1.34–2.22%).Conclusion: A HPLC method based on C8 column for analysis of Nicotine was developed and validated successfully.International Journal of Human and Health Sciences Vol. 07 No. 02 April’23 Page: 166-170