Application of the Ratio Subtraction Ultraviolet Spectrophotometry Method for Simultaneous Quantification of B1, B2, B3, B6, and B12 Vitamins in Health Supplement Syrups

Abstract

The present investigation aimed to simultaneously determine the concentrations of thiamine (B1), riboflavin (B2), nicotinamide (B3), pyridoxine (B6), and cobalamin (B12) in health supplement syrups, commonly used as multivitamins. Precise quantification of these vitamins is essential to guarantee product efficacy, safety, and compliance with quality standards. The analytical approach employed was based on ratio subtraction ultraviolet (UV) spectrophotometry, utilizing methanol as a solvent to resolve the overlapping UV absorption spectra of the vitamin mixture. Carefully optimized divisor concentrations 12 µg.mL–1 for B1, 22 µg.mL–1 for B2, 18 µg.mL–1 for B3, 11.5 µg.mL–1 for B6, and 25 µg.mL–1 for B12 were selected to ensure accurate spectral differentiation of each analyte. The method was rigorously validated according to standard parameters, including linearity, accuracy, precision, limit of detection (LOD), and limit of quantification (LOQ), all of which fulfilled established validation criteria. The quantified vitamin contents in health supplement syrup samples were 102.75% ± 2.20% for B1, 101.03% ± 2.17% for B2, 102.65% ± 2.21% for B3, 102.11% ± 2.16% for B6, and 101.26% ± 2.17% for B12, all of which conformed to the specifications outlined in the United States Pharmacopeia 38 and National Formulary 33. The proposed ratio subtraction ultraviolet spectrophotometry method offers a straightforward, cost-effective, and selective solution for concurrent estimation of these vitamins in complex pharmaceutical matrices, without the need for sophisticated instrumentation or elaborate procedures.