HPLC Analysis of Active Drug in a Formulation

HPLC method for separation of active ingredients of drug/supplemental composition was developed on an Obelisc R trimodal HPLC column. Compounds are retained by combination of reversed-phase, cation-exchange and anion-exchange mechanisms. Compounds are well separated, and method can be used for quantitation of pyridoxine, ascorbic acid, niacinamide, pantothenic acid, caffeine and riboflavin in a mixture or as separate compounds in various complex mixtures. Various detection techniques can be applied for quantitation (ELSD, UV, LC/MS, Corona). This HPLC method can be adopted as general approach for analysis of active drug components in various formulations.


Column Obelisc R , 4.6x150 mm, 5 µm, 100A
Mobile Phase MeCN/H2O  -5/95%
Buffer NaHPO4 pH 3.0 - 30 mM
Flow Rate 1.0 ml/min
Detection UV, 210 nm


Class of Compounds Drug, Vitamin B₆, Hydrophobic, Ionizable
Analyzing Compounds Pyridoxine, Ascorbic acid, Niacinamide, Pantothenic acid, Caffeine, Riboflavin

Application Analytes:

Ascorbic Acid
Pantothenic Acid
Riboflavin (Vitamin B2)
Vitamin B6 (Pyridoxine)