Lc 2010cht hplc system

The multi-vitamins stock solution was prepared by weighing in a 100 mL volumetric flask 5 mg of vitamin B12; 12.5 mg of vitamin B2; 25 mg each of vitamins B1, B6 and B3. Forty millilitres (40 mL) of water was then added and the solution was shaken vigorously before adding 4 mL of 2 M NaOH. After complete dissolution of the vitamins, 50 mL of 1 M phosphate buffer (pH 5.5) was added and the solution made up to the mark with water. Stock standard solutions were prepared daily. Different concentrations of the standards were injected into the HPLC to obtain the peak areas. Peak areas were plotted against concentration for each vitamin to make specific calibration curves. The Shimadzu LC-2010CHT HPLC system was used with conditions according to Moreno et al. [25 (link)]. A volume of 20 µL for each sample was injected into the HPLC equipped with a C18 reversed-phase column (250 × 4.6 mm, 4 μm), 0.05 M ammonium acetate (solvent A)–methanol (solvent B) 92.5:7.2 as mobile phase at 1 mL/minute flow rate. A diode array detector was used to scan from 200 to 500 nm and LC-solutions software (Shimadzu, Japan) was used to integrate the peak areas for each vitamin. After the run, the peak area of each unknown sample was obtained, and concentrations were calculated using the calibration curves.