HPLC Analysis of Water-Soluble B-Vitamins on Ascentis^® Express C18

column

Ascentis Express C18, 5 cm x 3.0 mm I.D., 2.7 μm particles (53811-U)

mobile phase

[A] 20 mM potassium phosphate, dibasic, pH 7; [B] methanol

gradient

0.5% B for 1.3 min, 0.5-30%B over 1.7 min; to 30% B in 1.4 min, 2 min equilibration at 0.5% B

flow rate

1.0 mL/min

pressure

4130 psi (285 bar)

column temp.

35 °C

detector

UV, 210 nm

sample

9 vitamins in water, 10 μg/mL for nicotinic acid and folic acid, 12 μg/mL for thiamine hydrochloride and cyanocobalamin, (20 μg/mL for pyridoxine, 30 μg/mL for riboflavin, 40 μg/mL for niacineamide, 100 μg/mL for calcium pantothenate, 120 μg/mL for biotin)