Liquid Chromatographic Method for Determination of Vanillin and Ethyl Vanillin in Imitation Vanilla Extract (Modification of AOAC Official Method 990.25): Collaborative Study

A collaborative study of a method for analysis of vanillin and ethyl vanillin in fortified and imitation vanilla flavors was performed. The method, which is an extension of AOAC Official Method 990.25, Vanillin, Vanillic Acid, p-Hydroxybenzaldehyde, and p-Hydroxybenzoic Acid in Vanilla Extract, Liquid Chromatographic Method, involves reversed-phase high-pressure liquid chromatography (LC) of the sample on a C8 column with a water-methanol-acetic acid (89 + 10 + 1) mobile phase and UV detection at 254 nm. The method yields good recoveries of vanillin at 484 and 723 mg/100 mL and of ethyl vanillin from 37 to 400 mg/100 mL. For vanillin, repeatability (r) values were 8 mg/100 mL at a level of 155 mg/100 mL, 12 mg/100 mL at a level of 484 mg/100 mL, and 31 mg/100 mL at a level of 723 mg/100 mL. Reproducibility (R) values were 20 mg/100 mL, 55 mg/100 mL, and 137 mg/100 mL over the same range. For ethyl vanillin, r values were 2.4 mg/100 mL at a level of 37 mg/100 mL, 3.2 mg/100 mL at a level of 74 mg/100 mL, and 8.6 mg/100 mL at a level of 180 mg/100 mL. R values were 6.4 mg/100 mL, 5.4 mg/100 mL, and 22.0 mg/100 mL over the same range. AOAC Official Method 990.25 has been modified to include determination of ethyl vanillin in vanilla extract and artificial vanilla flavor.