Determination of Anthocyanins in Cranberry Fruit and Cranberry Fruit Products by High-Performance Liquid Chromatography with Ultraviolet Detection: Single-Laboratory Validation

Abstract

Abstract A single-laboratory validation study was conducted on an HPLC method for the detection and quantification of cyanidin-3-O-galactoside (C3Ga), cyanidin-3-O-glucoside (C3Gl), cyanidin-3-O-arabinoside (C3Ar), peonidin-3-O-galactoside (P3Ga), and peonidin-3-O-arabinoside (P3Ar) in cranberry fruit (Vaccinium macrocarpon Aiton) raw material and finished products. An extraction procedure using a combination of sonication and shaking with acidified methanol was optimized for all five anthocyanins in freeze-dried cranberry fruit and finished products (commercial extract powder, juice, and juice cocktail). Final extract solutions were analyzed by HPLC using a C18 RP column. Calibration curves for all anthocyanin concentrations had correlation coefficients (r2) of 99.8. The method detection limits for C3Ga, C3Gl, C3Ar, P3Ga, and P3Ar were estimated to be 0.018, 0.016, 0.006, 0.013, and 0.011 g/mL, respectively. Separation was achieved with a chromatographic run time of 35 min using a binary mobile phase with gradient elution. Quantitative determination performed in triplicate on four test materials on each of 3 days (n 12) resulted in RSDr from 1.77 to 3.31. Analytical range, as defined by the calibration curves, was 0.5736.53 0000000g/mL for C3Ga, 0.159.83 g/mL for C3Gl, 0.2817.67 g/mL for C3Ar, 1.0164.71 g/mL for P3Ga, and 0.4227.14 g/mL for P3Ar. For solid materials prepared by the described method, this translates to 0.063.65 mg/g for C3Ga, 0.020.98 mg/g for C3Gl, 0.031.77 mg/g for C3Ar, 0.106.47 mg/g for P3Ga, and 0.042.71 mg/g for P3Ar.