Quantification of small quantities of a standardized Dry Grape Extract in complete feed using UHPLC-MS/MS

A specific and sensitive method for quantification of small quantities of a standardized Dry Grape Extract in complete feed based on UHPLC-MS/MS has been developed and validated. The standardized Dry Grape Extract studied is a natural commercial feed additive (Nor-Grape®, Nor-Feed, France) used in animal nutrition, consisting of a complex mixture of grape seed extracts and grape skin extracts in which malvidin-3-O-glucoside (M3OG) has been identified and quantified. M3OG was used as phytomarker for the quantification of the additive in feed. Key steps in the preparation of the sample are the use of several successive and selective extractions allowing firstly to eliminate a large number of undesirable molecules and in a second time to recover and concentrate the anthocyanins of the additive present in the sample. The use of an internal standard (cyanidin-3-O-sambubioside) makes it possible to compensate for the losses of the analyte during sample work-up. M3OG quantification is performed by the standard addition method to compensate ion suppression during HPLC-MS/MS analysis. The method was developed and validated in-house in accordance with the guidelines recommended by IUPAC (selectivity, calibration and linearity, trueness, precision, recovery, limit of quantification, measurement uncertainty) [1] for the quantification of a small dose of the standardized Dry Grape Extract in complete feed (30ppm). This patented method of analysis has all the prerequisites to be used by European authorities as part of the registration of a feed additive [2].

• References

• 1 Thompson M, Ellison SLR, Wood R. Harmonized guidelines for single-laboratory validation of methods of analysis (IUPAC Technical Report). Pure Appl Chem 2002; 74: 835-855
• 2 FEEDAP. . Guidance for the preparation of dossiers for zootechnical additives: Guidance on zootechnical additives. EFSA J 2012; 10: 2536