SPIFAN Nutrients ERP Book_9-29-15

Carn-06 (Choline Validation Report) FOR ERP USE ONLY DO NOT DISTRIBUTE

Eurofins Lab. LTD., Wolverhampton: SOP CHROM/341-Choline in pet foods by ion chromatography with conductivity detection.

The quantification by LC-MS/MS is based on the analytical concepts of the follow- ing article.

ANDRIEUX et AL. J.AOAC INT. 91,4,2008, 777-785. Simultaneous Determination of Free Carnitine and Total Choline by Liquid Chromatography/Mass Spectrome- try in Infant Formula and Health-Care Products: Single-Laboratory Validation.

Brief method description:

The sample is shaken in a mixture of water and methanol thereby extracting cho- line including their phosphorylated forms. An aliquot is treated with potassium hy- droxide solution at elevated temperature for 4 hours thereby converting the phos- phorylated forms to free choline. The mixture is neutralised and deuterered inter- nal standard is added. Quantification is done by LC-MS.

Statistics:

In the current validation report, a p-value < 0.05 is considered statistically signifi- cant. Consequently, confidence intervals (CI) are calculated at a confidence level at 95 %. Paired t-testing is performed on normal distributed raw data. Data which is not within the normal distribution is clearly marked within the text.

Matrices:

This method can be used for quantification of free and/or total choline in the fol- lowing matrices:

 Food and feed.  Supplements, premixes and raw products.  Milk powder and nutritional formulas.  Animal feed / pet food (dry and wet).

Specificity:

The specificity has been evaluated by examining chromatograms of natural sam-