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FOR EXPERT REVIEW PANEL USE
ONLY NOT FOR DISTRIBUTION
1
Expert Review Panel for Infant Formula and Adult Nutrition
Title:
Determination of Free and Total Choline and Carnitine in Infant Formula and Adult/Pediatric
Nutritional Formula by Liquid Chromatography/Tandem Mass Spectrometry (LC/MS/MS)
Author:
Covance; David Ellingson, Study Director
Summary of Method:
Method is applicable to the determination of free and total L-carnitine and choline. L-carnitine portion of
the method (2015.10) was approved to move to MLT in 9/2015. The SMPR for choline requires
“determination of total choline” so the method as it applies to free choline determination is not
considered.
A portion of reconstituted powder or liquid sample is accurately weighed into a microwave digestion
vessel. Labeled internal standards for both compounds of interest are added. The sample is acidified, the
vessel sealed, and it is processed through the microwave digestion unit at 120C for c 40 minutes. The
contents are then transferred to a polypropylene tube, and diluted with water, then a portion of the
resulting solution is diluted with ACN, centrifuged and filtered. A reagent blank and the assay calibrants
are processed in the same manner. Quantification is by LC/MS/MS (ESI+) w MRM using a multi-point
ISTD calibration for each compound.
Method Scope/Applicability:
General comments about the method:
Method Clarity:
Clearly written.
Method Safety Concerns:
None.
Pros/Strengths:
•
Recovery of two endogenous forms of choline was demonstrated (acetylcholine and
phosphatidylcholine)
Cons/Weaknesses
•
In the repeatability study, the validation report indicated that outliers had been removed and replaced,
but did not show the data.
Supporting Data
•
General Comment:
Evaluation of Method Chol-08 / Carn-07
#1
Chol-08 Review Forms