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FOR EXPERT REVIEW PANEL USE ONLY
NOT FOR DISTRIBUTION
1
Expert Review Panel for Infant Formula and Adult Nutrition
Evaluation of Method
Caro-03
Title:
Determination of β-Carotene, Lutein and Lycopene in Infant and Adult/Paediatric Nutritionals
Formula using HPLC with UV Detection
Author:
Abbott Nutrition
Summary of Method:
Carotenoids are extracted from samples with 75:25 hexane: MTBE following
saponification with a mix of KOH, THF and methanol. Following extraction a portion of the organic layer is
evaporated to dryness and the residue dissolved in 75:25 10% BHT:MTBE. Analysis is by HPLC with
separation of the cis and trans isomers, beta carotene and lycopene on a C30 carotenoid column with a
methanol: MTBE gradient and detection by UV spec at 445nm.
Method Scope/Applicability:
Applicable to the determination of
cis
and
trans
-lutein, cis and
trans
-β-
carotene, and
cis
and
trans
lycopene in infant formula and adult nutritionals.
General comments about the method:
Need to include scope/applicability – can use sentence from introduction in SLV report
Requires more detail on sample weigh – sample weight for liquid products included but no detail on
powder or reconstituted products. Assume 1 g is used for all but need to be stated.
To comply with SPIFAN guidelines, add sample preparation for homogeneous vs non-homogeneous
samples – 25g plus 200 mL water etc.
Add note indicating that liquid and reconstituted samples should be prepared fresh prior to analysis.
Add chromatograms to the method - (in the SLV report).
Suggest laying out standard and sample concentration calculations in a clearer format with glossary.
Consider two curves – one for low concentration and one for high carotenoid concentrations or allow for
optimizing curve to suit concentration of samples being analyzed.
Method Safety Concerns:
General safety section needs to be added to the method.
Pros/Strengths:
Good detail on materials, chemicals, reference standard and sample preparation.
Both UV Spectrophotometric and Chromatographic purity performed.