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Robustness:
The robustness of the method has been evaluated on the following critical pa-
rameters; extraction time and saponification time.
Extraction time:
The robustness test has been performed on a milk powder and a pet food con-
taining choline.
The sample is weighed into the extraction bottle, extraction solvents are added,
the bottle is closed and placed on the shaking table at 37°C. At the time t. the
sample is removed from the shaking table, centrifuged and placed on cold stor-
age until saponification.
t / hour
1 2 3 4 5 22
Choline:
From the data (see appendix 4) it is seen that an extraction time of 20 hours is
sufficient to extract all the choline from the sample.
Saponification time:
The robustness test has been performed on raw egg, pet food, milk powder (in-
fant formula), cornflakes and fish feed containing choline.
An aliquot of the extraction solution is transferred to the saponification bottle, po-
tassium hydroxide and water is added and the mixture is saponifyed at a 100°C.
At the time t. the sample is removed from the oven and neutralized with hydro-
chloric acid.
0
500
1000
1500
2000
2500
Choline (mg/kg)
Time (hour)
Ekstraktion 37°C
Pet food
Milk powder
Carn-06 (Choline Validation Report)
FOR ERP USE ONLY
DO NOT DISTRIBUTE