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Dual Lab Validation of a Method for the Determination of Fructans in Infant
Formula & Adult Nutritionals /
25 Jul 2016
49
1M hydrochloric acid:
add in a 1000 ml volumetric flask 500 ml water. Then add with a graduated glass
83 ml hydrochloric acid (36.5 – 38%). Make up with demineralized water to the mark and
homogenize.
80% Acetonitrile + 0.1% TFA, wash solution GC‐SPE
: add in a 200 ml volumetric flask 160 ml acetonitrile
and 100 μl trifluoroacetic acid. Make up with demineralized water to the mark and homogenize.
25% acetonitrile + 0.05% TFA
: add in a 1000 ml volumetric flask 250 ml acetonitrile. Add 100 μl
trifluoroacetic acid. Make up with demineralized water to the mark and homogenize.
Carrez I solution
: dissolve 106 g potassium hexacyanoferrate (II) trihydrate in 1000 ml demineralized
water and store in a brown bottle.
Carrez II solution
: Dissolve in a 1000 ml volumetric flask 220 g zinc acetate in 900 ml demineralized
wateradd 29 ml glacial acetic acid. Make up with demineralized water to the mark and homogenize.
1M sodium chloride
: dissolve in a 1000 ml volumetric flask 58.5 ± 0.1 g sodium chloride in 500 ml
demineralized water. Dissolve and homogenize. Make up with demineralized water to the mark and
homogenize.
Enzyme mix A (Sucrase/β‐amylase/pullulanase/malase)
: dissolve the content of bottle 1 in the
Megazyme fructan assay kit (K‐FRUC) in 22 ml 0.1M sodium maleate buffer pH = 6.5. Mix well and
distribute the obtained solution in 4 ml portions in 15 ml screw cap closed polypropylene tubes. Store
the tubes in the freezer at – 20°C until use.
Enzyme mix B (fructanase)
: dissolve the content of bottle 2 in the Megazyme fructan assay kit (K‐FRUC)
in 22 ml 0.1M sodium acetate buffer pH = 4.5. Mix well and distribute the obtained solution in 4 ml
portions in 15 ml screw cap closed polypropylene tubes. Store the tubes in the freezer at – 20°C until
use.
A‐6
Sample preparation
Reconstitution of (milk) powder products.
Dissolve 25.0 ± 0.1 g powder sample in 200 ± 0.2 g
demineralized water in an erlemeyer flask. The mixture is well homogenized at ambient temperature.
The reconstitution factor F can easily be calculated with the following formula:
In which:
F = factor reconstitution
w
1
= weight powder original
w
2
= weight added amount of water
The weight of the test sample depends on the expected fructan content in the sample to analyze. The
scheme of the weight of the test portion is given table 1.
Table 1
Scheme of the weight of the testportion dependent on the expected fructan content
Expected fructan content in the sample
Required test portion of the sample (g)
< 1% m/m
4 ± 0.1
reconstituted samples
4 ± 0.1
> 1% m/m
1 ± 0.1
1
2
1
/)
(
w ww F
VALIDATION REPORT
FOR ERP USE ONLY
DO NOT DISTRIBUTE