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Silliker

Chem, Res. Center Crete, IL – Report of A Validation of LC-MS/MS Method for Folate Analys

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about 10% of the total folates in the samples. One other sample, infant formula powder based on

partially hydrolyzed milk was found to contain a very small amount of minor folate which contributed

to 6.5% of total folate in the sample.

% Spike Recovery of total folate (sum analyzed folates as folic acid) after spike with the targeted

folate compounds and their analysis in SPIFAN II matrices:

Details of Reconstitution of the samples before the analysis:

All of the 5 samples received from SPIFAN for spiking purposes were analyzed in replicates by the

method for the targeted folate analytes with and without spike with the targeted folate compounds. The

powder sample were reconstituted by taking approximately 25 g of the powder into 200 mL of water.

Details of the samples and reconstitution are provided in the following Table 20.

There are total of five samples for spiking purposes out of which two are powders. All the powdered

samples were reconstituted as ready to feed beverages.

All samples were spiked in triplicates with targeted folate compounds at three different levels (spike

Level I, II and III). Samples with different spike levels were analyzed employing trienzyme extraction

including conjugase on different days. The samples were spiked with each targeted folate compound

and their sum as folic acid calculated as total folate spike level (folic acid). Spike levels are calculated

as mcg/100g in the reconstituted liquid and as is the RTF samples.

Table 20: Details of the spike levels and reconstitution of the SPIFAN samples processed for %

spike recovery evaluation

Matrix

Lot #/ID

Reconstitution of Powder in

to water

Analyzed as

Spike levels

(mcg/100g)*

Total Folate (T),

Individual Folate

Compound (I)

Powder (g)

In to water (g)

Level

1

Level

2

Level

3

Child Formula Powder

004011RF00

Recd 12/4/15

25.0083

200

Reconst.

Beverage

T=

500

I = 60-

80

T=120

I=20

T=60

I=10

Infant Formula Powder

00402RF00

Recd 12/4/15

25.0026

200

Reconst.

Beverage

Adult Nutritional RTF High

Protein

00821RF00

Recd 12/4/15

Recd as Beverage

As is

Adult Nutritional RTF High

Fat

00820RF00

Recd 12/4/15

Recd as Beverage

As is

Infant Formula Powder

Milk Based, SPIFAN Blank

Milk Form

EV4H2Q Recd

12/4/15

Recd as Beverage

As is

* Samples were spiked with each targeted folate compound. Total Folic calculated as sum of the folate

compounds after adjusting their values to folic acid.

2011.06/Fol-22 (February 2016)

FOR ERP USE ONLY

DO NOT DISTRIBUTE