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Determination of B-Vitamins in Infant Formula and Adult/Pediatric Formulas

ESI LC-MSMS

3

F. Procedure

a) Preparation of food samples:

Add 25.0 g of powdered infant formula/nutrtionals to 200 g of water

at 40

˚C. Mix until a homogenous suspension is obtained

.

b) Extraction:

1) Weigh 2.5 g aliquot of homogenized sample suspension into a 50 mL polypropylene

centrifuge tube. Add 25 mL of 1% Ascorbic acid solution in water and vortex for 10 minutes.

2) Adjust the pH to 4.8 ± 0.2 with aqueous ammonia.

3) Transfer quantitatively to a 50 mL volumetric flask and makeup to 50 mL with laboratory

water and mix.

4) Filter an aliquot through 0.22 μm PTFE syringe filter and inject into LC-MSMS.

c) Chromatographic Conditions:

Table 3:

Column

Zorbax RRHD, 1.8 μm, 3 x 100 mm

Mobile phase A

20 mM Ammomium Formate with 0.1% Formic

acid in water

Mobile phase B

20 mM Ammomium Formate with 0.1% Formic

acid in Methanol

Flow rate

0.5 mL/minute

Injection Volume

5.0 μL

Column Temperature

55°C

Total Run Time

8.0 minutes

Gradient Method

Time (minutes)

A %

B%

0.00

99.0

1.0

0.50

99.0

1.0

0.60

90.0

10.0

5.00

15.0

85.0

5.10

99.0

1.0

8.00

99.0

1.0

Source Parameters

Parameter

ESI (+)

Gas Temp (°C)

300

Gas Flow (l/min)

7

Nebulizer (psi)

35

SheathGasHeater

400

SheathGasFlow(l/min)

11

Capillary (V)

2500

VCharging

0

BVit-03 (February 2016)

FOR ERP USE ONLY

DO NOT DISTRIBUTE