149 / 696
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