TN-0019
APPLICATIONS
SPE and HILIC HPLC Strategy for Determination of Melamine
Terrell Matthews, Product Manager
Phenomenex, Inc., 411 Madrid Ave., Torrance, CA 90501 USA
Introduction
As the world faces its second melamine scare in two years, there method, which measures the amount of organic nitrogen in the
has been a global concern over international food product imports sample. This method is non-specific, so any other nitrogen present
and food safety testing. The compound itself has a relatively low in the sample will cause the test to show higher protein levels than
toxicity to humans. However, when melamine combines with cya- are actually present. By adding melamine to their products, food
nuric acid, it forms an insoluble salt crystal that causes kidney fail- producers have found a way to cover up diluting their products with
ure in animals and humans. the consumer suffering the consequences.
Nitrogen is a major component of amino acids, which are the The determination of melamine in food products is of great impor-
building blocks for proteins. With 6 nitrogen atoms in its structure, tance, especially with the recent discovery of melamine in baby
melamine has been added to some foods to falsely increase their formula. The following methodology can be used to determine the
protein content. This manipulation is possible because the stan- levels of melamine in food products. In this method, SPE clean up
dard method to determine protein content in food is the Kjeldahl of the sample is followed with analysis by LC/MS/MS.
SPE method - specific targeting using Strata™-X-C HPLC method using Luna® HILIC
SPE Method HPLC Method
Material: Strata-X-C, 100 mg/6mL Column: Luna 3 μm HILIC, 100x2.0 mm
Part Number: 8B-S029-ECH Part Number: 00D-4449-B0
Condition: 2 mL Methanol Flow rate: 0.4 mL/min
Equilibrate: 2 mL 0.1% TFA in water, pH<4 Mobile Phase: (90:10) Acetonitrile:100 mM Ammonium Formate,
pH 3.2
Load: Load sample and allow to slowly drip through cartridge
at a rate of 1-2 dps (drops per second) Analysis: Isocratic
Wash A: 2 mL 0.1% TFA in Water, pH<4 Run time: 3.5 minutes
Wash B: 2 mL Methanol Injection volume: 5 μL
Elution: 2 mL Methanol / 5% NH4 OH. Use a 2 stage elution Temperature: Ambient
(2 x 1 mL) MRM: m/z 127 – 85, 127 – 60
ESI Positive mode
For related information, see the FERN method for melamine analysis online at http://www.fsis.usda.gov/PDF/FERN_CHE_0003.pdf.
Page 1 of 3
TN-0019
APPLICATIONS
50 ng/ml Melamine standard, (95:5) Acetonitrile:Water
2.02
Max. 7613.3 cps.
0 1 2 3 min
2.00
Max. 9120.0 cps.
0 1 2 3 min
2.00
Max. 9126.7 cps.
0 1 2 3 min
2.02
Max. 8786.7 cps.
0 1 2 3 min
10 ng/ml Melamine standard (95:5) Acetonitrile:Water
2.05
Max. 2273.3 cps.
0 1 2 3 min
2.0 3
M a x. 21 40 .0 cp s.
0 1 2 3 min
For related information, see the FERN method for melamine analysis online at http://www.fsis.usda.gov/PDF/FERN_CHE_0003.pdf.
Page 2 of 3
TN-0019
APPLICATIONS
Australia
t: 02-9428-6444
f: 02-9428-6445
[email protected]
Austria
t: 01-319-1301
f: 01-319-1300
[email protected]
Belgium
t: +31 (0)30-2418700
f: +31 (0)30-2383749
[email protected]
Canada
t: (800) 543-3681
f: (310) 328-7768
[email protected]
Denmark
t: 4824 8048
f: 4810 6265
[email protected]
France
t: 01 30 09 21 10
f: 01 30 09 21 11
[email protected]
Germany
t: 06021-58830-0
f: 06021-58830-11
[email protected]
Ireland
t: 01 247 5405
f: +44 1625-501796
[email protected]
Italy
t: 051 6327511
f: 051 6327555
[email protected]
Luxembourg
t: +31 (0)30-2418700
f: +31 (0)30-2383749
[email protected]
Netherlands
t: 030-2418700
f: 030-2383749
[email protected]
New Zealand
t: 09-4780951
f: 09-4780952
[email protected]
Puerto Rico
t: (800) 541-HPLC
f: (310) 328-7768
[email protected]
United Kingdom
t: 01625-501367
f: 01625-501796
[email protected]
All other countries:
Corporate Office USA
t: (310) 212-0555
f: (310) 328-7768
[email protected] TN63080708
www.phenomenex.com
Trademarks
Phenomenex products are available worldwide. For the distributor in your country,
Luna is a registered trademark of Phenomenex, Inc. Strata-X is a trademark of Phenomenex,
contact Phenomenex USA, International Department at [email protected].
Inc.
Disclaimer
Subject to Phenomenex Standard Terms & Conditions, which may be viewed at
www.phenomenex.com/TermsAndConditions.
© 2008 Phenomenex, Inc. All rights reserved.
Page 3 of 3
