Rapid, Specific Analysis of Melamine and Cyanuric Acid in Infant Formula

This method is rapid and sensitive for the analysis of melamine and cyanuric acid simultaneously in infant formula. Using two Oasis solid-phase extraction protocols and the ACQUITY UPLC, the results are consistent with the published US FDA interim method, while demonstrating a reduced analysis time.

Figure 1: Chromatograms of melamine in dry infant formula at 500ppb and 2500ppb using ACQUITY BEH HILIC column.

Responding to recent worldwide concerns related to melamine in food, the US FDA issued an interim method for the determination of residual melamine and cyanuric acid in foods using LC–MS-MS. Based upon that work, this application note describes the rapid, specific analysis of melamine and cyanuric acid in infant formula by UPLC–MS-MS.

Two separate solid-phase extraction protocols are used: Oasis MCX for melamine and Oasis MAX for cyanuric acid. Two LC methods were developed on the ACQUITY UPLC system. The first HPLC method uses an Atlantic HILIC Silica column. The second method uses an ACQUITY UPLC BEH HILIC column coupled with the ACQUITY UPLC system to provide a significantly reduced cycle time.

Experimental

All standard and sample solutions were prepared per the FDA interim method.

SPE Procedure

Melamine SPE clean-up

Use Oasis MCX cartridge, 6 cc 150mg (part number 186000255). Condition (5 mL), equilibrate (5 mL), load (3 mL), wash (5 mL), and elute (4 mL). Filter the eluent into vials using 0.2 μm PTFE syringe filters and syringes. Pipette 950 μL of extract and 5 μL of water into autosampler vial.

Cyanuric acid SPE clean-up

Use Oasis MAX cartridges, 6 cc 150 mg (part number 186000370). Condition (5 mL), equilibrate (5 mL), load (3 mL), wash (5 mL), and elute (2 mL). Filter the eluent into vial using 0.2 μm PTFE syringe filters and syringes. 950 μL of extract and 5 μL of water into autosampler vial.

UPLC Conditions

LC system: ACQUITY UPLC system

Column: ACQUITY UPLC BEH HILIC

2.1 × 100 mm, 1.7 μm

Part number: 186003461

MS Conditions

MS system: Waters ACQUITY TQD

Software: MassLynx v.4.1

Ionization mode: ESI positive (melamine and ¹³ C₃¹⁵ N₃ melamine) ESI negative (cyanuric acid and ¹³ C₃¹⁵ N₃ cyanuric acid)

Conclusions

The results are consistent with the published results indicated in the FDA interim method. When applied for the detection of melamine in liquid infant formal, reliable detection was confirmed at an LOQ of 3 ppb on the ACQUITY BEH HILIC. The instrument analysis time using either method has been reduced compared to what is listed in the FDA method. The time savings is especially significant when using the ACQUITY BEH HILIC column coupled with the ACQUITY UPLC system.

For the complete UPLC application note and results from the HPLC method, visit www.waters.com/28651.

© 2009 Waters Corporation. Waters, The Science of What's Possible, ACQUITY, UPLC, ACQUITY UPLC, Oasis and MassLynx are trademarks of Waters Corporation.

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