Detection of Dimethyl Fumarate with a NovaChrom Nova Atom PC18 Column

Dimethyl fumarate also known as (E)-2-butenedioic acid dimethyl ester, trans-butenedioic acid dimethyl ester, alternatively referred to as dimethyl maleate, commonly known as moldproof preservative Moldkiller No. 1, abbreviated as DMF. It appears as white crystals or a crystalline powder at room temperature. It is soluble in ethyl acetate, chloroform, acetone, and alcohols, slightly soluble in ether, and very slightly soluble in water. DMF is toxic due to its corrosive and allergenic effects on the human body. Clinical trials have shown that ingestion of DMF can cause corrosive damage and allergic reactions in the intestines and internal organs. Furthermore, when this substance comes into contact with the skin, it can induce painful contact dermatitis, including symptoms such as itching, irritation, redness, and burns. It poses a significant threat to human health and is particularly detrimental to the growth and development of children. Dimethyl fumarate was once widely used for mold, decay, and insect prevention, as well as freshness preservation, in food, beverages, feed, traditional Chinese herbals, cosmetics, fish, meat, vegetables, fruits, etc. It is now classified as a non-edible substance and is prohibited for use in food products (including flour-based products).

Reference standard: NY/T 1723-2009 Determination of dimethyl fumarate in foods - High performance liquid chromatography.

1. Experiment Materials

1.1 Methanol: HPLC grade

1.2 Beverage sample

1.3 Ultrapure water

1.4 Dimethyl fumarate standard

1.5 Filter: pore size 0.22μm

1.6 Ultrasonic cleaner

1.7 2mL amber sample vial

1.8 2mL disposable syringe

1.9 Nova Pre - HLB 150mg/6mL

2. Experiment Pre-treatment

Prepare calibration curve concentration points at 5mg/L, 10mg/L, 15mg/L, 20mg/L, 25mg/L, and 50mg/L.

Clean-up using a NovaPre HLB SPE column (150mg/6mL):

2.1. Activation: Condition the HLB cartridge with 6mL of methanol followed by 6mL of water before use.

2.2. Loading: A 5mL aliquot of the beverage sample was passed through the cartridge, and the effluent was discarded.

2.3. Washing: Wash the cartridge with 5mL of a 5% methanol-in-water solution. Discard the effluent and dry the cartridge under vacuum.

2.4. Elution: The cartridge was eluted with 5mL of an 80% methanol-in-water solution, and the eluate was collected. (Throughout the entire loading and elution process, the flow rate was maintained at or below 1mL/min).

2.5. Instrumental Analysis: The eluate was filtered through a 0.22μm membrane and then submitted for instrumental analysis.

3. Test Conditions

4. Experiment Result

4.1 Standard Testing

4.2 Spiked Sample Chromatogram

Spiked Recovery Results

5. Conclusion

The analysis of dimethyl fumarate in beverages was achieved using a Wayeal LC3200 series liquid chromatograph with a NovaChrom Nova Atom PC18 column (4.6 × 250mm, 5μm), combined with a simple sample preparation method employing a NovaChrom NovaPre-HLB solid-phase extraction column (150mg/6mL). This method demonstrated an average recovery of 98.98% at a spiking level of 5mg/kg, with excellent peak shape.