Estimation of Vitamin D2 and D3 in Food using LC-QQQ

Most people get their vitamin D requirements through a combination of solar conversion and by maintaining a various and balanced diet. Liver, fish, eggs, and milk, as well as other dairy products are rich in vitamin D. In dairy products, vitamin D exists as cholecalciferol. To maintain the vitamin D requirements of the population, ergocalciferol and cholecalciferol are used to fortify food ingredients such as cooking oil. Edible oil is one of the best food commodities to serve as a vehicle to easily reach consumers and reduce issues related to vitamin deficiency. Analysis of ergocalciferol and cholecalciferol in edible oil is challenging because of the complexity of the matrix.

For the accurate and simultaneous quantitation of these vitamins, an analytical method was developed using the Agilent 6470 triple quadrupole LC/MS system with APCI ionization installed. Sample preparation is based on saponification, followed by extraction with hexane. The evaporated hexane residue was reconstituted in methanol before instrumental analysis.

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Overlay of various concentrations of Vitamins D2 and D3

Matrix-matched calibration curve for cholecalciferol

Highlights of the Application Note

  • Sample processing by saponification followed by simple extraction.
  • Derivatization of Vitamin D with PTAD for stability and increased sensitivity
  • Excellent linearity in the range from 0.4 to 40 ppb
  • Lower quantification limit of 0.4 ppb
  • Excellent recovery within 80 to 120 % of the spiked concentration for both Vitamin D2 & D3
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Application Note

Simultaneous Determination of Vitamin D2 and D3 in Milk