Application of Ultra-Performance Liquid Chromatography/Tandem Mass Spectrometry for the Measurement of Vitamin D in Foods and Nutritional Supplements

Abstract

Ultra-performance liquid chromatography (UPLC)/MS/MS was applied to measure vitamin D in various foods and nutritional supplements. The run-time of the chromatographic separation was cut from 20 min in HPLC/MS/MS to 10 min in UPLC/MS/MS, while equal or better separation efficiency was achieved to deal with complex food matrixes. Under the optimized conditions, all the previtamins of vitamin D3, D2, and isotope-labeled vitamin D3 were baseline-separated from their corresponding vitamins. It was also demonstrated that many sterol isomers in complex food matrixes that interfere in the analysis could be well-separated from the analytes. Accuracy of this method was evaluated by analysis of NIST SRM 1849 infant formula reference material. With eight replicates, the average vitamin D3 concentration was 0.251 + 0.012 mg/kg, an excellent agreement with the certified value of 0.251 + 0.027 mg/kg. In addition, spike recovery from a commercial infant formula matrix was in the range of 100 to 108% for both vitamins D3 and D2 at three spike concentration levels. The spike recovery for an even more complex matrix, pet food, was 101–105%. LOQ values were 0.026 and 0.033 IU/g, or 0.086 and 0.11 IU/mL in solution, for vitamins D3 and D2, respectively. The dynamic range had three orders of magnitude, which made the method flexible and useful to deal with the wide concentration range of vitamin D in various samples. The method was robust based on the results of changing the parameters of LC separation and MS measurement. This accurate and reliable vitamin D method increased instrument efficiency and analysis productivity significantly.

Application of Ultra-High-Performance Liquid Chromatography/ Tandem Mass Spectrometry for the Measurement of Vitamin D in Infant Formula and Adult/Pediatric Nutritional Formula: First Action 2011.11

“…The method was found suitable for the routine measurement of vitamin D in infant formula and adult/pediatric nutritional formula in a single-laboratory validation (SLV) study (2). The method provides a faster separation of analytes by using a column with sub-2 μm particle size.…”

“…The SLV provided information on the parameters, which included accuracy, precision, range, linearity, and robustness (2). Accuracy was achieved by bias and recovery.…”

“…n March 28, 2011, the AOAC Board of Directors approved an alternative path to achieve Offi cial First Action status. Following this process, an expert review panel (ERP) reviewed the method, "Application of Ultra-High-Performance Liquid Chromatography/Tandem Mass Spectrometry for the Measurement of Vitamin D in Foods and Nutritional Supplements" during the "Standards Development and International Harmonization: AOAC INTERNATIONAL Mid-Year Meeting," held on June 29, 2011 (2). After evaluation of the validation data submitted, an ERP determined that the method met the standard method performance requirements (SMPRs), as articulated by the Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN).…”

Mechanism of error caused by isotope-labeled internal standard: accurate method for simultaneous measurement of vitamin D and pre-vitamin D by liquid chromatography/tandem mass spectrometry

“…High‐performance liquid chromatography (HPLC) has been widely used for the measurement of vitamin D . However, with high sensitivity and high selectivity, liquid chromatography/mass spectrometry (LC/MS) has become more popular for vitamin D measurement recently . Heudi et al .…”

“…synthesized and used an isotope‐labeled vitamin D 3 to measure fat‐soluble vitamins in human breast milk. Recently, several authors have used deuterium‐labeled internal standards to measure vitamin D in a variety of foods and supplements by LC/MS/MS after a variety of sample preparation methods including overnight saponification at room temperature (25 °C), hot saponification, or direct extraction of vitamin D in infant formula followed by derivatisation …”