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United States Department of Agriculture

Agricultural Research Service

Research Project: MEASUREMENT SYSTEMS FOR DETERMINATION OF VITAMINS IN FOODS AND DIETARY SUPPLEMENTS

Location: Food Composition and Methods Development Lab

2011 Annual Report


1a.Objectives (from AD-416)
Objective 1: Evaluate existing measurement systems and determine the need for improved measurements for water and lipid soluble vitamins (e.g., vitamins D and E and selected B vitamins), for which significant public health concern and inadequate composition data exists in foods and dietary supplements. Sub-Objective 1.A: Systematically evaluate public health concerns and adequacy of food composition data for vitamins and matrices to prioritize improved measurement needs, including methods and available Reference Materials. Sub-Objective 1.B. Develop detailed evaluations of measurement systems for priority analytes and matrices. Sub-Objective 1.C. Develop specific purpose statements for development and/or updating of methods for measurements of single or multiple vitamins.

Objective 2: Develop and validate new or updated analytical methods using current technology to determine the levels of water-soluble vitamins (WSV), lipid soluble vitamins (LSV) and/or other components for foods and dietary supplements. Sub-Objective 2.A: Develop/update and optimize measurement procedures to establish validated capability for simultaneous measurements of multiple WSV (SimWSV). Sub-Objective 2.B: Develop and validate analytical methods for the determination and quantification of LSV (A, D, E, and K) and lipids in food matrices and dietary supplements. Sub-Objective 2.C Develop/update and optimize measurement procedures to establish validated capability for measurements of vitamin B12 in dietary supplements and foods. Sub-Objective 2.D Develop multivariate calibration methods for simultaneous determination of multiple vitamins in extracts with no prior chromatographic separations.

Objective 3: Develop and validate sample preparation procedures to optimize extraction, remove interferences, and/or to concentrate difficult to analyze vitamins in foods and dietary supplements.

Objective 4: Catalyze cooperative activities to identify and provide improved measurement systems and essential Reference Materials for vitamins in foods and dietary supplements. Provide analytical data to characterize the vitamin content of selected Reference Materials. Sub-Objective 4.A: Generate information with developed and validated methods to assign value-added information on vitamin content to available Reference Materials. Sub-Objective 4.B: Catalyze development of overall measurement systems for vitamins in foods and dietary supplements.


1b.Approach (from AD-416)
Objective 1: Evidence-based reviews will provide priorities for specific vitamins, define adequacy of existing data, and define quality of data required for future needs. The present state of the performance of individual labs and the adequacy of analysis of specific vitamin measurement systems will be evaluated from extensive data available from the USDA contract analyses conducted as part of NFNAP and DSID. Clearly defined purpose statements will be developed for specific applications. Objective 2: Improved procedures will be developed to simultaneously measure water soluble vitamins (SIMWSV) [thiamin, riboflavin, niacin, pyridoxine, folic acid, pantothenic acid, biotin, choline, and ascorbic acid] in foods and dietary supplements (DS). SIMWSV and LC-IDMS methods for DS will be extended to fortified foods. The additional challenges of natural levels of vitamins in unfortified foods require different approaches or compromise conditions to obtain acceptable analytical results. We will examine newer chromatographic separation modes such as hydrophilic interaction liquid chromatography (HILIC) and aqueous normal phase (ANP) chromatography to improve these separations. A defined protocol of intra-laboratory validation will be carried out following AOAC guidelines. The FCMDLVitamin D method is implemented for analysis of food samples, and additional foods, dietary supplements and reference materials will be analyzed. IDMS methods will be initiated for other lipid soluble vitamins A, E, and K. The FCMDL method to determine the low levels of B12 in vitamin supplements using dual column LC/UV will be extended to fortified foods using sensitive LC/MS techniques and collaboratively cross-validated to the microbiological method. The possibility to calibrate spectral fingerprints (information with no chromatographic separation) of extracts of various types of food and dietary supplement materials to obtain quantitative information about vitamin content will be explored. Calibration models will be developed and validated.

Objective 3: Multiple extractions for WSV with different buffers, pHs, and multiple extraction approaches (including classical and modern methods such as pressurized liquid extraction [PLE], microwave-assisted extraction [MAE], ultrasonic irradiation, stirring, shaking, and Soxhlet) will be systematically explored to ensure complete extraction and compare extraction efficiencies of different procedures. The lipid soluble vitamins (LSV) (A, D, E, K) would be similarily extracted with a organic solvents of different polarities.

Objective 4: FCMDL capability for high quality vitamin determinations will be applied to provide reference measurements to value to NIST SRMs such as the Adult/Infant Formula SRM and Fortified Cereal SRM. Through initiating and providing guidance for a number of nutrition metrology-related activities, FCMDL will catalyze improvement of the overall measurement system for vitamins. FCMDL will participate as collaborators in method validation studies as appropriate. FCMDL will continue to organize and advise the development and conduct of symposia, and other appropriate workshops.


3.Progress Report
A method was developed for the determination of vitamin B6 in vitamin and mineral supplements. A general extraction method was used that was modified slightly depending on whether the supplement was a tablet, soft gel, liquid, sustained release material, or a “gummy-bear” material. Chromatographic separation was based on both high performance liquid chromatography (HPLC) and ultra-HPLC (U-HPLC). U-HPLC separation was accomplished in less than 20 minutes. Vitamin B6 was detected using ultraviolet absorption, fluorescence, and mass spectrometry. Accuracy and precision of ±5% were achieved for all supplement materials.

A method was developed for the determination of vitamin D, as well as the excipient oil in dietary supplement gelcaps. This "dilute and shoot" method eliminates the labor- and resource-intensive sample pretreatment typically required for vitamin D analysis. The method uses six types of detectors, including ultraviolet detection, and three types of mass spectrometry, which provide not only quantification of vitamin D3 but also characterization of the triacylglycerols (TAGs) in the oil, and identification of TAG oxidation products and glycerin polyglycerols.


4.Accomplishments
1. Determination of Vitamin D in orange juice. Levels of vitamin D in natural and fortified foods have been questioned because of the inadequacy of the analytical methods. Levels in orange juice are particularly suspect because the fat-soluble vitamin D is incorporated into a water matrix. In this work, samples of vitamin D-fortified orange juice obtained from retail food stores were analyzed for vitamin D3 content using a method developed by combining the best features of two AOAC methods. The method was applied to 40 samples of American orange juice brands collected using a statistically designed sampling plan as part of the National Food and Nutrient Analysis Program. Vitamin D3 values ranged from 1.071 µg/100 g (43 IU/100 g) to 1.663 µg/100 g (67 IU/100 g), with an average across 55 samples analyzed, including duplicates, of 1.4 ± 0.1 µg/100 g (57 ± 5 IU/100 g, indicating that a typical 8 oz. (240 mL = 240 cm3) glass of orange juice provided 3.4 ± 0.3 µg (140 ± 10 IU) vitamin D3. This work provided analytical values for vitamin D to the USDA National Nutrient Databank for Standard Reference, used in estimating intakes of vitamin D for the Institute of Medicine Dietary Reference Intakes Committee review of Vitamin D reference values.


Review Publications
Patterson, K.K., Exler, J., Byrdwell, W.C., Phillips, K.M., Horst, R., Lemar, L.E., Holden, J.M. 2010. Vitamin D content and variability in fluid milk from a USDA nationwide sampling to update values in the National Nutrient Database for Standard Reference. Journal of Dairy Science. 93(11):5082-5090.

Phillips, K.M., Ruggio, D.M., Horst, R.L., Minor, B., Simon, R., Feeney, M., Byrdwell, W.C., Haytowitz, D.B. 2011. Vitamin D and sterol composition of ten types of mushrooms from retail suppliers in the United States. Journal of Agricultural and Food Chemistry. 59(14):7841-7853.

Byrdwell, W.C., Exler, J., Gebhardt, S.E., Harnly, J.M., Holden, J.M., Horst, R., Patterson, K.K., Phillips, K., Wolf, W.R. 2010. Ultraviolet and mass spectrometric analysis of vitamin D in retail fortified orange juice. Journal of Food Composition and Analysis. 24:299-306.

Last Modified: 8/22/2014
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