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United States Department of Agriculture

Agricultural Research Service


Location: Food Composition and Methods Development Laboratory

2010 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
Methods for water-soluble vitamins (WSVs) were developed based on LC with fluorescence and mass spectrometric detection (LC-F and LC-MS) and with stable isotope dilution with MS detection (IDMS). These methods were used to provide data for WSVs in Standard Reference Material (SRM) 1849 – Infant/Adult Nutritional Formula and SRM 3233 – Fortified Cereal. Multi-lab validation of the methods was achieved by comparing the results from FCMDL with those from NIST and other participating labs. Results from the LC-F method are being used in on-going research to develop a method to analyze the WSVs without LC separation. Methods for vitamin B12 and vitamin B6 were developed based on the WSV methods. The vitamin B6 method was selected by AOAC INTERNATIONAL for development as a Single Lab Validated Method (see 1235-52000-059-01R). FCMDL also compared a variety of extraction techniques for vitamins in dietary supplements and fortified cereals. Optimized extraction methods were used to analyze WSVs in series of vitamin-fortified cereal samples that were collected regionally. Initial results show that almost all cereals had vitamin levels significantly higher than label claim values, in line with the general manufacturing practice of over-fortifying to provide adequate shelf life. Development of methods and acquisition of data for vitamin D was extended to a second year. Improvements included: 1) a three-point calibration curve, 2) two columns in series, and 3) use of more potassium hydroxide in the initial step for a more thorough digestion of the sample. These improvements enhanced accuracy and eliminated interferences. This method was used to analyze seafood samples for the National Nutrient Database. The vitamin D method was further modified for the analysis of vitamin D2 in mushrooms. This method allowed both Vitamin D3 and D2 to be determined simultaneously. New efforts are underway to incorporate the analysis of 25-hydroxy vitamin D3, present in meats, to provide additional information on active forms of vitamin D in foods. A third modification to the vitamin D method, a “dilute-and-shoot” method, was developed for determination of vitamin D and triacylglycerols in gel cap dietary supplements. The saponification and extraction steps were completely eliminated, reducing sample preparation time and allowing quantification of intact triacylglycerols (normally be decomposed by saponification). The new method is being used to quantify vitamin D in numerous 1000 IU, 2000 IU, and 5000 IU gel cap dietary supplements. FCMDL scientists participated in advisory groups and expert committees on methods and metrology related to vitamins in foods, planned and participated in the ODS workshop on evaluation of vitamin methods for dietary supplements, were invited to the First International Vitamin Symposium and to the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN), and one was elected to the USP Expert Committee on Dietary Supplements.

4. Accomplishments

Review Publications
Chen, P., Wolf, W.R., Castanheira, I., Sanches-Silva, A. 2010. A LC/UV/Vis method for determination of cyanocobalamin in multivitamin dietary supplements with on-line sample clean-up. Analytical Methods. 2:1171-1175.

Byrdwell, W.C. 2010. Dual parallel mass spectrometry for lipid and vitamin D analysis. Journal of Chromatography A. 1217(25):3992-4003.

Goldschmidt, R.J., Wolf, W.R. 2010. Simultaneous determination of water-soluble vitamins in SRM 1849 Infant/Adult Nutritional Formula powder by liquid chromatography–isotope dilution mass spectrometry. Analytical and Bioanalytical Chemistry. 397(2):471-481.

Wolf, W.R. 2010. History of reference materials for food and nutrition metrology: As represented in the series of BERM Symposia. Analytical and Bioanalytical Chemistry. 397(2):413-421.

Last Modified: 10/19/2017
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