2007 Annual Report 1a.Objectives (from AD-416) To develop new analytical methods for water-soluble vitamins and vitamin D to replace existing, outdated methods. 1b.Approach (from AD-416) For the water-soluble vitamins, use liquid chromatography-mass spectrometry for optimized individual methods, multiple vitamin methods, and stable isotope dilution methods. For vitamin D, use liquid chromatography-mass spectrometry for an optimized individual method and a stable isotope dilution method. 3.Progress Report Development of a multivitamin method for seven water soluble vitamins in dietary supplements was completed and the manuscripts were accepted for publication. The new multivitamin method was used to assist in the characterization the new Multi-Vitamin Standard Reference Material 3280, which was jointly developed with National Institute of Standards and Technology and the Office of Dietary Supplements at the National Institutes of Health. This method is also being extended to fortified foods. An isotope dilution-liquid chromatography/mass spectrometric (ID-LC/MS) method for the analysis of individual water soluble vitamins in dietary supplements was completed and a manuscript was accepted for publication. Early availability of multiple isotope-labeled vitamins in FY 2005 was essential to the rapid progress made towards these milestones. The systematic development of individual ID-LC/MS methods for the determination of water soluble vitamins (thiamine, niacin, pyridoxine, folic acid, biotin, pantothenic acid, and ascorbic acid) in foods has been initiated. Two methods from the Association of Official Analytical Chemists (AOAC) (Method 2002.05 and Method 992.26) were evaluated for their usefulness in measuring vitamin D for a wide range of food sample matrices. Each method was found to have certain strengths, but also weaknesses. The best aspects of the two AOAC methods were incorporated into a single method that has been found to be effective for the wide range of samples matrices tested to date. Conditions for the two chromatographic separations in the method and for atmospheric pressure chemical ionization mass spectrometry (APCI-MS) of vitamin D have been optimized. The Food Composition and Methods Laboratory (FCML) participated in a round robin collaborative study for the analysis of vitamin D in five representative foods: skim milk, multi-grain cereal, orange juice, Alaskan red sockeye salmon, and processed cheese food, as well as a diluted vitamin D standard from the U.S. Pharmacopoiea. The analytical results from FCML were consistently close to the average and demonstrated the Lab’s ability to analyze samples for the National Nutrient Databank. Given adequate resources, FCML is now prepared to undertake analysis of samples obtained through the Nutrient Data Laboratory. It was determined that the vitamin D provided by commercial chemical suppliers is not in the chemical form defined as vitamin D, but is a precursor. The vitamin D from commercial suppliers undergoes a transformation during sample preparation and reverts to the precursor structure. This is the structural form that is commonly analyzed and reported. The results of this research have been presented in seminars given to the Office of Dietary Supplements (ODS), to the Beltsville Human Nutrition Research Center, and to the Food and Drug Administration. Results were also presented at the annual meeting of Experimental Biology (April 2007). A manuscript is being prepared. A second manuscript describing the results on the round robin analysis of the representative foods is also in preparation. 4.Accomplishments Simultaneous Determination of Multiple Water-Soluble Vitamins: Expediting the analysis of vitamins in foods is necessary to allow the USDA Nutrient Database to keep up with the changing food supply and to allow the Food Composition and Methods Laboratory (FCML) to support the development of vitamin reference materials. A method has been developed for the simultaneous determination of niacin, thiamine, pyridoxine, riboflavin, folic acid, biotin, pantothenic acid, and ascorbic acid in dietary supplements. This method uses reversed phase liquid chromatography (LC) and detection with a combination of photodiode array/fluorescence in tandem with mass spectrometry. Single laboratory method validation of this method has been conducted and the results are being prepared for publication. Having a simple, single-run LC method for determination of multiple vitamins in dietary supplements will have a significant impact on validating the capability of commercial laboratories to generate data to populate the USDA Dietary Supplement Ingredient Database currently being assembled in collaboration with the Office of Dietary Supplements (National Institutes of Health) and the Beltsville Nutrient Data Laboratory (ARS). This accomplishment addresses National Program 107 Component 1: Composition of Foods. 5.Significant Activities that Support Special Target Populations None 6.Technology Transfer Number of non-peer reviewed presentations and proceedings 7 Review Publications Chen, P., Wolf, W.R. 2006. LC/UV/MS-MRM for the simultaneous determination of water-soluble vitamins in multi-vitamin dietary supplements. Analytical and Bioanalytical Chemistry, 387(7):2441-2448. Phillips, K.M., Wolf, W.R., Patterson, K.K., Sharpless, K.E., Amanna, K.R., Holden, J.M. 2007. Quality and performance evaluation of reference materials for the determination of nutrients in foods. Accreditation and Quality Assurance: Journal for Quality, Comparability and Reliability in Chemical Measurement. 12(3-4):126-133. Goldschmidt, R.J., Wolf, W.R. 2007. Determination of niacin in food materials by isotope dilution mass spectrometry. Journal of Association of Official Analytical Chemists International. 90(4):1080-1089. Chen, P., Ozcan, M., Wolf, W.R. 2007. Simultaneous determination of selected B vitamins in the NIST SRM 3280 multivitamin/multielement tablets by liquid chromatography isotope dilution mass spectrometry. Analytical and Bioanalytical Chemistry. Available: doi:10.1007/s00216-007-1430-9. Phillips, K., Wolf, W.R., Sharpless, K., Holden, J.M. 2007. Reference materials for determination of the nutrient composition of foods: Results from USDA's National Food and Nutrient Analysis Program (NFNAP). Analytical and Bioanalytical Chemistry. Available: doi:10.2007/s00216-007-1366-0. Roseland, J.M., Holden, J.M., Andrews, K., Zhao, L., Schweitzer, A., Harnly, J.M., Wolf, W.R., Perry, C., Dwyer, J., Picciano, M., Betz, J., Saldanha, L., Yetley, E., Fisher, K., Sharpless, K. 2007. Dietary Supplement Ingredient Database (DSID): Preliminary USDA studies on composition of adult multivitamin/mineral supplements. Journal of Food Composition and Analysis. 21(1): S69-S77. Available: doi:10.1016/j.fca.2007.07.009. Dwyer, J.T., Picciano, M., Betz, J.M., Fisher, K.D., Saldanha, L.G., Yetley, E.A., Coates, P.M., Milner, J., Whitted, J., Burt, V., Radimer, K., Wilger, J., Sharpless, K.E., Holden, J.M., Schweitzer, A., Andrews, K., Roseland, J.M., Zhao, C., Harnly, J.M., Wolf, W.R., Perry, C.R. 2007. Progress in developing dietary supplement databases: The analytically validated Dietary Supplement Ingredient Database (DSID) and Dietary Supplement Label Databases (DSLD). Journal of Food Composition and Analysis. Available: doi:10.1016/j.jfca.2007.07.010.