Performing Department
Food Science
Non Technical Summary
Thiamin deficiencies impact populations in both developed and developing countries. In many cases, fortifying staple refined foods with thiamin can mitigate the deficiencies. To optimize the delivery of thiamin, it must be stable in the product throughout storage and use. Current US consumer demands (and therefore industry trends) include increasing both whole-grain (whole-food) and gluten-free products. More information is needed to document the impact of thiamin ingredient form, food matrix, and processing and storage conditions on the stability of thiamin throughout shelf-life in a variety of food products beyond refined and enriched wheat-based foods. The proposed study will document the magnitude of thiamin losses through production and storage scenarios, encompassing a variety of thiamin ingredient forms, model systems, staple refined foods (based on wheat, corn, and rice), and food products (including gluten-free). Not only will this information be useful to compare thiamin stability in whole and fortified foods made from staple crops throughout the world, but it will also be useful to support thiamin-containing gluten-free product development efforts. The ultimate goal is to improve the delivery of thiamin in a variety of foods to promote a safe (interpret this as gluten-free for Celiac patients), sufficient, and nutritious food supply for both developed and developing nations. This project will address the USDA program area priority in Program Area A1361 "Improving Food Quality" by comprehensively investigating thiamin stability in natural and synthetic ingredients and whole and fortified foods and then developing recommendations to stabilize and optimize the delivery of thiamin in both ingredients and foods. While research on thiamin stability is available in the literature, we believe that ours is a novel approach, combining an applied characterization of thiamin content and stability in foods and ingredients, including investigations of gluten-free products, with a fundamental extension of the latest in scientific advances on solid state architecture from the pharmaceutical arena to whole food applications. In particular, our proposed research has the following novel features: 1) A systematic investigation of natural and synthetic as well as crystalline and amorphous thiamin structures; 2) Development of a mechanistic understanding of how common food ingredients (starch, proteins, and gums) disrupt the molecular assembly of thiamin additives, resulting in amorphous structures with altered physical and chemical stability; 3) A direct comparison of the stability of natural and synthetic thiamin forms in food and model systems across relevant production and storage scenarios; and 4) An understanding of the synergistic or antagonistic formulation/process/environmental interactions on the kinetics of thiamin degradation through processing and storage of foods. The Specific Objectives are designed to enhance the fundamental understanding of the impacts of vitamin form, formulation, solid state properties, and storage treatment interactions on thiamin stability and to develop recommendations for improving the stability of thiamin in foods. The Potential Impact and Expected Outcomes of the fundamental new knowledge we will generate in manipulating thiamin ingredients in crystalline forms and amorphous dispersions include the development of recommendations and implementation of solid state strategies to enhance thiamin additive stability in both ingredients and foods. Scientifically sound recommendations developed from this work will enable selection of the optimal form of thiamin for different products, with the potential to improve delivery of thiamin in a wide variety of products and ultimately reduce rates of thiamin deficiencies in both developed and developing countries.
Animal Health Component
0%
Research Effort Categories
Basic
90%
Applied
10%
Developmental
(N/A)
Goals / Objectives
The long-term goal of this interdisciplinary effort is to improve the delivery of thiamin in whole foods and food ingredients. The objectives of this project are designed to enhance the fundamental understanding of the impacts of vitamin form, formulation, solid state properties, and storage treatment interactions on thiamin stability. Thiamin degradation will be modeled, and the optimal form of thiamin for different products will be identified based on the interplay between physical and chemical stability across formulation, production, and storage scenarios. The central hypothesis is that different thiamin forms will exhibit different stability traits, including response to food formulation and production scenarios, and therefore recommendations can be developed for selecting the optimum thiamin form for a particular product or process. The supporting objectives are to:Monitor the stability of thiamin (natural and synthetic forms, including different crystalline forms) through the production and storage of whole and refined grain flours and products (wheat, rice, corn) and model food systems, and identify formulation, matrix (crystalline and amorphous systems), processing, and storage factors (e.g., temperature and relative humidity, RH) that have a significant impact on thiamin degradation kinetics.Develop mathematical models and elucidate reaction constants that can be used to predict thiamin degradation in food ingredients and products based on the type of ingredient/food, process, storage conditions, and other significant factors identified in Objective 1, and document interactions between thiamin and food ingredients, as a basis for recommendations for the optimal form of thiamin for different ingredients, foods, and use scenarios.
Project Methods
Sample preparation for food systems: The stability of natural forms of thiamin will be determined in three staple grains (hard white wheat, dent corn, and brown rice) and their products, as well as in brewer's yeast. Each grain will be evaluated as a whole kernel, whole grain flour, refined flour, the separated bran and germ, and macronutrient fractions (starch, protein).Allalsoexcept the whole grain kernel will also be fortified with thiamin HCl and thiamin mononitrate (separately). Both physical blends and solid dispersions (lyophilized) will be studied.To compare thiamin degradation kinetics in a processed food product, the fortified and unfortified whole grain flour and refined flours from wheat, corn, and rice will be made into a standard cracker formulation following procedures identified in this reference. For the investigation of different crystalline salts, we will evaluate acids with pKa values lower than the pKa value of thiamin.Thiamin free base will be dissolved in aqueous ethanol and the counterion will be added in the appropriate stoichiometric ratio. A miscible antisolvent will be added to reduce the solubility of the formed salt, and the solution will be left to crystallize. Following formation of a crystalline solid, the XRPD pattern will be obtained to confirm the formation a unique crystal form. Solution state nuclear magnetic resonance will be used to confirm the stoichiometry of the salts. Infrared spectroscopy will be used to confirm proton transfer. The physicochemical properties of the salts will then be determined and will include measurement of melting point (and Tg) using DSC, hydration state using TGA, aqueous solubility determination using HPLC, and characterization of hygroscopicity using dynamic vapor sorption analysis. Only anhydrous salts with a low total moisture sorption over the range 0-95% RH will be further evaluated. All synthetic thiamin forms will also be studied in solutions stored at different temperatures (4-80°C) to document dissolution, solubility, and chemical stability over time.Cocrystal formation will be investigated with thiamin using the solvent assisted cogrinding method. Essentially, thiamin and the coformer are blended together in different stoichiometric ratios with a small amount of water and milled together. Following cogrinding, samples will be dried at ambient temperature and then XRPD will be used to seek evidence of cocrystal formation. Following identification of successful cocrystal reactions, the resultant cocrystals will be characterized as described above for the salts. For investigating amorphous thiamin forms, different synthetic thiamin forms (HCl, mononitrate, other salts) will be converted to the amorphous state using primarily lyophilization, with select spin coating or cryomilling to explore other dispersion techniques, following procedures we have described in more detail elsewhere. The Tg of different solid state forms of thiamin alone will be evaluated and related to the Tm. Different polymers will be used to explore factors that influence amorphization of thiamin. The resultant samples will be subjected to a number of solid state analyses in order to confirm the amorphous nature of the system, determine the mixture Tg and study the intermolecular thiamin-polymer interactions. The amount of polymer required to produce an amorphous system will be evaluated. Following initial characterization, samples will be stored at select temperature and RH conditions. Crystallization kinetics will be monitored using XRPD. A calibration curve will be constructed and used to determine % crystallinity as a function of time. Crystallization kinetics will be modeled using the Avrami equation and further evaluated by considering T-Tg, whereby Tg is expected to vary as a function of both polymer content and RH. Control samples will comprise the crystalline material alone and mixed with the polymer at the same ratios as used in the dispersions, stored under the same conditions. Additional chemical and physical stability studies, described below, will be undertaken as a function of time and storage conditions (temperature, RH). Storage treatments: All control, fortified, and model samples will be stored in the dark in environmental chambers at 4-80°C in desiccators above saturated salt solutions (0-97%RH), and solutions will be stored in nitrogen-flushed sealed glass vials at 4-80°C. Samples will be prepared so triplicates are available at each time point, with evaluations at least every week for the first 8 weeks (more frequently at higher temperatures and in solution), then once a month up to 6 months, and then every 2 months until 24 months is reached. Adjustments will be made as needed to ensure a minimum of 10 data points collected over time at each condition, leading up to 50% thiamin degradation, to support kinetic modeling.Chemical stability: The effects of formulation, treatments, and storage conditions on the chemical stability of thiamin over time will be determined in all food and model systems. Following the selected sequence of treatments, established HPLC techniques for analyzing thiamin in use in our laboratories will be adapted. Thiamin will be verified by the elution time of standards and quantitated by the corresponding multilevel calibration curves. The microenvironmental pH of each formulation will also be documented.Sensory analysis: A series of Duo-Trio balanced design difference tests will be conducted over time to determine whether or not an aroma difference exists between samples based on thiamin form (crystalline and amorphous, different crystalline forms, different polymer dispersion types) or thiamin degradation (after exposure to different storage treatments).Physical properties: The solid state properties of the thiamin forms and formulations will be determined by a number of methods commonly applied in our laboratories. Crystallinity and polymorphism will be determined by obtaining XRPD patterns. Tm or degradation temperature will be determined by hot stage microscopy in combination with TGA and DSC. DSC measurements will also be used to determine the Tg of amorphous systems, and thereby to evaluate the effect of formulation and absorbed moisture on Tg. Moisture sorption profiles of all samples will be generated using automated moisture sorption balances. The solubility of different thiamin forms and preparations will be determined by equilibrating with buffer of known pH, separation of the solid from the supernatant, followed by analyzing the supernatant with HPLC.Methematical Models: Mathematical models and reaction constants will be established that can be used to predict thiamin degradation in a variety of food systems. Degradation kinetics and kinetics models as a function of thiamin form, food formulation, process, and environmental factors will be developed, and interaction extent between thiamin and food ingredients will be quantified using multivariate modeling. Various kinetic models have been used to describe chemical degradation and these will be used to model thiamin degradation in our model systems and foods, including the Arrhenius , Prout-Tompkins, Avrami, Bawn, and Weibull equations. Using either mechanistic or empirical equations, reaction rate constants will be evaluated for the various food and model systems. Multivariate analysis will be used to determine synergistic and antagonistic effects between solid state form, formulation, and RH. Following this analysis, the mechanism of the enhanced degradation will be further elucidated by linking the reaction rates to physical property determinations described above.