SOYBEAN DIETARY FIBER: A FUNCTIONAL INGREDIENT FOR PREVENTION OF CHILDHOOD OBESITY

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: TERMINATED
• Funding Source: EVANS-ALLEN
• Reporting Frequency: Annual
• Accession No.: 0232322
• Project No.: TENX-1301-CO
• Project Start Date: Jan 1, 2013
• Project End Date: Sep 30, 2016

Project Director
Wu, YI.

Recipient Organization
TENNESSEE STATE UNIVERSITY
3500 JOHN A. MERRITT BLVD
NASHVILLE,TN 37209

Performing Department
Agricultural and Environmental Sciences

Non Technical Summary
Eating patterns and nutritional status are closely related to health. In the recent decades, the growing prevalence of overweight and obesity at early stage of childhood is a serious concern (Hill and Trowbridge, 1998). Obesity is the third most dominant risk factor for noncommunicable diseases such as heart disease in high-income countries (Lopez, Mathers, Ezzati, Jamison, & Murray, 2006). According to the data published by National Health and Nutrition Examination Survey (Ogden et al., 2012), in the year 2009-2010, the obesity rate was 16.9 percent among children and adolescents in the United States. Due to the steep increase of overweight and obesity rate and its related health risks, childhood obesity has become one of the most pressing public health concerns in the country (Koplan et al., 2005). Diet is one of the important factors considered as causes of childhood obesity (Ebbeling et al., 2002). Important epidemiological evidence support the concept that diets rich in fiber are associated with lower body weight or weight gain (Lairon, 2007). High fiber diets are important in the prevention and management of obesity and chronic diseases, including type2 diabetes, heart disease and cancer (Kendall et al., 2010). Dietary guidance universally recommends diets higher in fiber for improved health benefits (Slavin, 2005). Current recommendations from the United States National Academy of Sciences, Institute of Medicine, suggest that adults should consume 20-35 grams of dietary fiber per day, but the average American's daily intake of dietary fiber is only 12-18 grams. Similarly, the Academy of Nutrition and Dietetics (AND) also recommends a minimum of 20-35 g/day for a healthy adult. The recommended amount for children is the age plus 5 g/day (e.g., a 4 year old should consume 9 g/day). There are emerging needs for food products with higher dietary content which are readily available to consumers especially at younger ages. However, incorporation of fiber ingredients to food products is challenging due to the adverse effect on texture and sensory quality of the products. In this project, fractions of dietary fiber obtained from soybeans will be added into bread at various concentrations. In vitro and in vivo studies will be carried out to evaluate physiological effects of different fractions vs. concentration. Starch digestion rate and glucose releasing rate will be compared among treatments. Sensory evaluation on breads will be conducted for consumer acceptance. This project will provide healthier bread choices with higher dietary fiber content for consumers to prevent obesity at younger ages.

Animal Health Component

0%

Research Effort Categories

Basic

40%

Applied

30%

Developmental

30%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
502	1820	1010	100%

Knowledge Area
502 - New and Improved Food Products;

Subject Of Investigation
1820 - Soybean;

Field Of Science
1010 - Nutrition and metabolism;

Keywords

dietary fiber

obesity prevention

soybean polysaccharides

bread.

Goals / Objectives
The scope of the current project is to add value from farm to forks for the local and national wide agricultural economy, as well as bring a positive impact by using the agricultural byproduct for obesity prevention. Efforts were made to develop dietary fortified foods using various resources as dietary fiber ingredients. The State of Tennessee produces an abundance of foodstuffs suitable for dietary fibers. According to the report from Tennessee Department of Agriculture (Tennessee AG Insider, TNagriculture.com, 2012), Tennessee's top crops are soybeans, corn, cotton, tobacco and wheat. Based on the data published in U.S. Environmental Protection Agency (2012), corn and soybeans are the largest crops produced in the United States as well. Both corn bran and soybean polysaccharides are well recognized as sources of dietary fiber. Soybeans polysaccharides are agriculture byproducts and worth little value by themselves. However, after their potential health benefits are recognized and successful applied into food, the byproducts could bring more profit into the food production chain. The project will investigate physiological roll of soybean polysaccharides as dietary fiber. Chemical, physical and functional properties of soy polysaccharides have been investigated extensively in recent years. However, their physiological properties as dietary fiber in food products have not been well documented. This research will bring more scientific evidence for the beneficial rolls of different fractions of soybean polysaccharides as a source of dietary fiber to prevent obesity and other diseases. This study will provide information on the optimum combination of fraction and concentration of soy fibers for making good quality bread products with enhanced physiological effects. This information can be used as an industrial guide for bread production. Farmers and food processers can also benefit from the increased use of the agricultural byproduct to produce value-added food. The fully developed dietary-fortified foods would be available to consumers at younger ages to achieve prevention-based health outcomes in finding new ways to reduce childhood obesity. The objectives of the present study are: (1) evaluate the effects of soy fiber-fortified bread on obesity prevention; (2) compare the functional and physiological effects of the three soy fiber fractions, total, insoluble and soluble fractions and; (3) compare the physiological effects of soy fiber fractions with a FAO approved soluble dietary fiber, oat β-glucan and evaluate the effects of the two different fibers; (4) sensory evaluation will be conducted on the bread prepared from different soy fiber fractions.

Project Methods
1. Preparation of fiber fractions from soybean would follow the method described by Makamura et al. (2001) with slight modification. 2. Incorporation of dietary fiber fractions into food product: bread. The basic recipe is adopted from Cadioli et al. (2011) who conducted sensory evaluation after adding soy protein and fiber into the bread. Bread is freshly prepared using a bread maker. 3. Quantification of soluble and insoluble dietary fiber: A commercial kit, Megazyme Total Dietary Fiber, will be used for this analysis. This kit is based on the method of AOAC 991.43, AOAC 985.29, AACC 32-07.01 and AACC 32-05.01. 4. Examination of physical properties: viscosity of samples/digesta would be investigated on an ARES rheometer (TA Instruments). Emulsification properties (particle size and zeta-potential) of samples would be evaluated using Zetasizer Nano ZS (Malvern Instruments). 5. In vitro study of glucose/oil release: This study will follow the procedure described by Roberts (2012), which was developed based on the methods of Brighenti et al. (1995) and Brennan et al. (2004) with minor modifications. 6. Sensory evaluation of breads: The sensory evaluation will be carried out on the breads 12 h after baking as described by Chlopicka et al., 2012. More than 20 people will be recruited for evaluation of the samples. The scorecard will be with 10-point category scale (disliked = 0; extremely liked = 10), each testers will be asked to assess the breads for overall quality, based on the color, odor, consistency and taste of bread. The provided choices include: interesting, tasty, natural, strange taste, vapid, not to eat, bad taste, gummy, crusty, little pronounced, sweet, salty, tart, difficult to determine, delicate, milky. The possible differences between various breads will be analyzed using Kruskal-Wallis test.

Progress 01/01/13 to 09/30/16

Outputs
Target Audience:Food Scientist, Bakery Industry, Geneal Public Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This projecthas involved many undergraduate and graduate students (totally 5) from the beginning of the experiment, starting from extraction, receipe development, bread making, quality assessment, as well as in vitro digestion of the fiber-enriched bread. These students were trained with lab skills in Food Chemistry, Product Development and Evaluation, as well as skills in biochemistry and cell culture skills. How have the results been disseminated to communities of interest?1. Publication in peer-reviewed journals (1 published review paper and one original research manuscript) 2. Presentations in symposiums and conferences 3. Classroom discussions with undergraduate and graduate students 4. Presentation to the high school students in summer programs at campus What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? In the current study, three fractions of dietary fiber, namely soluble, insoluble and total dietary fiber, were extracted from defatted soy meal. Total dietary fiber accounted for 50% of the defatted soy meal. The total dietary fiber is consisted of 7% soluble dietary fiber and 43% insoluble fiber. Study 1. Impact of different fiber fractions on bread quality Three fiber fractions were incorporated into bread to replace wheat flour, and to achieve 5% and 10% of soy fiber in bread. A control bread with 0% of soy fiber was used as a comparison. The study on bread volume showed that the 5% and 10% soluble fiber enriched breads gave the highest bread volume, even higher than that of the control bread. The bread with 5% insoluble fiber exhibited a similar volume with the control bread. When the concentration increased to 10%, the bread volume decreased. The volume of total fiber breads showed lower volume than control bread. However, the 10% total fiber-enriched breads possessed a higher volume than that of 5% total fiber bread. This study showed that 10% soluble fiber may have a positive impact on bread volume. However, the color of dietary fiber-enriched bread is obviously darker than that of the control bread. Although slightly darker than the control bread, soluble fiber-enriched bread exhibited lighter color than that of insoluble and total fiber breads. The darker bread color may have negative impact on bread sensory quality. Additionally, all of the 10% fiber enriched breads had a sticky texture. 2. Effect of different fiber fractions on glucose release and predictive glycemic index (pGI) In vitro digestion model was used to estimate predictive glycemic index (pGI) of the fiber-enriched breads. The in vitro digestion test showed that the higher the fiber contents, the lower the glucose releasing profile. At the same fiber level, soluble fiber showed the best glucose attenuation effect, followed by total fiber. The insoluble fiber-enriched bread exhibited the least glucose attenuation effect. The in vitro hydrolysis rate of starch was plotted over the period of 5 hours with dialysate samples taken every 30 minutes. Area under the curve (AUC) was determined from starch hydrolysis plots of each sample. The hydrolysis index (HI) was calculated as: (AUC Control/AUC Sample) X 100. The predicted glycemic index (pGI) was calculated from HI values using the empiric formula proposed by Granfeldt (1994): pGI = 0.862 HI + 8.198. The predicted Glycemic Index of the breads followed the trend: 0% >5% insoluble > 5% total > 5% soluble >10% insoluble > 10% total > 10% soluble. Therefore, this study revealed that consumption of fiber-enriched bread can lower down the pGI. Soluble soy fiber at 10% exhibited the best result in this study. 3. Effect of fiber-enriched bread on lipid accumulation in cells Anti-obesity effect was evaluated using 3T3-L1 cells (ATCC, Manassas, VA, USA). The cells were cultured in Dulbecco's modified Eagle's medium (DMEM, Gibco) containing 10% fetal bovine serum (FBS, Corning) and 1% penicillin streptomycin. After cells were maintained at confluence for 2 days, cells then cultured with induction medium (day 0) containing 0.5 mM 3-isobutyl-1-methylxanthine, 1 μM dexamethasone and 10 μg/mL insulin DMEM containing 10% FBS and 1% PS. The cultures were continued for 3 days to induce adipocyte differentiation. Thereafter, cells were cultured in DMEM containing 10% FBS for the remainder of the differentiation process. All other treatments including solvent with/without digesta solution of breads after 5 h in vitro digestion were administered from day 0 until day 8 . Cells were stained on day 8 with Oil-red O and measured the absorbance at 490 nm. The results showed that the 5% fiber-enriched bread exhibited no effect on lipid accumulation in cells. However, the 10% soluble fiber-enriched breads can significantly reduce accumulation in cells. Although the 10% total fiber-enriched bread had a lower amount of lipid accumulation but the difference was not significant. Therefore, this study indicated that both fiber type, soluble vs. insoluble, and fiber concentration, 5% vs. 10%, will have effects on lipid accumulation. Conclusion The present study has revealed that soy dietary fiber could be added into bakery products at a certain percentage without exhibiting negative impact on product quality. With a higher soluble fiber content (10%) have resulted in decreased lipid accumulation, while this phenomenon only applied to the soluble fiber, not insoluble or total fiber fractions. However, reduced pGI and glucose releasing profiles were observed in all breads fortified with fiber, and the soluble fiber-enriched bread exhibited the lowest pGI and glucose releasing profile. This study has provided evidence on significant effect of soluble soy dietary fiber on health benefits reading reduced glucose attenuation and lipid droplet accumulation.

Publications

• Type: Journal Articles Status: Published Year Published: 2016 Citation: Perry, J. and Wu, Y.* (2016) A review of physiological effects of soluble and insoluble dietary fibers. Journal of Nutrition and Food Sciences 6:2, http://dx.doi.org/10.4172/2155-9600.1000476

Progress 10/01/14 to 09/30/15

Outputs
Target Audience:Food Scientists, Bakery industry experts, general public Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Undergraduate students and graduate students are trained with skills onbakery processigf, in vitro digestion methods, glucose releasing profile, protein analysis, rheological tests. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?Dough performance, obesity prevention using cell line.

Impacts
What was accomplished under these goals? 1. Three fiber fractions were obtained 2. Products using the three fractions were made. Breads were evaluated. 3. Glucose releasing profile was measuring in vitro using the fiber-enriched bread.

Publications

Progress 10/01/13 to 09/30/14

Outputs
Target Audience: professionals, industry, community and students Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? 2 undergraduate students and 1 freshly graduated undergraduated studentparticipated in this research. They have gained many hands on experiences. How have the results been disseminated to communities of interest? Yes. Presentations were made in University's Research Symposium. High school students were reached to give the knowledge on dietary fiber. They also evaluated the bread made from soy dietary fiber. What do you plan to do during the next reporting period to accomplish the goals? Products based on soy dietary fiber will be evaluated. Other sourse of fibers including beta-glucan, and psyllium gum, will be used in similar products. Physiological parameters will be compared with soy fibers.

Impacts
What was accomplished under these goals? (1) evaluated the effects of soy fiber-fortified bread on glucose attenuation; (2) compared the functional and physiological effects of the three soy fiber fractions, total, insoluble and soluble fractions;

Publications

Progress 01/01/13 to 09/30/13

Outputs
Target Audience: Targeting audiences are professional in food product development sectors food industries, especially in bakery industries with interest producing and developming healthy food products. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? This project has included2 undergraduate students with Food Science Concentration to participate some of the experiments. One of these students showed great interest in persuing a Graduate degree in Food Science. The students has learned from this project about the importance of healthy diet, basic food science lab skills, and the career opportunities for students with Food Science background. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? The plan for the next reporting perios: 1. complete the chemical analysis of each fiber fractions, including protein, ash, total polysaccharides' 2. provide detailed information on polysaccharides in soluble and insoluble fractions including their single sugar composition, molecular weight, viscosity, interactions with wheat starch, and their impact on bread volume. 3. study the physiological benefit of soy fiber enriched bread with emphasis on glycymic index.

Impacts
What was accomplished under these goals? Soy Dietary Fiber Fractions and Bread Quality There are few publications presenting systematic analysis on soybean dietary fiber fractions with specific emphasis on investigating the application of these fractions into final products and on evaluation of the corresponding physiological functions of the final products. In the current study, three soy dietary fiber fractions were extracted, fractionated and incorporated into bread. Bread qualities were compared after different percentage of fiber fractions were incorporated into the breads. Materials and Methods Material preparation: defatted soy flour was purchased from Sigma-Aldrich. Preparation of fiber fractions from soybean would follow the method described by Makamura et al. (2001) with modification. The detailed procedures are provided here: Weigh 200g of soy flour and add 2200 ml dH2O into a 2600 mL crystallizing dish. (the ratio of water to soy flour should be 11:1. Adjustments can be made with varying amounts of soy flour but should also be made correspondingly at step 5 with a 4:1 ratio.) Place the dish on a hotplate and add a stirring rod. Set at 70 °C and 400 RPM for 1 h. Allow mixture to cool to room temperature and carefully pour off liquid (containing soy protein). The soluble and insoluble polysaccharide (PSS) fractions will settle to the bottom of the dish. Rinse the PSS well with dH2O and slowly pour off liquid to remove additional protein. Repeat this step 4 times. Allow the PSS to completely settle in the dish each time (1-2 minutes) to avoid loss of any PSS during each rinse. This is the total extracted PSS containing both soluble and insoluble fractions. Separation of the extracted soluble and insoluble fractions occurs in the following steps. Return the PSS in the dish with stirring rod to the hot plate. Add 800mL dH2O to the mixture. Reduce pH to 3.0 with 5N HCL by adding drop wise with constant stirring at 400RPM while monitoring with pH meter (Jenway 351-101 Model 3510). Set the hotplate at 100 °C and 400 RPM. Place aluminum foil over the dish (puncture in several places to prevent overheating). Once the mixture reaches 100 °C, maintain constant temperature and stirring for 2 hours while closely monitoring. Allow the PSS mixture to cool to room temperature before proceeding. Transfer the PSS mixture to 50 mL centrifuge bottles. Do not fill each bottle more than halfway. After balancing the bottles, centrifuge for 10 minutes at 10,000xG. *Ensure that none of the PSS mixture remains in the bottle caps or necks to ensure full separation of the fractions during centrifugation. Pour the supernatant into a 3500 mL beaker (this is the soluble PSS fraction). Accumulate until all of the PSS mixture has been centrifuged and separated. Remove the remaining insoluble fraction from each centrifuge bottle to an aluminum tray. Place tray in oven at 60°C for overnight. To the supernatant in the 3500 mL beaker (soluble PSS fraction), add enough EtOH to bring the total concentration to 70% EtOH. This will cause the soluble PSS to precipitate from the solution. Allow 12 hours or overnight for the precipitation reaction to be complete. Pour the precipitated soluble PSS into clean 50 mL centrifuge bottles and centrifuge for 10 min at 10,000xG to separate the extracted PSS from the EtOH. Pour the EtOH from each centrifuged bottle into a waste container. Remove as much of the precipitated soluble PSS from each bottle to an aluminum tray. Place tray in 60C oven 12 hours or overnight until completely dry. Collect the dried insoluble and soluble fractions from the oven and weigh respectively. Mill each fraction into fine powder (IKA heavy-Duty Analytical Mill 115VAC/60Hz, 250 mL) and store in an airtight container for future applications. Bread preparation Soy fiber enriched bread was baked in Cuisinart® Stainless Steel Programmable Bread Maker (Model CBK-200). Recipes are listed in Table 1. Table1. Recipe of soy fiber enriched bread 0% Fiber Recipe: 448g (1lb.) 5% Fiber Recipe: 448g (1lb.) 10% Fiber Recipe: 448g (1lb.) Wheat Flour: 393.67g Sugar: 23.62g Salt: 9.45g Vegetable Oil: 15.75g Yeast Powder: 5.51g Total: 448g Water: 472.2mL Wheat Flour: 373.99g Sugar: 22.44g Salt: 8.98g Vegetable Oil: 14.96g Yeast Powder: 5.24g Fiber: 22.39g Total: 448g Water: 472.2mL Wheat Flour: 354.30g Sugar: 21.26g Salt: 8.51g Vegetable Oil: 14.18g Yeast Powder: 4.96g Fiber: 44.79g Total: 448g Water: 472.2mL Bread volume measurement Bread volume will be measured using a fixed volume container. Fixed amount of mustard seeds are used to fill out the whole space of the container. After bread was made and placed at room temperature for 2 h, the bread is placed in the container followed by filling the container with the seeds. The volume of the left over seeds will be measured using a volumetric beaker. This volume will be recorded as the bread volume. Bread appearance evaluation Bread appearance will be compared by cutting the bread in half after it is been placed in room temperature for 2 h. Pictures will be taken to compare the texture and color of the breads. Results Yield of different fiber fractions: The results showed that from each extraction there are about 7% soluble, 43% insoluble obtained from the defatted soy flour. There are totally 50% of total soy fiber fractions in the defatted soy flour. Totally 219.34g of soluble dietary fiber, 1437.17g insoluble dietary fiber and 463.78g total fiber were obtained after sequential fractionation. These fractions were stored for bread preparation. Evaluation of fiber enriched bread Bread volume Three soy fiber fractions, soluble, insoluble and total dietary fiber, were incorporated in the bread with the fiber percentage of 5% and 10% respectively. Bread with 0% fiber will be made as control. The volumes of various breads are presented in Fig. 1. It is interesting to observe that the 5% and 10% soluble fiber enriched breads gave the highest bread volume, even higher than that of the control bread. 5% insoluble fiber bread exhibited similar volume with control bread. When the concentration increased to 10%, the bread volume decreased. The volume of Total fiber breads showed lower volume than control bread. It is surprising to notice that 10% total fiber breads have higher volume that of 5% total fiber bread. It worth further investigation on the mechanism ofbread volume change after incorporating different fiber fractions. This study showed that adding up to 10% soy fiber fractions the bread volume is not greatly impacted. Bread appearance evaluation Pictures of fiber enriched breads are presented in Graph 1. Graph1. Comparison of soy fiber enriched bread with control bread. (a. 5% breads, b. 10% breads. IF: insoluble fiber; SF: soluble fiber; TF: total fiber). Graph 1 showed the texture and color of different fiber enriched breads. As shomn in Graph 1, soluble fiber bread exhibited lighter color than insoluble and total fiber breads. The darker bread color may have negative impact on bread quality. It is also observed that the holes on fiber enriched became larger. Another property observed is the 10% fiber enriched breads have sticker texture, especially the 10% soluble fiber bread. This may have negative impact on bread as well. Future studies The breads were cut into 1.5 mm cubics and dried in 60 C oven, followed by grounded into powder using a grinder. In vitro digestion will be performed to study the relationship between fiber concentration and fraction on glucose releasing profile. Interactions between wheat starch, soluble and insoluble fractions will also be investigated using ARES train-controlled rheometer.

Publications