Oligosaccharides from Rice Bran as Nutraceuticals.

OLIGOSACCHARIDES FROM RICE BRAN AS NUTRACEUTICALS.

Sponsoring Institution

State Agricultural Experiment Station

Project Status

TERMINATED

Funding Source

STATE

Reporting Frequency

Annual

Accession No.

0198614

Grant No.

(N/A)

Project No.

ARK02020

Proposal No.

(N/A)

Multistate No.

(N/A)

Program Code

(N/A)

Project Start Date

Jul 1, 2003

Project End Date

Jun 30, 2006

Grant Year

(N/A)

Project Director
Wang, Y. J.

Recipient Organization
UNIVERSITY OF ARKANSAS
(N/A)
FAYETTEVILLE,AR 72703

Performing Department
FOOD SCIENCE

Non Technical Summary
The health benefits of oligosaccharides from rice bran have not been studied and are expected to be functional prebiotic ingredients with potentials to improve human health. This study will study the composition, structure, and physicochemical properties of rice bran oligosaccharides and to efficiently purify them for future probiotic study.

Animal Health Component

(N/A)

Research Effort Categories

Basic

(N/A)

Applied

100%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
501	1530	2000	35%
502	1530	2000	65%

Knowledge Area
501 - New and Improved Food Processing Technologies; 502 - New and Improved Food Products;

Subject Of Investigation
1530 - Rice;

Field Of Science
2000 - Chemistry;

Goals / Objectives
The U.S. rice production was close to 210 million cwt. (rough rice basis) in 2001 and the state of Arkansas accounts for about 45% of the total rice production. During milling process, about 10% of rough rice is removed as rice bran to produce milled rice. Arkansas produced about 7.5 million cwt. of rice bran in 2001, which is used for rice bran oil manufacture or as a low-valued by-product, e.g. feed. Rice bran contains about 6.5% oligosaccharides, but little research has been conducted to understand the composition of rice bran oligosaccharides and their contribution to human health. Non-digestible oligosaccharides have been claimed to benefit the colon by selectively stimulating the growth of bifidobacteria and by decreasing the toxicity of the colon contents. Oligosaccharides are short-chain saccharides and usually composed of 2-10 glycosidically linked monosaccharides. Human can not digest oligosaccharides because our bodies do not produce the enzymes necessary for their digestion. Instead, they are fermented and consumed by the beneficial intestinal bacteria, bifidobacteria, in the colon. Generally, oligosaccharides are classified as "prebiotics" because they are preferentially chosen by beneficial bacteria, which is termed probiotics, to promote intestinal health, i.e., a balance between the beneficial and harmful microbes. In vitro experiments have shown that fructo-oligosaccharides are not equally well metabolized by all bacterial species. Fructo-oligosaccharides selectively promote the growth of bifidobacteria in the intestine, as they can use the fructo-oligosaccharides better than other intestinal competitors. The presence of fructo-oligosaccharides in the diet results in increased bifidobacterial metabolism. Trials in humans have fructo-oligosaccharides increase the bifidobacteria population to the point where they become the most prevalent intestinal population, to the detriment of other harmful bacteria such as Bacteroides, Clostridia and coliforms. Animal studies in the area of lipid metabolism have shown that the consumption of fructo-oligosaccharides results in a reduction of the total triglycerides and an improvement of the HDL (high-density lipoproteins)/LDL (low-density lipoproteins) ratio. Most of the currently health claims are based on studies of fructo-oligosaccharides. Additional benefits from other sources of oligosaccharides may also exist due to their differences in chemical structures and compositions. Therefore, oligosaccharides from rice bran present a tremendous potential to offer unique health benefits that are not present in current commercial products and thus warrants further investigation. The objectives of this study are to (1) Identify the structure and composition of oligosaccharides in rice bran by chemical analysis and high-performance liquid chromatography, (2) Develop methods to increase, concentrate, and purify oligosaccharides from rice bran by chemical, physical and enzymatic means to produce readily consumable and palatable nutraceutical products, and (3) Characterize the physicochemical properties of rice bran oligosaccharides.

Project Methods
Objective 1: Identify the structure and composition of oligosaccharides in rice bran Defatted rice bran will be obtained from a commercial rice miller. Rice bran will be mixed with water and boiled for a certain period of time with constant stirring. The mixture will be centrifuged, the supernatant will be filtered and further purified by preparative gel filtration chromatography. The following information need to be determined in order to illustrate the complete structure of oligosaccharides: (a) the number of monosaccharide units present in the oligomer molecule; (b) the nature of the monosaccharide monomer(s); (c) in the case of hetero-oligosaccharides, the monosaccharide sequence; (d) the position of linkage of the different monosaccharides; and (e) the anomeric configuration and the conformation of the monosaccharide units. Objective 2: Develop methods to increase, concentrate and purify oligosaccharides in rice bran by chemical, physical and enzymatic methods. Chemical, physical and enzymatic methods will be applied to increase the quantity of rice bran oligosaccharides. The resultants will be then concentrated and purified. Ultrafiltration will be evaluated to concentration oligosaccharides. Spray drying or freeze-drying will be studied to obtain products with good properties and stability. (a) Chemical method: Dilute hydrochloric acid will be used to partially hydrolyze the polysaccharide portion in the rice bran, which will in turn increase the amount of oligosaccharides. The concentration of acid and conditions, e.g. temperature and duration, will be studied. (b) Enzymatic method: Carboxymethyl cellulase and xylanase from different commercial sources will be used in the enzymatic procedure to disintegrate fiber components in rice bran, resulting in an increase of oligosaccharides and the release of protein in the rice bran from the oligosaccharides. The enzymes will be screened and the conditions will be optimized to maximize the recovery of oligosaccharides. (c) Sonication/enzymatic method: High-intensity ultrasound will be used together with the selected enzyme(s) to further increase the recovery of oligosaccharides. Objective 3: Characteristics of rice bran oligosaccharides. (a) Dietary fiber content: An AOCS standard method will be used. (b) Freezing temperature and maximum freeze-concentrated glass transition temperature: A Perkin-Elmer differential scanning calorimetry (DSC) (Pyris 1) equipped with a cooling system will be used to determine the freezing temperatures of oligosaccharide solutions at different concentrations. Sample will be heated from -50C to 25C at 5C/min. The onset temperature of water melting peak will be recorded as the freezing temperature and the peak of the second derivative will be the glass transition temperature. (c) Sweetness: A 10% oligosaccharide solution will be compared against sucrose solution to determine the sweetness. (d) Viscosity: a viscometer will be used to measured the viscosity at different concentrations. (e) Chemical stability: stability against high temperature, low and high pHs, and salts will be measured with HPLC.

Progress 07/01/03 to 06/30/06

Outputs
The growing concern on healthy eating among consumers has increased the value of oligosaccharides as a food ingredient. Non-digestible oligosaccharides are generally categorized as prebiotics and benefit the colon by stimulating the growth of beneficial microflora and repressing the growth of pathogenic ones. Rice bran is an inexpensive, underutilized bio-resource. Its inherent oligosaccharide content is about 5-6% and presents a tremendous potential to offer unique health benefits should extraction yields can be further increased. This project aimed to increase the yield of oligosaccharides from rice bran by physical, chemical, and enzymatic means. The use of dilute hydrochloric acid, ethanol, boiling, homogenization, ultrasonication, xyllanase, cellulase, and/or combination was investigated. The optimized extraction procedure made use 1.0% cellulase (based on bran weight), carrying out the extraction at 50oC for 1 hour, arresting the enzymatic reaction by boiling for 15 min, precipitating out residual starch and protein by the addition of equal volume of ethanol, centrifugation, and then drying the supernatant at 40oC. Extraction yield ranged from 13.4 to 39.9%. Total carbohydrates, reducing sugars, and residual protein were 44.5-77.4%, 13.1-62.3%, and 7.1-11.6%, respectively. As determined by high-performance anion-exchange chromatography with pulsed amperometric detector, the oligosaccharide components in the extracts with prebiotic significance were triose (raffinose, cellotriose, and maltotriose), tetraose, and pentaose. Glucose was still the predominant sugar in the extracts and may consists of inherent bran glucose and glucose produced by the degradation of cellulose-rich bran matrix. In vitro digestibility with salivary and pancreatic α-amylase was very low compared with the original bran sample and potato dextrin.

Impacts
Rice bran is an inexpensive, underutilized bio-resource. The extraction of high-value, nutritionally important food ingredient like oligosaccharides will therefore enhance the economic value of rice bran and will increase the benefits that can be derived from rice farming, thus a potential impact to the rice industry and health of consumers.

Publications

Wang, L., Patindol, J., and Wang, Y.J. 2005. Increasing oligosaccharides content in rice bran by physical and enzymatic means. Book of Abstracts. American Association of Cereal Chemists Annual Meeting.
Patindol, J., Wang, L., and Wang, Y.J. 2006. Cellulase-catalyzed extraction of oligosaccharides from defatted rice bran. Book of Abstracts. Institute of Food Technologists Annual Meeting and Exposition.

Progress 01/01/05 to 12/31/05

Outputs
Non-digestible oligosaccharides have been classified as "prebiotics" because they benefit the colon by selectively stimulating the growth of bifidobacteria and decreasing the toxicity of the colon contents. The growing concern on "healthy-eating" among consumers has increased the value of oligosaccharides as a food ingredient. This research focuses on the extraction and identification of oligosaccharides present in rice bran. Rice bran is an inexpensive, underutilized bio-resource. Oligosaccharides from rice bran has been reported to be about 5-6% and present a tremendous potential to offer unique health benefits if the oligosaccharides content can be further increased. This study has demonstrated that the yield of oligosaccharides from rice bran can be increased by physical and enzymatic means. When rice bran is extracted with 40-80% ethanol, the yield of oligosaccharides ranges 10-12%. When ultrasonication is combined with ethanolic extraction, no further increase in oligosaccharide yield is observed. The main oligosaccharide in the extract is raffinose as determined by high-performance anion-exchange chromatography with pulsed amperometric detection. The yields of soluble sugars are 13 and 16% for 1 h, and 14 and 24% for 16 h of stirring with xylanase or cellulose, respectively, when rice bran (10% solid in water) is treated with xylanase or cellulase at room temperature. When the mixture is blended at a high speed for 2 min after the cellulase treatment, the extract yield is increased to 33%. When rice bran is first boiled in water for 5 min before the cellulase hydrolysis and high-speed blending, the extract yield is increased to 49%. However, no significant improvement in the extract yield is noted for those treated with xylanase. A combination of boiling, cellulase and blending treatments is an effective way to increase the amount of extractable oligosaccharides from rice bran.

Impacts
Rice bran is an inexpensive, underutilized bio-resource. The extraction of high-value, nutritionally important food ingredient like oligosaccharide will therefore enhance its economic value and will increase the benefits that can be derived from rice farming, thus, a significant impact to the rice industry and health-conscious consumers.

Publications

Wang, L., J. Patindol, and Y.-J. Wang. 2005. Increasing oligosaccharides content in rice bran by physical and enzymatic treatments. (Abstract) American Association of Cereal Chemists Annual Meeting.

Progress 01/01/04 to 12/30/04

Outputs
The growing concern on healthy-eating among consumers has increased the value of oligosaccharides as a food ingredient. Oligosaccharides (3 to 20 monomers) are generally categorized as prebiotics due to their potential to promote human intestine health by stimulating the growth of beneficial intestinal microflora and repressing the growth of pathogenic ones. This research is focused on the extraction and identification of oligosaccharides present in rice bran. Rice bran is an inexpensive, underutilized bio-resource. The extraction of high-value, nutritionally important food ingredient like oligosaccharide will therefore enhance its economic value and will increase the benefits that can be derived from rice farming, thus, a significant impact to the rice industry and health-conscious consumers. Extraction experiments were done using deionized water, 80% ethanol, and dilute HCl (0.1N). Different extraction time and temperature combinations were tested for each solvent (overnight shaking at room temperature, heating in a 40oC water bath/shaker for 2 hours, heating in a water bath/shaker at 80oC for 2 hours, and boiling by reflux for 1 or 2 hours). High-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) was used to separate the components of the oligosaccharide extracts according to degree of polymerization. With deionized water as extractant, crude oligosaccharide yield ranged from 11 to 16%. HPAEC-PAD analysis revealed that the water-extracted oligosaccharides consisted of 55.0% sucrose, 13.7% glucose, 12.0% maltose, 10.8% raffinose, 5.8% fructose, and 2.8% melibiose. With 80% ethanol as extractant, crude oligosaccharide yield ranged from 9.0 to 9.5%. HPAEC-PAD analysis showed that the ethanol-extracted oligosaccharides consisted of 64.9% sucrose, 18.1% glucose, 8.0% raffinose, 6.9% fructose, and 2.2% melibiose. With dilute HCl as extractant, extraction yields were higher, ranging from 14.0 to 50.0%. HPAEC-PAD analysis showed complicated saccharide profiles, and components with a degree of polymerization of up to 50 were even detected. This result implies that the oligosaccharides obtained with dilute HCl were not natural and could be acid-degradation products from starch and fiber. Work continues to use enzymatic and sonication methods to enhance extraction of the more nutraceutically-relevant oligosaccharide fractions (DP 3 to 20).

Impacts
Rice bran is an inexpensive, underutilized bio-resource. The extraction of high-value, nutritionally important food ingredient like oligosaccharides will therefore enhance the value of rice bran and will increase the benefits that can be derived from rice farming, thus, a significant impact to the rice industry.

Publications

No publications reported this period

Progress 01/01/03 to 12/31/03

Outputs
The growing concern on 'healthy-eating' among consumers has increased the value of oligosaccharides as a food ingredient. Oligosaccharides as prebiotics promote human intestinal health by stimulating the growth of beneficial intestinal microflora and repressing the growth of pathogenic ones. This research project is focusing on the extraction and characterization the composition and structure of oligosaccharides from rice bran. Dilute hydrochloric acid (0.1N HCl), deionized water, and ethanol (90%) were used for extraction. Extraction time and temperature were varied for the optimization of the extraction procedure. The crude extract yield using hydrochloric acid, water, and ethanol was 14.0-50.0%, 11.5-16.0%, and 9.0-9.5%, respectively. Refluxing rice bran in boiling dilute HCl for 2 h gave the highest yield (50.0%). Reducing the acid concentration from 0.1 to 0.05N for a one-hour reflux extraction reduced the yield from 45.5% to 20.0%. With water, extraction through reflux also improved the yield. Extraction at 80 and 40 degrees C with water for 2 h were comparable with overnight room temperature extraction in terms of yield. With ethanol, extraction yield was not affected by extraction temperature and time. Chemical analyses of the crude extracts are in progress.

Impacts
Rice bran is an inexpensive, underutilized bio-resource. The extraction of high-value, nutritionally important food ingredient like oligosaccharides will therefore enhance the value of rice bran and will increase the benefits that can be derived from rice farming, thus, a significant impact to the rice industry.

Publications

No publications reported this period