85 J. Appl. Glycosci., 53, 85 89 (2006) C 2006 The Japanese Society of Applied Glycoscience Regular Paper Dietary Fiber Quantity and Particle Morphology of An (Bean Paste) Prepared from Starchy Pulses (Received July 2, 2004; Accepted December 7, 2005) Michiyuki Kojima,, Hideki Shimizu 2 and Kiyoshi Ohba 2 Department of Animal Science, Animal Products and Food Science Unit, Obihiro University of Agriculture and Veterinary Medicine (, Nishi 2 sen, Inada cho, Obihiro 080 8555, Japan) 2 Hokkaido Tokachi Area Regional Food Processing Technology Center (23 0, Kita 2 choume, Nishi 22 jou, Obihiro 080 2462, Japan) Abstract: The relationship between the amount of dietary fiber and particle morphology of an (bean paste) prepared from starchy pulses (Adzuki beans, tebou variety of kidney beans and kintoki variety of kidney beans) was clarified. Dietary fiber content in the three beans decreased with prolonged heating during an preparation. Differences in heating time did not affect the proportions of the general components of the prepared ans and had no effect on the distribution of an particle size. However, it was shown by scanning electron microscopy that many lesions existed on the surfaces of an particles heated for long periods of time. In fact prolonged heating increased the proportion of damaged particles and decreased the proportion of intact particles. There was a direct relationship between the amount of dietary fiber and the proportion of intact an particles. These results indicate that the principal reason why the amount of dietary fiber increased when the starchy pulse was heated was the production of the intact an particles on which the digestive enzymes could not easily act. Key words: dietary fiber, Adzuki bean, kidney bean, an (bean paste), intact particles The quantity of dietary fiber in three kinds of starchy pulses (Adzuki beans and two varieties of kidney beans tebou and kintoki is greater in heat treated beans than in dry beans. 3) The increase in dietary fiber caused by cooking has been attributed to both a property change of the pulse starch (to resistant starch, RS) and a strengthening of the starch granule. ) RS resists the action of amylase 4 8) unless the RS is dissolved in an aqeous solution of 2 N potassium hydroxide. 9 2) RS has been classified into three types (RS RS3) based on its physicochemical properties. 4,3) Starches of RS are not dissolved by amylase because, in aqueous solution, the matrix structure wraps around the starch granule preventing amylase access to it. RS2 is the group in which resistance derives from the indigestibility of the starch itself. The group has been divided into types A through C based on differences in crystal structure, as determined by X ray analysis. 4,3) RS3 is the category of retrogradation starches produced after the gelatinization of starch. In pulses (e.g., Adzuki beans, kidney beans and peas), several starch granules are enveloped by a storage protein and cell wall. 4,5) By heating a starchy pulse, this protein and cell wall covering undergo thermal denaturation and suppress any swelling of the starch, thus forming an particles. 6,7) The increase in the quantity of dietary fiber caused by heating indicates that an particles have formed. ) Using an prepared from starchy pulses, the relationship between the amount of dietary fiber produced and an particle morphology was examined in order to clarify the form and features of the Corresponding author (Tel. +8 55 49 5547, Fax. +8 55 49 5547, E mail: kojima obihiro.ac.jp). an particle produced by heating. MATERIALS AND METHODS Preparation of an. Samples were prepared using Adzuki beans and kidney beans (tebou and kintoki varieties), which were harvested in 998 in the Tokachi district. Three volumes of water were added to 300 g of dried beans, the mixture was boiled for 5 min, the same quantity of water was then added again, and the resulting mixture was heated for 0 min. The supernatant liquid was discarded, and five volumes of water were added. The mixture was boiled, and samples were heated for 60, 90 or 20 min. After grinding, the mixture was separated into an and residue using sieves (40 and 65 mesh). The twice sieved an, the sample, was washed with water twice, dehydrated and then freeze dried. Analysis of an components. The general composition of an was analyzed by routine procedures. 6 9) The moisture content was determined by 35 C normal pressure drying, protein content by the micro Kjeldahl method, lipid content by acid digestion, and ash content by the 550 C direct ashing method. Carbohydrate was calculated as the difference between dry weight and the other components. Dietary fiber was assessed by the Prosky method. It was measured using a kit (Total dietary fiber assay kit; TDF 00), α amylase (heat stable, from Bacillus ilicheniformis), protease (from Bacillus ilicheniformis), amyloglucosidase (from Aspergillus niger, Sigma). The starch content was analyzed by the alkali gelatinization glucoamylase glucose oxidase method. 20) The glucoamylase (from Rhizopus) and glucose oxidase (from Aspergillus niger,
86 J. Appl. Glycosci., Vol. 53, No. 2 (2006) type II S) were purchased from Sigma corporation. Measurement of particle size distribution. The particle size distribution of an dispersed in water was measured using a SALD 2000A (Shimadzu). The surface of the an particles was observed with a JSM 630F scanning electron microscope (SEM, JEOL). Using the gold evaporation technique, the sample was observed under an acceleration voltage of 5 kv. Using the cryo SEM method, the cotyledon cell of dry beans and cross sections of the an particle were observed with an S 2400 SEM (Hitachi). Assay of intact particles, damaged particles, and ruptured particles of an. Using the method of Kugimiya, 2,22) the proportions of intact, damaged and ruptured an particles were determined. RESULTS AND DISCUSSION General composition and dietary fiber content of an. The effect of heating time on the dietary fiber content of an was examined. The total dietary fiber (TDF) values of an prepared from tebou kidney beans were the highest, being 53% (w w, 60 min) and 48% (w w, 20 min), whereas those from kintoki kidney beans were 5% (w w, 60 min) and 44% (w w, 20 min), and those from Adzuki beans were 43% (w w, 60 min) and 28% (w w, 20 min) (Fig. ). The TDF content of all ans was higher than that reported for dry beans. ) However, the dietary fiber quantity in the an decreased when heating time was lengthened. A clear inverse relationship between TDF and heating time was observed. There was also a clear negative relationship between heating time and insoluble dietary fiber (IDF). In an prepared from kintoki kidney beans, a positive relationship between water soluble dietary fiber (SDF) and heating time was suggested. However, because the SDF content of this type of an was only 5% (w w) or less, the apparent relationship between SDF content and heating time may not be particularly meaningful. No remarkable differences in the general components (protein, ash content, lipid and carbohydrate) of an were observed with different heating times (Table ). The quantities of starch in an prepared from Adzuki beans, tebou kidney beans and kintoki kidney beans were about Table. Composition General composition of an. Adzuki bean Kidney bean (tebou) (g 00 g sample) Kidney bean (kintoki) 60 90 20 60 90 20 60 90 20 Protein 29.9 29.6 29.2 26.0 25.9 26.0 25.4 24.7 25.2 Ash 0.8 0.8 0.8 0.8 0.8 0.8 0.9 0.9 0.9 Lipid.0.0.0 2.0.9.9 2.0.9.9 Carbohydrate 68.3 68.6 69.0 7.2 7.4 7.3 7.7 72.5 72.0 Starch 6. 6.3 6.8 55.8 56.0 55.9 57.8 58.7 57.3 Dietary fiber (estimated) 7.2 7.3 7.2 5.4 5.4 5.4 3.9 3.8 4.7 The dietary fiber content was calculated by deducing the quantity of starch from the amount of carbohydrate. 6% (w w), 56% (w w) and 58% (w w), respectively. TDF can also be calculated by subtracting the amount of starch from the amount of carbohydrate. The calculated amounts of dietary fiber were about 7% (w w), 5% (w w) and 5% (w w) in an made from Adzuki beans, tebou kidney beans and kintoki kidney beans, respectively. However, actual TDF was considerably higher than the calculated value (Fig. ), with the differences between them being 20 36% (w w) for Adzuki beans, 28 38% (w w) for tebou kidney beans and 29 37% (w w) for kintoki kidney beans. The an particle formed by heating made it difficult for the digestives enzyme to access the starch, which is consistent with the increased TDF content found by the Prosky method. Indigestible protein in an. It was shown that the main components of an were starch and the protein that enfolds it and, that the proportion of indigestible protein is high. According to the Kjeldahl results, the total amount of indigestible protein was 60 70% (w w) of all the proteins of the an, afterit was digested in α amylase, glucoamylase and protease (Fig. 2). The amount of protein indigestible by protease in the insoluble fraction was higher than that of water soluble fraction. When the heating time was lengthened, Fig.. Dietary fiber content of an prepared from three kinds of heated bean. A, Total dietary fiber (TDF); B, insoluble dietary fiber (IDF); C, water soluble dietary fiber (SDF).
Dietary Fiber Quantity and Particle Morphology of An (Bean Paste) 87 the quantity of the indigestible protein in IDF and SDF decreased (Fig. 2). Particle size distribution and microscopy of an. Measurements of particle size distribution and SEM observations were carried out to compare the sizes and shapes of an particles produced by heating the bean paste for different lengths of time. No differences in particle size were detected by either method. The average particle sizes in an made from Adzuki beans, tebou kidney beans, and kintoki kidney beans were about 20, 23 and 27 µm, respectively. The surface of particles in Adzuki bean an (heated 20 min) had lifted off in some places, exposing the particle interior (Fig. 3). However, in undamaged areas, a wrinkled cell wall covered the particle surface (Fig. 3b, d). This was not observed in Adzuki bean an heated for 60 min. Similar heating time based differences in the an particle surfaces were observed in an prepared from tebou and kintoki kidney beans. Cryo SEM observation of raw and heated (60 and 20 min) starch granule cross sections of Adzuki bean seeds revealed several starch granules and storage proteins, covered by the cell wall (Fig. 4). When the an particle was prepared and heated, it swelled, and the starch granule Fig. 2. The indigestible protein after enzyme digestion of an prepared from three kinds of heated bean. A, Indigestible protein in IDF; B, indigestible protein in SDF. Fig. 3. Particle of an prepared from Adzuki beans in which the heating time differs. (a) 60 min ( 500), (b) 60 min ( 3300), (c) 20 min ( 500), (d) 20 min ( 3300).
88 J. Appl. Glycosci., Vol. 53, No. 2 (2006) Fig. 4. Cross sections of raw and heated cells of Adzuki beans. (a) Raw cells, (b) 60 min heated cells, (c) 20 min heated cells. Fig. 5. Percentage of intact particles, damaged particles and ruptured particles in an prepared from three kinds of heated beans. A, Ruptured particles; B, damaged particles; C, intact particles. Fig. 6. The correlation between dietary fiber content and intact particle., Adzuki bean paste;, Adzuki bean paste (grinding treatment);, kidney bean paste (tebou);, kidney bean paste (tebou, grinding treatment);, kidney bean paste (kintoki);, kidney bean paste (kintoki, grinding treatment). spread into the cell interior, to cover the thermally denatured protein. The extent of swelling was greater at longer heating times, the latter further increasing the swelling pressure. This swelling pressure may account for cell wall damage observed in some of the an particles. The composition of intact particles, damaged particles and ruptured particles of an. The an particle structure multiple starch granules and protein wrapped in a cell wall was examined. Kugimiya reported that an particles could be classified into three types: intact, damaged and ruptured particles. 2,22) These types can be determined quantitatively by an enzymatic method using α amylase. Structural features of the three types of particles are as follows. (A) The starch is completely contained within the intact particle by protein and the cell wall. The enzyme has little effect on the starch, which does not leach from the an particle during washing in water. (B) A part of the cell wall of the an particle is damaged. For this reason, part of the starch comes into contact with the enzyme and is dissolved. Repeated exposure to the amylase dissolves more and more of the remaining starch. As is true for intact particles, the starch from damaged particles does not leach out during washing in water. (C) The ruptured particle has almost completely lost the form typical of an an particle. Because the ruptured cell wall exposes all of the starch, most starch is dissolved by the enzyme. Starch of a ruptured particle easily leaches out when the particle is washed in water. In the Prosky method, substances that are not decomposed by α amylase, protease or glucoamylase are defined as dietary fiber. The quantity of dietary fiber may be different among an particles in different states. The proportions of intact, damaged and ruptured an particles prepared using different heating times were determined
Dietary Fiber Quantity and Particle Morphology of An (Bean Paste) 89 (Fig. 5). The proportion of ruptured particles was about % for all beans heated for any length of time. This low proportion suggests that the process of washing in water removes the ruptured particles. The proportion of damaged particles increased as the heating time was lengthened, and the proportion of intact particles decreased. These results confirm the SEM observations of the particle surfaces. When the proportion of intact particles was low, the dietary fiber value tended to be low. Samples of an in which the proportion of intact particles was intentionally decreased were prepared in order to examine this relationship. There was a clear positive correlation (correlation coefficient; r= 0.83) between the dietary fiber quantitative value and intact particle mass (Fig. 6). The increase in dietary fiber was related to the state of the an particles when the starchy pulse was heated. 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