Starch Starch is one of the main forms of carbohydrate in the diet. Starches are polysaccharides, i.e.

they are made up of a number of glucose molecules which are linked together. They can therefore be described as complex carbohydrates. Starches are found in plant sources including potatoes and cereal products (e.g. bread, pasta). In general, digestible starches are broken down by digestive enzymes in the small intestine into glucose molecules. The glucose is then absorbed into the blood and used to provide energy for the body. Resistant starch Resistant starch is generally considered to be represented by the total amount of starch and the products of starch degradation that are not digested in the small intestine, and pass into the large intestine (or the colon). Therefore, resistant starch can be regarded as a component of dietary fibre. Types of Resistant starch Class RS1 RS2 RS3 RS4 Types physically inaccessible granular, ungelatinised nongranular, indigestible chemically changed starch

Resistant starch not digested There are several reasons why resistant starch is not digested: * The starch may be physically inaccessible to the digestive enzymes such as in grains, seeds or tubers. * The starch granules themselves are structured in a way which prevents the digestive enzymes from breaking them down, e.g. raw potatoes and unripe bananas. * When starches are heated they gelatinise and become more easily digested. However, if these starch gels are cooled, starch crystals form in the food that are resistant to enzyme digestion. This form of retrograded starch is found in foods such as cornflakes or cooked and cooled potatoes. * Starches that have been chemically treated (etherisation, esterisation, cross-bonding) cannot be broken down by digestive enzymes. Sources of resistant starch in the diet Resistant starch is found in a wide range of foods including intact wholegrains, legumes, pasta, unripe bananas, raw potatoes, cooked and cooled potatoes, bread, cereals, some high fibre drinks and foods containing modified starches (some breads and cakes) . Properties of RS Low water holding capacity. Small particle size. Bland flavour with no energy. Stable suspension viscosity. White colour. High gelatinized temperature

Functional Properties of RS in food Functional fibre Texture modifier Crisping agent Colour modifier Reduction in hardness Improved expansion Reduction in dough rheology Bulking agent Thickner for health

Resistant starch in the gut Resistant starch reaches the large intestine (or the colon) virtually unchanged. However, it is then fermented by the microbial flora naturally present in the colon, to produce low levels of the gases carbon dioxide, methane and hydrogen. Additional fermentation products include organic acids and short chain fatty acids. Short chain fatty acids Short chain fatty acids (SCFA) are formed when polysaccharides are fermented by the anaerobic bacteria present in the large intestine. Many different forms of polysaccharides are present in the large intestine, including resistant starch. The main SCFAs produced in the human gut are butyrate, propionate and acetate. The concentrations of SCFAs in the large intestine vary depending on the types of polysaccharides although generally acetate is the most abundant and butyrate is the least abundant. Concentrations also vary in the different regions of the large intestine with higher concentrations detected in the area nearest the junction with the small intestine (70 140mM). Effect of short chain fatty acids in the large intestine SCFAs lower the pH of the contents of the large intestine. This is of benefit to health because the reduced pH creates an environment that prevents the growth of harmful bacteria. A lower pH also aids in the absorption of minerals such as calcium and magnesium. SCFAs increase the blood flow to the colon and provide the cells in the wall of the intestine with a metabolic fuel (mainly in the form of butyrate). Additionally, butyrate has been shown to induce programmed cell death (apoptosis) and exert a level of control over the cell cycle. This suggests that butyrate might play an important role in maintaining the integrity of the gut wall by preventing the uncontrolled proliferation of abnormal cells that occurs in the early stages of colorectal cancer. Health benefits of resistant starch Resistant starch contributes to the amount of fibre in the large bowel. Many health benefits have been attributed to foods providing resistant starch in the context of a high fibre diet. These include: a slower, more controlled release of glucose from the food into the blood stream (the glycaemic response see below); improved bowel health; improved blood lipid profile; an increased feeling of satiety and increased micronutrient absorption (magnesium, calcium) in the colon. These factors may affect the risk of developing diseases such as colorectal cancer, cardiovascular disease, osteoporosis and obesity and assist in the management of diabetes, impaired glucose tolerance, inflammatory bowel diseases, diverticulosis and constipation . Resistant starch improve the glycaemic response after a

meal? When most carbohydrates are consumed, the levels of glucose in the blood are raised, peaking 15 45 minutes after finishing your meal. Levels return to normal within two to three hours. The concentration of the hormone insulin in the blood also increases in response to the elevated glucose concentration. High levels of insulin in the blood inhibit the use of stored body fat, as well as modulating appetite and satiety signals. Rapidly digestible carbohydrates, such as those in white bread and confectionery, bring about a high glycaemic and insulinaemic response. In some instances, for example when you are being very active, this is of benefit as the glucose is rapidly absorbed into your blood stream to give you a fast sugar boost. However, carbohydrates that are broken down slowly, for example those from wholegrain foods or legumes, are generally better for keeping energy levels up throughout the day. This slower, more controlled glycaemic response is could be beneficial to all individuals as there is greater access to and use of stored fat, and hunger signals are suppressed. For people with diabetes or impaired glucose tolerance, increasing the amounts of resistant starch in the diet can help with the day-to-day management of blood glucose levels, as well as contributing to the bodys use of fat. Resistant starch has been shown to improve bowel health and function. How can this be assessed? There are several markers that can be used to assess the health and function of the bowel. The time it takes for food to pass through the entire gut is referred to as the transit time. Healthy bowels process food quickly and gut cells absorb just enough water to produce firm stools. This, along with other factors, including the amount of fibre in the diet and the numbers of bacterial cells in the gut, affects the faecal weight. Some compounds, such as ammonia, phenols or bile acids are produced in the gut which can be damaging to the cells lining the large intestine. Low levels of these compounds in faeces suggest that the bowel is healthy and that their effects in the gut are lessened . Improvements in bowel health Resistant starch, like other forms of dietary fibre, helps to prevents constipation by providing bulk to the faeces. Bulky faeces move through the gut faster and result in an increased stool weight. Diets high in resistant starch have been shown to reduce the luminal pH, limiting the growth of harmful bacteria in the gut. They also decrease the amounts of secondary bile acids and ammonia present in the large bowel. Secondary bile acids can be converted into active compounds by the microbial flora and these can damage DNA. Ammonia has been shown to increase the proliferation rate of the cells in the gut wall. It is therefore advantageous to limit the concentrations of these compounds in the colon to reduce the risk of developing colorectal cancer. Additionally, the increased faecal bulk dilutes the effect of any genotoxic agents in the large intestine, thereby reducing the extent of DNA damage to the cells lining the colon. Resistant starch can function as a prebiotic. Prebiotics are non-digestible food ingredients that stimulate the growth and activity of bacteria in the colon. Approximately 100 trillion bacteria live in our large intestines and they are essential for keeping our digestive system healthy. As resistant starch, along with other forms of dietary fibre, arrives in the large intestine unchanged, it provides metabolic substrates (fuel) for these colonic bacteria and promotes the growth of the beneficial strains of bacteria (e.g.

Bifidobacteria). A healthy bacterial flora further improves bowel health and function. Resistant starch used to prevent weight gain Due to its indigestible nature, resistant starch has a lower calorific value than that of digestible starch (16kJ/g or 4kcal/g). Therefore, foods high in resistant starch may provide less energy weight for weight, although this will depend on the amounts of other nutrients in the food. How much of a contribution should resistant starch make to the diet? In the UK, resistant starch makes up a very small proportion of total daily starch intake. On average, the intake of all starches is 130g per day whereas the average intake of resistant starch is only around 4g per day. This is very low compared to countries in the developing world where intakes of resistant starch are much higher (for example 10g per day in India). Intake of dietary fibre in the UK should increase in order to improve bowel health and increasing consumption of foods rich in resistant starch is one way of achieving this. However, current intakes of dietary fibre in the UK are already well below recommendations and it has been recognised that substantial dietary changes would need to be made to reach these targets . Is resistant starch safe? Resistant starch is well tolerated in the diet. Minor effects of very high levels of resistant starch consumption are consistent with those of high fibre intakes, such as flatulence, belching, bloating and stomach aches which can occur when large amounts of resistant starch are fermented in the large intestine. How can resistant starch be incorporated into meals? You can up your intake of resistant starch, along with other forms of dietary fibre, by increasing the amount of wholegrain foods in your diet, for example seeded or granary breads, wholemeal pasta or high fibre cereals. There are also commercially produced resistant starches, for example Hi-maizeTM or NOVELOSE 330TM which are sometimes used by food manufacturers to produce a food product that is high in fibre and which can also be lower in calories. Factors influencing the formation of RS. Inherent properties of starch Crystallinity of starch. One of the causes of resistance to enzymes is the crystallinity of native type B starch granules as observed in the case of amylomaize starch and also the encapsulation of starch within plant cell or tissue structures. X-ray diffraction and differential scanning calorimetry studies on crystalline residues from amylomaize starch samples have suggested that chain fragments packed in a type B crystalline structure with a slightly enlarged crystal lattice contribute to formation of RS from amylomaize starch. Any treatment that eliminates starch crystallinity (that is, gelatinization) or the integrity of the plant cell or tissue structure (that is, milling) increases enzyme availability and reduces the content of RS, whereas recrystallization and chemical modifications tend to increase the RS. The modified food starches are partially resistant to enzymes as a result of chemical modifications induced intentionally . Besides these, the cellular structure of plant foods influences the digestibility of starch in the small intestine as well as the intrinsic digestibility of a particular physical form of starch.

Granular structure. A large variability in susceptibility to amylases shown by raw starch granules also influences RS formation. Potato starch and high amylose maize starch are known to be very resistant in vitro and incompletely absorbed in vivo, whereas most cereal starches are slowly but virtually completely digested and absorbed in vivo . The smaller surface-to-volume ratio of the large potato granules is probably important. The nature of the granule surface also needs to be considered; an adsorbed layer of non-starch material would effectively impede the action of the enzyme . Raw tepary starch is found to be more resistant to hydrolysis than maize starch, perhaps due to differences in granule structure and amylose content . Amylose:amylopectin ratio. A higher content of amylose lowers the digestibility of starch due to positive correlation between amylose content and formation of RS . The importance of the amylose:amylopectin ratio in the postprandial glycaemic and insulinaemic responses to corn was studied in commonly consumed corn products . The meals containing high amylose (70%) corn flour had an RS of 20 g/100 g DM than that containing ordinary corn flour (25% amylose) that had RS of 3 g /100 g DM. Retrogradation of amylose. When heated to about 50 C, in the presence of water, the amylose in the granule swells; the crystalline structure of the amylopectin disintegrates and the granule ruptures. The polysaccharide chains take up a random configuration, causing swelling of the starch and thickening of the surrounding matrix such as, gelatinizationa process that renders the starch easily digestible. On cooling/drying, recrystallization (retrogradation) occurs. This takes place very fast for the amylose moiety as the linear structure facilitates cross linkages by means of hydrogen bonds. Figure 10 shows the formation of gel and micelle on cooling of a concentrated solution of amylose . The branched nature of amylopectin inhibits its recrystallization to some extent and it takes place over several days. Retrograded amylose in peas, maize, wheat, and potatoes was found to be highly resistant to amylolysis . The rate and extent to which a starch may retrograde after gelatinization essentially depends on the amount of amylose present. Repeated autoclaving of wheat starch may generate up to 10% RS. The level obtained appeared to be strongly related to the amylose content, and the retrogradation of amylose was identified as the main mechanism for the formation of RS that can be generated in larger amounts by repeated autoclaving . During storage, the dispersed polymers of gelatinized starch are said to undergo retrogradation to semicrystalline forms that resist digestion by pancreatic _-amylase. It forms a major portion of RS in wheat bread and corn flakes , whereas only 25% of the RS in cooked, cooled potatoes can be accounted for as retrograded amylose. The digestibility of legume starch is much lower than that of cereal starch, which is attributable to higher content of amylose in the former. The digestibility of high amylose cereal starch is reported to be significantly lower. Native high-amylose starch is known to be high in type II RS (RS 2) , which is defined as starch in its native granular state that is resistant to digestion in the small intestine. This after cooking and cooling gives high yields of type III RSor retrograded starch . Heating of RS preparation from amylomaize VII resulted in broad endothermic transition, which is ascribed to melting of amylose crystallites . Exothermic transitions during controlled cooling of isolated potato amylose fractions have been attributed to amylose chain association. The formation of RS likewise has been attributed to the ordering of amylose chains . Based on previous studies of amylose behavior, it has been suggested that the exotherms observed during the cooling of either amylose or a thermally treated RS preparation reflect chain association, which may involve amylose aggregation and gelation dominated by formation and subsequent lateral aggregation

of type B double helices in crystalline array . Gelatinized waxy corn starch stored at varying temperatures from 6 C to 60 C for 1 to 29 d also showed reduced enzyme susceptibility to pancreatic _-amylase and amyloglucosidase . Influence of amylose chain length. Influence of amylose chain length on enzyme RS formation was studied by hydrolyzing potato starch amylose to varying degrees by incubation with barley _-amylase for different periods, and monitored by measuring the number of average chain lengths or degree of polymerization (DPn). The DPn of RS varied between 19 and 26 and was independent of the chain length of the amylose (DPn 40 to 610) from which it was formed. Results suggested that RS might be formed by aggregation of amylose helices in a crystalline _-type structure over a particular region of the chain (about 24 glucose units). Linearization of amylopectin. Linearization of amylopectin occurs during the long lowtemperature baking process due to the prolonged activity of intrinsic amylases in the dough, and is prominent in the presence of certain organic acids that is, in bread products baked with added lactic acid . It has been reported to significantly increase RS formation during wetautoclaving . Heat and moisture Water content is an important factor that affects formation of RS. Repeated heat/moisture treatment is associated with a decrease in the hydrolysis limit of pancreatic _-amylase and increased formation of RS. Maximum RS yield was obtained at a starch:water ratio of 1: 3.5 (w/w) and a heat treatment at 18% moisture gave increased levels of the degree of crystallinity of normal and waxy starches and thus reduced enzyme susceptibility. However, at 27% moisture, starch degradation to some extent made areas of starch more accessible to enzyme attack. Thus, proper heat treatment could be used as a method of preparation of RS . In addition, higher temperature and less water results in type A configuration, whereas lower temperature and high water content results in type B configuration .Some scientist determined solubility, water vapor sorption, and swelling characteristics for RS prepared fromwheat starch and linterized wheat starch by autoclaving and cooling and by cross-linking The experimental RS made from wheat starch contained 10% to 73% RS versus 58% and 40% in commercial sources, Novelose 240 and 330 respectively, produced from high-amylose maize (corn) starch. In excess water, the experimental RS starches (except for the cross-linked wheat starch) gained 3 to 6 times more water than the commercial RS starches at 25 C, and 2 to 4 times more at 95 C. All starches showed similar water vapor sorption and desorption isotherms at 25 C and aw < 0.8. At aw 0.84 to 0.97, the RS made from wheat starch (except cross-linked wheat starch) showed approximately 10% higher water sorption than the commercial RS.

RS determined in several selected cereals, legumes, and tubers subjected to dry and wet heat treatment brought out higher RS contents in foods subjected to dry heat treatment compared with wet processed ones. Sorghum, green gram dhal, and green plantain showed highest RS content (5.51%, 5.81%, and 10.7%, respectively) . Interaction of starch with other components Interactions of starch with different components present in the food system are known to influence the formation of RS as follows. Protein. Starch-protein interaction has been believed to reduce RS contents as observed in case of potato starch and added albumin when autoclaved and subsequently cooled at 20 C . Dietary fiber. Insoluble dietary fiber constituents such as cellulose and lignin have been shown to have minimal effects on RS yields compared with other constituents such as potassium and calcium ions and catechin. Enzyme inhibitors. Polyphenols, phytic acid, and lectins present mainly in leguminous seeds, have been reported to inhibit in vitro starch hydrolysis and to lower the glycemic index. Tannic acid significantly inhibits both amylases and intestinal maltase activity . Indigestible residues from black beans (Phaseolus vulgaris cv. Tacari gua), green beans (P. vulgaris), carrots (Daucus carota), and rice bran (Oryza sativa) are all reported to inhibit pancreatic _-amylase in vitro . Since amylolysis is inhibited by phytic acid, a decrease in phytate content increases starch digestibility . Contradictory information exists in the literature on this aspect. The autoclaving and subsequent cooling of potato starch and catechin was found to significantly reduce the yields of RS, whereas the addition of phytic acid to potato starch reduced the RS contents to a minor extent compared with the RS formed from potato starch with no added constituent. The reasons for the same are still not clear. Ions. The yields of RS in potato starch gels decrease in the presence of calcium and potassium ions compared with those with no added constituent , presumably due to the prevention of formation of hydrogen bonds between amylose and amylopectin chains caused by adsorption of these ions. Sugars. The addition of soluble sugars such as glucose, maltose, sucrose, and ribose has been found to reduce the level of crystallization and subsequently reduce the yields of RS . The mechanism of retrogradation inhibition was considered as the interaction between sugar molecules and the starch molecular chains, which change the matrix of gelatinized starch (the sugars act as anti-plasticizers and increase the glass transition temperature). The role of sugars on the formation of RS in starch gels (RS type III) . Sugars influenced the RS levels in starch gels only when added in high concentration (final starch-water-sugar ratio of 1:10:5 w/w). In wheat starch gels, the RS yields decreased from approximately 3.4% to 2.8% in the presence of sucrose or glucose, and to 2.5% in the presence of ribose or maltose. An increase in RS yield was observed with high-amylose corn starch. The experiments showed that the differences in gelatinization temperature, lipid content, and apparent amylose content of the 2 starches were not the main causes of the different impact of sugars on RS yields.

Lipids, emulsifiers. In a study, amylomaize VII starch, autoclaved at 125 C, was reacted during cooling below 100 C with lysophosphatidyl choline (LPC), sodium stearoyl lactylate (SSL), and hydroxylated lecithin (OHL) . Differential scanning calorimetry95 C to 110 C indicated formation of amylose-lipid complexes, and at about 155 C indicated the presence of enzyme-resistant starch (RS). Yields of RS from complexed samples isolated by thermostable bacterial _-amylase or amyloglucosidase were lower than yields of RS from the autoclaved and cooled control. Formation of complexes competes with amylose chains involved in generation of RS. Amylose-lipid complexes are enzyme-degradable, and an increase in complexed amylose reduced yields of RS. Amylose recrystallization in RS formation is competitively affected by complexation of amylose with LPC and SSL. Results of X-ray diffraction powder crystallography were in agreement with DSC measurements. Complexes of amylose with LPC, SSL, and OHL gave type V patterns; enzymic hydrolysis of the complexes yielded type B RS structures. However, the viewpoint differs among scientists working in this area. While some workers believe amyloselipid complex to reduce the formation of RS, others believe the amylose-lipid complex itself to be a form of RS. Processing conditions Processing techniques may affect both the gelatinization and retrogradation processes, influencing RS formation. This fact is of great importance for the food industry since it offers the possibility of increasing the RS content of processed foods and foodstuffs. Baking, pasta production, extrusion cooking, autoclaving, and so forth are known to influence the yield of RS in foods. Highly processed cereal flours and foods made from the flours, such as pasta, contain much lower levels of RS, averaging only about 1.5% to 8% RS on a dry basis. Since the crystalline structure of starch in legumes (type C) is more stable compared with the crystal structure in cereal grains (type A), processing cereal grains results in a large decrease in RS content, while legumes are excellent sources of RS. Cooking under conditions of high moisture and temperature can significantly lower the RS content by disrupting crystalline

structure. Increasing the levels of RS can be done in other conditions, such as extrusion followed by cooling to induce crystallization The RS contents in various processed food samples have been reported . Thermal processing Steam cooking. Steam cooking helps in production of RS. Starches isolated from several steam-heated legumes were rich in indigestible RS (19% to 31%, DM basis), which was not observed in raw beans .Similarly, RS measured directly in conventionally and high-pressure steamed beans were 3 to 5 times higher than in the raw pulses, suggesting retrogradation to be mainly responsible for the reduction in digestibility. Prolonged steaming as well as short dry pressure heating decreased the enzymically assessed total starch content of whole beans by 2% to 3% (DM basis), indicating that these treatments may induce formation of other types of indigestible starch. Autoclaving. Autoclaving results in increase in RS. Autoclaved wheat starch has 9% RS compared with less than 1% in uncooked wheat starch. Autoclaved wheat starch contained 6.2% RS (of dm); this increased to 7.8% after 3 further reboiling/cooling cycles . In another study, white flour subjected to repeated autoclaving and cooling cycles showed an increase in total dietary fiber >3 times that of bread flours and 4 times that of pastry flours . The increase was primarily due to the formation of RS. Investigations on the formation of enzyme-resistant starch (RS) during autoclaving and cooling showed highest yield (21.3%) to be obtained from amylomaize VII starch (70% amylose). Formation of RS in amylomaize VII starch was affected by the starch/water ratio, autoclaving temperature and number of autoclaving-cooling cycles. The number of cycles exerted the most pronounced effect on RS; increasing the number of cycles to 20 raised RS level to >40%. Furthermore, the thermoanalytical data suggested that amylose-lipid complexes were not involved in the formation of

RS. Yields in excess of 20% RS can be obtained from autoclaved amylomaize starch containing 70% amylose. They can be raised to levels of 40% by increasing the number of autoclaving-cooling cycles up to 20. The extent of RS formation in commercially available autoclaved corn, potato, and leguminous products and in autoclaved purees intended for consumption by infants aged 3 to 8 Parboiling. Parboiling increases RS production. In studies on 5 rice varieties, differing in amylose content, the in vitro and in vivo RS levels were low and positively correlated with amylose content Higher RS starch levels were found incooked and parboiled-cooked rice than in raw rice; waxy rice had very low values. Higher contents of RS have been reported in parboiled rice than raw white rice, which also increased by cooling or freezing . Baking. Baking increases RS content. In a study to evaluate the effect of baking on RS formation, white bread was baked and divided into 3 fractions (crumb, inner crust, and outer crust) .Starch levels were found to be highest in dough and lowest in outer crust after baking for 35 min. RS levels were lowest in dough and highest in crumb after baking for 35 min. A low-temperature, long-time baked product contained significantly higher amounts of RS than bread baked under ordinary conditions . Addition of lactic acid increased RS recovery further whereas malt had no impact on RS yield. The highest level of RS was noted in long-time baked bread based on highamylose barley flour. RS isolated from wheat-based foods such as chapatti and phulka was structurally characterized as a linear 1, 4linked _-D-glucan essentially derived from retrograded amylose fraction, which was dependent on the severity of the processing treatments as well as the levels of gluten and damaged starch in the wheat flour . Extrusion cooking. Effect of extrusion cooking, at different temperatures (90, 100, 120, 140, or 160 C), moisture contents (20%, 25%, 30%, 35%, or 40%) and screw speeds (60, 80, or 100 rpm), was investigated on the formation of RS of type 3 (RS3) in hull-less barley flours from CDC-Candle (waxy) and Phoenix (regular). The RS3 content of the native flours, in general, decreased

by extrusion cooking, but not significantly. Storage of extruded flour samples at 4 C for 24 h before oven drying slightly increased RS3 content . With pearl barley used as the primary material in tests designed to optimize the production of RS by extrusion an extrusion temperature of 150 C and a barley moisture content from 17.5% and 22.5% moisture, followed by cold storage at 18 C gave the best results . Corn starches with and without guar gum [10% (w/w)] and 2% (w/w) of diacetyl tartaric acid ester of monoglyceride, sodium stearoyl-2-lactylate or citric acid, respectively, were extrusioncooked in a twin-screw extruder at 18% moisture, 150 C, and 180 rpm screw speed. The formation of RS in extruded corn starch was found to be strongly affected by the addition of gum and the different food additives. X-ray diffraction of the extruded starches gave a V diffraction pattern indicating the effect of extrusion cooking and amylose-lipid complexes. Enzymatic digestion did not affect the V structure, which could apparently be attributed to extrusion cooking. Purification of the isolated RS by size exclusionHPLC showed a dependence of molecular weight on the added additives. Results of differential scanning calorimetry and X-ray diffraction suggest that amylose-lipid complexes could also be involved in the formation of RS in extruded cornstarch. Pyroconversion. Pyroconversion of starch increases RS content. Lima bean (Phaseolus lunatus) starch was modified using pyroconversion, the optimum product being recovered from native starch treated with a 160:1 starch/HCl ratio, at 90 C for 1 h, resulting in starches containing 49.5% indigestible starch . Starch pyrodextrinization decreased the amount of enzymically available starch through formation of atypical glycosidic bonds that are not digested by the amylases and maltooligosaccharidases in the small intestine of humans. Microwave irradiation. Microwave irradiation improves the digestibility of tuber starches, which could be accompanied by physicochemical and structure changes. Microwave cooking of legumes such as chickpeas and common beans produced a redistribution of the insoluble nonstarch

polysaccharides to soluble fraction, although the total nonstarch polysaccharides were not affected. This was evaluated by assessing the physicochemical, nutritional, and microstructural modifications in starch and nonstarch polysaccharides . The RS level decreased from 32.5% of total starch in raw chickpeas and beans, respectively, to about 10% in cooked samples with a concomitant increase in the level of rapidly digestible starch from 35.6% and 27.5% to about 80%. Studies on effects of different heat treatments (cooking, microwave cooking, pressure cooking) on the rate of hydrolysis, hydrolysis index, and glycaemic index values of kudzu starch and cornstarch showed increase in digestible starch and decrease in RS following heat treatment. The rate of hydrolysis of kudzu starch and cornstarch increased following heat treatment, especially after microwaving . Miscellaneous treatments. Milling. Leguminous seeds, in which cell structures are preserved after cooking (that is, bread with whole seeds); bean flour with intact cells ,foods containing large particles such as bread with whole seeds have lower physical accessibility of starch to amylase action, and thereby contribute to higher RS contents. In some foods, physically inaccessible starch is likely to be an important fraction of the total starch that is resistant to digestion in vivo. found evidence of 20% starch malabsorption from a diet containing bean flour with intact cells. About half of the malabsorbed starch was retrograded amylose. The precooked flours (PCF) prepared from dried lentils and beans, rich in intact cells filled with starch granules, indicated that they contained important quantities of RS, such as retrograded amylose (3% to 9%, DM). Germination. Germination is shown to decrease the RS content in bengal gram, field beans, cow pea, and green gram . Fermentation. Fermentation reduces RS content. Flour from sorghum cv. Tabat was mixed with water and previously fermented dough starter, and fermented at 37 C for a maximum of 36 h

showed an increase in the in vitro starch digestibility and a decrease in the content of RS and total starch .RS formation has also been shown to decrease in the fermented products, idlis and dhoklas . Storage conditions Generally, RS increases on storage, especially low-temperature storage. Cold storage seems to support an increase in RS content. Whole corn bread and corn bread crumb, when stored at different temperatures (20 C, 4 C, or 20 C) for 7 d showed RS contents to reach a maximum between 2 and 4 d at all storage temperatures, after which they decreased . Lowest RS levels in whole corn bread were found after storage at 20 C (2.18 g/100 g) for 7 d. Preparation of RS RS can be prepared by using heat treatment, enzyme treatment, combined heat treatment and enzyme treatment, and chemical treatment. Heat treatment Heat treatment of starch to various extents leads to formation of RS. RS can be obtained by cooking the starch above the gelatinization temperature and simultaneously drying on heated rolls like drum driers or even extruders. The gelatinization of starch granules by heat processing strongly influences their susceptibility to enzymatic hydrolysis. In a high-moisture environment, amylose leaches from the granules, increasing the solubility of starch and thereby its susceptibility . Good yields of RS can be obtained by gelatinizing starch at 120 C for 20 min, followed by cooling to room temperature . The starch gels are then frozen overnight at 20 C and dried at 60 C before milling. Many combinations of time and temperature treatments have been used to make type III RS from various sources of native starch. Even for starches with normal amylose levels, it is recognized that cooking at >100 C can increase the yield of type III RS. The temperature treatments have included autoclaving the starch

at 110 C , at 121 C for periods ranging from 30min. to 1 hr. An enzyme-RS type III, which has a melting point or endothermic peak of at least about 140 C, as determined by differential scanning calorimetry (DSC) can be produced in yields of at least 25% by weight, based on the weight of the original starch ingredient . A gelatinization stage, nucleation/propagation stage, and preferably a heat-treatment stage are required to produce reduced calorie starch-based compositions that contain the enzyme-resistant starch. It is produced using crystal nucleation and propagation temperatures, which avoid substantial production of lower melting amylopectin crystals, lower melting amylose crystals, and lower melting amylose-lipid complexes. The nucleating temperature used is above the melting point of the amylopectin crystals. The propagating temperature used is above the melting point of any amylose-lipid complexes but below the melting point of the enzyme RS. The high melting point of the enzyme RS permits its use in baked good formulations. Partial acid hydrolysis (PAH) of a high-amylose corn starch (aeVII) enhances the effects of hydrothermal treatments used to produce granular RS, which is stable to further heat treatment at atmospheric pressure . PAH of ae-VII starch involved heating 35% (w/v) starch suspensions with 1% (w/w) HCl at 25 C for up to 78 h. PAH followed by heat moisture treatment tended to increase yield of boiling-stable granular RS to the maximum of 63.2%. Selective heat treatment of high amylose starch in the presence of agents inhibiting the swelling of starch like alkali and alkaline earth metal salts of halides, sulfates, and phosphates yield granular RS with high dietary fiber. Recently, pyrodextrinization has been recognized as a way of producing a RS that is water-soluble and has non-starch linkages . Pyroconversion refers strictly to the modification of dry starch through heat treatments, with or without addition of acids. Acids used include hydrochloric acid at 0.15% (based on starch dry weight) and orthophosphoric or sulfuric acids

at 0.17% . Commercial pyrodextrins are generally produced by heating dry, acidified starch in a reactor with agitation. Acid may be sprayed on the starch to facilitate hydrolysis and transglycosidation. Depending on reaction conditions, pyroconversion produces a range of products that vary in digestibility, available starch, viscosity, cold-water solubility, swelling power, color, and stability. The production of indigestible dextrins or pyrodextrins by heat-treating potato starch in the presence of an acid and then refining the product has been described . Enzymic treatment The possibility of preparing a RS concentrate from isolated pea starch was investigated, and sorption of hydrophobic substances (indicative of health-benefiting properties) by such a concentrate . By use of a thermally stable _-amylase, a preparation of up to 70% RS containing a mixture of mineral and organic N compounds was obtained. The pea RS concentrate had an affinity to bile acid, deoxycholic, and cholesterol; however, its affinity to cholesterol was not as efficient as that of native pea starch. The results concluded that the pea RS concentrate may be potentially used as a food component in special diets, or for preventive, prophylactic, and therapeutic purposes. Readily fermentable heat-stable RS of optimal chain length from poly-1,4-_-D-glucan useful in various functional foods can be obtained by in vitro synthesis by adding an enzyme extract containing the amylosucrase of Neisseria polysaccharea to sucrose solutions, followed by incubation at 37 C over several hours . A method has been discovered to produce an RS product that retains the same cooking quality as found in untreated rice starch or flour, but has a higher percentage of starch resistant to _-amylase digestion . This method uses a debranching enzyme, that is, pullulanase, to digest the starch, but does not require pretreating the starch source before enzymatic treatment. This method produced RS from low amylose starches, rice starch

(24%), and rice flour (20%). Surprisingly the RS product formed by this method retained the pasting characteristics of the untreated flour or starch and was heat stable. This method may also be used to produce RS from other botanical sources, that is, corn, wheat, potato, oat, barley, tapioca, sago, and arrowroot. Heat and enzyme treatment Preparation of RS to be used as a food-grade bulking agent, by retrogradation of starch followed by enzymatic or chemical hydrolysis to reduce or remove the amorphous regions of retrograded starch . RS can be prepared from high amylose starch by gelatinization followed by treating the slurry with debranching enzymes like pullulanase and isolating the starch product by drying/extrusion. Controlled heat treatment of starch so as to achieve swelling and at the same time retain its granular structure followed by enzymatic debranching and annealing at suitable temperature followed by drying produces RS. These RS find applications in a variety of foods and beverage products. Purified RS products having at least 50% RS content can be produced by forming a water-starch suspension wherein the ratio of starch to water is approximately 1:2 to 1:20, heating the waterstarch suspension in an autoclave at temperatures above 100 C. to ensure full starch gelatinization and then cooling to allow amylose retrogradation to take place. It is reported that best results were obtained at a temperature of 134 C, with 4 heating and cooling cycles and a starch:water ratio of 1:3.5. The RS was purified by comminuting the starch gel and mixing it with an amylase to digest non-RS fractions, leaving RS. The amylase is inactivated by heat treatment above 100 C . For the preparation of a fragmented starch precipitate for use in reduced-fat foods, a debranched amylopectin starch is precipitated and then fragmented. The debranched amylopectin starch may be derived from a starch that contains amylopectin, for example, common corn starch and waxy maize starch, by gelatinizing the starch, followed by treatment with a debranching enzyme, such as isoamylase or pullulanase, and precipitation of the debranched starch. To form the precipitate, the solution is cooled to ambient temperature, to reduce the solubility of the debranched starch. The

precipitate may then be heated to about 70 C, while in contact with a liquid medium, to dissolve at least a portion of the precipitate. Reprecipitation by cooling of the suspension/solution may then be employed. Repetition of the dissolving and the reprecipitation tends to improve the temperature stability of the resulting aqueous dispersion as was observed on repeating the cycle of heating and cooling, a total of 8 times . A process for increasing the amount of amylase-RS (to a minimum of 15%) in high amylose starch, such as Hylon V or Hylon VII consisted essentially of gelatinization of a starch slurry, enzymic debranching of the starch, and isolation of the starch product by extrusion or drying. A further increase in amylase-resistant starch was obtained by addition of an inorganic salt to debranched starch before isolation . Chemical treatment In type IV RS, the enzyme resistance is introduced by modifying the starch by crosslinking with chemical agents . Crosslinked starches are obtained by the reaction of starch with bi- or polyfunctional reagents like sodium trimetaphosphate, phosphorus oxychloride, or mixed anhydrides of acetic acid and dicarboxylic acids like adipic acid. Cross-linking carried out by sulphonate and phosphate groups between various starch molecules involves their hydroxyl group thus bringing resistance to amylolytic attack on the starch molecule. Figure 9 shows the preparation of distarch phosphate ester. Distarch phosphates with 0.4% to 0.5% phosphorus have been prepared and they contain both slowly digested starch (SDS) and RS4. The modified starches were obtained in quantitative yield, and provided 13% to 69% of SDS and 18% to 87% RS4. RS4 starches with low swelling power have also been prepared similarly from wheat, corn, waxy corn, high amylose corn, oat, rice tapioca, mung bean, banana, and potato starches. Phosphated di-starch phosphate, a modified RS made from high amylose maize starch, is currently used as food additive (E1413) in the EU.

Determination of RS In vitro methods The main step of any method to measure the content of RS in foods must first remove all of the digestible starch from the product using thermostable _-amylases . At present, the method is considered the most reproducible and repeatable measurement of RS in starch and plant materials, but it has not been shown to analyze all RS as defined (Champ and others 2003). It is based on the principle of enzymic digestion and measures the portions of starch resistant to digestion at 37 C that are typically not quantitated due to the gelatization at 100 C followed by digestion at 60 C. Two general methods specifically proposed to determine RS remove digestible starch using different amylases, and the residual fraction is quantified after solubilization in 2M KOH. The Siljestrom and Asp procedure includes preparation and quantification of dietary fiber residue before RS determination. This is usually done by drying the samples at 105 C. As heating influences the RS content in foods, results may be modified by this step. A modified method for measuring RS in dietary fiber residues from various sources involves mixing fiber residues with KOH, acetate buffer, and HCl. After incubation with amyloglucosidase samples are centrifuged and diluted with distilled water. RS is calculated as glucose (mg) 0.9. Advantages of the method are the use of small amount of sample, less reagents and elimination of drying. In vivo methods Different methods are used to analyze RS in vivo. One of the ways to assay RS physiologically is to determine starch in the undigested ileal content. Terminal ileal samples can be recovered by intubation or from ileostomy bags. This implies measuring the area under the curve (AUC) of the serum glucose concentration over the first 2 h after administering a starch and dividing

this by the serum glucose response after consumption of an equal amount of glucose. Determination of breath hydrogen (breath tests) can also be used as a semiquantitative measurement for RS. In a study on effect of RS on human colon, increased fermentation was verified by elevated breath hydrogen excretion . From the different animal models, the antibiotic-treated rat model is the one commonly used.

References: Champ, M.,Langkilde A.M, Brouns, F. and Kettlitz, B. (2003a).Advances in the dietary fibre characterization .1. Definition of dietary fibre, physiological relevance ,health benefits and analytical aspects Nutr. Res. Rev. 16: 71-82,http://journals .cambridge. org/action/display Abstract from pg=online&aid=607688. Champ, M.,Langkilde A.M, Brouns, F. and Kettlitz, B.(2003b). Advances in dietary fibre characterization. 2. Consumption, chemistry, physiology and measurement of resistant starch; implication for health and food labeling. Nutr. Res. Rev. 75:733-747. http://journals. Cambridge.org/action/display abstract from page & aid=online=607768. Champ,M. (2004). Resistant Starch,Starches in food, Elisson, Ann-Chalotte, ed., Boca Raton: CRC-Press: 560-574. Englyst, H.N. and Cummings, J.H. (1987). Digestion of the polysaccharides of potato in the small intestine. Amer. J. Clin. Nutr, 45: 423-431. Englyst, H.N., Kingsman S.M. and Cummins, J.H. (1992). Classification and measurement of nutritionally important starch fractions. Eur. J. Clin. Nutr. 46(2): 33-50. Hylla, S. Gostre. A. Duesl, G., Anger, H., Bartram, H.P., Christl, S.U., Kasper, H., Scheppach, W. (1998). Effects of resistant starch on the colon in the healthy volunteers: possible implications for cancer prevention. Amer. J. Clin. Nutr. 67: 136-142. Premavalli, K.S., Roopa, S. and Bawa, A. (2006). Resistant starch:A functional dietary fibre. Indian food Industry. 25(2): 40-45.

Credit seminar On Resistant starch: An alternative source of dietary Speaker: Monika Mathur Addm. 2006FST145M fibre.