Starch is widely applied in various industrial sectors, including the food industry. Starch is used as a thickener, stabilizer, or emulsifier. However, arrowroot starch generally has weaknesses, such as unstable under heating and acidic conditions, which are generally applied to processing in the food industry. Modifications were applied to improve the characteristics of native arrowroot starch. In this study, arrowroot starch was modified by heat-moisture treatment (HMT), octenylsuccinylation (OSA), and dual modification between OSA and HMT in a different sequence––namely, HMT followed by OSA, and OSA followed by HMT. This study aims to determine the effect of different modification methods on the physicochemical and functional properties of native arrowroot starch. The result shows that both single HMT and dual modification caused damage to native starch granules, such as the formation of cracks and roughness. For single OSA treatment, especially, there is no significant change in granule morphology after modification. All modification treatments did not change the crystalline type of starch but reduced the RC of native starch. Both single HMT and dual modifications  increased pasting temperature and setback, but, conversely, decreased the peak and the breakdown viscosity of native starch, whereas single OSA had the opposite trend compared with the other modifications. HMT played a greater role in increasing the thermal stability and the retrogradation ability of arrowroot starch. Both single modifications (HMT and OSA) increased the hardness and gumminess of native starch, and the opposite was true for the dual modifications. HMT had a greater effect on color characteristics, where the lightness and whiteness index of native arrowroot starch decreased. Single OSA modification increased swelling volume higher than dual modification. Both single HMT and dual modifications increased water absorption capacity and decreased the oil absorption capacity of native arrowroot starch. Maranta arundinacea L.) is a tuber plant with a rhizome root, which is elongated like an arrow. Arrowroot tubers have been considered inferior commodities and have not been utilized optimally. Arrowroot tuber is one of the commodity sources of carbohydrates, which can be processed into starch or flour. Starch can be utilized in many ways, such as a food thickener, stabilizer, and emulsifier [Arrowroot  cultivated in Pangandaran, West Java, Indonesia were used as the raw material for starch extraction. All chemicals used for the modification process were 2-octen-1-ylsuccinic anhydride (OSA) , hydrochloric acid, sodium hydroxide, ethanol 95%, acetic acid, isopropanol, AgNO3, and phenolphthalein, with specification analytical grade.Arrowroot tubers  starch suspension by dissolving the starch in distilled water. The pH of the starch suspension was then adjusted to pH 8 using 1 M NaOH before the addition of 3% OSA solution (w/w). The esterification was then carried out by stirring the solution for 3 h. During the modification process, the pH of the suspension was adjusted and maintained in the range of 8.0–8.5 using 1 M NaOH and 1 M HCl. After the reaction, the starch suspension was adjusted to pH 6.5 using 1 M HCl. Starch was centrifuged at 5000 rpm for 2–3 min. The precipitated starch was washed using distilled water and then centrifuged again, and this process was repeated 2–3 times. The starch was then dried in a drying oven at 50 °C for 24 h. The dried modified starch was ground and sieved (no. 100 mesh sieve).Preparation of OSA-starch refers to Marta et al.  with a sThe native starch of arrowroot was modified by HMT (as mentioned in The native starch of arrowroot was modified by OSA (as mentioned in The granular morphology of native and modified arrowroot starch was determined using scanning electron microscopy (SEM), a JEOL JSM-6360 LA at 15 kV . Mounted starch samples were coated with gold/palladium at 8–10 mA for 10–15 min under low pressure (less than 10 tors). Representative digital images of native and modified starch granules were obtained at 5000 and 10,000 magnifications.The crystallinity of native and modified arrowroot starch was measured using an X’Pert PRO series PW3040/60 X-ray diffractometer  that operated using Cu-K alpha radiation with a wavelength of 1.540 nm as an X-ray source at 40 kV and 30 mA. The diffraction angle (2θ) scanning was from 3.0° to 50.0° with a scanning rate time of 2.9 s. OriginLab Program was used to calculate the relative crystallinity of the starch samples.o), peak temperature (Tp), conclusion temperature (Tc), and enthalpy of gelatinization (∆H). Starch was made into a slurry, with the ratio of water and starch being 3:1. The slurry was then hermetically sealed using a DuPont encapsulation press before weighing. The starch samples were heated at a rate of 5 °C/min from 20 to 100 °C.Thermal properties of starch samples were measured using DSC-Q100, TA Instruments . The parameters observed were onset temperature  used throughout the analysis, and the total analysis time was 12 min. The pasting properties included the following parameters: pasting temperature (PT), peak viscosity (PV), hold viscosity (HV), final viscosity (FV), breakdown (BD), and setback (SB).The texture properties of starch gels were evaluated on a TA-XT express enhanced Stable Micro System . Exponent Lite Express software was used to collect and calculate the data obtained. The gelatinized starch in the canister after the RVA measurement was poured into cylindrical plastic tubes  and then kept at 4 °C for 24 h to form a solid gel. Each gel sample in the tube was penetrated with a cylindrical probe (P36/R) at a speed of 5 mm/s to a distance of 10 mm for two penetration cycles. The texture profile curves were used to calculate hardness, adhesiveness, springiness, cohesiveness, and gumminess.Swelling volume and solubility of arrowroot starch were measured referring to Marta et al. . A totalAfter being separated, the supernatant was dried in a drying oven for 24 h. Solubility was calculated using the equation:Water absorption capacity (WAC) and oil absorption capacity (OAC) were measured referring to Marta et al. . One graFreeze-thaw stability or syneresis was determined by a previous method  with a sThe starch color was measured using a CM-5 Spectrophotometer  with SpectraMagic NX version 3.0 Software. The samples were placed in a glass cell and above the light source. After that, measurements were taken at room temperature. The parameters measured were CIE-lab namely, the L*, a*, and b* values in each sample. L* value indicates the lightness (whiteness/darkness), representing dark (0) to light (100); a* value indicates the degree of red-green color ((+) redness/(−) greenness); and b* value indicates the degree of the yellow-blue color ((+) yellowness/(−) blueness). The whiteness index for native and modified starches was determined using the following equation :(6)WhiteTotal color differences between starch samples were calculated using the following equation :(7)ΔE=L*L*, a*, and b* were parameters for modified starch, and p < 0.05). The statistical software program IBM SPSS Statistics version 25 was used to examine all of the data.Data are displayed as the mean ± SD. Experiments were all performed in triplicate. One-way analysis of variance (ANOVA) was used to analyze all of the data, and the Duncan test was used to compare the sample mean at a significance level of 5%  Tp, or peak temperature, represents the the endothermic peak on the DSC thermogram; (3) Tc, or conclusion temperature, is the final temperature at which the sample is wholly gelatinized, and is sometimes known as the crystalline melting temperature ; and (4) ΔH or enthalpy (J/g) is calculated based on the DSC endotherm, expressing the energy required to break the double helix structure during starch gelatinization [DSC is used for thermal analysis in order to determine the transition of starch crystallinity caused by heating with the following parameters: (1) Tnization ,16.o than native starch, except for HMT starch. Tp native starch decreased after modification from 50.46 °C to 39.83–44.36 °C. Both single methods (OSA and HMT) decreased Tc, but, conversely, both dual modification methods increased the Tc of native starch. The temperature range (Tc–To) of modified starches was higher than native starch, except for HMT-starch, whereas the enthalpy of modified starch decreased significantly from 1117.08 (J/g) to 936.44–647.30 (J/g).The thermal properties of native and modified arrowroot starches are presented in The viscoamylograph and pasting properties parameters of native and modified arrowroot starches are presented in Texture properties parameters of native and modified arrowroot starches are presented in OSA modification, for both single OSA and dual HMT-OSA, significantly increased the SV of native starch, but we saw a converse trend for OSA-HMT. All modified starch has lower solubility and higher syneresis than native starch . HMT staWAC describes the amount of water available for gelatinization . WAC of The color parameters of native and modified arrowroot starches are L*, a*, and b*, whiteness index, and ΔE . HMT modNative arrowroot starch has a round, oval shape with a granular surface that is slightly textured without cracks, and this is in line with some of the previous studies ,19. The X-ray Diffraction (XRD) is a method for assessing and quantifying long-range crystalline order in starch . Figure Relative crystallinity (RC) was calculated based on the ratio of the diffraction peak area  to the total diffraction area . Arrowroo, Tp, and ΔH compared to native starch, which is in agreement with some of the previous studies [c and ΔH in HMT-treated starch [o, Tp, Tc) than native starch [o and Tp of HMT starch were not significantly different to native starch. Both dual-modified starch HMT-OSA and OSA-HMT starches tend to have thermal characteristics that are almost similar, where the To, Tp, Tc, and Tc–To are not significantly different from each other, whereas ΔH of OSA-HMT starch is significantly higher than HMT-OSA starch.Esterification using OSA on arrowroot starch causes a decrease in T studies ,37. The  studies , whereasd starch . Some pre starch ,30,40,41The increase in PT and the decrease in PV on HMT starch, both for single HMT and dual modifications (HMT-OSA and OSA-HMT), could be due to the partial breakage of the ordered chain structure and the rearranging of the broken molecules, facilitated by the high temperature (100 °C) and limited moisture content (30%) during HMT. As a result, the forces of the intra-granular bonds would be augmented, and the linkages between starch chains would be strengthened. The HMT-treated starch samples needed greater heat for structural breakdown and paste production, resulting in a lower paste viscosity . On the The significant difference in PV between HMT-OSA and OSA-HMT-treated starch is because HMT facilitated this OSA particle to attack more of the –OH group in starch. According to Park et al. , HMT befFrom the industrial point of view, the modification of arrowroot starch could give arrowroot more value, as the starch could be used more in the food industry. HMT and dual-modified starches (OSA-HMT and HMT-OSA starcher) have a higher thermal stability than native starch, as shown by the lower breakdown viscosity of the starches. Starch with good thermal stability could be used as a thickening agent for food products that need to be sterilized, e.g., sauce, paste, etc. , whereasTexture parameters in starch have an important role because starch can be a texturizer agent, such as a thickening and gelling agent. TPA  is used to observe texture parameters, such as hardness, adhesiveness, springiness, cohesiveness, and gumminess . Gel harSwelling volume (SV) measures the hydration capacity of starch molecules due to the presence of water trapped in granules . SV is rThe SV of HMT starch decreased when compared to native starch, which is in line with other studies ,12,52. FAll modified starches have lower solubility than native starch, and this is caused by amylose leaching during the starch gelatinization process. Amylose is in the crystalline region and is relatively small in size and linear shape, making it easier to leach out from the starch granules . HoweverAmong the modified starches, OSA starch has the lowest WAC (0.83 g/g db). The reduced WAC on OSA starch was due to the presence of a hydrophobic substituent group from OSA that replaces the hydroxyl group on starch granules, which causes an increase in the hydrophobicity of starch , whereasThe esterification process with OSA increased starch’s hydrophobicity, thereby increasing the OAC . On the Freeze-thaw stability was determined as syneresis (%). Syneresis releases water from gel or starch paste during cooling, storage, and freeze-thawing . High syHMT and OSA-HMT decrease SV and SOL and increase the WAC of native starch, which indicates that HMT and OSA-HMT starches can be applied in pasta and noodle formulations. Marta et al.  have repColor is a characteristic that has an important role in determining the quality and the level of consumer acceptance in selecting a product. In flour or starch products, consumers generally prefer products that are white or bright in color. To determine the color of both native and modified arrowroot starch objectively, color analysis was performed. Color testing of products, mainly arrowroot starch, can be carried out using a stand-alone, top-port color measuring instrument, which was the Spectrophotometer CM-5 in this research. The color of each starch sample was then interpreted by referring to the CIELAB systems through the L*. a*, and b* values .The highest lightness (L*) was shown by OSA starch (96.39), while HMT starch showed the lowest lightness (94.96). There was a significant difference in lightness, especially for HMT-modified starch. This is presumably because thermal treatment can cause the degradation of color pigments in starch. The redness (a*) in arrowroot starch with a negative value is indicated by both native and OSA starches, which indicated a tendency to be green in color . MeanwhiThe whiteness index (WI) is based on a scale of 0–100, with the highest value described as the highest level of lightness. The WI value is positively correlated with the L* value; the higher the WI, the higher the L* value. HMT-modified starch, for both the single HMT and dual-modified starches, had low WI. The HMT modification process that was carried out reduced the lightness level of starch. Total color difference (∆E) is a parameter used to assess how much change or difference can be seen in the Lab* value results in ingredients or food after specific treatments . Dual moAll modification treatments, both for the single and dual modifications, had significant effects on granule morphology, crystallinity, thermal, pasting, and functional properties, texture, and color characteristics of native arrowroot starch. The granules of arrowroot starch are spherical and ellipsoid to oval shaped. There was no damage on the surface of the starch granules after OSA. In HMT-modified starch, both for single and dual modifications , the damage occurred on the surface of the granules. Native and modified starches have a similar crystallinity pattern, which was an A-type crystalline pattern. All modification treatments decreased the RC of native starch, where HMT has a greater effect on RC than OSA. Both single HMT and dual modification  increased PT and SB, and, conversely, decreased PV and BD viscosity of native starch. All HMT treatments, both for single and dual modifications, can improve the thermal stability of native starch, especially the HMT single treatment, whereas OSA single treatment has the opposite trend compared with the other modification treatments. OSA starch has a firm texture characteristic, which is indicated by the high value of hardness and gumminess. Both single OSA treatment and HMT-OSA significantly increased the SV of native starch, but, conversely, trend for the OSA-HMT. All modified starches have lower solubility and higher syneresis than native starch. Both single HMT modification and dual modifications  increase WAC and decrease the OAC of native starch. HMT starch for both single and dual modifications have low L* and WI values, while OSA starch has the brightest color among other starches. Based on pasting and functional properties, both HMT and OSA-HMT starch can be applied in pasta and noodle formulation and also as a thickening agent, whereas OSA starch can be used as an ingredient in baby food and bakeries because it has a low ability to retrograde.


The results showed that DHT had changed the morphology of BH starch, and the diffraction pattern remained an “A”-type crystalline structure. However, with an extension of DHT temperature and time, the amylose content, gelatinization temperature, enthalpy value, swelling power, and pasting viscosity of modified starches decreased, while the light transmittance, solubility, and water and oil absorption capacities increased. Additionally, compared with native starch, the content of rapidly digestible starch in modified samples increased after DHT, whereas those of slowly digestible starch and RS decreased. Based on these results, the conclusion could be drawn that DHT is an effective and green way to transform multi-structures, physicochemical properties, and in vitro digestibility of BH starch. This fundamental information might be meaningful to enrich the theoretical basis of physical modification on BH starch and extend the applications of BH in the food industry.As a physical method for starch modification, dry heating treatment (DHT) at high temperatures  was applied to blue highland barley (BH) starch with different durations (2 and 4 h). The effects on its multi-structures, physicochemical properties, and  Hordeum vulgare L. var. nudum Hook. f) and is known as “Qingke” in Mandarin. In China, highland barley is mainly distributed on the Qinghai–Tibet Plateau . It has been the essential staple food crop and economic crop relied on by Tibetans since the fifth century AD  was purchased from Nanjing Duly Biotech Co., Ltd . All the other chemicals were of analytical grades.The BH grains (Ganqing 4#) with 14.6% moisture content were purchased from Qinghai Xinning Biotechnology Co., LTD. Standard amylopectin , amylose , and porcine pancreatic α-amylase  were purchased from Sigma–Aldrich Chemical Co. . The amyloglucosidase from The extraction of BH starch was performed according to the previously described literature by Zhao et al. . The obtThe NBHS sample  was weighted into a heat-proof dish and distributed evenly in a thin layer (~1 mm). The dish was covered with aluminum foil to ensure no loss of material. Then, the sample dishes were heated in the oven at 150 and 180°C for 2 or 4 h, respectively . The corA scanning electron microscope  was used to observe the morphology of different samples. Different samples were adhered to a double-sided adhesive tape on a metal stub. The samples were coated with 20 nm of gold under vacuum and observed at an acceleration potential of 20 kV.The native and modified samples were obtained by a CLSM according to the method reported by Gou et al. .The XRD pattern of different samples was determined by an X-ray diffractometer . The scanning angle (2θ) was from 5° to 60° at a scanning rate of 4°/min under 40 kV at 30 mA current. The relative crystallinity  was calculated with Jade 6.0 software .−1.The FT-IR spectra of samples were determined by a Vertex 70  in a range of wavenumber from 400 to 4,000 cmL*, a*, and b* color systems.Color measurement of samples was carried out with a Hunter Colorimeter . It was equipped with an optical sensor based on Amylose content (AMC) was determined by the iodine-binding procedure of Juliano et al. . The alkg for 15 min. The volume of decanted supernatant fluid was measured, and milliliters of water (or oil) retained per gram of different samples were calculated.The oil and water absorption capacities of samples were determined according to the method reported by Liu et al. . The sam1), and mixed with 5 ml of distilled water. The tubes were incubated in a shaking water bath at 50, 60, 70, 80, and 90°C for 30 min, respectively, then cooled to room temperature and centrifuged at 657 g for 15 min. The supernatant was carefully decanted, and the resulting precipitate was weighed (W2). Solubility and SP per 100 g of sample on db were calculated using the following equations:The solubility and SP of samples were determined following the method of Liu et al. . The samPaste clarity  of samples was measured with the method reported by Zou et al. . The traTo), peak temperature (Tp), conclusion temperature (Tc), and gelatinization enthalpy (ΔH) were determined.The gelatinization characteristics were assessed with a differential scanning calorimeter . The sample , mixed with 9 ml distilled water, was sealed and equilibrated at room temperature overnight. The sample pans were heated from 30 to 120°C at 10°C/min, and an empty pan was taken as control. Onset temperature  and distilled water (25 g) were dispersed evenly in an aluminum canister. A Rapid Visco Analyzer  was used for the evaluation of pasting properties. The testing cycle was set at 50°C for 1 min, ramped to 95°C in 3.7 min, held at 95°C for 2.5 min, cooled to 50°C in 3.8 min, and finally held at 50°C for 2 min. Setback viscosity (SB), breakdown viscosity (BD), peak viscosity (FV), pasting temperature (PT), and final viscosity (FV) were measured.In vitro digestibility of different samples was performed according to previous research by Gao et al.  by Duncan's multiple range test using SPSS version 19.0 software . The least significant difference (LSD) test was performed to determine differences among the treatments. The principal component analysis (PCA) was conducted with Minitab version 17 . Statistical significance was set at a level of The SEM images of different starches are shown in A ring structure of amylose and amylopectin in starch granules grows outward from the hilum. The hilum contained substantial quantities of moisture, resulting in softening to be easily eroded by heating. It explained that the hilum of NBHS was more vulnerable to being sunken and damaged during DHT. The leaching of amylose and the reorganization of amylose–amylopectin possibly contributed to these morphological changes . MeanwhiThe CLSM images of native and dry-heated samples are shown in The XRD pattern and RC of native and dry-heated starches are presented in Compared with the RC of NBHS (29.8%), the RC of modified starches was significantly decreased, ranging from 28.2 to 21.9% with the order of BH150-2 > BH150-4 > BH180-2 > BH180-4. The reduction in RC of modified starches was dependent on the temperature and duration of DHT, which might be attributed to the movement of double helical, partial gelatinization of starch granules, and even degradation of the crystalline region during DHT , 33. In −1 are shown in −1 are shown in The FT-IR spectroscopy represents the short-range ordered structure of starch . The FT-−1, respectively, represent the content of crystalline starch and amorphous starch. The absorbance ratio of 1,047/1,022 suggests the amount of ordered crystalline to amorphous domains in starch  to 89.3 (BH180-4), while increased the a* value from 0.14 (NBHS) to 1.05 (BH180-4) and b* value from 1.4 (NBHS) to 13.6 (BH180-4).The influence of DHT on the color parameters of starches is shown in L* value indicated that the NBHS turned darker after DHT and was pronouncedly followed by the temperature increase. The a* value is relevant to redness characteristics. The increase in a* value showed that the DHT led to more browning in modified starches, probably due to caramelization reactions  and enthalpy (ΔH) of native and DHT-modified starches are presented in o, Tp, Tc, and ΔH of modified starches significantly decreased along with increased DHT temperature and duration. The gelatinization temperatures are indicative of the crystalline perfection in starch, where the more perfect crystallites show a higher To , the disruption of double helices in the crystalline region would also cause an increase in RDS content  is rich in dietary nutrients, its health-promoting potential is yet to be explored. In this study, DHT, as a safe and maneuverable physical method for starch modification with limited by-products, was introduced to modify BH starch. The effect on structural, physicochemical, and digestive properties of BH starch showed a thermodynamic impact with heating temperature and duration variations. The DHT greatly increased potholes and fissures on the surface of BH starch, while decreasing the AMC, RC, and RThe original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.SL: conceptualization, investigation, methodology, formal analysis, validation, and writing-original draft. HL: supervision, formal analysis, writing-review and editing, and funding acquisition. SG: writing-review and editing and data curation. SsG: methodology. CZ: resources and funding management. All authors contributed to the article and approved the submitted version.


Resistant starch (RS) has become a popular topic of research in recent years. Most scholars believe that there are 5 types of RS. However, accumulating evidence indicates that in addition to starch–lipid complexes, which are the fifth type of RS, complexes containing starch and other substances can also be generated. The physicochemical properties and physiologic functions of these complexes are worth exploring. New physiologic functions of several original RSs are constantly being discovered. Research shows that RS can provide health improvements in many patients with chronic diseases, including diabetes and obesity, and even has potential benefits for kidney disease and colorectal cancer. Moreover, RS can alter the short-chain fatty acids and microorganisms in the gut, positively regulating the body’s internal environment. Despite the increase in its market demand, RS production remains limited. Upscaling RS production is thus an urgent requirement. This paper provides detailed insights into the classification, synthesis, and efficacy of RS, serving as a starting point for the future development and applications of RS based on the current status quo. This paper provides detailed insights into the classification, synthesis, and efficacy of resistant starch, guiding the future development and applications of resistant starch based on the current status quo.5 to 1 × 106. Further, each chain of amylose contains ∼200 to 700 glucose residues. In contrast, amylopectin is a much larger molecule than amylose, with a molecular weight of 1 × 107 to 1 × 109. Its degree of polymerization ranges from 9600 to 15,900 glucose units [Starch is a polymer composed of glucose subunits. Based on the type of polymerization, starch can be divided into 2 categories: amylose and amylopectin. Amylose has a linear structure consisting of D-glucose residues linked by α-1,4-glycosidic bonds and a molecular weight of ∼1 × 10As a carbohydrate, starch is a key source of energy for physiologic processes in the human body. Previously, it was believed that all starches can be enzymatically digested in the digestive system, before finally being absorbed into circulation. However, with additional research, a unique type of starch that cannot be digested and absorbed by the small intestine was discovered . In 1982In addition to the naturally occurring RS that was originally discovered by Englyst, different types of RS have been synthesized through the artificial modification of starch. These novel RSs have shown better performance than natural RS. The increase in the demand for RS has bolstered research on RS synthesis, with scientists hoping to mass-produce RS to meet consumer needs. As a natural ingredient, RS can have broad applications in the food and health industries. RS can be used to improve food processing given its low water-holding capacity and fine texture-related organoleptic properties . It alsoIn this review, we describe the recent classification of RS, its in vivo digestion, the current progress in RS synthesis methods, and the physiologic functions of RS in the human body. This summary of the latest findings on RS could inspire the practical application of RS.1, RS2, and RS3. Natural starch granules are usually encapsulated within plant components , forming a physically embedded starch called RS1. This encapsulated structure hinders contact between amylase and starch within the digestive system, resulting in enzymatic protection. RS1 is mainly found in natural grains and beans that have not been ground sufficiently. In contrast, RS2 is a natural raw starch granule with a special crystalline structure and high-starch density. This special property reduces its sensitivity to enzymes. Therefore, RS2 can resist the hydrolysis caused by digestive enzymes to a certain extent. Raw potatoes and bananas have a high content of RS2. However, this type of starch is easily destroyed by heat. When starch is heated to a temperature higher than its gelatinization temperature, its original crystal structure is destroyed. When the gelatinized starch is placed at a low temperature for a certain period of time, it undergoes a retrogradation process to produce retrograded starch, namely RS3.When Englyst et al.  first pr4—a class of chemically modified starches. RS4 is resistant to digestive enzymes because its original functional groups have been chemically modified or new functional groups have been introduced to produce carboxymethyl starch, starch ether, starch ester, and cross-linked starch [5 [Following in-depth research on RS, Englyst et al.  identifid starch . In additarch [5 .During food processing, starch often interacts with other ingredients. In addition to lipids, other nutrients, such as proteins, are also present. Several studies have demonstrated the formation of ternary starch–lipid–protein complexes that affect starch digestibility during processing . It rema1) improving blood glucose and insulin levels, 2) reducing blood lipid levels, and 3) improving defecation [Due to its complex molecular structure and different functions, the definition of dietary fiber has been debated for many years. According to most definitions, dietary fiber includes 4 major substances: resistant oligosaccharides, nonstarch polysaccharides, RS, and noncarbohydrate compounds . In termfecation . The Eurfecation ,32. Tradfecation . As a hefecation . Thus, ffecation , with fua substrate that is selectively utilized by host microorganisms, conferring a health benefit” [The definition of prebiotics has continuously been updated since it was first put forth by Gibson and Roberfroid  in 1995.benefit” . In recebenefit” . The latbenefit” ,42. In abenefit” .2, RS3, and RS4. The results show that the effects of RS differ among different microbial species, even within the same colony. However, RS generally increases the number of bacteria beneficial to the human body. Therefore, to understand whether RS can be converted into a prebiotic, different types of RS need to be studied.Compared with its dietary fiber-related properties, the prebiotic properties of RS are more challenging to observe. There is a great difference between the results obtained in vitro, in animal studies in vivo, and in clinical trials. To study the prebiotic properties of RS, carefully designed human studies are required. The fifth part of this review collates all current population-based randomized controlled trials on RS and its effects on the gut microbiota. These studies focus on RSOverall, dietary fiber, RS, and prebiotics appear to share some overlap. Although RS is a dietary fiber, all dietary fibers are not prebiotics, and all types of RS are also not prebiotics.The 4 kinds of traditional RSs resist decomposition by digestive enzymes in the small intestine and enter the large intestine. The large intestine has an anaerobic environment and various microorganisms, including beneficial bacteria.Ruminococcus bromii and Bifidobacterium adolescentis [Ruminococcus bromii plays an important role in degrading RS to produce SCFA, promoting the reproduction of other beneficial bacteria through cross-feeding [Bifidobacterium adolescentis and Ruminococcus bromii play a similar role, they release lactic acid and polysaccharides for cross-feeding through 2 different pathways, which can stimulate the production of butyrate by other intestinal bacteria [RS interacts with gut microbes to produce SCFAs and some gases. SCFAs include acetic acid, butyric acid, and propionic acid, and the latter 2 account for ∼80% of all gut SCFA. Moreover, the gases include carbon dioxide, methane, hydrogen, and hydrogen sulfide. Gut microbes have different sensitivities to different types of RS ,45. Correscentis . The met-feeding ,48. Althbacteria .It should be noted that the digestion of starch–lipid complexes differs from that of other RSs. Rice starch–oleic acid complexes are beneficial for the reproduction of butyrate-producing bacteria in rats . This coIn the small intestine, the original structure of starch is destroyed by pancreatic α-amylase, resulting in the release of amylose. Lipids are digested by lipase to produce FFAs. In the neutral environment of the small intestine, amylose encounters FFAs, which likely promotes the formation of starch–lipid complexes. However, in the small intestine, starch is broken down into monosaccharides, and FFAs are reassembled into micelles . These 2In vitro experiments to measure the content of RS have largely been performed using 2 main methods: the direct method and the indirect method. Direct measurement is performed by converting RS into glucose . First, Canna edulis has also been explored as a raw material for RS synthesis [Many factors affect the RS yield during the synthesis process. The most important of these factors is the raw material used. ynthesis .TABLE 1YCarbohydrates are an essential component of legumes, but their composition varies across different legume species. Generally, the content of RS and amylose in legumes is higher than that in wheat and potatoes. However, there are few studies on the synthesis of RS from legumes . The proIn recent years, traditional Chinese medicinal materials have gradually become more well-recognized. Many medicinal foods not only form a part of the daily diet but also play a role in nutraceuticals and health care. Lotus seed was categorized as a dual-purpose resource for food and drugs by the National Health and Planning Commission of China. Lotus seed starch contains 40% amylose and is thus a rich source of amylose . Puerari3 from different varieties of sweet potato and found that with the same synthesis method, purple sweet potatoes provided the highest RS3 content. Even the same plant variety can show different starch contents based on its place of origin.Even when the same plant material is used, the final yield of RS can differ based on the plant variety. For example, common corn starch produces lower RS yields than waxy or high-amylose corn starch. Trung et al.  studied 1 and RS2. However, these kinds of RS usually have low-thermal stability, are easily destroyed, and cannot be mass-produced, although a high content of RS in the raw material indirectly reflects the high proportion of amylose. As described in an earlier section, the higher the amylose content, the higher is the RS production. The stability of natural RS2 is poor. Hence, during RS3 synthesis, unstable RS2 needs to be converted to heat-resistant RS3. However, the proportion of different RSs in a product cannot be accurately measured with current detection methods, and only structural qualitative analysis can be used. Therefore, in some experiments, including the study by Wang et al. [The method of starch extraction can also affect the final yield of RS. For example, commercially purchased starch may be low in amylose because of processing. If starch is directly isolated and extracted from rhizomes, it may contain more RSg et al. , the conThe synthesis method is another important factor affecting the RS yield. Based on their principle, synthesis methods can be roughly divided into 3 categories: physical methods, chemical methods, and enzyme treatment methods . Physica3, RS4, and various complexes. Under external factors, such as autoclaving and microwaving, the chain structure of starch cracks, and the internal water is discharged. This leads to the formation of a left-handed hydrophobic helical cavity. On the one hand, a stable double helix structure, RS3, may be formed inside the amylose. On the other hand, other ligands—especially lipids with a hydrophilic head and hydrophobic tail—may also enter the cavity to form single helical structures [3 requires a low-temperature aging process. The formation of other complexes will eventually involve a cooling step, resulting in the generation of RS3. This review only focuses on the RS generated during the synthesis process and does not deeply analyze the thermal stability, thermodynamic properties, and particle morphology of different products. Further research is needed to examine the changes in the crystal structure and physicochemical properties of the products during the synthesis of novel complexes.Among the various starch complexes, the most well-studied is the starch–lipid complex. The starch–lipid complex has a conformational barrier that hinders the entry of digestive enzymes, and the lipids prevent the hydration of starch granules ,98. Simiructures . The masAs can be seen from 1, RS2, RS3, and RS4, whereas the forms of intervention included addition in the form of supplements or ingestion through food.2 can reduce blood urea nitrogen and serum creatinine concentrations in patients with kidney disease [Among the 5 articles describing changes in fasting plasma glucose (FPG) after RS treatment, 3 suggested that RS could significantly reduce FPG levels ,108,115. disease , whereas disease . A meta- disease .These meta-analyses reveal that RS is beneficial for managing chronic diseases and related indicators. However, there are some differences in the results of different studies. These differences can be attributed to the variances in the study population, study period, type of RS used in the study, and quality control during the study.A study indicated that the consumption of 15–60 g/d of RS is effective in improving glycemia, insulin sensitivity, and satiety in healthy adults . The RS Thus, our suggestions for future randomized controlled trials are as follows. If the study includes patients with diabetes, a smaller dose of RS may provide a health benefit, but it should not be <10 g/d. When participants are healthy or have other diseases, an RS dosage of ≥20 g/d will be more helpful to observe an improvement. Similarly, in high-dose population studies, it will be necessary to consider the impact on the digestive system when the dose exceeds 45 g/d, as well as the impact of the high dose on participant compliance. Meanwhile, for individuals in whom RS has been a part of the daily diet for a long duration, more long-term clinical evidence is required for the optimal intake dose.Among the included meta-analyses, only 2 articles found that the duration of treatment has a significant effect. The subgroup analysis conducted by Yuan et al.  showed t2DM) with obesity were better than those in T2DM alone. Xiong et al. [2. The baseline levels of study end points are higher in participants with diseases, providing greater room for improvement. This makes the effect of RS intervention more obvious. The improvement caused by RS is relatively weak in healthy people with normal baseline indicators. Further, dietary patterns differ among patients with different diseases. For example, participants with diabetes pay more attention to the intake of dietary fiber. During the experiment, if the diet of the control and experimental group is not comparable, confounding factors could eventually interfere with the effect of RS. It is better to measure relevant indicators before the experiment and then also compare the intervention and control groups at the end of the experiment. After study completion, this will not only enable the comparison of differences between the intervention and control groups but also those before and after intervention in the same group. Finally, a comprehensive analysis and discussion can be conducted.A study by Gao et al.  includedg et al.  found thg et al.  demonstr2. More rigorously designed research and studies on other types of RS are required. Of all meta-analyses, only 1 study showed that RS2 could significantly reduce appetite when compared with RS1.At present, the RS type adopted by most studies is RSBy modulating hepatic glycogen structure through the gut–liver axis, promoting the production of starch-degrading enzymes, modulating IR, and remodeling the intestinal barrier, RS could have potential applications in the management of diabetes mellitus and obesity ,120. It SCFAs can affect energy metabolism, liver fat metabolism, blood sugar regulation, mineral absorption, and anti-inflammatory mechanisms in humans ,123. SCFBifidobacterium and Ruminococcus bacteria and decreases the number of Firmicutes. These microbiota-related changes reduce intestinal permeability and increase anti-inflammatory capacity in the intestines. Bifidobacterium and Ruminococcus are 2 of the main dominant flora in the human intestinal tract, and they largely affect the function of the entire host microbiome. The increase in Ruminococcus is closely related to the increase in intestinal butyrate concentrations [Eubacterium rectale, which promotes butyrate production [Bacteroidetes, Actinobacteria, and Lactobacillus in the gut [Faecalibacterium may mediate an important anti-inflammatory effect in patients with chronic kidney disease after RS2 intervention. In addition to affecting the number of specific microbes, RS also affects the diversity, richness, and evenness of the gut microbiota [trations . At leasoduction ,132,136. the gut ,131. Laf the gut  suggestecrobiota . As a pr2 is different from their response to RS4. RS4 induces changes at the phylum level, increasing Actinobacteria and Bacteroidetes and decreasing Firmicutes. RS2 does not cause phylum-level changes, but it increases the proportion of Ruminococcus bromii and Eubacterium rectale. In other words, the range of intestinal microecological changes caused by RS4 is greater. Walker et al. [3 can increase R. bromii and Eubacterium rectale in the intestinal flora of overweight men. An in vitro experiment used a pig in vitro fermentation model to study the changes in intestinal microbial composition caused by 3 kinds of RS fermentation [The impact of RS on intestinal microecology can be divided into different parts. Martínez et al.  showed tr et al.  found thentation . The resBifidobacterium was observed [Even the same RS can have different effects on the intestinal microecology of different subpopulations. The changes in SCFA concentration and intestinal microflora after the intake of RS were studied in healthy adults in 2 studies ,136. Oveobserved . The 4 aFaecalibacterium and Roseburia, which are closely related to kidney disease. The studies by Walker et al. [Studies by Laffin et al.  and Kempr et al.  and Salor et al.  showed t2 in patients with irritable bowel syndrome and showed that a certain dose of RS was beneficial in this patient group. A randomized controlled study examined the effects of indigestible carbohydrate supplementation on miR-32 expression in colorectal cells and found that miR-32 levels were significantly elevated in the colorectal mucosa of healthy human participants after 50 d of RS+ polydextrose supplementation [Eubacterium rectum [In addition to the abovementioned physiologic functions of RS, its other potential effects have also been investigated. Most studies have focused on its application in the prevention and treatment of tumors, especially intestinal tumors. Sasidharan et al.  investigentation . In a prentation . Studiesm rectum . This com rectum . RS incrm rectum . It is pRS has characteristics such as a low water-holding capacity and starch-like texture, but it has the physiologic functions of dietary fiber. RS can be added to staple foods, such as bread, noodles, cakes, etc., to improve the taste of food and its nutritional value. In the future, RS can be used as a food additive to develop foods with a low GI and low-caloric value . It can 2DM [2, can be considered as dietary fiber in nutritional ingredient and supplement ingredient labels. We believe that the RS content should be included in nutrition labels to allow consumers to make more informed choices. This could serve as a strategy to prevent chronic diseases, such as diabetes, obesity, dyslipidemia, and colorectal cancer.The European Food Safety Authority declared that RS is beneficial for postprandial blood glucose levels . The FDA2DM . The FDAAll types of clinical research have the same ultimate goal, ie, improving the lives of human beings. Research on blood sugar-related indicators shows that RS can be added to food to control blood sugar levels and improve quality of life in patients with diabetes. Similarly, studies on blood lipids and inflammation indicate that RS can act as an adjuvant for the management of MetS and inflammation-related diseases. From a public nutrition perspective, the intake of reasonable doses of RS as part of the daily diet can provide health benefits. From a clinical nutrition perspective, RS can help in disease prevention and treatment through diet fortification. The selection of appropriate raw materials and synthesis methods, and the study of digestive processes and physiologic functions of RS are key aspects of research. For practical application, a series of problems need to be solved, including the dosage and type of RS, targeted user population, method of administration, and interaction with other foods. This is a long road, and more researchers should devote their efforts toward addressing these issues.The authors’ responsibilities were as follows – YW: conceptualization of the review; ZW, SW: writing; QX, QK, FL, LL, YX: responsible for the final content; and all authors: read and approved the manuscript.The authors report no conflict of interest.Anhui Provincial Natural Science Foundation (grant number: 2208085MH244) and the Major Difficult Diseases Key Project of Integrated Chinese and Western Medicine of Administration of Traditional Chinese Medicine of Anhui Province (2021zdynjb10).Supported by the


Resistant starch appears to have promising effects on hypertension, cardiovascular and enteric illness. The influence of resistant starch on intestinal physiological function has drawn great attention. In this study, we first analyzed the physicochemical characteristics, including the crystalline properties, amylose content, and anti-digestibility among different types of buckwheat-resistant starch. The influence of resistant starch on the physiological functions of the mouse intestinal system, contained defecation, and intestinal microbes were also evaluated. The results showed that the crystalline mold of buckwheat-resistant starch changed from A to B + V after acid hydrolysis treatment (AHT) and autoclaving enzymatic debranching treatment (AEDT). The amylose content in AEDT was higher than in AHT and raw buckwheat. Moreover, the anti-digestibility of AEDT was also stronger than that in AHT and raw buckwheat. The buckwheat-resistant starch can promote bowel intestinal tract movement. The quantity of intestinal microbe was regulated by buckwheat-resistant starch. Our research demonstrates an effective preparation method for improving the quality of buckwheat-resistant starch and found that buckwheat-resistant starch has the role of adjusting the distribution of the intestinal flora and maintaining the health of the body. Fagopyrum. Because of its cold, drought, and barren resistance, buckwheat is considered an important global grain crop [Buckwheat, a dicotyledonous plant, is a species within the genus ain crop . The conain crop ,3,4. Morain crop . Differeain crop ,7. Nicotain crop . Dietaryain crop .Additionally, abundant mineral elements are beneficial to the healthy function of buckwheat. Magnesium can inhibit atherosclerosis, hypertension, and hyperlipidemia by promoting fibrinolysis and suppressing thrombin formation . MagnesiBifidobacterium and Lactobacillus in the intestinal canal [Prebiotics containing functional oligosaccharides, polysaccharides, plant nature extractions, protein hydrolysate, and polyols refer to nondigestible food ingredients that have beneficial effects on human health . Prebiotal canal ,17. Furtal canal . Furtheral canal . Resistaal canal . Becauseal canal . The curCurrently, the most common approach for processing resistant starch is autoclaved debranched treatment as well as acid and enzyme hydrolysis treatment. Autoclaved debranched treatment gelatinizes starch molecules under high temperature and pressure, during which the released amylose can formulate a more solid crystalline structure . After aResistant starches have promising effects on intestinal health, while the difference in physicochemical characteristics among different resistant starch preparation technologies was seldom studied. Therefore, the aim of the present study was to analyze differences in the physicochemical characteristics of the starch, including the crystalline property, amylose content, and anti-digestibility after AEDT and AHT. We investigated the effect of resistant starch on intestinal physiological function, including intestinal tract movement and intestinal microbes. Our research suggests a processing method for improving the content of buckwheat-resistant starch and the application of buckwheat-resistant starch on intestinal health.Raw buckwheat-resistant starch with a content of 23.5% (RS), AHT-resistant starch with a content of 29% (RS1), and AEDT-resistant starch with contents of 31.7% (RS2), 35.6% (RS3), 39.7% (RS4) and 45.5% (RS5) were prepared following previously described standard methods and were used for the analysis of physicochemical characters.A stepping scanning method was used to analyze the X-ray diffraction molds of buckwheat-resistant starch samples. The start and termination angle at 2θ for scanning was 4° to 65.0°, respectively, with a scanning speed of 0.06°/s. The sampling time, divergence slit, scattering slit, tube current, and voltage for X-ray diffraction were 1 s, 1°, 1°, 20 mA, and 30 kV, respectively.Standard amylose with contents of 0, 10, 20, 25, 30, and 35% was used to construct a standard curve. One hundred mg of standard amyloses were put into 1 mL of absolute ethyl alcohol and 9 mL of NaOH. The mixture was then dispersed under boiling water for 10 min and adjusted to a final concentration of 1 mg/mL. Iodine was added into a standard amylose solution; the absorbance value was then measured under 720 nm using a spectrophotometer. The standard curve was generated by placing amylose content on the X-axis and the absorbance value on the Y-axis. The amylose content of RS1–RS5 was determined according to the standard curve.The anti-digestibility of RS1–RS5 was analyzed by an in vitro digestion model. Four hundred mg of resistant starch was added into sulfate buffer (pH 6.9) to a constant volume of 20 mL. α-Amylase was then added to the mixture and transferred to a dialysis tube. The dialysis tube was placed in the sulfate buffer (pH 6.9) at 37 °C. Digestive products were sampled at 0.5, 1, 1.5, 2, 3, 4, and 5 h. The sugar content was then determined by the phenol-sulfuric acid method . SamplesIn total, 70 healthy mice with a weight of 20–25 g were randomly divided into 7 groups: buckwheat starch group (BS); low dosage of buckwheat resistance starch group (0.03 g/(10 g·bw), LBS); middle dosage of buckwheat resistance starch group (0.06 g/(10 g·bw), MBS); high dosage of buckwheat resistance starch group (0.12 g/(10 g·bw), HBS); negative group (NG), positive group (0.7 mg/(10 g·bw) of defecation capsule, PG) and constipation group (0.05 mg/(10 g·bw), CG) of compound diphenoxylate tablets). All the treatment groups were gavaged once each day, and after 15 d, experiment mice from all the treatments were fasting for 16 h and then gavaged with carbon ink. The time the first black feces appeared during each treatment was recorded, and the number of black feces within 5 h of each treatment was also recorded.Six treatments containing BS, LBS, MBS, NG, and CG were used to analyze the effect of buckwheat resistance starch on mice defecation. All the treatments were gavaged for 15 d, and then all the mice were fasting for 16 h. All the treatments were then gavaged with (0.05 mg/(10 g·bw) compound diphenoxylate tablets. After 0.5 h gavaged with carbon ink, the intestinal canal of each treatment was isolated, and the intestinal length was measured. The ink intestinal propulsion rate was regarded as the influence of buckwheat resistance starch on mice defecation. The ink intestinal propulsion rate (%) = ink intestinal propulsion length/intestinal total length × 100 .In total, 125 mice were divided into five treatments containing BS, LBS, MBS, and NG to analyze buckwheat-resistant starch’s effect on mice gut microbes. All the treatments were gavaged for 14 d, 28 d, and 35 d, respectively. Microbes were isolated from the feces, colon, and caecum contents of mice from each treatment. All the isolated microbes were identified with 16S rRNA amplification primers.Buckwheat raw starch has obvious diffraction peaks with the diffraction angles at 15.10°, 17.14°, 17.92°, and 22.96°, which is characteristic of typical A-type crystals. AHT and AEDT significantly altered the X-diffraction molds of resistant starch. AHT resulted in strong diffraction peaks at 5.56°, 9.4°, 14.26°, 16.84°, and 19.86°, in which 5.56° is a typical type B crystal diffraction peak, and 14.26° and 19.86°crystal are typical type V crystal diffraction peaks. AEDT had highly similar diffraction peaks compared with AHT. AHT also exhibited a B + V type crystal structure, with the diffraction peaks at diffraction angles of 5.50°, 15.22°, 15.58°, 16.24°, 18.76°, and 19.84° .The amylose content in AHT (RS1) and AEDT (RS2–5) was significantly higher than that in raw buckwheat . CompareThe amount of starch digestion products was positively related to digestion time among all of the buckwheat-resistant starch . HoweverCompared with other groups, the time the first black feces appeared in the CG group was longer than in other treatments. The time the first black feces appeared in HBS, MBS, and LBS was shorter than in the BS group, which indicated that gavaged with high, middle, and low dosages of buckwheat-resistant starch could promote mice defecation.Compared with the BS treatment, the ink intestinal propulsion rate was higher in three buckwheat resistance starch treatment groups, especially in the high dosage of buckwheat resistance starch group, which indicated that buckwheat resistance starch could promote bowel intestinal tract movement.Citrobacter, Enterococcus, Bifidobacterium, and Lactobacillus, were isolated, identified, and selected for analysis.In total, four types of bacteria, including Citrobacter in the LBS, MBS, HBS, and BS groups was reduced in the colon and cecum tissues, as well as feces at 28 and 35 days compared with negative control (without gavaged of any types of starch). Compared with gavaged buckwheat starch, being gavaged with different dosages of buckwheat resistance starch could increase the quantity of Citrobacter in the colon and cecum tissues at 14, 28, and 35 days (The quantity of  35 days .Enterococcus. The quantity of Enterococcus was reduced after being gavaged with buckwheat starch and different dosages of buckwheat resistance starch at all time points. The quantity of Enterococcus was lower in the three buckwheat resistance starch groups than in the buckwheat starch group (Enterococcus.A similar phenomenon had been found in ch group , which mBifidobacterium and Lactobacillus, the quantity alteration was different with Enterococcus and Citrobacter. The quantity of Bifidobacterium and Lactobacillus in the buckwheat starch group and buckwheat resistance starch groups was higher than without gavaged of any types of starch. Compared with the buckwheat starch group, being gavaged with different dosages of buckwheat resistance starch could increase the quantity of Bifidobacterium and Lactobacillus (For bacillus .Resistant starch appears to play an important role in food applications and physiological functions. On the one hand, it is used to develop a variety of functional-resistant starch products in the food and pharmaceutical fields. The addition of resistant starch to flour can markedly improve the quality of noodles, including increasing their brightness and decreasing their digestibility and the flavor of bread ,31. On tMoreover, previous studies suggest that resistant starch may play important roles in physiological function and the management of body sugar and lipid metabolism. Resistant starch can regulate glucose and insulin levels, improving the balance between glucose and lipids in the body. The intake of food containing resistant starch could help control diabetes by reducing blood glucose and increasing insulin sensitivity and intestinal hormones . Hence, Bifidobacterium and Lactobacillus, but inhibit the growth of acidophilic bacteria, such as Escherichia coli, Clostridium perfringens and Bacteroides, which might stem from the structure of the resistant starch surface [As a prebiotic, resistant starch is vital to the regulation of gut microbes and intestinal health. Resistant starch can promote the growth and reproduction of beneficial intestinal microbes, such as  surface ,46. Barc surface . Resista surface . Lastly,Understanding the mechanism by which resistant starch promotes health is important for its future applications. The addition of resistant starch into an oral rehydration solution could significantly reduce fecal fluid losses and the duration of diarrhea with cholera . It coulThe formation of resistant starch was influenced by amylose; therefore, using buckwheat starch to prepare it could reduce the waste of resources and improve the utility and value of buckwheat. Hence, developing an effective method for improving the content of resistant starch is important. Previously, we used AHT and AEDT to prepare resistant starch; this study primarily analyzed the physicochemical characteristics. Finally, we confirmed that AEDT results in the highest resistant starch content and the most stable structure.Different preparation methods can affect the crystalline structure of resistant starch. In this study, the crystalline structure of raw buckwheat starch changed from A to B + V after AHT and autoclaving enzymatic debranching treatment. Similar observations have been made in waxy maize. For example, Shi found after pullulanase debranching that the crystalline structure mold of raw starch changed from A to B + V . Figure We also found that the content of resistant starch was positively related to amylose. This pattern might stem from the fact that the amylose molecules form a double helix structure based on intermolecular forces, which result in the generation of resistant starch. Guraya found that increasing amylose facilitated the linkage of amylose molecules and promoted the formation of resistant starch . HoweverCompared with buckwheat starch treatment, gavaged with buckwheat resistance starch could promote bowel intestinal tract movement, which might be due to the buckwheat resistance starch could ferment into short-chain fatty acids in the colon. These short-chain fatty acids could stimulate the intestinal wall, promote the intestinal absorption of water and cause intestinal movement, thereby shortening the retention time of the feces in the gut.Future work could focus on deeply investigating the application of the selected buckwheat-resistant starch on human health, such as regulating the metabolism of blood lipids, blood sugar, or cholesterol. Moreover, developing buckwheat-resistant starch products was also a concern.The physicochemical properties of buckwheat-resistant starch were altered after AHT and AEDT. The crystalline mold in buckwheat raw starch changed from A to B + V after AHT and AEDT. As the resistant starch content increased, the amylose content increased. In vitro digestion simulation experiments showed that the digestion rate and products were negatively related to the content of resistant starch. The buckwheat-resistant starch can promote bowel intestinal tract movement. The quantity of intestinal microbe was regulated by buckwheat-resistant starch. Our research suggests a novel approach for preparing resistant starch with high content and applying buckwheat-resistant starch to intestinal health.


Potatoes are among the leading staple crops due to nutritional value and high demand. The undersized and damaged potatoes are considered low grade and mainly dumped as a waste or used in animal feed. The study aimed to extract starch from low grade potatoes, its modification to improve the starch properties and formulation of gluten free cookies using modified starch (MS). The starch was extracted from low-grade potatoes of three varieties known as Asterix, Kruda and Mosaic, using the water steeping method. The native starch (NS) was modified using lintnerization and repetitive autoclaving. MS contains high amylose content which is associated with health benefits. NS and MS were characterized for amylose content, color attributes, granular morphology, water solubility index (WSI), water absorption index (WAI), thermogravimetric analysis (TGA) and Fourier transform infrared spectrometer (FTIR) analysis. Gluten-free cookies were formulated by adding potato NS and MS. The cookies were characterized by sensory evaluation, proximate and textural analysis. The starch yield extracted from three different varieties of potatoes i.e. Asterix, Kruda, Mosaic was 11.53%, 11.32% and 11.24%, respectively. The amylose content of potato starch was significantly (p < 0.05) increased for all varieties (33.61–37.74%) after modification of NS, which was in the range of 25.71–26.60% for different potato varieties. The granules of MS were observed as amorphous structures in comparison to NS granules with smooth surfaces. The addition of MS significantly (p < 0.05) decreased the hardness of the cookies in comparison to NS. Overall, no significant difference was observed in the sensory attributes of control, NS and MS containing cookies. Therefore, in comparison to other dietary fibers, MS can be used as a functional ingredient in food products without compromising the texture and sensory attributes. In terms of global production, the potato crop ranked third after wheat and rice. Food and Agriculture Organization (FAO) of the United Nation reported global production of potatoes over 370 million metric tons O bond) was changed after the structural modification of starch [−1 and 2929–2935 cm−1 correspond to O–H and C–H bonds, respectively. The intensity of these peaks was different for native and modified starches. The variations in FTIR spectra of native and modified starches confirmed the structural variations in starch as a result of modification treatment [FTIR spectra of native starches changed after the modification, which indicated structural variation among native and modified starches . Peaks bf starch . The peareatment .Fig. 2FT3.2.6The thermal decomposition of native and modified potato starches was determined by TGA . Initial3.2.7The proximate composition of gluten-free cookies fortified with modified starches is summarized in 3.2.8The addition of MS significantly (p < 0.05) decreased the hardness of the cookies in comparison to native starch . Cookies3.2.9Overall, there was no significant difference observed in the sensory attributes of control, native and MS containing cookies . However4Low-grade potatoes can be used to extract starch and develop a modified starch to use as dietary fiber in food products. The combination of acid hydrolysis and heat treatment significantly increase the amylose content of potato starch. The incorporation of modified starch in food products not only increases the functional value but also results in minimal changes in textural and sensory attributes. Therefore, modified starch can be used as a dietary fiber in food products to replace the conventional fibers, which significantly compromise texture and sensory attributes.Experiments were conducted according to established ethical guidelines, and informed consent obtained from the participants.Syed Mueez Ali: Performed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data.Yumna Siddique: Performed the experiments.Samina Mehnaz: Contributed reagents, materials, analysis tools or data.Muhammad Bilal Sadiq: Conceived and designed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data; Wrote the paperThis research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.Data will be made available on request.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


Qingke β-glucan (QBG) concentrations and molecular weights (MWs) on rice starch (RS). With the increasing concentrations and MWs, the pasting properties and gelatinization enthalpy of RS/QBG suspension decreasing was observed by using rheometer and differential thermal scanning analysis, respectively, which was consistent with the results of X-ray diffraction. In Infrared spectrum, QBG combined with leached amylose via hydrogen bonds, thus preventing the reaggregation of RS particles and inhibiting the short-term retrogradation of RS. The results of scanning electron microscopy and confocal laser scanning microscopy suggested that interaction between QBG and RS changed RS microstructure, reduced the leached amylose of the starch, and thus altered RS/QBG digestibility that the digestion rate of RS/QBG decreased with the incrementing QBG MWs at in vitro simulated experiments. These results provide further understanding and expand potential application to starch-based foods.This study aimed to investigate the impact of  Starch is a vital carbohydrate polymer in the food industry and has numerous applications, including health-promoting foods, food additives, cosmetics, and pharmaceuticals . Rice stNon-starch hydrocolloids are extensively used in starch-containing systems to improve the quality and stability of the final products because they can alter gelation and flow behavior . These hHordeum vulgare L. var. nudum Hook.), also called Qingke, is a kind of cultivated barley with naked grains. Qingke is the main staple food crop in Qinghai-Tibet Plateau. Nowadays, it is gaining popularity as “functional grain” because it contains various biologically active ingredients, including 4–11% Qingke β-glucan (QBG) (d-glucopyranose units connected by (1 → 3) and (1 → 4) glycosidic linkages  . Our prelinkages . QBG haslinkages , colon clinkages , and metlinkages . Moreovelinkages , enhancivitro digestion. The influence of β-glucan on the viscosity of oat flour by the content, molecular weight, and structural characteristics of β-glucan has been investigated -linked-d-glucopyranosyl, β-(1 → 4)-linked-d-glucopyranosyl. The high (HQBG), medium (MQBG), and low molecular weight (LQBG) β-glucans were 280 (±18), 190 (±11), and 155 (±12) KDa, respectively, and the macromolecular characteristics were characterized in our previous study . A double-enzymatic technique was used to extract β-glucan from ke flour , and acike flour . The QBGus study . GlucoseQBG solutions with different molecular weights  and concentrations  were prepared. Then RS  was added to the QBG solutions, the mixtures were swirled to create homogenous slurries. Then, the samples were split into groups named RS/5.0%HQBG, RS/5.0%MQBG, RS/5.0%LQBG, RS/2.5%HQBG, RS/2.5%MQBG, and RS/2.5 %LQBG.The pasting characteristics of the RS/QBG mixtures were determined using a Modular Compact 302 rheometer  equipped with an intelligent starch cell. The RS concentration was 5.0%, and the QBG was 2.5% and 5.0% of the RS dry weight, respectively. RS powders and RS/QBG mixtures in the starch cells were dispersed in 20 mL of distilled water. For measurement, the slurries were incubated at 28 °C for 2 min and then gradually heated from 28 °C to 95 °C at a heating rate of 5 °C/min. The samples were maintained at 95 °C for 30 min, cooled from 95 °C to 50 °C at a cooling rate of 5 °C/min, and held at 50 °C for 10 min. The stirring speed was set at 800 rpm for the initial 10 s, followed by 160 rpm to confirm the diffusion consistency. Pasting parameters, including peak viscosity (PV), trough viscosity (TV), final viscosity (FV), breakdown viscosity (BD), setback viscosity (SB), were recorded using pasting curves.The prepared RS and RS/QBG samples according to Section “RS and RS/QBG system preparation” were heated in a water bath at 95 °C for 20 min and then the RS and RS/QBG paste was cooled to room temperature for further analysis.Dynamic oscillatory measurements were performed using a previously described protocol with slight modifications . An MCR To), peak temperature (pT), end temperature (Tc), and enthalpy (ΔH) of gelatinization were determined using the software.The gelatinization properties of RS were analyzed by differential scanning calorimetry (DSC) . A 30.0% w/w (dry basis) mixture of RS and RS/QBG was selected by maintaining the QBG ratio at 2.5% and 5.0% RS. The thoroughly mixed solutions were initially hydrated for 24 h at room temperature. Subsequently, 10-15 mg of the solution was precisely weighed and loaded into an aluminum DSC pan, followed by immediate hermetical sealing to prevent moisture loss. An empty pan was used as a reference, and the sealed pan was heated from 20 to 100 °C at 10 °C/min. The onset temperature   fitted with Cu-K radiation (λ = 0.15406 nm). The samples were scanned within the 4° < 2θ < 50° range using a scanning speed of 5°/min and 10 s for each step, with a tube pressure and flow of 40 kV and 40 mA, respectively.−1 to 4000 cm−1.The samples used in this experiment were the same as those described in Section “RS and RS/QBG paste preparation”. After overnight storage at −80 °C, the samples were freeze-dried in a vacuum freeze-drier and passed through a 100-mesh sieve. The ratio of samples to KBr was approximately 1:120, and the samples were compressed into slices. A Nicolet IS50 Fourier transform infrared (FT-IR) spectrometer  was used to acquire FT-IR spectra. The test wavelengths ranged from 400 cmThe RS  and RS/QBG  mixtures used in this experiment were the same as those described in Section “RS and RS/QBG paste preparation”. The samples were stored at −80 °C overnight and freeze-dried in a vacuum freeze-drier. The granular starch morphology was observed using a Nova Nano SEM 230 field scanning electron microscope (SEM) (Czechia). The samples were gold-coated and fixed on conductive tape, operating at an accelerating voltage of 20 kV.The gel images were captured using a confocal laser scanning microscope (CLSM) . A previously described procedure with minor modifications was used . First, in vitro starch digestibility was measured using the Englyst method . Analysis of variance (ANOVA) was followed by the Turkey’s HSD test for pairwise multiple comparisons between treatments, and the significance level was set as The pasting curves and properties of RS and RS/QBG with different molecular weights and concentrations are shown in BD represents the variation between the PV and TV in the pasting curve, which is indicative of the degree of damage during gelatinization and the thermal stability of starch paste . After tSB is the difference between FV and TV in the gelatinization curve, and the starch paste's viscosity increases during the cooling process. As shown in The effects of QBG on the storage modulus (G') and loss modulus (G'') of RS gel are illustrated in The elastic modulus of the starch gel system containing QBG with different molecular weights and concentrations was lower than the pure RS gel system. This may be because QBG enhances the entanglement between β-glucan molecules and molecular starch chains in the gel system, increasing the network density of the gel system. As a result, the energy storage modulus [G'] and loss modulus [G''] were RS/LQBG > RS/MQBG > RS/HQBG. According to our previous study, QBG behaves in an aqueous solution as a pseudoplastic fluid with high viscosity, which may have caused a reduction in the elastic modulus and increased the viscosity of the gel system. However, the viscosity of all the RS/QBG samples was lower than RS alone. Possibly, the high viscosity of β-glucan inhibited the starch gelatinization. Furthermore, gelation is challenging because of the glucan chain's consecutive β-(1 → 4)-linkage blocks. This characteristic may influence the gelling capacity of the RS/QBG. Moreover, competition between QBG and water, then QBG wrapping around the starch granules may decrease RS's ability to gel, thus inhibiting the gelatinization and digestibility of rice starch. Amylose is the primary polymer used to create cross-linked networks. Therefore, we hypothesized that QBG was connected to leached amylose, which could slow the reaction between starch amylose molecules, subsequently delaying amylose re-aggregation and reducing the elasticity and viscosity of the starch paste.The XRD patterns of RS and RS/QBG gels are shown in Qingke β-glucan molecules on the binding of starch molecules, which consequently reduces the amylose content in the system and diminishes the interaction between starch molecules. The addition of β-glucan weakened the recrystallization of the starch molecular chains, consistent with the decrease in the gelatinization enthalpy (ΔH) of RS/QBG.The natural A-type RS pattern was lost during the pasting test, demonstrating that the RS in the pasta was entirely gelatinized. RS/QBG showed a weak absorption peak at near 20° and the diffraction was reduced and became wide compared with RS. This may be attributed to the inhibitory effect of −1 (stretching vibration of the hydrogen bonding –OH groups)  . The cryR1047/1022 and R995/1022 values of RS/QBG decreased  , RS/5.0 %MQG (7.1 J/g), RS/5.0 %LQBG (7.3 J/g), RS/2.5 %HQBG (7.7 J/g), RS/2.5 %MQBG (7.8 J/g) and RS/2.5 %LQBG (7.9 J/g) were lower than RS (8.1 J/g). The amount of heat required was reduced owing to the preservation of RS crystallization by β-glucan. The enthalpy (ΔH) order was RS/LQBG > RS/MQBG > RS/HQBG and RS/2.5 %QBG > RS/5.0 %QBG, suggesting that the tightness of starch granules wrapped by β-glucan could be increased by increasing the molecular weight and concentration of QBG. QBG may have promoted excellent bonding with the starch granules owing to its higher viscosity, especially at higher concentrations and molecular weights. This enhanced bonding likely improves QBG's ability to protect the starch crystallization zone. Moreover, the enhanced hydration capacity of QBG inhibits the movement of water molecules and prevents them from penetrating the starch granules  . Accordigranules . TherefoThe RS samples exhibited thicker and denser pore walls and considerably smaller pore sizes than the RS/QBG samples. The internal structure of RS/QBG was loose, less dense, and more brittle, with more cracks and disordered fragments as the molecular weight of QBG increased. Compared with the RS/MQBG and RS/LQBG gels, the pores in the RS/HQBG gels were uneven and large. These results show that RS/QBG with larger molecular weights has better van der Waals forces. Therefore, larger slices were observed for the RS/HQBG gel than for the RS/MQBG and RS/LQBG gels. It is possible that QBG absorbed water and expanded before starch gelation, thereby competing with the starch for water. A large amount of QBG was wrapped in starch, which damaged the stability of the starch gel network. This outcome is consistent with the findings of a previous study in which the addition of hydrophilic colloids inhibited water absorption expansion and the exudation of amylose from RS granules .Previous studies have explained how starch molecules in grains are interconnected via hydrogen bonds . In the The inhibition of the RS starch digestion rate was in the order of control < 2.5 %QBG < 5.0 %QBG. QBG affects the overall viscosity of the system, and an increase in QBG concentration can increase the viscosity of the entire system, thereby preventing starch form absorbing water H. J. . AnotherAt a constant concentration, the order of starch digestion was as follows: LQBG > MQBG > HQBG. It was found that incorporating QBG of higher molecular weight in mixture significantly increased the viscosity. This increase in viscosity may have reduced the water available for starch gelatinization, ultimately decreasing the susceptibility of the enzymes to digestion . MoreoveIn addition, the wrapping of RS granules by QBG made some starch granules difficult to digest during the pasting process. Moreover, QBG can compete with rice starch for water molecules, inhibiting the expansion of rice starch particles, and leading to incomplete gelatinization of some rice starch particles, thus reducing the digestion rate of rice starch that was observed in the pasting properties , rheologΔH) decreased with QBG addition. Compared with the original RS, the R1047/1022 values of RS/QBG reduced from 1.1138 to 1.0153 and R995/1022 values of RS/QBG reduced from 1.1743 to 1.0539, respectively, indicating that the addition of β-glucan could inhibit the formation of the short-range ordered structure of starch and delay aging. XRD analysis showed the peak intensity did not vary significantly after the increase in the molecular weight of QBG. SEM and CLSM analyses revealed that the leaching out of the starch components was inhibited by QBG in RS/QBG. In addition, Qingke β-glucan increased the SDS and RS contents, decreased the RDS content, and decreased the digestion rate of the RS/QBG mixture system. The content of slowly digestible starch and resistant starch of the RS/QBG with the 5.0% HQBG addition were 7.8% and 12.7% respectively, which were 1.5 times and 2.54 times of the RS, respectively. These results may provide a theoretical basis for developing functional foods based on a starch/non-starch polysaccharide blending system.This study demonstrated the effect of QBG with different molecular weights and concentrations on the pasting, rheological, structural, and digestion properties of RS/QBG gels. The peak, final, and setback viscosity values decreased when the β-glucan was added. This may be attributable to QBG wrapping around the starch granules, thereby preventing their gelatinization and expansion. According to rheological analysis, QBG reduced the dynamic modulus of the RS/QBG gels. The blended system's gelatinization enthalpy (Lan Zhao: Methodology, Visualization, Investigation, Writing – original draft. Xinyan Jin: Investigation. Jia Wu: Writing – review & editing. Huibin Chen: Supervision, Conceptualization, Writing – review & editing.The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Huibin Chen reports financial support was provided by Fujian Normal University.