Effects of silane hydrolysis time on the physicochemical properties of bioplastics based on starch and epoxidized soybean oil.

This work investigated the effects of 3-aminopropyl triethoxy silane (APTES) hydrolysis time on the physicochemical properties of the resulting starch/epoxidized soybean oil (ESO) bioplastics comprehensively. FTIR analysis confirmed that APTES hydrolyzed for 4 h had the best modification effect on starch. The results of XRD and TGA demonstrated the successful silylation of starch by APTES despite hydrolysis time. Silylation treatment reduced the thermal stability of starch slightly, but enhanced the thermal stability of the resultant bioplastics, revealing better interaction between silylated starch and ESO. The interfacial adhesion of starch and ESO in the bioplastics was obviously enhanced when APTES was hydrolyzed for 2-24 h. The bioplastics with APTES hydrolyzed for 2-4 h showed more desirable tensile properties as the silane hydrolysis was complete and self-condensation of hydrolyzed silanes was avoided. The bioplastics containing silylated starch still showed superior opacity and biodegradability.

Effect of sorghum bran addition on antioxidant activities, sensory properties, and in vitro starch digestibility of Chinese southern-style steamed bread.

BACKGROUND: Chinese steamed bread (CSB) is one of the most important staple foods in China and is also popular in South-East Asia. Developing functional CSB could improve people's resistance to inflammatory and non inflammatory diseases. This work investigated the effect of sorghum bran addition on antioxidant activities, sensory properties, and in vitro starch digestibility of Chinese southern-style steamed bread (CSSB). RESULTS: In this study, the enhanced CSSB with 0-200 g kg-1 of fine black and tannin (sumac) sorghum bran addition was developed. A small change in phenol content and antioxidant activity was observed at various stages in the processing procedure before steaming. Moreover, a high retention of antioxidant phenolics CSSB with sorghum bran addition was observed. Sorghum bran addition significantly increased the total phenol content and antioxidant activity of CSSB by 4.5-10 times, on average, relative to control. Sorghum bran addition significantly also increased the content of resistant starch, and significantly decreased in vitro starch digestibility in CSSB; these effects were likely due to the joint inhibitory effect of tannins and ferulic acid on starch digestibility. Interestingly, the sorghum bran breads scored higher or similar to control in sensory color preference and overall appearance, but lower on most textural and mouthfeel attributes. CONCLUSION: Sorghum bran addition significantly increased the antioxidant activity of CSSB and significantly decreased starch digestibility. Moreover, the color and appearance properties were maintained or improved. However, the sensorial textural attributes were negatively impacted by the sorghum bran substitutions. Strategies to improve the texture of bran-fortified breads would likely enhance their consumer acceptability. © 2024 Society of Chemical Industry.

Development of a Zn-Based Single-Atom Nanozyme for Efficient Hydrolysis of Glycosidic Bonds.

Hydrolytic enzymes are essential components in second-generation biofuel technology and food fermentation processes. Nanozymes show promise for large-scale industrial applications as replacements for natural enzymes due to their distinct advantages. However, there remains a research gap concerning glycosidase nanozymes. In this study, a Zn-based single-atom nanozyme (ZnN4-900) is developed for efficient glycosidic bond hydrolysis in an aqueous solution. The planar structure of the class-porphyrin N4 material approximatively mimicked the catalytic centers of natural enzymes, facilitating oxidase-like (OXD-like) activity and promoting glycosidic bond cleavage. Theoretical calculations show that the Zn site can act as Lewis acids, attacking the C─O bond in glycosidic bonds. Additionally, ZnN4-900 has the ability to degrade starch and produce reducing sugars that increased yeast cell biomass by 32.86% and ethanol production by 14.56%. This catalyst held promising potential for enhancing processes in ethanol brewing and starch degradation industries.

Facile preparation of interpenetrating network hydrogel adsorbent from starch- chitosan for effective removal of methylene blue in water.

Hydrogels based on biopolymers have attracted considerable interest in the last decades. Herein, an interpenetrating network hydrogel (IPN-Gel) adsorbent from starch-chitosan was fabricated facilely in one-pot through tandem Schiff base reaction and photopolymerization. First, aldehyde starch (DAS) was synthesized by the reaction of soluble starch with sodium periodate. Afterward, acrylamide (AM), 2-acrylamido-2-methylpropanesulfonic acid (AMPS), polyethylene glycol dimethacrylate (PEGDMA), photoinitiator, chitosan and DAS were dissolved in water to obtain a clear solution. Schiff base reaction between chitosan and DAS took place quickly to form the first network, and then photopolymerization of AM, AMPS, and PEGDMA occurred under ultraviolet radiation to form the second network. The preparation conditions of the as-prepared IPN-Gel were optimized with two indexes of gel mass fraction and swelling ratio. Its swelling behavior with pH and temperature change was explored. Finally, its adsorption performance was characterized with methylene blue (MB) as a model contaminant. The maximum adsorption capacity of IPN-Gel can reach 2039 mg·g-1 at pH =10. Its adsorption performance accords with Langmuir isothermal model and pseudo-second-order kinetic model and it was mainly controlled by chemisorption. This strategy is expected to found broad application prospects in the preparation of hydrogel adsorbents.

Design and decoration of copper nanoparticles into lignosulfonate-starch bionanocomposite: Characterization and evaluation of its therapeutic properties on burn wound.

In this report, eco-friendly synthesis for the production of copper nanoparticles by employing the sodium lignosulfonate (NaLS) mixed starch composite (NaLS-Starch/Cu NPs). NaLS-Starch mixed hydrogel has notable reducing and stabilizing potential for preparation of Cu nanoparticles. Characterization of NaLS-Starch/Cu NPs bionanocomposite was subjected to analysis of spectroscopic and microscopic techniques, including FE-SEM, TEM, EDS-elemental mapping, particle size distribution, XRD and ICP. TEM images displayed the spherical structured NaLS-Starch/Cu NPs, averaging 5-10 nm size. NaLS-Starch/Cu NPs were applied to cure the induced burn wounds in 60 Wistar rats. A group was considered as control group. The animals were treated with basal, tetracycline 3 % and NaLS-Starch/Cu NPs 3 % for 30 days and the treatment efficacy was determined according to the burn wound area reduction and molecular and histological characteristics. Taken together, these results support therapeutic use of NaLS-Starch/Cu NPs as potent ointment that may be proposed for burn wound healing. NaLS-Starch/Cu NPs ointment increased the levels of platelet-derived growth factors (PDGF) and fibroblast growth factor (bFGF). The mean wound surface, in all groups treated by NaLS-Starch/Cu NPs was larger than control group.

Preparation of multi-modified/carbonized/gelatinized starch and its de-risking effect on Cd(II) and hymexazol in wastewater.

In this study, starch (S) was gelatinized and carbonized to prepare carbonized/gelatinized S (CGS) as the research material. Then, peat extract (Pe) and surfactants with different ratios were single- and multi-modified on CGS, respectively, to prepare Pe-modified CGS (Pe-CGS) and multi-modified CGS, respectively. The microscopic morphology of multi-modified CGS was studied using various testing methods. The de-risking effect on Cd(II) and hymexazol in wastewater was investigated, and the effects of temperature, pH, and ionic strength were compared. The spheroidal structure of S was destroyed after carbonization, and Pe and surfactants were modified on the surface and changed the surface properties of CGS. The adsorption processes of Cd(II) and hymexazol were suitable to be described by the Langmuir and Freundlich models, respectively. The maximum adsorption capacities (qm) of Cd(II) and adsorption capacity parameter (k) of hymexazol on different modified CGSs presented the peak value at BS/Pe-CGS. With the increase in the modification ratio of Pe, BS, and SDS, qm and k increased, which showed a high value at 100 % modification. Increases in temperature and pH were beneficial to Cd(II) adsorption but were not conducive to hymexazol adsorption. The adsorption amount decreased for Cd(II) and increased first and then reduced for hymexazol with the rise in ionic strength. The adsorption process exhibited spontaneity, endothermic behavior for Cd(II), exothermic behavior for hymexazol, and an entropy-increasing reaction. The adsorption amount of Cd(II) and hymexazol by multi-modified CGS maintained approximately 81 % of the original sample after three rounds of regeneration.

Effect of different chain-length fatty acids on the retrogradation properties of rice starch.

In order to delay the retrogradation of rice starch, the effects of three different chain length fatty acids (lauric acid, myristic acid and palmitic acid) on rice starch were studied. The fatty acids with longer carbon chains had strong steric hindrance and hydrophobicity, which formed a more compact structure in the helical cavity of amylose, and significantly reduced degree of expansion, migration of water, short-range ordered structure, number of double helical structures and crystallinity. These structural changes endowed the rice starch-long chain fatty acid complexes with better gel viscosity, liquid fluidity and thermal stability than in the rice starch-short chain fatty acid complexes. The results showed that fatty acids with longer chain length inhibited the retrogradation of rice starch, most obviously when 5% palmitic acid was added. This study provides an important reference for the processing of rice starch-based foods.

Functional properties of starch cultivars of two Andean grains grown in Bolivia: Amaranth (Amaranthus caudatus) and canihua (Chenopodium pallidicaule).

The functional properties of Andean grain starches of two species, amaranth (Amaranthus caudatus) and canihua (Chenopodium pallidicaule), three cultivars each, were studied. The study focused on chemical composition, pasting properties, thermal properties, water solubility index (WSI), swelling power (SP), and granule morphology. All amaranth starches were waxy starches, with amylose content less than 5 %, which had some differences in chemical composition (p < 0.05). The pasting properties differed between the species, canihua showed more resistance, than amaranth, to heat and shear stress (higher cool paste (CPV) and lower breakdown (BD), ranged between 1250 and 1600 cP and -30 - 10 cP respectively. The amaranth starches presented only similar CPV with 800-1000 cP, while canihua cultivars presented similar PT and BD, and both species presented similar PV, around 1000 cP. Thermal properties (To, Tp, Tc, ΔH, and ΔT) differed among cultivars and species. These differences could be related to the homogeneity molecular structure and content of amylose in canihua cultivars and possibly to genotype factor. Polygonal shapes were the predominant shape of starch granules, ranged 1.0-1.4 µm and 0.8-0.9 µm, for amaranth and canihua starches respectively. Amaranth starches swelled quickly to disintegrate partially at the end, contrary to canihua starches. The thermal and pasting properties were correlated between them. SB, CPV, HPV, CS, were correlated to the content of amylose in canihua starches. One amaranth cultivar was significantly different from the others. Thus, according the functional properties differenced both species and some cultivars in each species. Additionally, the amaranth starch has the potential to be used in the food industry where heat and stress are applied such as extrusion, while canihua starches can be used in desserts or in cosmetic uses, based on their functional properties.

The solid state and nanostructure of starch: Effects on starch functionality.

In order to determine suitable end use applications for different starches, this review characterizes and differentiates the physical components, solid state, crystalline structures, and their effects on gelatinization, retrogradation, texture and functionality. There exist four crystalline packings of starch. A-, B- and C-type packings are attributed to amylopectin, and V-type which is attributed to amylose. B- and C- type crystallinity rely on water to help coordinate their crystal structures due to the congregation of water in the large intrahelical cavity of the B-type packings. The ratio of amylose to amylopectin content largely affects the textural and functional properties of starch. Amylose largely influences retrogradation, and thus can largely impact the crystallinity, strength, cohesion and brittleness of starch gel systems. Amylose has been found to crystallize prior to amylopectin, suggesting that amylose acts as a nucleation site for further radial crystallization of amylopectin. Processing treatments such as size reduction and drying, which are typically applied to all commercial starches, also impact the physiochemical and functional characteristics of the starch. These processes can cause damage to the starch granule while reducing crystallinity in the native starch, but also increasing retrogradation in gelatinized systems.

High-efficiency conversion of corn bran to ethanol at 150 L scale.

Fractionated corn bran was processed to maximize ethanol production from starch, cellulose, and xylan. After various bench-scale experiments, an optimized process with dilute acid pretreatment (1.5 % w/w H2SO4) at 90 °C for 60 min was utilized followed by enzymatic hydrolysis using cellulase and hemicellulase for 48 hr. After simultaneous saccharification (regarding starch) and fermentation at 150 L using an engineered yeast, which consumes both glucose and xylose to make ethanol, the 86 % total sugar conversion yield was achieved, including conversions of 95 % for starch, 77 % for cellulose and 77 % for xylan. Also, an accurate mass balance was formulated for ethanol-producing carbohydrates including starch, cellulose, and xylan from feedstock to final ethanol. A highly efficient process of converting corn fiber to ethanol was successfully scaled up to 150 L.

Combined effects of nitrogen limitation and overexpression of malic enzyme gene on lipid accumulation in Dunaliella parva.

Overexpression of Dunaliella parva (D. parva) malic enzyme (ME) gene (DpME) significantly increased DpME expression and ME enzyme activity in transgenic D. parva. Nitrogen limitation had an inhibitory effect on protein content, and DpME overexpression could improve protein content. Nitrogen limitation increased carbohydrate content, and Dunaliella parva overexpressing malic enzyme gene under nitrogen limitation (DpME-N-) group showed the lowest starch content among all groups. Dunaliella parva overexpressing malic enzyme gene under nitrogen sufficient condition (DpME) and DpME-N- groups showed considerably high mRNA levels of DpME. ME activity was significantly enhanced by DpME overexpression, and nitrogen limitation caused a smaller increase. DpME overexpression and nitrogen limitation obviously enhanced lipid accumulation, and DpME overexpression had more obvious effect. Compared with control (wild type), lipid content (68.97%) obviously increased in DpME-N- group. This study indicated that the combination of DpME overexpression and nitrogen limitation was favorable to the production of microalgae biodiesel.

Novel Botrytis cinerea Zn(II)2Cys6 Transcription Factor BcFtg1 Enhances the Virulence of the Gray Mold Fungus by Promoting Organic Acid Secretion and Carbon Source Utilization.

The Zn(II)2Cys6 zinc cluster protein family comprises a subclass of zinc-finger proteins that serve as transcriptional regulators involved in a diverse array of fugal biological processes. However, the roles and mechanisms of the Zn(II)2Cys6 transcription factors in mediating Botrytis cinerea, a necrotrophic fungus that causes gray mold in over 1000 plant species, development and virulence remain obscure. Here, we demonstrate that a novel B. cinerea pathogenicity-associated factor BcFTG1 (fungal transcription factor containing the GAL4 domain), identified from a virulence-attenuated mutant M20162 from a B. cinerea T-DNA insertion mutant library, plays an important role in oxalic acid (OA) secretion, carbon source absorption and cell wall integrity. Loss of BcFTG1 compromises the ability of the pathogen to secrete OA, absorb carbon sources, maintain cell wall integrity, and promote virulence. Our findings provide novel insights into fungal factors mediating the pathogenesis of the gray mold fungus via regulation of OA secretion, carbon source utilization and cell wall integrity.

Comprehensive lipidomic insights of differentially accumulating lipids in large pigmented rice sprout collection and the changes in the starch composition upon germination.

Germination represents a vital bioprocess characterized by numerous biochemical transformations that significantly influence the nutritional characteristics of rice. The mobilization of starch and lipids during germination plays a pivotal role in altering the dietary profile of rice, thus potentially addressing the nutritional requirements of populations heavily reliant on rice as a staple food. To explore this potential, a comprehensive analysis encompassing lipidomics and starch composition was conducted on a diverse collection of pigmented rice sprouts. High-resolution mass spectrometry unveiled substantial shifts in the lipidome of pigmented rice sprouts, showcasing a notable enrichment in carotenoids and unsaturated triglycerides, with potential human health benefits. Notably, purple rice sprouts exhibited heightened levels of alpha- and beta-carotene. Analysis of starch composition revealed slight changes in amylose and amylopectin content; however, a consistent increase in digestible carbohydrates was observed across all rice varieties. Germination also led to a reduction in resistant starch content, with purple rice sprouts demonstrating a pronounced two-fold decrease (p < 0.05). These changes were corroborated by a 1.33% decrease in gelatinization enthalpy and a 0.40% reduction in the melting of the amylose-lipid complex. Furthermore, pasting property analysis indicated a substantial 42% decrease in the complexation index post-germination. We posit that the insights garnered from this study hold significant promise for the development of novel products enriched with health-promoting lipids and characterized by unique flour properties.

Effect of ultrasonically stimulated potato germination during soaking on the physicochemical properties of starch and its use in edible films.

The aim of this work was to investigate the effects of ultrasonic treatment during soaking of potatoes on the physicochemical properties of starches obtained after 16 weeks of germination. The ultrasonic treatment showed a direct correlation between sprout length and ultrasonic time. The protein content decreased from 0.63 to 0.38 % and the fat content decreased significantly from 0.31 to 0.01 % after germination. The amylose content changed depending on the ultrasonic treatment, and increased from 36.27 to 40.92 % after 16 weeks of germination, which was related to the amylopectin debranching and the duration of the ultrasonic treatment. X-ray diffraction showed that the nanocrystals with hexagonal structure were not affected by the germination and the duration of ultrasonic treatment. Scanning electron microscopy showed that the surface of the starch granules was not affected by the enzymatic treatment. The sprouted potato starch resulted in films with better tensile strength and lower water vapor permeability (WVP) compared to the native potato starch films. In addition, the films produced with ultrasound stimulated potato starch exhibited better properties (high strength and low permeability), which is desirable when it comes to controlling moisture exchange between a food product and the surrounding atmosphere.

Effects of biodegradation of starch-nanocellulose films incorporated with black tea extract on soil quality.

This study aimed to investigate the biodegradation behaviour of starch/nanocellulose/black tea extract (SNBTE) films in a 30-day soil burial test. The SNBTE films were prepared by mixing commercial starch, nanocellulose (2, 4, and 6%), and an aqueous solution of black tea extract by a simple mixing and casting process. The chemical and morphological properties of the SNBTE films before and after biodegradation were characterized using the following analytical techniques such as field emission scanning electron microscopy (FESEM), energy-dispersive X-ray spectroscopy (EDX), and fourier transform infrared (FTIR). The changes in soil composition, namely pH, electrical conductivity (EC), moisture content, water holding capacity (WHC), soil respiration, total nitrogen, weight mean diameter (MDW), and geometric mean diameter (GMD), as a result of the biodegradation process, were also estimated. The results showed that the films exhibited considerable biodegradability (35-67%) within 30 days while increasing soil nutrients. The addition of black tea extract reduced the biodegradation rate due to its polyphenol content, which likely resulted in a reduction in microbial activity. The addition of nanocellulose (2-6% weight of starch) increased the tensile strength, but decreased the elongation at break of the films. These results suggest that starch nanocellulose and SNBTE films are not only biodegradable under soil conditions but also positively contribute to soil health, highlighting their potential as an environmentally friendly alternative to traditional plastic films in the packaging industry.

Dietary jack bean (Canavalia ensiformis L.) supplementation enhanced intestinal health by modulating intestinal integrity and immune responses of broiler chickens.

This study investigated the influence of supplementing with jack beans on jejunal morphology, cecal short-chain fatty acids production, gene expression both of pro- and anti-inflammatory cytokines and tight junctions. Four treatment groups including 288 Indian River chicks that were one day old were randomized at random. While the treatment groups received jack bean supplementation at levels of 5 %, 10 %, and 15 %, the control group (0 %) was given a basal diet. For 11-35 days, each treatment consisted of 8 pens with 9 birds each. Supplementing with jack beans significantly enhanced butyrate production (P < 0.001), while at 10 % supplementation did not differ from control. Villus height (VH) and the ratio (VH:CD) were significantly (P < 0.001) increased by dietary treatments, while villus width (VW) and crypt depth (CD) were significantly (P < 0.05) decreased. TLR-3, TNF-a, and IL-6 were all significantly (P < 0.001) increased by dietary supplementation. However, at 15 %, TLR-3 and IL-6 were same with control. IL-18 was significantly (P < 0.05) decreased at 15 %. IL-10 decreased significantly (P < 0.001), but at 10 % same with control. At 5 and 10 %, IL-13 increased significantly (P < 0.001), whereas dietary treatments decreased at 15 % compared to control. Although ZO1 decreased significantly (P < 0.001) and OLCN increased significantly (P < 0.001), both ZO1 and OCLN were not significantly different from the control at 15 %. Dietary treatments significantly (P < 0.001) increased CLDN1 but did not differ from the control at 10 %. JAM2 decreased significantly (P < 0.001) with dietary treatments. In conclusion, jack bean supplementation may increase broiler chicken performance and intestinal health due to butyrate production. It may affect intestinal morphology and integrity by upregulating a tight junction protein gene. Jack beans also impacted jejunum immune responses and inflammatory cytokine gene expression.

Important roles of Ruminococcaceae in the human intestine for resistant starch utilization.

Intricate ecosystem of the human gut microbiome is affected by various environmental factors, genetic makeup of the individual, and diet. Specifically, resistant starch (RS) is indigestible in the small intestine but nourishes the gut microbiota in the colon. Degradation of RS in the gut begins with primary degraders, such as Bifidobacterium adolescentis and Ruminococcus bromii. Recently, new RS degraders, such as Ruminococcoides bili, have been reported. These microorganisms play crucial roles in the transformation of RS into short-chain fatty acids (SCFAs), such as acetate, propionate, and butyrate. SCFAs are necessary to maintain optimal intestinal health, regulate inflammation, and protect against various illnesses. This review discusses the effects of RS on gut and highlights its complex interactions with gut flora, especially the Ruminococcaceae family.

Superabsorbent starch protective layer modulates zinc anode interface for long-life aqueous zinc ion batteries.

Aqueous zinc ion batteries (AZIBs) have attracted much attention for their safety, low cost and high theoretical capacity. Nevertheless, Zn dendrites and the adverse reactions such as corrosion, hydrogen evolution and passivation on the anode affect the cycle life and stability of AZIBs. Herein, superabsorbent starch (SS) was employed on Zn foil to form an artificial interface protection layer, which inhibited the formation of dendrites by guiding the uniform deposition of Zn2+. SS with a large amount of oxygen-containing functional group is superabsorbent, which can attract the active water around the hydrated Zn2+, promoting the desolvation process of the hydrated Zn2+ and significantly inhibiting the occurrence of hydrogen evolution reaction. In addition, the inherent pore structure of the SS artificial interfacial layer can induce uniform nucleation of Zn2+ and inhibit the dendrites growth. Moreover, compared to bare Zn//MnO2 cell (44.1 %), the capacity retention of Zn@SS//MnO2 cell remained as high as 87.8 % after 1000 cycles at 1.5 A g-1. The simple method provided a new method for the rapid development of AZIBs.

In situ-gelling hydrophobized starch nanoparticle-based nanoparticle network hydrogels for the effective delivery of intranasal olanzapine to treat brain disorders.

Intranasal (IN) delivery offers potential to deliver antipsychotic drugs with improved efficacy to the brain. However, the solubilization of such drugs and the frequency of required re-application both represent challenges to its practical implementation in treating various mental illnesses including schizophrenia. Herein, we report a sprayable nanoparticle network hydrogel (NNH) consisting of hydrophobically-modified starch nanoparticles (SNPs) and mucoadhesive chitosan oligosaccharide lactate (COL) that can gel in situ within the nasal cavity and release ultra-small penetrative SNPs over time. Hydrophobization of the SNPs enables enhanced uptake and prolonged release of poorly water soluble drugs such as olanzapine from the NNH depot through mucous and ultimately into the brain via the nose-to-brain (N2B) pathway. The hydrogel shows high in vitro cytocompatibility in mouse striatal neuron and human primary nasal cell lines and in vivo efficacy in an amphetamine-induced pre-clinical rat schizophrenia model, with IN-delivered NNH hydrogels maintaining successful attenuation of locomotor activity for up to 4 h while all other tested treatments (drug-only IN or conventional intraperitoneal delivery) failed to attenuate at any time point past 0.5 h. As such, in situ-gelling NNHs represent a safe excipient for the IN delivery of hydrophobic drugs directly to the brain using customized SNPs that exhibit high penetration and drug complexing properties to maximize effective drug delivery.

Heterologous expression, characterization, and application of recombinant thermostable α-amylase from Geobacillus sp. DS3 for porous starch production.

Novel Geobacillus sp. DS3, isolated from the Sikidang Crater in Dieng, exhibits promising characteristics for industrial applications, particularly in thermostable α-amylase production. Recombinant technology was used to express thermostable α-amylase in E. coli BL21(DE3) to overcome high-temperature production challenges. The study aimed to express, purify, characterize, and explore potential applications of this novel enzyme. The enzyme was successfully expressed in E. coli BL21(DE3) at 18 °C for 20 h with 0.5 mM IPTG induction. Purification with Ni-NTA column yielded 69.23 % from the initial crude enzyme, with a 3.6-fold increase in specific activity. The enzyme has a molecular weight of ±70 kDa (±58 kDa enzyme+11 kDa SUMO protein). It exhibited activity over a wide temperature range (30-90 °C) and pH range (6-8), with optimal activity at 70 °C and pH 6 with great stability at 60 °C. Kinetic analysis revealed Km and Vmax values of 324.03 mg/ml and 36.5 U/mg, respectively, with dextrin as the preferred substrate without cofactor addition. As a metalloenzyme, it showed the best activity in the presence of Ca2+. The enzyme was used for porous starch production and successfully immobilized with chitosan, exhibiting improved thermal stability. After the fourth reuse, the immobilized enzyme maintained 62 % activity compared to the initial immobilization.