α‐Amylase immobilization on functionalized nano CaCO3 by covalent attachment

In this study, α‐amylase was immobilized on glutaraldehyde activated silanized calcium carbonate nanoparticles by a using covalent binding method. The surface modified nano calcium carbonate (CaCO₃) were characterized using FTIR and SEM. Immobilization yield was found as 199.43 mg/g of calcium carbonate nanoparticles. The maximum activity was observed at pH 6.5. The immobilized enzyme had a higher activity at elevated temperature (50–90°C) than the free one. Reuse studies demonstrated that the immobilized enzyme could reuse 25 times while retaining 18.2% of its activity. Free enzyme lost its activity completely within 15 days. V_max values for the free and immobilized enzymes were calculated as 10 and 0.35 mg/mL/min, respectively.     

A more Efficient Starch Degradation by the Combination of Hydrolase and Transferase Activities of α‐Amylase and Cyclomaltodextrin Glucanotransferase

α‐Amylases catalyze the hydrolysis of internal α‐(1→4) linkages of glucose polymers as their main reaction; however, some α‐amylases catalyze transfer reactions in addition to hydrolysis. It has been observed that those α‐amylases capable of transferring glycoside residues are also those that generate low molecular weight products from their action on starch (i.e. saccharifying α‐amylases). In this paper the product profiles of a liquefying α‐amylase, a cyclomaltodextrin glucanotransferase and both enzymes acting together on starch and maltodextrins are compared. The increase in glucose and maltose concentration, when both enzymes act together, is due to the combined action of the transfer and hydrolytic activity of CGTase and the liquefying α‐amylase, respectively.     

Visual Observation of Hydrolyzed Potato Starch Granules by α‐Amylase with Confocal Laser Scanning Microscopy

Porous starch was produced by digestion of freeze‐dried potato starch with α‐amylase from Bacillus sp. The surface structure of the granules became perforated and in the interior of the granules a capsule‐like cavity was formed, i.e. the hydrolyzed starch can be used as an encapsulant. The structure change of the granules was observed with confocal laser scanning microscopy and scanning electron microscopy. The degree of starch hydrolysis could be correlated with the Avrami equation. The activation energy of starch hydrolysis by α‐amylase was 83 kJ/mol.     

Purification and physicochemical characterization of ovine β‐lactoglobulin and α‐lactalbumin

Ovine whey proteins were fractionated and studied by using different analytical techniques. Anion‐exchange chromatography and reversed‐phase high‐performance liquid chromatography (HPLC) showed the presence of two fractions of β‐lactoglobulin but only one of α‐lactalbumin. Gel permeation and sodium dodecyl sulfate (SDS)‐polyacrylamide gel electrophoresis allowed the calculation of the apparent molecular mass of each component, while HPLC coupled to electrospray ionisation‐mass spectrometry (ESI‐MS) technique, giving the exact molecular masses, demonstrated the presence of two variants A and B of ovine β‐lactoglobulin. Amino acid compositions of the two variants of β‐lactoglobulin differed only in their His and Tyr contents. Circular dichroism spectroscopy profiles showed pH conformation changes of each component. The thermograms of the different whey protein components showed a higher heat resistance of β‐lactoglobulin A compared to β‐lactoglobulin B at pH 2, and indicated high instability of ovine α‐lactalbumin at this pH.     

Characterization of an acidophilic and thermostable α‐amylase from Alicyclobacillus sendaiensis NUST

This paper describes the characterization of an acidophilic and thermostable α‐amylase from Alicyclobacillus sendaiensis NUST. The MW of this enzyme was estimated to be 56 kDa by SDS–PAGE. The enzyme was stable over a range of pH from 2.5 to 5.5 with an optimum around 3.5. Maximum activity of the α‐amylase was observed at pH 3.5 and 85°C in the presence of soluble starch as substrate. The enzyme activity was decreased by Mg²⁺, Cu²⁺, Zn²⁺, Al³⁺, K⁺, Li⁺, Ag⁺, urea, EDTA, trichloroacetic acid and Tween 60 and inhibited by Hg²⁺, Ce²⁺ and SDS, whereas the activity was increased by Mn²⁺, DTT, and β‐mercaptoethanol. Ca²⁺and Fe²⁺ did not affect the enzyme activity.     

Functional Expression of the Raw Starch‐Binding Domain of Bacillus sp. Strain TS‐23 α‐Amylase in Recombinant Escherichia coli

Two DNA fragments encoding the starch‐binding domain (SBD) of Bacillus sp. strain TS‐23 α‐amylase were prepared by polymerase chain reaction and cloned into the Escherichia coli expression vector, pQE‐30, to generate pQE‐N428/C607 and pQE‐N465/ C607. In isopropyl‐β‐D ‐thiogalactopyranoside (IPTG)‐induced E. coli strain M15 harboring these expression plasmids, the recombinant SBDs (N428/C607 and N465/C607) could comprise up to 20% of the total soluble proteins. The His‐tag/SBD fusion proteins were purified to homogeneity with a His‐bind affinity column and had molecular masses of approximately 22.6 and 16.5 kDa, respectively. Starch‐binding assays revealed that about 7.1 and 8.3 μg, respectively, of N428/C607 and N465/C607 were bound by 1 mg of raw corn starch, indicating that the SBD of Bacillus sp. strain TS‐23 α‐amylase retain sufficient function in the absence of a catalytic center.     

Inhibitory activities of kaempferol,galangin, carnosic acid and polydatin against glycation and α‐amylase and α‐glucosidase enzymes

The activities of four natural phenolics, kaempferol, galangin, carnosic acid and polydatin in scavenging free radicals, inhibiting advanced glycation end‐product (AGE) formation, α‐amylase and α‐glucosidase and trapping methylglyoxal (MGO), were evaluated in this study. Carnosic acid and galangin had the highest activity in scavenging free radicals. Kaempferol and galangin had the greatest activity in preventing bovine serum albumin (BSA) against glycation and reducing glycated proteins. Polydatin had the greatest performance in trapping MGO to reduce glycation reaction. However, there was no significant difference for kaempferol, galangin and carnosic acid in inhibiting AGE formation by BSA‐MGO reaction. Kaempferol, galangin and carnosic acid were the competitive inhibitors for α‐amylase, while kaempferol and carnosic acid were noncompetitive inhibitors for α‐glucosidase. However, polydatin showed as a mixed noncompetitive inhibitor for both α‐amylase and α‐glucosidase. The results indicated that the four natural phenolics have potential in inhibiting AGE production and the digestive enzymatic activity with different mechanisms.     

α‐Glucosidase and α‐amylase inhibitory activities of phloroglucinal derivatives from edible marine brown alga,Ecklonia cava

BACKGROUND: Diabetes mellitus (DM) is a chronic metabolic disorder characterized by defects in insulin secretion and action, which can lead to damaged blood vessels and nerves. With respect to effective therapeutic approaches to treatment of DM, much effort has being made to investigate potential inhibitors against α‐glucosidase and α‐amylase from natural products. The edible marine brown alga Ecklonia cava has been reported to possess various interesting bioactivities, which are studied here. RESULTS: In this study, five phloroglucinal derivatives were isolated from Ecklonia cava: fucodiphloroethol G ( 1 ), dieckol ( 2 ), 6,6′‐bieckol ( 3 ), 7‐phloroeckol ( 4 ) and phlorofucofuroeckol A ( 5 ); compounds 1, 3 and 4 were obtained from this genus for the first time and with higher yield. The structural elucidation of these derivatives was completely assigned by comprehensive analysis of nuclear magnetic spectral data. The anti‐diabetic activities of these derivatives were also assessed using an enzymatic inhibitory assay against rat intestinal α‐glucosidase and porcine pancreatic α‐amylase. Most of these phlorotannins showed significant inhibitory activities in a dose‐dependent manner, responding to both enzymes, especially compound 2 , with the lowest IC₅₀ values at 10.8 µmol L^?1 (α‐glucosidase) and 124.9 µmol L^?1 (α‐amylase), respectively. Further study of compound 2 revealed a non‐competitive inhibitory activity against α‐glucosidase using Lineweaver‐Burk plots. CONCLUSION: These results suggested that Ecklonia cava can be used for nutritious, nutraceutical and functional foods in diabetes as well as for related symptoms. Copyright © 2009 Society of Chemical Industry     

In vitro inhibitory effects of cranberry bean (Phaseolus vulgaris L.) extracts on aldose reductase, α‐glucosidase and α‐amylase

The in vitro inhibitory activities of different seed extracts prepared from cranberry bean mutant SA‐05 and its wild‐type variety Hwachia against aldose reductase, α‐glucosidase and α‐amylase were examined. The results indicated that the polyphenolics‐rich extracts obtained using 800 g kg^?1 methanol and 500 g kg^?1 ethanol demonstrated inhibitory activities against aldose reductase (IC₅₀ of 0.36–0.46 mg mL^?1) and α‐glucosidase (IC₅₀ of 1.32–1.94 mg mL^?1). The 500 g kg^?1 ethanol extracts also showed α‐amylase inhibitory activities (IC₅₀ of 70.11–80.22 μg mL^?1). Subsequent extracts, prepared further with NaCl and H₂O from precipitates of 800 g kg^?1 methanol or 500 g kg^?1 ethanol extracts, exhibited potent α‐amylase inhibitory activities (IC₅₀ of 17.68–38.68 μg mL^?1). A combination of 500 g kg^?1 ethanol extraction plus a subsequent H₂O extraction produced highest polyphenolics and α‐amylase inhibitors. The SA‐05 α‐amylase inhibitor extracts showed greater inhibitory activities than that of Hwachia. Thus, cranberry bean mutant SA‐05 is an advantageous choice for producing anti‐hyperglycaemic compounds.     

Inhibitory potential of phenylpropanoid glycosides from Ligustrum purpurascens Kudingcha against α‐glucosidase and α‐amylase in vitro

The leaves of Ligustrum purpurascens are used in a Chinese traditional tea called small‐leaved kudingcha, which is rich in phenylpropanoid glycosides (PPGs) and has many beneficial properties. Two critical exoacting glycoside hydrolase enzymes (glucosidases) involved in carbohydrate digestion are α‐glucosidase and α‐amylase. We investigated the properties of PPGs from L. purpurascens for inhibiting α‐amylase and α‐glucosidase activity in vitro and found IC₅₀ values of 1.02 and 0.73 mg mL^?1, respectively. The patterns of inhibiting both α‐amylase and α‐glucosidase were mixed‐inhibition type. Multispectroscopy and molecular docking studies indicated that the interaction between PPGs and α‐amylase and α‐glucosidase altered the conformation of enzymes, with binding at the site close to the active site of enzymes resulting in changed enzyme activity. Our studies may help in the further health use of small‐leaved kudingcha.     

Screening grape seeds recovered from winemaking by‐products as sources of reducing agents and mammalian α‐glucosidase and α‐amylase inhibitors

Grape seeds collected from vinification of various grape varieties were extracted by supercritical CO₂ for oil recovery. The defatted residues thus obtained were considered as a re‐utilisable co‐product and assessed for phenolic content, reducing capacity and inhibitory activities against mammalian α‐amylase and α‐glucosidase enzymes. Supercritical CO₂ treatment led to higher recovery of anthocyanins. Reducing capacity of phenolic extracts reached up to ~2200 mmolFe(II) kg^?1, much higher than that of various natural phenolic sources. The anthocyanin‐rich extracts showed the highest inhibitory effectiveness towards α‐glucosidase (I₅₀ value equal to ~40 μg gallic acid equivalents (GAE)/mL ~ half than acarbose). Inhibitory effectiveness towards α‐amylase activity was similar among grape varieties, with I₅₀ values comparable to that of acarbose and correlated with proanthocyanidin contents. These results could pave the way for an efficient processing of grapes, including cascade processes, namely: winemaking, oil extraction from recovered grape seeds and phenolic extraction from defatted grape seeds as potential cost‐effective nutraceuticals.     

Alterations of the Profiles of Iso‐α‐Acids During Beer Ageing,Marked Instability of Trans‐Iso‐α‐Acids and Implications for Beer Bitterness Consistency in Relation to Tetrahydroiso‐α‐Acids

Age‐induced decomposition of iso‐α‐acids, the main bittering principles of beer, determines the consistency of the beer bitter taste. In this study, the profiles of iso‐α‐acids in selected high‐quality top‐fermented and lager beers were monitored by quantitative high‐performance liquid chromatography at various time intervals during ageing. The degradation of the iso‐α‐acids as a function of time is represented by the ratio, in percentage, of the sum of the concentrations of trans‐isocohumulone and trans‐isohumulone to the sum of the concentrations of cis‐isocohumulone and cis‐isohumulone. This parameter is relevant with respect to the evaluation of bitterness deterioration in aged beers. Trans‐iso‐α‐acids having a shelf half‐life of less than one year proved to be significantly less stable than cis‐iso‐α‐acids, but it appears feasible to counteract degradation if a suitable beer matrix is available. The fate of the trans‐iso‐α‐acids in particular adversely affects beer bitterness consistency. In addition to using hop products containing low amounts of trans‐iso‐α‐acids, brewers may profit of the remarkable stability of tetrahydroiso‐α‐acids, even on prolonged storage, for the production of consistently bitter beers.     

Characterization of the Supermolecular Structure of Polydatin/6‐O‐α‐Maltosyl‐β‐cyclodextrin Inclusion Complex

Polydatin is the main bioactive ingredient in many medicinal plants, such as Hu‐zhang (Polygonum cuspidatum), with many bioactivities. However, its poor aqueous solubility restricts its application in functional food. In this work, 6‐O‐α‐Maltosyl‐β‐cyclodextrin (Malt‐β‐CD), a new kind of β‐CD derivative was used to enhance the aqueous solubility and stability of polydatin by forming the inclusion complex. The phase solubility study showed that polydatin and Malt‐β‐CD could form the complex with the stoichiometric ratio of 1:1. The supermolecular structure of the polydatin/Malt‐β‐CD complex was characterized by ultraviolet–visible spectroscopy (UV), Fourier transform infrared spectroscopy (FT‐IR), X‐ray diffractometry (XRD), thermogravimetric/differential scanning calorimetry (TG/DSC), and proton nuclear magnetic resonance (¹H‐NMR) spectroscopy. The changes of the characteristic spectral and thermal properties of polydatin suggested that polydatin could entrap inside the cavity of Malt‐β‐CD. Furthermore, to reasonably understand the complexation mode, the supermolecular structure of polydatin/Malt‐β‐CD inclusion complex was postulated by a molecular docking method based on Autodock 4.2.3. It was clearly observed that the ring B of polydatin oriented toward the narrow rim of Malt‐β‐CD with ring A and glucosyl group practically exposed to the wide rim by hydrogen bonding, which was in a good agreement with the spectral data.     

Combination of acid‐demineralization and enzymatic desizing of cotton fabrics by using industrial acid stable glucoamylases and α‐amylases

In this study, the possibility of simultaneous acid‐demineralization and enzymatic desizing of cotton fabric in acidic conditions (pH 2) by using industrial acid stable enzymes has been investigated. Acid‐demineralization is necessary to remove undesired cationic metals and earth alkalis. Our experiments showed that by use of a mixture of two appropriate enzymes, a glucoamylase (Multifect GA 10L) and an α‐amylase (Optisize Next) in a solution of citric acid and presence of a chelating agent, enzymatic desizing, and acid‐demineralization can be successfully carried out at the same time. Therefore, two processes of pretreatment were integrated into a single process, which can effectively reduce time and costs for textile industry.