Determination of total flavonoids content in buckwheat and inhibition of α-amylase activity

Diabetes is one of the most difficult chronic diseases to cure in the world,which seriously affects people’s health and quality of life.Flavonoids in buckwheat can regulate blood glucose levels by inhibitingα-amylase activity.Therefore,sweet buckwheat produced in Inner Mongolia was used as the research object,and buckwheat fl avonoids were extracted by ultrasonic-assisted extraction method.Total fl avonoids content was determined by ultraviolet-visible spectrophotometry.With acarbose as the positive control,the inhibition test ofα-amylase was carried out by DNS colorimetry to study the inhibition behavior of fl avonoids onα-amylase activity.The results showed that the extraction process of flavonoids was stable and reliable,and the established method for the determination of flavonoids was simple,accurate and reproducible.The total flavonoids content of buckwheat samples was 2.706 mg/g,buckwheat total fl avonoids extraction solution had an inhibitory eff ect onα-amylase,and its median inhibition concentration(IC_(50))was 38.53 mg/mL.The results of this experiment provide a technical reference for the development and utilization of fl avonoids in Inner Mongolia sweet buckwheat,and provide a theoretical reference for the development and application of flavonoid-rich hypoglycemic food.

Comparative analysis of physicochemical properties,ginsenosides content andα-amylase inhibitory effects in white ginseng and red ginsen

Ginseng(Panax ginseng C.A.Meyer)as a common dietary adjunct is widely applied in Traditional Chinese Medicine due to its health-promoting properties,but the differences between white ginseng and red ginseng was rarely studied.In the present study,color parameters and scanning electron microscope(SEM)were determined to evaluate the differences of ginseng color and microstructure induced by processing procedure.Quantitative analysis of multi-components by a single-marker(QAMS)method and anti-α-amylase activity test were used to assess variations of chemical ingredients and pharmacological activity between white and red ginseng.Finally,molecular docking studies were carried out to screen out the most effective compound againstα-amylase.Results indicated that processing had a significant impact on the physicochemical properties and pharmacological activity of white and red ginseng.After processing,the color value of L*declined significantly.Red ginseng sample displayed a compact structure and presented of a gel layer on the surface compared to white ginseng.Additionally,the content of ginsenosides and the activity of anti-α-amylase decreased.The contents of total ginsenosides were positively correlated with the anti-α-amylase activities of ginseng,and ginsenoside Rb1 might be the most effective compound to inhibit the activity ofα-amylase.

Chemical Constituents of Pu-erh Tea and Its Inhibition Effect on α-amylase in vitro

[Objective]The aim was to clarify the chemical substance basis of hypoglycemic and lipid-lowering effects of Pu-erh Tea. [Method]Pu-erh Tea was extracted with 95% ethanol,followed by petroleum ether,chloroform,ethyl acetate and n-butanol extraction,after the further purification and through the NKA-9 macroporous resin and many times of Sephadex column chromatography,two compounds were isolated,in the same time,the effect of Uracil and Gallic acid on α-amylase was studied. [Result]The Uracil and Gallic acid were isolated and identified respectively from Pu-erh Tea and the Uracil was firstly isolated from Pu-erh Tea; Gallic acid had strong inhibition on α-amylase. [Conclusion]It could provide some theories on the hypoglycemic and lipid-lowering effects of Pu-erh Tea.

Analysis of α-amylase Inhibitor Content Change in Pu-erh Tea During Pile-fermentation Process

The study was to investigate the changes of α-amylase inhibitor content in Pu-erh tea during pile-fermentation process. Pu-erh tea samples from two regions of Shuangjiang County and Jinggu Dai and Yi Autonomous County of Yunnan Province at various fermentation stages were used as experimental materials to investigate the effect of different fermentation stages on the inhibitory effect to α-amylase; and the change law of the inhibitory effect of c-amylase inhibitor during processing was meanwhile studied by determining the contents of tea polyphenol and amino acid. The results showed that crude meterial of Pu-erh tea presented strong inhibitory effect to α-amylase; this inhibitory effect assumed a de： creasing trend to the minimum at the middle stage of fermentation, whereafter it increased to some extent. Made tea also showed a strong inhibitory effect to α-amylase. During whole processing period, contents of tea polyphenol and amino acid generally assumed a remarkably decreasing trend. Our results provided references for further isolating co-amylase inhibitor from Pu-erh tea and discussing the mechanism of its health care function.

Studies on the Isolation and Screening of α-amylase Producing Strain and Their Enzymatic Properties

[Objective] The aim was to obtain α-amylase producing strains with some excellent properties like high temperature resistance,strong acid resistance,strong alkali resistance,etc..[Method] α-amylase producing strains were isolated and screened; furthermore their enzymatic properties were studied.[Result] 10 strains with an obvious starch hydrolysis cycle were screened out from starch screening plate coated by diluted sample,from which 3 strains with higher α-amylase activity were screened out,that was X6,X8 and X10.As for X6,X8 and X10,their optimum pH values all belonged to neutral,and their optimum temperatures were all 60 ℃.Meanwhile,Ca^2＋ could increase their enzyme thermal stability.When the concentration of Ca^2＋ was 0.02-0.04 mol/L,the enzyme thermal stability of X6 and X8 reached the highest; When the concentration of Ca^2＋ was 0.03-0.04 mol/L,that of X10 reached the highest; When the concentration of Ca^2＋ was increased continuously,those of the 3 strains all decreased.[Conclusion] The research provides theoretical basis for satisfying the demands of different industries for α-amylase with different characteristics.

Germination and Expression of α-Amylase of KF0680-1 and KF0680-2(Cultivars of Sweet Sorghum) under Salt Stress

[Objective] The aim was to study on germination and expression of α-amy- lase of KF0680-1 and KF0680-2 （cultivars of sweet sorghum） under salt stress. [Method] In the research, KF0680-1 and KF0680-2, cultivars of sweet sorghum, were used to measure related indices of germination and expression of co-amylase to discuss effect of salt stress on two cultivars. [Result] Germination of sweet sorghum would be promoted if treated with salt in low concentration and inhibited if treated in high concentration. In the latter condition, roots and seedlings were smaller than that of control group in length, but expression of s-amylase in the two cultivars was pro- moted under proper salt stress. [Conclusion] The research indicates that KF0680-1 and KF0680-2 are inhibited in growth by salt stress and the latter is stronger in salt resis- tance than the former under low salt concentration. In addition, expression of α-amy- lase could be promoted by salt in proper volume. The results provide references for selection of sweet sorghum which could be planted in northwestern areas.

In vitro α-amylase inhibitory activity and in vivo hypoglycemic effect of methanol extract of Citrus macroptera Montr.fruit

Objective:To investigate the therapeutic effects of methanol extract of Citrus macroptera Montr,fruit inα-amylase inhibitory activity(in vitro)and hypoglycemic activity in normal and glucose induced hyperglycemic rats(in vivo).Methods:Fruits of Citrus macroptera without rind was extracted with pure methanol following cold extraction and tested for presence of phytochemical constituents,α-amylase inhibitory activity,and hypoglycemic effect in normal rats and glucose induced hyperglycemic rats.Results:Presence of saponin,steroid and terpenoid were identified in the extract.The results showed that fruit extract had moderateα-amylase inhibitory activity[IC_(50)value=(3.638±0.190)mg/mL]as compared to acarbose.Moreover at 500 mg/kg and 1000 mg/kg doses fruit extract significantly(P<0.05 and P<0.01 respectively)reduced fasting blood glucose level in normal rats as compared to glibenclamide(5 mg/kg).In oral glucose tolerance test,500 mg/kg dose significantly reduced blood glucose level(P<0.05)at 2 h but 1000 mg/kg dose significantly reduced blood glucose level at 2 h and 3 h(P<0.05 and P<0.01 respectively)whereas glibenclamide(5 mg/kg)significantly reduced glucose level at every hour after administration.Overall time effect is also considered extremely significant with F value=23.83 and P value=0.0001 in oral glucose tolerance test.Conclusion:These findings suggest that the plant may be a potential source for the development of new oral hypoglycemic agent.

Comparisons of phaseolin type and α-amylase inhibitor in common bean(Phaseolus vulgaris L.)in China

The objective of this study was to characterize the phaseolin type and a-amylase(αAI) level in common bean(Phaseolus vidgaris L.) accessions deposited in the Chinese National Genebank.The 40 accessions sampled were common varieties originating in Asia,North America,South America,Europe,and Africa.No Inca(I-) phaseolin was observed in the accessions.Only four accessions contained Tendergreen(T-) phaseolin and the remaining36 contained Sanilac(S-) phaseolin.aAI proteins extracted from nine accessions showed higher a-amylase inhibitory activity than the control(Phase 2,IC_(50) = 0.65 μg).These common bean accessions have potential use as nutraceutical ingredients.

Relationship between malt qualities and β-amylase activity and protein content as affected by timing of nitrogen fertilizer application

The effects of different timing of N fertilizer application at the same rate on grain β-amylase activity, protein con- centration, weight and malt quality of barley were studied. Grain β-amylase activity and protein concentration were significantly higher in treatments where all top-dressed N fertilizer was applied at booting stage only or equally applied at two-leaf stage and booting stage than in the treatment where all top-dressed N fertilizer was applied at two-leaf age stage only. On the other hand, grain weight and malt extract decreased with increased N application at booting stage. There were obvious differences between barley varieties and experimental years in the grain and malt quality response to the timing of N fertilizer application. It was found that grain protein concentration was significantly and positively correlated with β-amylase activity, but significantly and nega- tively correlated with malt extract and Kolbach index. The effect of grain protein concentration on malt quality was predominant over the effect of grain β-amylase activity.

Effects of Ionic Surfactants on Bacterial Luciferase and α-Amylase

In order to study the effects of ionic surfactants on bacterial luciferase,the cationic surfactant dodecyltrimethylammonium biomide (DTAB) and anionic surfactant sodium dodecylsulfate (SDS) were chosen.For comparison with bacterial luciferase,α-amylase was used since these two enzymes have similar electrostatic potential and charged active sites.After the enzymes were treated with the surfactants,the catalytic properties of bacterial luciferase andα-amylase were assayed,and fluorescence spectroscopy and circular dichroism (CD) were used to analyze the alteration of the protein structure.The results showed that when the DTAB concentration was low,the cationic surfactant DTAB enhanced the enzymatic activities of bacterial luciferase andα-amylase.On the other hand,the anionic surfactant SDS did not alter the enzymatic activity.The main interaction of cationic surfactant DTAB and the negatively charged surface of the proteins was the ionic interaction,which could alter the environment for the enzyme to work when the DTAB/enzyme molar ratio was low.However,at high cationic surfactant concentration,the ionic interaction and hydrophobic interaction might destroy the secondary and tertiary structures of the proteins,leading to the loss of enzymatic activities.

Topographies of Organized Monolayer of α-Amylase Observed by Atomic Force Microscopy

In this paper, (-amylase organized monolayer was assembled on the surface of the PET-CO2- substrate in different conditions. The different topography of the (-amylase/PET monolayer was obtained by AFM in tapping mode.

Effect of microwave irradiation on secondary structure of α-amylase by circular dichroism

Based on the starch hydrolysis reaction accelerated by microwave irradiation with α-amylase, the circular dichroism （CD） and secondary structure changes of α-amylase under the condition of microwave irradiation and water bath were studied by circular dichroism spectra. The results showed that, both the peak heights （at 2=193 nm） of the CD spectra of the samples treated by microwave irradiation and water bath reduced. The reduced rate by microwave irradiation ranged from 140% to 220%, while the reduced rate by water bath ranged from 60% to 140%. The peak of the sample treated by microwave irradiation for 60 min disappeared at λ=193 nm, while the sample showed a wake peak by water bath. The peak position by microwave irradiation emerged a blue shift in the range of 5-8 nm at λ=204 nm and λ=220 nm, while it emerged in the range of 3-5 nm by water bath. With time going on, the microwave irradiation and water bath have prompted the secondary structure of α-helix, β-sheet, β-turn and the mutual transformations of random coil, but the trends were different.

The Dynamics of Changes in Starch and Lipid Droplets and Sub-Cellular Localization of β-Amylase During the Growth of Lily Bulbs

The ultra-structure of mother and outer daughter scales of Lilium Oriental hybrid Sorbonne were studied using transmission electron microscope to examine the sub-cellular localization of starch and lipid droplets during growth and development from shoot emergence to senescence.The contents of starch granules and lipid droplets in the cell of the mother scales decreased significantly from shoot emergence to anthesis,indicating that these scales served as a source for growth and development.After flowering,the number of starch granules and lipid droplets increased dramatically,and finally the cells were filled with the above molecules indicating that the bulb becomes a major sink during bulb enlargement.Ultrastructure observation also showed that symplastic pathway is the main pathway in cells in the exchange and transportation of material during bulb development.The activity of β-amylase,one of the key enzymes catalyzing starch breakdown,showed a similar trend.The enzyme sub-cellular localization via immune-gold electron-microscopy showed that βamylase was predominantly located together with starch granules,while the gold particles were scarcely found in other sub-cellular compartments.The result suggested that this enzyme is compartmented together with its functional substrate supporting its function in catalyzing starch breakdown in living plant cells.

The relationship between psychological distress with salivary <i>α</i>-amylase and stress related symptoms among breast cancer survivors

Background and Aims: This study examined the stress levels, the stressors and biomarker such as Alpha (α)-Amylase enzyme which is secreted under distress conditions. The aim was to determine the relationship between these three variables. Methods: The study respondents were made up of cancer outpatients from 3 hospitals namely the Hospital Kuala Lumpur, Seberang Jaya and Johor Baru, Pantai Ipoh and Putrajaya. The Personal Stress Inventory (PSI) questionnaire was used to identify stressors, while the Hospital Anxiety and Depression Scale (HADS) were used to determine the psychological distress levels. The Salimentary Oral Swab (SOS) Technique was used to collect the saliva and then the Salivary α-Amylase Assay Kit was used to analyse for α amylase. Results: The majority of respondents were stressed due to their sensitive emotion and nervousness in their daily lives (68.7%), they had poor memory and short attention spans of carrying out job tasks (67.3%) as well as they were emotionally depressed (65.3%). Their poor memory and short attention span (p = 0.037), heavy work load and poor task delivery (p = 0.008) were predictors for distress using HADS. The salivary α-amylase concentration was significantly related to the stress levels (p = 0.002). Conclusion: Breast cancer respondents with musculoskeletal system related symptoms, with heavy work load and poor task delivery, as well as had poor memory and short attention span were at higher risk of experiencing psychological distress. The salivary α-amylase which had significant relationships with psychological distress was therefore, a potential biological indicator for distress, while the musculoskeletal system related symptoms from the PSI questionnaire were predictors for distress.

Efficiency of Oryza punctata extract on glucose regulation: Inhibition of α-amylase and α-glucosidase activities

Red rice(Oryza punctata) is a type of unpolished rice which has higher nutritional value compared to white rice or even polished rice. Owing to higher nutritive content and metabolites, dieticians strongly advise red rice for peoples with metabolic disorders including diabetics. However, the mechanism of action and contents of secondary metabolites in Indian red rice variety not reported scientifically. Therefore, the present study aimed to evaluate its mechanism of action through inhibitory effect of α-amylase and α-glucosidase. Initially, the whole grain of red rice was macerated with methanol at room temperature for 2 weeks. Then, the dried and powdered, samples at different concentrationfi(2.5, 10, 40, and 80 μg/m L) were employed to nd out in vitro inhibitory effects on α-amylase and α-glucosidase. In addition, an enzyme kinetics of effective extract was calculated by Line-weaver Burk(LWB) plot analysis. Moreover,the valuable metabolites in the efficient methanolic extract were quantified using high performance liquid chromatography(HPLC). The results demonstrated that red rice methanolic extract(RRMEt) possess strong inhibitory activity onα-amylase and α-glucosidase compared with acarbose(P < 0.01). The IC50 values of RRMEt was found to be 29.7 ±7.43 μg/m L for α-amylase and 20.4 ± 0.25 μg/m L for α-glucosidase. LWB indicated that RRMEt is an uncompetitive inhibitor. Further, HPLC analysis revealed protocatechuic acid, catechin, and chlorogenic acids were more abundant in RRMEt among the fourteen metabolites. We conclude, the efficiency of enzyme inhibition through the influence of phenolic compounds in RRMEt.

Synthesis and Characterization of Zinc Oxide and Zinc Oxide Doped with Chlorine Nanoparticles as Novel <i>α</i>-Amylase Inhibitors

In this study we used a chemical solution method from oxalic acid (OX. acid) and zinc acetate (ZnAc) to prepare Zinc Oxide nanoparticles (ZnONPs) and Zinc Oxide nanoparticles doped with Chlorine (Cl:ZnONPs). The characterizations (FTIR, X-ray, SEM, TEM) of ZnONPs and Cl:ZnONPs were determined. Amylase inhibitors of ZnONPs and Cl:ZnONPs also were determined. SEM indicated that the ZnONPs and Cl:ZnONPs have an average particle size of 46.65 - 74.64 nm. TEM images of the ZnONPs and Cl:ZnONPs showed the round shaped. Compounds b, d and e exhibited significant inhibitory activity against amylase enzyme (from 69.21 ± 1.44 to 76.32 ± 0.78), respectively, and were comparable with that of acarbose (86.32 ± 0.63) at 1000 μg, thereby, projecting ZnONPs and Cl:ZnONPs as α-amylase inhibitors.

Purification and Characterization of Cold-active α-Amylase Excreted by A Strain of Marine Cold-adaptive Penicillia

The filamentous fungi from the Huanghai sea sludge were screened according to their ability to produce cold-active α-amylase. The strain with the highest amylase activity was identified as Penicillium species. The α-amylase purified by ammonium sulphate precipitation and column chromatography on DEAE-sepharose and sephadex G-100 shows a molecular weight of about 55000 and a pI of 4.38. The enzyme is stable in a pH range of 5.5—8.0 and has a maximum activity at pH 6.0. Compared with the α-amylase from mesophiles and thermophiles, the cold-active enzyme shows a high enzyme activity at lower temperatures and a high sensitivity at temperatures higher than 50 ℃. The optimal temperature is 40 ℃ and the activity decreases dramatically at temperatures above 50 ℃. Ca 2+ shows a significant effect on maintaining the structure and the activity of the enzyme. EDTA and Cu 2+ are its inhibitors. The products from the hydrolysis of soluble starch with the cold-active enzyme are maltose and other oligosaccharides.

Inhibition of α-amylase and α-glucosidase activities by ethanolic extract of Telfairia occidentalis (fluted pumpkin) leaf

Objective:To investigate the inhibitory effect of Telfairia occidentalis Hook f.(Curcubitaceae)(T.occidentalis)leaf on key enzyme linked to type-2 diabetes(α-amylase andα-glucosidase)as well as assess the effect of blanching(a commonly practiced food processing technique)of the vegetable on these key enzymes.Methods:Fresh leaves of T.occidentalis were blanched in hot water for 10 minutes,and the extracts of both the fresh and blanched vegetables were prepared and used for subsequent analysis.The inhibitory effect of the extract onα-amylase andα-glucosidase activities as well as some antioxidant parameter was determined in vitro.Results:The result revealed that unprocessed T.occidentalis leaf reduce Fe^(3+)to Fe^(2+)and also inhibitedα-amylase andα-glucosidase activities in a dose dependent manner.However,blanching of the leafy vegetables caused a significant(P<0.05)increase in the antioxidant properties but decrease their ability to inhibitα-amylase andα-glucosidase activities.Conclusions:This antioxidant properties and enzyme inhibition could be part of the mechanism by which they are used in the treatment/prevention of type-2 diabetes.However,the blanched vegetable reduces their ability to inhibit bothα-amylase andα-glucosidase activity in vitro.

Kinetics of medium-temperature α-amylase hydrolyzed <i>Huai yam</i>powder

In order to learn the enzymatic characteristics of Huai yam powder with medium-temperatureα-amylase, effects of substrate concentration,enzyme concentration, pH and temperature wereinvestigated. The Michealis-Menten equationwas used to fit the kinetics of the hydrolysis reaction. Experimental results indicate that maximum rate (Vm) is 3.1588 mg/mL·min under the condition of 70&#176;C, pH 7.0 and 0.0200 mg/mL of enzyme concentration. The Michealis constant (Km) is 6.6641 mg/mL. The kinetic model, including the factors such as substrate concentration, enzyme concentration and temperature, was established for the hydrolysis reaction under the temperature range from 40&#176;C - 70&#176;C.

Inheritance of Late Maturity α-amylase (LMA) in a Recombinant Inbreed Lines of Chuanmai 42 Crossed with Chuannong 16

[Objective] To study the inheritance and possible molecular marker of α-LMA and their relations to falling number. [Method] Recombined inbreed lines (RILS) of Chuanmai 42 (LMA genotype) crossed with Chuannong 16 (no LMA) were employed to test the late maturity α-amylase (LMA) by EL ISA kits,the Falling Number (FN),and three QTL,SSR markers (Xgwm 577,Xwmc276,Xwmc273) of LMA gene. [Result] Results suggested that LMA was controlled by a single,and the three SSR markers seemed to be not efficient to test LMA gene in this RILS. LMA was an important factor on FN,meanwhile FN was also affected by other factors. [Conclusion] The study could provide references for the breed of quality wheat.