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Distance-basedα-amylase biosensor fabricated with amylopectin-coated mesoporous membrane

Chinese Chemical Letters 2024年第1期35卷 446-450页

作者： Binglu Zhao Mashooq Khan Yulin Liu Wenjun Tai Chongyang Mu Wenli Wu Mei Zhao Yaohong Ma Li Yu Jin-Ming Lin Qiongzheng Hu Qilu University of Technology(Shandong Academy of Sciences) Shandong Analysis and Test CenterJi’nan 250014China School of Pharmaceutical Sciences Qilu University of Technology(Shandong Academy of Sciences)Ji’nan 250014China Key Laboratory of Colloid and Interface Chemistry Shandong UniversityMinistry of EducationJi’nan 250100China Key Laboratory for Biosensors of Shandong Province Biology InstituteQilu University of Technology(Shandong Academy of Sciences)Ji’nan 250353China Beijing Key Laboratory of Microanalytical Methods and Instrumentation MOE Key Laboratory of Bioorganic Phosphorus Chemistry&Chemical BiologyDepartment of ChemistryTsinghua UniversityBeijing 100084China

Paper-based biosensors are widely employed in point-of-care testing(POCT)due to their convenience,portability,low cost,and ease of use.This study reports an integrated distance-based paper biosensor fabricated with a mesoporous membrane coated with stimuli-responsive polymer.The detection ofα-amylase(AMY)using amylopectin-coated mesoporous membrane is demonstrated as an example.After introducing the AMY solution,it is observed that the aqueous solution flows along the paper strip due to AMY-catalyzed hydrolysis of amylopectin.The flow distance is proportional to the concentration of AMY with a detection limit as low as 4 mU/mL.In addition,the detection of AMY is demonstrated in human serum.Furthermore,the inhibitory effect of acarbose on AMY is evaluated.This reagent-free and disposable biosensor allows single-step rapid detection of the analyte.This approach is very promising for the development of user-friendly,equipment-free,and cost-effective biosensors with remarkable sensitivity and excellent selectivity for disease diagnosis and hypoglycemic drug screening.

关键词： Paper biosensor α-amylase Stimuli-responsive polymer Mesoporous membrane Distance Point-of-care testing

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

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Food Science and Human Wellness 2023年第1期12卷 14-27页

作者： Huairui Wang Yao Cheng Xue Zhang Yingping Wang Hui Zhao State Local Joint Engineering Research Center of Ginseng Breeding and Application College of Chinese Medicinal MaterialsJilin Agricultural UniversityChangchun 130033China School of Biotechnology and Food Science Tianjin University of CommerceTianjin 300134China School of Pharmaceutical Science and Technology Health Sciences PlatformTianjin UniversityTianjin 300072China

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.

关键词： Ginseng Color Microstructure Ginsenosides α-amylase

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Cloning,expression,and characterization of a porcine pancreatic α-amylase in Pichia pastoris

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Animal Nutrition 2018年第2期4卷 234-240页

作者： Lv-hui Sun Tao Qin Yan Liu Hua Zhao Xinjie Xia Xingen Lei Department of Animal Nutrition and Feed Science Huazhong Agricultural University International Center of Future Agriculture for Human Health Sichuan Agricultural University Wuhan Chopper Biology Co. LTD Department of Animal Science Cornell University

Pancreatic α-amylase(α-1, 4-glucan-4-glucanohydrolase, EC.3.***.1.1) plays a primary role in the intestinal digestion of feed starch and is often deficient in weanling pigs.The objective of this study was to clone,express, and characterize porcine pancreatic α-amylase(PPA).The full-length c DNA encoding the PPA was isolated from pig pancreas by RT-PCR and cloned into the pPICZαA vector.After the resultant pPICZαА-PPA plasmid was transferred into Pichia pastoris, Ni Sepharose affinity column was used to purify the over-expressed extracellular recombinant PPA protein(re PPA) that contains a His-tag to the C terminus and was characterized against the natural enzyme(α-amylase from porcine pancreas).The re PPA exhibited a molecular mass of approximately 58 kDa and showed optimal temperature(50℃),optimal pH(7.5), K_m(47.8 mg/mL), and V_(max)(2,783 U/mg) similar to those of the natural enzyme.The recombinant enzyme was stable at 40℃ but lost 60% to 90%(P amylase gene and over-expressed the gene in P.pastoris as an extracellular, functional enzyme.The biochemical characterization of the over-produced enzyme depicts its potential and future improvement as an animal feed additive.

关键词： Porcine α-amylase Pancreatic Gene expression Pichia pastoris

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

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The Crop Journal 2016年第1期4卷 68-72页

作者： Yang Yao Yibo Hu Yingying Zhu Yue Gao Guixing Ren Institute of Crop Science Chinese Academy of Agricultural Sciences

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.

关键词： Common bean Phaseolin α-amylase inhibitor Phenolic acid

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Study on the interaction mechanism of virgin olive oil polyphenols with mucin and α-amylase

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Food Bioscience 2022年第3期47卷 728-735页

作者： Gangcheng Wu Ruobing Mao Yiren Zhang Ling Zhu Emad Karrar Hui Zhang Qingzhe Jin Xingguo Wang State Key Laboratory of Food Science and Technology School of Food Science and TechnologyNational Engineering Research Center for Functional FoodInternational Joint Research Laboratory for Lipid Nutrition and SafetyCollaborative Innovation Center of Food Safety and Quality Control in Jiangsu ProvinceJiangnan UniversityChina International Joint Laboratory on Food Safety Jiangnan UniversityChina Department of Chemistry School of Physical ScienceUniversity of LiverpoolUK

The interaction mechanism of mucin and α-amylase with virgin olive oil (VOO) polyphenols (oleuropein (OL),tyrosol (TY),hydroxytyrosol (HT)) was analyzed by fluorescence,ultraviolet (UV) absorption,attenuated total reflection Fourier transform infrared (ATR-FTIR),circular dichroic (CD) spectroscopy and molecular docking.A total of 17 polyphenols have been identified in the selected VOO,and the TY,HT and OL were the main compounds.The quenching mechanism between mucin/α-amylase and three VOO polyphenols was static,mainly through van der Waals forces and hydrogen bonds.The results of UV absorption,ATR-FTIR,and CD spectroscopy revealed that the conformation of mucin changed after combining with the three polyphenols,while that of the α-amylase changed little.Molecular docking predicted the interaction sites of the three polyphenols with human salivary mucin and α-amylase.The present study could provide the theoretical foundation for further research on the interaction between human salivary protein and phenolic compounds in VOO.

关键词： Mucin α-amylase Spectroscopy Molecular docking Virgin olive oil polyphenols

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Optimization of Growth Conditions to Identify the Superior Bacillus Strain Which Produce High Yield of Thermostable Alpha amylase

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Advances in Enzyme Research 2022年第1期10卷 1-22页

作者： Witharanage Wasana Prasadini Rodrigo Lakmi Samodha Magamulla Morawakage Sajith Thiwanka Yapa Mudiyanselage Shashika Madhuwanthi Yapa Industrial Technology Institute Colombo Sri Lanka

Thermostable α-amylases hold a very important place in commercial industrial applications in Sri Lanka. Therefore, the main aim of this study was to identify superior Bacillus strain and optimize growth conditions that could yield high α-amylase production. Three Bacillus strains, B. amyloliquefaciens ATCC 23350, B. licheniformis ATCC 14580 and B. megaterium ATCC 14581 were used for the study. Shake flask culture experiments were conducted to identify the effect of various fermentation conditions such as growth temperature, incubation period, carbon source, nitrogen source, initial pH and carbon concentration on extracellular α-amylase production. DNSA assay was carried out to determine the enzyme activity. The highest temperature for enzyme activity was reported by B. licheniformis at 85°C, followed by B. amyloliquefaciens at 75°C and B. megaterium at 45°C. Both B. amyloliquefaciens and B. licheniformis were able to give their optimum enzyme production at 37°C, while B. megaterium at 30°C in 150 rpm with initial pH of 7. B. licheniformis and B. amyloliquefaciens gave their optimum yield of the enzyme after 48 h of incubation while B. megaterium gave after 24 h of incubation. Among the carbon sources tested cassava starch was able to give the highest enzyme production. For B. amyloliquefaciens, the highest yield of the enzyme was obtained with 2% of starch, tryptone as a nitrogen source and initial pH of 7. Maximum enzyme production for B. licheniformis was obtained with 1.5% of starch, KNO3 as a nitrogen source and initial pH of 6. For B. megaterium 1% of starch, tryptone and pH 7.5 induced the optimum α-amylase production. According to the results obtained, B. amyloliquefaciens is the highest thermostable alpha amylase producer. However, according to the industrial requirement, B. licheniformis can also be used as an enzyme producer due to its stability in higher temperatures.

关键词： Thermostable α-amylase Bacillus Strain Fermentation Incubation Period

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

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Asian Pacific Journal of Tropical Biomedicine 2012年第9期2卷 733-738页

作者： G Oboh AJ Akinyemi AO Ademiluyi Department of Biochemistry Federal University of TechnologyAkure

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(Pα-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.

关键词： Blanching Antioxidants Vegetables Telfairia occidentalis α-amylase α-glucosidase

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Expression of α-amylase Gene of Bacillus licheniformis in Saccharomyces cerevisiae

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Science China Chemistry 1994年第11期37卷 1321-1328页

作者：罗进贤黎杨李文清张添元何鸣 Department of Biology and Biotechnology Research Center Zhongshan UniversityGuangzhou 510275PRC

The secretive expression vector has been constructed using the promoter and signal se-quence of yeast MF-α1 factor,and the Bacillus licheniformis α-amylase gene without promoter and signal se-quence has been inserted into the downstream of the signal sequence on the vector.After the readjustment ofthe reading frame,the amylase gene was expressed in Saccharomyces cerevisiae and the product was secretedfrom it.The properties of enzymes secreted from yeast and B.subtilis are compared,and the mechanism ofthe gene expression and product secretion are discussed.

关键词： Bacillus licheniformis α-amylase gene expression Saccharomyces cerevisiae

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Effect of static magnetic field on activity of immobilized α-amylase

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Chinese Science Bulletin 1997年第2期42卷 127-130页

作者： YAN Liushui, ZHU Yuanbao, HE-Shuang’e and CAO Zhixiang1 Department of Chemical Engineering, Nanchang Institute of Aeronautical Technology, Nanchang 330034, China 2. Department of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, China Department of Chemical Engineering Nanchang Institute of Aenwututical Techndogy Nanchang China Department of Chemistry and Chemical Engineering Hunan University Changsha China

BIOLOGICAL effect of magnetic field is one of the three major subjects of contemporary biomagnetism, and is also the basis of applied biomagnetism. In order to explore magnetic biological effects, the influence of various sorts of magnetic field on many living systems as well as their different constructural levels has been extensively studied. However, only a few studies with in vitro enzyme have been reported, and few researches concerning the influence of magnetic field on immobilized enzyme reactions have been published. This note describes

关键词： static magnetic field immobilized α-amylase enzyme activity oscillatory behaviour.

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

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Asian Pacific Journal of Tropical Biomedicine 2014年第6期4卷 473-479页

作者： Nizam Uddin Rakib Hasan Monir Hossain Arjyabrata Sarker A.H.M.Nazmul Hasan A.F.M.Mahmudul Islam Mohd.Motaher H.Chowdhury Sohel Rana Laboratory of Natural Products Research Department of PharmacyJahangirnagar University

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.

关键词： Diabetes mellitus Hypoglycemic Citrus macroptera α-amylase OGTT Glibenclamide

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