amylase activity by dns method protocol

Epub 2015 Feb 16. One negative control (absence of inhibitor) was set up to obtain 100% enzyme activity. The test is done in microtitre plates with a total volume of 260L and an assay time of 40min including the pre-incubation steps. The enzyme concentration, incubation time, interaction between enzyme concentration and preincubation time, and the interaction between preincubation and boiling time showed a significant effect on the inhibition percentage. Pharmacol. 1 ml of D.N.S. The new PMC design is here! This evaluation revealed that minor modifications/changes in the assay conditions can lead to a significant variation in IC50 values (0.00068 to 47.5 mg mL-1) with acarbose used as a positive control. Master degree dissertation. Amylase level testing can be done with a blood or urine test. J Genet Eng Biotechnol 14:133141, CrossRef Five plant extracts were used to determine -amylase inhibitory activity using the proposed method. 1959, 31, 426. The absorbance of the product was recorded at 540 nm. In this experiment, acarbose at a concentration of 1.05 g mL-1 was used to record the desired maximum signal. Another potential application of the proposed method is the evaluation of the activity of amylolytic enzymes extracted from different natural matrices such as flours and microbial enzyme preparations. 16 United States Pharmacopeial Convention; The United States pharmacopeia: the national formulary; Rockville, Md., United States pharmacopeial convention, 1980. However, it showed a relatively higher -amylase activity in Fig 3 d. In additio n, Fig 3 d needs control to show equal amounts of proteins were loaded. The inhibition percentage was calculated. [Crossref], 4 Rahimzadeh, M.; Jahanshahi, S.; Moein, S.; Moein, M. R.; Iran. Amylase is an enzyme, or special protein, produced by your pancreas and salivary glands. Preparation of soluble starch standards for quantification of amylolytic enzyme activity. The absorbance of the well was monitored at 505nm, and the value recorded was 0.681. Simultaneously setup the color developed at 520nm. Then make it to 200ml with distilled water. The absorbance at 580nm was measured for both assay and control experiments for 10min, being the average values of 0.110 and 1.138, respectively. Most enzymes act specifically with only one reactant, called a substrate, to produce products. All experimental details are described in the section Amylolytic enzyme activity assay. government site. The monitoring of the activity of alpha-amylase and amyloglucosidase is an important tool for studying their role in the hydrolysis of starch. This study optimizes and validates microscale in vitro methods for the assessment of -amylase inhibition. Hugo M. Oliveira and Margarida R. G. Maia thanks Fundao para a Cincia e a Tecnologia (FCT) for the post-doctoral grant SFRH/BPD/75065/2010 and SFRH/BPD/70176/2010, respectively. (function(d){var s=d.createElement("script");s.setAttribute("data-account","s8XDZXWqQY");s.setAttribute("src","https://cdn.userway.org/widget.js");(d.body||d.head).appendChild(s);})(document)Please ensure Javascript is enabled for purposes of website accessibility. [Crossref]-1616 United States Pharmacopeial Convention; The United States pharmacopeia: the national formulary; Rockville, Md., United States pharmacopeial convention, 1980.,2727 Iversen, P.; Beck, B.; Chen, Y.; Dere, W.; Devanarayan, V.; Eastwood, B. J.; Farmen, M. W.; Iturria, S. J.; Montrose, C.; Moore, R. A.; Weidner, J. R.; Sittampalam, G. S. In HTS Assay Validation; Markossian, S.; Grossman, A.; Brimacombe, K.; Arkin, M.; Auld, D.; Austin, C. P.; Baell, J.; Chung, T. D. Y.; Coussens, N. P.; Dahlin, J. L.; Devanarayan, V.; Foley, T. L.; Glicksman, M.; Hall, M. D.; Haas, J. V.; Hoare, S. R. J.; Inglese, J.; Iversen, P. W.; Kales, S. C.; Lal-Nag, M.; Li, Z.; McGee, J.; McManus, O.; Riss, T.; Saradjian, P.; Sittampalam, G. S.; Tarselli, M.; Trask, O. J.; Wang, Y.; Weidner, J. R.; Wildey, M. J.; Wilson, K.; Xia, M.; Xu, X., eds. The most commonly used method to measure -amylase in wheat grain is the Hagburg Falling Number method but the values are also influenced by rheological properties of starch in the grain.1,2 The most specific method for the measurement of -amylase is the Ceralpha method which employs a defined substrate, 4,6-O-benzylidene--4-nitrophenyl- Add 20 ml of 2 N NaOH. J. Mol. The reagent has a dark orange colour and is stable for several days in room temperature. Amylase activity from partially purified samples was assayed by DNS method. You can decide for yourself whether you want to allow cookies or not. In the present method of Millers reaction, the product to be analyzed was obtained by heating it in a boiling water bath. System Appl Microbiol 41:516527, Shaik M, Sankar GG, Iswarya M, Rajitha P (2017) Isolation and characterization of bioactive metabolites producing marine Streptomyces parvulus strain sankarensis-A10. If not, the shelf-life of the reagent will be 2 weeks if preserved at 4C. Liu F, Zhu S, Ni L, Huang L, Wang K, Zhou Y. Endocrine. Determination of amylose/amylopectin ratio of starches. The pancreas is an organ located behind your stomach. LCF-TSN8valql#uy*|;IUV8,\d&sQ|`Pg&[#-&Jqb4!8RyQ31IlkfzJs! This discrepancy highlights the reason for not comparing the results obtained by different reports, although they might have used the same method but with different experimental conditions. J Genet Eng Biotechnol 15:8794, Zhao Y, Zhao Y, Zhang Z, Wei Y, Wang H, Lu Q, Li Y, Wei Z (2017) Effect of thermo-tolerant actinomycetes inoculation on cellulose degradation and the formation of humic substances during composting. Absorbance was recorded using a microplate reader (Multiskan FC, Thermo Scientific). 12 Stevenson, R. A.; American Laboratory, 2011, available at https://www.americanlaboratory.com/913-Technical-Articles/19109-Bioassays-2011/, accessed June 2022. Note: amylolytic enzyme working solutions should be prepared fresh daily and kept on ice at all times. (+)-Catechin hydrate (PubChem CID: 9064); 3,5-Dinitrosalicylic acid (PubChem CID: 11873); Amylase inhibitors; Anhydrous dextrose (PubChem CID: 5793); DNSA; Gallic acid (PubChem CID: 370); Glucose oxidase; Maltose; Maltose monohydrate (PubChem CID: 16211336); Potassium sodium tartrate tetrahydrate (PubChem CID: 2724148); Rutin hydrate (PubChem CID: 45479757); Sodium phosphate dibasic (PubChem CID: 24203); Sodium phosphate monobasic (PubChem CID: 23672064); Starch soluble (PubChem CID: 439341); -Amylase. [Crossref]. The reduced 3-amino-5-nitrosalicylic acid is red-brown in color and can be quantified spectrophotometrically with an absorption maximum at 540 nm. HCl 1.0molL1: add 8.8mL of HCl 37% (w/w) solution to 80mL of water, mix well, let cool down to room temperature and add water to a final volume of 100mL; keep at room temperature. Thank you Marsha for your replay, actually I am trying to calculate IC50 or EC50 from my samples (plant extracts) to see if it can use as inhibitor FOIA Furthermore, by performing the assay on a volume scale compatible with eppendorfs or vials, it is also possible to easily transfer the sample other subsequent experimental steps (e.g. 2017, Article ID 7319504. Pharmacol. Another import aspect was to maximize the information about enzyme role in the hydrolysis process, such as the monitoring of the release of glucose monomers from pure starches that can be obtained when the hydrolysis is catalyzed by amyloglucosidase. 2011, 136, 305. [1] Amylase was first described in the early 1800s and is considered one of the first enzymes in history to be scientifically investigated. For enzyme inhibition studies, the absorbance was recorded at a higher or maximum concentration of inhibitor (IC70). 8. Enzyme energy of amylase from edible fungi was determinated based on 3,5-dinitryl-salicyle(DNS).Taking czapek as induction medtum in whith the only carbon source was 2% soluble starch,and amylase energy ranged from 1.513 to 3.417 U/mL among Lentinula edodes,Pleurotus ostreatus,Pleurotus cornucopiae,so as to put forward a reference and analysis method for the edible fungistrain selection. 7 0 obj Phytochemical constituent analysis from the Zanthoxylum fagara and Chrysactinia mexicana plants using different solvents for extraction can be performed. Unfortunately, to date, many published studies cannot be reproduced; most of them are nonreplicable or simply erroneous. Saliva can be collected using a calibrated collection device, and the resulting sample should ideally be analyzed within 15 minutes after collection. Different experiments were performed to evaluate the robustness of the method. Chem. 3,5-Dinitrosalicylic acid ( DNS or DNSA, IUPAC name 2-hydroxy-3,5-dinitrobenzoic acid) is an aromatic compound that reacts with reducing sugars and other reducing molecules to form 3-amino-5-nitrosalicylic acid, which strongly absorbs light at 540 nm. Add 2.91mL of 85% (w/w) o-phosphoric acid, 2.86mL of acetic acid, and dissolve 3.09g of boric acid in water and complete to a final volume of 1000mL; keep at 4C. The most remarkable characteristic is that. One milligram of each extract was weighed and dissolved in 1 mL of phosphate buffer (100 mmol L-1, pH 6.9); some solutions were sonicated to achieve complete dissolution. PMC 29 Zhang, J. H.; Chung, T. D.; Oldenburg, K. R.; J. Biomol. The vial was then placed for 10min into a heating block at 45C. 2012, 60, 8924. Alpha-amylase is found in saliva and is responsible for the initial breakdown of starch into maltose. <>stream MTCC 5214 when grown in solid-state fermentation. and a-amylase activity. [Crossref]. So the final concentration is 0.1M of 200ml Phosphate buffer. Amylase is also used in brewing and baking, as it helps to convert starch into sugar. -Amylase from the porcine pancreas (15 U mg-1) was used instead of human amylase because of its cost and availability. 8600 Rockville Pike Plant extracts contain various phytochemicals, such as phenols, saponins, and alkaloids, that contribute to many observed pharmacologically relevant effects. 2014, 17, 465. The positive control used was acarbose (0.6 g mL-1). Like most things, amylase production slows as we age, which reduces the ability of the digestive system to properly absorb nutrients. 2011, 47, 270. [Crossref], 31 Quan, N. V.; Anh, H.; Lam, V. Q.; Takami, A.; Teschke, R.; Khanh, T. D.; Xuan, T. D.; Molecules 2022, 27, 774. In these experimental conditions, an illustrative calibration curve is abs505nm=0.042 (0.005)+0.0846 (0.0008) mglucose. An example of the application of this method was the determination of the glucose yield after the hydrolysis of four pure starches assisted by an amyloglucosidase from A. niger (Fig. 15 Markossian, S.; Coussens, N. P.: Dahlin, J. L.; Sittampalam, G. S.; SLAS Discovery 2021, 26, 1241. Block heater (Stuart SBH130D/3, Staffordshire, UK), for conducting the activity assay with the 4mL vials. 1.5 X 4 mg of Maltose formed / 0.3 ml of Enzyme / 1 hour, 1.5 X 4 X 3.3 mg of Maltose formed / 1ml of Enzyme / 1 hour, 1.5 X 4 X 3.3 X 100 mg of Maltose formed / 100ml of Enzyme / 1 hour. For measuring the amylase activity with DNS solution (Miller Method): 1. Salivary amylase is essential for the digestion of carbohydrates. This method is a redox reaction where DNS (yellow color) is reduced by reducing sugars to 3-amino-5-nitrosalicylic acid (red color) in an alkaline medium. [Crossref], 23 Ha, K. S.; Jo, S. H.; Kang, B. H.; Apostolidis, E.; Lee, M. S.; Jang, H. D.; Kwon, Y. I.; J. This method is a redox reaction where DNS (yellow color) is reduced by reducing sugars to 3-amino-5-nitrosalicylic acid (red color) in an alkaline medium. [Crossref], Stay informed of issues for this journal through your RSS reader, Text Warm the assay buffer to room temperature before use. Then, we have partially purified these amylases employing three conventional methods: (a) cold acetone precipitation, (b) ion-exchange chromatography, and (c) gel filtration chromatography. Dissolve 200.0mg of KI in 60mL of water, after complete dissolution, add 20.0mg of I2 and complete with water up to a total volume of 100mL; keep at room temperature protected from light. 2012, 60, 8924. Transfer 50L of sample supernatant (or standard), and add 150L of GOPOD reagent to each well of the microplate. Humana, New York, NY. To evaluate linearity, amylase inhibition studies were performed using different concentrations of the inhibitor acarbose (0.5 to 12 g mL-1). A uniformity assay is performed at maximum and minimum response levels, ensuring that the difference is adequate to detect active compounds or the influence of the inhibitor in the assay system. Testing. 26 Tundis, R.; Loizzo, M. R.; Statti, G. A.; Menichini, F.; Pharmazie 2007, 62, 473. Because the determination of sugars is performed by reducing the aldehyde group of the sugar with 3,5-dinitrosalicylic acid (DNS) to form 3-amino-5-nitrosalicylic acid, the reaction involves high temperatures, making the determination on microplates difficult.99 Miller, G. L.; Anal. (1) the calculated activity for this enzyme was 1123UmL1. See, The calibration curve for glucose was linear for masses of glucose up to 10g (equivalent to concentrations up to 200mgL, Determination of the alpha-amylase and amyloglucosidase activities, Enzyme activity, Starch-iodine, Glucose oxidase/peroxidase, Glucose yield. Before 2019 Mar 1;275:265-272. doi: 10.1016/j.foodchem.2018.09.114. This article will discuss the different applications of amylase. 6 This method, like the iodine procedure only measures ~-amylase activity and the results cannot be readily expressed in international units. Pol. 15 Markossian, S.; Coussens, N. P.: Dahlin, J. L.; Sittampalam, G. S.; SLAS Discovery 2021, 26, 1241. In that case, the sensitivity of the determination will be modified. Chang YX, Ge AH, Donnapee S, Li J, Bai Y, Liu J, He J, Yang X, Song LJ, Zhang BL, Gao XM. Amylase is naturally produced in humans by the salivary glands and pancreas. Likewise, actinobacteria have been shown to be excellent sources of enzyme production and, therefore, can be considered as good alternatives to the use of fungi to obtain enzymes of industrial interest. Evaluation of precision and accuracy was performed using the extracted sample of J. dioica in the presence or absence of acarbose (positive control). Starch is a widespread raw material with several application fields that cover food (including confectionery and drinks), feed, pharma, chemical, and paper industries [1]. The extracts were prepared using different solvents: methanol for aerial parts of the plant and a mixture of ethanol and water (90:10) for roots. KI/I2 solution: KI 2.0g L1, I2 200mgL1. As a unit of activity (unit, U) of the enzyme a-amylase, is arbitrarily appointed, the quantity of the enzyme required for the production of 1 mole of maltose in 1 min, when the enzyme is incubated along with the substrate at pH=4,9 and =40 C. Bookshelf The precision of the method was analyzed on the day of experimentation and on two different days. We adopted this strategy to separate the solid particles that remained after the hydrolysis of different pure starches mediated by amyloglucosidase (glucose yield protocol, step 5). Hence, the dilution factor corresponded to 2000. The proposed method relies on principles that allow a fast, simple, and versatile measurement of the activity of alpha-amylase or amyloglucosidase. [Crossref] Although other protocols are based either on the glucose oxidase reaction or on the determination by fluorescence, they are commonly used in microplates. Protocol Preparing Saliva 1. When compared with the original method, these new conditions lead to an extra dilution of the sample (a dilution factor of 5 compared to a dilution factor of 2) and to a longer optical path due to the higher volume placed at each well of the microplate (250L vs. 150L, and assuming the use of a classic cylindrical-shaped flat bottom microplate). Changes in conditions/parameters, such as enzyme concentrations (< 0.15 U mL 11 Bhat, M.; Zinjarde, S. S.; Bhargava, S. Y.; Kumar, A. R.; Joshi, B. N.; J. Evidence-Based Complementary Altern. [Crossref]. The degree of inhibition was evaluated in terms of the concentration of the inhibitor that inhibited half of the enzyme activity (IC50). Some more thoughts: Try boiling the DNS solution longer than 5 min. Even though the water bath is at 100C, the internal solution temperature may no The proposed optimized and validated method was applied to extracts of five plants. Sodium azide (1.0g) can be added as a preservative (antimicrobial agent). The % RSD was calculated for each solution. When an analytical method is considered, the matrix effect results in variation in the sensitivity, which is manifested by comparing the calibration curve of the standard (positive control) with that of the sample (plant extract). Learn more, Banerjee S, Maiti TK, Roy RN (2016) Identification and product optimization of amylolytic Rhodococcus opacus GAA 31.1 isolated from gut of Gryllotalpa africana. https://www.americanlaboratory.com/913-T https://www.fda.gov/files/drugs/publishe https://www.americanlaboratory.com/913-Technical-Articles/19109-Bioassays-2011/, https://www.fda.gov/files/drugs/published/Bioanalytical-Method-Validation-Guidance-for-Industry.pdf. Federal government websites often end in .gov or .mil. Biotechnol. Chem. By clicking accept or continuing to use the site, you agree to the terms outlined in our. Diabetes mellitus is a chronic metabolic disorder characterized by hyperglycemia (high blood sugar levels) due to insulin resistance or absolute lack of insulin production. Factors significantly contributing to the response were selected by the Pareto graph. We use cookies on our website. 1959, 31, 426. 5 ml of deionized water are added in each sample, followed by agitation. In a tube containing 1 mL of phosphate buffer (100 mmol L-1, pH 6.9), 100 L of -amylase (final concentration 0.15 U mL-1 in phosphate buffer) was added and incubated at 37 C for 7.2 min for preincubation. <>/ExtGState<>/XObject<>/ProcSet[/PDF/Text/ImageB/ImageC/ImageI] >>/MediaBox[ 0 0 595.32 841.92] /Contents 4 0 R/Group<>/Tabs/S/StructParents 0>> Springer Nature is developing a new tool to find and evaluate Protocols. The -amylase assay was performed using Millers method, i.e., the DNS method. Starch, acarbose, 3,5-dinitrosalicylic acid (DNS), sodium chloride, dibasic sodium phosphate, monobasic sodium phosphate, sodium, and potassium tartrate were procured from Sigma-Aldrich, USA. It was initially termed diastaste but was later renamed amylase in the early 20th Factors showing a significant effect on the inhibition percentage were selected and optimized using a basic sequential simplex method. Please note that if you reject them, you may not be able to use all the functionalities of the site. These inhibitors delay the rate of absorption of glucose, thereby maintaining glucose levels in hyperglycemic individuals. For calculation of enzyme activity base on (U/ml) try according to the following stages: 1. [Crossref], 20 Hou, W.; Li, Y.; Zhang, Q.; Wei, X.; Peng, A.; Chen, L.; Wei, Y.; Phytother. These bonds can be easily broken by water, so saliva is necessary for the enzyme to work. Your email address will not be published. 2017, Article ID 7319504. ; Humana Press: New York, 2016 [Crossref] Bioassays are challenging in terms of determining accuracy. [Link]5 Valentina, P.; Ilango, K.; Chander, S.; Murugesan, S.; Bioorg. [Crossref], 18 Apostolidis, E.; Lee, C. M.; J. Amylase is a digestive enzyme predominantly secreted by the pancreas and salivary glands and found in other tissues at very small levels. 2020-04-29T17:28:43+05:302017-05-24T07:28:08ZMicrosoft Office Word 2007 abs505nm=0.042 (0.005)+0.0846 (0.0008) mglucose), we obtained a mass of glucose of 7.56g that is contained in the 50L of sample transferred to the microplate. Different parameters, such as enzyme concentration, substrate concentration (starch), preincubation time, incubation time, incubation temperature, and time in the water bath, were evaluated. will also be available for a limited time. [Crossref] Additionally, it is necessary to validate the assay method to demonstrate that it is acceptable for determining the pharmacological activity of a new chemical entity by a detectable signal that allows the biological process to be quantified. Three different concentrations (1, 2.5, and 5 g mL 11 Bhat, M.; Zinjarde, S. S.; Bhargava, S. Y.; Kumar, A. R.; Joshi, B. N.; J. Evidence-Based Complementary Altern. [Link], 5 Valentina, P.; Ilango, K.; Chander, S.; Murugesan, S.; Bioorg. Those with normal plasma glucose had group I, those with impaired glucose tolerance had group II, and those who were diabetic had group III. Transfer 1500L of water and 1500L of universal buffer (control) or of amyloglucosidase solution (assay) into a 4mL amber glass vial. This was followed for a second dilution of 500L of the previous solution with 4500L of universal buffer at the desired pH (enzyme solution 2, dilution factor 10). By intercepting this absorbance values in the calibration curve (e.g. 27 Iversen, P.; Beck, B.; Chen, Y.; Dere, W.; Devanarayan, V.; Eastwood, B. J.; Farmen, M. W.; Iturria, S. J.; Montrose, C.; Moore, R. A.; Weidner, J. R.; Sittampalam, G. S. In HTS Assay Validation; Markossian, S.; Grossman, A.; Brimacombe, K.; Arkin, M.; Auld, D.; Austin, C. P.; Baell, J.; Chung, T. D. Y.; Coussens, N. P.; Dahlin, J. L.; Devanarayan, V.; Foley, T. L.; Glicksman, M.; Hall, M. D.; Haas, J. V.; Hoare, S. R. J.; Inglese, J.; Iversen, P. W.; Kales, S. C.; Lal-Nag, M.; Li, Z.; McGee, J.; McManus, O.; Riss, T.; Saradjian, P.; Sittampalam, G. S.; Tarselli, M.; Trask, O. J.; Wang, Y.; Weidner, J. R.; Wildey, M. J.; Wilson, K.; Xia, M.; Xu, X., eds. Amber glass vials (15mm45mm8mm, e.g. Glucose yield (average values from two determinations) measured by the described glucose oxidase/peroxidase assay that resulted from the hydrolysis of four different pure starches mediated by an amyloglucosidase from Aspergillus niger (PN: A9913, Sigma Aldrich). [. Additionally, analytical validation of the method it requires using appropriate positive and negative controls. Simple Protocol to Facilitate Students Understanding of the Effects of Enzyme Immobilization on Kinetics of Reaction and Mass Transfer. For the calculation of the glucose yield assay promoted by amyloglucosidase from Aspergillus niger (PN: A9913, Sigma Aldrich), 1500L of water were added to a vial containing of 3.00mg of a pure rice starch (mstarch) weighed in an analytical scale. Would you like email updates of new search results? Their age ranged between 25 65 years. For enzymes, denaturation can be defined as the loss of enough structure, rendering the enzyme inactive. Xiao et al. [Crossref], 24 Wang, Y.; Huang, S.; Shao, S.; Qian, L.; Xu, P.; Ind. 2000, 40, 133. Amylase - blood. For the GOPOD method under microplate format, we also performed a similar method characterization. 351 (2006) 146148]. where SD is the standard deviation of each measurement, and AVG is the average of such measurements. Note: pH value must be adjusted to the desired value with NaOH 4.0molL1 for each particular assay at room temperature. phenol). Based upon the plasma glucose value, all subjects were divided into three groups. The proposed method showed high precision in long-term use (RSD<6.3%). [Crossref], 30 Rahman, M. M.; Islam, M. R., Shohag, S.; Hossain, M. E.; Rahaman, M. S.; Islam, F.; Ahmed, M.; Mitra, S.; Khandaker, M. U.; Idris, A. M.; Chidambaram, K.; Emran, T. B.; Cavalu, S.; Molecules 2022, 27, 1713. Rev. Because crude extract contains a variety of bioactive metabolites, it is necessary to characterize the bioactive compound showing inhibition activity by purifying it to determine its IC50 value. Statistical analysis and validation were performed using Microsoft Excel 2010. We used a US bath to facilitate the complete solubilization of I2 and subsequent formation of triiodide (please note that the concentration of KI described in the original report of Xiao et al. 2010, 75, H97. Currently, there are different protocols and guidelines to perform the process of bioassay validation, such as Food and Drug Administration, USA (2018), Eli Lilly Guide (2007), and the 1033 Chapter of United States Pharmacopeia (1980).1414 Food and Drug Administration (USA); Bioanalytical method validation guidance for industry, US Department of Health and Human Services Food and Drug Administration Center for Drug Evaluation and Research and Center for Veterinary Medicine, 2018, available at https://www.fda.gov/files/drugs/published/Bioanalytical-Method-Validation-Guidance-for-Industry.pdf, accessed June 2022.https://www.fda.gov/files/drugs/publishe 50mL of concentrated solution for a final volume of 1000mL) in order to be used in the extraction protocol. Beta amylase is located in the pancreas and produces maltose from the hydrolysis of the maltose molecule. Home - Protocols - Quantitative Analysis - Assay of Salivary Amylase enzyme activity. Additionally, the significant factors were optimized using the rules of the basic sequential simplex method and the combination of conditions that maximized the inhibition percentage with the lowest photometric error. Nalin T., Sperb-Ludwig F., Venema K., Derks T.G.J., Schwartz I.V.D. If an amylase test finds too much amylase in your blood or urine, it may indicate a pancreas disorder or other health condition. Amylase is the hydrolytic enzyme that breaks down many polysaccharides like Starch, Amylose, dextrins, and yields a disaccharide i.e., Maltose. This study aimed at evaluating the efficacy of salivary amylase enzyme activity assay to detect diabetes mellitus. A total of 64 experiments were performed. Centrifuge (Astor 8, Astori Tecnica, Poncarale, Italy), for separating the solid particles in the glucose yield assay. official website and that any information you provide is encrypted In this study, the IC50 was determined following the method described by Tundis et al., and an IC50 value of 0.6 g mL-1 was obtained.2626 Tundis, R.; Loizzo, M. R.; Statti, G. A.; Menichini, F.; Pharmazie 2007, 62, 473. There are three basic types of amylase: alpha, beta, and gamma. Biocatal Agric Biotechnol 4:685693, Arumugam T, Kumar PS, Kameshwar R, Prapanchana K (2017) Screening of novel actinobacteria and characterization of the potential isolates from mangrove sediment of south coastal India. Evolution of sensitivity (represented by the slope of the calibration curve) along time for the glucose oxidase/peroxidase assay. The final aspect of the microplate, absorbance spectra, and illustrative calibration curve of different soluble starch standards (from left to right, respectively) of the starch-iodine quantitative assay used for the characterization of alpha-amylase and amyloglucosidase activities.

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