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Detection method of alpha-amylase inhibitor activity

A technology for amylase inhibitors and detection methods, which is applied in the field of determination of enzymes, can solve the problems of high purity requirements of α-amylase inhibitors, inaccurate detection results, low average recovery rate, etc., and achieve shortened binding time, Easy operation and high recovery rate

Inactive Publication Date: 2018-03-16
WEIHAI BAIHE BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The DNS colorimetric method calculates the activity of α-AI by detecting the content of reducing sugars. The defect of this method is that the reducing sugars in the reaction are not necessarily the reducing sugars that are hydrolyzed from starch. The detection of α-AI activity in samples with reducing sugars is inaccurate, and it is only suitable for pure α-amylase inhibitors. A purification step for α-amylase inhibitors is required before the detection, so this This method is complicated to operate and requires high purity of the α-amylase inhibitor, so it will cause disadvantages such as high cost and long time-consuming for the enterprise
The iodine colorimetric method can directly detect the activity of α-amylase inhibitor in the sample, and does not need to purify the α-amylase inhibitor. The operation is simple, the detection time is shortened, and the cost is greatly reduced. The requirements for the various conditions are strict, if the parameters of the reaction conditions are unreasonable, it will cause errors in the test results
The average recovery rate of the optimized parameter method given in the existing literature is low (99.03%), so it is necessary to optimize the parameters of each reaction condition of this method to make the detection results more accurate and increase the average recovery rate

Method used

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  • Detection method of alpha-amylase inhibitor activity
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  • Detection method of alpha-amylase inhibitor activity

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] A method for detecting the activity of an α-amylase inhibitor, which involves the following steps:

[0073] (I) establishment of starch standard curve;

[0074] (II) Preparation of amylase inhibitor test solution: accurately weigh 0.1 g of white kidney bean powder (containing α-amylase inhibitor) produced by Hanzhong Natural Valley into a volumetric flask, add water to 100ml, place for 30 minutes, fully Get the supernatant after dissolving as need testing solution;

[0075] (III) Preparation of α-amylase solution: Preparation of α-amylase solution: Accurately weigh porcine pancreatic amylase and add distilled water to make a 0.15 mg / ml solution, and shake well;

[0076] (IV) Starch hydrolysis test under the inhibitory condition of the test product: Precisely measure 5 mL each of the test product solution in step II and the porcine pancreatic amylase solution in step III, and preheat them in a water bath at 34.8°C for 8 minutes, mix and Continue to incubate for 5 minut...

Embodiment 2

[0090] A method for detecting the activity of an α-amylase inhibitor, which involves the following steps:

[0091] (I) establishment of starch standard curve;

[0092] (II) Preparation of amylase inhibitor test solution: accurately weigh 0.1 g of white kidney bean powder (containing α-amylase inhibitor) produced by Tianbaohua in Yunnan into a volumetric flask, add water to 100ml, place for 30 minutes, and fully Get the supernatant after dissolving as need testing solution;

[0093] (III) Preparation of α-amylase solution: Preparation of α-amylase solution: Accurately weigh porcine pancreatic amylase and add distilled water to make a 0.15 mg / ml solution, and shake well;

[0094] (IV) Starch hydrolysis test under the inhibitory condition of the test product: Precisely measure 5 mL each of the test product solution in step II and the porcine pancreatic amylase solution in step III, and preheat them in a water bath at 35°C for 8 minutes, mix and Continue to incubate for 8 minute...

Embodiment 3

[0108] A method for detecting the activity of an α-amylase inhibitor, which involves the following steps:

[0109] (I) establishment of starch standard curve;

[0110] (II) Prepare the test solution of amylase inhibitor: accurately weigh 0.1g of the control card white kidney bean solid beverage (containing α-amylase inhibitor) into a volumetric flask, add water to 100ml, place it for 30min, and dissolve it fully Get supernatant as need testing solution;

[0111] (III) Preparation of α-amylase solution: Accurately weigh porcine pancreatic amylase and add distilled water to make a 0.15 mg / ml solution, and shake well;

[0112] (IV) Starch hydrolysis test under the inhibitory condition of the test product: Precisely measure 5 mL each of the test product solution in step II and the porcine pancreatic amylase solution in step III, and preheat them in a water bath at 35°C for 8 minutes, mix and Continue to incubate for 10 minutes, add the soluble starch solution of step I and the p...

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Abstract

The invention discloses a detection method of alpha-amylase inhibitor activity, which includes steps of (1), building a standard curve; (II), preparing amylase inhibitor test sample solution; (III), preparing amylase solution; (IV), preheating test sample solution and pig pancreas amylase solution in a water bath; mixing and preserving temperature; adding soluble starch solution to react; adding mixed solution of iodine and potassium iodide, measuring the absorption value A2 under 660nm; (V), preheating the pig pancreas amylase solution in the water bath, adding soluble starch solution and phosphate buffer; adding distilled water to reach the same volume in step IV; after reaction, dropwise adding diluted hydrochloric acid; adding the mixed solution of iodine and potassium iodide; measuring the absorption value A1 under 660nm; (VI), calculating the activity. The detection method is convenient to operate, exact in detection, high in efficiency, wide in application range, and high in recycle rate; the detection method is applicable to detect healthcare food, drugs and alpha-amylase inhibitor activity in the raw material.

Description

technical field [0001] The invention relates to an assay method comprising enzymes, in particular to an assay method for the activity of α-amylase inhibitors. Background technique [0002] α-amylase inhibitor (α-amylase inhibitor, α-AI for short) is an inhibitor that inhibits glycoside hydrolase α-amylase. α-AI can inhibit the activity of saliva and pancreatic amylase in the gastrointestinal tract, hinder or delay the body's hydrolysis and digestion of the main carbohydrates in food, reduce the decomposition and absorption of starch carbohydrates in food, and reduce blood sugar, blood lipids and Inhibit the increase of blood sugar concentration, which is beneficial to the dietary treatment of diabetic patients. For obese patients, it can reduce the conversion of sugar to fat, delay intestinal emptying, increase fat consumption to reduce body weight, and α-amylase inhibitors have been used as weight loss health food in foreign countries. Therefore, α-AI can be used to preve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/31
CPCG01N21/31
Inventor 张伟丛芳王丽娜
Owner WEIHAI BAIHE BIOTECH