Detection method of angiotensin converting enzyme (ACE) inhibitory activity of antihypertensive peptide

An angiotensin and inhibitory activity technology, applied in measuring devices, instruments, scientific instruments, etc., can solve problems such as inability to detect ACE inhibitory activity, and achieve the effect of saving detection time and improving accuracy

Inactive Publication Date: 2012-07-25
THIRD INST OF OCEANOGRAPHY STATE OCEANIC ADMINISTATION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, some samples have weak absorption peaks or no absorption peaks in UV detection, and high performance liquid chromatography cannot detect the ACE inhibitory activity of these samples

Method used

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  • Detection method of angiotensin converting enzyme (ACE) inhibitory activity of antihypertensive peptide
  • Detection method of angiotensin converting enzyme (ACE) inhibitory activity of antihypertensive peptide
  • Detection method of angiotensin converting enzyme (ACE) inhibitory activity of antihypertensive peptide

Examples

Experimental program
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Effect test

example 1

[0027] 1. Dissolve a certain amount of fish scale antihypertensive peptide in pH 8.3, 0.1mol / L boric acid buffer, prepare a sample solution with a concentration of 1mg / ml, absorb 25μL of antihypertensive peptide, and add 75μL of 60mU·ml -1 The ACE was incubated at 37°C for 5min, and then 112μL of 5.0mmol / L HHL was added. In the blank control group, the volume of the antihypertensive peptide was replaced by the same volume of boric acid buffer solution, incubated in a constant temperature water bath at 37°C for 60min, and then 25μL of 0.1 %TFA terminated the reaction.

[0028] 2. LC-MS detection and analysis of reaction products, high performance liquid chromatography conditions: Ug120-C18 reversed-phase chromatographic column, mobile phase is acetonitrile: ultrapure water = 20:80 (containing 0.1% TFA, volume ratio), flow rate 1.0mL·min -1 , The detection wavelength is 228nm, and the injection volume is 10μL. The mass spectrometry conditions are: electrospray ionization posit...

example 2

[0031] 1. Dissolve a certain amount of peanut antihypertensive peptide in pH 8.3, 0.1mol / L boric acid buffer, prepare a sample solution with a concentration of 1mg / ml, absorb 25μL of antihypertensive peptide, and add 75μL of 60mU·ml -1 The ACE was incubated at 37°C for 5min, and then 112μL of 5.0mmol / L HHL was added. In the blank control group, the volume of the antihypertensive peptide was replaced by the same volume of boric acid buffer solution, incubated in a constant temperature water bath at 37°C for 60min, and then 25μL of 0.1 %TFA terminated the reaction.

[0032]2. LC-MS detection and analysis of reaction products, high performance liquid chromatography conditions: Ug120-C18 reversed-phase chromatographic column, mobile phase is acetonitrile: ultrapure water = 20:80 (containing 0.1% TFA) (volume ratio), flow rate 1.0mL·min -1 , The detection wavelength is 228nm, and the injection volume is 10μL. The mass spectrometry conditions are: electrospray ionization positive ...

example 3

[0035] 1、 Dissolve a certain amount of glycine-phenylalanine (Gly-Phe) in pH 8.3, 0.1mol / L boric acid buffer, prepare a sample solution with a concentration of 1mg / ml, absorb 25μL of antihypertensive peptide, add 75μL60mU·ml -1 The ACE was incubated at 37°C for 5min, and then 225μL of 5.0mmol / L HHL was added. In the blank control group, the same volume of boric acid buffer was used to replace the volume of the antihypertensive peptide, incubated in a constant temperature water bath at 37°C for 60min, and then 25μL of 0.1 %TFA terminated the reaction.

[0036] 2、 LC-MS detection and analysis of reaction products, high performance liquid chromatography conditions: Ug120-C18 reversed-phase chromatographic column, mobile phase is acetonitrile: ultrapure water = 20:80 (containing 0.1% TFA) (volume ratio), flow rate 1.0mL min -1 , The detection wavelength is 228nm, and the injection volume is 10μL. The mass spectrometry conditions are: electrospray ionization positive ion mode,...

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Abstract

A detection method of angiotensin converting enzyme (ACE) inhibitory activity of antihypertensive peptide comprises the following steps that: firstly, boric-acid buffer solution is prepared to perform an experiment on ACE activity inhibition of the antihypertensive peptide; secondly, reaction products are detected and analyzed by LC-MC (liquid chromatography-mass spectrometry), and the antihypertensive activity of the antihypertensive peptide is represented by detecting change of peak area of hippuric acid (Hip), wherein the purity of the Hip is ensured through monitoring of an HPLC (high performance liquid chromatography) ultraviolet detector and an electrospray ion source mass spectrum as well as with adoption of a total ion chromatogram and an ultraviolet detection chromatogram, and the peak area of the Hip is calculated by quantitatively analyzing the Hip peak area of an ion extract diagram of m / z180 in an SIM model; and thirdly, the peak area of the Hip of the reaction products in a reaction group and a no-treatment control group are obtained through the first and second steps, thereby obtaining the ACE inhibition ratio of a sample. In the method, the LC-MS technology is applied to screening activity of the antihypertensive peptide, so that the influence of impurities on activity detection is reduced; and the detection method has the characteristics of being high in precision, good in repeatability and high in sensitivity.

Description

technical field [0001] The invention relates to a method for detecting the inhibitory activity of angiotensin-converting enzyme (ACE) of antihypertensive peptide, which belongs to the field of biotechnology. Background technique [0002] Hypertension is a serious threat to human health. There are 180 million hypertensive patients in my country, and more than 1 million people die from hypertension every year. The body's pressor system, the renin-angiotensin system and the depressor system, the kallikrein-kinin system, play an important role in blood pressure regulation and cardiovascular function. ACE (angiotensin-converting enzyme) achieves the effect of raising blood pressure through the regulation of the above two systems. Antihypertensive peptides play a role in lowering blood pressure by inhibiting the activity of ACE. [0003] Antihypertensive peptides have the characteristics of high safety, no side effects, and obvious effects. They have become the research focus of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02
Inventor 陈俊德易瑞灶刘颖谢全灵徐建陈思谨陈晖张怡评孙继鹏郑美华许诺华余琳朱伟翔
Owner THIRD INST OF OCEANOGRAPHY STATE OCEANIC ADMINISTATION
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