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ACE Kit-WST

Item # Unit Size
A502-10
100 tests

For Research Use Only Products


  • Colorimetric microplate assay
  • Simple protocol
  • No organic solvent required
  • High reproducibility

    Application: Angiotensin-converting enzyme (ACE) activity detection.
                          Screening of ACE inhibitors

MSDS

Contents of the Kit:

Enzyme A  :2 tubes    Enzyme B  :2 tubes 
Enzyme C  :2 tubes    Coenzyme  :2 tubes 
Substrate buffer :1 ml x 2    Indicator solution  :5 ml x 2 

Storage Condition: 0-5°C
Shipping Condition: ambient temperature
 
Required Equipment and Materials
plate reader with 450 nm filter; 96-well culture plate, 2-20 μl, 20-200 μl, 100-1000 μl and multi-channel pipettes; 37ºC incubator, Disposable syringe (1 ml)


Product Description
The kit is used for the determination of ACE inhibition activity. ACE works in the Renin-Angiotensin system, which is one of the mechanisms of blood pressure control, to convert Angiotensin I to the vasopressor Angiotensin II. This enzyme also contributes to elevated blood pressure due to its role in breaking down the antihypertensive peptide Bradykinin. In recent years, food and supplements containing ingredients that block ACE have received attention for their use in preventing high blood pressure. The conventional method of measuring ACE inhibition employs the synthetic substrate Hippuryl-His-Leu. Hippuric acid from the synthetic substrate is extracted with ethyl acetate, condensed, redissolved, and then read at an absorbance of 228 nm. This method is cumbersome and measurement is subjected to error due to residual ethyl acetate. ACE inhibition Assay Kit enzymatically detects 3-Hydroxybutyric acid (3HB), which is made from 3-Hydryoxybutyryl-Gly-Gly-Gly (3HB-GGG). Using a 96-well format, it is possible to test multiple samples at one time. In addition, there is no need to use harmful organic solvents, resulting in a safe, simple, and highly reproducible assay.


Fig. 1
Principle of the assay system to determine ACE activity or inhibition activity.


User References
Sample Reference
15 kinds of DrugsAndrás Málnási-Csizmadia et.al., Experimental Confirmation of New Drug–Target Interactions Predicted by Drug Profile Matching. J. Med. Chem.201356 (21), 8377.
Protein fractions from the mycelia of G. lucidumMohamad Ansor et al., Anti-angiotensin converting enzyme (ACE) proteins from mycelia of Ganoderma lucidum (Curtis) P. Karst. BMC Complementary and Alternative Medicine 2013, 13, 256
Rat PlasmaNakamuta H., et al., Antihypertensive effects of continuous oral administration of nattokinase and its fragments in spontaneously hypertensive rats. Biol Pharm Bull. 2011, 34, 1696-701.
ϵ-Viniferin (Resveratrol)Miyazaki H., et al., ε-Viniferin is more effective than its monomer resveratrol in improving the functions of vascular endothelial cells and the heart. Biosci Biotechnol Biochem. 2012;76:954-60.
Edible MushroomsNorhaniza Aminudin, et al., Proteomic Analysis of Antihypertensive Proteins in Edible Mushrooms. J. Agric. Food Chem., 2012, 60, 12341–12348
Recycling of Bovine Meat and Bone Meal and 
Inactivation of BSE Prion by Sub-Critical Water Treatment
Recycling of Bovine Meat and Bone Meal and 
Inactivation of BSE Prion by Sub-Critical Water Treatment
Extract from MeatM. Yoshida, et al., Recycling of Bovine Meat and Bone Meal and  Inactivation of BSE Prion by Sub-Critical Water Treatment
Mongolian milk products
Kohji Yamaki, et al., Inhibition of Angiotensin-Converting Enzyme by Components of Traditional  Mongolian Fermented Milk Products. Food Sci. Technol. Res., 2011;17:567–572

References
1. L. H. Lam, et al., Assay of angiotensin I-converting enzyme-inhibiting activity based on the detection of 3-hydroxybutyric acid. Anal Biochem. 2007;364:104-111.
2. L. H. Lam, et al., Assay of angiotensin I-converting enzyme-inhibiting activity based on the detection of 3-hydroxybutyrate with water-soluble tetrazolium salt. Anal Sci. 2008;24:1057-1060.
3. L. H. Lam, et al., Flow injection analysis of angiotensin I-converting enzyme inhibitory activity with enzymatic reactors. Talanta. 2009;79:1130-1134.

Mitsugu FUJITA,
*
,a
Katsunori OHNISHI,
b
Shinsaku TAKAOKA,
b
Kazuya OGASAWARA,
b
Ryo FUKUYAMA,
a
and Hiromichi NAKAMUTAa

Assay Data


Fig. 2 Inhibition curves prepared by Alacepril and Captopril.
IC50 of Alacepril and Captopril are 3.62 μM and 2.14 nM, respectively. Both compounds are ACE inhibitors.



Fig. 3 Inhibition curves prepared by two beverages containing a valyltyrosine () or lacto tripeptide ().
IC50 of these beverages are 0.56% and 0.69%, respectively. It is known that these substances have antihypertensive effects.
*Concentration of the beverage in the sample solution.