Hippuryl-Arg-OH |
Catalog No.GA22584
One-Click Copy Product Info
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Hippuryl-Arg-OH is a fluorescent signal substrate used in enzyme activity assays for the identification and cleavage of peptide bonds containing arginine.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 744-46-7
Sample solution is provided at 25 µL, 10mM.
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Hippuryl-Arg-OH is a fluorescent signal substrate used in enzyme activity assays for the identification and cleavage of peptide bonds containing arginine. Hippuryl-Arg-OH can be used for enzyme activity studies such as carboxypeptidase B and aminopeptidase, as well as related research in the field of drug development[1-4].
References:
[1] Liu YH, Konermann L. Enzyme conformational dynamics during catalysis and in the 'resting state' monitored by hydrogen/deuterium exchange mass spectrometry. FEBS Lett. 2006 Oct 2;580(22):5137-42.
[2] Fricker LD, Devi L. Comparison of a spectrophotometric, a fluorometric, and a novel radiometric assay for carboxypeptidase E (EC 3.4.17.10) and other carboxypeptidase B-like enzymes. Anal Biochem. 1990 Jan;184(1):21-7.
[3] Song L, Fricker LD. Purification and characterization of carboxypeptidase D, a novel carboxypeptidase E-like enzyme, from bovine pituitary. J Biol Chem. 1995 Oct 20;270(42):25007-13.
[4] Casarini DE, Alves KB, Araújo MS, et al. Endopeptidase and carboxypeptidase activities in human urine which hydrolyze bradykinin. Braz J Med Biol Res. 1992;25(3):219-29.
Determination of TAFIa Activity in Plasma by the Hippuryl-Arg-OH Method[1]
This protocol is adapted from research data and provided for reference only. Adjustments may be necessary based on specific experimental requirements.
1. Reagent Preparation: Prepare a 20mM working solution of the Hippuryl-Arg substrate. Prepare working solutions of Thrombin at 5U/mL and Thrombomodulin at 7.5U/mL, then mix them to form a thrombin-thrombomodulin complex. Prepare a 60mM CaCl₂ solution. Prepare an activation buffer (20mM HEPES, 0.15M NaCl, 5mM CaCl₂, pH=7.4). Prepare a 1M HCl solution to stop the reaction. Prepare a 1M NaOH solution. Prepare a 1M phosphate buffer. Prepare a 3% cyanuric chloride solution in 1,4-dioxane.
2. TAFI Activation: Take 15μL of plasma sample and mix it with 12μL of the thrombin-thrombomodulin complex, 8.5μL of 60mM CaCl₂ solution, and 24μL of activation buffer. Incubate the mixture at room temperature for 10 minutes to activate TAFI in the plasma to TAFIa. Subsequently, add 20μL of Hirudin to terminate the activation reaction.
3. TAFIa Activity Detection: To the reaction mixture from the previous step, add 20μL of the 20mM Hippuryl-Arg substrate working solution and incubate at room temperature for 10 minutes. Then, add 20μL of 1M HCl solution to terminate the substrate reaction. Next, sequentially add 20μL of 1M NaOH solution and 25μL of 1M phosphate buffer to the reaction system. Finally, add 60μL of the 3% cyanuric chloride solution in 1,4-dioxane and let it stand for sufficient color development.
4. Detection and Calculation: Centrifuge the color-developed reaction mixture at 14,000g for 2 minutes. Take the supernatant and measure its absorbance at a wavelength of 405nm using a spectrophotometer. All samples and standard curve points should be diluted to 50% with heat-inactivated serum and measured in duplicate.
Precautions:
(1) For your safety and health, please wear a lab coat and disposable gloves during operation.
References:
[1] Santamaría A, Borrell M, Oliver A, et al. Association of functional thrombin-activatable fibrinolysis inhibitor (TAFI) with conventional cardiovascular risk factors and its correlation with other hemostatic factors in a Spanish population. Am J Hematol. 2004 Aug;76(4):348-52.
| Cas No. | 744-46-7 | SDF | |
| Formula | C15H21N5O4 | M.Wt | 335.36 |
| Solubility | 50 mg/ml in AcOH 80 % | Storage | Store at -20°C; protect from light |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.9819 mL | 14.9094 mL | 29.8187 mL |
| 5 mM | 596.4 μL | 2.9819 mL | 5.9637 mL |
| 10 mM | 298.2 μL | 1.4909 mL | 2.9819 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
g
μL
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >98.00% Appearance: A solid
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 14 reference(s) in Google Scholar.)