Suc-AAPF-pNA (Synonyms: SuccinylAlaAlaProPhepnitroanilide) |
| Catalog No.GC44960 |
Suc-AAPF-pNA (Suc-Ala-Ala-Pro-Phe-pNA) is a colorimetrically specific substrate for elastase. Under the action of enzymes, Suc-AAPF-pNA is hydrolyzed, releasing yellow p-nitroaniline (pNA), and the change in absorbance at 400-410 nm is measured to reflect enzyme activity through colorimetry.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 70967-97-4
Sample solution is provided at 25 µL, 10mM.
Suc-AAPF-pNA (Suc-Ala-Ala-Pro-Phe-pNA) is a colorimetrically specific substrate for elastase[1]. Under the action of enzymes, Suc-AAPF-pNA is hydrolyzed, releasing yellow p-nitroaniline (pNA), and the change in absorbance at 400-410 nm is measured to reflect enzyme activity through colorimetry[2]. Suc-AAPF-pNA also serves as a substrate for cathepsin G, bacillus protease, chymotrypsin, chymotrypsin, cyclophilin, and peptidyl-prolyl isomerase[3] .
The structure and group characteristics of Suc-AAPF-pNA are as follows:
(1)Succinyl group (Suc): Increases the water solubility and stability of the substrate.
(2)Amino acid sequence (Ala-Ala-Pro-Phe): This specific sequence mimics certain natural substrates and helps study the specificity of proteases towards different substrates.
(3)p-Nitroaniline (pNA): A common chromophoric group that releases a yellow product with absorbance characteristics upon hydrolysis, facilitating the monitoring of enzyme reactions through spectrometry.
References:
[1] Vinci V A. Biochemical and genetic analysis of serine proteases of Streptomyces spp[M]. The Ohio State University, 1988.
[2] Chaves-Pozo E, Valero Y, Lozano M T, et al. Fish granzyme A shows a greater role than granzyme B in fish innate cell-mediated cytotoxicity, Front. Immunol. 10 (2019) 2579[J]. 2019.
[3] Attucci S, Korkmaz B, Juliano L, et al. Measurement of free and membrane-bound cathepsin G in human neutrophils using new sensitive fluorogenic substrates[J]. Biochemical Journal, 2002, 366(3): 965-970.
This protocol only provides a guideline, and should be modified according to your specific needs.
1. Solution preparation
(1) Stock solution: Dissolve Suc-AAPF-pNA in DMSO to a final concentration of 10 mM.
Note: After unused storage solution is aliquoted, store it at -20℃ in the dark to avoid repeated freezing and thawing.
(2) Working solution: Dilute the stock solution with experimental buffer to the required working concentration, for example: 0.2 mM.
Note: Please adjust the optimal working concentration according to the actual situation or refer to the literature to set the gradient concentration by yourself. The working solution must be prepared and used immediately.
2. Sensitivity Determination of Suc-AAPF-pNA to Protease Cleavage[1] (from the literature, for reference only)
(1) Materials Preparation: Substrate: Suc-AAPF-pNA, Active enzyme, PBS buffer, 96-well microtiter plate, Microplate reader
(2) Reaction system: Add the chromogenic substrate Suc-AAPF-pNA (5 μl, final concentration 0.2 mM) and active enzyme (a few microliters, the amount depends on the enzyme activity) to each well, and add PBS to make the final volume reach 200 μl.
(3) Reaction conditions: Perform the reaction at a specific temperature, such as 20°C.
(4) Measure the absorbance spectrophotometrically at 405 nm using a microplate reader. Measure at the following time points: 0, 20, 40, 60, 120, 180, 240, 300, and 360 minutes. Control: Set up a blank well for each substrate without adding any enzyme.
(5) Data Collection and Analysis: Conduct reactions in triplicate. Subtract the blank measurement at each time point. Use the mean absorbance of the three reactions for plotting and analysis.
Notes:
This protocol provides guidance for determining sensitivity to protease cleavage using Suc-AAPF-pNA. It can be adapted based on other literature and specific experimental requirements.
References:
[1] Fu Z, Akula S, Qiao C, et al. Duodenases are a small subfamily of ruminant intestinal serine proteases that have undergone a remarkable diversification in cleavage specificity[J]. PLoS One, 2021, 16(5): e0252624.
| Cas No. | 70967-97-4 | SDF | |
| Synonyms | SuccinylAlaAlaProPhepnitroanilide | ||
| Canonical SMILES | OC(CCC(N[C@@H](C)C(N[C@@H](C)C(N1CCC[C@H]1C(N[C@@H](CC2=CC=CC=C2)C(NC3=CC=C([N+]([O-])=O)C=C3)=O)=O)=O)=O)=O)=O | ||
| Formula | C30H36N6O9 | M.Wt | 624.7 |
| Solubility | DMF: 5 mg/ml,DMSO: 5 mg/ml,DMSO:PBS (pH 7.2) (1:1): 0.5 mg/ml | Storage | Store at -20°C |
| 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 | |||
|
1 mg | 5 mg | 10 mg |
| 1 mM | 1.6008 mL | 8.0038 mL | 16.0077 mL |
| 5 mM | 0.3202 mL | 1.6008 mL | 3.2015 mL |
| 10 mM | 0.1601 mL | 0.8004 mL | 1.6008 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 ddH2O, 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%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 28 reference(s) in Google Scholar.)
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