| Size | Price | Stock | Qty |
|---|---|---|---|
| 5mg |
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| 10mg |
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| Other Sizes |
Purity: =99.67%
| Targets |
Trypsin[1]
Boc-Gln-Ala-Arg-AMC acetate targets trypsin and TMPRSS2 as a substrate. It is cleaved by these proteases at the Arg-AMC bond, releasing the fluorescent AMC moiety, which can be quantified to measure protease activity. |
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| ln Vitro |
Many laboratories routinely use the Boc-Gln-Ala-Arg-AMC (QAR-AMC) substrate to assay trypsin activity in pancreas homogenates. When used in low concentrations, this substrate is relatively specific for trypsin and does not react with thrombin. We compared this substrate to the Z-Gly-Pro-Arg-AMC (GPR-AMC) substrate in our assay and found that the background trypsin activity measured in the pancreas from mice given saline injections was significantly lower with the GPR-AMC substrate. As a result of the lower background activity, the fold increase in trypsin activity in mice given cerulein was about 3-fold higher when measured with GPR-AMC versus QAR-AMC. Based on these observations, we recommend the use of the GPR-AMC substrate to determine intrapancreatic trypsin activity. The trypsin activity from cerulein-treated mice measured with the GPR-AMC substrate can be completely blocked by 1–2 μM ecotin or aprotinin added simultaneously with the substrate (not shown). Since at the given concentrations these protease inhibitors do not affect thrombin (ecotin is completely ineffective and Ki for aprotinin is 61 μM), we can safely exclude the possibility that thrombin contamination would interfere with the assay.[1]
In vitro, Boc-Gln-Ala-Arg-AMC acetate is a fluorogenic substrate for trypsin and can be used for measuring the proteolytic activity of TMPRSS2. The release of AMC upon cleavage is monitored by fluorescence spectroscopy, providing a quantitative measure of protease activity. |
| ln Vivo |
Specific in vivo activity data for Boc-Gln-Ala-Arg-AMC acetate are not applicable, as it is a substrate used in in vitro assays rather than a bioactive compound with in vivo effects.
|
| Enzyme Assay |
Proteolytic activity assay- The preparation of recombinant TMPRSS2 was described in 4. The proteolytic activity of TMPRSS2 was examined by the measurement of a fluorescence resonance energy transfer (FRET) of a protease substrate (Boc-Gln-Ala-Arg-AMC).[2]
Non-cellular enzyme assays for Boc-Gln-Ala-Arg-AMC acetate involve incubating the substrate with the target protease (trypsin or TMPRSS2) in buffer. The release of fluorescent AMC is monitored over time using a fluorometer or fluorescence plate reader, and protease activity is calculated from the rate of fluorescence increase. |
| Cell Assay |
Cell-based proteolytic activity analysis- CWR22Rv1 was transiently transfected with pCMV-TMPRSS2 and PCDNA3.1-HAI-2 mutant plasmids using pCMV vector and pCDNA3.1 vector as control. CWR22Rv1 cells were seeded into 24-well-plate and transfected with pCMV-TMPRSS2 and PCDNA3.1-HAI-2 using Lipofectamine3000. After 48 hours, the proteolytic activity of CWR22Rv1 cells was examined by the measurement of a fluorescence resonance energy transfer (FRET) of a protease substrate (Boc-Gln-Ala-Arg-AMC).[2]
In vitro cellular assays for Boc-Gln-Ala-Arg-AMC acetate are not typically performed, as it is a substrate for biochemical assays rather than a cell-active compound. It may be used in cell lysates or conditioned media to measure protease activity. |
| Animal Protocol |
In vivo animal experiments are not applicable for Boc-Gln-Ala-Arg-AMC acetate, as it is a substrate used in biochemical assays rather than a compound with in vivo pharmacological activity.
|
| ADME/Pharmacokinetics |
Pharmacokinetic properties are not applicable for Boc-Gln-Ala-Arg-AMC acetate, as it is a substrate used in biochemical assays rather than a systemically administered compound.
|
| Toxicity/Toxicokinetics |
Toxicological data for Boc-Gln-Ala-Arg-AMC acetate are not provided in available sources. As a research substrate, its safety profile has not been extensively characterized. It is intended for research use only and not for human consumption.
|
| References | |
| Additional Infomation |
Activation of pancreatic digestive proteases (especially trypsin and chymotrypsin) is a hallmark of pancreatitis. In rodent models, fluorescent peptide substrates are commonly used to detect protease activation in pancreatic homogenates. This study investigated the optimal detection conditions for pancreatic trypsin and chymotrypsin activation following a single intraperitoneal injection of secretin in C57BL/6N mice. We found that using a smaller amount of pancreatic homogenate and removing bovine serum albumin from the detection buffer significantly improved the detection results for these proteases. In addition, the pancreatic homogenate must be freshly prepared and tested, as freeze-thaw cycles can stimulate protease activation. Finally, replacing the widely used Boc-Gln-Ala-Arg-AMC trypsin substrate with Z-Gly-Pro-Arg-AMC reduced the background activity in saline-treated control mice, thereby increasing the level of secretin-induced trypsin activation. Using the optimized protocol, we repeatedly measured that the activities of pancreatic trypsin and chymotrypsin increased by 20-fold and 200-fold, respectively, in mice given secretin. [1]
Boc-Gln-Ala-Arg-AMC acetate is a fluorogenic substrate for trypsin and TMPRSS2. It releases AMC upon proteolysis, enabling quantitative measurement of protease activity. It is supplied for research purposes only. |
| Molecular Formula |
C31H46N8O10
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|---|---|
| Molecular Weight |
690.74
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| Related CAS # |
Boc-Gln-Ala-Arg-AMC hydrochloride;201849-55-0
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| Appearance |
Typically exists as white to off-white solids at room temperature
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| Chemical Name |
tert-butyl N-[(2S)-5-amino-1-[[(2S)-1-[[(2S)-5-(diaminomethylideneamino)-1-[(4-methyl-2-oxochromen-7-yl)amino]-1-oxopentan-2-yl]amino]-1-oxopropan-2-yl]amino]-1,5-dioxopentan-2-yl]carbamate acetate
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO :~125 mg/mL (~180.97 mM)
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|---|---|
| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.4477 mL | 7.2386 mL | 14.4772 mL | |
| 5 mM | 0.2895 mL | 1.4477 mL | 2.8954 mL | |
| 10 mM | 0.1448 mL | 0.7239 mL | 1.4477 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution 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 stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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