| Size | Price | |
|---|---|---|
| 500mg | ||
| 1g | ||
| Other Sizes |
| Targets |
Trypsin[1]
|
|---|---|
| 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]
|
| 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]
|
| 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]
|
| References | |
| Additional Infomation |
Intrapancreatic activation of digestive proteases, trypsin and chymotrypsin in particular, is a hallmark of pancreatitis. In experimental rodent models, protease activation is routinely measured from pancreatic homogenates using fluorogenic peptide substrates. Here we investigated the optimal conditions for the determination of intrapancreatic trypsin and chymotrypsin activation elicited by a single intraperitoneal injection of cerulein in C57BL/6N mice. We found that these protease assays were significantly improved by using lower amounts of pancreatic homogenate and exclusion of bovine serum albumin from the assay buffer. Furthermore, pancreatic homogenates had to be freshly prepared and assayed; as freezing and thawing stimulated protease activation. Finally, replacement of 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 and thereby increased the extent of cerulein-induced trypsin activation. Using the optimized protocol, we reproducibly measured 20-fold and 200-fold increases in the intrapancreatic trypsin and chymotrypsin activity, respectively, in mice given cerulein. [1]
|
| Molecular Formula |
C29H43CLN8O8
|
|---|---|
| Molecular Weight |
667.153525590897
|
| Exact Mass |
666.289
|
| CAS # |
201849-55-0
|
| Related CAS # |
Boc-Gln-Ala-Arg-AMC acetate
|
| PubChem CID |
87167928
|
| Appearance |
Typically exists as solid at room temperature
|
| Hydrogen Bond Donor Count |
8
|
| Hydrogen Bond Acceptor Count |
9
|
| Rotatable Bond Count |
16
|
| Heavy Atom Count |
46
|
| Complexity |
1180
|
| Defined Atom Stereocenter Count |
3
|
| SMILES |
Cl.O(C(N[C@H](C(N[C@@H](C)C(N[C@H](C(NC1=CC=C2C(C)=CC(=O)OC2=C1)=O)CCC/N=C(\N)/N)=O)=O)CCC(N)=O)=O)C(C)(C)C
|
| InChi Key |
PJANUSPBJOFUEZ-BOKRKPFNSA-N
|
| InChi Code |
InChI=1S/C29H42N8O8.ClH/c1-15-13-23(39)44-21-14-17(8-9-18(15)21)35-26(42)19(7-6-12-33-27(31)32)36-24(40)16(2)34-25(41)20(10-11-22(30)38)37-28(43)45-29(3,4)5;/h8-9,13-14,16,19-20H,6-7,10-12H2,1-5H3,(H2,30,38)(H,34,41)(H,35,42)(H,36,40)(H,37,43)(H4,31,32,33);1H/t16-,19-,20-;/m0./s1
|
| 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;hydrochloride
|
| Synonyms |
Boc-Gln-Ala-Arg-AMC HCl; 201849-55-0; Boc-Gln-Ala-Arg-AMC (hydrochloride)
|
| HS Tariff Code |
2934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
|
| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
|
|---|---|
| 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 | 1.4989 mL | 7.4946 mL | 14.9891 mL | |
| 5 mM | 0.2998 mL | 1.4989 mL | 2.9978 mL | |
| 10 mM | 0.1499 mL | 0.7495 mL | 1.4989 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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