| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| 25mg |
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| 50mg |
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| 100mg |
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| 250mg |
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Purity: ≥98%
Leupeptin Hemisulfate (NK-381; N-acetyl-L-leucyl-L-leucyl-L-argininal) is a naturally occurring membrane-permeable, competitive, reversible inhibitor of cysteine and serine proteases that may have anti-inflammatory and antioxidant properties. With Ki values of 35 nM, 3.4 μM, 6 nM, and 72 nM, respectively, it inhibits human plasmin, bovine spleen cathepsin B, recombinant human calpain, and bovine trypsin. It was first separated from the Streptomyces species in order to investigate the protease activity. Because of its polar C-terminal, it had poor membrane permeability.
| Targets |
protease: Cathepsin B; Cathepsin L; Cathepsin H; Ser/Thr Protease; Mpro
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| ln Vitro |
Leupeptin, produced by a variety of actinomycetes, which effectively prevent proteolysis.[1] Tubulin purity is raised when leupeptin hemisulfate shields it from endogenous proteolytic activities during the isolation process.[2] Leupeptin hemisulfate has the potential to restore up to 50% of the expression of the hepatitis B surface antigen (HBsAg) in cell suspension cultures. [3]
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| ln Vivo |
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| Enzyme Assay |
Mpro enzyme activity inhibition test. [5]
A total of 20 mM leupeptin hemisulfate in deionized water was diluted to 2 mM to 31.25 μM with 25 mM Tris buffer (pH 8.0). A 30-μl inhibitor solution with a series of concentrations in 25 mM Tris buffer (pH 8.0) was first mixed with 10 μl 100 μM peptide substrate (Dabcyl-TSAVLQ↓SGFRKMK-Edans; GenScript). Next, 10 μl of a final concentration of 200 nM Mpro was added to the plate. The relative fluorescence unit (RFU) value was measured with an excitation wavelength of 360 nm and an emission wavelength of 490 nm at 37°C for 1 h by using a SpectraMax Paradigm multimode detection platform (Molecular Devices, USA). Experiments were performed in triplicate. The enzyme activity reaction rate and inhibition rate were calculated by using MS Excel. The inhibition curve was plotted by using GraphPad Prism 8.0. In vitro antiviral assays. [5] A total of 20 mM leupeptin hemisulfate in deionized water was diluted to 200 μM to 0.06 μM with DMEM containing 1% FBS. Vero cells cultured overnight in 96-well plates were infected by virus at a multiplicity of infection (MOI) of 0.01 for 2 h. The medium was removed, and fresh drug-containing medium was then added to the cells. After 48 h, the cells were lysed in lysis buffer. The viral RNA in 100 μl of the cell supernatant was quantified by reverse transcription-PCR (RT-PCR). Seventy-two hours later, the changes of cytopathic effect were also observed by microscopy. Experiments were performed in triplicate. The experimental results were processed using MS Excel and GraphPad Prism 8.0. |
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| Cell Assay |
Leupeptin inhibited human coronavirus strain 229E multiplication in MRC-C cell cultures. Leupeptin's IC50 value in plaque tests was 0.4 μg/mL, and at 50 μg/mL, it had no effect on the host cells' ability to grow. Leupeptin (100 μg/mL) only inhibited virus yield in single-cycle growth experiments when added within two hours of infection, suggesting that it acts on the early stages of virus replication.[5]
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| Animal Protocol |
C57BL/6NCrl male mice
20 mg/kg i.p. |
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| References |
| Molecular Formula |
C20H38N6O4.1/2H2SO4
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|---|---|
| Molecular Weight |
475.59
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| Exact Mass |
950.56
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| Elemental Analysis |
C, 50.51; H, 8.27; N, 17.67; O, 20.18; S, 3.37
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| CAS # |
103476-89-7
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| Related CAS # |
Leupeptin;55123-66-5;Leupeptin Ac-LL;24365-47-7
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| SequenceShortening |
Ac-LLR-CHO
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| Appearance |
White to off-white solid powder
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| Source |
Microbial Metabolite
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| tPSA |
421Ų
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| SMILES |
CC(C)C[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCN=C(N)N)C=O)NC(=O)C.CC(C)C[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCN=C(N)N)C=O)NC(=O)C.OS(=O)(=O)O
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| InChi Key |
CIPMKIHUGVGQTG-VFFZMTJFSA-N
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| InChi Code |
InChI=1S/2C20H38N6O4.H2O4S/c2*1-12(2)9-16(24-14(5)28)19(30)26-17(10-13(3)4)18(29)25-15(11-27)7-6-8-23-20(21)22;1-5(2,3)4/h2*11-13,15-17H,6-10H2,1-5H3,(H,24,28)(H,25,29)(H,26,30)(H4,21,22,23);(H2,1,2,3,4)/t2*15-,16-,17-;/m00./s1
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| Chemical Name |
(2S)-2-acetamido-N-[(2S)-1-[[(2S)-5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]-4-methylpentanamide;sulfuric acid
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| Synonyms |
NK-381; NK 381; Leupeptin hemisulfate; NK381;
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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. |
| 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) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: 100 mg/mL (210.27 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication (<60°C).
Solubility in Formulation 2: ~100 mg/mL (210.3 mM) in PBS; or ~83 mg/mL (175 mM) in H2O (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.1027 mL | 10.5133 mL | 21.0265 mL | |
| 5 mM | 0.4205 mL | 2.1027 mL | 4.2053 mL | |
| 10 mM | 0.2103 mL | 1.0513 mL | 2.1027 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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