E3132 Sigma-Aldrich


protease inhibitor

Synonym: trans-Epoxysuccinyl-L-leucylamido(4-guanidino)butane, L-trans-3-Carboxyoxiran-2-carbonyl-L-leucylagmatine, N-(trans-Epoxysuccinyl)-L-leucine 4-guanidinobutylamide

  • CAS Number 66701-25-5

  • Empirical Formula (Hill Notation) C15H27N5O5

  • Molecular Weight 357.41

  •  Beilstein/REAXYS Number 1405664

  •  MDL number MFCD00080261

  •  PubChem Substance ID 24278411

  •  NACRES NA.77



Related Categories A to C, Application Index, Biochemicals and Reagents, Bromelain, Cathepsin B,
Quality Level   200
biological source   synthetic (organic)
assay   ≥99% (HPLC)
form   powder
solubility   water: 20 mg/mL, clear, colorless to faintly yellow
storage temp.   2-8°C
SMILES string   CC(C)C[C@H](NC(=O)[C@@H]1O[C@H]1C(O)=O)C(=O)NCCCCNC(N)=N
InChI   1S/C15H27N5O5/c1-8(2)7-9(20-13(22)10-11(25-10)14(23)24)12(21)18-5-3-4-6-19-15(16)17/h8-11H,3-7H2,1-2H3,(H,18,21)(H,20,22)(H,23,24)(H4,16,17,19)/t9-,10+,11+/m0/s1


General description

E-64 is a cysteine protease inhibitor that was isolated from the mold Aspergillus japonicus TPR-64. E-64 is also known as N-[N-(L-3-trans-carboxyoxiran-2-carbonyl)-L-leucyl]-agmatine. E-64 effectively inhibits various cysteine proteases, in particular:
• cathepsin K
• cathepsin L
• cathepsin S
E-64 also acts against other enzymes, such as:
• calpain
• cathepsin B
• cathepsin H
• papain


E-64 is an effective ligand for affinity purification of cysteine proteases. When coupled to a thiolated affinity matrix, binding is no longer irreversible, but specificity is retained.


1, 5, 10 mg in glass bottle

25, 50 mg in poly bottle

Biochem/physiol Actions

E-64 is an irreversible, potent, and highly selective cysteine protease inhibitor. E-64 does not react with the functional thiol group of non-protease enzymes, such as L-lactate dehydrogenase or creatine kinase. E-64 will not inhibit serine proteases (except trypsin) like other cysteine protease inhibitors, leupeptin and antipain. The trans-epoxysuccinyl group (active moiety) of E-64 irreversibly binds to an active thiol group in many cysteine proteases, such as papain, actinidase, and cathepsins B, H, and L to form a thioether linkage. E-64 is a very useful cysteine protease inhibitor for use in in vivo studies because it has a specific inhibition, it is permeable in cells and tissues and has low toxicity.

Features and Benefits

This compound is also offered as part of Sigma′s Library of Pharmacologically Active Compounds (LOPAC®1280), a biologically annotated collection of high-quality, ready-to-screen compounds. Click here to learn more.

Preparation Note

E-64 is soluble in water. A 20 mg/ml solution can be prepared in water (heat may be needed). A suggested stock solution is a 1 mM aqueous solution. The effective concentration for use as a protease inhibitor is 1 to10 μM. Aqueous stock solutions are stable for months at -20 °C. Diluted solutions are stable for days at neutral pH. E-64 is stable from pH 2-10, but is unstable in ammonia or in HCl. E-64 is also soluble in DMSO, a 10 mM solution can be prepared in dry DMSO and stored at -20 °C. Subsequent dilutions were in culture medium. Solutions for injection were prepared by dissolving E-64 in 0.9% sodium chloride or in a minimum amount of saturated sodium bicarbonate followed by dilution with 0.9% sodium chloride (after adjusting the pH to 7.0 with acetic acid).

Legal Information

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Safety & Documentation

Safety Information

Personal Protective Equipment 
NONH for all modes of transport
WGK Germany 
Flash Point(F) 
Not applicable
Flash Point(C) 
Not applicable


Certificate of Analysis (COA)

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Certificate of Origin (COO)

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Protocols & Articles
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