Cyclohex-1,2-ylenediamine

Additional toxicological data

Administrative data

Endpoint:

additional toxicological information

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Vicinal Diamines as Pyrroloquinoline Quinone-directed Irreversible Inhibitors of Lysyl Oxidase

Author:

Stephen N. Gacheru, Philip C. Trackman, Susan D. Calaman, Frederick T. Greenaway, and
Herbert M. Kagan

Year:

1989

Bibliographic source:

THE JOURNAL OF BIOLOGICAL CHEMISTRY
Vol. 264, No. 22, Issue of August 5, pp. 12963-12969,1989

Reference 2

Reference Type:

review article or handbook

Title:

Enzymology of Lysyl Oxidase

Author:

Herbert M. Kagan, Stephen N. Gacheru, Philip C. Trackman, Susan D. Calaman
and Frederick T. Greenaway

Year:

1989

Bibliographic source:

J. A. Jongejan and 1. A. Duine (eds.), PQQ and Quinoproteins, 317-326.
© 1989 by Kluwer Academic Publishers.

Materials and methods

Principles of method if other than guideline:

The authors purified lysyl oxidase (LOX) from bovine aorta, defined one enzyme unit and quantified functional active site content. Afterwards, the enzyme was assayed against DCH by a peroxidase-coupled fluorescence method in a defined reaction mixture. Due to beginning enzyme inhibition at concentrations starting from 0.8 µM, steady-state kinetic analyses were restricted to concentrations not greater than 1.2 µM cis-1,2-diaminocyclohexane (0.2, 0.5, 0.8, 1.2 µM). The assays assessing inhibition of LOX activity against cis- and trans- isomers of 1,2-diaminocyclohexane contained enzyme active site concentrations of 0.02 µM and n-hexylamine as productive substrate. Initial rates of substrate oxidation were assessed from the slopes of fluorescence tracings versus time after addition of the enzyme (20 s to 3 min). Steady-state kinetic constants were calculated according to Michaelis-Menten equation using the Fortram program of Cleland (Cleland, W. W. (1979) Methods Enzymol. 63, 103-138) applying a least squares fitting procedure to the Michaelis-Menten equation. Additionally, enzyme inhibition in the absence of a productive substrate was assessed by incubation of 50 mM cis-DCH with LOX for 15 min at 37°C in the reaction mixture. Time dependence for the expression of irreversible inhibition was assessed in a further experiment as well.

GLP compliance:

not specified

Test material

Test material information

Test material form:

not specified

Details on test material:

Please refer to entry 'Specific details on test material used for the study'

Specific details on test material used for the study:

cis-1,2-diaminocyclohexane (CAS no.: 1436-59-5)

trans-1,2-diaminocyclohexane (CAS no.: 1121-22-8)

both substances obtained from Alfa products (Thermo Fisher Scientific, Waltham, MA, USA)

Results and discussion

Any other information on results incl. tables

Cis-1,2-diaminocyclohexane was shown to be a more potent inhibitor of elastin oxidation by LOX ( IC50 = 1.5 µM) than trans-1,2-diaminocyclohexane (IC50 > 10 mM, extrapolated). A Lineweaver-Burke plot showed that the inhibition is competitve with a K_I of 3.8 x10^-7M measured under the initial rate conditions (20s-3min) and at the low concentrations (0.2, 0.5, 0.8, 1.2 µM) used. After 15 min incubation of LOX with 50 mM cis-DCH in the absence of a productive substrate, LOX was completely and irreversibly inhibited. Time dependence assays showed that the rates of enzyme activity loss increased with increasing cis-DCH concentrations, following first order kintetics with k_inact = 18 min^-1.

Applicant's summary and conclusion

Conclusions:

Cis-1,2-diaminocyclohexane but not trans-1,2-diaminocyclohexane is a potent lysyl oxidase inhibitor.