-transparent.png){kind=logo}
The hepatic cytochrome P-450-mediated metabolism and metabolic activation of [chloroethyl-3H]cyclophosphamide [( chloroethyl-3H]CP) and [4-14C]cyclophosphamide [( 4-14C]CP) were investigated in vitro in the reconstituted system containing cytochrome P-450 isolated from phenobarbital-treated rats. In addition, hepatic microsomal binding and the hepatic microsome-mediated metabolism of [14C]acrolein, a metabolite of [4-14C]CP, were also investigated. The metabolism of [chloroethyl-3H]CP and [4-14C]CP to polar metabolites was found to depend on the presence of NADPH and showed concentration dependence with respect to cytochrome P-450 and NADPH:cytochrome P-450 reductase. Km and Vmax values were essentially similar (Km, 0.44 and 0.42 mM; Vmax, 4.8 and 7.0 nmol of polar metabolites formed/min/nmol of cytochrome P-450 for [4-14C]CP and [chloroethyl-3H]CP, respectively). The patterns of inhibition by microsomal mixed-function oxidase inhibitors, anti-cytochrome P-450 antibody, and heat denaturation of the cytochrome P-450 were essentially similar, with subtle differences between [4-14C]CP and [chloroethyl-3H]CP metabolism. The order of inhibition by various mixed-function oxidase inhibitors was SKF greater than alpha- and beta-naphthoflavones greater than metyrapone. The in vitro metabolic activation of CP in the reconstituted system demonstrated predominant binding of [chloroethyl-3H]CP to nucleic acids and almost exclusive binding of [4-14C]CP to proteins. Gel electrophoresis-fluorography of the proteins in the reconstituted system treated with [4-14C]CP demonstrated localization of the 14C label in the cytochrome P-450 region. To examine this association further, hepatic microsomes were modified with [14C]acrolein in the presence and the absence of NADPH. The results confirmed covalent association between [14C]acrolein and cytochrome P-450 in the microsomes and also demonstrated further metabolism of [14C]acrolein, apparently to an epoxide, which is capable of binding covalently to proteins. The results of these investigations not only confirm the significance of primary metabolism but also emphasize the potential role of the secondary metabolism of cyclophosphamide in some of its toxic manifestations.
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
March 1981,
Biochemical pharmacology,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
March 1980,
Biochemical pharmacology,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
June 1982,
Biochemical pharmacology,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
June 1978,
The Journal of biological chemistry,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
August 1977,
Archives of biochemistry and biophysics,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
August 1984,
Biochemical and biophysical research communications,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
October 1980,
Biochemical pharmacology,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
August 1976,
Chemico-biological interactions,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
January 1984,
Annals of the New York Academy of Sciences,
A J Marinello, and
S K Bansal, and
B Paul, and
P L Koser, and
J Love, and
R F Struck, and
H L Gurtoo
May 1977,
Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme,
