Mechanisms of the in vitro antimutagenic action of chlorophyllin against benzo[a]pyrene: studies of enzyme inhibition, molecular complex formation and degradation of the ultimate carcinogen.
Mechanisms of the antimutagenic action of chlorophyllin (CHL) towards benzo[a]pyrene (BP) were studied in vitro. In the Salmonella assay, CHL inhibited the mutagenic activity of BP in the presence of an S9 activation system and was particularly effective against the direct-acting ultimate carcinogen, benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE). Spectral studies indicated that the time-dependent hydrolysis of BPDE to tetrols was augmented in the presence of CHL concentrations on the order of 5 microM. Dose-related inhibition of several cytochrome P450-dependent enzyme activities was observed upon addition of CHL to in vitro incubations. Spectral changes for the interaction between CHL and cytochrome P450 indicated that CHL does not bind to the active site of the enzyme, but exerts its inhibitory effect indirectly. This was achieved by inhibiting NADPH-cytochrome P450 reductase (Ki approximately 120 microM with cytochrome c as substrate), and did not involve lowering of the effective substrate concentration by complex formation with the procarcinogen. It is concluded that the in vitro antimutagenic activity of CHL towards BP involves accelerated degradation of the ultimate carcinogen, with inhibition of carcinogen activation occurring only at high CHL concentrations. The latter mechanism is unlikely to occur in vivo following p.o. administration due to the limited uptake of CHL from the gut, but tissue concentrations may be sufficiently high to cause degradation of BPDE.
Tachino
N
N
Department of Environmental Biochemistry, University of Hawaii, Honolulu 96822.
Guo
D
D
Dashwood
W M
WM
Yamane
S
S
Larsen
R
R
Dashwood
R
R
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000378:metabolism; D000818:Animals; D016587:Antimutagenic Agents / Q000378:metabolism / Q000494:pharmacology*; D001564:Benzo(a)pyrene / Q000378:metabolism*; D001711:Biotransformation / Q000187:drug effects; D002273:Carcinogens / Q000378:metabolism; D002735:Chlorophyllides / Q000378:metabolism / Q000494:pharmacology*; D019363:Cytochrome P-450 CYP1A1; D065607:Cytochrome P-450 Enzyme Inhibitors; D004247:DNA / Q000378:metabolism; D018736:DNA Adducts; D006868:Hydrolysis; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors*; D009152:Mutagenicity Tests; D010088:Oxidoreductases / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D012486:Salmonella typhimurium / Q000187:drug effects / Q000235:genetics
D016587:Antimutagenic Agents; D002273:Carcinogens; D002735:Chlorophyllides; D065607:Cytochrome P-450 Enzyme Inhibitors; D018736:DNA Adducts; C043931:benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide-DNA; D001564:Benzo(a)pyrene; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D004247:DNA; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases; D019363:Cytochrome P-450 CYP1A1; C007020:chlorophyllin