Prevention by synthetic phenolic antioxidants of 2-amino-3, 8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)- or activated MeIQx-induced mutagenesis and MeIQx-induced rat hepatocarcinogenesis, and role of antioxidant activity in the prevention of carcinogenesis.

Effects of synthetic phenolic antioxidants 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), tert-butylhydroquinone (TBHQ) and propyl gallate (PG) on 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)- or activated MeIQx-induced mutagenesis and rat hepatocarcinogenesis were compared, and the association between antioxidative activity and inhibition of carcinogenesis was examined. When the antimutagenic activity of five antioxidants against MeIQx- or activated MeIQx-induced mutagenesis was compared in the Ames assay using the Salmonella strain TA 98, HTHQ showed the greatest effect, followed by BHA, BHT, PG and TBHQ, in that order. In a rat hepatocarcinogenesis study, 6-week-old male F344 rats were given a single i.p. injection of 200 mg/kg bw of diethylnitrosamine (DEN) and starting 2 weeks later, groups of 15 animals received a diet containing 0.03% MeIQx alone, MeIQx together with each antioxidant at a dietary dose of 0.25%, each antioxidant alone, or basal diet alone for 6 weeks. Three weeks after the DEN injection, animals were subjected to 2/3 partial hepatectomy. Liver tissues obtained at partial hepatectomy were processed for the measurement of 8-hydroxydeoxyguanine (8-OHdG) and lipid peroxidation. The average number and areas of glutathione S-transferase placental form (GST-P) positive foci were increased by the treatment with MeIQx (27.2 +/- 6.5 per cm2 and 3.17 +/- 0.96 mm2/cm2, respectively). A significant decrease in these parameters was found with the simultaneous antioxidant treatment, HTHQ demonstrating the greatest effect, followed by BHA, BHT and TBHQ, and PG. Without MeIQx, a weak increase in the number of foci was observed in the BHT treatment case. Examination of 8-OHdG levels in liver DNA, as well as malondialdehyde (MDA) and 4-hydroxyalkenals, did not reveal any inter-group variation. These results indicate that antimutagenic activity of antioxidants against MeIQx roughly parallels their anticarcinogenic activity, with HTHQ as the most powerful chemopreventor, but that oxidative stress and antioxidative activity may not be responsible for MeIQx-induced hepatocarcinogenesis and its inhibition, respectively.

Strong anti-mutagenic activity of the novel lipophilic antioxidant 1-O-hexyl-2,3,5-trimethylhydroquinone against heterocyclic amine-induced mutagenesis in the Ames assay and its effect on metabolic activation of 2-amino-6-methyldipyrido[1,2-a:3',2'-d] imidazole (Glu-p-1).

Antimutagenic effects of a novel lipophilic antioxidant, 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ), and other known antioxidants against heterocyclic amine- or other mutagen-induced mutagenesis were examined in the Ames assay using Salmonella strain TA 98 to access the chemo-preventive effects of antioxidants on heterocyclic amine-induced carcinogenesis. Further the mechanisms of inhibition by HTHQ were accessed. HTHQ was shown to potently inhibit mutagenesis induced by all of 8 different heterocyclic amines at rates between 100% and 63% in the presence of S9 mix. When the protection of HTHQ against 2-amino-6- methyldipyrido[1,2-alpha:3',2'-d]imidazole (Glu-P-1)-induced mutagenesis was compared with known antioxidants t-butylhydroquinone, propyl gallate, BHA, BHT and alpha-tocopherol, HTHQ showed the greatest effect. Among hexyl, butyl, ethyl and methyl derivatives of 1-O-alkyl-2,3,5-trimethylhydroquinone, HTHQ was the most effective in inhibiting Glu-P-1-, 3-amino-1-methyl-5-H-pyrido[4,3-b]indole (Trp-P-2)- or 2-amino-3-methylimidazo[4,5-f]quinoline (IQ)-induced mutagenesis. On the other hand, HTHQ did not inhibit mutagenic activity induced by other mutagens such as N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2) and benzo[a]pyrene. HTHQ weakly inhibited that due to direct mutagen 2-nitro derivative of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) only in the presence of S9 mix. No such influence on a 2-nitro derivative of 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) or 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced mutagenesis, was observed with or without the S9 mix. HTHQ slightly inhibited mutagenesis induced by activated Glu-P-1, a direct acting proximate metabolite of Glu-P-1, in the absence of the S9 mix. HPLC analysis revealed activated Glu-P-1 to be formed by incubating Glu-P-1 with the S9 mix, but this was considerably decreased by the addition of HTHQ. These results indicate that HTHQ is a powerful antimutagenic compound and specifically acts against heterocyclic amines. Its antimutagenic activity appeared to exert by both inhibiting metabolic activation of heterocyclic amines and action on activated N-hydroxy species.

Synthesis and anti lipid-peroxidation activity of hydroquinone monoalkyl ethers.

A series of hydroquinone monoalkyl ethers was synthesized and evaluated for anti lipid-peroxidation activity in rat liver microsomes. 4-Hexyloxy-2,3,6-trimethylphenol (9), having a low redox potential, as well as ascorbic acid exhibited the strongest anti lipid-peroxidation activity (IC50 = 4.2 x 10(-7) M). Structure-activity relationship studies demonstrated that the inhibitory effect of hydroquinone monoalkyl ethers on lipid peroxidation was increased by the acquisition of an optimum hydrophobicity and decreased by an insufficient or excessive hydrophobicity.

Synthesis of 4-alkoxyaryl beta-D-glucopyranosides and their inhibitory effects on histamine release from rat peritoneal mast cells induced by concanavalin A.

The inhibitory effects of newly synthesized 4-alkoxyaryl beta-D-glucopyranosides on histamine release from rat peritoneal mast cells induced by concanavalin A were examined. A plot of hydrophobicity (k') against inhibitory activity of the compounds showed a distinct maximum, and 4-decyloxy-2,3,6-trimethylphenyl beta-D-glucopyranoside was the most potent inhibitor among the tested compounds.

Protective effects of hydroxychalcones on free radical-induced cell damage.

A number of hydroxychalcones were synthesized to evaluate their protective effects against oxidative cell damage and the production of superoxide anion. The hydroxychalcones which have a 3,4-dihydroxycinnamoyl structure were potent inhibitors of lipid peroxidation in rat liver microsomes. In particular, we found that 2',4',3,4-tetrahydroxychalcone (3) exhibited a potent inhibitory effect on H2O2-induced hemolysis due to an antioxidant effect. In addition, this compound strongly inhibited CCl4-induced cytotoxicity in primary cultured hepatocytes and substantially decreased the production of superoxide anion by rat peritoneal exudate macrophages.

3,4-Dihydroxychalcones as potent 5-lipoxygenase and cyclooxygenase inhibitors.

A novel series of 3,4-dihydroxychalcones was synthesized to evaluate their effects against 5-lipoxygenase and cyclooxygenase. Almost all compounds exhibited potent inhibitory effects on 5-lipoxygenase with antioxidative effects, and some also inhibited cyclooxygenase. The 2',5'-disubstituted 3,4-dihydroxychalcones with hydroxy or alkoxy groups exhibited optimal inhibition of cyclooxygenase. We found that 2',5'-dimethoxy-3,4-dihydroxychalcone (37; HX-0836) inhibited cyclooxygenase to the same degree as flufenamic acid and 5-lipoxygenase, more than quercetin. Finally, these active inhibitors of 5-lipoxygenase inhibited arachidonic acid-induced mouse ear edema more than phenidone.

Strong inhibition of 2-amino-6-methyldipyrido[1,2-a:3',2'-d] imidazole-induced mutagenesis and hepatocarcinogenesis by 1-O-hexyl-2,3,5-trimethylhydroquinone.

The effects of 3-O-dodecylcarbomethylascorbic acid (3-O-DAsA), 3-O-ethylascorbic acid (3-O-EAsA) and 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ) on 2-amino-6-methyldipyrido[1,2-a:3',2'-d]-imidazole (Glu-P-1)-induced mutagenesis and hepatocarcinogenesis were examined. In a Salmonella assay, addition of 2.5 to 20.0 mg of HTHQ to Salmonella TA 98 in the presence of S-9 mixture dose-dependently inhibited Glu-P-1-induced mutagenesis. The highest dose showed a 99% reduction in revertants. 3-O-DAsA and 3-O-EAsA were without effect. In an animal study using the medium-term bioassay system for the detection of hepatocarcinogens or hepatopromoters in F344 male rats, treatment with Glu-P-1 alone was associated with a significant increase in the number and area of GST-P-positive foci (47.5 +/- 8.9 and 11.1 +/- 4.7, respectively). Combined treatment with 1.0% HTHQ significantly reduced the number and area of GST-P-positive foci (to 8.1 +/- 2.1 and 0.6 +/- 0.2) while 3-O-DAsA exerted marginal inhibition and 3-O-EAsA had no effect. On the other hand, all three of these compounds slightly enhanced the numbers and areas of foci when given alone. The results indicate that HTHQ is a potent chemopreventer of Glu-P-1-induced hepatocarcinogenesis.

3-O-alkylascorbic acids as free radical quenchers. 3. Protective effect on coronary occlusion-reperfusion induced arrhythmias in anesthetized rats.

Structural modification of ascorbic acid by substitution of the 3-hydroxy group with lipophilic moieties has allowed the development of agents for treating reperfusion injury. These ascorbic acid derivatives inhibited lipid peroxidation, and some of them also reduced coronary reperfusion-induced arrhythmias in anesthetized rats. We found that 3-O-[(dodecylcarbonyl)methyl]ascorbic acid (8) was protective against reperfusion injury without directly influencing hemodynamics. 2-O-Octadecylascorbic acid (19) and 5,6-O-dodecylideneascorbic acid (15) also exhibited a marked effect on reperfusion injury, but significantly reduced the arterial blood pressure and heart rate in rats.

3-O-alkylascorbic acids as free-radical quenchers: synthesis and inhibitory effect on lipid peroxidation.

A novel series of 3-O-alkylascorbic acids (3-RASA, 3a-n) was synthesized to act as radical scavengers for active oxygen species and free radicals, and their redox potentials and inhibitory effects on lipid peroxidation in rat liver microsomes were evaluated. The redox potentials of the 3-RASA compounds were increased by the substituent group to 90-190 mV above the potential for ascorbic acid (i.e., 3-RASA compounds were harder to oxidize). Although 3-O-dodecylascorbic acid (3c) and 3-O-(decylcarbomethyl)ascorbic acid (3i) differed in their redox potentials, they both markedly inhibited lipid peroxidation in rat liver microsomes to a similar extent (IC50 = 3.1 and 3.3 X 10(-6) M, respectively). Structure-activity relationship studies demonstrated that the anti lipid peroxidation activity of the 3-RASA compounds was markedly dependent upon their hydrophobicity.

3-O-alkylascorbic acids as free radical quenchers. II. Inhibitory effects on some lipid peroxidation models.

We previously found that 3-O-dodecylcarbomethylascorbic acid (3-RASA,3,HX-0112) exhibited a potent inhibitory effect on biochemical lipid peroxidation and that 3-RASA (3) alleviated myocardial lesions induced by ischemia-reperfusion treatment in rats. In this study we examined the mode of action of 3-RASA (3) on the inhibition of lipid peroxidation. There was no reducing activity by 3-RASA (3) (i.e., no oxide was produced) against ferric ions and superoxide anion radicals. The low reducing activity of 3-RASA (3) against a radical as compared to that of alpha-tocopherol was obtained by using a stable radical. However, 3-RASA (3) had a potent inhibitory effect, almost equal to that of alpha-tocopherol, in the model of lipid peroxidation dependent on enzymatic superoxide generation. 3-RASA (3) very strongly inhibited the chain-reaction of the peroxidation induced by Fe(2+)-linoleic acid hydroxyperoxide. On the basis of these findings, it appears that the anti-lipid-peroxidative effects of 3-RASA (3) are due to the inhibition of the radical chain-reaction, as a chain-breaking antioxidant.