In vitro effects of L-ascorbic acid (vitamin C) on aryl hydrocarbon hydroxylase activity in hepatic microsomes of mice.

When aromatic hydrocarbon (Ah)-responsive and -non-responsive strains of mice were pretreated with 3-methylcholanthrene (MC) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), vitamin C reduced the microsomal aryl hydrocarbon hydroxylase (AHH) activity. The AHH inhibitors 7,8-benzoflavone (7,8-BF) and 3-methylsulfonyl-3',4,4',5-tetrachlorobiphenyl (3-MSF-3',4,4',5-tetraCB) showed various inhibitory effects depending upon the types of microsomes, whereas vitamin C exhibited inhibition irrespective of the types of microsomes. 7,8-BF and 3-MSF-3',4,4',5-tetraCB as well as vitamin C suppressed the reverse mutation of the Salmonella typhimurium tester strains TA98 and TA100 induced by benzo[a]pyrene.     

Effects of various inducers on diethylstilbestrol metabolism, drug-metabolizing enzyme activities and the aromatic hydrocarbon (Ah) receptor in male Syrian golden hamster liver

In order to elucidate the role of metabolic activation of the synthetic estrogen, diethylstilbestrol (DES), in the mechanism of liver tumor formation in male Syrian golden hamsters observed after combined treatment with DES and 7,8-benzoflavone (7,8-BF), the metabolism of DES and the concentrations and activities of various drug-metabolizing enzymes were studied in hamster liver microsomes after various pretreatments. The levels of the hepatic aromatic hydrocarbon (Ah) receptor were also determined. Pretreatment with 7,8-BF increased both P450 and cytochrome b5 levels, whereas phenobarbital (PB) and 3-methylcholanthrene (MC) induced P450 but not cytochrome b5. 7,8-BF pretreatment increased 7-ethoxyresorufin-O-deethylase (EROD) 3-fold and 7-pentoxyresorufin-O-dealkylase (PROD) 2.5-fold, whereas aromatic hydrocarbon hydroxylase (AHH) and 7-ethoxycoumarin-O-deethylase (ECOD) activities were only slightly induced by 7,8-BF. MC pretreatment increased EROD 8-fold and PROD activity 7-fold, whereas PB pretreatment enhanced AHH 4.5-fold and PROD activity 4-fold. In contrast to PB, pretreatment with 7,8-BF and MC reduced the oxidative metabolism of DES in hepatic microsomes, but the pattern of metabolites was identical with that in untreated controls. Treatment of hamsters with the inducers changed the hepatic Ah receptor level. PB and MC-pretreatment resulted in an increase of the receptor level 1.5-fold and 1.3-fold, respectively, whereas 7,8-BF-pretreatment leads to a 1.5-fold decrease. The dissociation constant Kd is 170 nM for the reaction of 7,8-BF with the hamster Ah receptor compared to 70 nM for 5,6-BF and 38 nM for 2,3,7,8-tetrachlorodibenzofuran (TCDF). The Kd-value is 3.6 nM for TCDF with the rat receptor protein. It is concluded from these data that metabolic activation of DES is not involved in the mechanism of hepatocarcinogenesis in this animal tumor model.     

The effects of benzoflavones on polycyclic hydrocarbon metabolism and skin tumor initiation.

The effects of benzoflavones on skin tumor initiation by polycyclic hydrocarbons and epidermal aryl hydrocarbon hydroxylase were investigated. 7,8-Benzoflavone (7,8-BF) was found to be a potent inhibitor of the inhibition of skin tumors by 3-methylcholanthrene (MC) as well as 7,12-dimethylbenz(a)anthracene (DMBA). 5,6-Benzoflavone(5,6-BF) inhibited tumor initiation by MC and DMBA, but to a lesser degree than 7,8-BF. Dose-response studies of the capacity of 7,8-BF to inhibit DMBA tumor initiation revealed that 7,8-BF was an effective inhibitor at 2.5 microgram and a maximum inhibition of 90% occurred at 100 microgram of 7,8-FB. The tumor initiating ability of 7-hydroxymethyl-12-methylbenz(a)anthracene (7-OHMe-12MeBA) was not inhibited by 7,8-BF. Epidermal aryl hydrocarbon(benzo(a)pyrene hydroxylase(AHH) was increased by 5,6-BF and either had no effect or was slightly inhibited by 7,8-BF when given either topically or i.p. Both flavones when added directly to the assay tubes inhibited the in vitro epidermal AHH activity from control and MC pretreated mice by greater than 75%. When added in vitro, 7,8-BF and 5,6-BF inhibited epidermally mediated covalent binding of radioactive DMBA and dibenz(a,h)anthracene to DNA by 50% or more. The inhibition of skin tumor initiation by 7,8-BF and 5,6-BF appears to be partially related to its ability to inhibit the formation of electrophilic intermediates.     

Effect of pretreatment of male Syrian golden hamsters with 7,8-benzoflavone and with diethylstilbestrol on P-450 isoenzyme activities and on microsomal diethylstilbestrol metabolism

Combined treatment of male Syrian golden hamsters with the synthetic estrogen diethylstilbestrol (DES) and 7,8-benzoflavone (7,8-BF) gives rise to a high incidence of hepatocellular carcinomas, whereas no such tumors are formed with DES alone nor with 7,8-BF alone. To determine whether alterations in DES metabolism may account for the observed hepatocarcinogenicity, we have studied the effect of pretreatment with 7,8-BF alone, DES alone and 7,8-BF plus DES on the levels of hepatic P-450 and cytochrome b5, on the activities of various P-450 isoenzymes and on microsomal DES metabolism. Hepatic P-450 content was significantly increased after pretreatment with 7,8-BF and decreased after DES, while combined pretreatment led to levels similar to those in untreated control animals. Hepatic cytochrome b5 was also elevated in 7,8-BF-treated hamsters; DES pretreatment had no effect, and combined pretreatment led to a slight increase. Four different substrates were used to probe P-450 isoenzyme activity. Aryl hydrocarbon hydroxylase (AHH), 7-ethoxycoumarin-O-deethylase (ECOD), 7-ethoxyresorufin-O-deethylase (EROD) and 7-pentoxyresorufin-O-dealkylase (PROD) were all elevated after 7,8-BF-pretreatment, while DES led to a decrease in these activities with the exception of AHH, where a transient increase which was observed after 8 and 20 weeks of pretreatment was back to control levels after 32 weeks. Combined pretreatment with 7,8-BF and DES led to an intermediate response (slight increase) with AHH, EROD and PROD, but not with ECOD, where a full induction comparable with that observed after 7,8-BF alone was elicited. In spite of the modulation of enzyme levels and activities observed after the various pretreatments, the metabolism of DES in microsomes from pretreated animals was virtually identical with that from controls. Therefore it is concluded that modulation of hepatic DES metabolism is not the reason for the observed hepatotumorigenicity; instead, it is speculated that 7,8-BF is the carcinogenic agent in this tumor model, and DES may act as a promotor.     

Extensive biodegradation of highly chlorinated biphenyl and Aroclor 1242 by Pseudomonas aeruginosa TMU56 isolated from contaminated soils

A bacterial strain, designated TMU56, was isolated from soil that had been contaminated with electrical transformer fluid (Askarel) for over 35 years. The isolate was identified as Pseudomonas aeruginosa using its 16S rDNA sequence. This strain was found to grow on monochlorobiphenyls (CBs), including 2-chlorobenzoic acid and 4-chlorobenzoic acid. It was also found to grow on 2,4-, 2,5-, 2,2′-, and 4,4′-diCB, as well as on a wide range of other xenobiotic compounds. This is the first reported representative of the genus Pseudomonas that is capable of growing on 2,4,4′-triCB, 2,2′,5,5′-tetraCB and 2,2′,4,4′,5,5′-hexaCB as sole carbon sources. Washed benzoate-grown cells were able to degrade 89% and 56% of 2,4-diCB and 2,2′,4,4′,5,5′-hexaCB, respectively. Gas chromatography analysis of individual congeners in Aroclor 1242 (200 ppm) following a 4-day incubation showed 73.3% degradation of PCBs without the need for biphenyl as an inducer. The strain exhibited no noticeable specificity for the percentage of congener transformation or degree of chlorination.     

Mutagenicities of 2,4- and 2,6-dinitrotoluenes and their reduced products in Salmonella typhimurium nitroreductase- and O-acetyltransferase-overproducing Ames test strains

Mutagenicities of 2,4- and 2,6-dinitrotoluene (2,4-and 2,6-DNT), and reduced metabolites formed by the incubation of 2,4- and 2,6-DNT with Salmonella typhimurium TA98, were tested using S. typhimurium YG strains possessing high level of nitroreductase (NR) and/or O-acetyltransferase (OAT) activities. All compounds tested showed greatest mutagenic activities toward strains YG1041 and YG1042, which possess high levels of NR and OAT activities. The relative mutagenic activities of 2,4-DNT and its related compounds toward YG1041 and YG1042 were aminonitrotoluenes (2A4NT, 4A2NT)<2,4-DNT<2,2′-dimethyl-5,5′-dinitroazoxybenzene (2,2′-DM-5,5′-DNAOB)4-hydroxylamino-2-nitrotoluene (4HA2NT)4,4′-dimethyl-3,3′-dinitroazoxybenzene (4,4′-DM-3,3′-DNAOB), and aminonitrotoluenes (2A4NT, 4A2NT)<2,4-DNT<4HA2NT4,4′-dimethyl-3,3′-dinitroazoxybenzene (4,4′-DM-3,3′-DNAOB)<2HA4NT, respectively. In addition, the relative mutagenic activities of 2,6-DNT and its related compounds toward YG1041 and YG1042 were 2,6-DNT<2-hydroxylamino-6-nitrotoluene (2HA6NT)<2,2′-dimethyl-3,3′-dinitroazoxybenzene (2,2′-DM-3,3′-DNAOB), and 2-amino-6-nitrotoluene (2A6NT)<2,6-DNT<2HA6NT, respectively. These results, together with previous findings, suggested that aminohydroxylamino dimethylazoxybenzenes or aminohydroxylamino dimethylazobenzenes produced either by the reduction of hydroxylaminonitrotoluenes or by the reduction of dimethyl dinitroazoxybenzenes are active metabolites responsible for the mutagenic activities of 2,4- and 2,6-DNT.     

Comparative biochemical studies of carotenoids in sea cucumbers

The results of an investigation of the carotenoids in the seven species of sea cucumber (Stichopus japonicus, Holothuria leucospilota, H. moebi and H. pervicax of the order Aspidochirotida, Cucumaria japonica, C. echinata and Pentacta australis of the order Dendrochirotida), from the comparative biochemical point of view, are reported. β-Carotene, β-echinenone, canthaxanthin, phoenicoxanthin and astaxanthin were common in all the sea cucumbers examined. A series of novel marine carotenoids (cucumariaxanthin A, B and C) was obtained from the sea cucumbers of the order Dendrochirotida, while they could not be found from those of the order Aspidochirotida. Significant differences in the carotenoid patterns of the two orders were also observed. The structures of cucumariaxanthin A, B and C have been determined, by chemical and spectroscopic investigations, to be (9Z,9′Z)-5,6,5′,6′-tetrahydro-β,β-carotene-4,4′-dione, (9Z,9′Z)-4′-hydroxy-5,6,5′,6′-tetrahydro-β,β-caroten-4-one, and (9Z,9′Z)-5,6,5′,6′-tetrahydro-β,β-carotene-4,4'-diol, respectively. From the experimental results of carotenoids in the sea cucumbers examined, an oxidative metabolic pathway for β-carotene to astaxanthin, and a new reductive and isomeric metabolic pathway for canthaxanthin to cucumariaxanthin C (via cucumariaxanthin A and B) are proposed.     

PAF-antagonistic bicyclo[3.2.1]octanoid neolignans from leaves of Ocotea macrophylla Kunth. (Lauraceae)

Di-nor-benzofuran neolignan aldehydes, Δ⁷-3,4-methylenedioxy-3′-methoxy-8′,9′-dinor-4′,7-epoxy-8,3′-neolignan-7′-aldehyde (ocophyllal A) 1, Δ⁷-3,4,5,3′-tetramethoxy-8′,9′-dinor-4′,7-epoxy-8,3′-neolignan-7′-aldehyde (ocophyllal B) 2, and macrophyllin-type bicyclo[3.2.1]octanoid neolignans (7R, 8R, 3′S, 4′S, 5′R)-Δ⁸′-4′-hydroxy-5′-methoxy-3,4-methylenedioxy-2′,3′,4′,5′-tetrahydro-2′-oxo-7.3′,8.5′-neolignan (ocophyllol A) 3, (7R, 8R, 3′S, 4′S, 5′R)-Δ⁸′-4′-hydroxy-3,4,5′-trimethoxy-2′,3′,4′,5′-tetrahydro-2′-oxo-7.3′,8.5′-neolignan (ocophyllol B) 4, (7R, 8R, 3′S, 4′S, 5′R)-Δ⁸′-4′-hydroxy-3,4,5,5′-tetramethoxy-2′,3′,4′,5′-tetrahydro-2′-oxo-7.3′,8.5′-neolignan (ocophyllol C) 5, as well as 2′-epi-guianin 6 and (+)-licarin B 7, were isolated and characterized from leaves of Ocotea macrophylla (Lauraceae). The structures and configuration of these compounds were determined by extensive spectroscopic analyses. Inhibition of platelet activating factor (PAF)-induced aggregation of rabbit platelets were tested with neolignans 1–7. Although compound 6 was the most potent PAF-antagonist, compounds 3–5 showed some activity.     

Prenylated flavanones from the twigs of Dorstenia mannii

Investigation of the twigs of Dorstenia mannii gave 6,8-diprenyl-5,7,3′4′-tetrahydroxyflavanone and four new prenylated flavanones, named dorsmanins E-H and characterized as 5,6-7,8-bis-(2,2-dimethylchromano)-3′,4′-dihydroxyflavanone, 7,8-[2″-(1-hydroxy-1-methylethyl)-dihydrofurano]-6-prenyl-5,3′,4′-trihydroxyflavanone, 6,7-[2″-(1-hydroxy-1-methylethyl)dihydrofurano]-8-prenyl-5,3′,4′-trihydroxyflavanone and 6-prenyl-8-(2-hydroxy-3-methylbut-3-enyl)-5,7,3′,4′-tetrahydroxyflavanone, respectively, on the basis of spectral analysis and chemical evidence for the chromano derivative.     

Effect of various chemicals on the metabolism of benzo(a)pyrene by cultured rat colon

The effect of various co- and anti-carcinogens of colon carcinogenesis on the metabolism of benzo(a)pyrene (BP) in cultured rat colon is reported. Rat colon enzymatically converted BP into metabolites which bind to cellular macromolecules i.e., DNA and protein. Activity of aryl hydrocarbon hydroxylase (AHH) activity and binding levels of BP to macromolecules were higher in the descending colon when compared to other segments. The major metabolites of BP, extractable with ethylacetate, were quinones, tetrols, 7,8-diol and a peak containing 9,10-dihydroxy-9,10-dihydrobenzo(a)pyrene and 7,8,9-trihydroxy-7,8-dihydrobenzo(a)pyrene. The binding levels of BP to DNA and protein in the explant was lowered by co-incubation with 7,8-benzoflavone (7,8-BF) (3.6 and 18.0 μM), a known inhibitor of AHH, and with disulfiram (100 μM), an anti-oxidant. The absence of vitamin A in the media also resulted in a lower level of BP binding to DNA and protein and in lower activity of AHH. Pretreatment with known inducers of AHH such as phenobarbital (PB) or benz(a)anthracene (BA), did not have any significant effect on the binding levels of BP to DNA or on the AHH activity. of the bile acids investigated only taurodeoxycholic acid significantly increased the binding level of BP to DNA.     

Optical resolution of hexamethylbiphenol by cholesterol esterase and porcine pancreas lipase

Both enantiomers of 2,2′-dihydroxy-4,4′,5,5′,6,6′-hexamethybiphenyl (2), a potentially useful chiral synthon, were obtained with >99% ee in high enantioselectivity by cholesterol esterase or porcine pancreas lipase (PPL)-mediated hydrolysis of the corresponding (±)-dipentanoate or (±)-dihexanoate, respectively. Absolute configuration of (S)-3-bromo-2,6′-dimethoxy-4,5,6,2′,3′,4′-hexamethyl-biphenyl (2h) was determined by X-ray analysis.     

Stimulation of adenosine 3′,5′-monophosphate formation in mononuclear leukocytes by toad venoms

The content of adenosine 3′,5′-monophosphate in human mononuclear leukocytes was enhanced 3–5-times by venoms obtained from African toad (Bufo africanus), American toad (Bufo americanus), Colorado river toad (Bufo arenarum) and Marine toad (Bufo marinus) at 25 μg/ml for 5 min of incubation at 37°C. The maximum stimulation was observed after 1–5 min of incubation. The half-maximal stimulation was observed at 0.1 μg/ml venom obtained from Colorado river toad (Bufo arenarum). The increased content of adenosine 3′,5′-monophosphate in the mononuclear leukocytes persisted without significant change for at least 30 min of incubation at 37°C.     

Differences in liver homogenates from Donryu, Fischer, Sprague-Dawley and Wistar strains of rat in the drug-metabolizing enzyme assay and the salmonella/hepatic S9 activation test

Comparison studies for detecting differences between liver microsome and S9 preparations from 4 strains (Donryu, Fischer, Sprague-Dawley, Wistar) of young male rats were carried out with pretreatment of the animals by inducers such as PCBs and PB plus 5,6-BF. Each microsome fraction was assayed for the enzymic activity of metabolism of model substrates such as aniline, benzophetamine, BP, DMN and 7-ethoxycoumarin. The hepatic S9 sample was also compared, as regards its metabolizing ability to activate 9 pre-mutagens (2AA, AAF, o-AAT, BP, DAB, DMBA, DMN, m-PDA, quinoline) to directly acting mutagens in the Salmonella/hepatic S9 activation test by using TA98, TA100 and TA1537 strains with or without cytochrome P450 inhibitors (SKF-525A, metyrapone, 7,8-benzoflavone).In the enzymic assay with PCBs-induced microsomes, BP hydroxylation revealed a strain-specific difference: the microsomes from Fischer and Wistar rats were more effective for metabolizing BP than those from the other strains of rat. The effect of induction by PB plus 5,6-BF for Fischer rats showed relatively higher enzymic activity in the same induction group. Other microsomes prepared from rats with and without induction by PB plus 5,6-BF did not show a clear-cut strain dependency in the enzymic activities assayed.In the mutation experiments with hepatic S9 samples, the examination of DAB and quinoline revealed a marked strain difference when S9 samples prepared from PCBs-pretreated and PB-plus-5,6-BF-induced rats were used: the S9 sample from Fischer rats was available for activating the two pre-mutagens to directly acting mutagens. No marked difference in the metabolic activation of the remaining 7-pre-mutagens was observed on other S9 preparations.In examinations of mutagenicity activities with the use of three inhibitors, the two S9 preparations made with the two induction methods showed inhibition profiles closely similar to each other. However, there were minor differences in the profiles by these inhibitors.From these findings it was concluded that Fischer rat-liver S9 is useful for detecting mutagens in the metabolic activation test, when induction by PB plus 5,6-BF was used in the Ames Salmonella test.     

Inhibition of hepatic aryl hydrocarbon hydroxylase by 3-methylcholanthrene, 7,8-benzoflavone and other inducers added in vitro.

A close correlation has been observed between the ability of aromatic polycyclic hydrocarbons and 7,8-benzoflavone (7,8-BF) to induce hepatic aryl hydrocarbon hydroxylase (AHH) in vivo and to inhibit the induced enzyme system in vitro. The activity of this mono-oxygenase was measured by the conversion of 14C-labeled dimethylbenz(a)anthracene (DMBA) or benzo(a)pyrene (BP) to water-soluble products by rat liver preparations (8000 X g supernatant). DMBA as substrate had the advantage over BP in giving a wider range of ethyl acetate-soluble metabolites and allowing the observation of changes in the pattern of these products following injection or addition of the inducing agents. This property was used to detect low concentration (0.1 muM) of polycyclic hydrocarbons which are strong AHH inducers and which may also be carcinogenic. The liver preparation was active for several months when stored at --20 degrees. A possible mechanism of action for the in vitro behaviour of polycyclic hydrocarbons and 7,8-BF towards AHH is proposed.     

Hydroxylation of naringin by Trichoderma harzianum to dramatically improve its antioxidative activity

Transforming naringin using the mycelium of Trichoderma harzianum CGMCC 1523 produces two metabolites, 3′,4′,5,7-tetrahydroxy flavanone-7-rhamnoglucoside (3′-OHN) and 3′,4′,5′,5,7-pentahydroxy flavanone-7-rhamnoglucoside (3′,5′-DOHN), both of which were characterized by ESI–MS, ¹H NMR and ¹³C NMR analyses. The time course of the biotransformation by T. harzianum showed that 3′-OHN and 3′,5′-DOHN appeared simultaneously at 6 h, and the conversion yield (32.6%) of 3′,5′-DOHN was higher (10.6%) than that of 3′-OHN at 56 h. The optimal biotransformation temperature was 30 °C, the optimal pH was 5.0, and the optimal concentration of naringin was 400 mg/l. The bigger volume of biotransformation mixture and lower shaking speed did not favor hydroxylation reactions. The radical scavenging activity of naringin at 2000 μM was 11.1%, whereas activity of 3′-OHN at 100 μM could reach 38.4%, which is 68.6 times more than naringin. Antioxidative activity of 3′,5′-DOHN was increased 13.5% at 100 μM compared to 3′-OHN.     

Cyclic 3′,5′-AMP phosphodiesterase in Physarum polycephalum: II. Kinetic properties

The effect of several inhibitors of the enzyme cyclic 3′,5′-AMP phosphodiesterase as chemoattractants in Physarum polycephalum was examined. Of the compounds tested, 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (Roche 20-1724/001) and 1-ethyl-4-(isopropylidinehydrazino)-1H-pyrazolo-(3,4-b)-pyridine-5-carboxylic acid ethyl ester, hydrochloride (Squibb 20009) were the most potent attractants. 3-Isobutyl-1-methyl xanthine, theophylline, and morin (a flavanoid) were moderate attractants and sometimes gave negative chemotaxis at high concentrations. Cyclic 3′,5′-AMP was an effective, but not potent attractant. A repellent effect following the positive chemotactic action was sometimes observed with cyclic 3′,5′-AMP at concentrations as high as 1 · 10⁻² M. Dibutyryl cyclic AMP appeared to be a somewhat more potent attractant than cyclic 3′,5′-AMP. The 8-thiomethyl and 8-bromoderivatives of cyclic AMP, which are poorly hydrolyzed by the phosphodiesterase, were not attractants in Physarum. Possible participation of cyclic 3′,5′-AMP in the directional movement in P. polycephalum is discussed.     

Induction of SCEs in CHL cells by dichlorobiphenyl derivative water pollutants, 2-phenylbenzotriazole (PBTA) congeners and river water concentrates

We recently identified dichlorobiphenyl (DCB) derivatives and 2-phenylbenzotriazole (PBTA) congeners as major mutagenic constituents of the waters of the Waka River and the Yodo River system in Japan, respectively. In this study we examined sister chromatid exchange (SCE) induction by two dichlorobiphenyl derivatives, 3,3′-dichlorobenzidine (DCB, 4,4′-diamino-3,3′-dichlorobiphenyl) and 4,4′-diamino-3,3′-dichloro-5-nitrobiphenyl (5-nitro-DCB); three PBTA congeners, 2-[2-(acetylamino)-4-[bis(2-methoxyethyl)amino]-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-1), 2-[2-(acetylamino)-4-[N-(2-cyanoethyl)ethylamino]-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-2), and 2-[2-(acetylamino)amino]-4-[bis(2-hydroxyethyl)amino]-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-6); and water concentrates from the Waka River in Chinese hamster lung (CHL) cells. Concentration-dependent induction of SCE was found for all DCBs and PBTAs examined in the presence of S9 mix, and statistically significant increases of SCEs were detected at 2 μg per ml of medium or higher concentrations. SCE induction of MeIQx was examined to compare genotoxic activities of these water pollutants. According to the results, a ranking of the SCE-inducing potency of these compounds is the following: 5-nitro-DCB ≈ MeIQx > PBTA6 > PBTA-1 ≈ PBTA-2 > DCB.Water samples collected at a site at the Waka River showed concentration-related increases in SCEs at 6.25–18.75 ml-equivalent of river water per ml of medium with S9 mix. The concentrations of 5-nitro-DCB and DCB in the river water samples were from 2.5 to 19.4 ng/l and from 4100 to 18,900 ng/l, respectively. However, these chemicals showed only small contribution to SCE induction by the Waka River water.     

The genetics of aryl hydrocarbon hydroxylase induction in mice: A single gene difference between C57BL/6J and DBA/2J

Twenty-one inbred strains of mice were surveyed for inducibility of hepatic aryl hydrocarbon hydroxylase (AHH) activity by the carcinogen 3-methylcholanthrene (MC). In 11 strains given MC, AHH activity increased 1.3- to 5-fold (inducible), whereas ten strains responded with a less than 0.5-fold increase (noninducible). Neither the inducible nor the noninducible class was homogeneous, and in each considerable variation was found in both the basal activity of AHH and the response to MC. Strains DBA/2J and C57BL/6J were chosen to represent the noninducible and inducible classes, respectively. In the crosses (C57BL/6 × DBA/2)F₁ × DBA/2 and (C57BL/6 × DBA/2)F₂, inducibility segregated as a single autosomal dominant gene. The gene symbols Ahh ⁱ and Ahh ⁿ are proposed for the alleles present in C57BL/6J and DBA/2J, respectively. No genetic linkage was found between the Ahh locus and the following loci: b, d, Es-1, Es-3, Gpd-1, Hbb, Id-1, Pgm-1, and sex. Some implications of this work in the study of mammalian enzyme induction and chemically induced carcinogenesis are discussed. There is a positive correlation between AHH inducibility and the development of an inflammatory response to the topical application of the carcinogen 7,12-dimethylbenzanthracene.     

Methylenedioxy- and methoxyflavones from Melicope coodeana syn. Euodia simplex

Three new natural products, 3,8-dimethoxy-5,7-dihydroxy-3′,4′-methylenedioxyflavone, 3,6,8-trimethoxy-5,7-dihydroxy-3′,4′-methylenedioxyflavone and 3,6,8,3′,4′-pentamethoxy-5,7-dihydroxyflavone were isolated from Melicope coodeana syn. Euodia simplex (Rutaceae) along with 3,6,3′-trimethoxy-5,7,4′-trihydroxyflavone and 3,3′-dimethoxy-5,7,4′-trihydroxyflavone. The structural assignments are based on ¹H and ¹³C NMR data, including discussion of the chemical shifts of C-2 in 3,5-dihydroxy- and 3-methoxy-5-hydroxyflavones. The presence of highly methoxylated and methylenedioxyflavones is characteristic of the genus Melicope, and the present findings support the recent transfer of Euodia simplex to Melicope.