Antimutagenic and mutagenic activities of some terpenes in the bacterial reverse mutation assay

The mutagenic and antimutagenic effects of linalool, linalyl acetate and beta-caryophyllene were evaluated by the bacterial reverse mutation assay on Salmonella typhimurium TA 98 and TA 100, and on Escherichia coli WP2uvrA strains. Neither linalool nor beta-caryophyllene showed mutagenicity, but linalyl acetate induced a statistically significant increase in the number of revertant colonies in WP2uvrA, both with and without S9 mixture. Linalool was devoid of antimutagenic activity against 2-nitrofluorene (2NF), sodium azide (SA), methyl methane sulfonate (MMS) and 2-aminoanthracene (2AA). In contrast, beta-caryophyllene showed a strong antimutagenic activity against 2NF: at the maximum concentration tested (6.40mg/plate) the number of 2NF-induced revertant colonies was reduced by 83.9%. beta-Caryophyllene also showed to counteract the mutagenicity of SA (in TA 100), MMS and 2AA (in WP2uvrA): the effect was weak against SA (inhibition lower than 25%) and moderate against MMS and 2AA (up to 30.5%). The antimutagenic activity of beta-caryophyllene observed here suggests further studies to evaluate its possible chemopreventive properties.     

Antimutagenicity of some citrus fruits in Salmonella typhimurium

The antimutagenic effect of 10 citrus fruit juices was observed against the mutagenicity of N-nitro-o-phenylenediamine (NPD) in TA97a and sodium azide in TA100 tester strains of Salmonella typhimurium using the Ames test. It was noticed that the juices of all these fruits reduced significantly the NPD and sodium azide induced revertant colonies. The inhibitory activity was enhanced if the mutagen and juice were co-incubated for about 30 min at 37 degrees C prior to performing the mutagenicity assay. Dilution with distilled water led to the reduction in the inhibitory activity. The antimutagenic activity of synthetic ascorbic acid or citric acid or combined ascorbic acid and citric acid was also seen. But the results with fruit juices tempted us to believe that in addition to ascorbic acid and citric acid, the presence of other factor(s) possessing antimutagenic properties cannot be ruled out.     

An investigation on the antimutagenic properties of South African herbal teas

The antimutagenic properties of South African herbal teas were investigated using the Salmonella typhimurium mutagenicity assay. Aqueous extracts of fermented and unfermented rooibos tea (Aspalathus linearis) and honeybush tea (Cyclopia intermedia) both possess antimutagenic activity against 2-acetylaminofluorene (2-AAF) and aflatoxin B(1) (AFB(1))-induced mutagenesis using tester strains TA98 and TA100 in the presence of metabolic activation. A far less inhibitory effect was noticed against the direct acting mutagens, methyl methanesulfonate (MMS), cumolhydroperoxide (CHP), and hydrogen peroxide (H(2)O(2)) using TA102, a strain designed to detect oxidative mutagens and carcinogens. Depending on the mutagen used, the unfermented tea exhibited the highest protective effect. A similar response regarding the protection against mutagenesis was obtained when utilising different variations of the double layer Salmonella assay. The double layer technique proved to be more effective to detect the protective effect of the different tea preparations against the direct acting mutagens. With respect to indirect mutagens, the highest protection was noticed when the carcinogen was metabolically activated in the presence of the tea extract as compared with when the tea extract was incubated in a separate layer with the bacteria. The current data suggest that two mechanisms seem to be involved in the antimutagenicity of the tea extracts towards carcinogens that require metabolic activation: (i) the tea components may interfere with cytochrome P450-mediated metabolism of these mutagens and (ii) the direct interaction between the tea constituents, presumably the polyphenolic compounds, with the promutagens and/or the active mutagenic metabolites. However, the mild and/or lack of protection and in some cases even enhancement of mutagenesis induced by direct acting or oxidative mutagens, provide new perspectives regarding the role of the polyphenolic compounds known to exhibit antioxidant properties, in the protection against mutagenesis in the Salmonella assay. The present study provides the first evidence on the antimutagenic activity of honeybush tea and further evidence on the antimutagenicity of rooibos tea.     

Antimutagenicity of propionic acid bacteria.

The antimutagenic effect of dialysed cell extracts of 4 strains of propionic acid bacteria was examined against the mutagenicity of sodium azide in the TA1535 tester strain of Salmonella typhimurium using the Ames test. It was noted that dialysates of 2 strains of Propionibacterium shermanii, P. pentosaceum and P. acnes, significantly reduced sodium azide-induced revertants. The dialysate of propionic acid cocci did not show an antimutagenic effect. The inhibitory activity was enhanced if the mutagen and extract were coincubated for 20 min prior to performing the mutagenicity assay. Antimutagenicity of dialysates from P. shermanii VKM-103 against MNNG and 9-aminoacridine was shown in S. typhimurium strains TA1535 and TA97. The antimutagenic activity was found in the protein fraction of the cell extract of P. shermanii. The proteins of the dialysate of P. shermanii were separated using a Toyopearl gel column into 3 main peaks according to their molecular weights. The antimutagenic activity towards sodium azide was found in the second and the third peaks. We suggest that dialysates of the cells of propionic acid bacteria contain several kinds of antimutagenic substances with different molecular weights.     

Antimutagenic effects of mushrooms

A heat-resistant factor in ethanol extracts of the fungus Craterellus cornucopioides completely inhibited the mutagenicity of aflatoxin B1, benzo[a]pyrene, the acridine half mustard ICR-191 and 2-nitrofluorene in a forward-mutation system using Salmonella typhimurium TM677 (screening for 8-azaguanine resistance). There was no inhibitory effect on the mutagenic activity of 4-nitroquinoline-N-oxide, methyl methanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine. Experiments performed to elucidate the mechanism of the antimutagenic effect showed that neither an alteration of cell viability nor an interference with the excision-repair and the inducible SOS-repair system was involved. The conceivable mechanisms for the antimutagenicity of the ethanol extract include direct chemical interaction with the mutagen and/or inhibition of the activation process in the case of the promutagens. The antimutagenic activity of Craterellus cornucopioides is not unique among mushroom species. The ethanol extracts of 6 other mushrooms showed a similar antimutagenic activity.     

Antimutagenic activity of methanolic extracts of four ayurvedic medicinal plants

Methanolic extracts of Acorus calamus (Rhizome), Hemidesmus indicus (Stem), Holarrhena antidysenterica (Bark) and Plumbago zeylanica (Root), were tested for their antimutagenic potential. These extracts, at tested concentrations, showed no sign of mutagenicity to Salmonella typhimurium tester strains. The extracts of the plants exhibited varying level of antimutagenicity. At a dose of 100 microg/plate, the extracts exhibited the inhibition of His+ revertants from 18.51% to 82.66% against direct acting mutagens, methyl methanesulphonate (MMS) and sodium azide (NaN3) induced mutagenicity in Salmonella tester strains TA 97a, TA 100, TA 102 and TA 104. However, at lower concentrations (25 and 50 mcirog/plate) of the plant extracts, a decrease in antimutagenic activity was recorded. Dose dependent antimutagenic activity of the extracts is also evident from linear regression analysis of the data. The over all antimutagenic potential of above four extracts was found to be in order of A. calamus > H. indicus > H. antidysenterica > P. zeylanica. Further, total phenolic content of these extracts did not correlate with its antimutagenic activity in A. calamus and P. zeylanica.     

Sesamol exhibits antimutagenic activity against oxygen species mediated mutagenicity

The effects of sesamol, a phenolic compound responsible for the high resistance of sesame oil to oxidative deterioration as compared with other vegetable oils, have been investigated after mutagen treatment in various strains of Salmonella typhimurium. Sesamol was shown to exhibit strong antimutagenic effects in the Ames tester strains TA100 and TA102. The TA102 strain has been shown to be highly sensitive to reactive oxygen species. Mutagenicity was induced by the generation of oxygen radicals by tert-butylhydroperoxide (t-BOOH) or hydrogen peroxide (H(2)O(2)); therefore, the antimutagenic property of sesamol was attributed to its antioxidant properties. The superoxide and hydroxyl radical scavenging capabilities have further been elucidated using in vitro test systems. It was further shown to have a desmutagenic effect on t-BOOH-induced mutagenesis in TA102 strain. Sesamol also inhibited the mutagenicity of sodium azide (Na-azide) in TA100 tester strain while it had no effect on nitroquinoline-N-oxide (NQNO)-induced mutagenesis in TA98 strain of Salmonella typhimurium. Since active oxygen species are involved in multiple stage processes of carcinogenicity, this compound may also exhibit anticarcinogenic properties.     

Evaluation of antioxidant activities and antimutagenicity of turmeric oil: a byproduct from curcumin production

Curcumin removed turmeric oleoresin (CRTO) was extracted with hexane concentrated to get turmeric oil, and that was fractionated using silica gel column chromatography to obtain three fractions. These fractions were analyzed by GC and GC-MS. Turmeric oil contained aromatic turmerone (31.32%), turmerone (15.08%) and curlone (9.7%), whereas fractions III has aromatic turmerone (44.5%), curlone (19.22%) and turmerone (10.88%) as major compounds Also, oxygenated compounds (5,6,8-10) were enriched in fraction III. Turmeric oil and its fractions were tested for antioxidant activity using the beta-carotene-linoleate model system and the phosphomolybdenum method. The fraction III showed maximum antioxidant capacity. These fractions were also used to determine their protective effect against the mutagenicity of sodium azide by means of the Ames test. All the fractions and turmeric oil exhibited a markedly antimutagenicity but fraction III was the most effective. The antioxidant effects of turmeric oil and its fractions may provide an explanation for their antimutagenic action.     

Chemical investigation of different extracts and essential oil from the tubers of (Tunisian)Cyperus rotundus. Correlation with their antiradical and antimutagenic properties

The mutagenic potential of aqueous, Total Oligomers Flavonoids (TOF), ethyl acetate, and methanol extracts as well as essential oil (EO) obtained from tubers ofCyperus rotundus L. was assessed by “Ames assay”, usingSalmonella tester strains TA98 and TA100, and “SOS chromotest” usingEscherichia coli PQ37 strain with and without an exogenous metabolic activation system (S9). None of the different extracts showed a mutagenic effect. Likewise, the antimutagenicity of the same extracts was tested using the “Ames test” and the “SOS chromotest”. Our results showed thatC. rotundus extracts have antimutagenic effects withSalmonella typhimurium TA98 and TA100 strains towards the mutagen Aflatoxin B1 (AFB1), as well as withE. coli PQ37 strain against AFB1 and nifuroxazide mutagens. A free radical scavenging test was used in order to explore the antioxidant capacity of the extracts obtained from the tubers ofC. rotundus. TOF, ethyl acetate and methanol extracts showed an important free radical scavenging activity towards the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical. These extracts showed IC50 values of respectively 5, 20 and 65 μg/ml. The beneficial effects of TOF, ethyl acetate, methanol and essential oil extracts ofC. rotundus have been assessed by antioxidant and antimutagenic activities.     

Antimutagenic effects of natural phenolic compounds in beans

Polyphenols in fruits, vegetables (e.g., flavonols like quercetin) and tea (e.g., catechins such as epigallocatechin gallate) are good antioxidants with antimutagenic and anticarcinogenic properties. In the present study, the Salmonella typhimurium tester strain YG1024 was used in the plate-incorporation test to examine the antimutagenic effect of phenolic compounds, extracted from common beans (Phaseolus vulgaris), on 1-NP and B[a]P mutagenicity. Dose-response curves for 1-NP and B[a]P were obtained; the number of net revertants/plate at the peak mutagenic dosage were 880 for 1-NP and 490 for B[a]P. For the antimutagenicity studies doses of 0.1 microg/plate and 2 microg/plate for 1-NP and B[a]P, respectively, were chosen. We obtained a dose-response curve of ellagic acid (EA) against B[a]P and 1-NP mutagenicity. To test the bean extract, a dose of 300 microg/plate of EA was chosen as the antimutagenic control. The EA and bean extracts were not toxic to the bacteria at the concentrations tested. The inhibitory effects of the bean extracts and EA against B[a]P mutagenicity were dose-dependent. The percentages of inhibition produced against B[a]P (2 microg/plate) using 300 microg/plate of EA and for the extracts 500 microg equivalent catechin/plate were 82%, 83%, 81% and 83% for EA, water extract, water/methanol extract and methanol extract, respectively. However, for 1-NP mutagenicity, only the methanolic extract from beans showed an inhibitory effect. These results suggest that common beans, as other legumes, can function as health-promoting foods.     

Chemical composition, antioxidant and antimicrobial activities of essential oil of Thymus algeriensis wild-growing in Libya

The composition of essential oil isolated from Thymus algeriensis growing wild in Libya was analyzed by GC and GC-MS. The essential oil was characterized with thymol (38.50%) as the major component. The oil was screened for antioxidant activity using DPPH assay, and compared to thymol and carvacrol. Antioxidant activity was high, with the IC₅₀ of 0.299 mg/ml, compared to 0.403 and 0.105 mg/ml for thymol and carvacrol, and 0.0717 mg/ml for BHA. In addition, antimicrobial activity was tested against eight bacteria and eight fungi. T. algeriensis oil showed inhibitory activity against tested bacteria at 0.001–0.05 mg/ml, while bactericidal activity (MBC) was achieved at 0.0025–0.05 mg/ml. For antifungal activity MICs ranged 0.0005–0.025 mg/ml and MFC 0.001–0.05 mg/ml. High antimicrobial activity against the fungi in particular suggests that the essential oil of Thymus algeriensis could have a useful practical application.     

The antimutagenicity of 2-substituted selenazolidine-4-(R)-carboxylic acids

Selenium can have cancer chemopreventive activity, although the mechanism of action has not been well defined. Selenazolidine-4-(R)-carboxylic acids (SCAs) were devised as prodrugs of L-selenocysteine, to provide selenium in a form and at a concentration commensurate with cancer chemopreventive activity. In the present study, a series of selenazolidines has been evaluated in the Salmonella typhimurium TA98 tester strain and all were found to possess antimutagenic activity. There was little difference between the seven selenazolidines in their effectiveness against either benzo[a]pyrene (B[a]P) or 3,6-bis(dimethylamino)acridine (acridine orange), agents which differ in their requirement for mammalian enzyme bioactivation for mutagenicity. Antimutagenic activity against acridine orange was dependent on selenazolidine concentration, and EC50 values were in the 5-10 microM range. At 25 microM, the concentration tested in common for the two mutagens, the selenazolidines were more effective antimutagens against acridine orange than against B[a]P, with reductions in mutant frequency ranging from 54 to 71% for B[a]P and 79 to 93% for acridine orange. Efficacy against B[a]P was not enhanced when the concentration was increased to 50 microM. The similarity in efficacy among the selenazolidines against B[a]P mutagenicity, contrasted with inter-compound differences in their ability to inhibit S9 CYP1A activity. The CYP1A Ki values ranged from a low of 63 microM (2-[2'-hydroxyphenyl]SCA) to a high of 1.1mM (2-cyclohexylSCA), but all were above the concentration required to inhibit mutagenicity by 50%. Thus, all the SCAs possess antimutagenic activity against both B[a]P and acridine orange, the efficacy varies little between the individual selenazolidines, and for B[a]P, the efficacy is not proportional to the inhibitory effect on the mutagen bioactivating enzyme.     

Antimutagenic effects of the mushroom Agaricus blazei Murrill extracts on V79 cells

Agaricus blazei Murrill, a native mushroom in Brazil, has been widely consumed in different parts of the world due to its medicinal power. Its anticarcinogenic activity has been shown in experimental animals, and antimutagenic activity has been demonstrated only in Salmonella. In this work, the mutagenic and antimutagenic activities of mushroom teas of strains AB96/07, AB96/09 and AB97/11 were evaluated in Chinese hamster V79 cells, using the comet assay and the micronucleus test. The cells were treated with three different concentrations (0.05, 0.1 and 0.15) of teas prepared from a 2.5% aqueous solution, under three different temperatures: (1) room (20-25 degrees C); (2) ice-cold (2-8 degrees C); and (3) warm (60 degrees C). The teas were applied in co-, pre- and post-treatments in combination with the mutagen methyl methanesulfonate (MMS; 1.6x10(-4) and 4x10(-4)M). The duration of the treatment was 1h in the comet assay and 2h in the micronucleus test. The results showed that the mushroom was not mutagenic itself. Nevertheless, the mushroom is an efficient antimutagen against the induction of micronuclei by MMS in all concentrations and preparations tested. The observed reductions in the frequencies of micronuclei ranged from 61.5 (room temperature 0.1% tea in post-treatment) to 110.3% (co-treatment with warm and ice-cold 0.15% tea). In the comet assay, the antimutagenic activity was detected only when the cells were pre-treated with the following teas: warm 0.1 and 0.15%, room temperature 0.05% and ice-cold 0.1%. The results indicate that the mushroom A. blazei extracts are antimutagenic when tested in V79 cells.     

A monitoring technique of piezoelectric impedance analysis used in tea polyphenols (TP) antimutagenic characteristics

A new method was proposed for studying antimutagenic characteristics of Tea Polyphenols (TP) using piezoelectric impedance analysis during the Salmonella typhimurium strain TA100 growth process in the presence of mutagen 4-methylnitrosamino-1-(3-pyridyl)-1-butanone (abbreviated NNK). Compared with the general method of antimutagenic investigation, the proposed method provided real-time, multidimensional response information about the antimutagenic characteristics during the bacterial growth process. The results showed that TP was allowed to be antimutagenic at the lowest concentration of 0.25 μg/ml, and the inhibitory effect of TP presented a linear relationship with its dosage in the range of 0.25–2.5 μg/ml. And the relationship between the bacterial growth kinetic parameters and the TP dosage was also obtained. The present method provided a new approach for studying microbial antimutagenic characteristics.     

Assessment of the relationship between the antimutagenic action of riboflavin and glutathione and the levels of antioxidant enzymes

In this study the role of antioxidant enzymes on the antimutagenic actions of riboflavin and reduced glutathione against mutagenic potentials of 4-nitroquinoline 1-oxide and mitomycin C have been investigated. For this purpose the activities of catalase and superoxide dismutase enzymes have been determined in Salmonella typhimurium TA102 and TA100 strains preincubated with different combinations of 4-nitroquinoline 1-oxide, mitomycin C, riboflavin and reduced glutathione for thirty minutes. Also in part of the same samples, the mutagenicity has been determined for each combination of chemicals by using Salmonella preincubation test. The correlation between the levels of antioxidant enzymes and mutagenicity and antimutagenicity has been investigated.While riboflavin displayed a weakly antimutagenic effect on 4-nitroquinoline 1-oxide mutagenicity in TA102 and TA100 (0.25, 0.35 inhibition respectively), it did not have any effect on the strong mutagenicity of mitomycin C in both strains. Reduced glutathione, a well known antioxidant, had no antimutagenic effect against the mutagenicity of both compounds in TA102 and TA100 strains. The antioxidant enzymes, catalase and superoxide dismutase, seemed to have no direct effect on the antimutagenic action of riboflavin and mutagenic action of 4-nitroquinoline 1-oxide and mitomycin C because no change in the activities of catalase and superoxide dismutase was detected in relation to antimutagenicity of riboflavin and mutagenicity of 4-nitroquinoline 1-oxide and mitomycin C in both strains. It should be noted that many antimutagens have more than one mechanism of action and their effect depends on the mutagens being tested.     

The metabolic activation of 4-nitro-o-phenylenediamine by chlorophyll-containing plant extracts: the relationship between mutagenicity and antimutagenicity

Chlorophyllin, the sodium and copper salt of chloropyll, and chlorophyll a, and chlorophyll b were tested for their ability to inhibit the mutagenic activity of the direct-acting mutagen 4-nitro-o-phenylenediamine (NOP) and its plant-activated mutagenic enhancement. All three forms of chlorophyll were antimutagenic against both NOP and its plant-activated product, with chlorophyllin proving most effective. Chlorophyll-containing plant extracts, however, proved very efficient at activating NOP into a mutagen of greater potency. When these extracts were assayed for total chlorophyll content it was found that they contained far less chlorophyll than was required for an antimutagenic effect to occur. Thus, the balance between chemical mutagen activation and/or enhancement by chlorophyll-containing plant extracts and the potential antimutagenicity of these plant extracts is a function of chlorophyll concentration. The data presented here indicate that this balance must be taken into consideration in future studies investigating the efficacy of complex natural plant extracts as antimutagenic substances.     

The metabolic activation of 4-nitro-o-phenylenediamine by chlorophyll-containing plant extracts: The relationship between mutagenicity and antimutagenicity

Chlorophyllin, the sodium and copper salt of chloropyll, and chlorophyll a, and chlorophyll b were tested for their ability to inhibit the mutagenic activity of the direct-acting mutagen 4-nitro-o-phenylenediamine (NOP) and its plant-activated mutagenic enhancement. All three forms of chlorophyll were antimutagenic against both NOP and its plant-activated product, with chlorophyllin proving most effective. Chlorophyll-containing plant extracts, however, proved very efficient at activating NOP into a mutagen of greater potency. When these extracts were assayed for total chlorophyll content it was found that they contained far less chlorophyll than was required for an antimutagenic effect to occur. Thus, the balance between chemical mutagen activation and/or enhancement by chlorophyll-containing plant extracts and the potential antimutagenicity of these plant extracts is a function of chlorophyll concentration. The data presented here indicate that this balance must be taken into consideration in future studies investigating the efficacy of complex natural plant extracts as antimutagenic substances.     

Antifungal, phytotoxic and insecticidal properties of essential oil isolated from Turkish Origanum acutidens and its three components, carvacrol, thymol and p-cymene

The chemical composition of essential oil isolated by hydrodistillation from the aerial parts of Origanum acutidens was analyzed by GC-MS. Carvacrol (87.0%), p-cymene (2.0%), linalool acetate (1.7%), borneol (1.6%) and beta-caryophyllene (1.3%) were found to be as main constituents. Antifungal, phytotoxic and insecticidal activities of the oil and its aromatic monoterpene constituents, carvacrol, p-cymene and thymol were also determined. The antifungal assays showed that O. acutidens oil, carvacrol and thymol completely inhibited mycelial growth of 17 phytopathogenic fungi and their antifungal effects were higher than commercial fungicide, benomyl. However, p-cymene possessed lower antifungal activity. The oil, carvacrol and thymol completely inhibited the seed germination and seedling growth of Amaranthus retroflexus, Chenopodium album and Rumex crispus and also showed a potent phytotoxic effect against these plants. However, p-cymene did not show any phytotoxic effect. Furthermore, O. acutidens oil showed 68.3% and 36.7% mortality against Sitophilus granarius and Tribolium confusum adults, respectively. The findings of the present study suggest that antifungal and herbicidal properties of the oil can be attributed to its major component, carvacrol, and these agents have a potential to be used as fungicide, herbicide as well as insecticide.     

Antimutagenic and antioxidant/prooxidant activity of quercetin

The present study has been performed to evaluate the antimutagenic activity of quercetin, ascorbic acid and their combination against an oxidative mutagen. An effort was also made to correlate this activity to the in vitro antioxidant activity of these agents. Antimutagenicity testing was done in Ames Salmonella Assay system using Salmonella typhimurium TA102 against t-butylhydroperoxide as an oxidative mutagen. In vitro antioxidant scavenging activity was tested for DPPH free radical, superoxide anion, hydrogen peroxide and hydroxyl radical in their specific test systems. Quercetin (0.5-8 nmole/plate) and ascorbic acid (0.1-100 micromole/plate) showed significant effect. Quercetin (4 and 8 nmole/plate) when combined with ascorbic acid (500 nmole/plate) showed an increase in the antimutagenic activity. In vitro antioxidant activity of quercetin was better than ascorbic acid in all the test systems used. The study indicated that the antimutagenic activity of quercetin was not solely accountable by its antioxidant nature. However, in vitro free radical scavenging activity of quercetin correlated well with the antimutagenic activity.     

Evaluation of the antimutagenic potential of chitosan oligosaccharide: Rec, Ames and Umu tests

Two types of chitosan oligosaccharides (COS), COS I (1-kDa < MW < 3-kDa) and COS II (3-kDa < MW < 5-kDa), were tested for antimutagenic activities against chemical mutagens using Umu gene expression, Ames, and Bacillus subtilis Rec mutagenicity tests. At the highest chitosan oligosaccharide dose (1 mg) tested, mutagenic activity of indirect-acting mutagen was inhibited by 50% in the Umu gene expression system and in the Ames test. Chitosan oligosaccharide (0.01, 0.1 and 1 mg) also suppressed 4-nitroquinoline-N-oxide (NQO)-induced mutagenicity in the B. subtilis Rec^– assay.