A phytotherapeutic extract of Equisetum myriochaetum is not genotoxic either in the in vivo wing somatic test of Drosophila or in the in vitro human micronucleus test

Equisetum myriochaetum is a Mexican plant used in folk medicine to treat kidney diseases and type 2 diabetes mellitus. The main constituents of the phytoextract are flavonol glycosides (kaempferol), phytoesterols and carbohydrates. In this study, phytotherapeutic extracts from Equisetum myriochaetum were investigated for genotoxicity in the in vivo wing spot test in Drosophila melanogaster and in the in vitro human micronucleus test. No acute toxicity of the phytoextract could be determined in Drosophila or in human lymphocytes in culture, ranging from 0.78 microg/ml to 3700 microg/ml for the wing assay and between 12.5 microg/ml and 500 microg/ml for the micronucleus test. The Drosophila wing somatic mutation and recombination test (SMART) was applied in the standard version with basal biotransformation activity as well as in a variant version with increased cytochrome P450-dependent bioactivation capacity. The ranges of exposure concentrations for these genotoxicity experiments were between 0.78 microg/ml and 500 microg/ml. The human micronucleus test in vitro was performed with cultured lymphocytes obtained from four healthy donors. The concentrations assayed for these experiments ranged from 12.5 microg/ml to 500 microg/ml. No statistically significant increase was observed between treated series when compared with a concurrent negative (water solvent) control series in either assay. The results demonstrate clearly that the phytotherapeutic extract from Equisetum myriochaetum, under the experimental conditions tested, is not genotoxic in the in vivo experiments or in the in vitro studies.     

Hypoglycemic effect of Cecropia obtusifolia Bertol aqueous extracts on type 2 diabetic patients

Cecropia obtusifolia is widely used in Mexican traditional medicine due to its reputed hypoglycemic effect. During a period of 32 weeks, aqueous extracts of the leaves of this plant were administrated daily to 12 recently diagnosed type 2 diabetic patients, controlled only with diet and exercise. Serum glucose, cholesterol, triglyceride and insulin levels were determined every 15 days; HbA1c, ALT, AST and ALKP were measured every month. A significant reduction of glucose was detected after 4 weeks of administration but the reduction was significant and sustained after 18 weeks of administration. The HbA1c was also significantly reduced after 6 weeks of treatment. No significant changes on cholesterol, triglycerides ALT, AST, ALKP or insulin could be detected. No collateral effects were observed. After suspending the administration of the extract, and a follow-up of 34 weeks, the glucose and HbA1c levels increased reaching levels higher than the basal ones. It can be concluded that the aqueous extracts of Cecropia obtusifolia have a significant hypoglycemic effect with no adverse effects and that the mechanism of action is not brought about by stimulating the insulin secretion. The results support the fact that the extracts of Cecropia obtusifolia have a great potential to be further developed into a phytomedicine.     

A pharmacological study of Cecropia obtusifolia Bertol aqueous extract.

Cecropia obtusifolia (Cecropiaceae) is a species from tropical America and its leaves are used in traditional medicine for the treatment of diabetes and as an anti-inflammatory agent. In the present study, the analgesic, anti-inflammatory and central nervous system depressant effects of the aqueous extract from the leaves of C. obtusifolia were investigated in different experimental models, with the purpose of validating its ethnomedical uses. The results obtained with the extract from the leaves of C. obtusifolia reflect a low toxicity, a substantial central depressor effect and analgesic activity and significant motor incoordination and muscle relaxant activity. Concerning the analgesic activity, using the hot plate test, the extract did not produce any effect, however it showed a significant effect on the pain induced by chemical stimuli (acetic and formalin test); this suggests the peripheral analgesic effect of the extract. The extract also showed a topical and systemic anti-inflammatory effect. Thus this work could justify the popular use of C. obtusifolia in rheumatic and kidney inflammation pathologies and reveals that this plant is an interesting species.     

in vitro genotoxic and mutagenic evaluation of the aqueous extract of Codiaeum variegatum and its amoebicidal sub-fraction

Ethnopharmacological relevance

Codiaeum variegatum, grown in many varieties, has been widely used as a houseplant based on its brightly decorative foliage. In addition, a variety of this plant has been used for a long time against bloody diarrhea by the local population in Cameroon.

Aim of the study

In our previous study, the aqueous extract of this plant and an isolated sub-fraction exhibited significant anti-amoebic activity on axenic culture of Entamoeba histolytica. Due to the medicinal value of these extracts, we promptly initiated to investigate their genotoxic and mutagenic potential in order to assure their safe and rationale usage in traditional healthcare system.

Material and methods

Both extracts were incubated with L5178Y mouse lymphoma cells, primary hepatic cells and HepG2 human hepatocellular carcinoma cells and their genotoxicity and mutagenicity were evaluated by quantifying DNA damage and chromosomal aberrations through comet assay, micronucleus assay and mouse lymphoma mutation assay.

Results

The aqueous extract of Codiaeum variegatum is not cytotoxic up to 2000 µg/mL while the amoebicidal fraction is significantly cytotoxic (≤ 40–55%) on L5178Y mouse lymphoma and HepG2 cells at concentrations higher than 500 µg/mL. Besides, no significant DNA damage and induction of micronucleus formation were identified at concentrations up to 2000 µg/mL. Moreover, the mutagenic potential of these extracts after short (4 h) and long term (24 h) treatment, revealed no significant gene mutation induction.

Conclusion

The aqueous extract of Codiaeum variegatum and the amoebicidal fraction SF9B are neither genotoxic on non-competent or metabolic competent cell lines, nor mutagenic in mouse lymphoma mutation assay and therefore they could be safely used at lower doses for medicinal purpose.     

Evaluation of the genotoxicity of Orthosiphon stamineus aqueous extract

Aim of the study

Orthosiphon stamineus, Benth, also known as Misai Kucing in Malaysia and Java tea in Indonesia, is traditionally used in Southeastern Asia to treat kidney dysfunctions, diabetes, gout and several other illnesses. Recent studies of Orthosiphon stamineus pharmacological profile have revealed antioxidant properties and other potentially useful biological activities thereby lending some scientific support to its use in folk medicine. So far the genotoxicity of Orthosiphon stamineus extracts has not been evaluated. In this study the genotoxic potential of Orthosiphon stamineus aqueous extract was investigated by the Salmonella/microsome mutation assay and the mouse bone marrow micronucleus test.

Materials and methods

Chemical composition of Orthosiphon stamineus aqueous extract was analyzed by High Performance Liquid Chromatography-Diode Array Detector (HPLC-DAD). The Salmonella/microsome assay (TA97a, TA98, TA100 and TA1535; plate incorporation method) was performed in the presence or in the absence of extrinsic metabolic activation (S9 mixture). In the mouse micronucleus assay, Orthosiphon stamineus extract was administered by gavage (0, 500, 2000 and 4000 mg/kg body weight/day for 3 days) to male and female Swiss Webster mice (N = 6 per dose per sex) and bone marrow cells were harvested 24 h after the last dose. Ethoxy-resorufin-O-dealkylase (EROD) and benzyloxy-resorufin-O-dealkylase (BROD) activities were determined in mouse liver microsomes.

Results

The chemical analysis revealed that the Orthosiphon stamineus extract contained flavonoids (sinensetin, eupatorin), caffeic acid, and rosmarinic acid (44.00 ± 1.879 μg/mg), the latter seemed to be one of its major constituent. Tested at doses up to 5000 μg/plate, the Orthosiphon stamineus extract was not toxic to Salmonella tester strains and did not increase the number of revertant colonies over the background incidence. In the mouse bone marrow assay, the extract did not alter the polychromatic:normochromatic erythrocytes (PCE:NCE) ratio, nor did it increase the incidence of micronucleated polychromatic erythrocytes (MNPEs). No overt toxicity and no change of CYP1A (EROD) and 2B9/10 (BROD) activities were noted.

Conclusions

Based on the aforementioned findings, it is concluded that the use of Orthosiphon stamineus in the traditional medicine poses no genotoxic risk.     

Mutagenic and genotoxic effects of Baccharis dracunculifolia (D.C.)

Baccharis dracunculifolia (D.C.) (Asteraceae), a native plant to Brazil known as “vassourinha” or “alecrim-do-campo”, is the most important botanical source of a Brazilian propolis called green propolis. The leaf extracts of this plant have been used to treat liver and digestive disorders. It has been recognized that green propolis can induce mutagenic effects at high doses, but no study reporting possible mutagenic effects by Baccharis dracunculifolia extracts in the maximum tolerated dose has been conducted. The aim of the present study was to investigate the genotoxic and mutagenic effects of this plant in vivo. Adult CF-1 mice were treated with 0.5 g/kg, 1.0 g/kg or 2.0 g/kg of an aqueous extract of Baccharis dracunculifolia by gavage for 3 consecutive days. Blood and liver samples were collected to detect DNA damage using the comet assay, while bone marrow samples were used to assess chromosome mutations by the micronucleus test. The extract increased the DNA damage in blood and liver tissues and the frequency of micronucleus in bone marrow. These findings suggest genotoxic and mutagenic effects of Baccharis dracunculifolia comparable to green propolis in mice.     

Genotoxic and antigenotoxic properties of Baccharis trimera in mice

Ethnopharmacological relevance

Baccharis trimera (Less.) (Asteraceae) is a native plant from Brazil. Also known as “carqueja”, it is popularly used to treat liver diseases, diabetes, as well as digestive disorders, mainly by women with lower socioeconomic status.

Aim of the study

The aim of the present study was to investigate the in vivo genotoxic/antigenotoxic and mutagenic potential of this plant, using the comet and the micronucleus assays.

Material and methods

Female adult mice were treated with 500, 1000 or 2000 mg/kg of B. trimera aqueous extract (Bt-AE) by gavage for three consecutive days.

Results

Independently of the dose, no genotoxic effect was observed in blood and liver samples analyzed by the comet assay. Conversely, B. trimera showed an antigenotoxic effect in blood from treated mice, thus protecting cells against oxidative DNA damage induced by the ex vivo treatment with hydrogen peroxide. In addition, Bt-AE showed in vitro antioxidant activity, assessed by DPPH and xanthine oxidase assays, suggesting that the observed antigenotoxic effects might be related to its antioxidant properties.

Conclusions

However, the extract increased the frequency of micronucleus in bone marrow of treated mice, indicating a chromosomal mutagenic activity. Thus, medicines prepared from this plant should be used with caution, although the results also suggest antigenotoxic effects for B. trimera aqueous extract.     

Hypoglycemic effect of Cecropia obtusifolia on streptozotocin diabetic rats.

The hypoglycemic effects of water and butanolic extracts prepared from leaves of Cecropia obtusifolia (Cecropiaceae) were examined in streptozotocin induced diabetic rats. A single oral administration of a water extract at doses of 90 and 150 mg/kg and of a butanol extract at doses of 9 and 15 mg/kg significantly (P<0.05) lowered the plasma glucose levels in diabetic rats after 3 h administration. Glibenclamide was used as reference and showed similar hypoglycemic effect to the tested extracts at a dose of 3 mg/kg. The flavone, isoorientin and 3-caffeoylquinic acid (chlorogenic acid), were isolated as the important constituents of the plant and were identified as the main constituents in both extracts, too.     

Genotoxicity assessment of Garcinia achachairu Rusby (Clusiaceae) extract in mammalian cells in vivo

Ethnopharmacological relevance

Garcinia achachairu Rusby (Clusiaceae) is popularly known as “achachairu”, and is used in Bolivian folk medicine for its healing, digestive, and laxative properties, and in the treatment of gastritis, rheumatism and inflammation. Despite its widespread therapeutic use, there is a lack of data regarding its in vivo genotoxic effects. Therefore, in this study, we used the comet assay and the micronucleus test, respectively, to evaluate the possible genotoxic and clastogenic effects of Garcinia achachairu seed extract (GAE) on different cells of mice.

Material and methods

The GAE was administered by oral gavage at doses of 500, 1000 and 2000 mg/kg. For the analysis, the comet assay was performed on the leukocytes (collected 4 and 24 h after treatment), liver, bone marrow and testicular cells (collected 24 h after treatment), and the micronucleus test (MN) on bone marrow cells. Cytotoxicity was assessed by scoring 200 consecutive polychromatic (PCE) and normochromatic (NCE) erythrocytes (PCE/NCE ratio).

Results and conclusion

The results showed that GAE did not induce significant DNA damage in leukocytes (4 h and 24 h samples), liver, bone marrow and testicular cells (24 h samples). GAE also did not show any significant increase in micronucleated polychromatic erythrocytes (MNPCEs) at the three tested doses. The PCE/NCE ratio indicated no cytotoxicity. Under our experimental conditions, the data obtained suggest that a single oral administration of G. achachairu extract does not cause genotoxicity and clastogenicity in different cells of mice.     

Cyto-/genotoxic effects of the ethanol extract of Chan Su,a traditional Chinese medicine,in human cancer cell lines

Ethnopharmacological relevance

Chan Su, an ethanolic extract from skin and parotid venom glands of the Bufo bufo gargarizans Cantor, is widely used as a traditional Chinese medicine for cancer therapy. Although the anti-cancer properties of Chan Su have been investigated, no information exists regarding whether Chan Su has genotoxic effects in cancer cells. The aim of the present study was to examine the cyto-/genotoxic effect of Chan Su in human breast carcinoma (MCF-7 cells), human lung carcinoma (A-549 cells), human T cell leukemia (Jurkat T cells), and normal human lymphocytes.

Materials and methods

Effects on the viability of MCF-7, A-549, Jurkat T cells, and normal lymphocytes were evaluated by Trypan blue exclusion assays. The DNA content in the sub-G1 region was detected by propidium iodide (PI) staining and flow cytometry. The genotoxicity of Chan Su was assessed by single-cell gel electrophoresis (comet assay) and the cytokinesis-block micronucleus assay (CBMN assay).

Results

Chan Su significantly inhibited the viability of MCF-7, A-549, and Jurkat T cells dose dependently, but had no effect on normal human lymphocytes. Apoptotic death of the cancer cells was evident after treatment. Chan Su also induced genotoxicity in a dose-dependent manner, as indicated by the comet and cytokinesis-block micronucleus assays.

Conclusions

These findings suggest that Chan Su can induce apoptotic death of, and exert genotoxic effects on, MCF-7, A-549, and Jurkat T cells.     

Genotoxic and clastogenic activity of saponins extracted from Nauclea bark as assessed by the micronucleus and the comet assays in Chinese Hamster Ovary cells

Ethnopharmacological relevance

Bark extracts of Nauclea latifolia, Nauclea diderrichii, Nauclea pobeguinii and Nauclea vandergutchii are used in traditional medicine in West and South Africa for the treatment of fevers, diarrhea and malaria.

Aim of the study

To estimate the possible long-term toxicity and genotoxicity of plant extracts (dichloromethane, methanol, water/methanol, water) and saponins.

Materials and methods

The clastogenicity of plant extracts and saponins was assessed by the micronucleus assay performed on Chinese Hamster Ovary cells. The DNA-damaging activity of saponin mixture was assessed by the comet assay on Chinese Hamster ovary cells.

Results

Hydromethanolic extracts from Nauclea latifolia, Nauclea diderrichii and Nauclea pobeguinii exhibited a significant clastogenic/aneugenic activity without S9 mix. The hydromethanolic extract from Nauclea diderrichii was the most clastogenic/aneugenic fraction with a Minimal Active Concentration (MAC) of 23.1 μg mL⁻¹. It was submitted to a separation step leading to six main saponins identified as quinovic acid glycosides (saponins A, D, E, G, J, K). None of the isolated saponins exerted a significant clastogenic/aneugenic activity by the micronucleus assay, however a mixture made with equal quantities of each of the six saponins exhibited a direct genotoxic/clastogenic activity as assessed by both the micronucleus assay and the comet assay on Chinese Hamster Ovary cells.

Conclusion

Saponins present in the hydromethanolic extracts of Nauclea induced synergistic in vitro DNA-damage and chromosome mutations in mammalian cells. This genotoxic activity was probably due to the capacity of Nauclea saponins to reduce cell defense against oxidative stress through the inhibition of glutathione-S-transferase activity.     

Evaluation of the genotoxic and antigenotoxic potential of Brassica oleracea L. var. acephala D.C. in different cells of mice

Ethnopharmacological relevance

Brassica oleracea L. var. acephala D.C. has been extensively used in Brazilian traditional medicine to treat gastric ulcer.

Aim of the study

This study was conducted to evaluate the in vivo genotoxic and/or antigenotoxic potential of a Brassica oleraceae hydroalcoholic extract obtained from the leaves, in different cells of mice.

Materials and methods

Analyses were performed using the comet assay, on leukocytes (collected 4 and 24 h after treatment), liver, brain, bone marrow and testicular cells (collected 24 h after treatment), and using the micronucleus test (MN) in bone marrow cells. Eight groups of albino Swiss mice were treated (N=6): control (C), positive control (doxorubicin 80 mg/kg (DXR)), and six experimental groups, which received 500, 1000 and 2000 mg/kg of Brassica oleraceae extract alone by gavage, while a further three groups received the same doses plus DXR (80 mg/kg). We calculated the damage scores, and their averages were compared by ANOVA followed by the Tukey test for multiple comparisons.

Results

The results demonstrated that none of the tested doses of Brassica oleraceae extract showed genotoxic effects by the comet assay, or clastogenic effects by the MN test. On the other hand, for all cells evaluated, the three tested doses of the Brassica extract promoted inhibition of DNA damage induced by DXR.

Conclusions

Under our experimental conditions, Brassica oleraceae leaf extract showed no genotoxic or clastogenic effects in different cells of mice. However, it did show a significant decrease in DNA damage induced by doxorubicin. It is suggested that the antigenotoxic properties of this extract may be of great pharmacological importance, and may be beneficial for cancer prevention.     

Determination of genotoxic, antigenotoxic and cytotoxic potential of the extract from lichen Cetraria aculeata (Schreb.) Fr. in vitro

Naturally occurring compounds can have protective effects towards mutagens and carcinogens as shown by numerous studies. In the present study, the genotoxic/antigenotoxic activities of the extract from lichen Cetraria aculeata (Schreb.) Fr., which has been identified as an antibacterial agent in former studies, were investigated against known mutagens such as 4-nitro-o-phenylenediamine (4-NPD) and 2-aminofluorene (2-AF) in TA98 and TA100 strains of Salmonella typhimurium in the presence or absence of metabolic activity. Further genotoxicity/antigenotoxicity of the extract against mitomycin C for micronucleus formation was also evaluated in human lymphocytes. The cytotoxic effects of the extract on mammalian cells were investigated in three different cell line types by MTT assay. The results obtained show that the extract of C. aculeata has a significant antigenotoxic activity in bacterial systems, but not in mammalian cells and cytotoxic activity in some mammalian cancer cells. However, it was not mutagenic in all systems. The findings suggest that the lichen extract may have a possible therapeutic potential and therefore this must be further investigated by other multiple in vitro bioassays for the development of therapeutic agents.     

An in vitro Study on the DNA Damaging Effects of Phytochemicals Partially Isolated from an Extract of Glinus lotoides

An extract of Glinus lotoides, a medicinal plant used in Africa and Asia for various therapeutic purposes, was recently shown to cause DNA damage in vitro. To further explore the potential genotoxicity of this plant, fractionation of the crude extract was performed using reverse phase solid‐phase extraction and a stepwise gradient elution of methanol in water. Four fractions were collected and subsequently analysed for their DNA damaging effects in mouse lymphoma cells using an alkaline version of the comet assay. To identify potential genotoxic and non‐genotoxic principles, each fraction was then subjected to liquid chromatography coupled to mass spectrometry, LC‐MS/MS. 1D and 2D nuclear magnetic resonance analyses were used to confirm the identity of some saponins. Although fractions containing a mixture of flavonoids and oleanane‐type saponins or oleanane‐type saponins alone produced no DNA damage, those containing hopane‐type saponins exhibited a pronounced DNA damaging effect without affecting the viability of the cells. To conclude, even if this study presents evidence that hopane‐type of saponins are endowed with a DNA damaging ability, further studies are needed before individual saponins can be cited as a culprit for the previously reported genotoxicity of the crude extract of G. lotoides. Copyright © 2012 John Wiley & Sons, Ltd.     

Hypoglycemic effects of Cecropia pachystachya in normal and alloxan-induced diabetic rats

Ethnopharmacological relevance

Leaves of Cecropia pachystachya are described in the folk medicine as possessing antitusive, expectorant, antiasthmatic and hypoglycemic effects.

Aim of the study

To investigate the hypoglycemic and antioxidant effects of methanolic extract from the leaves of Cecropia pachystachya. The total amount of phenolic and flavonoids compounds was determined and the qualitative composition of the extract was analyzed.

Materials and methods

The hypoglycemic effect of the extract was tested in normal, glucose loading and alloxan-induced diabetic rats. The antioxidant activity was assessed by DPPH free radical scavenging and reduction power assays. The total amount of phenolic and flavonoids compounds was determined by Folin-Denis and AlCl₃ reagent method, respectively. The qualitative composition of the extract was analyzed using a HPLC-DAD system.

Results

The glucose tolerance test showed that in diabetic rats, the extract caused a significant hypoglycemic effect with a blood glucose reduction of 68% after 12 h. The administration of the extract in alloxan-induced diabetic rats also produced a significant reduction in the blood glucose levels at all points being more pronounced at 90 min (reduction of 60%). After 120 min, no significant difference was observed between the blood levels of the rats treated with the extract and those treated with the standard drugs (metformin and glibenclamide). The extract also presented relevant antioxidant activity with IC₅₀ = 3.1 μg/ml (DPPH assay) and EC₅₀ = 10.8 μg/ml (reduction power). Results were compared with the reference antioxidants quercetin, rutin, and ascorbic acid. The content of flavonoids was 83 mg/g plant and that of phenolics was 326 mg/g plant. Chlorogenic acid and the C-glycosylated flavones, orientin and isoorientin, were identified in the extract.

Conclusions

In conclusion, the findings showed that the folk medicinal plant Cecropia pachystachya possesses hypoglycemic and antioxidant effects which confirmed the traditional use of the plant in the treatment of diabetes. Chlorogenic acid and the C-glycosylated flavonoids may explain these activities.     

Dragon's blood Croton palanostigma induces genotoxic effects in mice

Ethnopharmacological relevance

Dragon's blood is a dark-red sap produced by species from the genus Croton (Euphorbiaceae), which has been used as a famous traditional medicine since ancient times in many countries, with scarce data about its safe use in humans. In this research, we studied genotoxicity and clastogenicity of Croton palanostigma sap using the comet assay and micronucleus test in cells of mice submitted to acute treatment.

Material and methods

HPLC analysis was performed to identify the main components of the sap. The sap was administered by oral gavage at doses of 300 mg/kg, 1000 mg/kg and 2000 mg/kg. For the analysis, the comet assay was performed on the leukocytes and liver cells collected 24 h after treatment, and the micronucleus test (MN) on bone marrow cells. Cytotoxicity was assessed by scoring 200 consecutive polychromatic (PCE) and normochromatic (NCE) erythrocytes (PCE/NCE ratio).

Results and conclusion

The alkaloid taspine was the main compound indentified in the crude sap of Croton palanostigma. The results of the genotoxicity assessment show that all sap doses tested produced genotoxic effects in leukocytes and liver cells and also produced clastogenic/aneugenic effects in bone marrow cells of mice at the two higher doses tested. The PCE/NCE ratio indicated no cytotoxicity. The data obtained suggest caution in the use of Croton palanostigma sap by humans considering its risk of carcinogenesis.     

吴茱萸及其主要成分的遗传毒性研究

目的:研究吴茱萸及其主要成分吴茱萸碱、吴茱萸次碱和柠檬苦素的遗传毒性,为有毒中药吴茱萸的开发利用提供依据。方法:选用了Ames试验、体外CHL细胞染色体畸变试验和小鼠骨髓嗜多染红细胞微核试验。Ames试验选用TA97、TA98、TA100、TA102、TA1535组氨酸缺陷型菌株。吴茱萸碱、吴茱萸次碱和柠檬苦素分别设5个剂量组:0.000 5、0.005、0.05、0.5、5mg/皿;体外CHL细胞染色体畸变试验,吴茱萸碱、吴茱萸次碱和柠檬苦素的剂量分别为0.005 mg/mL、0.05/mL和0.5 mg/mL,受试药物与CHL细胞接触时间分别为4 h和24 h;吴茱萸醇提物小鼠骨髓微核试验,设吴茱萸醇提物0.88、3.52、10.55 g(生药)/kg 3个给药剂量组。连续给药4 d,每天给药1次。结果:吴茱萸碱、吴茱萸次碱和柠檬苦素各剂量组Ames试验结果为阴性。CHL试验中,吴茱萸碱的试验结果为阴性;吴茱萸次碱24 h组细胞染色体畸变率略有增加,差异有统计学意义(P0.05);柠檬苦素4 h和24 h组细胞染色体畸变率都略有增加,0.05 mg/mL、0.5 mg/mL组与阴性对照组比较,差异有统计学意义(P0.05),而0.005 mg/mL组与阴性对照比较,差异无统计学意义(P0.05)。微核试验中,吴茱萸醇提物的小鼠骨髓细胞微核试验结果为阴性。结论:综合以上3个试验结果,认为在本实验室条件下,吴茱萸醇提物无遗传毒性。但在体外试验中,吴茱萸次碱和柠檬苦素有致突变性。为进一步确认吴茱萸及其主要成分的遗传毒性,可进一步研究和探讨。     

Assessment of the genotoxic risk of Punica granatum L. (Punicaceae) whole fruit extracts

Punica granatum L. (Punicaceae) whole fruit extracts, have been used in Cuban traditional medicine as an effective drug for the treatment of respiratory diseases. This species showed interesting anti-viral activity, e.g. aqueous or hydroalcoholic extracts of whole fruits have proved highly active against the influenza virus. However, some toxic properties of this extract have also been reported and, to date, very little is known about its genotoxic properties. In the present study, the genotoxicity of a Punica granatum (pomegranate) whole fruit extract was assessed using different in vitro and in vivo assays that detect DNA damage at different expression levels. Results from reversion and gene-conversion test in microorganisms, sister chromatid exchanges, micronuclei and sperm-shape abnormality assays in mice, clearly showed that the hydroalcoholic extract of P. granatum whole fruits is genotoxic when tested both in vitro and in vivo.     

Evaluation of genotoxic and antigenotoxic effects of hydroalcoholic extracts of Zuccagnia punctata Cav

Zuccagnia punctata Cav. (Fabaceae), a widely used plant species in Argentine folk medicine, has been shown to have a broad spectrum of antibacterial, antifungal, antioxidant and cytoprotective activities. In this study, the hydroalcoholic extract of Zuccagnia punctata and 2',4'-dihydroxychalcone isolated from it were investigated for genotoxicity/antigenotoxicity in the in vitro comet assay test on human hepatoma HepG2 cells. No acute toxicity of the extract could be determined. HepG2 cells were treated with three different concentrations (2.5, 5.0 and 10.0 microg/mL) or 2',4'-dihydroxychalcone (0.01, 0.10 and 1.00 microg/mL). To explore the potential mechanisms of action, two approaches were followed: co-treatment with 4-nitroquinoline-N-oxyde (4-NQO), a direct genotoxic compound, and a pre-treatment protocol with benzo[a]pyrene (B[a]P), an indirect genotoxic compound. The natural products neither affected cell viability nor induced DNA damage in the concentration range tested. Zuccagnia punctata tinctures were able to diminish the DNA damage induced in HepG2 cells by 4-NQO and B[a]P in 31% and 10%, respectively at 10 microg/mL. Pre-treatment of HepG2 cells with 2',4'-dihydroxychalcone was highly effective in decreasing B[a]P-induced DNA damage at a statistically significant level, with an almost clear dose-response relationship. The inhibition values were 28.2-43.9% for the tested concentrations of 0.01-1 microg/mL, respectively. The results clearly indicate that the phytoextract from Zuccagnia punctata, under the experimental conditions tested, is not genotoxic and that 2',4'-dihydroxychalcone contributes to a high degree to the antigenotoxic effects of Zuccagnia punctata tincture.