Antimutagenicity of rosmarinic acid in Swiss mice evaluated by the micronucleus assay

Rosmarinic acid (RA) is a natural phenolic compound which presents different biological activities such as antitumor, antibacterial, anti-inflammatory, hepatoprotective and cardioprotective properties. In view of its important biological activities, the study of the effects of RA on genetic material becomes relevant. Thus, the objective of the present study was to evaluate the mutagenic and/or antimutagenic potential of RA on peripheral blood cells of Swiss mice using the micronucleus assay. Three doses of RA (50, 100 and 200 mg/kg body weight, b.w.) were used for the evaluation of its mutagenic potential. In the antimutagenicity assays, the different concentrations of RA were combined with the chemotherapeutic agent doxorubicin (DXR, 15 mg/kg b.w.). Peripheral blood samples were collected 24, 48 and 72 h after treatment for the evaluation of micronucleated polychromatic erythrocytes (MNPCEs). The results of the mutagenicity assays showed no increase in the frequency of micronuclei in animals treated with different concentrations of RA when compared to the negative controls. Treatment with different concentrations of RA combined with DXR revealed a significant reduction in the frequency of micronuclei compared to animals treated with DXR only. Although the mechanisms underlying the protective effect of RA are not completely understood, the putative antioxidant activity of RA might explain its effect on DXR mutagenicity.     

Antimutagenicity of rosmarinic acid in Swiss mice evaluated by the micronucleus assay

Rosmarinic acid (RA) is a natural phenolic compound which presents different biological activities such as antitumor, antibacterial, anti-inflammatory, hepatoprotective and cardioprotective properties. In view of its important biological activities, the study of the effects of RA on genetic material becomes relevant. Thus, the objective of the present study was to evaluate the mutagenic and/or antimutagenic potential of RA on peripheral blood cells of Swiss mice using the micronucleus assay. Three doses of RA (50, 100 and 200 mg/kg body weight, b.w.) were used for the evaluation of its mutagenic potential. In the antimutagenicity assays, the different concentrations of RA were combined with the chemotherapeutic agent doxorubicin (DXR, 15 mg/kg b.w.). Peripheral blood samples were collected 24, 48 and 72 h after treatment for the evaluation of micronucleated polychromatic erythrocytes (MNPCEs). The results of the mutagenicity assays showed no increase in the frequency of micronuclei in animals treated with different concentrations of RA when compared to the negative controls. Treatment with different concentrations of RA combined with DXR revealed a significant reduction in the frequency of micronuclei compared to animals treated with DXR only. Although the mechanisms underlying the protective effect of RA are not completely understood, the putative antioxidant activity of RA might explain its effect on DXR mutagenicity.     

Inhibition of doxorubicin-induced mutagenicity by Baccharis dracunculifolia

Baccharis dracunculifolia DC (Asteraceae), a native plant from Brazil, have been used as an antipyretic, stomachic and health tonic in Brazil. The objective of the present study was to investigate the potential mutagenic effect of B. dracunculifolia ethyl acetate extract (Bd-EAE) and its influence on the mutagenicity induced by the chemotherapeutic agent doxorubicin (DXR) using the rat bone marrow and peripheral blood micronucleus test. Wistar rats were divided into 10 treatment groups. Five groups received DXR (90 mg/kg body weight, b.w., intraperitoneally) to induce mutagenicity and three of these groups received a single oral dose of Bd-EAE at a concentration of 6, 12 or 24 mg/kg b.w. prior to DXR administration. A vehicle-treated control group and Bd-EAE control groups were also included. The results showed that Bd-EAE itself was not mutagenic, in the rat micronucleus assay. In animals treated with Bd-EAE and DXR, the number of MNPCEs was significantly decreased compared to animals receiving DXR alone. HPLC analysis of the extract obtained permitted the identification of the following phenolic compounds: caffeic acid, p-coumaric acid, aromadendrin-4'O-methyl ether, 3-prenyl-p-coumaric acid (drupanin), 3,5-diprenyl-p-coumaric acid (artepillin C) and baccharin. The putative antioxidant activity or the interference of one or more of the active compounds of Bd-EAE with mutagenic metabolic pathways may explain its effect on DXR mutagenicity.     

Inhibition of doxorubicin-induced mutagenicity by Baccharis dracunculifolia

Baccharis dracunculifolia DC (Asteraceae), a native plant from Brazil, have been used as an antipyretic, stomachic and health tonic in Brazil. The objective of the present study was to investigate the potential mutagenic effect of B. dracunculifolia ethyl acetate extract (Bd-EAE) and its influence on the mutagenicity induced by the chemotherapeutic agent doxorubicin (DXR) using the rat bone marrow and peripheral blood micronucleus test. Wistar rats were divided into 10 treatment groups. Five groups received DXR (90 mg/kg body weight, b.w., intraperitoneally) to induce mutagenicity and three of these groups received a single oral dose of Bd-EAE at a concentration of 6, 12 or 24 mg/kg b.w. prior to DXR administration. A vehicle-treated control group and Bd-EAE control groups were also included. The results showed that Bd-EAE itself was not mutagenic, in the rat micronucleus assay. In animals treated with Bd-EAE and DXR, the number of MNPCEs was significantly decreased compared to animals receiving DXR alone. HPLC analysis of the extract obtained permitted the identification of the following phenolic compounds: caffeic acid, p-coumaric acid, aromadendrin-4′O-methyl ether, 3-prenyl-p-coumaric acid (drupanin), 3,5-diprenyl-p-coumaric acid (artepillin C) and baccharin. The putative antioxidant activity or the interference of one or more of the active compounds of Bd-EAE with mutagenic metabolic pathways may explain its effect on DXR mutagenicity.     

Induction of chromosome aberrations, micronucleus formation and sperm abnormalities in mouse following carbofuran exposure

Carbofuran was tested to study in vivo cytogenetic effects in mouse bone marrow cells and morphological alterations in sperms. The acute oral and intraperitoneal (i.p.) LD(50) of carbofuran was determined to be 9.5 or 2.0 mg/kg b.w. in mice, respectively. The animals were orally administered 1.9, 3.8 or 5.7 mg/kg b.w. (20, 40 and 60% of LD(50)) of carbofuran for 24 h or 1.9 mg/kg b.w. for 4 consecutive days (cumulative 7.6 mg/kg or 80% of LD(50)) to analyse chromosome aberrations (CAs). For micronucleus test (MT) animals were orally exposed to 5.7 mg/kg b.w. for 24 and 48 h or 1.9 mg/kg b.w. for 4 consecutive days. For reference mice were exposed to peanut oil (negative control) and cyclophosphamide (20 mg/kg) or ethyl methanesulfonate (EMS: 100 mg/kg) positive control for CAs and MT respectively. To analyse the effect on sperm morphology mice were exposed to single i.p. dose of 1 and 2 mg/kg b.w. of carbofuran and repeatedly to 0.5 mg/kg for 5 consecutive days. Cytogenetic analysis revealed that all the test doses induced mitotic inhibition, CAs, micronucleus (MN) formation and sperm abnormalities in a dose dependent manner. Present observations concurrent with earlier reports substantiate the genotoxic potential of carbofuran and possible risk to human beings.     

In vivo analgesic and anti-inflammatory activities of ursolic acid and oleanoic acid from Miconia albicans (Melastomataceae)

The aim of this work was to use in vivo models to evaluate the analgesic and anti-inflammatory activities of ursolic acid (UA) and oleanoic acid (OA), the major compounds isolated as an isomeric mixture from the crude methylene chloride extract of Miconia albicans aerial parts in an attempt to clarify if these compounds are responsible for the analgesic properties displayed by this plant. Ursolic acid inhibited abdominal constriction in a dose-dependent manner, and the result obtained at a content of 40 mg kg(-1) was similar to that produced by administration of acetylsalicylic acid at a content of 100 mg kg(-1). Both acids reduced the number of paw licks in the second phase of the formalin test, and both of them displayed a significant anti-inflammatory effect at a content of 40 mg kg(-1). It is noteworthy that the administration of the isolated mixture, containing 65% ursolic acid/35% oleanolic acid, did not display significant analgesic and anti-inflammatory activities. On the basis of the obtained results, considering that the mixture of UA and OA was poorly active, it is suggested that other compounds, rather than UA and OA, should be responsible for the evaluated activities in the crude extract, since the crude extract samples displayed good activities.     

Protective effect of CardiPro against doxorubicin-induced cardiotoxicity in mice

The effect of CardiPro, a polyherbal formulation, with an antioxidant property, has been studied on doxorubicin (DXR)-induced cardiotoxicity in mice. CardiPro (150 mg/kg b.w., twice daily was administered orally for 7 weeks along with four equal injections (each containing 4.0 mg/kg b.w., DXR) intraperitoneally, once weekly (cumulative dose 16 mg/kg). After a 3-week post DXR treatment period, cardiotoxicity was assessed by noting mortality, volume of ascites, liver congestion, changes in heart weight, myocardial lipid peroxidation, antioxidant enzymes and histology of heart. DXR-treated animals showed higher mortality (50%) and more ascites. Myocardial SOD and glutathione peroxidase activity were decreased and lipid peroxidation was increased. Histology of heart of DXR-treated animals showed loss of myofibrils and focal cytoplasmic vacuolization. CardiPro significantly protected the mice from DXR-induced cardiotoxic effects as evidenced by lower mortality (25%), less ascites, myocardial lipid peroxidation, normalization of antioxidant enzymes and minimal damage to the heart histologically. Our data confirm the earlier reports that DXR cardiotoxicity is associated with the free radical-induced tissue damage. Administration of CardiPro, with an antioxidant property, protected the DXR-induced cardiotoxicity in mice.     

Antimutagenic effect of Agaricus blazei Murrill mushroom on the genotoxicity induced by cyclophosphamide

Agaricus blazei Murrill extracts have previously been shown to have anticarcinogenic and antimutagenic properties. These results suggest that antimutagenic activity, besides the modulation of the immune system, might be involved in the anticarcinogenic action of A. blazei. To investigate the possible antimutagenic effect of A. blazei in vivo, we evaluated its effect on clastogenicity induced by cyclophosphamide (CP) in mice, using the micronucleus test in bone marrow (MNPCE) and in peripheral blood (MNRET). Male Swiss mice were treated with CP (25 or 50mg/kg i.p.) or with CP plus mushroom solution at three different temperatures: 4, 21, and 60 degrees C. Aqueous solution of a mixture from various lineages of the mushroom inhibited induction of micronuclei by CP in bone marrow and in peripheral blood of mice. In contrast to the mixture of lineages, a single isolated lineage did not lead to a reduction of CP-induced MN frequencies in either bone marrow or blood cells of mice. The results suggest that under certain circumstances these mushrooms exhibit antimutagenic activities that might contribute to an anticarcinogenic effect.     

In Vivo Assessment of Genotoxic,Antigenotoxic and Anticarcinogenic Activities of Solanum lycocarpum Fruits Glycoalkaloidic Extract

The fruits of Solanum lycocarpum, known as wolf-fruit, are used in folk medicine, and because of that we have evaluated both the genotoxic potential of its glycoalkaloidic extract (SL) and its influence on the genotoxicity induced by methyl methanesulfonate. Furthermore, the potential blocking effect of SL intake in the initial stage of colon carcinogenesis in Wistar rats was investigated in a short-term (4-week) bioassay using aberrant crypt foci (ACF) as biomarker. The genotoxic potential was evaluated using the Swiss mice peripheral blood micronucleus test. The animals were treated with different doses of SL (15, 30 and 60 mg/kg b.w.) for 14 days, and the peripheral blood samples were collected at 48 h, 7 days and 14 days after starting the treatment. For antigenotoxicity assessment, MMS was administered on the 14^th day, and after 24 h the harvesting of bone marrow and liver cells was performed, for the micronucleus and comet assays, respectively. In the ACF assay, male Wistar rats were given four subcutaneous injections of the carcinogen 1,2-dimethylhydrazine (DMH, 40 mg/kg b.w.), twice a week, during two weeks to induce ACF. The treatment with SL (15, 30 and 60 mg/kg b.w.) was given for four weeks during and after carcinogen treatment to investigate the potential beneficial effects of SL on DMH-induced ACF. The results demonstrated that SL was not genotoxic in the mouse micronucleus test. In animals treated with SL and MMS, the frequencies of micronucleus and extensions of DNA damage were significantly reduced in comparison with the animals receiving only MMS. Regarding the ACF assay, SL significantly reduced the frequency of ACF induced by DMH.     

Relationship between experimental results in mammals and man. I. Cytogenetic analysis of bone marrow injury induced by a single dose of cyclophosphamide.

The extrapolation of experimental results to man was studied by cytogenetic bone marrow analysis and micronucleus test in mice, rats and Chinese hamsters. Furthermore, the frequency of chromosomal aberrations was compared with the frequencies of polychromatic erythrocytes containing micronuclei. Cyclophosphamide (CY) was given intraperitoneally at the doses of 5, 10, 20, 40 and 80 mg/kg b.w. to ICR mice and Wistar rats and at the doses of 10, 20, 40, 80, 120 and 160 mg/kg b.w. to Chinese hamsters. Five patients with various types of malignancies until then medically untreated, were i.v. administered 40 mg CY/kg b.w. Bone marrow cells were examined 24 h after the administration. CY induced in all rodents a clear-cut dose-effect relationship in the frequency of breaks, abnormal metaphases as well as in the frequency of micronuclei in polychromatic erythrocytes. When comparing the results in rodents and man at the dose of 40 mg CY/kg b.w., the sensitivity pattern of species was mice greater than rats greater than Chinese hamsters greater than man. From this aspect the possible differences in the metabolism of CY in analysed species are discussed. The presented results tend to a conclusion that micronucleus testing may be a very suitable method used for screening purpose, however, the method of classical cytogenetic analysis, especially the evaluation of breaks, still remains the most exact and reliable technique.     

Protective effects of ursolic acid and oleanolic acid in leukemic cells

Ursolic acid (UA) and oleanolic acid (OA) have similar chemical structures but differ in the position of one methyl group on the ring E. We investigated protective effects of these two triterpenoic acids against H₂O₂-induced DNA damage in leukemic L1210, K562 and HL-60 cells using single-cell gel electrophoresis (SCGE). We compared their protective effects (antioxidant activities) with respect to the different position of the methyl group in their chemical structures. After 24 h pre-treatment of cells both compounds investigated inhibited significantly the incidence of DNA single strand breaks induced by H₂O₂. The concentration range of UA and OA was in all experiments 2.5–10 μmol/l. The antioxidant activity of OA determined by SCGE was significantly higher compared to UA in L1210 (⁺P < 0.05) and K562 cells (⁺⁺⁺P < 0.001). Significant difference of the antioxidant activities of the two compounds was evidently connected with the different position of the methyl group. The protective effect of OA was in HL-60 cells slightly lower compared to the activity of UA, but the difference between the protective effects of UA and OA was not significant. In conclusion we can say that both natural pentacyclic triterpenoic acids investigated, UA and OA, manifested potent antioxidant effects. The different position of one methyl group in their chemical structures caused moderately different biological activities of these compounds on three leukemic cell lines. To explore their mechanisms of action further investigation seems to be therefore worthwhile.     

Anti-genotoxic effect of Aloysia triphylla infusion against acrylamide-induced DNA damage as shown by the comet assay technique

Aloysia triphylla a perennial, bushy plant originally from South America has long been used in traditional medicine. Its aqueous extract contains considerable amounts of polyphenolic compounds, namely flavonoids and phenolic acids. In view of the interest in natural phenolic compounds as antioxidant in preventive medicine, this study was undertaken to investigate the chemoprotective effects of cedron leaves infusion against the genetic damage induced by acrylamide (AA) by using the alkaline version of the comet assay technique. Mice were separated in nine groups (eight animals each): (I) untreated, (II) negative control, (III) treated with infusion of cedron leaves 5%, 20 days twice a day, (IV) treated with AA (5 mg/kg b.w.), (V) treated with AA (20 mg/kg b.w.), (VI) treated with AA (30 mg/kg b.w.), (VII) treated with AA (50 mg/kg b.w.), (VIII) pretreated with infusion and treated with AA (50 mg/kg b.w.) and (IX) positive control (cyclophosphamide, 20 mg/kg b.w.). Three hundred blast cells were digitally evaluated per animal from three different slides (100 each). Media of tail moment (TM) values were analyzed by ANOVA test. No statistical differences (p>0.05) were found between untreated animals, negative control and infusion-treated mice. A single dose of AA-induced genetic damage as revealed by a statistically significant increase in TM values (p<0.01). Pretreatment with infusion prior to AA injection significantly reduces the capacity of AA to induce genetic damage. In these conditions, tail moments values did not differ from data obtained in negative control (p>0.05) and exhibit statistical differences from animals treated only with AA (p<0.01). Cell viability was at least 90% in all cases as measured by the trypan blue exclusion method. The ferric reducing ability of plasma (FRAP) method reveals that the plasma of infusion-treated mice has a significantly higher antioxidant capacity than plasma from controls (p<0.01). The results suggest that the infusion could exerts an in vivo chemo protective action, probably due to its scavenging potency towards free radicals.     

Assessment of the mutagenic, antimutagenic and cytotoxic activities of ethanolic extract of araticum (Annona crassiflora Mart. 1841) by micronucleus test in mice.

A typical Brazilian plant, araticum (Annona crassiflora Mart.), is widely used in humans as therapeutic medicine to treat several diseases such as diarrhea, rheumatism and syphilis. It contains acetogenins which present cytotoxic, antitumogenic, and antiparasitic properties. In this study, mutagenic, antimutagenic and cytotoxic effects of araticum leaves ethanolic extract were evaluated by micronucleus test in mice. To evaluate the mutagenic activity, animals were treated with ethanolic extract of araticum (EEA) using 10, 20, 50, 100 and 160 mg.kg(-1). For all doses, micronucleated polychromatic erythrocytes (MNPCE) frequency was evaluated at 24, 48 and 72 hours after treatment. To evaluate the antimutagenic activity, animals were treated with 10, 20, 50 and 100 mg.kg(-1) of EEA and 4 mg.kg(-1) of MMC simultaneously. The frequency of MNPCE was evaluated 36 hours after exposure. Cytotoxicity was evaluated by the polychromatic and normochromatic erythrocytes ratio (PCE/NCE). In the mutagenicity assessment, all doses of EEA resulted in no significant increase of MNPCE (P > 0.05), compared to solvent- control group. Regarding administration time, no significant difference among three evaluation periods was observed (P > 0.05). Such results indicate that EEA did not exert mutagenic activity. Cytotoxicity was evident in doses of 50, 100 and 160 mg.kg(-1) at 24 and 48 hours after exposure. Concerning antimutagenicity, except the 10 mg.kg(-1) co-administered with 4 mg/kg of MMC, all doses reduced significantly the frequency of MNPCE compared to the positive control group (P < 0.05). These results, therefore, indicate an antimutagenic activity of the EEA. Cytotoxicity was significantly increased (P < 0.01) at 100 mg.kg(-1) EEA doses co-administered with 4 mg.kg(-1) of MMC.     

Effect of glutathione on mitomycin C-induced micronuclei in bone marrow erythrocytes of Swiss albino mice

The antimutagenic potential of glutathione (GSH) on mitomycin C (MMC)-induced micronuclei was evaluated in Swiss albino mice using the in vivo bone marrow micronucleus test. Six groups of animals were maintained simultaneously. The first group received distilled water only, the second group of animals received 2 mg/kg MMC and the third group was administered 4 doses of GSH, i.e., 20, 40, 80 and 160 mg/kg. The fourth group of animals received GSH and MMC simultaneously. The fifth and sixth groups received a cumulative dose of GSH followed by MMC after 24 h. The fifth group of animals were killed 6 h after the administration of MMC, while the sixth group were killed 24 h after the administration of MMC. The results clearly show a statistically significant increase in micronuclei in MMC-treated animals and also in animals that received GSH followed by MMC. However, there was a decrease in micronuclei in animals that received GSH and MMC simultaneously. The results clearly indicate that GSH exhibits an antimutagenic property in the presence of MMC. It is also observed the treatment with GSH prior to MMC does have some protective effect.     

Evaluation of the genotoxic and antigenotoxic effects after acute and subacute treatments with açai pulp (Euterpe oleracea Mart.) on mice using the erythrocytes micronucleus test and the comet assay

Açai, the fruit of a palm native to the Amazonian basin, is widely distributed in northern South America, where it has considerable economic importance. Whereas individual polyphenolics compounds in açai have been extensively evaluated, studies of the intact fruit and its biological properties are lacking. Therefore, the present study was undertaken to investigate the in vivo genotoxicity of açai and its possible antigenotoxicity on doxorubicin (DXR)-induced DNA damage. The açai pulp doses selected were 3.33, 10.0 and 16.67 g/kg b.w. administered by gavage alone or prior to DXR (16 mg/kg b.w.) administered by intraperitoneal injection. Swiss albino mice were distributed in eight groups for acute treatment with açai pulp (24 h) and eight groups for subacute treatment (daily for 14 consecutive days) before euthanasia. The negative control groups were treated in a similar way. The results of chemical analysis suggested the presence of carotenoids, anthocyanins, phenolic, and flavonoids in açai pulp. The endpoints analyzed were micronucleus induction in bone marrow and peripheral blood cells polychromatic erythrocytes, and DNA damage in peripheral blood, liver and kidney cells assessed using the alkaline (pH >13) comet assay. There were no statistically significant differences (p > 0.05) between the negative control and the groups treated with the three doses of açai pulp alone in all endpoints analyzed, demonstrating the absence of genotoxic effects. The protective effects of açai pulp were observed in both acute and subacute treatments, when administered prior to DXR. In general, subacute treatment provided greater efficiency in protecting against DXR-induced DNA damage in liver and kidney cells. These protective effects can be explained as the result of the phytochemicals present in açai pulp. These results will be applied to the developmental of food with functional characteristics, as well as to explore the characteristics of açai as a health promoter.     

Evaluation of the mutagenicity and antimutagenicity of Ziziphus joazeiro Mart. bark in the micronucleus assay

The aim of this study was to evaluate the mutagenicity (clastogenicity/aneugenicity) of a glycolic extract of Ziziphus joazeiro bark (GEZJ) by the micronucleus assay in mice bone marrow. Antimutagenic activity was also assessed using treatments associated with GEZJ and doxorubicin (DXR). Mice were evaluated 24–48 h after exposure to positive (N-nitroso-N-ethylurea, NEU - 50 mg.kg⁻¹ and DXR - 5 mg.kg⁻¹) and negative (150 mM NaCl) controls, as well as treatment with GEZJ (0.5–2 g.kg⁻¹), GEZJ (2 g.kg⁻¹) + NEU and GEZJ (2 g.kg⁻¹) + DXR. There were no significant differences in the frequencies of micronucleated polychromatic erythrocytes in mice treated with GEJZ and GEJZ + DXR compared to the negative controls, indicating that GEZJ was not mutagenic. Analysis of the polychromatic:normochromatic erythrocyte ratio revealed significant differences in the responses to doses of 0.5 g.kg⁻¹ and 1–2 g.kg⁻¹ and the positive control (NEU). These results indicated no systemic toxicity and moderate toxicity at lower and higher doses of GEZJ. The lack of mutagenicity and systemic toxicity in the antimutagenic assays, especially for treatment with GEZJ + DXR, suggested that phytochemical compounds in Z. joazeiro bark attenuated DXR-induced mutagenicity and the moderate systemic toxicity of a high dose of Z. joazeiro bark (2 g.kg⁻¹). Further studies on the genotoxicity of Z. joazeiro extracts are necessary to establish the possible health risk in humans and to determine the potential as a chemopreventive agent for therapeutic use.     

Cardiotonic and antidysrhythmic effects of oleanolic and ursolic acids, methyl maslinate and uvaol.

The cardiotonic and antidysrhythmic effects of four triterpenoid derivatives, namely oleanolic acid (OA), ursolic acid (UA), and uvaol (UV), isolated from the leaves of African wild olive (Olea europaea, subsp. africana) as well as methyl maslinate (MM) isolated from the leaves of Olea europaea (Cape cultivar) were examined. The derivatives showed low toxicity on brine shrimp test. They displayed significant, dose-response vasodepressor effect and sinus bradicardia, most prominent for OA and MM. The derivatives acted as beta-adrenergic antagonists, blocking the effect of adrenaline and isoprenaline. The established positive inotropic and dromotropic effects were most distinctive for OA and MM. The antidysrhythmic effects were evaluated on CaCl2- and adrenaline-induced chemical arrhythmias, and on ischemia-reperfusion arrhythmia. OA and UA displayed antidysrhythmic effects on both types of chemical arrhythmia; OA and UV in dose 40 mg/kg conferred significant antidysrhythmic activity on ischemia and reperfusion arrhythmias. The effect was comparable to that of propranolol and suggestive of beta-adrenergic antagonistic activity. On the basis of the vasodepressor, cardiotonic and antidysrhythmic effects of these compounds, it was concluded that OA and UV isolated from wild African olive leaves, or crude extract containing all components, can provide a cheap and accessible source of additive to conventional treatment of hypertension, complicated by stenocardia and cardiac failure.     

Anesthetic Effects of a Mixture of Medetomidine,Midazolam and Butorphanol in Two Strains of Mice

The combination of ketamine and xylazine is a widely used anesthetic for laboratory animals. However, due to an abuse problem in Japan, ketamine has been specified as a narcotic since 2007. Instead of using ketamine, Kawai et al. reported an injectable formula with an equivalent effect to the mixture of ketamine and xylazine [11]. The mixture of 0.3 mg/kg body weight (b.w.) medetomidine (Med.), 4.0 mg/kg b.w. midazoram (Mid.), and 5.0 mg/kg b.w. butorphanol (But.) produced an anesthetic duration of around 40 min in outbred ICR mice. However, the anesthetic effect of the mixture for inbred mice strains remains unknown. Therefore, we examined anesthetic effects of the mixture of Med., Mid., and But. in the BALB/c and C57BL/6J strains. After intraperitoneal injection into mice, right front paw, left hind paw, and tail pinch reflexes as well as corneal and righting reflexes were observed. Every 5 min, we scored each reflex category as 0 for reaction or 1 for no reaction. As long as the total score was at least 4 out of 5, we considered the mixture as putting a mouse in a surgical anesthetic state. The mixture produced an anesthetic duration of more than 45 min in both strains of mice. These results indicate that the mixture of Med., Mid., and But. can be a useful and effective anesthesia for the BALB/c and C57BL/6J strains of inbred mice as well as outbred ICR mice.     

In vivo cytogenetic effects of a commercially formulated mixture of cypermethrin and quinalphos in mice

In vivo cytogenetic effects of commercially formulated cypermethrin (CYP, synthetic pyrethroid insecticide) and/or quinalphos (QUI, organophosphate insecticide), generally used in combination, were examined through chromosomal aberrations (CA) and micronucleus test (MT) in mice. Male mice were orally gavaged to a single dose of CYP/QUI commercial mixture (22, 44 or 67 mg/kg b.wt.) for 24h (CA) or 48 h (MT). Based on the concentrations of active ingredients of CYP and QUI present in the test doses of CYP/QUI mixture, mice were orally exposed to 0.66, 1.32 and 2 mg/kg of CYP or 4.4, 8.8 and 13.4 mg/kg of QUI. For reference, a group of five mice was intraperitoneally administered to cyclophosphamide (20 or 50 mg/kg) or orally gavaged to peanut oil for vehicle control. Exposure of CYP/QUI mixture inhibited the mitotic index (MI) and induced CA in a dose-dependent manner at 24 h; however, significant (p<0.01 or 0.001) frequencies of CA were observed at 44 mg/kg onwards, whereas inhibition of MI at 67 mg/kg. Independent exposure of QUI at 8.8 mg/kg onwards also significantly (p<0.01 or 0.001) inhibited MI and induced CA, whereas CYP at 2 mg/kg (highest concentration in CYP/QUI mixture) inhibited MI significantly but failed to induce CA. Chromatid breaks and fragments found to be frequent aberrations in all the test groups. Treatment of CYP/QUI mixture also induced micronucleus formation dose-dependently at 48 h, yet statistically significant (p<0.001) frequencies of micronucleated polychromatic erythrocytes (MNPCE) were observed at 44 mg/kg onwards. QUI (8.8 and 13.4 mg/kg) alone also induced significant frequencies of MNPCE, whereas frequencies of MNPCE observed with the CYP even at 2 mg/kg were comparable to that of vehicle control. Present findings indicate the genotoxicity potential of CYP/QUI mixture and suggest that the simultaneous presence of the toxic doses of CYP and QUI can lead to synergistic genotoxicity in mice and may pose mutagenic risk in human beings.     

Antibacterial activity of oleanolic and ursolic acids and their derivatives

Bacterial resistance to antibiotics is increasing at an alarming rate and many commonly used antibiotics are no longer effective. Thus, there is considerable interest in investigating novel antibacterial compounds, such as the plant-derived pentacyclic triterpenoids, including oleanolic acid (OA), ursolic acid (UA) and their derivatives. These compounds can be isolated from many medicinal and crop plants and their antibacterial, antiviral, antiulcer and anti-inflammatory effects are well documented. OA and UA are active against many bacterial species, particularly Gram-positive species, including mycobacteria. They inhibit bacterial growth and survival, and the spectrum of minimal inhibitory concentration (MIC) values is very broad. In addition, OA, UA and their derivatives display potent antimutagenic activity. Studies to identify the cellular targets and molecular mechanisms of OA and UA action were initiated a few years ago and it has already been demonstrated that both acids influence bacterial gene expression, the formation and maintenance of biofilms, cell autolysis and peptidoglycan turnover. Before these compounds can be used clinically as antimicrobial agents, further extensive studies are required to determine their cytotoxicity and the optimum mode of their application.