Mouse Micronucleus Research Articles

In vitro and in vivo assessment of the mutagenic activity of N-[ N-[3-(3-hydroxy-4-methoxyphenyl) propyl]-α-aspartyl]- l-phenylalanine 1-methyl ester, monohydrate (advantame)

Advantame (N-[ N-[3-(3-hydroxy-4-methoxyphenyl) propyl]-α-aspartyl]- l-phenylalanine 1-methyl ester, monohydrate), an N-substituted analogue of aspartame, has been developed as a high-intensity sweetener. It is approximately 100 and 20,000 times sweeter than aspartame and sucrose, respectively. In this study the safety of advantame has been evaluated using a series of in vitro and in vivo genotoxicity assays including, bacterial mutation, mammalian cell mutation, and mouse micronucleus tests. Advantame did not induce reverse mutations in Salmonella typhimurium and Escherichia coli at concentrations of up to 5000 μg/plate. In the mammalian cell mutation assay, advantame did not induce mutation at the Hprt locus of L5178Y mouse lymphoma cells in two independent experiments, either in the absence or presence of S9. In vivo, there was no effect on the incidence of micronucleated immature or mature erythrocytes in bone marrow after oral administration of the test substance at any dose level (up to 2000 mg/kg body weight) or sampling time (24 and 48 h). The results of these studies demonstrate that advantame is without genotoxic potential.

English

The aim of this study was to examine the mechanism of green tea in treating a broad spectrum of human diseases and to develop a safe and effective agent for antimicrobial and anti-inflammatory application. Catechin-enriched green tea extract (CEGTE) was prepared containing 83.15 ± 0.79% of total catechins. In vitro antimicrobial and in vivo anti-inflammatory effects were investigated and safety was evaluated in vivo with relevant positive controls. CEGTE was found to have significant bacteriostatic effects in all the bacteria isolates tested and inhibitory effects on influenza virus A3 (H3N2) and coxsackie virus B4. CEGTE showed inhibitory effects both on carrageenin-induced paw edema in rats and xylene-induced ear edema in mice and exhibited an inhibitory response against acetic acid-induced writhing in mice. The oral LD50 of CEGTE in both Sprague Dawley (SD) rats and imprinting control region (ICR) mice was determined to be greater than 20000 mg/kg body weight. The bacterial Ames test and mammalian micronucleus and sperm abnormality assay in mice showed no mutagenic properties with CEGTE. CEGTE can be applied as a safe and effective antimicrobial and anti-inflammatory agent with advantages over medicines which treat bacterial and viral infection or inflammation alone. Key words: Catechin-enriched green tea extract (CEGTE), antimicrobial, anti-inflammatory, safety evaluation.

Genotoxicity assessment of ethylenediamine dinitrate (EDDN) and diethylenetriamine trinitrate (DETN)

Ethylenediamine dinitrate (EDDN) and diethylenetriamine trinitrate (DETN) are relatively insensitive explosive compounds that are being explored as safe alternatives to other more sensitive compounds. When used in combination with other high explosives they are an improvement and may provide additional safety during storage and use. The genetic toxicity of these compounds was evaluated to predict the potential adverse human health effects from exposure by using a standard genetic toxicity test battery which included: a gene mutation test in bacteria (Ames), an in vitro Chinese Hamster Ovary (CHO) cell chromosome aberration test and an in vivo mouse micronucleus test. The results of the Ames test showed that EDDN increased the mean number of revertants per plate with strain TA100, without activation, at 5000μg/plate compared to the solvent control, which indicated a positive result. No positive results were observed with the other tester strains with or without activation in Salmonella typhimurium strains TA98, TA1535, TA1537, and Escherichia coli strain WP2 uvrA. DETN was negative for all Salmonella tester strains and E. coli up to 5000μg/plate both with and without metabolic activation. The CHO cell chromosome aberration assay was performed using EDDN and DETN at concentrations up to 5000μg/mL. The results indicate that these compounds did not induce structural chromosomal aberrations at all tested concentrations in CHO cells, with or without metabolic activation. EDDN and DETN, when tested in vivo in the CD-1 mouse at doses up to 2000mg/kg, did not induce any significant increase in the number of micronuclei in bone marrow erythrocytes. These studies demonstrate that EDDN is mutagenic in one strain of Salmonella (TA100) but was negative in other strains, for in vitro induction of chromosomal aberrations in CHO cells, and for micronuclei in the in vivo mouse micronucleus assay. DETN was not genotoxic in all in vitro and in vivo tests. These results show the in vitro and in vivo genotoxicity potential of these chemicals.

Genotoxicity potential of a new natural formicide

Assessment of environmental impacts from pesticide utilization should include genotoxicity studies, where the possible effects of mutagenic/genotoxic substances on individuals are assessed. In this study, the genotoxicity profile of the new formicide Macex® was evaluated with two genotoxicity tests, namely, the micronucleus test with mouse bone marrow and Vicia faba, and a mutagenicity test using the Ames Salmonella assay. The bacterial reverse mutation test (Salmonella typhimurium strains TA97, TA98, TA100, TA102, and TA1535), the Vicia root tip and mouse micronucleus tests were conducted according to published protocols. In the range of the formicide Macex® concentrations tested from 0.06 to 1.0 g L⁻¹ (or mgkg⁻¹ in the mouse test), no genotoxicity was observed in the prokaryotic or eukaryotic test organisms. However, at Macex® concentrations of 0.5 g L⁻¹ and above a significant decrease in the mitotic index (P ≤ 0.05) in the V. faba was observed. Micronucleus formation was likewise increased in the test organism at concentrations starting at 2.0 g L⁻¹. These data allow us to classify this natural formicide preparation as a product with no geno-environmental-impact when applied at recommended concentrations.

Erratum for: Antimutagenic Potential ofSolanum lycocarpumagainst Induction of Chromosomal Aberrations in V79 Cells and Micronuclei in Mice by Doxorubicin

Correction to: Antimutagenic Potential of Solanum lycocarpum against Induction of Chromosomal Aberrations in V79 Cells and Micronuclei in Mice by DoxorubicinPlanta Med 2011; 77(13): 1489-1494DOI: 10.1055/s-0030-1270886

In vitro and in vivo assessment of the genotoxic activity of aloesin

Aloesin is a chromone that is a component of Aloe spp. It may have potential as a functional food ingredient as it has been shown to likely have beneficial effects in persons in a pre-diabetic state or who have metabolic syndrome. In this study the safety of aloesin has been evaluated using a series of in vitro and in vivo genotoxicity assays including, bacterial mutation, mammalian cell cytogenetic, and mouse micronucleus tests. Aloesin did not induce reverse mutations in Salmonella typhimurium and Escherichia coli at any of the tested dose levels up to 10,000μg/plate. Similarly, aloesin did not increase the incidence of chromosome aberrations when incubated with Chinese hamster lung cells at any of the tested concentrations up to 10,000μg/mL. In vivo, there was no effect of aloesin on the incidence of micronucleated erythrocytes following oral administration on two consecutive days at doses up to 5000mg/kg body weight. There was no evidence of toxicity to bone marrow. The results of these studies demonstrate that aloesin is without genotoxic potential.

Safety evaluation of steroidal saponin DT-13 isolated from the tuber of Liriope muscari (Decne.) Baily

Steroidal saponin DT-13 (25 (R, S)-ruscogenin-1-O-[β-d-glucopyranosyl - (1→2)] [β-d-xylopyranosyl-(1→3)]-β-d-fucopyranoside) is the main active component of the tube of Liriope muscari (Decne.) Baily and has been studied as a candidate drug for cancer metastasis. The objective of this study was to evaluate the safety of DT-13 systematically by genotoxicity and acute oral toxicity and subchronic 90-day oral gavage toxicity. Results of Ames test confirmed that DT-13 did not induce mutations in histidine auxotrophs Salmonella typhimurium (TA 97, TA 98, TA 100 and TA 102) both in the presence and absence of metabolic activation system at the doses of 0.05–500μg/plate. Meanwhile, DT-13 did not induce clastogenicity at doses of 1250, 2500 and 5000mg/kg in mouse micronucleus test. And the single oral dose of DT-13 at 5000mg/kg did not produce mortality or significant changes in the general behavior and gross appearance of the internal organs of mice. In subchronic toxicity study, DT-13 was administrated to Sprague–Dawley rats via oral gavage at doses of 10, 60 and 360mg/kg for 90days. Necropsy, hematological and biochemical analysis, and histopathological examination did not reveal any remarkable and treatment related changes. In conclusion, DT-13 is of low toxicity at the tested doses.

Micronucleus Test of Picrorrhiza Rhizoma Aqueous Extract in Bone Marrow Cells of Male ICR Mice.

In this research, the genotoxic effect of Picrorrhiza Rhizoma (PR) aqueous extract was evaluated using the mouse micronucleus test. PR extract was administered once a day for 2 continuous days by oral gavage to male ICR mice at doses of 2000, 1000 and 500 mg/kg. Cyclophosphamide was used as a known genotoxic agent in a positive control. The appearance of a micronucleus (MN) in polychromatic erythrocyte (PCE) is used as an index for genotoxic potential, and PCE ratio is used as an index of cytotoxicity. Although significant (p < 0.01) increase of the number of PCE with one or more nuclei (MNPCE) was detected in cyclophosphamide treated groups, no significant increases of MNPCE numbers were observed in all three different dosages of PR extracts treated mice with over 0.39 of the individual polychromatic erythrocyte ratio in all mice used in this study. The results obtained indicated that PR extract shows no genotoxicity effects up to 2000 mg/kg dosing levels.

Overexpression of eIF-5A2 in mice causes accelerated organismal aging by increasing chromosome instability

BackgroundAmplification of 3q26 is one of the most frequent genetic alterations in many human malignancies. Recently, we isolated a novel oncogene eIF-5A2 within the 3q26 region. Functional study has demonstrated the oncogenic role of eIF-5A2 in the initiation and progression of human cancers. In the present study, we aim to investigate the physiological and pathological effect of eIF-5A2 in an eIF-5A2 transgenic mouse model.MethodsAn eIF-5A2 transgenic mouse model was generated using human eIF-5A2 cDNA. The eIF-5A2 transgenic mice were characterized by histological and immunohistochemistry analyses. The aging phenotypes were further characterized by wound healing, bone X-ray imaging and calcification analysis. Mouse embryo fibroblasts (MEF) were isolated to further investigate molecular mechanism of eIF-5A2 in aging.ResultsInstead of resulting in spontaneous tumor formation, overexpression of eIF-5A2 accelerated the aging process in adult transgenic mice. This included decreased growth rate and body weight, shortened life span, kyphosis, osteoporosis, delay of wound healing and ossification. Investigation of the correlation between cellular senescence and aging showed that cellular senescence is not required for the aging phenotypes in eIF-5A2 mice. Interestingly, we found that activation of eIF-5A2 repressed p19 level and therefore destabilized p53 in transgenic mouse embryo fibroblast (MEF) cells. This subsequently allowed for the accumulation of chromosomal instability, such as errors in cell dividing during metaphase and anaphase. Additionally, a significantly increase in number of aneuploidy cells (p < 0.05) resulted from an increase in the incidences of misaligned and lagging chromosomal materials, anaphase bridges, and micronuclei in the transgenic mice.ConclusionThese observations suggest that eIF-5A2 mouse models could accelerate organismal aging by increasing chromosome instability.

Melamine induces sperm DNA damage and abnormality, but not genetic toxicity

Melamine is a category III carcinogen. However, its illegal addition to milk and other protein products has led to the pet melamine poisoning in USA and infant renal calculus cases in China. Its long-term toxicity is not clear and needs to be investigated urgently to appease the public panic. In this study, the effects of melamine on mutagenesis in prokaryotes and eukaryotes were tested by Ames test, in vitro mammalian chromosome aberration test, mouse micronucleus test and sperm abnormality test; the effects of melamine on DNA damage in sperm cells were observed by single cell gel electrophoresis, and the effects on malignant transformation were examined by malignant transformation experiment of normal human liver cell line L02. The results show that melamine has no mutagenic function in prokaryotes and eukaryotes in vitro and in vivo, and does not induce malignant cell transformation after long-term exposure. However, it has ability to increase sperm abnormality rate and DNA damage.

Antimutagenic Potential ofSolanum lycocarpumagainst Induction of Chromosomal Aberrations in V79 Cells and Micronuclei in Mice by Doxorubicin

Solanum lycocarpum A. St. Hil. (Solanaceae) is a hairy shrub or small much-branched tree of the Brazilian Cerrado. S. lycocarpum fruits are commonly used in traditional medicine in powder form or as folk preparations for the treatment of diabetes and obesity, as well as for controlling cholesterol levels. The aim of the present study was to chemically characterize the hydroalcoholic extract (SL) of S. lycocarpum by determination of total flavonoids and total poyphenols and quantification of steroidal alkaloids, as well as to evaluate its mutagenic and/or antimutagenic potential on V79 cells and Swiss mice using chromosomal aberrations and bone marrow micronucleus assays, respectively. Three concentrations of SL (16, 32, and 24 µg/mL) were used for the evaluation of its mutagenic potential in V79 cells and four doses (0.25, 0.50, 1.0, and 2.0 g/kg body weight) were used for Swiss mice. In the antimutagenicity assays, the different concentrations of SL were combined with the chemotherapeutic agent doxorubicin (DXR). HPLC analysis of SL gave contents of 6.57 % ± 0.41 of solasonine and 4.60 % ± 0.40 of solamargine. Total flavonoids and polyphenols contents in SL were 0.04 and 3.60 %, respectively. The results showed that not only SL exerted no mutagenic effect, but it also significantly reduced the frequency of chromosomal aberrations induced by DXR in both V79 cells and micronuclei in Swiss mice at the doses tested.

Antigenotoxicity of artepillin C in vivo evaluated by the micronucleus and comet assays

Artepillin C (3,5-diprenyl-p-coumaric acid), a major compound found in Brazilian green propolis and Baccharis dracunculifolia, shows anti-inflammatory, antibacterial, antiviral, antioxidant and antitumoral activities, among others. The aim of this study was to evaluate the genotoxic potential of artepillin C and its ability to prevent the chemically induced chromosome breakage or loss and the primary DNA damage using the micronucleus and comet assays in male Swiss mice, respectively. The animals were treated by gavage with different doses of artepillin C (0.4, 0.8 and 1.6 mg kg(-1) b.w.). For the antigenotoxicity assays, the different doses of artepillin C were administered simultaneously to doxorubicin (DXR; micronucleus test; 15 mg kg(-1) b.w.) and to methyl methanesulfonate (MMS; comet assay; 40 mg kg(-1) b.w.). The results showed that artepillin C itself was not genotoxic in the mouse micronucleus and comet assays. In the animals treated with artepillin C and DXR, the number of micronucleated reticulocytes was significantly lower in comparison with the animals treated only with DXR. Regarding antigenotoxicity, artepillin C at the tested doses significantly reduced the extent of DNA damage in liver cells induced by MMS.

Abstract B61: Chemoprotective properties of some African spices on sodium arsenite-induced toxicities in albino Wistar rats

Abstract Certain types of cancer have been linked to the consumption of arsenic contaminated water. Increases in arsenic-induced carcinogenesis were associated with the presence of inorganic trivalent arsenic as observed during previous epidemiological studies. Consumption of chemoprotective plants food products rich in antioxidants have been suggested and shown to hold promise in mitigating disease onset and chemical-induced carcinogenesis. We investigated the effect of some commonly consumed African spices and nuts — Afromomum longiscapum (AL), Keayodendron bridelioides (KB), Monodora myristica (MM) and Juglans nigra (JN) — on sodium arsenite (SA) a known clastogen induced toxicty in Albino Wistar rats for five weeks. Phytochemical profiling of aqueous extracts of these spices indicated non-specifically the presence and absence of these pharmacologically active compounds: tanins, saponins, cardenolides, anthraquinones, flavonoids and alkaloids. Our findings indicate that MM and AL alone induce the formation of micronucleated polychromatic erythrocytes (mPCEs) in the rat bone marrow significantly (p&amp;lt;0.05) while KB and JN alone did not using the mouse micronuclei assay. Co-administration of AL, MM, KB and JN individually in addition to SA decreased (p&amp;lt;0.05) SA-induced mPCEs formation compared with SA only. KB and JN effectively reduced mPCEs formation (50 and 75% respectively) than AL and MM (38.5 and 31% respectively) over SA alone. However, AL and MM did not additively increase SA-induced mPCEs formation but appears to act in an antagonistic manner. Relative to control, SA increased serum levels of hepatic transaminases (p&amp;lt;0.05) while all extract co-treatments with SA resulted in a decrease (p&amp;lt;0.05) in serum transaminases. AL, KB, MM, JN and SA treatment decreased sperm count and motility (p&amp;lt;0.05) with no observable effect on sperm viability compared with the control. All parameters measuring morphological abnormalities in semen showed no difference (p&amp;gt;0.05) across the treated groups. Limited hepatic histopathology showed mild mononuclear cellular infiltration in the control groups. Compared with the control, severe hepatic degeneration and necrosis was observed in SA treated group with no visible lesions in the AL, KB and JN treatments. However, diffused vascular degeneration of hepatocytes was common in the MM treated group. Mild hepatic degeneration and congestion of the portal vessels were seen in AL+SA and MM+SA treated groups. KB treatment largely conferred protection on SA-induced hepatotoxicity. Overall, some extracts (JN and KB) exhibited an higher anticlastogenic and hepatoprotective potentials, while others (MM and AL) did to a lesser extent confered such properties, bordering on being toxic. Furthermore, the extracts reduced sperm count and motility with no significant effect on viability or morphology. In conclusion, the aqueous extracts of these spices exhibited chemoprotective properties, with the potential of mitigating the effect of arsenic-induced clastogenicity. Citation Information: Cancer Prev Res 2010;3(12 Suppl):B61.

Evaluation of the genotoxicity of Orthosiphon stamineus aqueous extract

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

Genotoxicity and subchronic toxicity evaluation of Active Hexose Correlated Compound (AHCC)

Active Hexose Correlated Compound (AHCC), a mushroom extract rich in α-1,4 linked glucans, is associated with immunostimulatory effects. AHCC is used in Japan as a dietary supplement to boost immune function and it also is purported to improve the symptoms of cancer and liver disease patients. A series of toxicological studies were conducted on a freeze dried preparation of AHCC (AHCC-FD) to further develop the body of evidence supporting the safety of this ingredient. AHCC-FD was not mutagenic to Salmonella typhimurium and did not exhibit clastogenicity in a mouse micronucleus assay. In a 90-day study, Sprague–Dawley rats were administered 1000, 3000, or 6000mg/kg body weight/day by gavage. No changes attributable to AHCC-FD treatment were observed in overall condition, body weight, food consumption, ophthalmology findings, hematology and clinical chemistry parameters, and absolute and relative organ weights. Changes in urinary pH values observed in high-dose animals and mid-dose females were considered physiological rather than adverse effects given the acidic nature of AHCC-FD. Urinary protein also was increased in the same dose groups. As this finding was associated with decreased urinary pH and no evidence of kidney dysfunction was observed, it was considered of no toxicological significance. Histopathological changes related to AHCC-FD administration were observed in the limiting ridge of the stomach and in the liver of the high-dose group. The NOAEL was considered to be 3000mg/kg body weight/day.

Safety assessment of EPA-rich triglyceride oil produced from yeast: Genotoxicity and 28-day oral toxicity in rats

The 28-day repeat-dose oral and genetic toxicity of eicosapentaenoic acid triglyceride oil (EPA oil) produced from genetically modified Yarrowia lipolytica yeast were assessed. Groups of rats received 0 (olive oil), 940, 1880, or 2820mg EPA oil/kg/day, or fish oil (sardine/anchovy source) by oral gavage. Lower total serum cholesterol was seen in all EPA and fish oil groups. Liver weights were increased in the medium and high-dose EPA (male only), and fish oil groups but were considered non-adverse physiologically adaptive responses. Increased thyroid follicular cell hypertrophy was observed in male high-dose EPA and fish oil groups, and was considered to be an adaptive response to high levels of polyunsaturated fatty acids. No adverse test substance-related effects were observed on body weight, nutritional, or other clinical or anatomic pathology parameters. The oil was not mutagenic in the in vitro Ames or mouse lymphoma assay, and was not clastogenic in the in vivo mouse micronucleus test. In conclusion, exposure for 28days to EPA oil derived from yeast did not produce adverse effects at doses up to 2820mg/kg/day and was not genotoxic. The safety profile of the EPA oil in these tests was comparable to a commercial fish oil.

Safety Evaluation of Zinc Threoninate Chelate

The acute toxicity of zinc threoninate chelate was assessed. The oral lethal dose 50% (LD(50)) was 2710 mg/kg in female rats and 3160 mg/kg in male rats. Genotoxicity was assessed by Ames test in Salmonella typhimurium strains TA97, TA98, TA100, and TA102, by bone marrow mouse micronucleus test and a sperm abnormality test with mice. Thirty-day repeat dose toxicity study was conducted at oral daily doses of 0, 42, 169, and 675 mg/kg in rats. Teratogenicity was assessed at the same daily dose in pregnant rats by gavage. No significant changes in body weight, food consumption, organ weight, relative organ weight, hematology, blood biochemistry, histopathology, behavior, mortality, sperm abnormality, mutagenicity, and micronucleus formation were observed and no clinical signs or adverse effects were detected. Zinc threoninate chelate had no significant teratogenic effect at a daily dose of 42 mg/kg.

Intestinal α-glucosidase inhibitory activity and toxicological evaluation of Nymphaea stellata flowers extract

Aim of the study Nymphaea stellata willd. flowers (NSF) are used as a traditional medicine in India and Nepal to treat diabetic disease. Different works have demonstrated that NSF extract showed antihyperglycemic effect on alloxan-induced diabetic rats. In the present work we evaluated in vitro intestinal α-glucosidase inhibition as the possible mode of action of NSF extract on suppressing postprandial hyperglycemia for curing diabetic mellitus. In addition, NSF extract was studied to assess its possible acute oral toxicity and genotoxicity. Materials and methods Rat intestinal crude enzyme preparation and Caco-2 monolayer were used to evaluate α-glucosidase inhibitory activity of NSF extract. The main α-glucosidase inhibitors were detected by HPLC. For acute toxicity test, NSF extract was administered at doses of 2000, 5000 and 10,000 mg/kg body to three groups of 10 ICR mice each, and then clinical symptoms including mortality, clinical sign and gross findings were observed once a day for 14 days. In Ames test, histidine-dependent auxotrophic mutants of Salmonella typhimurium (strains TA97, TA98, TA100, TA102 and TA1535) were used and incubated in the presence and absence of S9 metabolic activation using NSF extract with concentrations of 150–5000 μg/plate. The chromosome aberration test was conducted with Chinese hamster lung (CHL) cells treated with NSF extract at doses of 150–5000 μg/ml in the presence and absence of S9 metabolic activation. In the in vivo mouse micronucleus assay, 9-week-old male and female ICR mice ( n = 90, 25–30 g) were administered daily by oral gavage at doses of 2.5, 5.0 and 10.0 g/kg body for 1 or 2 days. Bone marrow smears were prepared from each treatment group 24 h after last administration and then polychromatic erythrocytes (PCEs) and normochromatic erythrocytes (NCEs) were identified. Results NSF extract showed potent rat intestinal α-glucosidase inhibitory activity for maltose hydrolysis with ED 50 value of 0.1 mg/ml. In Caco-2 monolayer, α-glucosidase activity for the maltose hydrolysis was down-regulated by NSF extract at a concentration of 0.05 mg/well level, showing 74% inhibition compared to the saline treated control. NSF was rich in phenol contents and the main α-glucosidase inhibitor, 1,2,3,4,6-penta-O-galloyl-β- d-glucose, was identified together with two phenolic compounds of gallic acid and corilagin. In acute toxicity test, NSF extract did not produce any toxic signs or deaths and the LD 50 value of this extract could be greater than 10,000 mg/kg body weight. These results of genotoxicity assessment showed that NSF extract did not cause genotoxic effects in Ames test, in the in vitro chromosomal aberration assay and in the in vivo micronucleus assay. Conclusion The current study shows that the extract from Nymphaea stellata flowers exhibits significant intestinal α-glucosidase inhibitory activity, without showing any acute toxicity or genotoxicity, which may be useful in suppressing postprandial hyperglycemia in diabetics. The results presented here suggest that the use of NSF in folk medicine as a natural antidiabetic treatment could be safe as well as beneficial.

Safety evaluation of an açai-fortified fruit and berry functional juice beverage (MonaVie Active ®)

The safety of an açai ( Euterpe oleracea Mart.) pulp enriched fruit and berry juice, MonaVie Active ®, fortified with the functional ingredient, glucosamine, was studied. The beverage was found not to be mutagenic, clastogenic, cytotoxic, or genotoxic, as determined by the bacterial reverse mutation assay, chromosomal aberration assay, mouse micronucleus assay, and mammalian cell gene mutation (L5178Y) assay. The single dose LD50 based on a 14-day acute oral toxicity study is greater than 20,000 mg/kg bw, the highest dose tested. In a repeat dose 90-day oral subchronic toxicity study by gavage, 220 animals were randomly assigned to a control group, an untreated group, or one of three experimental groups (10, 20 and 40 g/kg bw). No treatment-related significant changes in body weight, food and water consumption, ophthalmology, organ weights, urinanalysis, hematological and clinical chemistry, or gross pathology, were observed in surviving animals compared to the control groups. Three animals died midway through the observation period (male, 20 g/kg bw/day; male 40 g/kg bw/day; and, female, 10 g/kg bw/day). These animals died without preceding clinical symptoms, histopathological lesions, or evidence of injury to tissue or organs except for signs of suffocation/aspiration congestion, which was concluded to be due to problems with the gavage administration of the fluid test article, and not due to the test article itself. The NOEAL was determined to be 40 g/kg bw/day for male and female rats, which was the highest dose tested. Phylloquinone (vitamin K1) content averaged 21.7 μg/100 g, comparable to amounts found in iceberg lettuce. In conclusion, the results provide additional experimental evidence that MonaVie Active ® juice is non-toxic.

Genotoxicity assessment of two hypergolic energetic propellant compounds

Recognition of the occupational hazards from exposure to the propellants hydrazine and monomethylhydrazine (MMH) has led to research into less toxic alternatives. Two hypergolic compounds, dimethylamino-2-ethylazide (DMAZ) and N,N,N′,N′-tetramethylethanediamine (TMEDA), have been identified as possible replacements for MMH. We have obtained genotoxicity data for these compounds from in vitro and in vivo studies. DMAZ did not produce any mutagenic effects at concentrations up to 5 mg/plate in the TA98 and TA1537 strains of Salmonella typhimurium and in an Escherichia coli (WP2 uvrA) strain, with or without metabolic activation, but did produce a positive response in the TA100 and TA1535 strains, both with and without metabolic activation. TMEDA was found not to be mutagenic in any of the bacterial strains tested ( Salmonella TA98, TA100, TA1535, TA1537 and E. coli, WP2 uvrA), with or without metabolic activation. DMAZ did not induce structural chromosomal aberrations at levels up to 5 mg/mL in Chinese hamster ovary (CHO) cells, with or without metabolic activation. TMEDA produced a positive response in this system, with or without metabolic activation, but only at the highest concentration, 5 mg/mL. However, according to the OECD guideline TG 473, the compound is considered to be negative in the CHO chromosomal aberration assay, since the compound was not clastogenic at 0.01 M (1.140 mg/mL). DMAZ and TMEDA, when tested in vivo in the CD-1 mouse at doses up to 500 and 250 mg/kg, respectively, did not induce micronuclei in bone marrow erythrocytes. These studies demonstrate that DMAZ is mutagenic in specific strains of Salmonella. However, both compounds were negative for induction of chromosomal aberrations in CHO cells in vitro and in the in vivo mouse micronucleus assay.