In Vivo Micronucleus Assay and GST Activity in Assessing Genotoxicity of Plumbagin in Swiss Albino Mice

Information available on the mutagenicity of a large number of indigenous drugs commonly employed in the Siddha and Ayurveda systems of medicine is scanty. In this context, the current investigation on plumbagin, 5-hydroxy-2methyl-1,4-napthoquinone, an active principle in the roots of Plumbago zeylanica used in Siddha and Ayurveda for various ailments, was carried out; 16 mg/kg b.w. (LD₅₀) was fixed as the maximum dose. Subsequent dose levels were fixed as 50% and 25% of LD₅₀ amounting to 8 mg and 4 mg/kg b.w., respectively, and given orally for 5 consecutive days in 1% Carboxyl Methyl Cellulose (CMC) to Swiss albino mice weighing 25–30 g. The micronucleus assay was done in mouse bone marrow. Plumbagin was found to induce micronuclei at all the doses studied (4 mg/kg, 8 mg/kg, 16 mg/kg b.w.), and it proves to be toxic to bone marrow cells of Swiss albino mice. Animal treated with cyclophosphamide (40 mg/kg b.w.) served as positive control. In addition, glutathione S-transferase (GST) activity was observed in control, plumbagin (4 mg, 8 mg, 16 mg/kg b.w., respectively), and genotoxin-treated experimental group of animals. No significant change in GST activity was observed with plumbagin dose of 4 mg/kg b.w., whereas GST activity was significantly inhibited by higher doses of plumbagin (8 mg and 16 mg/kg b.w.) and cyclophosphamide.     

Restoration of histoarchitecture in the paracetamol-induced liver damaged rat by earthworm extract, Lampito mauritii (Kinberg)

Lampito mauritii, an earthworm widely used in Siddha as well as Ayurveda, has been reported to have anti-inflammatory, anti-ulceral and anti-oxidative properties. The present investigation shows its hepatoprotective role. Whole tissue extract of Lampito mauritii protect the paracetamol (2 g/kg body weight. p.o.) induced liver damage rat as a dose dependent manner (100, 200 and 300 mg/kg body weight p.o.) by significantly (P < 0.05) decreasing the hepatic marker enzymes--aspartate transaminase, alanine transaminase and alkaline phosphatase similar to silymarin (150 mg/kg b.w. p.o.). Histopathological observations of liver tissues corroborated these findings.     

Anticlastogenic activity of morin against whole body gamma irradiation in Swiss albino mice

Anticlastogenic activity of morin was explored against whole body gamma radiation, at a dose rate of 1.66 Gy/min in Swiss albino mice pretreated intraperitoneal or orally. Pretreatment with morin 10, 25, 50, 75, 100, 125, and 150 mg/kg, i.p. delayed and reduced percentage mortality and increased mean survival times in mice irradiated with 10 Gy gamma radiation. Intraperitoneal route was found superior to oral route. An i.p. dose of 100 mg/kg was found to be the most effective dose in preventing radiation-induced weight loss, increasing the mean survival times and reducing percentage mortality. Morin (100 mg/kg) pretreatment effectively maintained spleen index (spleen weight/body weight x 100) and stimulated endogenous spleen colony forming units. Pretreatment with morin (100 mg/kg) significantly reduced dead, inflammatory, and mitotic cells in irradiated mice jejunum along with a significant increase in goblet cells and rapidly multiplying crypt cells. Morin (100 mg/kg) also maintained the villus height close to normal, prevented mucosal erosion and basement membrane damage in irradiated jejunum. Nuclear enlargement in epithelial cells of jejunum was lower in morin treated mice compared to radiation control. Morin (100 mg/kg) also significantly elevated the endogenous antioxidant enzymes viz. glutathione S transferase (GST), superoxide dismutase (SOD) and reduced glutathione (GSH), in normal mice at 2, 4 and 8 h post treatment. Drastic decrease in endogenous enzymes (GSH, GST, catalase and SOD) and total thiols was observed in irradiated mice at 2, 4 and 8 h post irradiation, while pretreatment with morin (100 mg/kg) prevented this decrease. Morin (100 mg/kg) also elevated radiation LD(50) from 9.2 to 10.1 Gy, indicating a dose modifying factor (DMF) of 1.11.     

Assessment of acetamiprid-induced genotoxic effects in bone marrow cells of Swiss albino male mice

Acetamiprid (ACE), a neonicotinoid insecticide, is widely used in agriculture either alone or in combination with other insecticides. A combined approach employing micronucleus test (MNT) and chromosomal aberrations (CA) assay was utilized to assess the genotoxic effects of ACE in bone marrow of Swiss albino male mice. Acetamiprid was administered i.p. daily at 4.6 and 2.3?mg/kg/day along with 3% gum acacia as negative control for 60 and 90?days and cyclophosphamide (50?mg/kg b.wt.) as positive control. ACE treatment resulted in a dose-dependent increase in the frequencies of micronuclei per cell and chromosomal aberrations in bone marrow cells. The increased micronuclei formation in total erythrocyte cells (immature PCEs and mature NCEs) was observed only at higher dose level (4.6?mg/kg b.wt.) administered for 90?days. The test also indicated the cytotoxic effect of higher dose level of pesticide by PCE/NCE ratio. The number of chromosomal aberrations were increased in the pesticide treated group compared to the negative control group, although significant increase was observed only in the group exposed to higher dose level of pesticide for both 60 and 90?days. Thus, daily exposure of ACE at a dose level of 4.6?mg/kg body weight for 60 and 90?days caused genotoxic and cytotoxic effects on the somatic cells of Swiss albino male mice.     

Aegle marmelos (L.) Correa inhibits the proliferation of transplanted Ehrlich ascites carcinoma in mice

The anticancer effect of hydroalcoholic extract of Aegle marmelos (AME) was studied in the Ehrlich ascites carcinoma bearing Swiss albino mice. The spatial effect of various AME administration schedules showed that six-day administration increased the survival of tumor bearing mice. The best antineoplastic action of AME was obtained when AME administered through intraperitoneal route than the oral route at equimolar dose. Administration of AME once daily for six consecutive days to the tumor bearing mice caused a dose dependent remission of the tumor at 400 mg/kg body weight, where the greatest antitumor effect was observed and the higher doses showed toxic manifestations. A 24-d lengthening in life span was observed in EAC animals treated with 400 mg/kg AME. This dose of 400 mg/kg was considered as the best dose, where the animals survived up to 43 d post-tumor-cell inoculation as against no survivors in the saline treated control group. The antitumor activity when tested for different schedules for triple administrations, the best effect was observed for 1-2-3, followed by 1-3-5 and 1-5-9 days, respectively. Stage specific evaluation of AME inhibited the increase in body weight gain in animals due to tumor development during early stages only. The AME treatment resulted in a dose dependent elevation in the median survival time (MST) and average survival time (AST) up to 400 mg/kg AME and decline thereafter. The effective dose of 400 mg of AME is 1/6th of the LD50 dose, which increased the MST and AST up to 29 and 27 d, respectively. The acute toxicity study of AME showed that the drug was non-toxic up to a dose of 1750 mg/kg b. wt. The LD10 and LD50 was found to be 2000 and 2250 mg/kg.     

Role of curcumin on chloroquine phosphate-induced reproductive toxicity

The present study was conducted to screen the efficacy of curcumin against chloroquine phosphate (CQ)-induced reproductive toxicity in adult male Swiss albino mice. Animals were given oral doses of 100, 200, 300?mg/kg body weight (b.w.), and high dose of CQ (300?mg/kg b.w.) + curcumin (80?mg/kg b.w.) for 45 days. Animals of the withdrawal group were given high dose of CQ (300?mg/kg b.w.) for 45 days and, at day 46, were kept for another 45 days. Effects were observed on some key enzymes, such as alkaline phosphatase, which was found to be decreased, whereas acid phosphatase was increased and succinate dehydrogenase and adenosine triphosphatase were decreased. Oxidative parameters, such as superoxide dismutase declined, whereas thiobarbituric acid-reactive substances were found to be elevated. Protein level was also decreased. Gravimetric indices were also recorded. Results obtained indicated adverse effects of CQ in a dose-dependent manner. The presence of curcumin with CQ alleviated its toxic effects. Hence, it can be concluded that curcumin has beneficial influences and appears able to ameliorate CQ toxicity.     

Role of curcumin on chloroquine phosphate-induced reproductive toxicity

The present study was conducted to screen the efficacy of curcumin against chloroquine phosphate (CQ)-induced reproductive toxicity in adult male Swiss albino mice. Animals were given oral doses of 100, 200, 300?mg/kg body weight (b.w.), and high dose of CQ (300?mg/kg b.w.) + curcumin (80?mg/kg b.w.) for 45 days. Animals of the withdrawal group were given high dose of CQ (300?mg/kg b.w.) for 45 days and, at day 46, were kept for another 45 days. Effects were observed on some key enzymes, such as alkaline phosphatase, which was found to be decreased, whereas acid phosphatase was increased and succinate dehydrogenase and adenosine triphosphatase were decreased. Oxidative parameters, such as superoxide dismutase declined, whereas thiobarbituric acid-reactive substances were found to be elevated. Protein level was also decreased. Gravimetric indices were also recorded. Results obtained indicated adverse effects of CQ in a dose-dependent manner. The presence of curcumin with CQ alleviated its toxic effects. Hence, it can be concluded that curcumin has beneficial influences and appears able to ameliorate CQ toxicity.     

In vitro activity of protoanemonin, an antifungal agent

Protoanemonin, the lactone of gamma-hydroxy-vinylacrylic acid, isolated from Pulsatilla alpina has in vitro activity against fungi. The MIC is 15 micrograms/ml and RNA inhibition seems to be the first target of the drug. The LD50 of protoanemonin in male Swiss albino mice was 190 mg/kg.     

Study on dermal toxicity and urinary metabolites of the new insect repellent N,N-diethylphenylacetamide in rabbits

Dermal toxicity of the new multi-insect repellent N,N-diethylphenylacetamide (DEPA) was studied in female rabbits. LD50 of DEPA was estimated to be 3505 mg/kg b.w. On daily topical application for 21 d at a dose of 50 mg/kg b.w., dermal irritancy score and blood chemistry changes indicated that the compound is non-irritant and non-toxic. N-Ethylphenylacetamide and conjugated phenylacetic acid were identified as the urinary metabolites of DEPA by gas-liquid chromatography.     

The evaluation of the acute toxicity and long term safety of hydroalcoholic extract of Sapthaparna (Alstonia scholaris) in mice and rats

The acute and sub-acute toxic effects of various doses of hydroalcoholic extract of Alstonia scholaris (ASE) was studied in mice and rats. The acute toxicity in mice depended on the season of collection of plant. The highest acute toxicity was observed in the ASE prepared from the summer collection followed by winter. The least toxicity was observed in the extract prepared from the bark of A. scholaris collected in the monsoon season. The administration of different doses of ASE showed a dose dependent increase in the toxicity in all species of mice. The Swiss albino mice were found to be the most sensitive followed by the DBA and C(57)BL. The crossbred mice were resistant when compared to the pure inbred strains. The oral administration of ASE was non-toxic up to a dose of 2000 mg/kg b. wt., while maximum number of animals succumbed to death after administration of 1100 mg/kg ASE by intraperitoneal route. The rats were more sensitive than the mice as the LD(50) dose of ASE was lesser for the former than the latter. The sub-acute toxicity in the rats was carried out with 120 and 240 mg/kg b. wt. ASE (1/10th and 1/5th of the LD(50) dose of ASE). The 240 mg was observed to be more toxic than 120 mg/kg ASE since it caused mortality and deformity in various organs of the recipient animals. The various biochemical parameters like AST, ALT, ACP, ALP, CK, LDH, creatinine, urea, ammonia, glucose and LPx were higher at 240 mg/kg ASE when compared with the 120 mg and the non-drug treated animals. In contrast, the total protein, albumin, DNA, RNA, cholesterol, glucose, glutathione, total thiols declined in the 240 mg/kg ASE treated animals when compared with non-drug treated controls. The hematological analysis showed a dose dependent decrease in the RBC, WBC, hemoglobin, neutrophils and monocytes, while a significant increase in the lymphocytes, eosinophils and basophils was observed. The observed toxic effect of ASE may be due to the presence of echitamine. Our studies shows that at high doses, A. scholaris exhibited marked damage to all the major organs of the body.     

Inhibition of genotoxicity by saffron (Crocus sativus L.) in mice

Experiments were carried out to ascertain whether or not saffron (dried stigmas of Crocus sativus L.), a commonly used agent for flavoring and coloring food can exert modulatory effects on the in vivo genotoxicity of cisplatin (CIS), cyclophosphamide (CPH), mitomycin C (MMC) and urethane (URE). For this purpose, Swiss albino mice were pretreated for five consecutive days with three doses (20, 40 and 80 mg/kg body weight) of the aqueous extract of saffron. Genotoxic effects were assessed in the mouse bone marrow micronucleus test. The results obtained suggest that pretreatment with saffron can significantly inhibit the genotoxicity of CIS, CPH, MMC and URE. This inhibitory effect was not always dose-dependent. In addition, the hepatic glutathione S-transferase (GST) activity was assessed in the control and treated animals. No significant change in GST activity was observed after pretreatment with saffron alone. Treatment with the genotoxins alone significantly inhibited GST activity. Saffron pretreatment attenuated the inhibitory effects of the genotoxins on GST activity.     

活性氯化稀土的毒性研究

本文对活性氯化稀土进行了大鼠急性、蓄积毒性及90天喂饲试验。活性氯化稀土大鼠经口LD_(50)为2553mg/kg,属低毒类物质,对大鼠无明显蓄积作用;90天喂饲试验结果表明,剂量为250mg/kgb.w.及以下时,各项生长发育、血液学、生化学及病理组织学指标均未见不良改变,活性氯化稀土混合物大鼠最大无作用剂量为250mg/kg b.w.,人的日容许摄入量建议值为2.5mg/kgb.w.。     

Modification of Cytotoxic Effects of Inorganic Arsenic by a Crude Extract of Allium sativum L. in Mice

Abstract

A crude extract of Allium sativum (100 mg/kg b.w./day) was administered orally to Swiss albino mice with a normal diet for 30 days. Sodium arsenite, a known cytotoxic agent, was given subcutaneously in normal saline to mice (0.1 mg/kg b.w. = 1/50 of LD₅₀) on days 7,14,21 AND 30 of experiments. Chromosomal studies were conducted on bone marrow preparations following the colchicine-air-drying Giemsa schedule. The frequency of chromosomal aberrations was significantly lower in animals maintained on crude plant extract as a dietary supplement during exposure to sodium arsenite as compared to those treated with arsenite alone. A crude extract of Allium sativum thus protects against the clastogenicity of sodium arsenite.     

Amelioration of cyclophosphamide-induced hepatotoxicity by the root extract of Decalepis hamiltonii in mice

Hepatoprotective potential of the aqueous extract of the roots of Decalepis hamiltonii (DHA) against cyclophosphamide (CP)-induced oxidative stress has been investigated in mice. Administration of CP (25 mg/kg b.w., i.p) for 10 days induced hepatic damage as indicated by the serum marker enzymes aspartate and alanine transaminases (AST, ALT), alkaline phosphatase (ALP) and lactate dehydrogenase (LDH). Parallel to these changes CP induced oxidative stress in the liver as evident from the increased lipid peroxidation (LPO), reactive oxygen species (ROS), depletion of glutathione (GSH), and reduced activities of the antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione-S-transferase (GST). Treatment with DHA (50 and 100 mg/kg b.w., po) mitigated the CP-induced oxidative stress. Moreover, expression of genes for the antioxidant enzymes, were down-regulated by CP treatment which was reversed by DHA. Our study shows the DHA protected the liver from toxicity induced by CP and therefore, it could be serve as a safe medicinal supplement during cyclophosphamide chemotherapy.     

Inhibitory effect of Emblica officinalis on the in vivo clastogenicity of benzo[a]pyrene and cyclophosphamide in mice

Benzo[a]pyrene (B[a]P) and cyclophosphamide (CP) are potent carcinogens/mutagens. Effect of Emblica officinalis extract administration on the in vivo genotoxicity of B[a]P and CP was studied using bone marrow chromosomal aberration and micronucleus induction tests in mice. Three doses (50, 250 and 500 mg/kg body weight) of the plant extract were administered orally for 7 consecutive days prior to the administration of single dose of mutagens (B[a]P 125 mg/kg oral; CP 40 mg/kg i.p.). It was found that administration of 250 and 500 mg/kg of E. officinalis extract significantly inhibited the genotoxicity of B[a]P as well as CP in both the assay systems. Administration of 50 mg/kg of the plant extract had no inhibitory effect. Vitamin C, a major constituent of E. officinalis when administered at dose level of 9 mg/kg b.w. (the approximate estimated amount present in the highest dose of plant extract, i.e. 500 mg) for 7 days did inhibit chromosomal aberrations and micronuclei induction, but not in a significant manner. Effect of administration of the abovementioned effective doses (250 and 500 mg/kg oral for 7 days) of plant extract and vitamin C (9 mg/kg oral for 7 days) on the hepatic activation and detoxification enzymes was also studied. Significant induction in the levels of glutathione content (GSH) and of antioxidant and detoxification enzymes viz., glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S transferase (GST) resulted from plant extract treatment to animals. On the other hand, cytochrome P 450 level was significantly decreased in the plant-extract-treated animals. There was no significant change in cytochrome P 450, GSH contents and activities of enzymes on treatment with vitamin C. The data indicate that the possible mechanism of inhibition by plant extract is mediated by its modulatory effect on hepatic activation and disposition processes.     

Genotoxic effect of hydroquinone on the cultured mouse spleenocytes

The daily administration of 0, 0.5, 1, 2, 4 and 8 mg/kg b. wt. of hydroquinone (HQ) to Swiss albino mice, consecutively for 6 days resulted in a dose dependent elevation in the frequency of micronucleated binucleate spleenocytes (MNBNS). This increase in the frequency of MNBNS was significantly higher than that of non-drug treated controls (0 mg/kg). The daily administration of various doses of HQ for 6 days not only increased the frequency of binucleated spleenocytes bearing one micronuclei (MN), but also the BNS bearing two, three, and four MN. The study demonstrates that the daily administration of various doses of HQ for 6 days increases the genotoxic and mutagenic potential in mice.     

Uric acid as an inhibitor of biochemical changes induced by cyclophosphamide in mice.

The biochemical changes induced by uric acid, cyclophosphamide and uric acid plus cyclophosphamide were evaluated in Swiss albino male mice. Uric acid dissolved in water was administered orally in different doses for 7 days. Some mice from each group were injected intraperitoneally with cyclophosphamide (25 mg/kg) and sacrificed after 30 h. The blood of all animals was analyzed for uric acid levels. Uric acid was found not to affect the biosynthesis of nucleic acids and proteins in liver, testes and brain at 10-100 mg/kg/d. Pretreatment with uric acid provided significant protection against cyclophosphamide-induced impairment of DNA, RNA and protein biosynthesis.     

Micronucleus induction by diuron in mouse bone marrow

Diuron, a widely used substituted urea herbicide, induced the formation of micronuclei in bone marrow cells of Swiss mice. A single i.p. dose of 340 mg/kg b.w. diuron which is maximum tolerated dose (MTD) increased significantly the number of micronuclei at 30 h and 48 h time period. The dose of 170 mg/kg b.w. also induced the micronuclei formation in the above time period. However, a dose of 85 mg/kg b.w. was ineffective at the time periods studied. No induction of micronuclei was observed at 72 h time period after all the doses of diuron studied as compared to the solvent control. The diuron-induced frequency of micronucleated erythrocytes was independent of the sex of the test animals.     

Attenuation of cyclophosphamide-induced pulmonary toxicity in Swiss albino mice by naphthalimide-based organoselenium compound 2-(5-selenocyanatopentyl)-benzo[de]isoquinoline 1,3-dione

Abstract

Context: The widely used antineoplastic drug cyclophosphamide causes pulmonary toxicity by inducing oxidative stress. Selenium, a dietary micronutrient, has been found to protect various organs from oxidative injuries.

Objective: This study was designed to investigate the protective efficacy of an organoselenium compound 2-(5-selenocyanato-pentyl)-benzo[de]isoquinoline 1,3-dione against cyclophosphamide-induced pulmonary toxicity in Swiss albino mice.

Materials and methods: Cyclophosphamide (25?mg/kg b.w.) was administered intraperitoneally for 10?d and the organoselenium compound (3?mg/kg b.w.) was given by oral gavage in concomitant and pretreatment schedules. Various biochemical parameters related to oxidative stress and antioxidant enzymes along with histology of lungs were evaluated to assess the effect of the test compound.

Results: The oral LD₅₀ of the test compound was more than 1000?mg/kg b.w. in Swiss albino mice. The test compound substantially ameliorated cyclophosphamide-induced pulmonary injury by reducing the levels of reactive oxygen species, reactive nitrogen species, and lipid peroxidation, respectively, by 14.88, 18.54, and 21.10% in concomitant treatment schedule and by 23.89, 35.73, and 30.76% in the pretreatment schedule as well as by restoring the level of reduced glutathione and activities of glutathione-S-transferase, superoxide dismutase, catalase, and glutathione peroxidase, respectively, by 36.88, 42.43, 38.0, 35.0, and 34.06% in the concomitant treatment schedule and by 66.02, 59.29, 57.23, 71.59, and 57.22% in the pretreatment schedule. The test compound also attenuated cyclophosphamide-induced histological alterations of lung tissue.

Discussion and conclusion: The test compound emerged as an efficient antioxidant protecting lungs tissue from cyclophosphamide-induced injury.