Early embryogenesis study on a dimethoate containing formulation and Cd-sulphate in chicken embryos.

The toxic effects of a widely used organophosphate insecticide (BI 58 EC containing 38% dimethoate as active ingredient) applied alone or in combination with cadmium sulphate modelling the heavy metal load of the environment were studied on chicken embryos in the early phase of development. Solutions and emulsions of different concentrations were made from the test materials and injected in 0.1 ml volume into the air space of eggs on the first day (day 0) of incubation. Subsequently, on days 2 and 3 of incubation permanent preparations were made from the embryos in order to study the early developmental stage. Embryos fixed on slides and stained with osmium tetroxide solution were studied under light microscope. Summarising the findings, it can be established that the embryotoxicity increased after the simultaneous administration of Cd-sulphate and 38% dimethoate containing insecticide formulation compared to the control or the individually treated groups.

In vitro antioxidant activity of Anthriscus cerefolium L. (Hoffm.) extracts.

Standardised aqueous extracts of chervil (Anthriscus cerefolium L. Hoffm.) (Apiacae) were investigated for antioxidant effect. Numerous in vitro test methods were used to determine whether the extracts, from different vegetative parts (root, herb) had H-donor, metal binding, reductive, free radical scavenging and membrane protective activity. Apiin was used as a reference material. The herb extract showed better activity in all experiments than the root extract. The present results underline that the wateric chervil extracts have antioxidant and anti-lipoperoxidant activity.

One-generation reproduction toxicity study of Dithane M-45 (mancozeb) and lead acetate.

The reproductive toxicity of lead acetate and of a fungicide formulation (Dithane M-45) containing 80% mancozeb was studied on rats. Lead acetate was applied in the feed in the following dose groups: control, 1,000, 5,000 and 10,000 mg/kg of diet. The three treatment groups received, in addition to the above doses of lead acetate, 4,500 mg/kg Dithane M-45 in the diet. The method was based on the OECD Guideline for Testing of Chemicals No. 415 (1981). Clinical symptoms and mortality were not found in the parent generation. The body weight of female animals decreased significantly before the pregnancy period. This tendency was also seen in males after the combination treatment. At the two high dose levels a remarkable body weight increase was seen in the female animals during the lactation period. As a result of treatment, decreased body weight of offspring was measured during the lactation period. No gross pathological changes were seen. Histological examination showed general tubulonephrosis in the experimental animals. It can be established that the administration of Dithane M-45 did not enhance the reproductive toxicity of lead acetate.

[Investigation of the in vitro antioxidant effect of Petroselinum crispum (Mill.) Nym. ex A. W. Hill]. / A Petroselinum crispum (Mill.) Nym. ex A. W. Hill. in vitro antioxidáns hatásának vizsgálata.

Free radical reactions have excited excessive research in the past two decades. Since then it has been proven that these mechanisms may be important in the pathogenesis of certain diseases and aging. Many synthetic antioxidant components have shown toxic and/or mutagenic effects, which have directed most of the attention on the naturally occurring antioxidants. Their use has mainly centered around prevention, and the maintenance of health. Parsley, Petroselinum crispum (Mill.) Nym. ex A. W. Hill belonging to the Apiaceae family, is a well-known spice and vegetable. Its herb and root are widely known for their effects on digestion, stomach, kidney, blood, and liver. The essential oil obtained from the fruit has also strong action on the central nervous system. Characteristic constituents are: flavonoids (apiin, luteolin-, apigenin-glycosides), essential oil (apiol, miriszticin), cumarines, (bergapten, imperatorin) and vitamin C. In our experimental work, various extracts prepared from different vegetative organs of parsley have been investigated. The chemical composition of the extracts and fractions were analyzed by chromatographic (GC, HPLC) and spectroscopic (UV, UV-VIS) techniques. We intended to provide evidence for the antioxidant activity of vegetable drugs and also studied the free radical scavenger activities by means of spectrophotometry (H-donor activity, reducing capability, chelat formation) and chemiluminometry. To prove the free radical scavenging effect of the extracts, the reduced intensity of the H2O2/.OH-isoluminol, microperoxidase system was studied in vitro. The highest correlation was found between the chemical property and the antioxidant effect of the flavonoid rich samples. At the same time it was also observed that the essential oil plays a significant role in the scavenging effect as well. More experiments are in progress to study the most promising compounds in the vegetative and generative organs of parsley.

Comparing two methods of examination in the interaction study of a pesticide and a heavy metal.

The use of pesticides in field application involves the risk of poisoning wild animals. The reproduction period of pheasant takes place at the same time as the spraying time of pesticides, which justifies, that we evaluate in a point of the ecotoxicologic view the influence of the pesticide on progressive avian embryo. The most frequent technical way is injecting the exam stuffs to the some part of the embryonated eggs under the bird teratological trials. The advantage of this method is that it can be injected in a correct measured dose into the optional part of eggs. The disadvantage of this method is that it can't model properly the influence on the environment. If adverse effect of the embraced chemical substance on the embryo is experienced under the study, it will be necessary to use an immersion treatment. This procedure shows only the possible indirect influence of the pesticide on the embryo but it can suitably model its influence in plant protection practice. Treatment was done on day 12 of incubation. Applied concentration of heavy element (Cd sulphate) was 0.01% and the concentration of pesticide (Dithane M-45) was 0.2%. Evaluation was done on day 19 of incubation. Injection treatment: the simultaneous administration of Cd sulphate and the 80% mancozeb containing fungicide formulation on day 12 of incubation did not result in a significant decrease in the average body weight of embryos compared to neither the control nor the pesticide individually treated group. At the same time the body weight of embryos significantly decreased because of combined administration as compared to the Cd sulphate treated group. The embryo mortality and the incidence of developmental anomalies markedly increased after the simultaneous administration. Immersion treatment: the combined administration of Cd sulphate and the mancozeb containing fungicide formulation on day 12 of incubation did not result in a significant decrease in the average body mass of embryos compared to neither the control nor the individually treated groups. The number of embryo mortality was very high after the simultaneous administration. The incidence of developmental anomalies was sporadic.

Toxicity of a mancozeb containing formulation and Cd-sulphate to chicken embryos after administration as single compounds or in combination.

Environmental pollution of metal modelled by cadmium-sulphate and a 80% mancozeb containing fungicide formulation (Dithane M-45) were studied on chicken embryos after administration as a single compounds or in combination. The test materials were injected into the air-chamber in a volume of 0.1 ml/egg on day 0 of incubation. The concentration of cadmium-sulphate was 0.01%. The applied concentration of Dithane M-45 fungicide was 0.2%. Evaluation was done on day 19 of the hatching period. The individual administration of cadmium-sulphate and the 80% mancozeb containing fungicide formulation did not cause a significant reduction in body weight as compared to the control data. Embryonic mortality increased at all individual treated groups and reached almost a 35% rate. After the individual administration of pesticide, the number of chicken embryos with developmental anomalies did not differ markedly from the control. After the combined administration of cadmium-sulphate and the 80% mancozeb containing fungicide formulation (Dithane M-45) on day 0 of the hatching period embryonic mortality markedly increased. 88% of the treated embryos were dead. Results from the combined administration of cadmium-sulphate and an 80% mancozeb containing fungicide formulation (Dithane M-45) caused higher embryomortality with respect to individual toxicity test of cadmium-sulphate and fungicide in our study.

Embryonic toxicity of a mancozeb containing fungicide formulation and Cu-sulphate in pheasant after individual or combined administration.

Cu-sulphate and a 80% mancozeb containing fungicide formulation (Dithane M-45) were studied in pheasants after administration as single compounds or in combination. The test materials were injected directly into the air-chamber of eggs on day 12 of the hatching period and evaulation was carried out on day 23 of incubation. The pheasant embryos were examined for the following: rate of embryo mortality, body mass, type of developmental anomalies, light microscopic examination. After the administration of copper-sulphate on day 12 of incubation, the average body weight of pheasant embryos significantly did not decrease as compared to the control. The embryonic mortality was 68%. After the administration of a mancozeb containing fungicide formulation (Dithane M-45) on day 12 of incubation, the average body weight of embryos did not decrease as compared to the control. The embryonic mortality was 50%. After the individual administration of pesticide, the incidence of developmental anomalies was very high. After the combined administration of copper-sulphate and the 80% mancozeb containing fungicide formulation (Dithane M-45) on day 12 of incubation the embryonic mortality markedly increased. The rate of embryo mortality was 93%. We did not find any degenerative change in the liver tissue in either the control group or the treated groups by light microscopic examination. In summary, it can be established that the simultaneous administration of Cu-sulphate and mancozeb containing fungicide formulation caused high toxicity compared with the individual toxicity of test materials.

Teratogenicity test of dimethoate containing insecticide formulation and Cd-sulphate in chicken embryos after administration as a single compound or in combination.

Cd-sulphate and a 38% dimethoate containing insecticide formulation (BI 58 EC) were used as test material in a teratogenicity test in chicken after administration as a single compounds or in combination. The incubated chicken eggs were directly exposed to the applied test materials with injection into the air-chamber in a volume of 0.1 ml/egg before the starting of incubation. Applied concentration of Cd-sulphate was 0.01% and the concentration of pesticide was 0.1%. Evaluation was done on day 19. In test of individual toxicity after injection on day 0 of incubation Cd-sulphate did not cause a significant reduction in body mass of embryos. The rate of embryonic mortality was 26%. After the administration of dimethoate containing insecticide formulation on day 0 of incubation no decrease occurred in embryonic body mass. The rate of embryonic mortality was 31%. The developmental anomalies observed in the different treatment groups occurred sporadically. The simultaneous administration of Cd-sulphate and the dimethoate containing insecticide formulation on day 0 of incubation resulted in expressed embryonic mortality. The rate of embryonic mortality was 93%. In summary, it can be established that the simultaneous administration of Cd-sulphate and the dimethoate containing insecticide formulation on day 0 in studies of chemical interaction exerted an adverse effect on embryonic mortality.

Toxicity and degradation of metolachlor (Dual Gold 960 EC) in chicken embryos.

The herbicide formulation Dual Gold 960 EC (960 g/l metolachlor) was applied, ROSS 308 embryonated hen eggs were treated on day 12 of incubation period. The pesticide was diluted in water to a concentration level 0.3% and the emulsion was injected into the air space in a volume of 0.1 ml/egg, or hen's eggs were treated by the immersion technique (30 min). Residues of metolachlor were measured by GC in 14 collected embryo samples on days 13, 15 and 19 of the incubation of chicken embryos, and macro- and microscopic morphological examinations of 49 embryos were performed simultaneously on day 19. Body mass of embryos was weighed on 13th, 15th and 19th day of incubation. After the both treatments the mortality rate of embryos was similar. The average data of body mass showed a significant decrease compared to the control in the immersion study on day 15 and 19 of the hatching period. This phenomenon may only be in connection with the presence of metolachlor over the limit of quantification (LOQ) on day 13 of incubation period. The macroscopic deformations were sporadic in the embryos. No histologically detected alterations were seen.

Irritative effects of some pesticides and a technical component on tissue structure of the chorioallantoic membrane.

The chorioallantoic membrane (CAM) is a complete tissue that responds to injury with a complete inflammatory reaction, this process similar to that induced by chemicals in the conjunctival tissue of the rabbit eye. During the study chemicals are placed directly onto the chorioallantoic membrane and the occurrence of vascular injury or coagulation in response to a compound is as an indication of the potential of a chemical to damage mucous membranes. In our study irritant pesticides (Fusilade S, Karathane LC) and a technical pesticide component (Trend) were tested and their effects on the tissue structures of CAM were examined. After treatment with the test materials, first lysis and then haemorrhage were observed macroscopically on CAM. In histological pictures stained with H-E the rupture of the blood vessel wall was seen and blood was observed around the blood vessels in the middle layer. The histological findings correlated well with the macroscopic appearance in this study. In general a good correlation was found between the HET-CAM results and reported data from Draize test. The subjective nature of the evaluation is reduced through the histological examination of treated CAM. The HET-CAM test can be a useful component of a battery of tests needed for replacing the Draize rabbit eye irritation test.

Toxicity of a dichlorvos containing insecticide formulation and an atrazine containing herbicide formulation in chicken embryos after individual administration.

A 50% dichlorvos containing insecticide formulation (Unifosz 50 EC) and a 50% atrazine containing herbicide formulation (Hungazin PK 50 WP) were studied in chicken embryos after administration as single compounds. Applied concentrations of dichlorvos were 0.1% (corresponding to the plant protection practice), 0.05%, 0.02%, 0.01%. Applied concentrations of atrazine were 0.66% (corresponding to the plant protection practice), 0.33%, 0.132%, 0.066%. The test materials were injected directly into the air-chamber of eggs on day 0 of the hatching period and evaulation was carried out on day 19 of incubation. The chicken embryos were examined for the following: rate of embryo mortality, body mass, type of developmental anomalies. After the single administrations of dichlorvos containing insecticide formulation and atrazine containing herbicide formulation on day 0 of incubation, the average body weight of chicken embryos significantly did not decrease as compared to the control. After the individual administrations of pesticides the incidence of developmental anomalies was sporadic. The embryonic mortality markedly increased at the highest concentrations of pesticides. The rate of embrio mortality were 61% (dichlorvos insecticide containing formulation) and 52% (atrazine containing herbicide formulation). In summary, the 50% dichlorvos containing insecide formulation (Unifosz 50 EC) and the 50% atrazine containing herbicide formulation (Hungazin PK 50 WP) were toxic to the developing chicken embryos at the highest concentration in our study. The toxic effect was expressed in the high rate of embrio mortality.

Teratogenicity testing of dimethoate containing insecticide formulation (BI 58 EC) in chicken embryos.

BI 58 EC insecticide formulation was tested for teratogenicity in chicken embryos after the treatment of embryonated eggs. The pesticide was diluted in water to 0.8% concentration level, and the emulsion was injected into air space in a volume of 0.1 ml/egg or hen eggs were treated by the immersion technique. The morphological examinations were done on the days 13, 15 and 19 of incubation of chicken embryos. BI 58 EC produced an increased embryo mortality after the treatment which was the most expressive on day 15 of incubation. The trend of embryo weight showed similarity in the control and treated groups after both treatments. The developmental anomalies were sporadic and dose-effect dependency was not seen. Light microscopic findings exhibited a degenerative change in the liver tissue of both treated groups. In summary, the 38% dimethoate containing pesticide formulation (BI 58 EC) was toxic to the developing embryo at 0.8% concentration in our study.

One-generation reproduction toxicity study of mancozeb and lead acetate.

The reproduction toxicity of lead acetate and 80% mancozeb containing fungicide formulation (Dithane M-45) were studied on rats. The lead acetate was applied in diet at the following dose groups: Control-1,000-5,000-10,000 mg/kg. Three treatment and a control groups were applied, 4,500 mg/kg Dithane M-45 was administered in all the dose levels simultaneously in diet. The basis of the method was the OECD Guideline for Testing of Chemicals No. 415. Clinical symptoms and mortality were not found in the parent generation. The body weight of female animals diminished significantly before the pregnancy period. This tendency was also seen on males after the combination treatment. Remarkable body weight growth of female animals was observed during lactation period at the two high dose levels. Diminished body weight data of offsprings were measured after treatment at the end of the lactation period. The histological examination showed a general tubulonephrosis in the trial. Summing up, it can be established the administration of fungicide Dithane M-45 did not increase the toxicity of lead acetate.

Teratogenicity test of dimethoate containing insecticide formulation and heavy elements (Cu, Cd) in chicken embryos after administration as single compounds or in combination.

The teratogenic effects of heavy elements (Cu-sulphate, Cd-sulphate) and a 38% dimethoate containing insecticide formulation (Bi 58 EC) were studied on chicken after administration as a single compounds or in combination. The test materials were injected directly into the airchamber with an injector on day 12 of incubation. Applied concentrations of heavy elements (Cu-sulphate, Cd-sulphate) were: 0.01-0.001% and the concentration of pesticide was 0.1%. Final volume was 0.1 ml/egg. Evaluation was done on day 19 of incubation. After the administration of heavy elements (Cu-sulphate, Cd-sulphate) on day 12 of incubation, the average body mass of embryos significantly decreased at all the treated dose groups as compared to the control. Embryonic mortality did not increase at all the two dose levels of Cu-sulphate and at 0.001% conc. of Cd-sulphate, while the highest concentration of Cd-sulphate killed 20% of embryos treated. The number of embryos with developmental anomalies did not differ markedly from the control. The simultaneous administration of heavy elements (Cu-sulphate, Cd-sulphate) and dimethoate containing insecticide a statistically significant reduction in embryonic body mass occurred at all the treated dose groups. The incidence of developmental anomalies was markedly higher after the simultaneous use of heavy elements (Cu-sulphate, Cd-sulphate) and dimethoate containing formulation than in tests based on the separate use of heavy elements. Embryonic mortality increased at all the treated dose groups and reached almost 20% rate. In summary, it can be established that the simultaneous administration of heavy elements (Cu-sulphate, Cd-sulphate) and the dimethoate containing insecticide caused higher toxicity with respect to test of individual toxicity of heavy elements.

Free radical scavenging and membrane protective effects of methanol extracts from Anthriscus cerefolium L. (Hoffm.) and Petroselinum crispum(Mill.) nym. ex A.W. Hill.

The antilipoperoxidant activity of Anthriscus cerefolium L. (Hoffm.), chervil, Petroselinum crispum (Mill.) Nym. ex A. W. Hill., parsley extracts were evaluated with ascorbic acid induced lipid peroxidation on rat brain homogenates. These results are completed by the antiradical potential of these extracts against a solution of OH. radical. In all cases luteolin-7-O-glucoside was used as a reference material.

Embryonic toxicity of a dimethoate containing insecticide formulation and Cu-sulphate in chicken after individual or combined administration.

The aim of this study was to determine the individual and combined toxic effects of a dimethoate containing insecticide formulation and Cu-sulphate on the development of chicken embryos. The test materials were injected directly into the air-chamber in a volume of 0.1 ml/egg, or eggs were treated by immersion technique on day 12. Applied concentration of Cu-sulphate was 0.01% and the concentration of insecticidE was 0.1%. After the injection treatment of a dimethoate containing insecticide on day 12 of incubation, the average body mass of embryos significantly decreased. The simultaneous injections of Cu-sulphate and a dimethoate containing insecticide a statistically significant reduction in embryonic body mass occurred. Embryonic mortality did not increase after the individual injection of test materials, while the combined injection of Cu-sulphate and a dimethoate containing insecticide killed 30% of embryos treated. After the individual and combined immersion treatment of Cu-sulphate and a dimethoate containing insecticide, the average body mass of embryos did not decrease significantly as compared to the control. After the combined immersion treatment the incidence of embryonic mortality and the number of embryos with developmental anomalies did not differ markedly from the control. In summary, it can be established that the combined injection treatment of Cu-sulphate and a dimethoate containing insecticide caused higher embryotoxicity with respect to the test of the combined immersion treatment of test materials.

Toxicity of a mancozeb containing fungicide formulation and CU-sulphate to chicken embryos after administration as single compounds or in combination.

Environmental pollution of metal modelled by copper-sulphate and a 80% mancozeb containing fungicide formulation (Dithane M-45) were studied on chicken embryos after administration as a single compounds or in combination. The test materials were injected into the air-chamber in a volume of 0.1 ml/egg on day 12 of incubation. The concentration of copper-sulphate was 0.01%. The applied concentration of Dithane M-45 fungicide formulation was 0.2%. Evaluation was done on day 19 of the hatching period. The combined administration of copper-sulphate and the fungicide formulation did not cause a significant reduction in body weight as compared to the control data and the results from individual toxicity study of the test materials. After the combined administration of copper-sulphate and the fungicide formulation the rate of embryomortality was 40%. The incidence of developmental anomalies were sporadic. Light microscopic findings exhibited a degenerative change in the liver tissue of combined administration group. Activities of GPT and GOT enzymes increased markedly in the combined administration group. In summary, it can be established that the interaction of copper-sulphate and an 80% mancozeb containing fungicide formulation (Dithane M-45) caused higher embryomortality with respect to the test of individual toxicity of copper-sulphate and fungicide in our study.