New simple liquid chromatographic method for the determination of trimethoprim, sulfadiazine and N4-acetylsulfadiazine in plasma of broilers.

A new method for simultaneous quantification of trimethoprim, sulfadiazine and N4-acetylsulfadiazine in plasma of broilers at levels down to 13-16 ng/ml has been developed. Samples were deproteinized with acetonitrile, defatted with hexane, and extracted with dichloromethane. Chromatographic analysis was carried out on a C18 column in the presence of tetrabutylammonium hydrogen sulfate, a competing base, while detection was performed at 240 nm for trimethoprim, and 270 nm for both sulfadiazine and N4-acetylsulfadiazine. Accuracy and precision data showed recoveries and relative standard deviation values better than 87.3% and 3.1%, respectively, for all three analytes. The good analytical characteristics of the method could allow limits of detection in the low ng/ml range to be realised. The method was successfully applied to determine drug concentrations in plasma samples from broilers administered a combination of sulfadiazine and trimethoprim.

Influence of inositolhexaphosphori acid (phytic acid) on the copper distribution in tissues and the excretion of copper in rats.

The aim of the present study was to investigate the effect of the administration of phytic acid on copper (Cu) concentrations in several different rat tissues. The animals used were divided into three groups: Group A (received a diet supplemented with 2% phytic acid), group B (received a diet supplemented with 10% phytic acid) and group C (control). At the end of the experiment, the animals were sacrificed and the concentration of copper was determined in the different tissues. Phytic acid significantly increased Cu concentration in the duodenum of the animals of both groups as well as in the lungs and blood of the animals of group A. The copper concentration was also increased in the uterus and bone of the animals of group B. On the other hand, the stomach copper concentration of the animals of both groups, the heart and lung copper concentrations of the animals of group B as well as the jejunum, colon and hair copper concentrations of the animals of group A were significantly decreased. Copper excretion through feces was significantly decreased in the animals of both groups, while the excretion through urine was not significantly affected by the administration of phytic acid. In conclusion, the administration of phytic acid can produce translocation and/or elimination of copper in various tissues of rats.

Effects of cadmium chloride administration on the macroscopic and microscopic characteristics of ejaculates from Chios ram-lambs.

The effects of cadmium chloride on the volume of the ejaculate, semen density, total number of spermatozoa per ejaculate, viability, grade of motility, and morphological abnormalities were studied in 3-month-old ram-lambs of the Chios breed. Two groups of seven animals each were used. For a period of 7 months, one group was treated with a daily oral dose (3 mg/kg b.w.) of cadmium chloride and the other group received the corresponding volume of doubly distilled water. Blood samples were collected for cadmium determinations, whereas semen was collected weekly. In the cadmium-treated animals, cadmium concentration in the whole blood was increased and the testes weight was lower. The volume of the ejaculate, the semen density and the total number of spermatozoa were significantly reduced by the administration of cadmium chloride. No differences were observed in the viability, the grade motility of spermatozoa, or the percentage of dead and morphologically abnormal spermatozoa between the control and the cadmium-treated animals. Histopathological examination in the cadmium-treated animals revealed the presence of lesions in the Sertoli cells, the seminiferous tubules, the primary and the secondary spermatocytes and the spermatides, whereas in the Leydig cells no significant lesions were evident.

Influence of zinc oxide and eugenol sealer on concentration of zinc, calcium and copper in rat tissues.

A zinc oxide and eugenol root canal sealer (Roth 811) and sterile saline solution were injected into the dorsal thoracic midline of 70 male Wistar-Furth rats. Every day for the next 7 days, 10 animals were sacrificed by either inhalation. The liver, heart, kidneys and brain were removed from the animals and analysed for zinc, calcium and copper concentrations by flame atomic absorption spectrophotometry. The tissue around the injection site was also surgically removed and prepared for histological evaluation under a microscope. The injection of Roth 811 significantly affected the concentrations of zinc, calcium and copper in some of the examined organs, especially on the 4th and 5th day. The inflammatory reaction adjacent to the material was severe during the first 3 days while on the 7th day the presence of connective tissue with collagen formation was observed.

[Effect of estradiol valerate on the male reproductive organs and various semen parameters in rats]. / Uber den Einfluss von Estradiolvalerat auf die männlichen Geschlechtsorgane und verschiedene Samenparameter von Ratten.

The effect of the administration of estradiol valerate on the male reproductive organs, on their histological structure, and several semen parameters of Wistar rats was studied. In experiment A and B 140 micrograms estradiol valerate/kg b.w. were administered once a week for 4 weeks to 14 weeks old rats by s.c. injection. One week after the 4th injection the rats of experiment A were sacrificed, while the rats of experiment B lived 5 weeks without treatment for recovery. In both experiments, suppression of body weight, food consumption, decreases in absolute and relative weights of testes, epididymides, prostate and semen vesicles were observed along with testis, epididymis, seminal vesicle and prostate atrophy. The absolute and relative weights of adrenals and pituitary revealed a tendency for increase in both treated groups. The histopathological examination of the testes revealed degeneration of spermatozytes in experiment A, and degeneration of spermatozytes, spermatides, spermatozoa, Sertoli and Leydig cells in experiment B. In experiment A the motility, and number of sperms was significantly decreased, the sperm abnormalities were significantly increased. In experiment B the motility of sperms was slightly, the number significantly decreased and the abnormalities slightly increased.

Influence of 3,4-dihydroxybenzoic acid on metal ion concentrations in guinea pig tissues after copper intoxication.

3,4-Dihydroxybenzoic acid (3,4-dhbH3 or protocatechuic acid) is a copper chelator which has potential as an agent for the treatment of copper-overload disease (Wilson's disease). The present investigation describes the fluctuation in copper, magnesium, zinc and calcium (Cu, Mg, Zn, Ca) concentrations in tissues of guinea pigs intoxicated with Cu after the administration of 3,4-dhbH3. We investigated the efficacy of 3,4-dhbH3 to eliminate Cu from poisoned guinea pigs, as well as to assess the changes in concentrations of Zn, Ca and Mg which normally occur in the tissues of experimental animals. The results are in agreement with other experimental data when we administered drugs capable of forming complexes with metal ions. Although 3,4-dhbH3 is capable of forming in vitro complexes with Cu, it can not be used successfully for chelation therapy of Cu intoxication, but its effectiveness as a ligand for Ca, Zn and Mg mobilization is discussed.

[Effects of captopril on the male reproductive organs and various semen parameters of rabbits]. / Uber den Einfluss von Captopril auf die männlichen Geschlechtsorgane und verschiedene Samenparameter von Kaninchen.

The effect of the administration of captopril on the concentrations of Zn, Cu, Mg and Ca into different organs, on their histological structure and several semen parameters of male rabbits was studied. For 9 weeks 6.5 mgs captopril/kg b.w. were administered daily to 7 months old White New Zealand rabbits p.o. Semen samples were collected at the beginning of the experiment and after 4 and 9 weeks. The animals were sacrificed 9 weeks after the beginning of the experiment and organ samples were collected for histological examination and for the determination of the Zn, Cu, Mg and Ca concentrations in several tissues and the semen samples. The absolute and relative weight of the right and left testes of the test animals revealed a tendency for increase. Absolute and relative weight of the right epididymis and the relative weight of the left epididymis were significantly increased. The concentration of Zn in the blood, of Cu and Ca in the epididymis and of Mg in the testes of the test animals were significantly decreased. A significant increase was observed of the Cu and Mg concentrations in the adrenals. In the semen Cu concentration was significantly increased 9 weeks after the beginning of the experiment. Mg concentration was significantly decreased 9 weeks as compared with 4 weeks after the beginning of the experiment. Histological examination of tissue specimens of brain, liver, kidney, adrenal glands, testes, epididymis, ductus deferens and seminal vesicles from all experimental animals didn't reveal any remarkable lesion under the light microscope. The other semen parameters like volume, motility, sperm number and morphology had not changed. As the values of alcalic and acid phosphatase and ASAT in the semen samples showed many variations, statistical analysis could not be performed.

Fluctuation of zinc, copper, magnesium and calcium concentrations in guinea pig tissues after administration of captopril (SQ 14225).

The effect of the administration of captopril on Zn (zinc), Cu (copper), Ca (calcium) and Mg (magnesium) concentrations in guinea pig tissues was studied. For nine weeks 2 mg captopril per kg b.w. were administered daily to adult male guinea pigs intraperitoneally. The concentrations of the studied metals were determined in several tissues. Captopril significantly decreased Zn concentration in liver, Cu concentration in liver, adrenals, jejunum, urine and hair and Mg concentrations in blood and urine. A significant increase was observed in testicular and epididymal Zn, in heart, epididymal and fecal Cu, in Mg concentration of lung, kidney, adrenals, jejunum, epididymis and hair and in Ca concentrations in brain, heart, lung, kidney, spleen and stomach. No significant changes were observed in the colon and the thigh bone concentrations of the various elements tested. In conclusion Captopril treatment can produce translocation and/or elimination of Zn, Cu, Mg and Ca ions in various tissues of guinea pigs.

Experimental study of the biocompatibility of four root canal sealers and their influence on the zinc and calcium content of several tissues.

Four root canal sealers (AH-26, Roth 811, CRCS, and Sealapex) were tested for tissue biocompatibility in rat connective tissue. Each sealer was placed in Teflon tubes and implanted subcutaneously in Wistar-Furth rats. The implants were removed after 7, 14, and 21 days, fixed, and histologically prepared for microscopical evaluation. Brain, liver, kidneys, and uterus were removed from the animals killed at the first experimental period (7 days) and analyzed for zinc and calcium concentration by flame atomic absorption spectrophotometry. In total, 100 specimens were examined. At the seventh day, the most irritant material was seen to be AH-26, but this inflammatory reaction decreased with time. Roth 811 and Sealapex caused moderate-to-severe inflammatory reaction, whereas CRCS caused mild to moderate. CRCS and Roth 811 induced redistribution of zinc, whereas AH-26 induced changes in calcium content in some organs.

The influence of Datura ferox alkaloids on egg-laying hens.

Seeds of the weed Datura ferox are frequent contaminants of raw materials used for animal feed. In this study a mixture of scopolamine and hyoscyamine (98:2), the 2 main alkaloids of Datura ferox seeds, was incorporated at 4 total alkaloid levels (1.5, 15, 75 or 150 mg/kg feed) into a control diet fed to 100 egg-laying hens for 3 mo. Alkaloid doses of 150 mg/kg feed reduced egg production for the first 5-6 w of feeding, whereas lower doses had no effect. Egg weight, eggshell thickness and body weight of hens were unaffected at all doses. Doses of 150 mg/kg feed produced significant increases in the cardiac rate of hens after 5 w. Breathing frequency at all doses was unaffected. Determination of plasma aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and leucine aminopeptidase activities, as well as autopsy and histological examinations, revealed no obvious alkaloid-related toxic effects. It was concluded that a total alkaloid dose as high as 75 mg/kg feed can be safely administered to egg-laying hens.

Toxicity study of the main alkaloids of Datura ferox in broilers.

Seeds of the weed Datura ferox are frequent contaminants of raw materials used for animal feed. These seeds produce various toxic effects and contain mainly the alkaloids scopolamine and hyoscyamine. In this 3-month toxicity study, a mixture of scopolamine and hyoscyamine (98:2) was incorporated at four total alkaloid levels (1.5, 15, 75 or 150 mg/kg feed) into a control diet fed to 100 broilers. Alkaloid feeding caused significant reductions in the body weight gain of birds, especially of those fed a dose of 150 mg alkaloid/kg feed. Growth-retarding effects, however, were transient, as no changes in body weight gain were noted after 52 days of alkaloid feeding. Alkaloid-treated broilers showed no significant differences from controls with respect to the cardiac rate and breathing frequency nor in relation to plasma aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase activities. In contrast, plasma leucine aminopeptidase activity was significantly reduced after 3 months in all alkaloid-fed birds. Autopsy and histological examination of tissues by light and electron microscopy revealed no pathological changes associated with alkaloid feeding. Broilers appeared generally healthy and behaved normally throughout. These data should be considered in the formulation of new, improved regulations defining the maximum allowable alkaloid content of D. ferox seeds contaminating raw materials destined for use as broiler feed.

Teratogenic effects of a mixture of scopolamine and hyoscyamine in chick embryos.

The effect of a mixture of scopolamine and hyoscyamine (98:2) on developing chick embryos was investigated. The embryos were exposed at 2 stages of development to various doses of this mixture via yolk sac injection. This resulted in deaths at different stages of prenatal development, as well as various malformations and reduction deformities. These findings suggest that the studied mixture has a teratogenic effect when injected into the yolk sac of developing chick embryos.

Influence of naproxen on the zinc, copper, magnesium and calcium content of guinea pig tissues.

The effect of the administration of chronic doses of naproxen on the zinc, copper, magnesium and calcium concentrations in guinea pig tissues was studied. One hundred mg naproxen/kg body weight/week were administered intraperitoneally to adult female guinea pigs in two doses twice a week for eight weeks. A statistically significant decrease was found of the zinc content in the brain, lung, heart, liver, kidney, spleen and adrenals, of the copper content in the liver, kidney and adrenal and of the magnesium content in the spleen and heart. A significant increase was observed of the calcium content in the adrenals, heart, spleen and uterus of the treated animals, while no significant changes were observed in the thigh bone concentrations of the various ions tested.

Caffeine, aminoimidazolecarboxamide and dicoumarol, inhibitors of NAD(P)H dehydrogenase (quinone) (DT diaphorase), prevent both the cytotoxicity and DNA interstrand crosslinking produced by 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB 1954) in Walker cells.

A form of NAD(P)H dehydrogenase (quinone) (DT diaphorase, menadione reductase (NMOR), phylloquinone reductase, quinone reductase, EC 1.6.99.2) has been isolated from Walker 256 rat carcinoma cells. This enzyme can convert 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB 1954) to a cytotoxic DNA interstrand crosslinking agent by reduction of its 4-nitro group to the corresponding hydroxylamino species (Knox et al. Biochem Pharmacol, 37: 4661-4669 and 4671-4677, 1988). 2-Phenyl-5(4)-aminoimidazole-4(5)-carboxamide and AICA [5(4)-aminoimidazole-4(5)-carboxamide] have previously been reported to be antagonists of the anti-tumour effects of CB 1954. We have shown that both these compounds are inhibitors of the above enzyme and that AICA protects against both the cytotoxicity and the formation of DNA interstrand crosslinks, produced by CB 1954 in Walker cells. Similarly, known inhibitors of NAD(P)H dehydrogenase (quinone) such as dicoumarol, also reduced the cytotoxicity and DNA-interstrand crosslinking of CB 1954 in Walker cells. Caffeine was shown to be a novel inhibitor of NAD(P)H dehydrogenase (quinone) and also elicited the above protective effects. All of the above inhibitors were also shown to potentiate the toxic effects of menadione against the Walker cell. This quinone is known to be detoxified by NAD(P)H dehydrogenase (quinone) and thus emphasises the ability of these compounds to inhibit this enzyme within the cell.

Potentiation of sulphur mustard or cisplatin-induced toxicity by caffeine in Chinese hamster cells correlates with formation of DNA double-strand breaks during replication on a damaged template.

Caffeine inhibited the elongation of nascent DNA and induced breaks in the template DNA of sulphur mustard-treated Chinese hamster cells. The sizes of template and nascent DNAs, as indicated by alkaline sucrose gradient sedimentation, were similar suggesting that incision of template DNA occurred opposite gaps formed in nascent DNA by the action of caffeine, forming, effectively, double-strand breaks in DNA. Double-strand break formation was demonstrated, by means of elution of labelled DNA through polycarbonate filters at neutral pH, in both sulphur mustard- and cisplatin-treated cells when they were incubated in the presence of caffeine for 24 h. Double-strand breaks were only formed in that DNA which had been replicated in the presence of caffeine after treatment with sulphur mustard or cisplatin. Non-toxic concentrations of cycloheximide abolished the potentiation by caffeine of sulphur mustard-induced toxicity to Chinese hamster cells and at the same time abolished the formation of the low molecular weight nascent DNA, and as a consequence of its inhibitory effect on DNA synthesis, and the formation of double-strand breaks in DNA. Potentiation of the lethal and clastogenic effects of genotoxic agents by caffeine is therefore due to effects on the rate and mode of DNA synthesis which lead finally to double-strand breaks in DNA.