bluuebunny/crossref_metadata_2025 huggingface dataset:
Mutagenic activity of acetonitrile and fumaronitrile in three short term assays with special reference to autoinduction
Authors: Schlegelmilch R. , Krug A. , Wolf H. U.
Published: October 2006

Two aliphatic nitriles, acetonitrile and fumaronitrile were tested for their genotoxic potential in three mutagenicity test systems: the Salmonella/microsome‐assay, an assay using Saccharomyces cerevisiae (strain D7), and the bone marrow micronucleus test.

Both compounds were tested with and without metabolic activation in the yeast and the bacterial test systems using S9 preparations from phenobarbitone‐pretreated and autoinduced rats. Autoinduction was performed by chronic (7 days) application of a dose equivalent to a 5% oral LD50‐value of the respective compound.

With yeast strain D7 both nitriles induced low levels of gene conversion in the presence of phenobarbitone‐induced liver homogenate. An increase in the number of ile+‐revertants was not detectable under any condition.

Neither of the compounds showed mutagenic activity in the Ames test with or without metabolic activation.

A weak positive effect of acetonitrile could be detected in the micronucleus test 24 h after i.p.‐injection of the compound using a dose of 60% LD50. Fumaronitrile showed positive results with a 50% LD50 dose 48 h after administration to mice not preinduced. After 1 week of autoinduction these effects did not appear anymore, with the exception of acetonitrile 72 h after application of a dose amounting to 60% of the oral LD50‐value.
Effect of four‐week repeated inhalation exposure to unconjugated azodicarbonamide on specific and non‐specific airway sensitivity of the guinea pig
Authors: Gerlach Robert F. , Medinsky Michele A. , Hobbs Charles H. , Bice David E. , Bechtold William E. , Cheng Yung‐Sung , Gillett Nancy A. , Birnbaum Linda S. , Mauderly Joe L.
Published: October 2006

Reports of respiratory problems among industrial workers exposed repeatedly by inhalation to azodicarbonamide (ADA) raised concern that ADA might be a pulmonary sensitizer. We used a non‐invasive method for measuring specific airway conductance to evaluate the potential for repeated inhalation of unconjugated ADA to cause specific or non‐specific pulmonary sensitization in the guinea pig. Two groups of male Hartley guinea pigs were exposed 6 h/day, 5 days/week for 4 weeks to aerosolized ADA at 51 or 200 mg/m3, or to filtered air as controls. One group was tested for specific sensitization to ADA by measuring specific airway conductance during inhalation challenge with ADA before and on the third day after the 4‐week ADA exposure. The ADA concentrations for the challenges were identical to the repeated exposure concentrations (51 or 200 mg/m3, 200 mg/m3 for controls). The other group was tested for non‐specific airway sensitization by inhalation challenge with aerosolized histamine before and after the 4‐week ADA exposure. Histamine was administered in stepwise increasing concentrations to elicit an airway response in each guinea pig. Skin tests for immunological responses to ADA, body weight and histopathology of the respiratory tract and skin test sites were also evaluated. The 4‐week exposure to ADA did not result in either specific or non‐specific airway sensitization. The ADA exposure did not induce positive skin reactions, influence body weight or cause histopathological responses. These results indicate that ADA, acting alone (i.e. not conjugated to a protein), is not a pulmonary sensitizer in the guinea pig exposed repeatedly for 4 weeks and challenged to simulate a ‘Monday morning’ exposure.
Influence of particulate trap oxidizers on emission of mutagenic compounds by diesel automobiles
Authors: Rasmussen Ronald E. , Devillez Gary , Smith Lawrence R.
Published: October 2006

Diesel exhaust particles are known to contain mutagenic and carcinogenic chemicals. The aim of this study was to determine whether, and to what extent, catalytic particulate trap oxidizers on light‐duty diesel engines may reduce the emission of particle‐associated mutagenic chemicals into the environment. Exhaust particles were collected from Mercedes Benz and Volkswagen diesel automobiles, equipped with or without the manufacturer's exhaust traps, while running on a chassis dynamometer under specified load conditions. Exhaust particles were collected from a dilution tunnel onto 20″ × 20″ Teflon®‐coated fiberglass filters. Mutagenesis tests of dichloromethane (DCM) extracts of the particles were conducted using the Ames Salmonella bacterial test system. The mutation rate was calculated in terms of histidine revertants per mile of travel during a set of standard test cycles. With both vehicles the traps produced an 87–92% reduction in the total amount of particulate material collected by the filters. There was no significant change in the specific mutagenic activity (revertants per microgram of DCM particle extract) with or without the traps. These studies support the notion that installation of exhaust traps which reduce particulate emission on diesel‐powered vehicles will also reduce the emission of particle‐associated mutagenic and carcinogenic materials into the environment.
Hypertrophy and hyperplasia of the rat pancreas produced by short‐term dietary administration of soya‐derived protein and soybean trypsin inhibitor
Authors: Smith J. C. , Wilson F. D. , Allen P. V. , Berry D. L.
Published: October 2006

Feeding soy protein concentrate to weanling rats over a one‐week period produced a dose‐related increase in pancreatic weight due to an increase in acinar cell size. Hyperplastic changes occur simultaneously, as evidenced by an increase in mitotic activity after two days on the test diet. Similar changes were also obtained by feeding soybean Kunitz trypsin inhibitor over the same time period. The results suggest that this approach may be useful as a model to investigate the effect of plant‐derived material on the pancreas, in particular proliferative lesions.
Peroxidative Damage and Nephrotoxicity of Dichlorovinylcysteine in Mice
Authors: Beuter W. , Cojocel C. , Müller W. , Donaubauer H. H. , Mayer D.
Published: October 2006

Male NMRI mice were treated i.p. with dichlorovinylcysteine (DCVC) in a dosage of 2.5‐500 mg/kg−1 and renal cortical slices from naive mice were incubated with 0–125 μg/ml−1 DCVC. The effects of DCVC on blood urea nitrogen (BUN), reduced glutathione (GSH) content, malondialdehyde (MDA) production, p‐aminohippuric acid (PAH)‐ and tetraethylammoniun (TEA)‐accumulation and glucose synthesis (gluconeogenesis) were measured.

DCVC depleted GSH in a time‐ and dose‐dependent manner. Depletion of renal cortical GSH by DCVC was more pronounced in the kidney cortex than in the liver. DCVC caused a dose‐dependent increase of ethane exhalation and of MDA production in the renal cortex. When animals were kept in a closed system, decrease in oxygen concentration increased the peroxidative damage. No increase of MDA concentration was observed in the liver. Treatment of mice with DCVC induced a dose‐dependent increase in BUN and decreased the accumulation of PAH and TEA in renal cortical slices. Pretreatment of mice with aminooxyacetic acid (AOAA) and (+) cyanidanol‐3 (CY) caused a significant reduction in DCVC‐induced lipid peroxidation and nephrotoxicity. In vitro incubation of renal cortical slices of naive mice with DCVC resulted in a concentration‐dependent increase in MDA and a concentration‐dependent decrease in the accumulations of PAH, TEA and of gluconeogenesis.

In conclusion, the interaction of DCVC and/or its metabolites with membrane lipids may be responsible for lipid peroxidation and nephrotoxicity. The formation of lipid peroxidation products was greater under hypoxic conditions and appeared to be related to the DCVC‐induced nephrotoxicity. This data suggests lipid peroxidation as a possible mechanism of DCVC‐induced nephrotoxicity.
Evaluation of the teratogenic potential of the rubber accelerator N‐cyclohexyl‐2‐benzothiazylsulfenamide in rats
Authors: Ema Makoto , Murai Toshimi , Itami Takafumi , Kawasaki Hironoshin , Kanoh Seizaburo
Published: October 2006

The teratogenicity of N‐cyclohexyl‐2‐benzothiazylsulfenamide (CBS) was studied in Wistar rats. Pregnant rats were given CBS at a dosage of 0.001, 0.01, 0.1 or 0.5% in the diet from Day 0 to Day 20 of pregnancy. Daily intakes of CBS were 0.7 mg kg−1 for the 0.001% group, 7.1 mg kg−1 for the 0.01% group, 69.6 mg kg−1 for the 0.1% group and 288.8 mg kg−1 for the 0.5% group. Maternal body weight gain during pregnancy in the 0.1 and 0.5% groups was significantly lowered. Food consumption during pregnancy in the CBS‐treated groups, except for the 0.5% group, did not differ from that in the control group. Neither death nor clinical signs of toxicity were noted in the pregnant females of any group. Lowered weight in fetuses and the placentae were observed in the 0.5% group. There were no significant compound‐related effects on the incidences of pre‐ and post‐implantation losses and the number and ratio of live fetuses. Morphological examinations of the fetuses revealed no evidence of teratogenesis. It could be concluded that CBS possesses no adverse effects on the prenatal development of the offspring in rats at doses employed in the present study.
Characterization of toxicity involving haemorrhage and cardiovascular failure, caused by parenteral administration of a soluble polyacrylate in the rat
Authors: Hicks R. , Satti A. K. , Leach G. D. H. , Naylor I. L.
Published: October 2006

Adverse effects, involving a combination of hypotension and widespread haemorrhages, as well as evidence of vascuiiar and other tissue dissociation, associated with various systemic malfunctions (cyanosis, cardiac arrhythmias, neuromuscular disorder and death) were observed in the rat, from parenteral administration of a high‐molecular‐weight soluble polymer, sodium polyacrylate (EN21). Experiments to elucidate the mechanism of this effect involved comparisons with a low‐molecular‐weight sodium polyacrylate (EN5), either drug being given to groups of 5–8 rats, male or female, in doses of 5–100 mg/kg i.v., i.p. or s.c., or up to 100 mg/kg p.o. Effects were observed visually up to the death of the animals, and these plus survivors killed after 10 h were prosected for macroscopic and histopathological examination of internal organs. Characteristic EN21 effects were only observed in animals treated by the i.p. and i.v. routes, death occurring in 4–10 h or rapidly (1 h) after i.v. dosage. No such effects were observed from ENS by any dose or route. There were no differences between effects in male and female animals.

In rats anaesthetized with sodium pentobarbitone and cannulated for blood pressure recording (arterial and venous), no hypertensive effects were observed to explain haemorrhagic effects. Instead, i.v. and i.p. injections gave depressor effects, insidious in the latter case, with a precipitous fall only in the delayed terminal stage. Effects were accompanied by cardiac arrhythmias. Depressor effects were prevented by prior treatment of the animals with calcium chloride solution. These effects could not be evoked by EN5, nor by injection of methylcellulose solutions of equivalent high viscosity. It is suggested that, in the absence of pressor effects, haemorrhages must be explained by vascular fragility (angiostaxis), associated with disintegration of other tissues (e.g. intestinal villi). Other malfunctions, too rapid to be dependent on loss of blood, could be explained in terms of hypocalcaemia.

It is suggested that localization of this viscous, high‐molecular‐weight material in vascularized compartments, together with its known ability to form insoluble precipitates with calcium, leads to the operation of a calcium ‘sink’. Resulting hypocalcaemia would explain neuromuscular and cardiac malfunctions, while radical loss of tissue calcium could lead to decrease in cellular adhesion. Resultant loss of tissue integrity could be consistent with observed haemorrhage and disintegrative lesions.
Adrenal‐dependent leucopenia after short‐term exposure to various airborne irritants in rats
Authors: Brondeau M. T. , Bonnet P. , Guenier J. P. , Simon P. , de Ceaurriz J.
Published: October 2006

Leucopenia without any change in differential or red blood cell counts was observed in rats exposed for 4 h to increasing concentrations of the following airborne irritants: acetic acid, benzyl chloride, 1,2‐dichlorobenzene, ethyl acetate, ethyl acrylate, formaldehyde, isophorone, mesityl oxide, phenol, styrene, toluene diisocyanate and vinyl toluene. This effect was abolished by adrenalectomy. Vinyl toluene and 1,2‐dichlorobenzene induced leucopenia at levels as low as their current occupational standards of 50 ppm. With the exception of 1,2‐dichlorobenzene, the tested compounds caused leucopenia only when exposure reached the irritant level. It is discussed that stress associated with the irritative effect of chemicals can confound specific hacmatological effects.
Oxime‐induced reactivation of acetylcholinesterase inhibited by organophosphinates
Authors: Hanke D. W. , Beckett M. S. , Overton M. A. , Burdick C. K. , Lieske C. N.
Published: October 2006

The comparative potency of oximes for reactivation of inhibited eel acetylcholinesterase (AChE) in vitro is dependent on the organophosphinate inhibitor. Some of the data, dealing with a reference organophosphonate, support the conclusion of other investigators that the oxime potency order is also dependent on the inhibiting phosphonate. This work was done to identify more clearly the nature of phosphinylated AChE with regard to oxime reactivation potency and the potential of phosphinates as pretreatment drugs to protect AChE against organophosphonate poisoning. We have determined the reactivation potency of four oximes—2‐PAM, HI‐6, TMB‐4 and toxogonin—against four phosphinates—4‐nitrophenyl methyl(phenyl)phosphinate (PMP), 4‐nitrophenyl chloromethyl(phenyl)phosphinate (CPMP), 4‐nitrophenyl trifluoromethyl(phenyl) phosphinate and 4‐nitrophenyl bis(2‐thienyl)phosphinate. For comparison, the phosphonate sarin (GB, isopropyl methylphosphonofluoridate) was included. Incubation of the inhibited enzyme (I‐AChE) at 25°C was with 0.30 μM oxime for PMP, 3.0 μM oxime for sarin and CPMP and 100 μM oxime for the two remaining phosphinates. AChE activity was assayed spectrophotometrically for 3.0 min at 272.5 nm at 25°C in 0.10 M MOPS buffer (pH 7.60) using phenyl acetate as substrate. When sarin was the inhibitor (0% spontaneous recovery after a 2‐h incubation), the order of oxime reactivation was 2‐PAM (46%) ⩾ toxogonin (33%) = TMB‐4 (31%) > HI‐6 (9%) after 2‐h incubations. For PMP (12% spontaneous recovery after a 2‐h incubation) the oxime order was toxogonin (67%) > TMB‐4 (53%) > 2‐PAM (40%) after 2‐h incubations. For CPMP (8% spontaneous recovery after a 30‐min incubation) the order was toxogonin (65%) = TMB‐4 (59%) = HI‐6 (57%) > 2‐PAM (12%) after 30‐min incubations. For the trifluorophosphinate the order was toxogonin (18%) = TMB‐4 (17%) > HI‐6 (12%) ⩾ 2‐PAM (3%) after 5‐min incubations. For the bis‐thienylphosphinate the order was HI‐6 (38%) > 2‐PAM (20%) > TMB‐4 (0.4%) > toxogonin (0%) after 5‐min incubations. Despite some uncertainty in the oxime orders, these results support the conclusion that oxime potency is dependent on the organophosphinate used as the AChE inhibitor. The results also broaden the database that underpins the generally accepted conclusion, that for a given oxime its reactivation potency is dependent on the organophosphorous compound used to inhibit AChE.
Desferrioxamine‐induced alterations in hepatic iron distribution, DNA damage and lipid peroxidation in control and 2, 3, 7,8‐tetrachlorodibenzo‐p‐dioxin‐treated rats
Authors: Wahba Z. Z. , Murray W. J. , Stohs S. J.
Published: October 2006

Desferrioxamine (DFX), an iron chelator, has been used to inhibit in vitro redox cycling of the transition iron ions that may interfere with free‐radical formation, and is used clinically to treat conditions associated with iron overload. The effect of DFX on hepatic iron distribution has not been examined. Therefore, we have examined the effect of DFX on iron content and distribution, lipid peroxidation and DNA single‐strand breaks in liver. Since 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) enhances hepatic lipid peroxidation and induces DNA damage, the ability of DFX to modulate these effects was also examined. Female Sprague‐Dawley rats received 200 mg DFX kg−1 i.p. every 12 h for 10 days and 50 μg TCDD kg−1 p.o. on Day 4 as a single dose. All animals were killed 7 days after TCDD administration. The results demonstrate that DFX alone did not decrease hepatic iron content, but enhanced the iron distribution in mitochondria, microsomes and nuclei. A 10‐day administration of DFX resulted in enhanced hepatic lipid peroxidation and DNA single‐strand breaks, consistent with the alterations in iron distribution. TCDD administration produced large increases in lipid peroxidation in microsomes, nuclei and whole‐liver homogenates, as well as enhanced DNA damage. However, the marked increases in lipid peroxidation produced by TCDD were attenuated by DFX, while the DNA damaging effects of TCDD and DFX were additive. The results clearly demonstrate that a 10‐day administration of DFX does not result in hepatic iron depletion and the DFX‐associated increases in hepatic lipid peroxidation and DNA single‐strand breaks may reflect altered iron distribution or the formation of an iron‐catalysed nitroxide free‐radical of DFX.
The design of an experiment using statistical power with a startlechamber study as an example
Authors: Bittman Richard M. , Carniello Michael L.
Published: October 2006

Using a startle chamber experiment as a case study, it is shown how the sensitivity of a study's design can be quantified by using the concept of statistical power and how the design can be planned to achieve the power desired.

The purpose of the experiment was to compare background responses in three empty startle chambers. The study design nested groups of noise events into trials, entailing two sources of experimental error, variation within trials and between trials. For this nested design, the proper statistical analysis and calculation of power are described. It is shown how the power depends on the numbers of trials and events per trial (sampling effort), magnitudes of the sources of variability and background differences to be detected. A worked example shows the power associated with several sampling alternatives. Associated implications for cost and benefit are also discussed.
Effects of 1,4‐naphthoquinone derivatives on red blood cell metabolism
Authors: Kruger‐Zeitzer Ellen , Sullivan Stephen Gene , Stern Arnold , Munday Rex
Published: October 2006

The effect on red blood cell metabolism of a series of substituted 1,4‐naphthoquinones has been investigated. 2‐Methoxy‐1,4‐naphthoquinone was found to be a potent oxidative compound, generating hydrogen peroxide in erythrocytes and causing both methemoglobin formation and glutathione depletion in the absence of glucose. Flux of glucose through both glycolysis and the hexose monophosphate shunt was stimulated, 2‐Hydroxy‐ and 2,3‐dihydroxy‐1,4‐naphthoquinone were less oxidative. Both compounds caused oxidation of glutathione and formation of hydrogen peroxide with corresponding stimulation of the hexose monophosphate shunt, but did not cause methemoglobin formation, 2‐Hydroxy‐3‐alkyl‐1,4‐naphthoquinones were not oxidative but did increase the flux of glucose through glycolysis, possiMy reflecting membranal damage. The in vitro oxidative effects of these substances do not correlate with their hemolytic activity in rats, indicating that factors other than oxidative damage are important in mediating the in vivo toxicity of these substances.
Acute toxicity induced by 2‐aryl‐N‐methylsuccinimides
Authors: Rankin Gary O. , Shih Hsien‐Cheng , Teets Vonda J. , Nicoll Derek W. , Brown Patrick I.
Published: October 2006

Phensuximide (PSX) is a 2‐arylsuccinimide useful in the treatment of absence seizures. PSX is a mild urotoxicant and is structurally related to N‐phenylsuccinimide (NPS) and its antifungal derivatives. Since substitution of the phenyl ring of NPS with chloro or tert‐butyl groups can produce compounds with enhanced nephrotoxic potential, it was felt that similar substitutions on the phenyl ring of PSX also might produce derivatives with enhanced nephrotoxic potential. Three derivatives of PSX were prepared and tested: 2‐(3‐chlorophenyl)‐N‐methylsuccinimide (CPMS); 2‐(4‐tert‐butylphenyl)‐N‐methylsuccinimide (BPMS) and 2‐(3,5‐dichlorophenyl)‐N‐methylsuccinimide (DPMS). In one set of experiments, male Fischer 344 rats were administered a single intraperitoneal (i.p.) injection of a succinimide (0.4 or 1.0 mmol kg−1) or vehicle (sesame oil, 2.5 ml kg−1) and renal function monitored at 24 and 48 h. Only minor changes in renal function were noted with the PSX derivatives. BPMS and DPMS (1.0 mmol kg−1) treatment induced mild renal tubular necrosis and thickening of the glomerular membranes. However, no significant morphological changes were noted in ureters, bladder or liver in any treatment group. In a second set of experiments, rats were pretreated with phenobarbital (75 mg kg−1 day−1, i.p., 3 days) followed by a single i.p. injection of DPMS (0.4 or 1.0 mmol kg−1) or DPMS vehicle. Renal function was monitored as before. Phenobarbital pretreatment did not markedly enhance the functional nephrotoxicity induced by DPMS (0.4 mmol), but tubular necrosis was greater than observed in non‐phenobarbital‐pretreated rats receiving DPMS (1.0 mmol kg−1). In addition, hepatotoxicity was observed as the appearance of numerous non‐staining vacuoles in hypertrophied hepatocytes. In the phenobarbital plus DPMS (1.0 mmol kg−1) treatment group, all rats died by 48 h. Prior to death, rats exhibited increased proteinuria (+3), hematuria (+3) and blood urea nitrogen concentration. At 24 h, kidneys from rats treated with phenobarbital plus DPMS (1.0 mmol kg−1) exhibited extensive proximal tubular necrosis and numerous glomeruli with thickened membranes. Hepatotoxicity was more pronounced than with phenobarbital plus DPMS (0.4 mmol kg−1) at 48 h and urinary bladders had focal areas of erythrocytes pooling below the epithelial lining. These results demonstrate that although NPS and PSX are structural analogs, chemical substitutions that enhance the nephrotoxic potential of NPS do not have a similar effect on PSX. In addition, DPMS can induce urotoxicity in a manner similar to that observed for PSX and probably induces toxicity via one or more metabolites.
Mutagenicity and genotoxicity of ethylvinyl ketone in bacterial tests
Authors: Deininger Christoph , Eder Erwin , Neudecker Tilmann , Hoffman Christian
Published: October 2006

The mutagenic and genotoxic effects of ethylvinyl ketone were investigated. This α, β‐unsaturated carbonyl compound is widely distributed in the environment, in particular in food. Whereas ethylvinyl ketone shows only weak genotoxicity in the SOS Chromotest with Escherichia coli PQ37, it was distinctly mutagenic per se in the Salmonella preincubation assay with TA100. Using SKF 525 (an inhibitor of microsomal monooxygenase) and trichloropropene oxide (an inhibitor of epoxide hydrolase) we found indication for additional activation via epoxidation by S9 mix. The need for further investigation of the genotoxic, mutagenic and carcinogenic effects of this compound is strongly indicated.
Effects of in ovo injection of carbamates on chick embryo hatchability, esterase enzyme activity and locomotion of chicks
Authors: Farage‐Elawar Miranda
Published: October 2006

Carbaryl and aldicarb, two carbamate pesticides used extensively throughout the United States, are known to act as acetylcholinesterase inhibitors. We have demonstrated previously that exposure to carbaryl and aldicarb in young chicks caused persistent locomotion alterations with no correlation to esterase inhibition. In this study, we investigated the effects of these carbamates when injected in ovo to chick embryos, at two time periods (days 5 and 15) during incubation. Carbaryl dosed at 45 mg kg−1 egg weight was extremely toxic to the embryos on day 5 of incubation. Hatchability was reduced to 0% as compared to 80% when carbaryl was injected on day 15 of incubation. Aldicarb at 1.5 mg kg−1 egg weight had no major effect on hatchability when injected either on day 5 or day 15 of incubation (hatchability = 90 and 100%, respectively). Plasma, liver and brain esterases were measured in the chick at different time points during incubation and after hatching. Brain acetylcholinesterase (AChE) and liver cholinesterase (ChE) were inhibited significantly during incubation in embryos dosed on day 15 with both carbaryl and aldicarb. Liver carboxylesterase was inhibited significantly during incubation with only the carbaryl treatment. All esterase enzyme activities returned to normal after hatching. Plasma ChE and carboxylesterase levels were not affected with either carbaryl or aldicarb treatment from 8 until 47 days after hatching. Neither carbamate had any effect on brain neuropathy target esterase (NTE) activity either during incubation or after hatching. The locomotion of chicks was affected in both treatment groups until 47 days after hatching. This study indicates that carbaryl and aldicarb may cause long‐term delayed alterations in the chicks.
Acute effects of the antiepileptic succinimides on the urinary tract and potentiation of phensuximide‐induced urotoxicity by phenobarbital
Authors: Rankin Gary O. , Teets Vonda J. , Nicoll Derek W. , Brown P. I.
Published: October 2006

Phensuximide (PSX), methsuximide (MSX) and ethosuximide (ESX) are succinimide antiepileptic agents used worldwide in the treatment of absence seizures. A previous study from our laboratory demonstrated that PSX (0.3 or 0.6 mmol kg−1 day−1, i.p.) induced urotoxicity following daily administration for 5–7 days in Fischer 344 rats, but PSX (1.25 mmol kg−1, i.p.) induced only minimal urotoxicity following acute administration. The purpose of this study was to determine the acute nephrotoxic potential of MSX and ESX in male Fischer 344 rats and if antiepileptic succinimide‐induced urotoxicity is potentiated by phenobarbital pretreatment. In one set of experiments, rats (four rats per group) were administered a single intraperitoneal (i.p.) injection of a succinimide (0.4 or 1.0 mmol kg−1) or vehicle (sesame oil, 2.5 ml kg−1), and the renal function was monitored at 24 and 48 h. Neither ESX or MSX induced substantial changes in renal function or morphology, which suggests that neither compound is acutely nephrotoxic. Similar results were obtained with PSX, which supported our earlier findings with this antiepileptic agent. In a second set of experiments, rats (four rats per group) were pretreated for 3 days with phenobarbital (75 mg kg−1 day−1, i.p.) prior to receiving a succinimide (0.4 or 1.0 mmol kg−1, i.p.) or vehicle (sesame oil, 2.5 ml kg−1, i.p.). Renal function was monitored at 24 and 48 h after the last injection. Phenobarbital pretreatment had only minor effects on ESX‐ or MSX‐induced renal effects, with no significant morphological changes detected between treated and pair‐fed control groups. However, PSX‐induced urotoxicity was potentiated by phenobarbital. Marked hematuria and proteinuria were observed following PSX (1.0 mmol kg−1) administration to phenobarbital‐pretreated rats. Bladders from these animals contained bloody urine and exhibited hemorrhagic areas among the bladder epithelial cells. Proximal tubular cells exhibited mild damage, with some swelling and blebbing at the brush border membrane. These results suggest that phenobarbital potentiates PSX‐induced urotoxicity and that the resulting hemorrhagic cystitis is mediated by one or more PSX metabolites.
Assessment of renal function and damage in animal species. A review of the current approach of the academic, governmental and industrial institutions represented by the animal clinical chemistry association
Authors: Stonard M. D.
Published: October 2006

There are a wide variety of laboratory tests available to assess damage to and functional impairment of the kidneys, although the effectiveness of these tests varies greatly depending upon the site specificity of the damage and to a lesser extent upon the animal species involved.

Several traditional tests of renal dysfunction and damage, including plasma creatinine and urea, and urinalysis (dipstick and/or quantitative protein), can be used in the first instance to detect nephrotoxicity. A second tier of specific, targetted indicators (concentration test, urinary enzymes, clearance of analytes, specific proteins, etc.) may then be applied to identify further the site of the lesion and the functional status of the kidneys.

The glomerular filtration rate (GFR) may be estimated from the clearance of exogenous and endogenous substances. The difficulty in obtaining accurately timed urine samples limits the value of these tests in small animals, although methods that do not involve urine collection are available.

The kidney is the origin of several enzymes found in urine that can be used to monitor the toxic effects of chemicals and therapeutic substances. Selective measurement of enzyme activities in urine can be used to detect the site of the renal lesion after traditional tests have established the presence of renal injury. Separation of proteins in urine by electrophoretic techniques may also be used to discriminate damage to different parts of the nephron.

Renal cell excretion in urine is a sensitive but unreliable indicator of acute damage to the proximal tubule. The rate of cell excretion is not a good predictor of the severity of tubular injury.

The tests available, and their use in toxicological investigations, have been reviewed in the light of current practice in the UK.
The role of membrane composition in ATPase activities of cirrhotic rat liver: Effect of silymarin
Authors: Muriel Pablo , Mourelle Marisabel
Published: October 2006

The activities of Ca2+‐ and Na+, K+ ‐ATPases were studied in liver plasma membranes from CCI4‐cirrhotic rats and from livers of rats treated with silymarin in addition to CCI4. CCI4 chronic treatment produced significant decreases in Na+, K+‐ and Ca2+‐AT Pase activities; however, the animals treated with silymarin along with CCI4 showed no differences in ATPase activities as compared to controls. The lipid analysis performed in plasma membranes revealed increases in the cholesterol/phospholipid (CH/PL) and sphingomyelin/phosphatidylcholine (SM/PC) ratios in the cirrhotic group. Again, the membranes isolated from rats receiving CCI4 + silymarin showed normal CH/PL and SM/PC values. Considering that CH/PL and SM/PC ratios are related to membrane microviscosity, this study suggests that a lower fluidity of the membrane may be responsible for the observed decreases in ATPase activities in the cirrhotic group. Additionally, the role of silymarin to improve liver function in CCI4‐cirrhosis can be attributed partially to its action at membrane level by preventing the increases in CH/PL and SM/PC ratios.
Role of glutathione in vanadate reduction in young and mature rats: Evidence for direct participation of glutathione in vanadate inactivation
Authors: Kretzschmar M. , Bräunlich H.
Published: October 2006

The influence of renal glutathione content, modulated by the glutathione synthesis inhibitor buthionine sulphoximine or by glutathione infusion, on the polyuric and natriuretic effects of i.v. administered vanadate was investigated in 20‐ and 5‐day‐old rats. The modulation of renal glutathione content led to significant changes in urine volume and sodium excretion, independently of age.

A decrease in the renal glutathione level led to intensification and prolongation of the diuretic effects of vanadate in 55‐day‐old animals. Treatment with glutathione abolished and treatment with buthionine sulphoximine increased the polyuric effect of vanadate. These observations indicate a role for renal glutathione in vanadate inactivation. Age‐dependent differences in the polyuric and natriuretic effectiveness of vanadate are caused by differences in renal glutathione content during maturation of the kidney.
The role of iron in t‐butyl hydroperoxide‐induced lipid peroxidation and hepatotoxicity in rats
Authors: Younes Maged , Wess Alexander
Published: October 2006

Treatment of rats with 100 mg kg−1 t‐butyl hydroperoxide led to an enhanced ethane exhalation as a marker of in vivo lipid peroxidation, as well as a moderate hepatoxicity as evidenced by a rise in plasma activities of liver‐specific enzymes (glutamate‐pyruvate transaminase and sorbitol dehydrogenase) and an increase in hepatic calcium content. Furthermore, a depletion of hepatic glutathione by 17% was observed. Apart from the loss of glutathione, all these effects were antagonized by pretreatment of rats with the potent iron chelator deferrioxamine and potentiated by pretreatment with low concentrations of FeSO4 having no pro‐oxidant activity per se; this was also the case in rats under conditions of iron overload (experimental haemochromatosis). These data indicate a close relationship between t‐butyl hydroperoxide‐induced lipid peroxidation and its hepatotoxicity, and point out the importance of iron in catalysing reinitiation (propagation) reactions of lipid peroxidation in vivo.
Embryotoxicity and teratogenicity study in rats dosed epicutaneously with dimethylformamide (DMF)
Authors: Hansen Ernst , Meyer Otto
Published: October 2006

The reproductive effect of N,N‐dimethylformamide (DMF) administered epicutaneously to rats was examined. The rats were dosed on gestation days 6–15 or on gestation days 1–20 at dose levels of up to 2 ml DMF kg−1 body weight. Body weight, weight gain and pregnancy rate were reduced in those rats receiving 2 ml DMF kg−1 body weight per day on days 6–15. A reduction in the number of live fetuses and in fetal weight, as well as an increase in postimplantation loss, were also observed at this dose level. Similar but more pronounced effects were observed in rats dosed on days 1–20 with 2 ml DMF kg−1 body weight. The lowest effect level was 1 ml DMF kg−1 body weight. No other dose‐related effects were found in this study.
Evaluation of the teratogenic potential of the plasticizer butyl benzyl phthalate in rats
Authors: Ema Makoto , Murai Toshimi , Itami Takafumi , Kawasaki Hironoshin
Published: October 2006

The teratogenicity of butyl benzyl phthalate (BBP) was studied in Wistar rats. Pregnant rats were given BBP at a dosage of 0, 0.25, 0.5, 1.0 or 2.0% in the diet from day 0 to day 20 of pregnancy. Daily intakes of BBP were 185 mg kg−1 for the 0.25% group, 375 mg kg−1 for the 0.5% group, 654 mg kg−1 for the 1.0% group and 974 mg kg−1 for the 2.0% group. Adjusted maternal body weight gain (body weight gain excluding the gravid uterus) during pregnancy in the 1.0 and 2.0% groups was significantly lowered. Food consumption during pregnancy in the 0.25 and 0.5% groups did not differ from that in the control group. No death was noted in the pregnant females of any group. There was no significant compóund‐related effects on the incidence of prcimplantation loss. All dams given 2.0% BBP exhibited complete resorption of all the implanted embryos. Morphological examinations of the fetuses revealed no evidence of teratogenesis. It could be concluded that the no‐observable‐effect‐levels (NOEL) in rats were 0.5 and 1.0% BBP in the diet for maternal and embryofetal toxicity, respectively.
No evidence of toxicity or carcinogenicity of pentaerythritol tetranitrate given in the diet to F344 rats and B6C3F1 mice for up to two years
Authors: Bucher John R. , Huff James , Haseman Joseph K. , Eustis Scot L. , Lilja Herman S. , Murthy A. S. K.
Published: October 2006

Toxicology and carcinogenesis studies of pentaerythritol tetranitrate (PETN), an organic nitrate used in explosives and as a therapeutic agent for angina pectoris, were conducted by administering diets containing PETN,NF (National Formulary Grade, a 1:4 mixture of PETN and lactose) to both sexes of F344 rats and B6C3F1 mice in 14‐day, 13‐week and 2‐year studies. PETN was found to be essentially non‐toxic in 14‐day and 13‐week studies at dietary concentrations as high as 10 000 ppm; the weight gain of female rats was lower than that of controls at 5000 and 10 000 ppm in the 13‐week study. In the 13‐week studies, one in ten high‐dose female rats had an adenoma of the Zymbal gland and one in ten high‐dose female mice had a hepatocellular adenoma. Dietary concentrations chosen forthe 2‐year studies were 5000 and 10 000 ppm for male rats and male and female mice, and 1240 and 2500 ppm for female rats. Inthe 2‐year studies, there were no adverse effects on survival or body weight gains in either sex of rats or mice. No neoplastic or non‐neoplastic lesions were considered to be related clearly to PETN administration. Neoplasms of the Zymbal gland occurred at low incidences in PETN‐exposed groups of both sexes of rats inthe 2‐year study.
Effect of calcium channel blocking agents on the reductive metabolism of halothane
Authors: Kawahara Michio , Akita Susumu , Fujii Kohyu , Morio Michio
Published: October 2006

The effect of calcium channel blocking agents on the reductive metabolism of halothane in liver microsomes of guinea pigs was investigated. The reaction mixture for the measurement of the end products consisted of microsomal suspension, 5 mM NADPH, calcium channel blocking agents (verapamil, diltiazem, nicardipine and nifedipine) and halothane in 0.1 M phosphate buffer (pH 7.4). The reductive metabolism of halothane was inhibited competitively by verapamil, diltiazem and nicardipine. The binding spectra for the interaction of these three drugs with cytochrome P‐450 in microsomes were investigated. Verapamil caused the reverse type I difference spectrum and diltiazem caused the type I difference spectrum. However, the change caused by nicardipine was not observed by the presence of its specific spectra. NADPH–cytochrome P‐450 reductase activity in microsomes did not change by the addition of these three drugs. These results suggest that these three calcium channel blocking agents inhibit the production of radical intermediates during the reductive metabolism of halothane.
A comparison between the isovolume and the end‐inflation occlusion methods for measurement of lung mechanics in rats
Authors: Cardoso Wellington Veras , Saldiva Paulo Hilário Nascimento , Martins Patricia , Criado Pereira , Sakae Regina Silvia , Böhm György Miklós , Zin Walter Araújo
Published: October 2006

The isovolume method is one of the most used techniques to assess the mechanical properties of the lungs in ordinary challenge tests and in toxicological studies. It has been considered that isovolume lung resistance (RL) could assess central airway calibre in such tests, whereas dynamic elastance (EL,dyn) would reflect the elastic properties of lung parenchyma. However, peripheral phenomena, such as Pendelluft and stress relaxation, can interfere with RL measurements, especially in disease. In order to investigate the contribution of such phenomena to isovolume RL and EL,dyn, we compared the results provided by isovolume technique with those given by the end‐inflation occlusion method (EIOM) in 12 normal Wistar rats. We found similar values for total lung resistance (RL,max) and isovolume RL. Homogeneous resistance (RL,min), which describes airway calibre, comprised about 30% of RL in the tidal volume range. Values of EL,dyn calculated by the EIOM were approximately 20% greater than the corresponding value measured during spontaneous breathing. Our data indicate that isovolume RL is significantly affected by tissue forces and cannot be interpreted as a pure index of airway size.
Haemolytic activity and nephrotoxicity of 2‐hydroxy‐1,4‐naphthoquinone in rats
Authors: Munday R. , Smith B. L. , Fowke E. A.
Published: October 2006

The short‐term toxicity of 2‐hydroxy‐1,4‐naphthoquinone (lawsone) and 2‐methyl‐1,4‐naphthoquinone (menadione) has been compared in rats.

2‐Methyl‐1,4‐naphthoquinone has been shown previously to cause haemolytic anaemia in animals, and this was confirmed in the present experiment. 2‐Hydroxy‐1,4‐naphthoquinone was found also to cause haemolysis, in a dose‐dependent manner, as reflected by decreased blood packed cell volumes and haemoglobin levels and by histopathological changes in spleen, liver and kidney. With both naphthoquinones, the haemolysis was of the oxidative type, characterized by the presence of Heinz bodies within erythrocytes.

Haemolysis was the only toxic change identified in rats dosed with 2‐methyl‐1,4‐naphthoquinone. In contrast, 2‐hydroxy‐1,4‐naphthoquinone was not only a haemolytic agent but also a nephrotoxin, causing renal enlargement, elevated plasma levels of urea and creatinine and histologically‐identified tubular necrosis, largely confined to the distal segment of the proximal convoluted tubules.

The relationship between the in vivo toxic effects of these naphthoquinones and previously‐reported data on their in vitro cytotoxic action is discussed.
Gene mutation assay of acrolein in the CHO/HGPRT test system
Authors: Parent Richard A. , Caravello Halina E. , Harbell John W.
Published: October 2006

The mutagenic potential of acrolein has been studied with a wide range of in vitro and in vivo genetic toxicity assays. The data often have been conflicting, especially with the Ames assay. This study was undertaken to assess the mutagenic potential of acrolein using the CHO/HGPRT assay, both with and without metabolic activation. This assay system was chosen because it provides eukaryotic DNA as the target and is capable of detecting a range of mutational events. Because of its considerable toxicity, acrolein was tested over a very narrow dose range of 0.2–2 nl ml−1 without exogenous activation and 0.5–8 nl ml−1 with rat S‐9 activation. Multiple assays were performed under both conditions. The results indicated that while acrolein was clearly very cytotoxic, it did not induce a significant mutagenic response in the presence or absence of metabolic activation.
Globin adducts of benzo[a]pyrene: Markers of inhalation exposure as measured in F344/N rats
Authors: Bechtold W. E. , Sun J. D. , Wolff R. K. , Griffith W. C. , Kilmer J. W. , Bond J. A.
Published: October 2006

We have explored methods for determing benzo[a]pyrene (BaP) dosimetry by measuring adduction levels to F344/N rat blood hemoglobin, and have refined and validated an assay that measures the in vivo binding of the 7, 8‐diol 9, 10‐epoxide metabolite of BaP to globin. The assay for BaP‐globin adducts was based on the release of tetrahydroxy‐BaP (BaP‐tetrols) from globin by mild acid hydrolysis. After extensive isolation, BaP‐tetrols were quantitated by high‐pressure liquid chromatography using fluorescence detection. BaP‐tetrol levels were measured in rats dosed intraperitoneally with 242, 71 and 24 μmol BaP kg−1 body weight in corn oil. The formation of BaP‐tetrols was not linear with dose. The lowest dose yielded adduct levels that represented the limits of sensitivity for the method, as performed in this laboratory. Once this limit of sensitivity was established, the potential use of the assay was assessed by measuring the radiochemical binding of inhaled [14C]BaP or its metabolites to the globin of F344/N rats. Rats were exposed for 4 h per day, 1 day per week, for 12 weeks to pure aerosols of [14C]BaP at a level of 2 mg m−3. At the conclusion of exposure, rats were sacrificed and globin was isolated. The extent of [14C]BaP binding to the globin was determined by liquid scintillation spectrometry. Rats exposed to aerosols of [14C]BaP had statistically increased levels of binding to globin, and the levels were comparable to those observed previously after intragastric administration. Based on the spectroscopic assay data, the sensitivity of the proposed analytical method was projected to be insufficient to measure BaP‐tetrol levels after inhalation exposures similar to those described above. The use of the globin‐BaP adduct approach may be limited by the small fraction of BaP‐tetrols that can be released by acid hydrolysis.
A CCl4/CHCl3 interaction study in isolated hepatocytes: Non‐induced and phenobarbitalpretreated cells
Authors: O'Hara Todd M. , Sheppard Melissa A. , Clarke Elizabeth C. , Borzelleca Joseph F. , Gennings Chris , Condie Lyman W.
Published: October 2006

The purpose of this study was to evaluate an isolated hepatocyte model for predicting the in vivo hepatotoxicity of carbon tetrachloride (CCl4) and chloroform (CHCl3), alone and in combination. Response surface methodology (RSM) was used to analyze and describe the data. The interaction was evaluated for % initial K+ (cell injury) and % LDH leakage (cell death) in non‐induced (untreated) and phenobarbital‐pretreated suspended hepatocytes. CCl4 and CHCl3 were delivered alone and in combination in dimethyl sulfoxide (DMSO) to suspended hepatocytes. The maximum observed no‐effect level (MONEL) for CCl4 in non‐induced cells was 1.0 mM (LDH and K+). In induced cells, the MONEL was 0.25 mM (K+) and 0.5 mM (LDH). The MONEL for CHCl3 in non‐induced cells was 5.0 mM (LDH and K+) and in induced cells was 0.5 mM (K+) and 1.0 mM (LDH). Phenobarbital pretreatment enhanced the toxicity of both CCl4 and CHCl3, alone and in combination. RSM analysis of the % initial K+ and % LDH for CCl4 and CHCl3 in combination in non‐induced and induced cells showed a greater than additive interaction. The isolated hepatocyte model appears to be a promising system for evaluating the toxicity of chemical mixtures and predicting their in vivo effects.
Chinese hamster ovary cell assays for mutation and chromosome damage: Data from non‐carcinogens
Authors: Oshiro Y. , Piper C. E. , Balwierz P. S. , Soelter S. G.
Published: October 2006

In vitro genotoxicity tests are employed to screen chemicals for their capability to cause various DNA and chromosomal alterations, and the results are often used to predict their potential for carcinogenicity. However, there is controversy regarding the apparent low specificity of some in vitro genotoxicity assays, which result in a high false positive rate. Since we use and rely upon in vitro assays for risk assessment and prediction of carcinogenicity, this specificity issue is of serious concern to us. Hence, we selected ten compounds deemed non‐carcinogens in the literature to test for the induction of gene mutation and chromosomal damage using the Chinese hamster ovary cell/hypoxanthine–guanine phosphoribosyl transferase (CHO/HGPRT) mutation assay performed concurrently with a CHO micronucleus assay. The chemical exposures for the two end‐points were done simultaneously. The protocol for the two end‐points was developed using the carcinogens N‐methyl‐N'‐nitro‐N‐nitrosoguanidine, 3‐methylcholanthrene, cyclophosphamide and 7, 12‐dimethylbenzanthracene. The non‐carcinogens chosen were 4‐nitro‐o‐phenylenediamine, p‐phenylenediamine dihydrochloride, 3‐nitropropionic acid, dichlorvos, 2‐(chloromethyl)pyridine, N‐(1‐naphthyl)ethylenediamine 2HCl, O‐anthranilic acid, 4‐nitroanthraniic acid, anilazine and triphenyltin hydroxide. Each of these chemicals had been reported positive in the Ames test and/or the mouse lymphoma TK+/‐ mutation assay. In addition, eight of them were also reported positive in in vitro assays for chromosome aberrations and/or sister chromatid exchange (SCE). We found four of the ten chemicals negative for gene mutation and micronucleus induction without and with activation in the CHO/HGPRT mutation and CHO micronucleus assays. However, one of these four chemicals may be a potential carcinogen according to other carcinogenicity reviewers. Four other chemicals that induced only micronuclei were negative for gene mutation. Dichlorvos was positive for gene mutation and micronucleus induction without and with activation. This chemical has been shown recently to cause various tumors in rodents. One of the non‐carcinogens was positive in the micronucleus test and equivocally positive in the mutation test. These results indicate that the CHO/HGPRT mutation assay may provide more relevant results than the CHO micronucleus assay, the mouse lymphoma mutation assay, or in vitro SCE and chromosome aberration assays when screening chemicals for potential carcinogenicity.