Genetic Variations in the Promoter of the APE1 Gene Are Associated with DMF-Induced Abnormal Liver Function: A Case-Control Study in a Chinese Population.

Acute or long-term exposure to N,N-dimethylformamide (DMF) can induce abnormal liver function. It is well known that DMF is mainly metabolized in the liver and thereby produces reactive oxygen species (ROS). The base excision repair (BER) pathway is regarded as a very important pathway involved in repairing ROS-induced DNA damage. Several studies have explored the associations between GSTM1, GSTT1, CYP2E1 polymorphisms and DMF-induced abnormal liver function; however, little is known about how common hOGG1, XRCC1 and APE1 polymorphisms and DMF induce abnormal liver function. The purpose of this study was to investigate whether the polymorphisms in the hOGG1 (rs159153 and rs2072668), XRCC1 (rs25487, rs25489, and rs1799782), APE1 (rs1130409 and 1760944) genes in the human BER pathway were associated with the susceptibility to DMF-induced abnormal liver function in a Chinese population. These polymorphisms were genotyped in 123 workers with DMF-induced abnormal liver function and 123 workers with normal liver function. We found that workers with the APE1 rs1760944 TG/GG genotypes had a reduced risk of abnormal liver function, which was more pronounced in the subgroups that were exposed to DMF for <10 years, exposed to ≥10 mg/m³ DMF, never smoked and never drank. In summary, our study supported the hypothesis that the APE1 rs1760944 T > G polymorphism may be associated with DMF-induced abnormal liver function in the Chinese Han population.

Metabolite profiles of rats in repeated dose toxicological studies after oral and inhalative exposure.

The MetaMap(®)-Tox database contains plasma-metabolome and toxicity data of rats obtained from oral administration of 550 reference compounds following a standardized adapted OECD 407 protocol. Here, metabolic profiles for aniline (A), chloroform (CL), ethylbenzene (EB), 2-methoxyethanol (ME), N,N-dimethylformamide (DMF) and tetrahydrofurane (THF), dosed inhalatively for six hours/day, five days a week for 4 weeks were compared to oral dosing performed daily for 4 weeks. To investigate if the oral and inhalative metabolome would be comparable statistical analyses were performed. Best correlations for metabolome changes via both routes of exposure were observed for toxicants that induced profound metabolome changes. e.g. CL and ME. Liver and testes were correctly identified as target organs. In contrast, route of exposure dependent differences in metabolic profiles were noted for low profile strength e.g. female rats dosed inhalatively with A or THF. Taken together, the current investigations demonstrate that plasma metabolome changes are generally comparable for systemic effects after oral and inhalation exposure. Differences may result from kinetics and first pass effects. For compounds inducing only weak changes, the differences between both routes of exposure are visible in the metabolome.

Transdermal delivery of lercanidipine hydrochloride: effect of chemical enhancers and ultrasound.

The effects of permeation enhancers and sonophoresis on the transdermal permeation of lercanidipine hydrochloride (LRDP) across mouse skin were investigated. Parameters including drug solubility, partition coefficient, drug degradation and drug permeation in skin were determined. Tween-20, dimethyl formamide, propylene glycol, poly ethylene glycol (5% v/v) and different concentration of ethanol were used for permeation enhancement. Low frequency ultrasound was also applied in the presence and absence of permeation enhancers to assess its effect on augmenting the permeation of drug. All the permeation enhancers, except propylene glycol, increased the transdermal permeation of LRDP. Sonophoresis significantly increased the cumulative amount of LRDP permeating through the skin in comparison to passive diffusion. A synergistic effect was noted when sonophoresis was applied in presence of permeation enhancers. The results suggest that the formulation of LRDP with an appropriate penetration enhancer may be useful in the development of a therapeutic system to deliver LRDP across the skin for a prolonged period (i.e., 24 h). The application of ultrasound in association with permeation enhancers could further serve as non-oral and non-invasive drug delivery modality for the immediate therapeutic effect.

Anti-nociceptive activity and toxicity evaluation of Cu(II)-fenoprofenate complexes in mice.

The proposed curative properties of copper(II)-non-steroidal anti-inflammatory drugs (NSAIDs) have led to the development of numerous copper(II)-NSAID complexes with enhanced anti-inflammatory activity. In this work, the antinociceptive and toxic effects of two new coordination complexes: Cu2(fen)4(caf)2 [fen: fenoprofenate anion; caf: caffeine] and Cu2(fen)4(dmf)2 [dmf: N-N'-dimethylformamide] were evaluated in mice. The antinociceptive effect was evaluated with two models: acetic acid-induced writhing response and formalin test. For the sub-acute exposure, the complexes were added to the diet at different doses for 28days. Behavioral and functional nervous system parameters in a functional observational battery were assessed. Also, hematological, biochemical and histopathological studies were performed. Cu2(fen)4(caf)2 and Cu2(fen)4(dmf)2 significantly decreased the acetic acid-induced writhing response and the licking time on the late phase in the formalin test with respect to the control and fenoprofen salt groups. The sub-acute exposure to Cu2(fen)4(caf)2 complex increased the motor activity, the number of rearings and the arousal with respect to the control and fenoprofen salt groups. These impaired parameters in mice exposed to Cu2(fen)4(caf)2 can be attributable to the presence of caffeine as stimulating agent. On the other hand, all exposed groups decreased the urine pools in the functional observational battery and increased the plasmatic urea. These effects could be due to the decrease in the glomerular filtration caused by NSAIDs. In conclusion, both complexes Cu2(fen)4(dmf)2 and Cu2(fen)4(caf)2 were more potent antinociceptive agents than fenoprofen salt. Sub-acute exposure to different doses of these complexes did not produce significant changes in the parameters that evaluate toxicity.

Dimethylformamide as a cryoprotectant for canine semen diluted and frozen in ACP-106C.

The objective was to assess the effect of adding various concentrations of dimethylformamide on characteristics of canine semen diluted in powdered coconut water (ACP-106C; ACP Biotecnologia, Fortaleza, CE, Brazil) and frozen at -196°C. Fifteen ejaculates were collected by manual stimulation from five adult Boxer dogs. The sperm-rich fraction was diluted in ACP-106C (ACP Biotecnologia) containing 10% egg yolk and divided into four aliquots. The cryoprotectants used for each aliquot were 6% glycerol (control group; CG) or 2%, 4%, or 6% dimethylformamide (DF2, DF4, and DF6, respectively). After thawing, total motility (mean ± SEM) for CG (58.4 ± 24.6) was higher (P < 0.05) than that of the other groups (2% dimethylformamide, 24.4 ± 12.3; 4% dimethylformamide, 26.5 ± 16.1; and 6% dimethylformamide, 21.7 ± 17.9). Furthermore, there was a greater percentage of fast, average, and slow moving sperm (assessed with computer-aided semen analysis; CASA) in CG in comparison with the other three groups. Therefore, based on concentrations tested in this study, dimethylformamide, together with ACP-106C (ACP Biotecnologia) and 10% egg yolk as a diluent, yielded unsatisfactory in vitro results for freezing canine semen.

Differences in ability of jennies and mares to conceive with cooled and frozen semen containing glycerol or not.

A suitable method for the cryopreservation of donkey semen would be very valuable for the ex situ management of genetic diversity in this species. This report uses a variety of observation and trials to evaluate the effect of cryoprotectants in per-cycle pregnancy rates (PC) in equids females (jennies (donkey) and mares (horse)). This was explored by (1) comparing the results of insemination of jennies and mares with cooled or frozen donkey semen, (2) examining the possible toxic effect of the cryoprotectant (CPA) glycerol in these two species and (3) studying alternative solutions. Donkey and horse semen was either used immediately, or cooled according to some steps of the pre-freezing procedure or frozen and thawed. The pre-freezing procedure included semen dilution, centrifugation, resuspension in milk or in INRA82+2% egg yolk+various % CPA (expressed as final concentrations in extended semen (v/v)) and then cooling to 4 degrees C. PC was similar in mares and jennies inseminated with donkey semen cooled to 4 degrees C in milk. However, the PC was significantly higher in mares than in jennies when donkey semen was frozen with 2.2% glycerol (36%, n=50 cycles vs. 11%, n=38 cycles; P<0.01). Increasing the concentrations of glycerol (0, 2.2, 3.5, 4.8%) before cooling stallion semen resulted in a progressive decrease in mare PC (87, 53, 53, 13% (n=15 cycles for each concentration); P<0.0001). The addition of 2.2% glycerol before cooling donkey semen decreased the PC measured in jennies to 0. The replacement of glycerol by 2% dimethylformamide increased the fertility obtained in jennies with cooled donkey semen (PC: 67%, n=12 cycles) but did not increase the fertility obtained with frozen-thawed donkey semen (PC: 11%, n=28 cycles with dimethylformamide vs. 0%, n=16 cycles with glycerol). In conclusion, this study clearly shows that the ability of jennies to conceive after AI with donkey frozen semen is lower than that of mares. Glycerol affects the fertility of donkey and stallion spermatozoa as early as during the pre-freezing procedure. In consequence, the glycerol level must be low in frozen equine semen to provide good fertility. The toxic dose of glycerol for donkey spermatozoa seems to be almost half that for stallion spermatozoa. Whether this greater sensitivity of donkey spermatozoa to glycerol is responsible for the low success of semen cryopreservation in jennies is not so obvious because replacement of glycerol by dimethylformamide was not much more effective in terms of fertility.

Enhanced proliferative response of hepatocytes to combined inhalation and oral exposures to N,N-dimethylformamide in male rats.

Male Wistar rats were exposed by inhalation to N,N-dimethylformamide (DMF) at 0 (control), 200 or 400 ppm (v/v) for 6 hr/day, 5 days/week and 4 weeks, and each inhalation group received DMF-formulated drinking water at 0, 800, 1,600 or 3,200 ppm (w/w) for 24 hr/day, 7 days/week and 4 weeks. Both the combined inhalation and oral exposures and the single-route exposure through inhalation or ingestion induced centrilobular hypertrophy and single-cell necrosis of hepatocytes, increased plasma levels of alanine aminotransferase (ALT), increased percentage of proliferating cell nuclear antigen (PCNA)-positive hepatocytes without glutathione-S-transferase placental form (GST-P)-positive liver foci, and increased relative liver weight. Those hepatic parameters of the DMF-induced effects were classified into hypertrophic, necrotic and proliferative responses according to the pathological characteristics of affected liver. While magnitudes of the hypertrophic and necrotic responses were linearly increased with an increase in amounts of DMF uptake in the single-route exposure groups, those dose-response relationships tended to level off in the combined-exposure groups. Saturation of the hypertrophic and necrotic responses at high dose levels might be attributed to suppression of the metabolic conversion of DMF to its toxic metabolites. Percentage of PCNA-stained hepatocytes classified as the proliferative response was increased more steeply in the combined-exposure groups than in the single-route exposure groups. It was suggested that the proliferative response of hepatocytes to the combined exposures would be greater than that which would be expected under an assumption of additivity for the component proliferative responses to the single-route exposures through inhalation and ingestion.

Potential mechanisms by which a single drink of alcohol can increase transdermal absorption of topically applied chemicals.

BACKGROUND: Both chronic and acute ethanol consumption increase transdermal penetration of topically applied xenobiotics. The mechanisms by which this enhancement occurs are unknown. We hypothesized that either the vasodilatory effects of ethanol or its ability to disrupt the lipid bilayer via lipid peroxidation, may be contributing to the increased transdermal absorption observed in alcohol consuming animals. METHODS: Male Wistar rats were gavaged with 1.5, 3, 4.3, 6 or 10 g/kg ethanol or saline control or were treated with either the vasoconstrictor epinephrine or with the vasodilator prilocaine. Dermal blood flow, transepidermal water loss (TEWL), and skin moisture were non-invasively measured. Transdermal penetration was then determined for four xenobiotics (paraquat, dimethyl formamide (DMF), 2,4-dichlorophenoxyacetic acid (2,4-D) and N,N-diethyl-m-toluamide (DEET)). Lipid peroxidation was also determined by monitoring the formation of malondialdehyde. RESULTS: Dermal blood flow increased by approximately 27% (p<0.05), TEWL increased 1.12+/-0.2-fold while skin lipid peroxidation increased 1.4-fold (p<0.05) 2h after gavage with 10 g/kg alcohol. Transdermal penetration of paraquat was increased by prilocaine (ER=2.1+/-0.4, p<0.05), but the absorption of DEET, 2,4-D and DMF were not influenced by greater blood flow. Reducing dermal blood flow with epinephrine did not cause any significant changes in transdermal penetration. CONCLUSIONS: Vasodilation triggered by a single episode of ethanol ingestion is not responsible for the observed increase in transdermal absorption. Ethanol induced changes in lipid peroxidation and TEWL demonstrate that drinking alcohol induces transdermal absorption of xenobiotics.

Evaluation of the effectiveness of personal protective equipment against occupational exposure to N,N-dimethylformamide.

The objectives of this study were to evaluate the protective effectiveness of various personal protective equipment and the respective exposure contributions from respiratory and skin exposures of N,N-dimethylformamide (DMF) with a self-comparison study design. Two high-, four intermediate- and four low-DMF exposure workers from a synthetic leather factory were monitored in airborne DMF concentrations and N-methylformamide (NMF) concentrations in urine across four consecutive days. The workers were designated to wear no personal protective equipment on the first day. The barrier cream, rubber gloves and rubber gloves plus respirator were used on the second, third and fourth days, respectively. Person-to-personal observation was performed in the field to record all high and low exposure tasks during work for each subject. Protective effectiveness index (PEI) was used to evaluate different glove effectiveness. We concluded that the direct skin contact to the strong skin penetrates like DMF could be a more significant exposure source than the respiratory exposure in the actual occupational environment. The provision of protective equipment from skin exposure could be more important than that from respiratory exposure. The application of barrier cream could be as effective as wearing impermeable rubber gloves in the prevention from the skin penetrate in the occupational settings.

Carcinogenicity and chronic toxicity after inhalation exposure of rats and mice to N,N-dimethylformamide.

Carcinogenicity and chronic toxicity of N,N-Dimethylformamide (DMF) were examined by inhalation exposure of groups of 50 rats and 50 mice of both sexes to DMF vapor at a concentration of 0, 200, 400 or 800 ppm (v/v) for 6 h/d, 5 d/wk, for 104 wk. In rats, incidences of hepatocellular adenomas and carcinomas significantly increased in the 400 and 800 ppm-exposed groups and in the 800 ppm-exposed group, respectively. The hepatocellular adenoma did not increase significantly in the 400 ppm-exposed female rats, but its incidence exceeded a range of historical control data in the Japan Bioassay Research Center (JBRC). In mice, incidences of hepatocellular adenomas and carcinomas significantly increased in all the DMF-exposed groups. Incidence of hepatoblastomas significantly increased in the 200 and 400 ppm-exposed male mice, and 4 cases of hepatoblastomas in the 400 ppm-exposed female mice and the 800 ppm-exposed male mice exceeded the range of historical control data of the JBRC. Incidences of altered cell foci increased in the liver of exposed rats and mice in an exposure concentration-related manner, and those foci were causally related to the hepatocellular tumors. Liver weights increased in both rats and mice exposed to DMF at 200 ppm and above. Increased levels of gamma-GTP, ALT, AST and total bilirubin in exposed rats of both sexes and AST and ALT in exposed mice of both sexes were noted. It was concluded that 2-yr inhalation exposure to DMF increased incidences of hepatocellular adenomas and carcinomas in rats and incidences of hepatocellular adenomas, carcinomas and hepatoblastomas in mice, and that hepatocarcinogenicity of DMF was more potent in mice than in rats.

Thirteen-week inhalation toxicity of N,N-dimethylformamide in F344/N rats and B6C3F1 mice.

Male and female F-344 rats and B6C3F1 mice (10/sex/group) were exposed to N,N-dimethylformamide (DMF) by whole body inhalation exposure at 0, 50, 100, 200, 400, or 800 ppm, 6 h/day, 5 days/week, for 13 weeks. A concentration-dependent depression in body weight occurred in rats of both sexes at 400 (6-11%) and 800 ppm (20-22%). In contrast, all weight changes in both sexes of mice were within 10% of controls. No rats died, while 5 mice died from nonexposure-related causes. Relative liver weights were significantly increased at all DMF concentrations in both sexes and both species. Activities of serum sorbitol dehydrogenase (SDH) were statistically increased in male and female rats (200 to 800 ppm) on study days 4, 24, and 91 (13 weeks). Activities of alanine aminotransferase (ALT) and isocitrate dehydrogenase (ICD) were statistically increased in both sexes of rats exposed to 800 ppm DMF at all time points. Cholesterol (CHOL) levels were statistically increased in male and female rats (50-800 ppm) at all sampling time points. Levels of total bile acids (TBA) were statistically increased in both sexes of rats (400-800 ppm) on days 24 and 91. Centrilobular hepatocellular necrosis (minimal to moderate) was seen in rats of both sexes exposed at 400 and 800 ppm, with the lesions more severe in females. Centrilobular hepatocellular hypertrophy (minimal to mild) was found in all groups of DMF-exposed male mice, and in female mice exposed at 100-800 ppm. For male and female rats the no-observed-adverse-effect concentration (NOAEC) for microscopic liver injury was 200 ppm. The NOAEC was 50 ppm for female mice, but an NOAEC based upon the absence of microscopic liver injury was not determined in male mice.

Enhancer aided in vitro permeation of atenolol and prazosin hydrochloride through mice skin.

Effect of penetration enhancers were studied on the permeation of antihypertensive drugs prazosin hydrochloride and atenolol through full thickness skin of swiss albino mice. Atenolol was delivered to skin from saturated alcoholic solution containing 5% of 1-decanol and alcohol alone, while prazosin hydrochloride was saturated in dimethyl formamide(DMF, 5% v/v in water) and dimethyl sulfoxide(DMSO, 5% v/v in water). Atenolol permeation was augmented significantly in decanolic solution and also in pure alcohol. In case of prazosin hydrochloride, significant enhancement of permeation was shown by DMSO but not by DMF.

Does the polymorphism of cytochrome P-450 2E1 affect the metabolism of N,N-dimethylformamide? Comparison of the half-lives of urinary N-methylformamide.

The aim of this study was to clarify whether phenotypic variation exists when subjects with different genotypes of cytochrome P450 2E1 (CYP2E1) are exposed to N,N-dimethylformamide (DMF). The genotypes of CYP2E1 were confirmed in 123 healthy male volunteer subjects. Of the 123 subjects, the numbers of c1 homozygotes, c2 heterozygotes, and c2 homozygotes were 77, 45, and 1, respectively. Seven of the c1 homozygotes, five of the c2 heterozygotes, and the one c2 homozygote (mean age: 22.7 years, range: 20-27 years) were exposed to DMF vapor twice, once via the skin and once via the lung, for a total of 8 h per subject at a concentration below 10 ppm, the occupational exposure limit recommended by the Japan Society for Occupational Health, the American Conference of Governmental and Industrial Hygienists, and Deutsche Forschungsgemeinschaft, at 27 degrees C and 44% relative humidity. Exposure levels were 6.2+/-1.0 ppm in dermal exposure and 7.1+/-1.0 ppm in inhalation exposure. Urine samples were collected until 72 h after exposure. The half-lives of urinary N-methylformamide (NMF) were obtained as the phenotype. The average urinary NMF half-lives of the c1 homozygotes, the c2 heterozygotes, and the c2 homozygote were 3.86+/-1.90, 4.38+/-1.53, and 4.2 h after dermal exposure, and 1.58+/-0.42, 1.84+/-0.61, and 3.2 h after respiratory exposure. The NMF half-lives of the c1 homozygotes were not significantly different from those of the c2 heterozygotes, and there were no differences between the NMF half-lives on the subjects with and without the c2 allele. Even though the data were obtained from only one c2 homozygote, it is noteworthy that the NMF half-life of this subject was slightly less than that of the c1 homozygotes after respiratory exposure.

Characterization of the antinociceptive and sedative effect of amitraz in horses.

Amitraz, an acaricide used to control ectoparasites in animals has a complex pharmacological activity, including alpha2-adrenergic agonist action. The purpose of this research was to investigate the possible antinociceptive and/or sedative effect of amitraz in horses. The sedative effect of the intravenous (i.v.) injection of dimethylformamide (DMF, 5 mL, control) or amitraz (0.05, 0.10, 0.15 mg/kg), was investigated on the head ptosis test. The participation of alpha2-adrenergic receptors in the sedative effect provoked by amitraz was studied by dosing yohimbine (0.12 mg/kg, i.v.). To measure the antinociception, xylazine hydrochloride (1 mg/kg, i.v., positive control) and the same doses of amitraz and DMF were used. A focused radiant light/heat directed onto the fetlock and withers of a horse were used as a noxious stimulus to measure the hoof withdrawal reflex latency (HWRL) and the skin twitch reflex latency (STRL). The three doses of amitraz used (0.05, 0.10 and 0.15 mg/kg) provoked a dose-dependent relaxation of the cervical muscles. The experiments with amitraz and xylazine on the HWRL showed that after i.v. administration of all doses of amitraz there was a significant increase of HWRL up to 150 min after the injections. Additionally, there was a significant difference between control (DMF) and positive control (xylazine) values up to 30 min after drug injection. On the other hand, the experiments on the STRL show that after administration of amitraz at the dose of 0.15 mg/kg, a significant increase in STRL was observed when compared with the control group. This effect lasted up to 120 min after injection. However, no significant antinociceptive effect was observed with the 0.05 and 0.10 mg/kg doses of amitraz or at the 1.0 mg/kg dose of xylazine.

Assessment of the developmental toxicity, metabolism, and placental transfer of N,N-dimethylformamide administered to pregnant rats.

This study evaluates the developmental toxicity and placental and milk transfer of N,N-dimethylformamide (DMF) in rats. Sprague-Dawley rats were given 0, 50, 100, 200, and 300 mg DMF/kg/day, by gavage, on Gestational Days (GD) 6 through 20. Maternal toxicity was indicated by depressions in weight gain and food consumption at doses >/=100 mg/kg. Fetal toxicity was indicated by decreased fetal body weight at doses >/=100 mg/kg, and by increased incidences of two skeletal variations (absent or poorly ossified supraoccipital and sternebrae) at 200 and 300 mg/kg. Thus, the maternal and developmental no-observed-adverse-effect level was 50 mg/kg/day. The time course disposition of [14C]DMF was examined over a 48-hr period in GD12- and GD18-pregnant rats after a single oral dose of 100 mg [14C]DMF/kg. Peak concentrations of radiocarbon occurred within 1 hr after dosing. Embryonic (GD12) and fetal (GD18) tissues accounted for 0.15 and 6% of the administered dose, respectively. Levels of radiocarbon in embryonic and fetal tissues were equal or slightly less than in maternal plasma up to 8 and 24 hr, respectively, and higher thereafter. HPLC analysis performed at intervals from 1 to 8 hr on GD12 and 1-24 hr on GD18 indicated that unchanged DMF and metabolites were readily transferred to the embryonic and fetal tissues, where their levels were generally equal to those in maternal plasma. The parent compound accounted for most of the radioactivity until 4-8 hr and then decreased. N-Hydroxymethyl-N-methylformamide (HMMF) and N-methylformamide (NMF) were the predominent metabolites and increased with time. Much lower concentrations were found for formamide and N-acetyl-S-(N-methylcarbamoyl)cysteine. Transfer of radioactivity into milk was studied in dams given a single oral administration of 100 mg [14C]DMF on Lactation Day 14. DMF, HMMF, and NMF were found in the milk at concentrations equal to those in plasma.

Hepatotoxicity of N, N-dimethylformamide in rats following intraperitoneal or inhalation routes of administration.

Male Sprague-Dawley rats were administered a single intraperitoneal injection of N, N-dimethylformamide (DMF, 0.01-1.5 g kg-1) or were exposed for 4 h to DMF vapours (75-900 ppm). The serum activities of the enzymes sorbitol deshydrogenase and glutamate deshydrogenase were used as indicators of liver damage, and were determined at 24, 48 or 72 h post-treatment. Following either route of administration DMF caused concentration-dependent elevations in enzyme activities, the maxima of which occurred later after administration of higher concentrations of DMF than after lower concentrations.