Persistence and efficacy of a new formulation based on dog allomonal repellents against Rhipicephalus sanguineus sensu lato tick / Persistência e eficácia de uma nova formulação baseada em repelentes alomônios de cães contra o carrapato Rhipicephalus sanguineus sensu lato

Abstract Beagles are less susceptible to Rhipicephalus sanguineus sensu lato tick due to the production of the allomones benzaldehyde and 2-hexanone. Our previous published work showed that these compounds can reduce tick burden on susceptible dogs. Here we tested the hypothesis that an increase in repellent dose and release rate could increase repellent efficacy and persistence. Slow-release formulations of these compounds, with higher doses and release rates, were tested on artificially-infested dogs. Ten dogs were randomly assigned to two groups with five dogs each. The treated group received collars with slow-release formulations of the compounds attached, while the control group received collars with clean formulations attached. Five environmental infestations were performed, with the number of ticks (at all stages) on the dogs being counted once a day for 40 days. No significant increase in repellent efficacy was observed with the higher doses and release rates, whereas a greater persistence in repellent activity was observed. Treatment with the formulations resulted in a two-to-three-fold reduction in the number of immature stage ticks for up to three weeks. However, the number of adults was similar in both groups. Loss of repellent activity after the third week of testing coincided with a marked change in the relative release rates for the two compounds. It is hypothesized that relative amounts, rather than absolute amounts, of repellent release from slow-release formulations are important for repellent activity. We also hypothesize that the avoidance of less-preferred hosts by ticks relies on olfactory-mediated perception of specific blends of volatile cues from less preferred hosts.

Resumo Beagles são menos suscetíveis ao carrapato Rhipicephalus sanguineus sensu lato devido à produção de benzaldeído e 2-hexanona. Nosso trabalho anterior já publicado mostrou que esses compostos podem reduzir a carga de carrapatos em cães suscetíveis. Aqui testamos a hipótese de que um aumento na dose destes repelentes e na taxa de liberação poderia aumentar a eficácia e a persistência do efeito repelente. As formulações de liberação lenta destes compostos, com doses e taxas de liberação mais elevadas foram testadas em cães infestados artificialmente. Dez cães foram distribuídos aleatoriamente em dois grupos com cinco cães cada. O grupo tratado recebeu colares contendo formulações de liberação lenta dos compostos, enquanto o grupo controle recebeu colares com formulações limpas. Cinco infestações ambientais foram realizadas, com o número de carrapatos (em todas as fases) nos cães sendo contados, uma vez ao dia, por 40 dias. Não se observou aumento significativo na eficácia do repelente com doses e taxas de liberação mais elevadas e, enquanto observou-se maior persistência na atividade repelente. O tratamento com as formulações resultou em uma redução de duas a três vezes no número de carrapatos dos estágios imaturos, por até três semanas. No entanto, o número de adultos foi semelhante em ambos os grupos. A perda de atividade repelente após a terceira semana de teste coincidiu com uma mudança nas taxas de liberação relativa para os dois compostos. A hipótese é que as quantidades relativas, ao invés das quantidades absolutas de liberação lenta, são importantes para a atividade repelente. Então, a hipótese é de que a repelência apresentada por hospedeiros menos susceptíveis aos carrapatos depende da percepção pelos carrapatos de misturas específicas de voláteis liberados por estes hospedeiros.

Persistence and efficacy of a new formulation based on dog allomonal repellents against Rhipicephalus sanguineus sensu lato tick.

Beagles are less susceptible to Rhipicephalus sanguineus sensu lato tick due to the production of the allomones benzaldehyde and 2-hexanone. Our previous published work showed that these compounds can reduce tick burden on susceptible dogs. Here we tested the hypothesis that an increase in repellent dose and release rate could increase repellent efficacy and persistence. Slow-release formulations of these compounds, with higher doses and release rates, were tested on artificially-infested dogs. Ten dogs were randomly assigned to two groups with five dogs each. The treated group received collars with slow-release formulations of the compounds attached, while the control group received collars with clean formulations attached. Five environmental infestations were performed, with the number of ticks (at all stages) on the dogs being counted once a day for 40 days. No significant increase in repellent efficacy was observed with the higher doses and release rates, whereas a greater persistence in repellent activity was observed. Treatment with the formulations resulted in a two-to-three-fold reduction in the number of immature stage ticks for up to three weeks. However, the number of adults was similar in both groups. Loss of repellent activity after the third week of testing coincided with a marked change in the relative release rates for the two compounds. It is hypothesized that relative amounts, rather than absolute amounts, of repellent release from slow-release formulations are important for repellent activity. We also hypothesize that the avoidance of less-preferred hosts by ticks relies on olfactory-mediated perception of specific blends of volatile cues from less preferred hosts.

Spontaneous calcium transients in the immature adult-born neurons of the olfactory bulb.

Spontaneous neuronal activity and concomitant intracellular Ca2+ signaling are abundant during early perinatal development and are well known for their key role in neuronal proliferation, migration, differentiation and wiring. However, much less is known about the in vivo patterns of spontaneous Ca2+ signaling in immature adult-born cells. Here, by using two-photon Ca2+ imaging, we analyzed spontaneous in vivo Ca2+ signaling in adult-born juxtaglomerular cells of the mouse olfactory bulb over the time period of 5 weeks, from the day of their arrival in the glomerular layer till their stable integration into the preexisting neural network. We show that spontaneous Ca2+ transients are ubiquitously present in adult-born cells right after their arrival, require activation of voltage-gated Na+ channels and are little sensitive to isoflurane anesthesia. Interestingly, several parameters of this spontaneous activity, such as the area under the curve, the time spent in the active state as well as the fraction of continuously active cells show a bell-shaped dependence on cell's age, all peaking in 3-4 weeks old cells. This data firmly document the in vivo presence of spontaneous Ca2+ signaling during the layer-specific maturation of adult-born neurons in the olfactory bulb and motivate further analyses of the functional role(s) of this activity.

Brown dog tick, Rhipicephalus sanguineus sensu lato, infestation of susceptible dog hosts is reduced by slow release of semiochemicals from a less susceptible host.

Domestic dog breeds are hosts for the brown dog tick, Rhipicephalus sanguineus sensu lato, but infestation levels vary among breeds. Beagles are less susceptible to tick infestations than English cocker spaniels due to enhanced production of 2-hexanone and benzaldehyde that act as volatile tick repellents. We report the use of prototype slow-release formulations of these compounds to reduce the burden of R. sanguineus s. l. on English cocker spaniel dogs. Twelve dogs were randomly assigned to two groups with six dogs each. The treated group received collars with slow-release formulations of the compounds attached, while the control group received collars with clean formulations attached. Five environmental infestations were performed, with the number of ticks (at all stages) on the dogs being counted twice a day for 45days. The counts on the number of tick stages found per dog were individually fitted to linear mixed effects models with repeated measures and normal distribution for errors. The mean tick infestation in the treated group was significantly lower than in the control group. For larvae and nymphs, a decrease in tick infestation was observed at the fifth count, and for adults, lower average counts were observed in all counts. The compounds did not interfere with the distribution of the ticks on the body of the dogs, as a similar percentage of ticks was found on the anterior half of the dogs (54.5% for the control group and 56.2% for the treated group). The biological and reproductive parameters of the ticks were not affected by the repellents. This study highlights for the first time the potential use of a novel allomone (repellent)-based formulation for reduction of tick infestation on susceptible dogs.

Methyl isobutyl ketone-induced hepatocellular carcinogenesis in B6C3F1 mice: A constitutive androstane receptor (CAR)-mediated mode of action.

In a National Toxicology Program (NTP) chronic inhalation study with methyl isobutyl ketone (MIBK), increases in hepatocellular adenomas and hepatocellular adenomas and carcinomas (combined) were observed in male and female B6C3F1 mice at 1800 ppm. A DNA reactive Mode-of-Action (MOA) for this liver tumor response is not supported by the evidence as MIBK and its major metabolites lack genotoxicity in both in vitro and in vivo studies. Constitutive androstane receptor (CAR) nuclear receptor-mediated activation has been hypothesized as the MOA for MIBK-induced mouse liver tumorigenesis. To further investigate the MOA for MIBK-induced murine liver tumors, male and female B6C3F1, C57BL/6, and CAR/PXR Knockout (KO) mice were exposed to either 0 or 1800 ppm MIBK for 6 h/day, 5 days/week for a total of 10 days. On day 1, mice were implanted with osmotic mini-pumps containing 5-Bromo-2-deoxyuridine (BrdU) 1 h following exposure and humanely euthanized 1-3 h following the final exposure. B6C3F1 and C57BL/6 mice had statistically significant increases in liver weights compared to controls that corresponded with hepatocellular hypertrophy and increased mitotic figures. Hepatocellular proliferation data indicated induction of S-phase DNA synthesis in B6C3F1 and C57BL/6 mice exposed to 1800 ppm MIBK compared to control, and no increase was observed in MIBK exposed CAR/PXR KO mice. Liver gene expression changes indicated a maximally-induced Cyp2b10 (CAR-associated) transcript and a slight increase in Cyp3a11(PXR-associated) transcript in B6C3F1 and C57BL/6 mice exposed to 1800 ppm MIBK compared to controls, but not in Cyp1a1 (AhR-associated) or Cyp4a10 (PPAR-α-associated) transcripts. CAR/PXR KO mice exposed to 1800 ppm MIBK showed no evidence of activation of AhR, CAR, PXR or PPAR-α nuclear receptors via their associated transcripts. MIBK induced hepatic effects are consistent with a phenobarbital-like MOA where the initiating events are activation of the CAR and PXR nuclear receptors and resultant hepatocellular proliferation leading to rodent liver tumors.

Methyl isobutyl ketone (MIBK) induction of alpha2u-globulin nephropathy in male, but not female rats.

Male F-344 rats were administered corn oil (vehicle control), d-limonene (positive control, 300mg/kg), or MIBK (1000mg/kg) and female F-344 rats corn oil (vehicle control) or MIBK for 10 consecutive days by oral gavage. Approximately 24h after the final dose the kidneys were excised and the left kidney prepared and evaluated for histological changes including protein (hyaline) droplet accumulation, immunohistochemical staining for alpha2u-globulin (alpha2u), and proliferating cell nuclear antigen (PCNA) to quantitate renal cell proliferation. The right kidney was prepared for quantitation of total protein and alpha2u using an ELISA. MIBK elicited an increase in protein droplets, accumulation of alpha2u, and renal cell proliferation in male, but not female rats, responses characteristic of alpha2u-mediated nephropathy. MIBK produced identical histopathological changes in the male rat kidney when compared to d-limonene, an acknowledged inducer of alpha2u-nephropathy except that the grade of severity tended to be slightly lower with MIBK. MIBK did not induce any effects in female rats. Therefore, renal histopathology, along with the other measures of alpha2u accumulation, provides additional weight of evidence to support the inclusion of MIBK in the category of chemicals exerting renal effects through a alpha2u-nephropathy-mediated mode-of-action.

Simulation of repeated dose kinetics of methyl isobutyl ketone in humans from experimental single-dose inhalation exposure.

Methyl isobutyl ketone (MIBK) is a solvent used in numerous products and processes and may be present in the air of the workplace as a vapor. The American Conference of Governmental Industrial Hygienists (ACGIH) threshold limit value-time-weighted average (TLV-TWA) and TLV-short term exposure limit (TLV-STEL) for MIBK are 50 and 75 ppm, respectively. These workplace air concentration limits were set to protect workers from irritation, neurasthenic symptoms and possible adverse effects to their livers and kidneys. A recent revision of the ACGIH limit value has been proposed, to reduce the current TLV-TWA to 30 ppm. This article predicts the kinetics and accumulation of MIBK in humans exposed repeatedly in various exposure scenarios (8, 12, and 24h/day for 7 days) to the current ACGIH TLV-TWA of 50 ppm. The kinetic parameters of the model were derived from published human time-course blood MIBK data from a single 2h inhalation exposure to 48.9 ppm MIBK. The model correctly simulated single exposure experimental data with a rapid rise in blood concentration to 1.06 microg/ml within 1h and approached >or=99% steady-state blood level in 4h of exposure. MIBK was predicted to be rapidly eliminated from blood after terminating the exposure, reaching 0.53 microg/ml and 0.13 microg/ml within 0.5 and 2h post-exposure, respectively. Within 4h after the termination of exposure, blood concentration would be expected to <1% of the steady-state concentration. On the basis of these results, it is concluded that accumulation of MIBK in workers due to repeated inhalation exposure is not likely to occur at the current TLV-TWA concentration of 50 ppm.

Inhalation two-generation reproductive toxicity study of methyl isobutyl ketone in rats.

To evaluate whether methyl isobutyl ketone (MIBK) affects reproductive performance, a two-generation reproduction study was conducted. MIBK was administered to 30 Sprague-Dawley rats/sex/group via whole-body inhalation at concentrations of 0, 500, 1000, or 2000 ppm, 6 h daily, for 70 days prior to mating. F(0) and F(1) females were exposed from mating through gestation day 20 and from postnatal day 5; F(2) litters were maintained through postnatal day 21. No treatment-related mortality of adult animals occurred. There was a dose-related increase in adult animals with no or a decreased response to a sound stimulus at 1000 and 2000 ppm; however, no adverse clinical signs occurred 1 h after exposure, suggesting this was a transient sedative effect. Clinical signs of central nervous system (CNS) depression in the pups were observed and one F(1) pup died after initial exposure to 2000 ppm on postnatal day 22; subsequently exposure was delayed until postnatal day 28. Decreased body weight gain and slight decreased food consumption were observed during the first 2 weeks of exposure in both generations at 2000 ppm. There were no adverse effects on male and female reproductive function or landmarks of sexual maturation. Increased F(0) and F(1) liver weights with associated centrilobular hypertrophy occurred in rats at 2000 ppm, indicative of an adaptive response. Increased male kidney weights at all exposure concentrations, associated with hyaline droplets, were indicative of male rat-specific nephropathy. Other than acute sedative effects, the no-observed-adverse-effect level (NOAEL) for parental systemic effects (excluding male rat kidney) was 1000 ppm, based on transient decreased body weight and food consumption; for reproductive effects, 2000 ppm, the highest concentration tested; and for neonatal toxicity, 1000 ppm (based on acute CNS depressive effects).

Comparative acute toxicity of o-chlorobenzylidene malononitrile (CS) and oleoresin capsicum (OC) in awake rats.

Tear gases are largely used to control civil unrest. Their incapaciting effects involve eyes, skin and respiratory tract. This study was performed to compare acute respiratory effects of o-chlorobenzylidene malononitrile (CS), oleoresin capsicum (OC) and their respective solvents in awake rats, using an integrated system of nose-only exposure and multiple monitoring of breathing. Aerosols were generated by a Collison Nebulizer from the solutions held in tear gas sprays. The reduction of minute ventilation, observed during a 5 min exposure, was significantly more important with CS than with OC: minute ventilation represented 29+/-8 and 50+/-6% of pre-exposure minute ventilation respectively (P<0.05). The reduction of minute ventilation observed with CS and OC solvents alone was not significantly different from that observed with the tear gases themselves. The decrease in minute ventilation observed, between the second and the fifth minute of exposure, was of the same level for repeated exposure separated by 24 h. Time necessary to recover to 80% of pre-exposure minute ventilation was not significantly different between the two tear gases: 722+/-272 and 691+/-262 s for CS and OC respectively (NS). Histological analysis of the trachea, performed at the end of exposures, revealed an increase in mucus secretion after exposure to OC and cytoplasmic vacuoles in epithelial cells after exposure to CS. In the lungs, interstitial oedema was observed after exposure to OC and emphysema after exposure to CS.

Ketone potentiation of intrahepatic cholestasis: effect of two aliphatic isomers.

Occupational exposure to methyl isobutyl ketone (MiBK) or methyl n-butyl ketone (MnBK) normally occurs by inhalation. The present study reports that exposure to both ketones can potentiate cholestasis experimentally induced by taurolithocholic acid (TLC, 30 mumol/kg) or by a combination of manganese (Mn, 4.5 mg/kg) and bilirubin (BR, 25 mg/kg). Male Sprague-Dawley rats were exposed for 3 d, 4 h/d, to MiBK or MnBK vapors using 0.5, 1, 1.5, or 2 times the minimal effective concentration (MEC). The estimated MiBK or MnBK MEC for potentiating TLC- or Mn-BR-induced cholestasis were 400 and 150 ppm, respectively. Eighteen hours after ketone exposure, rats were injected i.v. with TLC or Mn-BR. Bile flow was measured from 15 to 150 min after the cholestatic regimen. Rats exposed to MiBK or MnBK exhibited an enhanced diminution in bile flow compared to controls that was dose-dependent with the inhaled ketone dose. The dose-effect characteristics of the potentiation phenomenon were established. Results indicate that MiBK or MnBK inhalation potentiated both TLC and Mn-BR cholestasis in a dose-related fashion. Quantitative differences, however, exist between both ketones with respect to their ability to potentiate both models. Comparison between the two isomers was established, and MnBK was found to be more potent than MiBK.

Altered cholesterol synthesis as a mechanism involved in methyl isobutyl ketone-potentiated experimental cholestasis.

Mechanisms by which ketones potentiate manganese-bilirubin (Mn-BR)-induced cholestasis are unknown. The purpose of the present study was to investigate the effect of methyl isobutyl ketone (MiBK), a widely used ketonic solvent, at the level of the bile canalicular membrane (BCM) and to verify if altered membrane lipid dynamics could be involved in MiBK-potentiated Mn-BR cholestasis. Male Sprague-Dawley rats were exposed 4 hr/day for 3 days to MiBK vapors (200 or 600 ppm). Eighteen hours after the last exposure, manganese (Mn, 4.5 mg/kg) was given i.v. followed 15 min later by bilirubin (BR, 25 mg/kg). Rats were killed 30 min after BR; liver cell plasma membranes (bile canalicular and sinusoidal), microsomes, mitochondria, and cytosol were isolated by differential centrifugation. Lipids were extracted and cholesterol was measured in each fraction. After Mn-BR and MiBK exposure (600 ppm), results indicated a marked increase in BCM cholesterol content compared to rats exposed to air only. This increase was greater than that due to Mn-BR or MiBK given alone. Also, results indicated that cholesterol increased in a dose-related fashion in BCM after MiBK exposure, whereas PM cholesterol remained unaltered. To identify the source of the increased BCM cholesterol and to permit distinction between de novo cholesterol synthesis and subcellular shifts, the hepatic lipid pool was labeled in vivo with [3H]-cholesterol and [2-14C]-mevalonic acid, a cholesterol synthesis precursor. Results showed that after 600 ppm MiBK exposure, 14C-labeled cholesterol was greater than 3H-labeled cholesterol, indicating that the contribution of de novo cholesterol synthesis to the total cholesterol content of the various isolated hepatocellular fractions was more important than the contribution of intracellular pools. Therefore, increased BCM cholesterol content and enhanced accumulation of newly synthesized cholesterol appear to be involved in MiBK potentiation of Mn-BR-induced cholestasis.

Ketone potentiation of haloalkane-induced hepato- and nephrotoxicity. I. Dose-response relationships.

Carbon tetrachloride (CCl4) induced hepatotoxicity and chloroform (CHCl3) induced nephrotoxicity were evaluated in male Sprague-Dawley rats pretreated with acetone (A), methyl ethyl ketone (MEK), and methyl isobutyl ketone (MiBK). Dose-response relationships for A, MEK, and MiBK potentiation of CCl4-induced hepatotoxicity and CHCl3-induced nephrotoxicity were compared. A, MEK, and MiBK pretreatment at a dosage of 6.8 mmol/kg, given daily for 3 d, markedly potentiated CCl4-induced liver toxicity as indicated by a decrease in the CCl4 ED50 to 3.4, 4.6, and 1.8 mmol/kg, respectively, compared to vehicle-pretreated rats (17.1 mmol/kg). Similarly, pretreatment with these ketones (13.6 mmol/kg) potentiated CHCl3 kidney toxicity but to a lesser degree; CHCl3 ED50 values for vehicle-, A-, MEK-, and MiBK-pretreated rats were 3.4, 1.6, 2.1, and 2.2 mmol/kg, respectively. Our results indicate a potency ranking profile for the potentiation of CCl4 hepatotoxicity of MiBK > A > MEK and of A > MEK > or = MiBK for CHCl3 nephrotoxicity. These dissimilar ranking profiles could be due to differences in mechanisms of action for the two target sites.

Different contributions of cytochrome P450 2E1 and P450 2B1/2 to chloroform hepatotoxicity in rat.

The contribution of cytochrome P450 isozymes CYP2E1 and CYP2B1/2 to chloroform-induced hepatotoxicity taken at 18 hr after the treatment was investigated in rats treated with n-hexane as an inducer of CYP2E1, 2-hexanone as an inducer of CYP2E1 and CYP2B1/2, and phenobarbital (PB) as an inducer of CYP2B1/2. Hepatic damage was evaluated by gross measurement of plasma alanine aminotransferase activity and histopathological examination. All treatments potentiated chloroform-induced hepatic damage. In n-hexane-pretreated rats, the damage was maximal with the middle dose of chloroform (0.2 ml/kg), whereas the damage increased with dose in rats treated with 2-hexanone or PB. The degree of hepatic damage induced with the three pretreatments was in the following order: n-hexane > 2-hexanone = PB with the middle dose of chloroform and PB >> 2-hexanone > n-hexane with the high dose (0.5 ml/kg); little difference among the pretreatments was seen with the low dose (0.1 ml/kg). These findings suggest that CYP2E1 is a low Km isoform and CYP2B1/2 a high Km isoform for chloroform activation. CYP2E1-dependent hepatic damage was characterized by ballooned hepatocytes, which were restricted to the centrilobular area; with CYP2B1/2, more necrotic than ballooned hepatocytes were seen and the necrotic hepatocytes were found not only in the centrilobular but also in the midzonal and periportal areas. Chloroform treatment did not affect the activity of N-nitrosodimethylamine N-demethylase in pretreated rats; the high dose increased the activity in control rats. In contrast, the high dose of chloroform decreased the activity of 7-pentoxyresorufin O-depentylase in all induced rats but not in controls. Immunoinhibition and immunoblot analyses showed that the high dose of chloroform induced CYP2E1 in control rats but decreased CYP2B1/2 in all pretreated rats. These results suggest that although both CYP2E1 and CYP2B1/2 contribute to chloroform-induced hepatic damage, they do so quite differently.

Tissue concentrations of methyl isobutyl ketone, methyl n-butyl ketone and their metabolites after oral or inhalation exposure.

Quantitative relationships between plasma, liver and lung methyl isobutyl ketone (MiBK) and methyl n-butyl ketone (MnBK) concentrations after oral or inhalation exposure were established. Their respective metabolites (4-methyl-2-pentanol, 4-hydroxy-methyl isobutyl ketone, 2-hexanol, and 2,5-hexanedione) were also quantified. Male Sprague-Dawley rats were exposed for 3 days to MiBK or MnBK vapors (4 h/day) or treated orally for 3 days with a MiBK- or MnBK-corn oil solution. Both ketones and their respective metabolites in plasma or tissue concentrations were determined by gas chromatography. MiBK and MnBK plasma and tissue concentrations increased in a dose-related manner with the administered dose irrespective of the route of administration. Metabolite concentrations, however, were influenced by the route of administration.

Plasma concentrations in methyl isobutyl ketone-potentiated experimental cholestasis after inhalation or oral administration.

In studies of methyl isobutyl ketone (MiBK)-potentiated cholestasis induced by taurolithocholic acid (TLC) or manganese-bilirubin (Mn-BR) combinations, MiBK is usually given by gavage whereas industrial exposure to MiBK normally occurs by inhalation. The present study was conducted to verify if the route of administration could influence the potentiation. Male Sprague-Dawley rats were treated with MiBK for 3 days orally or by inhalation (4 hr/day). The minimal effective doses (MED) for potentiating both models of intrahepatic induced cholestasis were estimated to be 3 mmol/kg or 400 ppm for the oral or inhalation route, respectively. Groups of rats were treated with 0.5, 1, or 2 times the MED. Half of each group was sacrificed after the last MiBK administration to determine plasma concentrations of MiBK and its metabolites by gas-liquid chromatography. The other half was challenged 18 hr later with TLC (30 mumol/kg) or a combination of manganese (4.5 mg/kg) and bilirubin (15 mg/kg). Bile flow was measured from 15 to 135 min after the cholestatic challenge. Rats exposed to MiBK orally or by inhalation exhibited an enhanced diminution in bile flow that was dose-dependent. With dosages of 3 mmol/kg po or 400 ppm by inhalation or more, diminution in bile flow was significantly different from control values. Comparisons between maximal bile flow decrease and MiBK plasma concentration showed that the severity of the hepatotoxic response was dependent on the plasma MiBK concentration, irrespective of the route of administration.

Modulation of hexachlorobenzene-induced hepatic porphyria by methyl isobutyl ketone in the rat.

Potential toxic interaction between hexachlorobenzene (HCB) and methyl isobutyl ketone (MiBK) was investigated using two different schedules of toxicant administration. The first schedule involved simultaneous administration of HCB (50 mg/kg/d, p.o. in 10 ml/kg corn oil at 10.00 a.m. for 5 d/wk) and MiBK (7.5 mmol/kg/d, p.o. in 10 ml/kg corn oil at 4.00 p.m. for 3 d/wk) for 6 weeks. The second schedule involved an initial dosing of 25 or 50 mg HCB/kg/d for 12 consecutive days, followed by the administration of 7.5 mmol MiBK/kg every other day for 27 days. When administered simultaneously, MiBK reduced the severity of HCB-induced porphyria, but when given sequentially after HCB accumulation, it enhanced the porphyrinogenic response. These results suggest that the effect of combined exposure to HCB and MiBK on hepatic porphyria depends on the sequence of the administration of both chemicals, and that the mechanism involved in this interaction may invoke both the induction and inhibition of specific hepatic isoenzymes by MiBK.

Percutaneous uptake and kinetics of methyl isobutyl ketone (MIBK) in the guinea-pig.

Continuous intravenous infusion of 0.478 mumol/min methyl isobutyl ketone (MIBK) was performed for 30 min in pentobarbital-anesthetized guinea-pigs. Epicutaneous exposure for 150 min was carried out 2.5 h later after administration of MIBK to a sealed glass ring on the clipped back of the animals. Arterial blood was analyzed for MIBK by gas chromatography. Blood clearance averaged 201 ml.min-1.kg-1 body wt. A maximum percutaneous uptake of 1.1 mumol.min-1.cm-2 was reached 10-45 min after the onset of exposure and decreased to 0.56 mumol.min-1.cm-2 during the latter part of exposure.

Induction of cytochrome P450 isozymes by simultaneous inhalation exposure of hens to n-hexane and methyl iso-butyl ketone (MiBK).

Chickens were exposed simultaneously to the industrial hexacarbon solvents n-hexane and methyl iso-butyl ketone (MiBK). n-Hexane has been shown to be neurotoxic in both humans and other vertebrates. While MiBK is not neurotoxic, it has been shown to greatly synergize the clinical appearance of neurotoxicity in animals exposed to both of these solvents. Groups of hens were exposed for 29 days in inhalation chambers to 1000 ppm n-hexane in combination with 10, 100, 250, 500, or 1000 ppm MiBK. Other groups received either 1000 ppm n-hexane, 1000 ppm MiBK, or ambient air and served as controls. A dose-dependent decrease in body weight and an increase in clinical effects were noted for the highest exposure groups (1000 ppm n-hexane combined with 1000, 500 or 250 ppm MiBK). There was an MiBK dose-dependent increase in cytochrome P450 content and benzphetamine N-demethylase activity, but there was no distinct pattern for ethoxyresorufin O-deethylase or cytochrome c reductase activities. Mixed-function oxidase levels and activities (cytochrome P450 content and benzphetamine N-demethylase) were elevated significantly (P less than 0.05) over controls even in the lowest MiBK group (10 ppm), although there were no clinical signs of neurotoxicity. Four different isozymes of cytochrome P450 were measured immunologically. There was a dose-dependent increase in three of the isozymes, two of which were phenobarbital inducible and one of which was induced by beta-napthoflavone. Quantitatively, the largest increase was in the PB-A isozyme, a phenobarbital-inducible isozyme which accounted for approximately 70% of the cytochrome P450 present in animals treated with MiBK. The results suggest that MiBK selectively induces cytochrome P450 isozymes leading to the metabolic activation of the weak neurotoxicant n-hexane to the potent neurotoxicant 2,5-hexanedione (2,5-HD).

Mechanism of transport and distribution of organic solvents in blood.

Little is known about the mechanism of transport and distribution of volatile organic compounds in blood. Studies were conducted on five typical organic solvents to investigate how these compounds are transported and distributed in blood. Groups of four to five rats were exposed for 2 hr to 500 ppm of n-hexane, toluene, chloroform, methyl isobutyl ketone (MIBK), or diethyl ether vapor; 94, 66, 90, 51, or 49%, respectively, of these solvents in the blood were found in the red blood cells (RBCs). Very similar results were obtained in vitro when aqueous solutions of these solvents were added to rat blood. In vitro studies were also conducted on human blood with these solvents; 66, 43, 65, 49, or 46%, respectively, of the added solvent was taken up by the RBCs. These results indicate that RBCs from humans and rats exhibited substantial differences in affinity for the three more hydrophobic solvents studied. When solutions of these solvents were added to human plasma and RBC samples, large fractions (51-96%) of the solvents were recovered from ammonium sulfate-precipitated plasma proteins and hemoglobin. Smaller fractions were recovered from plasma water and red cell water. Less than 10% of each of the added solvents in RBC samples was found in the red cell membrane ghosts. These results indicate that RBCs play an important role in the uptake and transport of these solvents. Proteins, chiefly hemoglobin, are the major carriers of these compounds in blood. It can be inferred from the results of the present study that volatile lipophilic organic solvents are probably taken up by the hydrophobic sites of blood proteins.