Developmental toxicity studies in Crl:CD (SD) rats following inhalation exposure to trichloroethylene and perchloroethylene.

The potential for trichloroethylene (TCE) and perchloroethylene (PERC) to induce developmental toxicity was investigated in Crl:CD (SD) rats whole-body exposed to target concentrations of 0, 50, 150 or 600 ppm TCE or 0, 75, 250 or 600 ppm PERC for six hours/day, seven days/week on gestation day (GD) 6-20 and 6-19, respectively. Actual chamber concentrations were essentially identical to target with the exception of the low PERC exposure level, which was 65 ppm. The highest exposure levels exceeded the limit concentration (2 mg/L) specified in the applicable test guidelines. Maternal necropsies were performed the day following the last exposure. Dams exposed to 600 ppm TCE exhibited maternal toxicity, as evidenced by decreased body weight gain (22% less than control) during GD 6-9. There were no maternal effects at 50 or 150 ppm TCE and no indications of developmental toxicity (including heart defects or other terata) at any exposure level tested. Therefore, the TCE NOEC for maternal toxicity was 150 ppm, whereas the embryo/fetal NOEC was 600 ppm. Maternal responses to PERC were limited to slight, but statistically significant reductions in body weight gain and feed consumption during the first 3 days of exposure to 600 ppm, resulting in a maternal NOEC of 250 ppm. Developmental effects at 600 ppm consisted of reduced gravid uterus, placental and fetal body weights, and decreased ossification of thoracic vertebral centra. Developmental effects at 250 ppm were of minimal toxicological significance, being limited to minor decreases in fetal and placental weight. There were no developmental effects at 65 ppm.

Inhalation exposure to methylene chloride does not induce systemic immunotoxicity in rats.

Methylene chloride (dichloromethane) is used in a variety of industrial applications. To date, there has been no formal assessment of immunotoxicity attributed to methylene chloride. Studies were undertaken to examine whether methylene chloride has any potential to influence the integrity of immune function. For this purpose, Sprague-Dawley rats of both genders were exposed by inhalation to a single high dose (5000 ppm) of methylene chloride for 6 h/d, 5 d/wk for 28 d. This was considered the relevant route of administration, as not only is inhalation a primary route for human exposure to methylene chloride, but, also, the chemical is absorbed rapidly via the lungs. Under these conditions of exposure, methylene chloride failed to influence absolute or relative thymus weights in either gender and produced a significant reduction in relative, but not absolute, spleen weight in female rats only. Immunocompetence was measured as a function of the ability of treated animals to mount immunoglobulin M (IgM) antibody responses to sheep red blood cells (SRBC) as determined by enzyme-linked immunosorbent assay (ELISA). Exposure to methylene chloride did not affect antibody production. Evidence indicates that under these conditions of exposure, methylene chloride did not compromise immune function.

In vitro absorption of some o-phthalate diesters through human and rat skin.

The absorption of undiluted phthalate diesters [dimethyl phthalate (DMP), diethylphthalate (DEP), dibutyl phthalate (DBP) and di-(2-ethylhexyl)phthalate (DEHP)] has been measured in vitro through human and rat epidermal membranes. Epidermal membranes were set up in glass diffusion cells and their permeability to tritiated water measured to establish the integrity of the skin before the phthalate esters were applied to the epidermal surface. Absorption rates for each phthalate ester were determined and a second tritiated water permeability assessment made to quantify any irreversible alterations in barrier function due to contact with the esters. Rat skin was consistently more permeable to phthalate esters than the human skin. As the esters became more lipophilic and less hydrophilic, the rate of absorption was reduced. Contact with the esters caused little change in the barrier properties of human skin, but caused marked increases in the permeability to water of rat skin. Although differences were noted between species, the absolute rates of absorption measured indicate that the phthalate esters are slowly absorbed through both human and rat skin.

Absorption through the skin: theory, in vitro techniques and their applications.

Human exposure to chemicals often occurs by skin contact and estimations of percutaneous absorption play an important part in the assessment of the risks involved. It is well established that the rate determinant step in absorption through skin is diffusion across the outermost layer, the stratum corneum, a process that is known to be passive in character. This latter property, combined with the stability of the stratum corneum, makes possible meaningful in vitro measurements of skin permeability in diffusion cells. The use of diffusion cells has permitted the development of a thorough understanding of the physical chemistry of the absorption process. Various parameters can be established in vitro that allow assessments of absorption to be calculated for a range of exposure situations. Another major advantage is that human skin samples may be used in vitro and, since no animal model has been found to be adequate for all penetrant chemicals, this has proved to be the most relevant tissue for assessing absorption in practical circumstances.

A model for quantifying absorption through abnormal skin.

Techniques are available for quantitatively studying factors governing absorption through normal skin (in vivo and in vitro) but relatively little is known about the permeability of abnormal skin. We have designed and evaluated an in vivo model for quantifying absorption through abnormal skin. Absorption of [3H]mannitol and [14C]octyl benzoate was studied through altered rat skin. [3H]Mannitol penetrated normal skin much more slowly than did [14C]octyl benzoate. Abnormal skin was more permeable to [3H]mannitol and [14C]octyl benzoate, absorption was greater than 100X and greater than 2X greater, respectively, than normal. The in vivo model has been successfully used to quantify absorption through abnormal skin.

Permeability of abnormal rat skin.

We have measured the permeability (to water in vivo and in vitro) and examined the histology of rat skin after mild, superficial epidermal alterations: scalpel blade (Cat I) and sandpaper abrasion (Cat II), adhesive tape stripping (Cat III), and suction blister top removal (Cat IV). After each alteration the permeability was increased (Cat IV greater than Cat III greater than Cat II greater than Cat I) and the epidermis regenerated in a distinct, biphasic manner, as indicated by the permeability and histology data. The rapid first phase corresponded with a decrease in permeability and the development of a scab (the greater the increased permeability, the slower the rate of regeneration). The second phase was more gradual (with a similar rate of regeneration after each alteration) and corresponded with a return to normal permeability and gradual thickening of the stratum corneum (return to normal corresponded with degree of initial stratum corneum removal). A similar, though slower biphasic regeneration has been reported to occur in human skin following similar types of alterations. It is concluded that abnormal rat skin is suitable for quantifying absorption through abnormal epidermis.

A comparison of the in vitro permeability properties of human and some laboratory animal skins.

Synopsis The in vitro permeability of human and some common laboratory animals'skin has been measured to two chemicals with different physico-chemical properties: [(3)H] water, a small polar molecule and [(14)C]paraquat dichloride (the ion), a fully ionised divalent cation. All the skins examined had similar permeabilities to [(3)H]water: relative to human skin the greatest difference (x4.8) was measured through guinea pig skin. However, relative to human skin, all the animals were much more permeable to [(14)C]paraquat, with factors of difference ranging between 40 (haired rat) and 1460 (hairless mouse). The in vitro technique allows the permeability of human and animal skin to be compared, facilitating extrapolation to Man of effects seen in animals during dermal exposure studies.

Vapour and liquid diffusion of model penetrants through human skin; correlation with thermodynamic activity.

This work investigates vapour and liquid permeation through human skin of model penetrants benzyl alcohol, benzaldehyde, aniline, anisole and 2-phenylethanol applied in model vehicles butanol, butyl acetate, isophorone, isopropyl myristate, propylene carbonate, toluene, n-heptane and water. Vapour permeation was a linear function of thermodynamic activity as measured by headspace gas chromatography, except when the vehicle was n-heptane. Liquid permeation did not always follow simple thermodynamic predictions, e.g. for the penetrant, benzyl alcohol, when the vehicle damaged the skin (toluene, n-heptane) or when propylene carbonate produced low fluxes and isopropyl myristate, high values. At comparable thermodynamic activities, liquid fluxes were often ten-fold higher than vapour fluxes, and these differences were reflected by the partition coefficients and the amount of penetrant entering the stratum corneum membrane. The conclusion was that liquid fluxes were membrane controlled, whereas an interfacial effect probably contributed to low vapour permeation.

Correlation of thermodynamic activity and vapour diffusion through human skin for the model compound, benzyl alcohol.

This work tested the potential for predicting percutaneous absorption rates of a volatile penetrant from any vehicle by using thermodynamic activity measurements. Benzyl alcohol was chosen as a non-ideal, hydrogen bonding, volatile model penetrant. A manual headspace gas chromatography method measured benzyl alcohol vapour concentrations and thermodynamic activities above binary mixtures with vehicles: butanol, butyl acetate, isopropyl myristate, isophorone, toluene and propylene carbonate. Benzyl alcohol vapour diffusion through human, abdominal skin was also measured in-vitro for these mixtures. The benzyl alcohol vapour flux was linearly related to the activity, suggesting that percutaneous absorption is controlled by thermodynamic activity when the vehicle has no effect on the stratum corneum barrier.

Absorption of some glycol ethers through human skin in vitro.

To assist evaluation of the hazards of skin contact with selected undiluted glycol ethers, their absorption across isolated human abdominal epidermis was measured in vitro. Epidermal membranes were set up in glass diffusion cells and, following an initial determination of permeability to tritiated water, excess undiluted glycol ether was applied to the outer surface for 8 hr. The appearance of glycol ether in an aqueous "receptor" phase bathing the underside of the epidermis was quantified by a gas chromatographic technique. A final determination of tritiated water permeability was compared with initial values to establish any irreversible alterations in epidermal barrier function induced by contact with the glycol ethers. 2-methoxyethanol (EM) was most readily absorbed (mean steady rate 2.82 mg/cm2/hr), and a relatively high absorption rate (1.17 mg/cm2/hr) was also apparent for 1-methoxypropan-2-ol (PM). There was a trend of reducing absorption rate with increasing molecular weight or reducing volatility for monoethylene glycol ethers (EM, 2.82 mg/cm2/hr; 2-ethoxyethanol, EE, 0.796 mg/cm2/hr; 2-butoxyethanol, EB, 0.198 mg/cm2/hr) and also within the diethylene glycol series: 2-(2-methoxyethoxy) ethanol (DM, 0.206 mg/cm2/hr); 2-(2-ethoxyethoxy) ethanol (DE, 0.125 mg/cm2/hr) and 2-(2-butoxyethoxy) ethanol (DB, 0.035 mg/cm2/hr). The rate of absorption of 2-ethoxyethyl acetate (EEAc) was similar to that of the parent alcohol, EE. Absorption rates of diethylene glycol ethers were slower than their corresponding monoethylene glycol equivalents. Combination of intrinsic toxicity and ability to pass across skin contribute to assessment of hazards of contact with undiluted glycol ethers.

Effects of freezing on human skin permeability.

The percutaneous absorption of water was measured in-vitro at 30 degrees C for pale caucasian abdominal skin which had been stored at -20 degrees C for up to 466 days and compared with fresh skin. Prolonged freezing of the skin did not affect the absorption of water which had a mean permeability coefficient of 1.71 +/- 0.62 X 10(-3) cm h-1 (180 diffusion experiments with 39 skin specimens). No significant difference was found between the absorption of water through human skin which was fresh or had been frozen. The mean permeability coefficient for skin which had not been frozen was 1.30 +/- 0.55 X 10(-3) cm h-1 for 6 skin specimens.

A comparison of techniques for the measurement of transepidermal water loss.

An Evaporimeter and a ventilated chamber technique have been compared in their ability to measure transepidermal water loss (TEWL) through rat skin. These techniques measure TEWL under very different conditions; the Evaporimeter measures the net TEWL under ambient relative humidity (RH) whereas the ventilated chamber employs a constant atmosphere, usually of low RH and thus measured the uni-directional diffusion of water. Paired Evaporimeter and ventilated chamber measurements were made of TEWL through normal skin and through skin whose barrier properties had been altered by tape-stripping (15 applications) or single applications of n-hexadecane (28.4 mumol cm-2). Both measuring techniques indicated the same level of TEWL through normal skin (mean 0.3 mg cm-2 h-1) and during increases in TEWL induced by n-hexadecane (max TEWL c 3.5 mg cm-2 h-1). However, the Evaporimeter was found to underestimate the higher rates of TEWL induced by tape-stripping, ie above TEWL raters of 7.5 mg cm-2 h-1. The Evaporimeter is portable, easy to use and suitable for measurements of net water loss up to 7.5 mg cm-2 h-1; it can only be used for comparative assessment of epidermal barrier function if used at a particular ambient RH. The more cumbersome ventilated chamber is to be preferred for accurate assessments of barrier function where high rates of TEWL occur.

Non-ionic surfactants and gastric mucosal transport of paraquat.

The influence of a range of polyoxyethylated non-ionic surfactants upon the transport of [14C]-paraquat dichloride across rabbit isolated gastric mucosa was investigated. Paraquat was shown to cross the mucosal epithelium by passive diffusion. Certain members of the surfactant series enhanced transmucosal paraquat transfer at low surfactant concentrations (e.g. 0.001%) but the occurrence and magnitude of this effect was not dependent in a simple manner upon surfactant structure of physiochemical properties. At micellar concentrations the increase in paraquat transport was greatest with those surfactants possessing both ethylene oxide chains of 10-20 units and alkyl chains longer than C7-C9. The most effective absorption promoter was found to be Brij 36T (C12E10). At micellar surfactant concentrations the enhancement of paraquat transfer appeared, from histological evidence, to be related to the ability of the surfactants to solubilize membrane components and disrupt epithelial cells.