Contaminated soils (I): In vitro dermal absorption of benzo[a]pyrene in human skin

Dermal absorption of the lipophile and potential carcinogen benzo[a]pyrene (BaP) in soils from contaminated sites was simulated in vitro using human skin exposed to 14C-BaP-spiked soil. This study is the first in a series of tests at Health Canada with several soil contaminants spanning a wide range of lipophilicity conducted with viable human skin. Breast skin was obtained fresh from a local hospital and dermatomed to a thickness of 0.4-0.5 mm. Teflon Bronaugh diffusion cells were perfused with HEPES buffered Hanks saline (pH 7.4) with 4% bovine serum albumin (BSA) and fractions were collected at 6-h intervals for up to 24 h exposure either to 14C-BaP applied in acetone or spiked in a commercial gardening soil. As skin depot 14C levels were still high at 24 h, the study was repeated for up to 42 h to examine skin depot bioavailability. Skin was washed with soapy water at 24 h in both the 24- and 42-h studies. Exposure to 14C-BaP both with and without soil was conducted in triplicate with skin specimens from at least 4 patients. In the 24-h exposure tests including the skin depot there was 15 and 56% absorption with and without soil, respectively. The lower total percent absorption from the spiked soil applied to skin resulted from lower depot absorption of 8% with and 45% without soil. Data for 42-h studies were similar and revealed no significant decrease in skin depot levels. Including the 42-h depots there was 16 and 50% absorption with and without soil, respectively, with respective depots of 7 and 39%. As there was no significant difference between the 24- and 42-h depots both with and without soil, the data suggest the depot for BaP was not bioavailable for at least the additional 18-h post soap wash exposure. The bioavailability of BaP is discussed in relation to previous in vitro and in vivo studies in perspective with dermal exposure to contaminated soils.     

Mixture component effects on the in vitro dermal absorption of pentachlorophenol

Interactions between chemicals in a mixture and interactions of mixture components with the skin can significantly alter the rate and extent of percutaneous absorption, as well as the cutaneous disposition of a topically applied chemical. The predictive ability of dermal absorption models, and consequently the dermal risk assessment process, would be greatly improved by the elucidation and characterization of these interactions. Pentachlorophenol (PCP), a compound known to penetrate the skin readily, was used as a marker compound to examine mixture component effects using in vitro porcine skin models. PCP was administered in ethanol or in a 40% ethanol/60% water mixture or a 40% ethanol/60% water mixture containing either the rubefacient methyl nicotinate (MNA) or the surfactant sodium lauryl sulfate (SLS), or both MNA and SLS. Experiments were also conducted with 14C-labelled 3,3',4,4'-tetrachlorobiphenyl (TCB) and 3,3',4,4',5-pentachlorobiphenyl (PCB). Maximal PCP absorption was 14.12% of the applied dose from the mixture containing SLS, MNA, ethanol and water. However, when PCP was administered in ethanol only, absorption was only 1.12% of the applied dose. There were also qualitative differences among the absorption profiles for the different PCP mixtures. In contrast with the PCP results, absorption of TCB or PCB was negligible in perfused porcine skin, with only 0.14% of the applied TCB dose and 0.05% of the applied PCB dose being maximally absorbed. The low absorption levels for the PCB congeners precluded the identification of mixture component effects. These results suggest that dermal absorption estimates from a single chemical exposure may not reflect absorption seen after exposure as a chemical mixture and that absorption of both TCB and PCB are minimal in this model system.     

Fate of chemicals in skin after dermal application: does the in vitro skin reservoir affect the estimate of systemic absorption?

Recent international guidelines for the conduct of in vitro skin absorption studies put forward different approaches for addressing the status of chemicals remaining in the stratum corneum and epidermis/dermis at the end of a study. The present study investigated the fate of three chemicals [dihydroxyacetone (DHA), 7-(2H-naphtho[1,2-d]triazol-2-yl)-3-phenylcoumarin (7NTPC), and disperse blue 1 (DB1)] in an in vitro absorption study. In these studies, human and fuzzy rat skin penetration and absorption were determined over 24 or 72 h in flow-through diffusion cells. Skin penetration of these chemicals resulted in relatively low receptor fluid levels but high skin levels. For DHA, penetration studies found approximately 22% of the applied dose remaining in the skin (in both the stratum corneum and viable tissue) as a reservoir after 24 h. Little of the DHA that penetrates into skin is actually available to become systemically absorbed. 7NTPC remaining in the skin after 24 h was approximately 14.7% of the applied dose absorbed. Confocal laser cytometry studies with 7NTPC showed that it is present across skin in mainly the epidermis and dermis with intense fluorescence around hair. For DB1, penetration studies found approximately 10% (ethanol vehicle) and 3% (formulation vehicle) of the applied dose localized in mainly the stratum corneum after 24 h. An extended absorption study (72 h) revealed that little additional DB1 was absorbed into the receptor fluid. Skin levels should not be considered as absorbed material for DHA or DB1, while 7NTPC requires further investigation. These studies illustrate the importance of determining the fate of chemicals remaining in skin, which could significantly affect the estimates of systemically available material to be used in exposure estimates. We recommend that a more conclusive means to determine the fate of skin levels is to perform an extended study as conducted for DB1.     

Cutaneous corticosteroid-induced glaucoma

Skin contact with nonylphenol ethoxylates (NPE), a group of widely used surfactants, is the primary source of human exposure. Previous studies have shown that the absorption of NPE through human and animal skin in vitro is limited (<1% over 8 hr) [Monteiro-Riviere et al. Toxicol Indust Health 2000; 16:49–57]. The purpose of this study was to examine the percutaneous absorption of NPE and the chemical precursor, nonylphenol (NP), in the isolated perfused porcine skin flap (IPPSF) model for comparison to the in vitro porcine skin flow through (PSFT) diffusion studies. The IPPSF model is considered to accurately predict absorption of chemicals through human skin. The IPPSF was dosed with 100 μl of 1% ¹⁴C ring-labeled NP, ¹⁴C ring-labeled NPE-4, or ¹⁴C ring-labeled NPE-9 in aqueous polyethylene glycol (PEG-400) solution and perfused for 8 hr. All three chemicals were minimally absorbed, with only approximately 0.1% of the applied dose found in the perfusate over the 8-hr collection. This absorbed material represents the systemic exposure expected following skin contact in humans. In addition, less than 1% of the applied dose penetrated into the stratum corneum and underlying dermis, but remained within the skin and did not go through to the perfusate. Thus, the overall potential systemic exposure to these chemicals from skin contact, using a model considered similar to human skin in vivo, is less than 1%. The absorption results of this study were consistent with previous studies in the PSFT model. The penetration of NPEs and NP in the IPPSF was less than the PSFT and is probably more predictive of in vivo human absorption as this model is physiologically closer to human skin. This suggests that the overall potential for skin absorption of these chemicals in humans is even lower than previous estimates.     

In vitro dermal absorption of flame retardant chemicals

Flame retardant chemicals may be used in furniture fabric in the future to reduce the flammability of the fabric. As a part of the process to evaluate the potential for exposure to these chemicals, this study examined the in vitro dermal absorption of two flame retardant chemicals. The chemicals were [¹⁴C]decabromodiphenyl oxide (DBDPO) and [¹⁴C]tris-(1,3-dichloro-2-propyl)phosphate (TDCP). Skin from the adult hairless female mouse (SKH1) was removed and mounted in flow-through diffusion cells. The chemicals, at three dose levels (DBDPO: 6, 30 and 60 nmol; TDCP: 20, 100 and 200 pmol), were applied in a volatile vehicle (tetrahydrofuran for DBDPO; acetone for TDCP) to the skin. Fractions of receptor fluid, pumped below the skin, were collected over a 24-h period. The skin was washed with solvent (tetrahydrofuran for DBDPO; ethanol for TDCP) to remove unabsorbed chemical 24 h after application. The receptor fluid, skin wash and skin were analyzed for chemical-derived radioactivity. The skin from the high-dose group of both chemicals, and the receptor fluid from TDCP high-dose samples, were analyzed for parent compound and metabolites by HPLC. The 24-h cumulative percent of the dose of DBDPO in the receptor fluid was very low (0.07–0.34%). The applied dose of DBDPO detected in the skin ranged from 2 to 20%. The lowest dose of DBDPO had the highest percentage of the dose (20%) in the skin. The major portion of the applied dose was removed by washing the skin 24 h after application of DBDPO, and ranged from 77 to 92%. HPLC analysis of homogenate extract prepared from the high-dose of DBDPO-treated skin showed the presence of DBDPO and a minor unknown peak. TDCP was readily detected in the receptor fluid; 39–57% of the applied dose of TDCP was in the receptor fluid by 24 h. The solvent wash removed 11–25% of the dose from the skin and 28–35% remained in it. HPLC analysis of the skin homogenate extract and receptor fluid extract from the TDCP high-dose treated samples showed the presence of parent compound and a minor unknown peak. TDCP more readily penetrated hairless mouse skin and diffused into the receptor fluid than DBDPO. TDCP has a lower molecular weight and log octanol:water partition coefficent than DBDPO. The differences in the physico-chemical properties of these two chemicals most likely explains their dissimilar absorption through hairless mouse skin.     

Assessing the safety of cosmetic chemicals: Consideration of a flux decision tree to predict dermally delivered systemic dose for comparison with oral TTC (Threshold of Toxicological Concern)

Threshold of Toxicological Concern (TTC) aids assessment of human health risks from exposure to low levels of chemicals when toxicity data are limited. The objective here was to explore the potential refinement of exposure for applying the oral TTC to chemicals found in cosmetic products, for which there are limited dermal absorption data. A decision tree was constructed to estimate the dermally absorbed amount of chemical, based on typical skin exposure scenarios. Dermal absorption was calculated using an established predictive algorithm to derive the maximum skin flux adjusted to the actual ‘dose’ applied. The predicted systemic availability (assuming no local metabolism), can then be ranked against the oral TTC for the relevant structural class. The predictive approach has been evaluated by deriving the experimental/prediction ratio for systemic availability for 22 cosmetic chemical exposure scenarios. These emphasise that estimation of skin penetration may be challenging for penetration enhancing formulations, short application times with incomplete rinse-off, or significant metabolism. While there were a few exceptions, the experiment-to-prediction ratios mostly fell within a factor of 10 of the ideal value of 1. It can be concluded therefore, that the approach is fit-for-purpose when used as a screening and prioritisation tool.     

In vitro dermal absorption of pesticides: V. In vivo and in vitro comparison of the herbicide 2,4-dichlorophenoxyacetic acid in rat, guinea pig, pig, human and tissue-cultured skin

In vitro dermal absorption tests were conducted with the ¹⁴C-ring-labelled herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), dissolved in acetone and applied to dermatomed skin (0.5 mm) of a number of species at dose rates of 7–8 μg/cm². Skin absorption was determined for 48 hr after exposure using an in vitro flow-through system. Skin absorption was calculated from the sum of the percentage recovery of ¹⁴C activity in the receiver solution and the percentage recovery in the methanol washes of the skin at 48 hr and the skin digest samples. Two receiver solutions, Ringer's saline, and Hanks' HEPES buffered saline with 4% serum albumin were used. Listed in decreasing order, the total percentage in vitro dermal absorptions obtained by 48 hr after exposure for the five skin types were: 47 ± 4.3% [tissue cultured Testskin; Hanks' receiver (HR)], 40 ± 4.5% (rat; HR), 19 ± 1.8% (human; HR), 14 ± 2.3% (hairless guinea pig; HR), 14 ± 8.8% (pig; Ringer's receiver). The percentage recovery of the radiolabel in soapy water skin washes at 24 hr, methanol washes and skin digests at 48 hr, and of ¹⁴C-labelled volatiles collected in air traps at 48 hr after exposure are reported. Comparative in vivo studies were conducted for 14 days after exposure and demonstrated 32 ± 3.9 and 28 ± 7.8% recovery of ¹⁴C in the urine of rats (dose rate, 3 μg/cm²) and guinea pigs (dose rate, 4 μg/cm²), respectively. Total faecal recovery was 2 ± 0.3 and 9 ± 3.5% for rats and guinea pigs, respectively. Analysis of tissue taken at autopsy 14 day after dosing demonstrated a total tissue recovery of ¹⁴C activity of 1 ± 0.1 and 2 ± 0.5% in rats and guinea pigs, respectively. Including the ¹⁴C activity extracted from the skin removed from the dose site at 14 days after exposure, the total recovery of dermally absorbed residues was 49 ± 10.4 and 40 ± 9.9% in rats and guinea pigs, respectively. Recovery of ¹⁴C activity from soapy water skin washes conducted at 24 hr after exposure was 28 ± 8.1 and 43 ± 9.0% for rats and guinea pigs, respectively. Recovery in skin patches was 18% (guinea pigs) and 26% (rats). In summary, the in vitro/in vivo concordance for the rat dermal absorption data was good but the in vitro data for hairless guinea pigs underestimated the in vivo absorption, and therefore for 2,4-D, rat skin may provide a better model of percutaneous absorption.     

PENETRATION OF CHLOROFORM,TRICHLOROETHYLENE, AND TETRACHLOROETHYLENE THROUGH HUMAN SKIN

In vitro dermal absorption was measured for three volatile organic compounds in dilute aqueous solution through freshly prepared and previously frozen human skin. The permeability coefficients at 26 C for chloroform (0.14 cm/h) and trichloroethylene (0.12 cm/h) were similar but much larger than that for tetrachloroethylene (0.018 cm/h). Storage of the skin at-20 C did not significantly affect the penetration of these chemicals. The dermal absorption of chloroform through freshly prepared human skin was not changed significantly by pretreatment of the skin with commonly used consumer products (moisturizer, baby oil, insect repellent, sunscreen); however, the permeability coefficient was found to increase from 0.071 cm/h at 11 C to 0.19 cm/h at 50 C. These data suggest that exposure estimates for chloroform and other contaminants in water should consider the appropriate exposure scenario to properly assess the dermal dose.     

Dermal absorption of chemicals: effect of application of chemicals as a solid, aqueous paste, suspension, or in volatile vehicle.

The purpose of this study was to investigate the dermal absorption of chemicals in different physical forms when applied to female F344 rats. Chemicals were applied either as a solid, aqueous paste, suspension, or dissolved in the volatile vehicle ethanol. The chemicals investigated were [14C]-2-sec-butyl-4,6-dinitrophenol (DNBP, 4.2 mumol), 2,4,5,2',4',5'-[14C]-hexachlorobiphenyl (HCB, 2.3 mumol), and 3,4,3',4'-[14C]-tetrachlorobiphenyl (TCB, 0.5 mumol). The chemicals were applied on the clipped mid-dorsal region of the rat over a 2.54-cm2 treatment area, which was then occluded. Urine and feces were collected and assayed for radioactivity. Twenty-four hours post-application, the treated skin was washed with a mixture (1:1) of soap and water, dried, and reoccluded. The animals were sacrificed at 120 h by exsanguination under ether anesthesia. Radioactivity in the blood, skin (treated and untreated), and carcass was assayed. Dermal absorption of DNBP-derived radioactivity was approximately 50% of the recovered dose after application in the four physical forms, and the major route of excretion was via the urine. Twelve percent of the absorbed dose of DNBP was retained in the body. Dermal penetration of HCB-derived radioactivity was 5-8% of the recovered dose after application in the four forms, and the major route of excretion was via the feces. Greater than 90% of the absorbed dose of HCB-derived radioactivity was retained in the body. Dermal penetration of TCB-derived radioactivity was 6-8% of the recovered dose in the four forms, and the major route of excretion was via the feces. Approximately 21% of the absorbed dose was retained in the body at 120 h. Absorption of each chemical applied either as solid, aqueous paste, or suspension was compared to the absorption of the same chemical in ethanol. Absorption of HCB applied as a solid was significantly higher (p less than or equal to .05) as compared to HCB applied in ethanol. There were no other significantly differences in the comparisons of absorption. The data indicate that the chemicals examined in this study can penetrate the skin as readily when applied either as a solid, aqueous paste, or suspension, as when applied in the volatile vehicle ethanol.     

In vivo percutaneous absorption,skin barrier perturbation,and irritation from JP-8 jet fuel components

JP-8 jet fuel has been reported to cause systemic and dermal toxicities in animal models and humans. There is a great potential for human exposure to JP-8. In this study, we determined percutaneous absorption and dermal toxicity of three components of JP-8 (i.e., xylene, heptane, and hexadecane) in vivo in weanling pigs. In vivo percutaneous absorption results suggest a greater absorption of hexadecane (0.43%) than xylene (0.17%) or heptane (0.14%) of the applied dose after 30 min exposure. Transepidermal water loss (TEWL) provides a robust method for assessing damage to the stratum corneum. Heptane showed greater increase in TEWL than the other two chemicals. No significant (p < 0.05) increase in temperature was observed at the chemically treated site than the control site. Heptane showed greater TEWL values and erythema score than other two chemicals (xylene and hexadecane). We did not observe any skin reactions or edema from these chemicals. Erythema was completely resolved after 24 h of the patch removal in case of xylene and hexadecane.     

The within- and between-laboratories reproducibility and predictive capacity of Amino acid Derivative Reactivity Assay using 4 mM test chemical solution: Results of ring study implemented at five participating laboratories

Amino acid derivative reactivity assay (ADRA) for skin sensitization was adopted as an alternative method in the 2019 OECD Guideline for the Testing of Chemicals (OECD TG 442C). The molar ratio of the nucleophilic reagent to the test chemicals in the reaction solution was set to 1:50. Imamura et al. reported that changing this molar ratio from 1:50 to 1:200 reduced in false negatives and improved prediction accuracy. Hence, a ring study using ADRA with 4 mM of a test chemical solution (ADRA, 4 mM) was conducted at five different laboratories to verify within- and between-laboratory reproducibilities (WLR and BLR, respectively). In this study, we investigated the WLR and BLR using 14 test chemicals grouped into three classes: (1) eight proficiency substances, (2) four test chemicals that showed false negatives in the ADRA with 1 mM test chemical solution (ADRA, 1 mM), but correctly positive in ADRA (4 mM), and (3) current positive control (phenylacetaldehyde) and a new additional positive control (squaric acid diethyl ester). The results showed 100% reproducibility and 100% accuracy for skin sensitization. Hence, it is clear that the ADRA (4 mM) is an excellent test method in contrast to the currently used ADRA (1 mM). We plan to resubmit the ADRA (4 mM) test method to the OECD Test Guideline Group in the near future so that OECD TG 442C could be revised for the convenience and benefit of many ADRA users.     

In vitro dermal absorption rate testing of certain chemicals of interest to the Occupational Safety and Health Administration: summary and evaluation of USEPA's mandated testing

In 2004, the United States Environmental Protection Agency (USEPA) published a final test rule in the US Federal Register requiring in vitro dermal penetration rate testing for selected industrial chemicals. The test rule described procedures for determining a permeability coefficient (Kp) and two short-term dermal absorption rates at 10 and 60min using human cadaver skin mounted in an in vitro diffusion cell model. According to the USEPA announcement, the selected chemicals were to be spiked with their radiolabeled form and tested in either water, isopropyl myristate (IPM) or neat depending on their physical character at room temperature, their aqueous solubility, their potential to damage the skin and their ability to achieve the study endpoints as prescribed. Overall, and for the majority of chemicals, the short-term absorption rates were higher at 10min than at 60min and the portion of applied dose remaining in the skin at the end of the exposure period was greater than the portion of dose that had penetrated through the skin and was detected in the receptor solution. In contrast to this, the amount of chemical in the receptor solution at study termination for the Kp (steady-state) experiments was greater than the amount remaining in the skin. For the Kp experiments, which lasted from 2 to 48h, a majority of skins exposed to neat chemical exhibited a reduced barrier function. However, integrity was mostly unaltered for skins from the short-term experiments and Kp experiments using chemicals applied either in water or IPM. Quantitative structure activity relationship (QSAR) model-predicted Kp values were in fair agreement with experimental data for those chemicals that were applied in a water vehicle when the integrity of the skin was not compromised. However, QSAR-derived Kp values were not predictive for those chemicals when applied in IPM vehicle or neat. Absorption predictions, based on the measured Kp and steady-state flux data for chemicals applied in water or neat, respectively, were comparable to measured values at both 10 and 60min. Kp data for chemicals applied in water and the flux values for neat chemicals will be useful for making estimates of skin absorption in occupational settings. Kp measurements for chemicals applied in IPM vehicle are not envisioned to provide useful data for estimating the risk from dermal exposure to chemicals in the workplace. When available, in vitro dermal flux measurements should be combined with toxicity information in order to improve the utility of chemical skin notations.     

Penetration of chloroform, trichloroethylene, and tetrachloroethylene through human skin.

In vitro dermal absorption was measured for three volatile organic compounds in dilute aqueous solution through freshly prepared and previously frozen human skin. The permeability coefficients at 26 degrees C for chloroform (0.14 cm/h) and trichloroethylene (0.12 cm/h) were similar but much larger than that for tetrachloroethylene (0.018 cm/h). Storage of the skin at -20 degrees C did not significantly affect the penetration of these chemicals. The dermal absorption of chloroform through freshly prepared human skin was not changed significantly by pretreatment of the skin with commonly used consumer products (moisturizer, baby oil, insect repellent, sunscreen); however, the permeability coefficient was found to increase from 0.071 cm/h at 11 degrees C to 0.19 cm/h at 50 degrees C. These data suggest that exposure estimates for chloroform and other contaminants in water should consider the appropriate exposure scenario to properly assess the dermal dose.     

In Vivo Percutaneous Absorption,Skin Barrier Perturbation,and Irritation from JP-8 Jet Fuel Components

Abstract

JP-8 jet fuel has been reported to cause systemic and dermal toxicities in animal models and humans. There is a great potential for human exposure to JP-8. In this study, we determined percutaneous absorption and dermal toxicity of three components of JP-8 (i.e., xylene, heptane, and hexadecane) in vivo in weanling pigs. In vivo percutaneous absorption results suggest a greater absorption of hexadecane (0.43%) than xylene (0.17%) or heptane (0.14%) of the applied dose after 30 min exposure. Transepidermal water loss (TEWL) provides a robust method for assessing damage to the stratum corneum. Heptane showed greater increase in TEWL than the other two chemicals. No significant (p<0.05) increase in temperature was observed at the chemically treated site than the control site. Heptane showed greater TEWL values and erythema score than other two chemicals (xylene and hexadecane). We did not observe any skin reactions or edema from these chemicals. Erythema was completely resolved after 24 h of the patch removal in case of xylene and hexadecane.     

Comparative dermal absorption of 2,3,7,8-tetrachlorodibenzo-p-dioxin and three polychlorinated dibenzofurans

Polychlorinated dibenzodioxins (PCDDs) and dibenzofurans (PCDFs) are toxic environmental contaminants which have the potential to accumulate in human tissues. In order to examine the potential for systemic exposure following dermal exposure, the absorption, distribution, and elimination of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,7,8-tetrachlorodibenzofuran (TCDF), 1,2,3,7,8-pentachlorodibenzofuran (1PeCDF), and 2,3,4,7,8-pentachlorodibenzofuran (4PeCDF) were evaluated in male F344 rats. TCDD (0.00015, 0.001, 0.01, 0.1, 0.5, and 1.0 mumol/kg) and the three PCDFs (0.1, 0.5, and 1.0 mumol/kg) were applied to a preclipped region on the back of the rat and covered with a perforated cap. The rats were held in individual metabolism cages for 3 days. In animals administered 0.1 mumol/kg, the absorption of TCDF was greater than that of 4PeCDF, 1PeCDF, and TCDD. Relative absorption (percentage of administered dose) declined with increasing dose while the absolute absorption (microgram/kg) increased nonlinearly with dose. Absorption of TCDF at 0.1 mumol/kg was 48% of the administered dose which was significantly greater than that of the other compounds. At this dose, absorption of 4PeCDF was greater than that of TCDD. Absorption at the higher doses was similar for all four compounds. Maximum relative absorption of TCDD (approximately 40% of the administered dose) was obtained at 0.001 and 0.00015 mumol/kg. Major tissue depots for these four chemicals included liver, adipose, skin, and muscle tissue; however, the liver:fat ratio for 4PeCDF was approximately fourfold higher than that for the other three compounds. When normalized to 100% of dose absorbed, the distribution of 4PeCDF-derived radioactivity in liver and adipose tissue was similar to that previously observed after oral and iv administration. In animals administered 0.1 mumol TCDF or 1PeCDF/kg, 56 and 32% of the respective absorbed dose was excreted as polar metabolites within 3 days. Very little of the absorbed dose of either TCDD (approximately 10%) or 4PeCDF (approximately 2%) was eliminated. Results indicate that the dermal absorption of these compounds is incomplete and that systemic toxicity following acute dermal exposure to levels found in the environment is unlikely.     

Absorption of lawsone through human skin

Lawsone (2-hydroxy-1,4-naphthoquinone) is the principal color ingredient in henna, a color additive approved with limitations for coloring hair by the Food and Drug Administration (FDA) under 21 CFR 73.2190. In 2002, the scientific committee on cosmetics and non-food products (SCCNFP), now known as the scientific committee for consumer products (SCCP), evaluated the safety of lawsone as a coloring agent in hair dye products of the European Union (EU). The SCCNFP concluded that lawsone was mutagenic and not suitable for use as a hair coloring agent. As a result, studies were conducted to measure the extent of lawsone absorption through human skin. Lawsone skin absorption was determined from two hair coloring products and two shampoo products, all containing henna. [(14)C]-Lawsone (sp. act. 22.9 mCi/mmol) was added to each commercial product and the products were applied to dermatomed, nonviable human skin mounted in flow-through diffusion cells perfused with a physiological buffer (HEPES-buffered Hanks' balanced salt solution, pH 7.4). Products remained on the skin for 5 minutes (shampoos) and 1 hour (hair color paste). For the henna hair paste products, 0.3 and 1.3% of the applied dose was absorbed into the receptor fluid in 24 hours while 2.2 and 4.0% remained in the skin. For both henna shampoo products, 0.3% of the applied dose was absorbed into the receptor fluid at 24 hours while 3.6 and 6.8% remained in the skin. For all products, most of the lawsone applied was washed from the surface of the skin (83-102%) at the end of the exposure period. Extended absorption studies were conducted for 72 hours to determine if skin levels of lawsone in the 24 hour studies might eventually be percutaneously absorbed. These studies determined that the majority of the lawsone remained in the skin with only a small but significant increase (for three out of four products) in receptor fluid values. Therefore, it appears that receptor fluid values would give a good estimate of lawsone absorption for an exposure estimate and that skin levels of lawsone need not be included.     

In vitro dermal absorption of pesticides: IV. In vivo and in vitro comparison with the organophosphorus insecticide diazinon in rat, guinea pig, pig, human and tissue-cultured skin

In vitro skin absorption tests are currently being developed as an alternative to in vivo animal tests for predicting the degree of occupational exposure to pesticides. In the study reported here, in vitro percutaneous absorption tests were conducted with the ¹⁴C-ring-labelled pesticide, diazinon, dissolved in acetone and applied to the dermatomed skin (0.5 mm) of a number of species at a dose rate of 9.5–16.7 μg/cm². Skin permeation was determined for 48 hr after exposure using an in vitro flow-through system. Skin permeation was calculated from the sum of the percentage recovery of ¹⁴C activity in the receiver solution and the percentage recovery obtained in methanol washes of the skin at 48 hr and in skin digests. Listed in decreasing order, the total percentage in vitro dermal absorptions (mean ± SD) obtained by 48 hr after exposure for the five skin types were: 47 ± 3.4% (rat), 36 ± 0.9% (tissue cultured Testskin), 33 ± 2.8% (hairless guinea pig), 20 ± 3.1% (human) and 15 ± 13.1% (pig). The percentage recoveries in soapy water skin washes at 24 hr, in methanol washes and skin digests at 48 hr and of ¹⁴C-labelled volatiles collected in air traps at 48 hr after exposure are reported. Comparative in vivo studies demonstrated 37 ± 0.8 and 24 ± 5.7% recovery of ¹⁴C in the urine of rats (dose rate, 6 μg/cm²) and hairless guinea pigs (dose rate, 5 μg/cm²), respectively, by 14 days after exposure. Total faecal recovery 14 days after exposure was 18 ± 0.4 and 4 ± 0.9% for rats and guinea pigs, respectively. Analysis of tissue taken at autopsy 14 days after exposure demonstrated a total tissue recovery of 0.6 ± 0.1% [¹⁴C]diazinon in rats and 1 ± 0.2% in hairless guinea pigs. The total recovery in skin removed from the dose site at 14 days after exposure was 0.2 ± 0.02% and 0.1 ± 0.05% in rats and hairless guinea pigs, respectively. Recovery of radioactivity from soapy water skin washes conducted at 24 hr after exposure was 21 ± 3.8% for rats and 2 ± 0.1% for hairless guinea pigs. Recovery in skin patches was 23 ± 5.4% and 73 ± 2.9% in rats and hairless guinea pigs, respectively. The in vitro data for dermal absorption of [¹⁴C]diazinon for rats (47 ± 3.4%) and hairless guinea pigs (33 ± 2.8%) were in good agreement with the data observed for rats (56 ± 1.03%) and hairless guinea pigs (28 ± 6.0%) in vivo. This study supported the use of in vitro skin absorption tests as an alternative to in vivo animal testing.     

In vitro and in vivo percutaneous absorption of catechol.

The Cosmetic Ingredient Review Expert Panel found insufficient data to conclude that catechol could be used safely in permanent hair dye products. Information was lacking on the extent of oxidation and skin absorption of remaining catechol. In vitro percutaneous absorption studies were conducted in human and rat skin using a consumer permanent hair dye spiked with 0.6% catechol. A 30-min application demonstrated 0.4% of the applied dose was absorbed through human skin and 0.2% through rat skin. The minimal absorption observed was due to the short exposure time and to partial oxidation of catechol by the dye developer. The fate of catechol remaining in rat skin after exposure in vitro and in vivo was investigated with additional absorption studies using catechol in ethanol. At 72 h, 24-h application of 4% catechol resulted in skin absorption of 81% of the applied dose in vitro and 53% in vivo. Skin levels measured at 24 h remained unchanged after 72 h. Therefore the skin reservoir did not contribute to the estimated systemic absorption. A deconvolution technique employed to predict skin absorption using plasma levels from intravenous and dermal administration overestimated in vivo skin absorption due to volatility of catechol in an ethanolic vehicle.     

A comparative 90-day toxicity study of allyl acetate, allyl alcohol and acrolein

Allyl acetate (AAC), allyl alcohol (AAL), and acrolein (ACR) are used in the manufacture of detergents, plastics, pharmaceuticals, and chemicals and as agricultural agents. A metabolic relationship exists between these chemicals in which allyl acetate is metabolized to allyl alcohol and subsequently to the highly reactive, alpha,beta-unsaturated aldehyde, acrolein. Due to the weaker reactivity of the protoxicants, allyl acetate and allyl alcohol, relative to acrolien we hypothesized the protoxicants would attain greater systemic exposure and therefore deliver higher doses of acrolein to the internal organs. By extension, the higher systemic exposure to acrolein we hypothesized should lead to more internal organ toxicity in the allyl acetate and allyl alcohol treated animals relative to those treated with acrolein. To address our hypothesis we compared the range of toxicities produced by all three chemicals in male and female Fischer 344/N rats and B6C3F1 mice exposed 5 days a week for 3 months by gavage in 0.5% methylcellulose. Rats (10/group) were dosed with 0-100mg/kg allyl acetate, 0-25mg/kg allyl alcohol, or 0-10mg/kg acrolein. Mice (10/group) were dosed with 0-125mg/kg allyl acetate, 0-50mg/kg allyl alcohol, or 0-20mg/kg acrolein. The highest dose of allyl acetate and acrolein decreased survival in both mice and rats. The primary target organ for the toxicity of all three chemicals in both species and sexes was the forestomach; squamous epithelial hyperplasia was observed following exposure to each chemical. In both species the highest allyl acetate dose group exhibited forestomach epithelium necrosis and hemorrhage and the highest dose of acrolein led to glandular stomach hemorrhage. Liver histopathology was the most apparent with allyl acetate, was also observed with allyl alcohol, but was not observed with acrolein. All chemicals had effects on the hematopoietic system with allyl acetate having the most pronounced effect. When dosed at quantities limited by toxicity, allyl acetate and allyl alcohol produce higher levels of urinary mercapturic acids than the minimally toxic dose of acrolein. This observation is likely due to biotransformation of allyl acetate and ally alcohol to acrolein that occurs after absorption and suggests that these chemicals are protoxicants that increase systemic exposure of acrolein. Increased systemic exposure to acrolein is likely responsible for the differences in hepatic toxicological profile observed with these chemicals.     

Enhanced systemic tissue distribution after dermal versus intravenous 3,3',4,4'-tetrachlorobiphenyl exposure: limited utility of radiolabel blood area under the curve and excretion data in dermal absorption calculations and tissue exposure assessment.

As a dioxin-like polychlorinated biphenyl (PCB), 3,3',4,4'-tetrachlorobiphenyl (TCB) is receiving increasing research and regulatory interest due to its high toxicity and persistence in the environment. (14)C-TCB was administered at an identical dose of 300 microg via the intravenous (iv) or dermal route to swine to examine the exposure route dependency of the relationship between tissue exposure and blood area under the curve (AUC) and the relationship between dermal absorption and excretion of radiolabel. After iv and dermal exposure, blood, urine, and feces samples were collected during the 11-day in vivo studies. At the end of the experiments, full mass balance studies were conducted to characterize tissue distribution of label. On average, over 70% of the applied dermal and iv doses were recovered. As expected, more than a 10-fold increase in blood AUC (0.49 vs 0.031, h x % dose/ml), plasma AUC (0.40 vs 0.038, h x % dose/ml), urine excretion (29 vs 2.3% of the applied dose), and fecal (30 vs 3.0% of the applied dose) excretion was determined after iv exposure compared to dermal exposure. However, we unexpectedly found that the tissue residue following iv exposure (8.0% of the applied dose) was only half that following dermal exposure (16% of the applied dose). Significantly larger (20- to 30-fold) ratios of blood AUC:tissue residue and excretion:tissue residue were observed after iv exposure compared to dermal exposure. This may indicate a route-related concentration-dependent blood-to-tissue partition process of pooled label, unique skin metabolism, or saturable hepatic metabolism of TCB. Thus, a long-term, low-input exposure pattern similar to this dermal exposure could be more harmful to systemic tissues than a short-term, high-dose exposure similar to this iv exposure. One should be aware that greater absorption, higher blood concentrations, greater blood and plasma AUCs, and greater excretion of label do not necessarily result in a greater overall tissue exposure and that some conventional approaches using label determination in blood and excreta without full mass balance studies may underestimate dermal absorption of chemicals similar to TCB.