The stratum corneum is the skin’s primary barrier against the environment. Therefore, for our experiments, the dermis was removed when skin tissue was used for in vitro permeation studies. However, full thickness Kp values were also measured and compared with the Kp values of dermatomed skin. Whole skin studies are useful in situations where dermatoming or using other methods to separate the epidermis from the dermis is ineffective such as when course hair leaves holes in the epidermis when separated from the dermis.
Quantification of Kp and lag time
Six 12-hr steady-state experiments (3 dermatomed and 3 whole skin samples) were used to obtain Kp values and lag times for the HANs and CH. (). The Kp values for CAN, DCAN, BCAN and DBCN for dermatomed skin ranged from 0.099 to 0.167 cm/hr, and the value for whole skin ranged from 0.04 to 0.048 cm/hr. The Kp values for CH were 0.0039 cm/hr for dermatomed skin and 0.0017 cm/hr for whole skin. Only a range of Kp values could be determined for trichloroacetonitrile (TCAN), from 0.039 to 0.14 cm/hr using dermatomed skin and 0.0011 to 0.0072 cm/hr using whole skin. One caveat to applying these Kp values to calculating the dose from dermal contact is even though the procedures used are the accepted method for measuring skin permeability there may be a difference in the absolute permeability value between hydrated skin as used in the in-vitro experiments and non-hydrated skin that would result from baths and showers of typical durations. It is expected that rank order of absorption would be similar.
Permeability coefficients (cm/hr) and lag times of DBPs determined at steady-state conditions.
The donor concentrations used to calculate Kp were the average of the values measured immediately before and after the steady-state exposures. Significant TCAN donor concentration losses of 45–82% were observed from the initial concentration of 0.47±0.06 g/L, measured at the start of the experiments (). The percent difference between the pre- and post-concentration levels in the donor cells for the remaining compounds were within expected experimental variations of ± 20% ().
Chemical properties and donor concentration changes for the six steady-state experiments (3 dermatomed and 3 whole skin samples).
At steady-state conditions, both the flux and donor concentrations should remain constant to calculate Kp. If the donor concentration decreases, then a decreasing flux is expected. This is seen in for TCAN. Therefore, only a range of possible Kp values could be calculated using the initial and final donor concentrations (). A lag time could also not be determined for TCAN.
The decreasing flux over time through the skin for TCAN from a representative steady-state experiment using dermatomed skin.
The decreases in donor concentrations of TCAN is likely due to a degradation of TCAN over time as TCAN can be unstable at pH values outside of 4.5–5.5 and the donor solutions was maintained near neutral to slightly basic pH to reflect typical drinking and pool water conditions. Losses due to evaporation is unlikely since the same closed system was used for all HANs and no losses were observed for the other compounds
The minimum time for steady-state conditions to be reached has been suggested to be between 2.4 and 3 times the lag time (Crank, 1975
). For the HANs for dermatomed skin, the mean lag times were between 6 and 7 min implying that the upper end estimate where steady-state can be reached is between approximately 14 min (2.4 × (6 min- the lag)) and 21 min (3 ×(7 min- the lag)). The mean lag time calculated for CH for the dermatomed skin was not different from zero, but the standard deviation was ±12 min suggesting that the maximum time required to reach steady-state for CH is no greater than 36 min. All compounds measured reached steady-state for the dermatomed skin within an hour. Therefore, the data points from 1 to 12 hours were used to calculate Kp. The permeation rate of whole skin is much lower than dermatomed skin. Thus it takes longer for the compounds to reach steady-state. The lag times were much longer in whole skin with the mean lag times ranging from 90 to 157 minutes for HANs and 208 minutes for CH. The time it takes to reach steady-state for HANs was between 216 and 471 minutes and 500 minutes for CH for whole skin.
When all the data points were used for steady-state linear regression of cumulative dose (mg/cm2) vs. time, slight variations near the end of the experiments can drastically change the calculated lag times. This is the probable reason why the percent standard deviations in the whole skin were near 25% rather than the less than 10% calculated for the dermatomed skin. Very high variability may also be common when the lag times are very short as with CH in dermatomed skin. Data points from 0.167 to 1 hour were used to calculate the lag times from the dermatomed steady-state experiments as opposed to the 1 to 12 hour data points used for whole skin.
The stratum corneum (only about 10 to 40 µm thick) represents the most efficient barrier against hydrophilic compounds and the viable epidermis (about 100 µm thick), and the dermis underneath (about 10 to 40 µm thick) provides a barrier to lipophilic compounds (U.S. EPA, 1992
). It is therefore not surprising that whole skin with the epidermis and dermis intact has lower Kp values and longer lag time values for the moderately lipophilic compounds HANs and CH compared to Kp values from dermatomed skin (). This decrease in Kp is more noticeable in the more lipophilic haloacetonitriles than with chloral hydrate, which is the least lipophilic of the compounds studied. Most absorbed compounds are quickly taken up into the bloodstream through a capillary network above the dermis. Thus, using dermatomed skin, where the dermis layer was cut away, provides a better estimate of the dermal absorption in viable skin than using whole skin since in vivo
compounds do not need to transverse the dermis to enter the bloodstream.
Simulated Internal Dose Estimates for Haloacetonitriles and Chloral Hydrate
The monthly mean, median and range of ingestion doses across gender and for adults and children for the HANs and for CH based on the Monte Carlo Simulations are given in . Since a definitive Kp could not determined for TCAN it was not included in the simulations. It was assumed that all of the consumed drinking water was unfiltered tap water to provide an estimate of the maximum ingestion dose. The variation across compounds is a function of the different drinking water concentrations typically found for each compound in the distribution system. The potential mean, median and range of dermal doses for HANs and CH from the subset of the population that takes baths over a monthly period is given in . The variation in dermal dose across compounds is a function of both the water concentration and the lipophilic property of each compound. These calculations have the caveat that the Kp values calculated using the hydrated skin are applicable to exposures encountered during showering, bathing and swimming when the skin may not become fully hydrated. A sensitivity analysis of the Monte Carlo indicated that the most important input variables were the same across gender and age and for all compounds, which could be a function of using the actual concentration in the water delivered rather than changes in water concentration during use which would vary with volatility of the compounds. For ingestion, the key input variables were drinking water concentration followed by ingestion rate with a minor contribution by body weight. For bathing, the key variables were duration of bath and drinking water concentration. For swimming, the key variable was duration of swim with lag-time through the skin also important for chloral hydrate. It is likely that pool water concentration was not included since few concentrations were available in pool water so the actual range used under-represented the true range and for chloral hydrate and dichloroacetonitrile were single values, thus those could not be evaluated in the sensitivity analyses.
Estimated population drinking water ingestion dose distributions/month (from Monte Carlo simulations).
Estimated dermal dose distributions/month from bathing (from Monte Carlo simulations) and the ratio of dermal to ingestion dose.
The dermal doses are 0.39 to 0.747 times the monthly ingestion doses for the HANs and 0.0160 to 0.0195 times the ingestion dose for CH. The additional dermal dose for the subset of the population that swims in pools over a monthly period from pool water is given in . Data on pool water concentration was available for only DCAN, DBAN and CH. The DCAN and DBAN mean pool derived dermal doses are 0.304 to 2.25 times the monthly ingestion doses and for CH 0.192 to 0.245 times the monthly ingestion dose. Since the dermal doses are a function of the drinking water concentrations and the Kp of each DBP, the higher water concentration of CH, when compared to the HANs, does not result in a high dermal dose for CH due to its low Kp value (0.0039 (cm/hour)).
Estimated dermal pool dose distributions/month (from Monte Carlo simulations) and the ratio of dermal to ingestion dose.
The dermal bath and pool water exposures for the children were higher than that of the adults on a per weight basis. This is due to children having a higher surface area per kilogram body mass than adults. A smaller individual will have a higher dose of DBPs per kilogram. This is also the reason for higher dermal doses per body mass of women compared to men and higher doses per body mass for boys compared to girls since girls 5–11 years of age are on average larger than boys of the same age.
The dermal doses are a significant fraction of the ingestion doses for the HANs. For a population that routinely swims in chlorinated swimming pools, the dermal dose during swimming is an important contributor to the total dose of these DBPs because of the order of magnitude higher pool water concentrations compared to drinking water levels. For DCAN, the dermal dose while swimming is greater than the dose received from other dermal contributions or ingestion of chlorinated drinking water. Therefore, estimating dermal absorption of HANs and CH from water is important since dermal contact is a potentially significant route of exposure due to their high Kp values. In addition, Kp in vitro data is essential in estimating potential internal doses of in vivo dermal exposures.