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The influence of physicochemical properties on the internal dose of trihalomethanes in humans following a controlled showering exposure


Although disinfection of domestic water supply is crucial for protecting public health from waterborne diseases, this process forms potentially harmful by-products, such as trihalomethanes (THMs). We evaluated the influence of physicochemical properties of four THMs (chloroform, bromodichloromethane, dibromochloromethane, and bromoform) on the internal dose after showering. One hundred volunteers showered for 10 min in a controlled setting with fixed water flow, air flow, and temperature. We measured THMs in shower water, shower air, bathroom air, and blood samples collected at various time intervals. The geometric mean (GM) for total THM concentration in shower water was 96.2 μg/l. The GM of total THM in air increased from 5.8 μg/m3 pre shower to 351 μg/m3 during showering. Similarly, the GM of total-blood THM concentration increased from 16.5 ng/l pre shower to 299 ng/l at 10 min post shower. THM levels were significantly correlated between different matrices (e.g. dibromochloromethane levels) in water and air (r=0.941); blood and water (r=0.845); and blood and air (r=0.831). The slopes of best-fit lines for THM levels in water vs air and blood vs air increased with increasing partition coefficient of water/air and blood/air. The slope of the correlation plot of THM levels in water vs air decreased in a linear (r=0.995) fashion with increasing Henry's law constant. The physicochemical properties (volatility, partition coefficients, and Henry's law constant) are useful parameters for predicting THM movement between matrices and understanding THM exposure during showering.


Water disinfection is important to control infectious disease outbreaks such as typhoid, hepatitis, Giardia infection, and cholera arising from the public water supply. However, potentially harmful chemicals, such as trihalomethanes (THMs), can be formed as by-products. THMs are halogenated organic compounds formed during disinfection by the reaction of chlorine with naturally occurring organic matter such as humic and fulvic acids. Chloroform, bromodichloromethane (BDCM), dibromochloromethane (DBCM), and bromoform are the primary THMs found in tap water in the United States.1

Epidemiological studies suggest that exposure to THMs increases the risk of various adverse health outcomes in humans. Long-term exposure to THMs may lead to increased risk of cancers in the bladder, stomach, pancreas, kidney, and rectum.2, 3, 4 THM exposure may also increase the risk of Hodgkin's and non-Hodgkin's lymphoma.2, 3, 4, 5 Furthermore, exposure to disinfection by-products (DBP), including THMs, may be associated with adverse reproductive outcomes such as reduced gestational age, intrauterine growth retardation, and birth defects.6, 7 Although the mechanisms by which DBPs might cause adverse health effects are not well understood, a more accurate assessment of exposure will improve the precision of epidemiological studies by reducing exposure misclassification.

The presence of THMs in the domestic water supply leads to exposure to THMs from daily water-related activities,8, 9, 10, 11, 12, 13 such as drinking and bathing/showering. THMs in tap water are inhaled following aerosolization or vaporization during showering. In addition, THMs are absorbed dermally during showering and bathing. Because THMs are lipophilic and metabolized rapidly, the levels in blood may vary depending on the time of sample collection after exposure.14

Because of prevalent exposure to THMs from the use of tap water,15 the US Environmental Protection Agency (US EPA) established regulatory limits for THM levels in tap water. These regulatory changes appear to have resulted in decreased THM levels in both domestic water supply and in US residents using that water.16 Estimating human exposure to THMs is complicated owing to variation in routes of exposure (e.g. ingestion, inhalation, and dermal), absorption, distribution, metabolism, and excretion. Measuring metrics of internal dose (e.g. THM concentrations in blood) is therefore a useful approach for assessing recent human exposure to THMs.17 Internal dose of THMs is related to many factors, including tap water THM levels; duration and frequency of water use activity; route of exposure; genetics; metabolic factors; and THM physicochemical properties (e.g. volatility, partition coefficient, half-life in blood).

To better understand how physicochemical properties affect THM uptake during showering, we evaluated THM levels in shower water, shower air, and blood of 100 study participants who showered under controlled conditions (e.g. 10-min shower, shower temperature, shower water flow). A summary of our findings has been published.10 In this paper, we examine the relationships between THM physicochemical properties (Henry's law constant, volatility, and partition coefficients) and post-showering blood THM levels.


Human Subjects

The institutional review boards of the Centers for Disease Control and Prevention (CDC), the National Institutes of Health, the General Clinical Research Center (GCRC) at the University of Pittsburgh, and Battelle Memorial Institute approved this study protocol. We complied with all applicable requirements for protection of human subjects. Study participants gave written informed consent before the study.

Study Design and Participants

Pittsburgh was chosen for this study, because the tap water there typically contains measurable levels of the four DBPs of interest. Brominated DBPs can form during disinfection of water containing natural organic material and bromide. Significant levels of bromide are typically found in the Allegheny River in Western Pennsylvania, possibly because of a combination of natural and anthropogenic sources.18 The Allegheny River was the source for water for the utility that serviced the GCRC at the University of Pittsburgh, and thus initial tap water samples contained measurable levels of all four target THMs.

We measured THM levels in shower water, shower air, bathroom air, and blood of 100 study participants who showered under controlled conditions (10-min shower, 40 °C shower temperature, 5.6–6.7 l/min shower water flow). Briefly, we recruited adults from a panel who volunteered to participate in a research project at the GCRC and conducted the study in July–September 2004. Eligible subjects (18–45 years of age) had a normal blood screen, were not pregnant, and were willing to provide blood by venipuncture. Questionnaire data were collected by direct face-to-face interview on the day of the showering event. Occupation and data on recent THM exposure activities (bathing, swimming, using hot tub or sauna, washing clothes or dishes) were collected for each participant just before the exposure activities began. To limit the exposure to THMs unrelated to the showering, participants were asked not to flush the toilet or run tap water while in the study area, to dry off and dress within 5 min, and to stay in a separate room, away from the shower room, while waiting for the 30 min post-shower blood sample to be drawn. We provided study participants with THM-free bottled drinking water to be consumed while at the study site to minimize/eliminate oral THM exposure. Further details of this study design were reported by Backer et al.10

Biological Specimen

Whole-blood samples for THM analysis were collected by a certified phlebotomist using vacutainer tubes processed to remove THM contamination.19 A baseline blood sample was collected shortly before shower activity began. Blood samples were collected at 10 and 30 min after the end of the 10-min shower. Participants stayed in the shower room for 5 min after the shower while they were getting dressed. Therefore, study participants had additional THM exposure by inhalation during the 5 min post shower in the bathroom, and thus the total exposure period was actually 15 min. One can note that the blood samples were collected 5 and 25 min after the end of the total exposure period, which is the same as 10 and 30 min, respectively, after showering ended. It is important to note that both the 10- and 30-min post-shower samples were collected during the elimination phase for THMs, and thus the samples do not represent the peak blood concentrations but rather occur in the downslope of the blood concentration vs time curve.20 Samples were analyzed for THM levels using headspace solid-phase micro-extraction (SPME) coupled with gas chromatography (GC) and high-resolution mass spectrometry (MS). Analyte quantification was based on stable isotope dilution.21

Air Samples

We collected three air samples near the subject's breathing zone: a pre-exposure instantaneous sample collected in the bathroom (hereafter referred to as the “pre-shower” air samples), a 10-min time-integrated sample collected in the shower stall during the entire 10-min showering period (hereafter referred to as the “during shower” air samples), and a 5-min time-integrated post-shower sample collected in the unventilated bathroom, starting immediately after the shower ended (hereafter referred to as the “post-shower” air samples). Air samples were collected remotely using evacuated stainless steel canisters and a continuously flowing sampling line (copper tubing, ¼″ outer diameter). Filled canisters were sealed and shipped to Battelle (Columbus, OH, USA) for analysis. We analyzed the samples for THMs by automated GC/MS using a modified version of US Environmental Protection Agency method TO-14.22 Full details of air sampling and analysis procedures are available elsewhere.23

Water Samples

The shower head was modified to allow remote water sampling. Duplicate samples were collected 5 min after each shower began. Participants were instructed to set the shower water temperature between 40 and 41 °C. Shower water temperature was monitored by the participant via a digital thermometer in the shower stall, and remotely by the study staff via wireless transmission to a display outside the bathroom. Water samples were collected in borosilicate glass vials containing sodium thiosulfate to quench further THM formation and phosphate buffer to standardize pH between 6.0 and 6.5. We analyzed water samples for THM levels using headspace SPME–GC/MS with quantification based on stable isotope dilution.24

Statistical Analysis

Unweighted estimates (geometric mean, mean, median, SD, SE, and the range) were calculated using JMP (John's Macintosh Project) statistical software, version 8.0 (SAS Institute, Cary, NC, USA). Correlation analyses for distribution of THMs in water vs shower air, blood vs shower air, and blood vs water were performed using JMP. The slope and the Pearson's correlation coefficient (r) for linear regression for all THMs for 100 study participants were obtained using JMP.


We measured four THMs in water, air, and blood samples collected before, during, and after controlled showers for 100 participants. Distribution of THM levels (geometric mean and 95% confidence interval) in shower water collected 5 min after the shower began is presented in Table 1. The shower water THM geometric means across the full study period were as follows: chloroform (GM 64.1 μg/l); BDCM (GM 19.9 μg/l); DBCM (GM 8.3 μg/l); and bromoform (GM 0.760 μg/l) (Table 1). The geometric mean for total THMs was 96.2 μg/l (Table 1). We observed a decrease in brominated THM levels over the study period (July to September 2004) (Supplementary Figure S7). Relative percent differences between duplicate water samples for a subset (30 out of 100 participants) of showers were <0.65% for all THMs (Supplementary Table S1), indicating uniform sampling and precise THM measurement (Supplementary Figure S1).

Table 1 Geometric mean (±95% confidence interval (CI)) of THMs for tap water samples collected 5 min after shower began and air samples collected 10-min time-integrated during shower and 5-min time-integrated post-shower.

Results from the analysis of the during-shower and post-shower samples for THMs in air are presented in Table 1. The geometric mean of pre-shower air levels of total THMs was 5.8 μg/m3. Similar to water, chloroform was the most abundant THM (GM=243 μg/m3), followed by BDCM (GM=70.9 μg/m3), DBCM (GM=25.0 μg/m3), and bromoform (GM=2.67 μg/m3) for the samples collected during the shower. Post-shower air samples had higher total-THM levels (GM=367 μg/m3) than the during-shower air samples (GM=351 μg/m3).

For some participants, duplicate air samples were collected. Relative percent differences for post-shower duplicate air samples (8 out of 100) were <0.91%, for all THMs (Supplementary Table S1), indicating uniform sampling and precise THM measurement (Supplementary Figure S2). THM levels in air samples collected during and post showering are well correlated (Supplementary Figure S3). Relative percent differences for the duplicate integrated air samples (8 out of 100) during and post (7 out of 100) showering were <1.31% for all THMs (Supplementary Table S1).

Table 2 presents the distributions of blood THM levels for samples collected before shower, 10 min after shower, and 30 min after shower. The relative quantities of THMs in baseline blood followed the same pattern found in water and air: chloroform (GM=11.0 ng/l); BDCM (GM=2.25 ng/l); DBCM (GM=1.30 ng/l); and bromoform (GM=0.960 ng/l, Table 3). In the samples collected 10 min after shower, GM of total THM concentrations in blood was increased 18-fold (GM=299 ng/l) from GM of baseline levels. Blood total-THM levels subsequently decreased in samples collected 30 min after shower (GM=152 ng/l, Table 2). Similarly, compared with baseline, the GM of blood chloroform collected 10 min after the shower were increased 18-fold from 11.0 to 197 ng/l, GM of BDCM increased 29-fold from 2.25 to 64.8 ng/l, GM of DBCM increased 21-fold from 1.3 to 27.9 ng/l, and GM of bromoform increased 3.5-fold from 0.96 to 3.42 ng/l. The GM of blood levels of all THMs 30 min after shower were lower than in blood samples collected 10 min after the shower (Table 2).

Table 2 Geometric mean (±95% confidence interval) of THMs for blood samples collected before shower, 10 min after shower, and 30 min after shower for study participants.
Table 3 Simple linear regression parameters (slope and Pearson's correlation coefficient) for THM distribution between water/air, blood/air, and blood/water.

Post-shower blood levels of each of four THMs correlated well with THM levels in during-shower air samples (r>0.574, Table 3) and during-shower water samples (r>0.583, Table 3). Scatter plots for DBCM in blood, shower air, and water are presented in Figure 1 and other THMs in Supplementary Figures S4–S6 for blood samples collected 10 min after shower, and air and water samples collected during shower. Regression parameters (slope, SE for slope, and Pearson's correlation coefficient) for the simple linear regression analysis of all THMs in water vs shower air, blood vs air, and blood vs water are presented in Table 3.

Figure 1

Scatter plot for dibromochloromethane (DBCM) in whole blood, shower water, and shower air samples. Blood samples were collected 10 min after the shower, water samples were collected 5 min after shower began, and air samples were collected during the shower (10-min time-integrated).

The partitioning of each THM between water and shower air (regression slope of water vs air concentrations) remained nearly constant between samples collected during and after the shower (Table 3). Figure 2 demonstrates the relationship between Henry's law constants and the regression slopes for the water/air partition for each THM during the shower. Regression slopes decreased linearly with increasing Henry's law constants.

Figure 2

Relationship between partition of trihalomethanes (THMs) in shower water/shower air (regression slope of scatter plot) and Henry's law constant (at 25°C) in water for chloroform (CHCl3), bromodichloromethane (BDCM), dibromochloromethane (DBCM), bromoform (CHBr3).

Regression slopes of THMs in water/air and blood/air during the shower correlated well with the partition coefficient (K) of THMs (Figure 3). Regression slopes increased linearly with increasing partition coefficients in water/air (Kwater/air) and blood/air (Kblood/air).

Figure 3

Relationship between distribution of trihalomethanes (THMs) in water/air and blood/air (regression slope of scatter plot) and partition coefficient in water/air and blood/air for chloroform (CHCl3), bromodichloromethane (BDCM), dibromochloromethane (DBCM), and bromoform (CHBr3) during showering.


We observed significant exposure to THMs resulting from showering in chlorinated tap water. Volatility and partition coefficients of THMs influence the amount of THM transferred into the air during showering, and thus the amount of THM that can be inhaled by the person showering. THM levels in shower air decreased with decreasing volatility of THM and increasing water/air partition coefficient (Kwater/air) (Tables 1 and 4, and Figure 3). Of the four THMs we measured, chloroform has the lowest boiling point. The boiling point and Kwater/air of the THMs increases with increasing substitution of bromines in the THM molecule (Table 4), leading to decreased partitioning from shower water into shower air. Therefore, the ratio of geometric mean THM levels in shower water (μg/l) to shower air (μg/m3) increases from chloroform (0.26) to BDCM (0.28) to DBCM (0.33). Furthermore, the slope of the best-fit regression line for the paired water vs post-shower air THM level increases with increasing bromine content (chloroform, slope=0.174; BDCM, slope=0.219; DBCM, slope=0.321; bromoform, slope=0.376 (Table 3)). Thus, as THM volatility increases and Kwater/air decreases, partitioning into air during showering increases and likely leads to larger relative contributions of inhalation compared with dermal exposure. Dermal exposure likely remains an important exposure route for all four THMs,25 with skin-permeability coefficient increasing26 with increasing bromine content (Table 3).

Table 4 Relevant physicochemical properties (boiling point, Henry's law constant, and partition coefficient) of trihalomethanes in water, air, and human blood.

We observed a substantial increase in all THM blood levels at 10 min post shower compared with pre-shower levels (Table 2). This significant increase in blood THMs after showering is consistent with the published literature; showering in THM-containing water leads to a substantial increase in THM levels in blood.8, 9, 10, 27 THM uptake from tap water into blood is affected by shower water temperature.28 For this reason shower water temperature was held at 40.7 °C±0.9 °C, thus allowing us to focus on the contribution of physicochemical properties in modulation of THM exposure. Study participants showered in 40 °C water; however, we found no published work on Henry's law constants measured at that temperature. Therefore, we related THM partitioning in our study to Henry's law constants that were previously measured at 25 °C (Figure 2). These empirically determined Henry's law constants were used rather than constants predicted from lower temperature measurements,29 because of uncertainties introduced by extrapolating from data collected at much lower temperatures. As shown in Figure 2, Henry's law constants were linearly related with the regression slopes for the water/air partition for each THM; regression slopes decreased linearly with increasing Henry's law constants. Thus, we found the expected relationship between published work on Henry's law constants and empirically measured THM levels in shower air and water. This finding indicates that our measurements were well controlled. As an additional test of this relationship, predicted Henry's law constants at 40 °C were also linearly related with the regression slopes for THM water/air partitioning (Supplementary Figure S9).

For all THMs studied, the levels correlated well among water, air, and blood. Scatter plots of THMs (Figure 1 and Supplementary Figures S4–S6) graphically display correlations of each THM in water, shower air, and post-shower blood. For all four THMs, the post-shower air levels increased linearly with the water levels (all Pearson's correlation coefficients >0.808, Table 3). The tight correlations observed likely result from tightly controlling variables that impact transfer of THMs from water to air (e.g. water temperature, shower spray parameters, showering time, and shower stall ventilation), leading to reproducible partitioning of THMs from water to air. The consistency of the air THM data also indicates complete mixing of air between shower stall and bathroom over the course of exposure period. The minimal variability in the experimental system is also supported by the high degree of correlation between during- and post-shower air samples (Supplementary Figure S3). In Table 3, THM slopes increased for water/air, blood/air, and blood/water plots with increasing bromine content (i.e., chloroform<BDCM<DBCM<bromoform); the same trend was observed for the Pearson's correlation, with the exception of the BDCM water–air correlation. These trends are likely influenced by the physical and chemical properties of the THMs: increasing the bromine content leads to decreased volatility and increased lipophilicity, which result in increased skin permeability and greater dermal absorption, and thus increased water/air, blood/air, and water/blood slopes and Pearson's correlations.

Although blood THM concentrations increased linearly with THM levels in water and shower air, correlations between THM levels in water and air were stronger than correlations of THM levels between blood and either shower air or water. The stronger correlation between THM levels in water and air matrices likely results from a variety of factors. Shower water and air in this study were tightly controlled to minimize variability in temperature and ventilation. Conversely, THM levels in blood were affected by numerous factors that vary between study participants, including genetic and physiological variables (e.g. THM exhalation rate, lung capacity, metabolism, blood lipids, BMI, and cardiac parameters). Correlations were weaker for chloroform and BDCM compared with DBCM and bromoform, likely due to differences in volatility, partitioning characteristics, and exposure pathways. Specifically, dermal absorption likely becomes increasingly more important with increasing bromine substitution, and may be a less variable exposure route than inhalation. In summary, this study shows that exposure to chloroform, BDCM, DBCM, and bromoform does occur during showering, but the extent of exposure is influenced by physicochemical and physiological parameters.

Although each participant was exposed to chlorinated tap water under controlled conditions, the reported individual level of water, air, and blood THM levels varied widely. Total water THM levels during the study period ranged from 75.0 to 130 μg/l most likely owing to variation of THM concentration in the water distribution system. As a result, THM levels in shower air ranged from 184 to 542 μg/m3 (10-min time-integrated sample during shower), and 256–542 μg/m3 (5-min time-integrated sample post shower). Total blood THMs (10 min post shower) ranged from 47.5 to 615 ng/l in the study participants, due at least in part to differences in water and shower air THM levels during and after showering. An additional source of variability of blood THM levels is most likely due to interindividual differences in metabolism and physiology.10

The physicochemical properties of the THMs impacted the rate at which THMs were eliminated from the blood. From 10 min post shower to 30 min post shower, blood THM levels decreased. The percent decrease of THMs in blood from 10 min post shower to 30 min post shower was 49.6%, 49.7%, 46.2%, and 31.0% (calculated from GMs listed in Table 2) for chloroform, BDCM, DBCM, and bromoform, respectively. Compared with the other THMs, bromoform levels in blood decreased at a slower rate. THM elimination is predicted by physiological half-life (t1/2). The t1/2 of THMs is dependent on physical, chemical, and metabolic characteristics. Because of their high volatility, THMs may be eliminated via exhalation from the lungs as a function of Kblood/air. The greater lipid solubility of bromoform may lead to retention in poorly perfused adipose tissue and thus slower elimination than the less lipophilic THMs.30

We chose the study site (Pittsburgh, PA) because bromide present in the raw source water would lead to formation of brominated THMs during chlorination.31 Therefore, we expected to find measurable levels of brominated THMs in the shower water. The shower water of the first eight participants contained relatively high bromoform levels (>6 μg/l) and had a relatively high ratio of the sum of the molar concentrations of bromide in each individual THM species to the total concentration of four THMs (bromine incorporation factor (BIF)).31 Median BIF for shower water was 0.649 for these eight study participants. However, the BIF dropped substantially (Supplementary Figure S7) for the subsequent 92 study participants (median BIF=0.295). This reduction in BIF may have been caused by heavy rain in the area that diluted bromide levels in the source water. Blood BIFs in samples collected 10 and 30 min after the shower are presented in Supplementary Figure S8. BIFs of THMs in blood samples are highly correlated with BIF in water samples with a higher correlation coefficient (r=0.978) for samples collected 10 and 30 min after shower than for baseline samples (r=0.733), similar to the findings of Miles et al.31

We compared the ratio of median THM levels in blood to water in this study with results reported in other studies (Table 5), because THMs in water drive post-showering blood levels. Table 5 presents the ratio of 10-min post-shower blood levels to water levels for our study and three other studies. Previous studies evaluated showering exposures in a smaller number of subjects (N≤25) compared with the current study (N=100). Study parameters such as water temperature, duration of shower, and other activities related to exposure were controlled in all of these studies. The ratio of the chloroform levels in blood to water in the current study was 3.03, which agreed well with Backer et al.9 (4.29),27 high THM site (3.29), and Nuckols et al.12 (1.89–2.25). The ratio of blood to water levels of less volatile THMs (e.g. BDCM and DBCM) are in good agreement with Backer et al.9, 27 and Nuckols et al.12 (Table 5). In the present study, higher blood to water ratios were observed for the brominated THMs compared with chloroform.

Table 5 Comparison of ratios of THMs in 10 min post-shower blood to shower water for participants in four controlled showering studies.

In conclusion, exposure to THMs occurs during showering with chlorinated tap water. As expected, THM levels in post-shower blood are highly correlated with THM levels in water and air during showering. Under these controlled exposure conditions (temperature, duration, air circulation) THMs transferred from shower water into showering individuals are consistent with their physicochemical properties, most importantly, Henry's law constant, volatility, and partition coefficients. Therefore, physicochemical properties of THMs must be considered when predicting THM exposures in epidemiological studies where conditions cannot be controlled.


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We acknowledge Mitchell Smith for unconditional laboratory assistance, Stephen Stanfill for technical editing, and John Morrow for data analysis. The use of trade names and commercial sources is for identification purposes only and does not imply endorsement by the U.S. Department of Health and Human Services or the Centers for Disease Control and Prevention.

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Correspondence to Lalith K Silva.

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The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.

Supplementary Information accompanies the paper on the Journal of Exposure Science and Environmental Epidemiology website

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Supplementary Table S1 (DOC 28 kb)

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Silva, L., Backer, L., Ashley, D. et al. The influence of physicochemical properties on the internal dose of trihalomethanes in humans following a controlled showering exposure. J Expo Sci Environ Epidemiol 23, 39–45 (2013).

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  • trihalomethane exposure
  • water disinfection by-products
  • human blood
  • showering
  • partition coefficients
  • Henry's law constants

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