CHAPTER 2HEALTH EFFECTS

2.1. INTRODUCTION

The primary purpose of this chapter is to provide public health officials, physicians, toxicologists, and other interested individuals and groups with an overall perspective on the toxicology of bromodichloromethane. It contains descriptions and evaluations of toxicological studies and epidemiological investigations and provides conclusions, where possible, on the relevance of toxicity and toxicokinetic data to public health. When available, mechanisms of action are discussed along with the health effects data; toxicokinetic mechanistic data are discussed in Section 3.1.

A glossary and list of acronyms, abbreviations, and symbols can be found at the end of this profile.

To help public health professionals and others address the needs of persons living or working near hazardous waste sites, the information in this section is organized by health effect. These data are discussed in terms of route of exposure (inhalation, oral, and dermal) and three exposure periods: acute (≤14 days), intermediate (15–364 days), and chronic (≥365 days).

As discussed in Appendix B, a literature search was conducted to identify relevant studies examining health effect endpoints. Figure 2-1 provides an overview of the database of studies in humans or experimental animals included in this chapter of the profile. These studies evaluate the potential health effects associated with inhalation, oral, or dermal exposure to bromodichloromethane, but may not be inclusive of the entire body of literature. A systematic review of the scientific evidence of the health effects associated with exposure to bromodichloromethane was also conducted; the results of this review are presented in Appendix C.

Summaries of the human observational studies are presented in Table 2-1. Animal inhalation studies are presented in Table 2-2 and Figure 2-2, and animal oral studies are presented in Table 2-3 and Figure 2-3; no dermal data were identified for bromodichloromethane.

Levels of significant exposure (LSEs) for each route and duration are presented in tables and illustrated in figures. The points in the figures showing no-observed-adverse-effect levels (NOAELs) or lowest-observed-adverse-effect levels (LOAELs) reflect the actual doses (levels of exposure) used in the studies. LOAELs have been classified into “less serious” or “serious” effects. “Serious” effects are those that evoke failure in a biological system and can lead to morbidity or mortality (e.g., acute respiratory distress or death). “Less serious” effects are those that are not expected to cause significant dysfunction or death, or those whose significance to the organism is not entirely clear. ATSDR acknowledges that a considerable amount of judgment may be required in establishing whether an endpoint should be classified as a NOAEL, “less serious” LOAEL, or “serious” LOAEL, and that in some cases, there will be insufficient data to decide whether the effect is indicative of significant dysfunction. However, the Agency has established guidelines and policies that are used to classify these endpoints. ATSDR believes that there is sufficient merit in this approach to warrant an attempt at distinguishing between “less serious” and “serious” effects. The distinction between “less serious” effects and “serious” effects is considered to be important because it helps the users of the profiles to identify levels of exposure at which major health effects start to appear. LOAELs or NOAELs should also help in determining whether or not the effects vary with dose and/or duration, and place into perspective the possible significance of these effects to human health. Levels of exposure associated with cancer (Cancer Effect Levels, CELs) of bromodichloromethane are indicated in Table 2-3 and Figure 2-3.

A User’s Guide has been provided at the end of this profile (see Appendix D). This guide should aid in the interpretation of the tables and figures for LSEs and MRLs.

The health effects of bromodichloromethane have been evaluated in epidemiological and laboratory animal studies. As illustrated in Figure 2-1, most of the health effects data come from oral exposure studies in animals. Animal data are available for each health effect category and exposure duration category. The most examined endpoints were body weight (approximately 70% of the animal studies examined this endpoint), hepatic (approximately 50%), and renal (approximately 50%). Only five animal studies evaluated toxicity following inhalation exposure and these studies examined a limited number of endpoints (body weight, hepatic, renal, ocular, and other noncancer). The small number of available observational epidemiological studies only examined hepatic, immunological, reproductive, developmental, and cancer endpoints. Although some epidemiological studies suggest associations between bromodichloromethane exposure and an adverse health outcome, most of the studies are cross-sectional in design and do not establish causality. The epidemiological studies used several biomarkers of exposure including levels of bromodichloromethane measured in municipal water, blood bromodichloromethane levels, and levels of bromodichloromethane in exhaled breath. These biomarkers assess recent exposure, particularly the blood and exhaled breath since bromodichloromethane is rapidly excreted; most studies did not evaluate historical exposures. Another limitation of the epidemiological studies is that they involve co-exposure to other disinfection byproducts, which have similar targets of toxicity. Most studies did not statistically adjust for co-exposure to other compounds (e.g., chloroform, dibromochloromethane); thus, it is difficult to evaluate whether the observed effect was related to bromodichloromethane exposure or total exposure to disinfection byproducts, including other trihalomethanes.

The human and animal studies suggest several sensitive targets of bromodichloromethane toxicity:

• Hepatic Endpoints: Hepatic effects are a presumed health effect for humans based on limited evidence in humans and strong evidence in mice following acute inhalation exposure and in rats and mice following acute, intermediate, and chronic oral exposure. The liver effects include increases in serum enzymes, increases in liver weight, hepatocellular degeneration, and bile duct damage.
• Developmental Endpoints. Developmental effects are a presumed health effect for humans based on strong evidence from acute and intermediate oral exposures in rats. The most sensitive developmental endpoint was full-litter resorption in rats acutely administered bromodichloromethane via gavage. Inconsistent results have been observed in epidemiology studies, with some studies finding decreases in birth weight and increased risk of small for gestational age, and other studies not finding developmental effects.
• Renal Endpoints: Renal effects are a suspected health effect for humans based on moderate evidence in rats and mice following inhalation and oral exposure. The main effect observed was renal tubular degeneration; high acute oral doses also reported increases in blood urea nitrogen, urinary glucose, and urinary protein levels.
• Immune Endpoints. Immunological effects are a suspected health effect for humans based on moderate evidence in rats following acute and intermediate oral exposure. Decreased immune responses to stimulants were observed in rats.
• Reproductive Endpoints. Data are inadequate to conclude whether reproductive effects will occur in humans. Inconsistent results have been observed in animal studies examining potential reproductive endpoints, with some studies reporting effects (alterations in reproductive hormone levels and decreases in sperm velocity) and others reporting no effects (no alterations in histopathology, no changes in sperm motility, and no alterations in fertility in a 2-generation rat study).
• Other Endpoints. Alterations in body weight and gastrointestinal, hematological, ocular, endocrine, and neurological effects have also been observed in inhalation and/or oral exposure studies in laboratory animals; however, these do not appear to be sensitive targets of bromodichloromethane toxicity.

2.2. DEATH

Deaths have been reported in laboratory animals following acute or intermediate inhalation exposure and acute, intermediate, and chronic oral exposure. Increases in mortality were observed in two strains of mice exposed to 30 ppm bromodichloromethane vapor for 1 week (Torti et al. 2001). Deaths were also observed at 30 ppm in a similar 3-week study, but only in one of the two mouse strains tested (Torti et al. 2001).

Oral LD₅₀ values of 916 and 969 mg/kg were calculated in male and female rats (Chu et al. 1980). Deaths were also noted in rats receiving a single dose of 600 mg/kg (NTP 1987), but not in rats dosed for 14 days with 600 mg/kg/day (NTP 1987). However, another study reported 33% mortality in rats administered 300 mg/kg/day for 5 days (Thornton-Manning et al. 1994). The differences between the two studies may be due to the gavage vehicle used, oil in the NTP study versus an aqueous solution in the Thornton-Manning study. In contrast to the lack of sex differences observed in rats, male mice appear to be more sensitive to the lethal effect of bromodichloromethane than female mice. LD₅₀ values of 450 and 900 mg/kg were calculated in males and female mice, respectively (Bowman et al. 1978). NTP (1987) reported 100% mortality in male mice administered 600 mg/kg once or 150 mg/kg/day for 14 days; in females, 40% mortality occurred at 600 mg/kg and no deaths occurred at 150 or 300 mg/kg/day in the repeated exposure study.

Most intermediate- and chronic-duration studies did not test lethal doses. NTP (1987) reported increases in mortality in male and female rats administered 300 mg/kg for 13 weeks. No deaths were observed in studies testing lower doses in rats or mice (Aida et al. 1989, 1992; Chu et al. 1982; Hooth et al. 2002; Lock et al. 2004; McDorman et al. 2003; NTP 2006) or in female mice administered 400 mg/kg (NTP 1987). No deaths were noted in rats administered ≤190 mg/kg/day for chronic durations (Aida et al. 1992; George et al. 2002; NTP 1987, 2006; Tumasonis et al. 1985). In mice, decreases in survival were observed in female mice administered 75 or 150 mg/kg for 2 years (NTP 1987); no deaths were observed in mice chronically exposed to lower doses (George et al. 2002; NTP 2006).

2.3. BODY WEIGHT

No human studies have evaluated the effect of bromodichloromethane exposure on body weights. In general, alterations in body weight do not appear to be a sensitive indicator of bromodichloromethane toxicity in laboratory animals. In C57BL/6 mice, inhalation exposure to ≥30 ppm for 1 week resulted in decreases in body weight gain (Torti et al. 2001); increases in mortality were also observed at these concentrations. No alterations in body weight gain were observed when the mice were exposed for 3 weeks or in another mouse strain exposed for 1 or 3 weeks (Torti et al. 2001).

Several acute-duration oral studies have reported decreases in body weight gain in rats administered doses ≥300 mg/kg (Chu et al. 1982; Lilly et al. 1996, 1997; NTP 1987; Thornton-Manning et al. 1994); other rat studies utilizing doses ≤400 mg/kg, did not find body weight alterations (Keegan et al. 1998; Lilly et al. 1994). Two mouse studies evaluated body weight, one found a significant decrease at 250 mg/kg (Munson et al. 1982), and the other reported no effect at 148 mg/kg (Condie et al. 1983). Several studies have reported decreases in maternal weight gain following acute-duration oral exposure to ≥25 mg/kg (Bielmeier et al. 2001; Narotsky et al. 1997; Ruddick et al. 1983).

In intermediate-duration oral studies, 12–30% decreases in body weight gain were observed in rats administered 138–180 mg/kg bromodichloromethane (Aida et al. 1989, 1992; NTP 1987). A 12–17% decrease was also observed in rats exposed to 38 mg/kg/day bromodichloromethane in drinking water; however, significant decreases in water consumption were also observed at this dose level and the decrease in body weight is likely to be secondary to the decreased water intake (NTP 2006). No alterations in body weight were observed in rats administered 35 or 45 mg/kg (Chu et al. 1982; Lock et al. 2004) or in mice administered 50–400 mg/kg (Lock et al. 2004; NTP 1987, 2006). Decreases in body weight were also observed in rats and mice following chronic-duration exposure to ≥90 mg/kg/day (Aida et al. 1992; NTP 1987; Tumasonis et al. 1985), but not at lower doses (George et al. 2002; NTP 2006).

2.4. RESPIRATORY

The respiratory tract has not been examined in the available inhalation exposure studies in mice (Torti et al. 2001). No respiratory effects have been reported in animal oral exposure studies (Aida et al. 1989, 1992; George et al. 2002; NTP 1987, 2006; Ruddick et al. 1983).

2.5. CARDIOVASCULAR

No human studies have evaluated the cardiotoxicity of bromodichloromethane. No histological alterations were observed in the hearts of rats and mice orally administered bromodichloromethane at doses as high as 200 mg/kg/day (Ruddick et al. 1983), 400 mg/kg/day (Aida et al. 1989, 1992; NTP 1987, 2006), or 138 mg/kg/day (Aida et al. 1992; George et al. 2002; NTP 1987, 2006) for acute-, intermediate-, or chronic-durations, respectively.

2.6. GASTROINTESTINAL

No human studies have evaluated the gastrointestinal toxicity of bromodichloromethane. No non-neoplastic alterations have been observed in the gastrointestinal tract in most acute- (Ruddick et al. 1983), intermediate- (Aida et al. 1989, 1992; Hooth et al. 2002; NTP 1987, 2006), or chronic-duration (Aida et al. 1992; George et al. 2002; NTP 1987, 2006) oral studies in rats and mice. The NOAEL values for each duration category are 200, 400, and 138 mg/kg/day, respectively. McDorman et al. (2003) found an increase in the number of Eker rats having aberrant crypt foci in the colon following a 10-month exposure to 6.5 or 48.0 mg/kg/day in bromodichloromethane in drinking water. However, there were no significant increases in the total number of aberrant crypt foci, mean per colon, total number of crypts with aberrant foci, or distribution of aberrant foci in the different regions of the colon. The investigators considered aberrant crypt foci to be a preneoplastic lesion.

2.7. HEMATOLOGICAL

No studies examining hematological indices in humans were identified. Erythrocyte counts and hematocrit were significantly reduced in male rats 14 days after administration of a single dose of ≥390 mg/kg, and hemoglobin was significantly reduced in males and females at ≥546 mg/kg (Chu et al. 1982). No other acute (Munson et al. 1982; Ruddick et al. 1983), intermediate (Aida et al. 1989, 1992; Chu et al. 1982; NTP 2006), or chronic (Aida et al. 1992) oral studies reported erythrocyte or hemoglobin alterations, although the doses tested were lower than those in the Chu et al. (1982) acute study. The only other hematological alteration observed was a decrease in fibrinogen levels in female mice administered 125 mg/kg/day and male and female mice administered 250 mg/kg/day for 14 days (Munson et al. 1982).

2.8. MUSCULOSKELETAL

No studies evaluated the potential of bromodichloromethane to induce musculoskeletal alterations in humans. No histological alterations were observed in skeletal muscle of pregnant rats administered 200 mg/kg/day bromodichloromethane on GDs 6–15 (Ruddick et al. 1983). No longer-term studies examining musculoskeletal endpoints were identified.

2.9. HEPATIC

Information on the hepatoxicity of bromodichloromethane in humans is limited to a study which utilized NHANES data and did not find an association between bromodichloromethane blood levels and aspartate aminotransferase levels (Burch et al. 2015); this study is described in greater detail in Table 2-1. Animal studies provide strong evidence of the hepatotoxicity of bromodichloromethane. Based on a systematic review of the human and animal data, it is concluded that the liver is a presumed target of bromodichloromethane in humans (see Appendix C for additional information). The available animal data for bromodichloromethane and animal studies for two related compounds (bromoform and dibromochloromethane) (ATSDR 2005) provide evidence that oral exposure to bromodichloromethane results in an accumulation of fat in the liver as evidenced by increases in liver weight, centrilobular swelling, vacuolization, and fatty degeneration. Bromodichloromethane also appears to damage the bile duct. The animal studies also demonstrate vehicle-specific differences in hepatotoxicity, with greater toxicity associated with oil vehicles than aqueous vehicles.

A single dose of bromodichloromethane administered via gavage resulted in liver damage at doses as low as 74 mg/kg (Condie et al. 1983). At this dose, centrilobular pallor was observed in mice. At ≥81.9 mg/kg, marked increases in alanine aminotransferase, aspartate aminotransferase, and sorbitol dehydrogenase were observed in rats and mice (Condie et al. 1983; Keegan et al. 1998; Lilly et al. 1994, 1996). At 400 mg/kg, mild centrilobular vacuolar degeneration and minimal centrilobular hepatocellular necrosis were observed in rats (Lilly et al. 1994, 1996). The toxicity of bromodichloromethane was greater when it was administered in a corn oil vehicle than when administered in an aqueous vehicle (Lilly et al. 1994). The magnitude of the increases in alanine aminotransferase and aspartate aminotransferase was greater for the corn oil vehicle, particularly 48 hours after administration when the enzyme levels were at least twice as high in the corn oil vehicle group compared to the aqueous vehicle group. Similarly, the incidences of hepatocellular necrosis 48 hours post-administration were 5/6 in the oil vehicle group and 2/6 in the aqueous vehicle group. Bromodichloromethane was more toxic following repeated acute exposure (5–14 days), with increases in alanine aminotransferase and aspartate aminotransferase observed at ≥250 mg/kg/day (Munson et al. 1982; Thornton-Manning et al. 1994) and centrilobular hepatocellular vacuolar degeneration at ≥150 mg/kg/day (Thornton-Manning et al. 1994).

Intermediate-duration studies have reported hepatic effects ranging from increases in liver weight to fatty degeneration. There is a considerable amount of overlap between the NOAEL and LOAEL values for hepatotoxicity between studies, which may be due to differences in study durations and/or administration route. Administration of bromodichloromethane via gavage with an oil vehicle resulted in vacuolization in rats exposed to ≥60 mg/kg/day for 1 month (Aida et al. 1989) and centrilobular degeneration in rats exposed to 300 mg/kg for 3 months (NTP 1987); the NOAELs for these studies were 20 and 150 mg/kg, respectively. Microencapsulating bromodichloromethane dissolved in oil and adding it to the diet resulted in hepatocellular vacuolization, swelling, and necrosis in rats exposed to 180 mg/kg/day for 1 month (Aida et al. 1989) and fatty degeneration in male rats exposed to ≥6.1 mg/kg/day for 6 months (LOAEL in females was 31.7 mg/kg/day) (Aida et al. 1992). Two studies administering bromodichloromethane in drinking water did not find increases in liver lesions at the highest doses tested, 45 mg/kg/day for 28 days (Chu et al. 1982) and 71 mg/kg/day for 22 days (NTP 2006). However, a third study identified a LOAEL of 35 mg/kg/day for centrilobular swelling in rats exposed to 35 mg/kg/day for 4 or 10 months (Hooth et al. 2002). The Aida et al. (1989) studies allow for a direct comparison between exposure routes since the gavage and dietary studies utilized the same rat strain (Wistar), dose levels, and exposure duration (1 month). The gavage study identified a lower LOAEL (60 mg/kg/day) for vacuolization than the dietary study (180 mg/kg/day). Enlarged hepatocytes with vacuolization were also observed in female mice administered via gavage ≥200 mg/kg bromodichloromethane in corn oil (NTP 1987); no liver effects were observed in a 13-week drinking water study in which mice were exposed to doses as high as 51 mg/kg/day (NTP 2006).

Eight studies have evaluated the chronic toxicity of bromodichloromethane in rats and mice (Aida et al. 1992; George et al. 2002; NTP 1987, 2006; Tumasonis et al. 1985). With the exception of the lifetime drinking water exposure study conducted by Tumasonis et al. (1985), the other studies involved a 2-year exposure to bromodichloromethane administered via gavage with a corn oil vehicle (NTP 1987), in drinking water (George et al. 2002; NTP 2006), or in the diet (Aida et al. 1992). The Aida et al. (1992) study identified the lowest LOAEL for hepatic effects; at ≥6.1 mg/kg/day, fatty degeneration was observed in the liver of male rats exposed for 12, 18, or 24 months; the LOAEL in the female rats was 31.7 mg/kg/day after 12 and 18 months and 8.0 mg/kg/day after 24 months of exposure. Fatty metamorphosis was observed in rats administered ≥50 mg/kg (lowest dose tested) for 2 years (NTP 1987). In drinking water studies, no histological alterations were observed in the liver of rats exposed to 56.3 mg/kg/day (George et al. 2002). NTP (2006) noted that minimal to mild liver inflammation of questionable significance was observed at 12 and 25 mg/kg/day; the biological relevance of the lesion was questioned since the lesion morphology is consistent with spontaneous inflammation observed in aging rats, which is considered to be due to bacterial showering from the intestinal tract. Tumasonis et al. (1985) reported an increase in the incidence of hepatic adenofibrosis in female rats exposed to 190 mg/kg/day. In a mouse gavage study, an increase in fatty metamorphosis was observed in males at 50 mg/kg/day, but no lesions were observed in females at 150 mg/kg (NTP 1987). A drinking water study by George et al. (2002) did not find liver effects at the highest dose tested (43.3 mg/kg/day).

In addition to the hepatocellular effects noted in rats and mice, intermediate- and chronic-duration exposure has resulted in damage to the bile duct. Bile duct proliferation and cholangiofibrosis was observed in rats exposed to 138.0 (males)/168.4 (females) mg/kg/day for 6, 12, 18, and 24 months (Aida et al. 1992) and mild bile duct hyperplasia was observed in rats administered 300 mg/kg for 13 weeks (NTP 1987).

There are limited data on the mechanisms of bromodichloromethane hepatotoxicity. The available data suggest that its toxicity is due to the production of reactive intermediates. As reported by Thornton-Manning et al. (1994), pretreatment of rats with the cytochrome P450 inhibitor, 1-aminobenzotriazole, significantly reduced the hepatic toxicity of bromodichloromethane and pre-treatment with acetone, a CYP2E1 inducer, greatly increased its toxicity. Additionally, pretreatment with the glutathione synthesis inhibitor butathione sulfoxime (BSO) increased bromodichloromethane’s toxicity (Gao et al. 1996). Similarly, adding glutathione to hepatic microsomes under anaerobic conditions decreased binding of [14C]bromodichloromethane to lipids (Gao et al. 1996). These data demonstrate a protective role of glutathione that is consistent with metabolism of bromodichloromethane to one or more reactive species.

2.10. RENAL

No studies have evaluated the renal toxicity of bromodichloromethane in humans. However, based on the available animal studies, the kidney is a suspected target in humans (see Appendix C for more information on the systematic review of these data).

In inhalation studies, renal tubular degeneration was observed in mice exposed to ≥10 ppm bromodichloromethane for 1 or 3 weeks (Torti et al. 2001); the NOAELs identified in these studies were 1 and 3 ppm, respectively. Increased incidence of nephrosis was also observed at 10 ppm in a 13-week study (Torti et al. 2001); the NOAEL was 3 ppm.

In single-dose oral studies, mild to marked renal tubule degeneration and minimal-to-moderate renal tubule necrosis were observed in rats following administration via gavage with corn oil or aqueous vehicles at 200 mg/kg (Lilly et al. 1996) and/or 400 mg/kg (Lilly et al. 1994). Increases in serum urea nitrogen, urinary glucose, and urinary protein levels were observed at 400 mg/kg/day (Lilly et al. 1994), and decreases in urinary pH and osmolarity were observed at ≥200 mg/kg (Lilly et al. 1994). Similarly, renal tubule degeneration and tubular regeneration were observed in rats administered ≥150 mg/kg/day for 5 days (Thornton-Manning et al. 1994), and tubular necrosis and increases in serum creatinine and urea nitrogen were observed at 300 mg/kg/day. The acute studies provide some suggestive evidence of species differences in that no renal effects have been observed in mice administered bromodichloromethane for 5 days at doses as high as 150 mg/kg/day (Thornton-Manning et al. 1994). Another study found intratubular mineralization and epithelial hyperplasia at 148 mg/kg/day in mice exposed for 14 days (Condie et al. 1983), but did not report tubular degeneration or regeneration.

Similar renal effects have been reported in rats and mice in intermediate- and chronic-duration studies. Degeneration of proximal tubular epithelial cells were observed in rats administered 300 mg/kg for 13 weeks (NTP 1987) and nephrosis was observed in rats (females only) administered 100 mg/kg for 2 years (NTP 1987). Another rat study reported renal tubular cell hyperplasia in rats exposed to 36.3 mg/kg/day for 2 years (George et al. 2002). Other intermediate and chronic studies did not find histological alterations in the kidneys at doses as high as 180 mg/kg/day (Aida et al. 1989, 1992; Chu et al. 1982; Lipsky et al. 1993; Lock et al. 2004; NTP 2006). As noted in NTP (2006), differences in the route of administrations (gavage versus feed versus water) and stability of the bromodichloromethane in water and feed may have accounted for the overlap between the NOAEL and LOAEL values. In mice, proximal tubular focal necrosis was observed in males administered 100 mg/kg for 13 weeks (NTP 1987), but no effects were observed in females at doses as high as 400 mg/kg. An increase in the incidence of renal tubular epithelial cell cytomegaly was also observed in mice at 25 mg/kg for 2 years (NTP 1987). No renal effects were observed in mice administered via gavage 50 mg/kg/day for 4 weeks (Lock et al. 2004) or exposed to 36 mg/kg/day in drinking water for 2 years (NTP 2006).

2.11. DERMAL

No human or animal studies have evaluated the dermal toxicity of bromodichloromethane.

2.12. OCULAR

No human studies examined potential ocular effects following inhalation, oral, or direct contact exposure to bromodichloromethane. Mild eye irritation was noted in mice exposed to ≥30 ppm bromodichloromethane vapors for 1 week (Torti et al. 2001); the investigators did not report incidence data. Eye irritation was not noted in a 3-week study conducted by this group.

2.13. ENDOCRINE

In general, endocrine tissues do not seem to be a target of bromodichloromethane toxicity; see Section 2.16 for a discussion of alterations in reproductive hormone levels. Human studies evaluating endocrine endpoints following exposure to bromodichloromethane were not identified. No histological alterations were observed in rats following exposure to ≤200 mg/kg/day on GDs 6–15 (Ruddick et al. 1983), intermediate-duration exposure of rats to ≤300 mg/kg/day (Aida et al. 1989, 1992; NTP 1987, 2006) or mice to ≤400 mg/kg/day (NTP 1987, 2006), or chronic-duration exposure of rats to ≤138 mg/kg/day (Aida et al. 1992; NTP 1987, 2006). Thyroid follicular cell hyperplasia was observed in male mice administered via gavage 50 mg/kg or in females administered ≥75 mg/kg (NTP 1987); no endocrine effects were observed in mice exposed to ≤36 mg/kg/day in drinking water (NTP 2006).

2.14. IMMUNOLOGICAL

Immunotoxicity is a suspected health effect for humans based on a systematic review of several studies examining immunological endpoints in laboratory animals orally exposed to bromodichloromethane (see Appendix C for more information). Epidemiological data are limited to a study examining immune markers following a 40-minute swim in a chlorinated pool (Vlaanderen et al. 2017). Decreases in C-X-C motif chemokine 10, C-C motif chemokine 22, C-reactive protein, and vascular endothelial growth factor and increases in interleukin-1rA were associated with exhaled breath bromodichloromethane levels.

Acute exposures have resulted in decreased responses to humoral and cell-mediated immune stimulants in rats administered ≥75 mg/kg/day for 5 days (French et al. 1999) or mice administered 250 mg/kg/day for 14 days (Munson et al. 1982). Following a 26-week exposure to 49 mg/kg/day, an impaired response to the mitogen concanavalin A was observed in splenic lymphocytes, but there was no altered response in the lymph node lymphocytes or responses by either type of lymphocyte to other mitogens or to Salmonella tymphimurium (French et al. 1999).

The available data provide some suggestive evidence that rats may be more sensitive to the immunotoxic effects of bromodichloromethane than mice. No alterations in immune function were observed in mice exposed to 62 mg/kg/day in drinking water for 14 days (French et al. 1999) or administered 125 mg/kg/day via gavage with an aqueous vehicle for 14 days (Munson et al. 1982) or 250 mg/kg/day for 16 days (French et al. 1999). These NOAELs are higher than LOAEL values in rats. Although bromodichloromethane results in impaired immune function, no histological alterations were observed in lymphoid tissues following acute (Ruddick et al. 1983), intermediate (NTP 1987, 2006), or chronic (NTP 1987, 2006) exposure; the lymphoid atrophy observed at 300 mg/kg in the NTP (1987) intermediate-duration rat study was likely secondary to a decrease in body weight rather than a direct effect on the lymphoid tissue.

2.15. NEUROLOGICAL

No studies were located regarding neurological effects in humans exposed to bromodichloromethane. Balster and Borzelleca (1982) performed a series of tests in mice ≥24 hours after the last of a series of doses of bromodichloromethane. Exposure to doses of 1.2–11.6 mg/kg/day for 14–90 days had no effect on tests of coordination, strength, endurance, or exploratory activity, and 90-day exposure to 100 mg/kg/day did not affect passive avoidance learning. Exposure to 100 or 400 mg/kg/day for 60 days did result in an acute effect on operant behavior learning, but this change tended to diminish over the exposure period, suggesting that there was no progressive effect and that partial tolerance developed. One other study evaluating neurological function did not find alterations in performance on functional observational battery tests in rats exposed to 71.7 mg/kg/day for 6 months (Moser et al. 2007). No histological alterations in the brain and/or peripheral nerves were observed in rats or mice exposed to bromodichloromethane for acute (Ruddick et al. 1983), intermediate (Aida et al. 1992; Moser et al. 2007; NTP 1987, 2006), or chronic (Aida et al. 1992; NTP 1987, 2006) durations.

2.16. REPRODUCTIVE

In a systematic review of the available reproductive toxicity data for bromodichloromethane, it was determined that hazard identification for reproductive toxicity potential could not be classified due to the inconsistent results found in epidemiology and laboratory animal studies (see Appendix C for more information). A small number of human (Table 2-1) and laboratory animal (Table 2-2) studies evaluated the reproductive toxicity of bromodichloromethane; the studies examined potential effects on sperm parameters, menstrual cycle, fertility, hormone levels, and reproductive organ pathology. Three epidemiological studies examined reproductive endpoints associated with environmental exposure to bromodichloromethane. Interpretation of the study results is limited by the lack of confirming studies and potentially confounding exposure to other compounds, particularly other disinfection byproducts. Zeng et al. (2013) did not find a significant association between blood bromodichloromethane levels and sperm concentration, sperm count, or sperm motility in men. Associations between exposure to bromodichloromethane in drinking water and decreasing overall menstrual cycle length and follicular phase length specifically, as measured by urine estrogen and progesterone metabolite levels, were found in women participating in a reproductive health study (Windham et al. 2003). In a large prospective cohort study, a decreased time to pregnancy was associated with an estimate of the amount of bromodichloromethane ingested from tap water (MacLehose et al. 2008); however, no associations were found for other bromodichloromethane dose metrics. A fourth study examined possible interactions between CYP2E1, GSTZ1, and GSTT1 polymorphisms and bromodichloromethane levels in drinking water on sperm motility, sperm count, and sperm concentration (Yang et al. 2016). The only observed association was found in men with blood bromodichloromethane levels ≥1.70 µg/mL and a CYP2E1 rs2031920 CC polymorphism.

Most studies evaluating the histopathology of the testes and uterus did not find alterations (Aida et al. 1992; NTP 1987, 2006; Ruddick et al. 1983). One study did find mild to moderate atrophy of the seminal vesicles and/or prostate in rats administered a lethal dose of 300 mg/kg for 13 weeks (NTP 1987). A 2-generation reproduction study in rats did not find any alterations in reproductive parameters at the highest dose tested (51.7 mg/kg/day) (Christian et al. 2001b). No alterations in the percentage of motile or progressively motile sperm were observed in rats exposed to doses of 39 mg/kg/day for 52 weeks (Klinefelter et al. 1995); however, the study did find significant decreases in sperm velocity at 39 mg/kg/day. A study in rats found a diminished responsiveness to luteinizing hormone when 75 mg/kg bromodichloromethane was administered on GDs 8–10 (Bielmeier et al. 2001, 2004, 2007).

2.17. DEVELOPMENTAL

The available human and animal studies provide evidence that developmental toxicity is a presumed health effect of bromodichloromethane in humans (see Appendix C for information on the systematic review).

A number of epidemiology studies have examined the association between exposure to trihalomethanes, bromodichloromethane among them, and developmental effects in humans (Table 2-1). Specific endpoints examined have included birth weight and length, small for gestational age (SGA), various birth defects, gestational age, preterm delivery, spontaneous abortion, stillbirth, and incidence of hypospadias. Overall, these studies provide limited evidence for an association between bromodichloromethane and developmental effects, possibly due to the main limitation of non-differential misclassification of individual exposure. In addition, the various studies have used different approaches to assess exposure, including blood levels of bromodichloromethane, bromodichloromethane in water supplied to the places of residence, and total dose (measured concentration of bromodichloromethane in water plus estimates of water ingestion combined with inhalation and dermal exposure through showering and bathing, and other activities). There is considerable uncertainty due to self-recollection of water use and due to spatial and seasonal variation of disinfection byproducts within a distribution system.

Mixed results have been reported in studies examining the potential effect of bromodichloromethane exposure and birth weight. Birth weight was not significantly associated with bromodichloromethane levels in blood during late pregnancy (median 2.5 ng/L) in a case-control study of pregnant women in China (Cao et. al. 2016), with daily doses ≤0.34 µg bromodichloromethane/day during the entire pregnancy or individual trimesters in a nested-case-control study of pregnant women in Lithuania (Danileviciute et al. 2012), or with bromodichloromethane levels in water (Hoffman et al. 2008). In a retrospective cohort study of 196,000 live births in Massachusetts between 1995 and 1998, exposure to water containing ≥5 µg bromodichloromethane/L during the third trimester of pregnancy was associated with a reduction in birth weight of 12 g (Wright et al. 2004). A more recent study of the same population, that included evaluation of 672,120 live births, confirmed the earlier observations and reported that exposure to a mean concentration of 6.1 µg bromodichloromethane/L in water during the third trimester was associated with reductions in birth weight of 49–63 g in unadjusted models; the association remained significant in adjusted models, but the magnitude of the reductions in birth weight were considerably lower (Rivera-Núñez and Wright 2013).

Evaluations of small for gestational age (SGA) have also provided seemingly inconsistent results. SGA was not associated with exposure to bromodichloromethane assessed by measuring its concentration in blood (Cao et al. 2016), assessed as total intake via multi-route exposure (Danileviciute et al. 2012), or by average water concentration (Hoffman et al. 2008). In contrast, SGA was associated with third trimester bromodichloromethane water supply levels of ≥19 μg/L in a retrospective cohort study of 341,982 live births in Australia (Summerhayes et al. 2012). In general, larger associations were seen in nonsmokers than in smokers, which the investigators attributed to the relatively large smoking effect on SGA possibly masking the effects of subtle risk factors such as trihalomethane exposure on SGA. An association between SGA and ≥5 µg bromodichloromethane/L in water during the third trimester was reported in the earlier study of women in Massachusetts (Wright et al. 2004); bromodichloromethane was also associated with longer gestational age (0.5–0.6 days) in this study. The most recent study of this population did not find an association after adjustments for confounding variables (Rivera-Núñez and Wright 2013); gestational age was not evaluated. In the two studies of women in Massachusetts, preterm delivery was not associated with bromodichloromethane levels in the water supply (Rivera-Núñez and Wright 2013; Wright et al. 2004).

Four studies evaluated associations between exposure to bromodichloromethane and risk of congenital anomalies. Mean bromodichloromethane levels in water during pregnancy were associated with an increase in risk of neural tube defects in a prospective cohort study of residents of Nova Scotia, Canada (Dodds and King 2001); no associations were found for cardiovascular defects or cleft defects. A study of women from Massachusetts did not find associations between bromodichloromethane water levels and the risk of cardiovascular defects (Wright et al. 2017). In a study of women from Lithuania, internal bromodichloromethane dose during the first month of pregnancy was associated with an increased risk of heart anomalies in comparisons of the third tertile (0.051–0.436 µg/day) versus the first tertile (0.000–0.013 µg/day) (Grazuleviciene et al. 2013); no associations were found for musculoskeletal or urogenital anomalies. An intake of ≥6 µg bromodichloromethane/day (combined estimates of water consumption, dishwashing, showering, and swimming during the first trimester) was associated with an increased risk of hypospadias in male offspring in a small case-control study in England (Iszatt et al. 2011); notably, the concentration of bromodichloromethane in water was not associated with hypospadias. However, elevated risk of hypospadias was associated with consumption of cold tap water at home, total water, bottled water, and total fluid (the concentrations of bromodichloromethane in water was not provided, but mean total trihalomethanes ranged from 15 to 51 µg/L).

As with other effects, mixed results have been found in studies examining the possible association between bromodichloromethane and the risk of stillbirth or spontaneous abortions. In a prospective cohort study of Canadian women, exposure to exposure to ≥20 µg bromodichloromethane/L in the water during pregnancy almost doubled the risk of stillbirth (King et al. 2000). Analysis of risk in a continuous representation showed a 29% increase in risk with each 10 µg bromodichloromethane/L. Risk of unexplained stillbirth was not associated with bromodichloromethane, but risk of stillbirth caused by asphyxia was increased 32% per 10 µg/L bromodichloromethane. In contrast, a study of women living in Massachusetts did not find an association between bromodichloromethane levels in municipal water and all causes of stillbirths, but did find an association with unexplained stillbirths (Rivera-Núñez et al. (2018). A large prospective study of pregnant women in California found a doubling of the risk of spontaneous abortion among women with high personal exposure to bromodichloromethane in the tap water (Waller et al. 1998); the risk was further increased after adjustment for high exposure to other trihalomethanes.

A study of 3-day-old infants found an inverse association between maternal blood bromodichloromethane levels and neonatal neurological assessment test scores (Chen et al. 2019); no other epidemiological studies evaluated potential neurodevelopmental effects.

Several studies provide information regarding the developmental effects of bromodichloromethane in laboratory animals following oral exposure. With the exception of one study in rabbits, all have been conducted in rats. The results of these studies indicate that: (1) F344 rats are considerably more susceptible than Sprague-Dawley rats, particularly for the endpoint of full-litter resorptions; (2) mode of administration of bromodichloromethane, gavage vs. drinking water, and the vehicle influence the toxicity; (3) bromodichloromethane is not teratogenic; and (4) effects occur in animals at exposure levels significantly higher than what humans normally encounter through residential or environmental exposures to bromodichloromethane.

The lowest LOAEL for developmental effects in animals was 50 mg/kg/day for full-litter resorptions in F344 rats dosed by gavage on GDs 6–15; no significant resorptions occurred at 25 mg/kg/day (Narotsky et al. 1997). A significantly higher resorption rate was reported when doses of 75 mg/kg/day were administered in an oil vehicle (83%) than when given in an aqueous vehicle (8%). The difference may have been due, at least in part, to a slower measured elimination rate of bromodichloromethane when administered in the oil vehicle compared to the aqueous vehicle. Comparative evaluation of F344 rats and Sprague-Dawley rats showed that full-litter resorptions occurred in the former at a rate of 62% (8/13) following dosing with 75 mg/kg/day, whereas the rate was 0% in the latter strain dosed with ≤100 mg/kg/day (Bielmeier et al. 2001). The investigators noted that it was not clear whether the difference in sensitivity was due to strain differences in reproductive physiology or toxicokinetics.

Studies in F344 rats indicate that the early gestation window as the most sensitive time period for bromodichloromethane-induced full-litter resorptions. Bielmeier et al. (2001) observed 75 and 50% full-litter resorption rates when rats were administered 75 mg/kg/day doses on GDs 6–10 and 6–15, respectively, while administration on GDs 11–15 resulted in 0% full-litter resorptions. It should be noted that in these studies, doses of bromodichloromethane that induced full-litter resorptions (≤100 mg/kg/day) also significantly reduced maternal body weight gain during gestation; however, there was no significant effect on pup viability or neonatal body weight in pregnancies with live litters sacrificed on postnatal day (PND) 6 (Bielmeier et al. 2004).

Additional studies that examined a wide range of developmental endpoints in Sprague-Dawley rats and New Zealand white rabbits exposed during gestation to ≤200 mg/kg/day did not report full-litter resorptions (Christian et al. 2001a, 2001b; Ruddick et al. 1983).

Studies in rats reported minor delays in ossification of the forelimbs and hindlimbs following maternal doses of 82 mg/kg/day in drinking water on GDs 6–21 (Christian et al. 2001a) and of the sternebrae of fetuses from dams dosed with 200 mg/kg/day by gavage on GDs 6–15 (Ruddick et al. 1983). The respective NOAELs were 45 and 100 mg/kg/day. However, no developmental abnormalities were reported in fetuses from rabbits following maternal doses of ≤55.3 mg/kg/day in the drinking water on GDs 6–29 (Christian et al. 2001a). Other endpoints evaluated in these studies included number of corpora lutea, implantation sites, live and dead fetuses and early and late resorptions, fetal body weight, sex ratios, and external and soft tissue abnormalities; none were significantly affected by exposure to bromodichloromethane.

Bromodichloromethane was also tested in a 2-generation reproductive toxicity study in rats (Christian et al. 2001b). The most significant effect was a 14% reduction in body weight in pups from the F1 generation on PND 21; the maternal dose estimated by the investigators during lactation days 1–15 was 94.2 mg/kg/day. The decrease in pup body weight began when the pups started drinking water containing bromodichloromethane and there was a 20% decrease in water intake in this group which was attributed to taste aversion. Thus, the decrease in body weight was considered to be secondary to taste aversion and was not considered toxicologically relevant. Relative spleen weight was also significantly reduced in F1 pups on PND 21 (10–28%). Small but significant delays in preputial separation in F1 males and in vaginal patency in F1 females were reported. However, the differences lost significance when the effects were analyzed using body weight at weaning as covariate. Histological evaluation of unspecified tissues of weanling F1 or F2 pups did not show treatment-related alterations.

Support for the developmental toxicity of bromodichloromethane come from several in vitro studies. In vitro studies by Chen et al. (2003, 2004) provide some support for the association between bromodichloromethane exposure and increases in spontaneous abortion risks. These studies found bromodichloromethane-induced decreases in the secretion of chorionic gonadotrophin in cultured human placental trophoblasts. It is noted that trophoblasts are the sole source of chorionic gonadotrophin in humans and play a major role in maintenance of the conceptus. In porcine embryos, exposure to bromodichloromethane resulted in decreases in blastocyst rate and alterations in hormonal response (Pagé-Larivière et al. 2016). The study also found gene alterations that are consistent with cardiac anomalies.

2.18. OTHER NONCANCER

The available studies in laboratory animals provide suggestive evidence that oral exposure to bromodichloromethane may result in a decrease in blood glucose levels. Decreases in blood glucose levels were observed in rats exposed to bromodichloromethane in the diet for 1, 6, or 18 months (Aida et al. 1989, 1992). However, these data are inconsistent and there is overlap between the NOAEL and LOAEL values. Following 6 months of exposure, the LOAEL was 25.5 (males)/31.7 (females) mg/kg/day and the NOAEL was 6.1/8.0 mg/kg/day; however, after 18 months, only females were affected and the NOAEL and LOAEL values were 31.7 and 168.4 mg/kg/day. This study (Aida et al. 1992) also reported significant increases in blood glucose levels in males exposed to 6.1 or 25.5 mg/kg/day, but not 138.0 mg/kg/day, for 12 months. Acute, single administration studies did not find significant alterations in blood glucose levels (Chu et al. 1982; Lilly et al. 1994, 1996).

No histological alterations were observed in the urinary bladder of rats exposed to 48.0 mg/kg/day bromodichloromethane for 10 months (McDorman et al. 2003).

2.19. CANCER

Information on the carcinogenicity of bromodichloromethane is limited to oral exposure studies in humans and animals. Numerous epidemiological studies indicate that there may be an association between ingestion of chlorinated drinking water (which typically contains bromodichloromethane) and increased risk of cancer in humans (e.g., Cantor et al. 1998; Gottlieb et al. 1981; Kanarek and Young 1982; Marienfeld et al. 1986), but such studies cannot provide information on whether any effects observed are due to bromodichloromethane or to one or more of the hundreds of other byproducts that are also present in chlorinated water. Three studies (Bove et al. 2007; Jones et al. 2019; Min and Min 2016) evaluated risk by individual trihalomethane. No associations were found between bromodichloromethane levels in public water supplies and rectal cancer risk (Bove et al. 2007) or between whole blood bromodichloromethane levels and total cancer deaths (Min and Min 2016). The third study (Jones et al. 2019) found an association between bromodichloromethane levels in municipal water and an increased risk of rectal cancer, but no association with colon cancer.

Several chronic oral studies in laboratory animals have examined the carcinogenic potential of bromodichloromethane. Gavage exposure studies have found significant increases in the incidence of neoplastic lesions in rats and mice. Administration of bromodichloromethane in corn oil for 2 years resulted in increases in the incidence of adenocarcinomas in the large intestine of male rats administered 50 mg/kg and male and female rats administered 100 mg/kg (NTP 1987). Increases in the incidences of renal tubular cell adenocarcinomas and/or combined incidence of adenoma and adenocarcinomas were observed in male and female rats at 100 mg/kg (NTP 1987) and in male mice administered 50 mg/kg (NTP 1987). In female mice, increases in the incidences of hepatocellular adenomas and/or carcinomas were observed at 75 and 150 mg/kg (NTP 1987). Tumasonis et al. (1985) also reported a significant increase in hepatic neoplastic nodules (no additional information was provided) in female rats exposed to 190 mg/kg/day bromodichloromethane in drinking water over a lifetime. Increases in the incidence of skin squamous cell papilloma and/or carcinoma were observed in male rats administered 50 mg/kg, but not at 100 mg/kg or in females at either dose (NTP 1987). Drinking water studies testing lower doses (≤36.3 mg/kg/day in rats and ≤43.3 mg/kg/day in mice) did not find dose-related increases in neoplastic lesions (George et al. 2002; NTP 2006); one study (George et al. 2002) found a significant increase in hepatocellular adenomas and carcinomas in male rats exposed to 3.9 mg/kg/day, but not in groups exposed to 20.6 or 36.3 mg/kg/day. Another study did not find significant increases in neoplastic lesions in male and female rats exposed to doses as high as 138.0 or 168.4 mg/kg/day, respectively, bromodichloromethane microencapsulated and added to the diet (Aida et al. 1992).

NTP (2006) explored possible differences in organ dosimetry between drinking water or dietary administration and gavage administration using physiologically based pharmacokinetic (PBPK) modeling to predict neoplasm incidences in the kidney and large intestine in rats exposed to bromodichloromethane in drinking water. Given the water concentrations used, the model predicted kidney cancer rates of <1%, which is consistent with the empirical incidence of 0/50 in the NTP (2006) drinking water study, suggesting that the difference between the 1987 and 2006 studies was due to organ dosimetry. However, predicted incidences of large intestine neoplasms (3.5–10% depending on the dose metric used) were higher than the observed incidences (2% at 12 mg/kg/day and 0% at 6 and 25 mg/kg/day). NTP (2006) noted that the difference in large intestine tumors between the studies may have also been due to differences in fiber content of the diet used in each study (higher fiber content in the 2006 study compared to the 1987 study).

NTP, EPA, and IARC have classified bromodichloromethane as reasonably anticipated to be a human carcinogen (NTP 2016), a probable human carcinogen (Group B2) (IRIS 2002), or possibly carcinogenic to humans (Group 2B) (IARC 2016), respectively. The cancer classifications are based on inadequate data in humans and sufficient evidence in animal studies.

2.20. GENOTOXICITY

This section is divided into two subsections. The first subsection discusses the results of in vitro and in vivo studies evaluating the genotoxicity of bromodichloromethane. The second subsection presents the results of in vivo studies evaluating epigenetic DNA alterations.

Genotoxicity. Bromodichloromethane has displayed mixed results for genotoxic activity in a variety of in vivo and in vitro tests with organisms ranging from bacteria to humans. As summarized in Table 2-4, bromodichloromethane produced mixed results in gene mutation studies using Salmonella typhimurium (Mortelmans et al. 1986; NTP 1987; Simmon et al. 1977; Sofuni et al. 1996; Varma et al. 1988; Zeiger 1990). Negative results were reported with and without metabolic activation in three studies (Mortelmans et al. 1986; NTP 1987; Zeiger 1990). Varma et al. (1988) reported positive results with metabolic activation in two strains and with or without activation in another two strains; Simmon et al. (1977) also reported an increase in gene mutations when tested with metabolic activation, but only when the assay was performed under a desiccator. Inconclusive results were reported by Sofuni et al. (1996), as only one study out of three produced an increased mutation frequency in the presence of activation only. A weakly positive result was reported in Saccharomyces cerevisiae in the absence of metabolic activation only (Nestmann and Lee 1985). Positive results for gene mutations were also found in mouse lymphoma cells with metabolic activation (McGregor et al. 1988; NTP 1987).

DNA damage was observed in human cell lines (Geter et al. 2004; Zhang et al. 2012), rat hepatocytes (Geter et al. 2004), and human and rat kidney cells (Robbiano et al. 2004), all tested without metabolic activation. A toxicogenomic genotoxicity assay using S. cerevisiae provided evidence of DNA damage (Lan et al. 2018). Inconsistent results have been found in clastogenicity assays. Increases in micronuclei formation were observed in human and rat kidney cells (Robbiano et al. 2004). Four studies found negative results for chromosomal aberrations and/or sister chromatid exchanges (Anderson et al. 1990; Morimoto and Koizumi 1983; NTP 1987; Tucker et al. 1993). However, other studies have found positive results for chromosomal aberrations (Ishidate et al. 1988; Matsuoka et al. 1996) or sister chromatid exchanges (Fujie et al. 1993).

The in vivo genotoxicity of bromodichloromethane has been evaluated in humans, rats, and mice (Table 2-5). In a human study, a 1 μg/m³ increase in bromodichloromethane levels in expired air was associated with an increase in frequency of micronucleated peripheral blood lymphocytes; however, bromodichloromethane only accounted for 10% of the increase in micronuclei formation (Kogevinas et al. 2010). No significant associations were found for micronuclei formation in exfoliated urothelial cells (assessed 2 weeks postexposure), DNA damage in peripheral blood lymphocytes, or reverse mutations in a urine mutagenicity assay (Kogevinas et al. 2010).

Inconsistent results have been found in studies examining the potential of bromodichloromethane to cause DNA damage. Although Robbiano et al. (2004) found a significant increase in single strand breaks in kidney cells of rats administered a single dose of bromodichloromethane; studies by Geter et al. (2004) and Potter et al. (1996) did not find increases in kidney, liver, or duodenum epithelial cells of rats following single dose or repeated oral exposure. No increases in unscheduled DNA activity were observed in the livers of rats administered a single gavage dose of bromodichloromethane (Stocker et al. 1997). In general, positive results have been observed in several studies evaluating bromodichloromethane-induced clastogenic alterations. A weak induction of micronuclei was observed in mature red blood cells of mice exposed to 15 ppm bromodichloromethane vapor for 13 weeks (Torti et al. 2002). A significant increase in micronuclei in bone marrow cells was also observed in mice exposed to 100 ppm for 1 week, but the increase was not statistically significant at the next highest concentration (150 ppm); no significant increases in bone marrow nuclei were observed following a 3-week exposure to ≤15 ppm (Torti et al. 2002). Significant increases in micronuclei formation were also observed in kidney cells of rats administered via gavage 458 mg/kg bromodichloromethane (Robbiano et al. 2004). A dose-related increase in the frequency of chromosomal aberrations was observed in bone marrow cells of rats administered bromodichloromethane via intraperitoneal injection (Fujie et al. 1990); a weakly positive result was also reported in this study for rats receiving bromodichloromethane via gavage for 5 days. Increases in the frequency of sister chromatic exchanges were observed in mice administered bromodichloromethane for 4 days (Morimoto and Koizumi 1983). Although there are inconsistencies in the findings, overall the available data provide suggestive evidence that bromodichloromethane has the potential to damage DNA and chromosomes.

Epigenetic DNA Alterations. Two studies evaluated the potential of bromodichloromethane to induce epigenetic DNA alterations. A study of pregnant women found no associations between maternal blood bromodichloromethane levels (measured in late pregnancy) and DNA methylation in Alu and long interspersed nucleotide element-1 repetitive elements in cord blood after adjustments for prenatal body mass index (BMI), infant sex, passive smoking, and marital status (Yang et al. 2017). A second study (Tao et al. 2005) found significant reductions in DNA methylation in renal cells of mice administered bromodichloromethane via gavage in corn oil or in drinking water and rats administered bromodichloromethane via gavage (Tao et al. 2005). The decreases in DNA methylation were dose-related.