Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 1 Draft, do not cite or quote Chapter III. Toxicokinetics A. Absorption Trichloroacetic acid Older short term studies with mongrel dogs ( Hobara, 1988a) and male B6C3F1 mice ( Styles, 1991) indicate that most of the orally administered dose of TCA is rapidly absorbed.

TCA concentrations in the plasma or liver peak in the first hour following oral dosing. Similar observations are reported in the more recent studies summarized here.

Quantitative evidence for systemic absorption of TCA following oral dosing was provided in a toxicokinetic study by Schultz ( 1999). Male F344 rats were given single oral or intravenous ( IV) doses of 500 µ mol/ kg ( 82 mg/ kg) of TCA. Concentrations of the parent compound were monitored in blood at various times for up to 48 hours. Concentrations of the parent compound in the urine and feces were measured at 48 hours after dosing. Key results from this study are presented in Table III 1. The oral bioavailability of the administered compound was determined from the ratio of the blood area under the curve ( AUC) for the oral and IV doses. Based on this measurement, the oral bioavailability was reported by the study authors as 116% for TCA.

Because blood levels were measured from the jugular vein, any first pass metabolism following oral dosing would not be accounted for in the oral bioavailability calculation. The fact that the Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 2 Draft, do not cite or quote oral bioavailability is high suggests that TCA is not extensively metabolized via first pass metabolism. The AUC for oral dosing was slightly greater than that following IV dosing, but the degree of absorption cannot be greater by the oral route since IV dosing presumes 100%

absorption. Thus, the reported oral bioavailability of 116% likely reflects measurement or statistical variability and/ or differences in clearance rate by the two routes of administration. As a measurement of absorption rate, the mean time to peak blood concentration was determined, and found to be 1.55 hours following oral dosing. The mean absorption time, which was determined as the difference in the mean residence time in blood following dosing via oral and IV routes, was reported as 6 hours for TCA. The mean absorption time is dependent on clearance from the blood as well as the absorption rate; therefore, the longer mean absorption time as compared to time to peak blood concentration of 1.55 hours might also reflect slower clearance following oral dosing. Taken together, the data from this study show that TCA is readily absorbed following a single oral bolus dose. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 3 Draft, do not cite or quote Table III 1. Toxicokinetic Data for TCA in F344 Ratsa Parameters determined following IV dosing with 500 µ mol/ kg TCA ( 82 mg/ kg)

Area under blood concentration time curve AUC ( µ M h) 5406 ± 144b Amount excreted in urine in 24 h (% Dose) 48.5 ± 13.0 Steady state apparent volume of distribution ( mL/ kg) 782 ± 117 Total body clearance ( mL/ hr kg) 92.5 ± 2.5 Renal clearance ( mL/ hr kg) 42.1 ± 9.9 Mean residence time ( hr) 8.5 ± 1.6 Elimination half life ( hr)  total time course 8.0 ± 2.4 Unbound fraction in plasma ( f u) 0.53 ± 0.04 Blood/ plasma concentration ratio 0.76 ± 0.16 Parameters determined following oral dosing with 500 µ mol/ kg TCA ( 82 mg/ kg)

Area under blood concentration time curve AUC ( µ M hr) 6304 ± 1361 Maximum concentration in blood ( µ M) 340 ± 17 Mean residence time ( hr) 14.5 ± 4.7 Time to peak blood concentration ( hr) 1.5 ± 0.3 Mean absorption time ( hr) c 6.0 Oral Bioavailability (%) d 116e aAdapted from Schultz, 1999 bMean ± standard deviation cCalculated as the difference between the mean residence time following IV versus oral dosing dThe ratio of the mean values for AUC for oral versus IV dosing x 100%

eAlthough the study authors reported estimated values of 116% for TCA, this value likely reflects either measurement or statistical variability, and/ or differences in clearance rate between oral and intravenous routes of administration, as oral bioavailability cannot actually be greater than 100%. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 4 Draft, do not cite or quote TCA also appears to be readily absorbed through the skin. Kim and Weisel ( 1998)

investigated the potential for dermal absorption of TCA by evaluating the correlation between exposure to TCA ( swimming pool water concentration × exposure duration) and urinary excretion of TCA in four human volunteers ( two/ sex). Swimming pool water concentrations were measured before and after volunteers either walked or swam in the pools for 30 minutes.

TCA concentrations in the swimming pool water varied from 57 to 871 µ g/ L with a mean of 420 µ g/ L and a median of 278 µ g/ L. In one set of exposures, the four subjects simultaneously walked around in the pool ( dermal exposure only), submerging the entire body exclusive of the head for 30 minutes. In another set of exposures, the same four subjects swam for 30 minutes ( resulting in both dermal exposure and presumed oral exposure from incidental ingestion of pool water).

Entire urine voids were collected for at least 24 hours before exposure, and 20 40 hours following exposure, at approximately 3 hour intervals. Additional urine samples were collected 5 10 minutes before and after exposures. During the 24 hours prior to and following exposure,

subjects avoided activities, such as drinking chlorinated tap water or visiting the dry cleaners,

which might result in urinary TCA excretion. Background levels of TCA were calculated from the amount of urinary TCA excreted in the urine void during the 3 hours prior to pool water exposure. The amount of urinary TCA associated with exposure was estimated by subtracting background levels from TCA levels in the urine void collected 5 to 10 minutes following the exposure period. The results showed that urinary TCA levels were elevated in the 10 minute period following exposure, and generally returned to pre exposure levels within 3 hours.

Postexposure urinary excretion of TCA was highly variable, ranging from approximately 1.1 fold to Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 5 Draft, do not cite or quote 3.9 fold greater than background excretion levels. Higher exposures resulted in higher amounts of urinary TCA adjusted to the subjects' body surface area, suggesting a dose response relationship. The correlation coefficient for TCA exposure and amount excreted was 0.80 ( p=

0.00005).

In another study by the same authors ( Kim and Wiesel, 1998), one male and one female volunteer ingested 500 mL of chlorinated drinking water containing less than 10 µ g/ mL of TCA,

and urine was collected for the following 24 hours. No increase in TCA levels were observed following ingestion, which the authors suggested was due either to variability in background excretion rates or to TCA not being excreted with urine within the time period that urine samples were collected. The rapid appearance of TCA in urine following dermal exposure in swimmingpool water suggested that dermal absorption of TCA was rapid. Skin permeability was not estimated for TCA.

These studies confirm earlier findings and demonstrate that TCA is readily absorbed by the oral and dermal routes. No new studies were identified on the degree or rate of TCA absorption following inhalation exposure.

Monochloroacetic acid Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 6 Draft, do not cite or quote Multiple cases of human systemic poisoning and/ or lethality following acute dermal exposure to MCA, via accidental splashing with molten or concentrated MCA solution ( up to 90% MCA), have been reported, and demonstrate rapid absorption and systemic distribution of this compound following direct skin contact covering at least 10% of the skin surface ( Millischer,

1988; Kusch, 1990; Kulling, 1992).

No toxicokinetic studies in animals that described the degree or rate of absorption of MCA following any route of exposure were identified in the literature. However, the ability of MCA to induce systemic toxicity following oral dosing in animals ( e. g., Berardi, 1987; NTP,

1992; DeAngelo, 1997) shows that MCA is absorbed by the gastrointestinal tract.

B. Distribution Trichloroacetic acid Oral gavage studies in rodents revealed that orally administered TCA is available for systemic distribution in the plasma of animals. Styles ( 1991) reported that in male B6C3F1 mice administered a single oral dose of 500 mg/ kg [ 2 14C] TCA, 43% of the administered radioactivity was found in the liver after 24 hours. The study authors considered the apparent binding in liver tissue to be the result of the metabolism of TCA and subsequent incorporation of metabolites into cellular macromolecules. In another single dose gavage study, male F344 rats and B6C3F1 mice Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 7 Draft, do not cite or quote were administered 20 or 100 mg/ kg ( 4/ dose group) [ 14C] TCA radiolabeled at both carbons ( Larson and Bull, 1992). The majority of the radiolabel in the plasma was not associated with plasma protein, suggesting that most of the TCA distributed in the blood would be available for tissue uptake. TCA can also be recirculated systemically and has been reported to undergo cholecystohepatic circulation as well as reabsorption from the urinary bladder ( Hobara, 1987a;

Hobara, 1988b).

Numerous recent animal studies have been conducted to assess the distribution of TCA.

Schultz ( 1999) administered male F344 rats a single oral or IV dose of 500 µ mol/ kg ( 82 mg/ kg)

of TCA and measured the parent compound in venous blood at various times for up to 48 hours.

The fraction of TCA in plasma not bound to plasma protein ( the unbound fraction) was estimated to be 0.53. The blood/ plasma concentration ratio for TCA was 0.76, indicating some propensity for TCA to partition to the plasma, and was consistent with the ability of TCA to bind plasma proteins. Tissue concentrations were not measured in this study, but based on the similarity between the apparent volume of distribution and the total body water volume in rats, TCA appeared to be widely distributed. The calculated steady state apparent volume of distribution was 782 mL/ kg for TCA, while the authors reported the total body water volume for rats as approximately 660 mL/ kg. Further evidence supporting wide tissue distribution of TCA in total body water is the low lipophilicity of TCA at physiological pH. The octanol buffer partition coefficient ( Log D) at pH 7.4 was reported to be  1.47 ( Schultz, 1999). This highly negative Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 8 Draft, do not cite or quote value suggests that at physiological pH, TCA would have little propensity for accumulation in lipid rich tissues and, thus, would likely be distributed in body water.

The dose dependent partitioning of TCA between blood and liver was examined by Templin ( 1993). Male B6C3F1 mice were administered TCA via a single oral dose of 0.03, 0.12,

or 0.61 mmol/ kg ( corresponding to 5, 20, or 100 mg/ kg), and blood samples were taken at 1, 2,

4, 6, 9, 12, 18, and 24 hours following treatment. Four mice per treatment group were euthanized at each time point and all blood samples were analyzed separately. A pharmacokinetic analysis was conducted to determine the elimination rate constants, area under the blood concentration time curve ( AUC), and clearance values; TCA plasma protein binding was also assessed.

Based on both peak values and totals ( AUC), TCA distribution favored the blood over the liver, and the partitioning of TCA into the blood increased with increasing dose of TCA in a nonlinear manner. Dosing with 0.03, 0.12, or 0.61 mmol/ kg TCA resulted in peak blood concentrations of approximately 50, 250, or 475 nmol/ mL, respectively, and peak concentrations of TCA in liver of approximately 50, 125, or 175 nmol/ mL, respectively, as estimated from a figure presented in the paper. Partitioning to blood was also favored, based on AUC measurements. For example, the liver AUC to blood AUC ratio was approximately 0.75 for a peak TCA blood concentration of 50 nmol/ mL, and approximately 0.45 for a peak blood concentration of 450 nmol/ mL ( estimated from a figure in the paper). The degree of plasma Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 9 Draft, do not cite or quote protein binding was also concentration dependent. The amount of TCA bound to plasma constituents was estimated in vitro by incubating [ 14C] TCA ( position of carbon radiolabel not specified) with plasma, and determining the amount of unbound radioactivity and bound radioactivity ( radioactivity added minus unbound radioactivity). At TCA concentrations in plasma equal to or below 306 nmol/ mL, approximately 50 57% of the TCA was bound to plasma constituents, while plasma constituent binding decreased with plasma TCA concentration at higher TCA concentrations. Approximately 41, 34, and 23% of TCA was bound to plasma constituents at plasma TCA concentrations of 306, 612, and 1224 nmol/ mL, respectively. The decrease in the percent of the bound TCA with increasing plasma concentration was consistent with the binding parameters for TCA estimated by Scatchard plot analysis of these in vitro data.

The estimated K D ( the plasma concentration of TCA resulting in half maximal binding) was 248 nmol/ mL and the estimated B max ( the plasma concentration of TCA resulting in maximal binding)

was 310 nmol/ mL. Thus, plasma TCA concentrations of 306, 612, and 1224 nmol/ mL equaled or exceeded the binding capacity of the plasma, and a lower percent of the TCA in the plasma was bound to plasma constituents. Similarly, based on the determined binding parameters, oral doses of TCA between 20 and 100 mg/ kg/ day, which resulted in peak blood concentrations of 250 and 475 nmol/ mL, respectively, would be expected to result in nearly half maximal to maximal plasma constituent binding in the mouse, and there would be more free TCA present at the higher dose.

The concentration dependent plasma binding is toxicologically significant because it determines the distribution of TCA from blood to target tissues. In addition, plasma binding also would be expected to sequester free TCA and thus compete with TCA metabolism. Therefore, as plasma Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 10 Draft, do not cite or quote binding capacity is exceeded at high doses, more TCA would be available for uptake by tissues.

The role plasma protein binding plays in the distribution of TCA may be of added significance for risk assessment because of potential species differences. In a recent review, Lash ( 2000) noted that TCA is bound more efficiently to plasma proteins in the mouse than in humans, but quantitative differences were not presented.

Toxopeus and Frazier ( 1998) investigated the kinetics of TCA in isolated perfused rat liver ( IPRL), using male F344 rats. The IPRL system was dosed with either 5 or 50 µ mol of TCA, and TCA concentrations were monitored in perfusion medium and bile for 2 hours. Liver viability was assessed by measuring lactate dehydrogenase ( LDH) leakage into perfusion medium and by the rate of bile production. At the end of the exposure period, the concentration of TCA in liver was measured. In the study with 50 µ mol TCA, the total TCA concentration ( free and bound) in perfusion medium decreased slightly during the first 30 minutes and then remained constant for the duration of the exposure period; the total TCA concentration in the perfusion medium was relatively constant in the study with 5 µ mol TCA. At the high dose, approximately 93% TCA was bound to albumin, and the free TCA concentration averaged 15.4 µ M at 5 minutes of exposure and 14.9 µ M at 120 minutes. At the low dose, 96% of the TCA was bound to protein,

and the free TCA concentration was approximately constant at 0.9 to 1 µ M over the study period.

The calculated free TCA concentration in the liver intracellular space was higher than the free

TCA concentration in the perfusion medium. Enzyme leakage and bile flow were similar at both TCA exposure levels to that in the control liver, indicating the absence of hepatotoxicity. The Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 11 Draft, do not cite or quote authors concluded that the binding of TCA to albumin in perfusion medium limits the uptake of TCA by the liver, and that TCA is virtually unmetabolized by the liver. These findings are consistent with those from in vivo mouse studies ( e. g., Templin, 1993) demonstrating TCA binding to serum protein, and suggest that TCA kinetics are determined by plasma protein binding.

Yu ( 2000) studied the tissue distribution of TCA in male F344 rats injected IV with radiolabeled [ 1 14C] TCA at doses of 0, 6.1, 61, or 306 µ mol/ kg ( 0, 1, 10, or 50 mg/ kg). The radiolabeled 14C in blood and tissues was measured at various time points for up to 24 hours post injection and the concentration of TCA equivalents was determined. Following IV injection, the concentration of TCA equivalents in various tissues peaked rapidly. Overall kinetic behaviors were similar at all three doses. No TCA metabolites were measured in plasma, urine, or tissue samples. At early time points, the highest TCA concentrations were measured in plasma, followed by kidney, red blood cells ( RBC), liver, skin, small intestine, large intestine, muscle and fat; the relative order of these concentrations remained unchanged up to 3 hours following dosing. For example, at 3 hours after administration of 306 µ mol/ kg ( 50 mg/ kg) TCA, the TCA equivalent concentrations were roughly: 550 µ mol/ L in plasma, followed by 400 µ mol/ kg in the kidney, 350 µ mol/ kg in the liver, 250 µ mol/ kg in RBCs, 200 µ mol/ kg in skin, 200 µ mol/ kg in the small intestine, 200 µ mol/ kg in the large intestine, 120 µ mol/ kg in muscle, and 30 µ mol/ kg in fat. Thus,

the initial distribution of TCA in tissues appears to be independent of dose, although at the high dose, some nonlinear behavior was noted. However, at 24 hours following dosing, the Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 12 Draft, do not cite or quote distribution pattern was markedly different, reflecting plasma and tissue differences in terminal disappearance rate constants. Disappearance of TCA equivalents from RBC, muscle, and fat was similar to, or faster than, plasma at all doses; disappearance rate constants for kidney and skin were slightly lower than plasma, whereas liver small intestine, and large intestines demonstrated significantly slower elimination. The most notable difference at 24 hours postexposure was between plasma and the liver, when the total concentration of TCA equivalents in liver greatly exceeded that in plasma, perhaps reflecting the slower terminal elimination rate constant from liver compared to plasma due to biological incorporation of TCA metabolites into hepatic intracellular components.

To more fully explain the differences of kinetics of TCA in the plasma and liver, the authors studied the binding characteristics of TCA in plasma and liver. Based on in vitro experiments, the authors concluded that there is much stronger binding of TCA in the plasma than in the liver. However, the authors also noted that it was not possible at the present time to determine whether TCA or its metabolite( s) binds covalently with macromolecules or whether radiolabeled carbon derived from TCA is metabolically incorporated into macromolecules. In addition, based on the rate of formation of extractable and non extractable radioactivity, only limited TCA metabolism was observed. As an alternative to tissue covalent binding or metabolic incorporation into liver cells to explain the slower elimination rate of TCA from liver as compared with plasma, the authors investigated the hepatic intracellular accumulation of TCA. They Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 13 Draft, do not cite or quote determined that TCA binding in the liver was negligible, found that the concentration of unbound TCA in the intracellular space was significantly higher ( p < 0.05) than the biliary concentration of free TCA. Since the difference could not be attributed to TCA binding in the liver, the authors concluded that the slower elimination rate of TCA in the liver results from TCA being transported into hepatic cells faster than it is transported out of these cells.

In agreement with the results of Yu ( 2000), Abbas and Fisher ( 1997) had previously determined partition coefficient values for TCA in B6C3F1 mouse tissues by a centrifugation method. The tissue to blood partition coefficients were 1.18 for the liver, 0.88 for the muscle,

0.74 for the kidney, and 0.54 for the lung. These data support the conclusion that TCA distributes preferentially to the liver in rats and mice. In contrast to the partition coefficients determined for the mouse, the tissue: blood partition coefficients for humans did not show this effect. Using PBPK models developed by Fisher and his colleagues and incorporating whole blood and plasma TCA measurements taken from human volunteers exposed to trichloroethylene.

, Fisher ( 1998) reported human tissue: blood partition coefficients of 0.66 for the liver, 0.66 for the kidney, 0.47 for the lung, and 0.52 for muscle. Thus, tissue: blood partitioning may differ among species. Based on a review of limited data from animal studies and in vitro assays, Lash et al. ( 2000) have proposed that there may also be species differences in plasma protein binding of TCA between humans and mice. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 14 Draft, do not cite or quote No additional studies were identified that might confirm the nature and extent of species differences in TCA distribution. Indirect evidence in humans, primarily from studies involving exposure to chlorinated solvents, suggests that TCA is widely distributed. TCA is a metabolite of trichloroethylene, and has been frequently measured in the urine or blood of humans exposed to trichloroethylene as a result of environmental contamination ( Ziglio, 1981; Ziglio, 1983;

Vartiainen, 1993; Skender, 1994; Bruning, 1998) and in human volunteer studies ( NIOSH, 1973;

Brashear, 1997; Fisher, 1998). TCA is also found in the blood and urine of humans without known chlorinated solvent exposures ( Hajimiragha, 1986) and in individuals exposed to low concentrations of TCA in swimming pool and drinking water ( Kim and Weisel, 1998). These studies demonstrate that TCA, whether it is absorbed from external sources or is formed as a downstream metabolite of other compounds, appears in the blood and urine and is thus available for systemic distribution in humans.

No studies investigating the toxicokinetics or degree of maternal to fetus or blood

tobreast milk transfer of TCA were located.

Monochloroacetic Acid No toxicokinetic studies of MCA in humans have been identified. However, the appearance of damage to multiple organ systems in humans following accidental acute dermal exposure to molten or concentrated MCA solution demonstrates systemic distribution of this Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 15 Draft, do not cite or quote compound following dermal exposure ( Millischer, 1988; Kusch, 1990; Kulling, 1992). As further support for this conclusion, detectible levels of MCA were observed in the blood of a dermal poisoning patient in one reported case ( Kulling, 1992).

Toxicokinetic studies in animals revealed wide tissue distribution of MCA. One study found that liver and kidney levels of radioactivity were greater than levels in the plasma, brain, or heart 32 minutes following subcutaneous administration of 53 or 162 mg/ kg [ 14C] MCA ( position of carbon radiolabel not specified) to male Sprague Dawley rats ( Hayes, 1973). An IV dosing study revealed initially high levels of radioactivity in the liver of male Sprague Dawley rats that decreased over 4 hours, while central nervous system, thymus, and pancreas levels increased ( Bhat, 1990).

Kaphalia ( 1992) administered a single oral gavage dose of 100 µ mol/ kg ( about 10 mg/ kg)

of [ 1 14C] MCA to 15 male Sprague Dawley rats. Urine was collected at 4, 8, 12, 24, and 48 hours following treatment. Three rats at each time point were sacrificed and distribution of radioactivity in plasma, red blood cells, plasma proteins, and major organs and tissues was determined. To study the effect of treatment with a higher dose on the distribution of radioactivity, a second group of 3 animals was administered a single oral gavage dose of approximately 1000 µ mol/ kg ( 100 mg/ kg) [ 1 14C] MCA and sacrificed at 24 hours. A third group was treated with the higher dose ( 100 mg/ kg) daily for 3 days, and sacrificed at 24 hours following the last dose to evaluate the potential for bioaccumulation of MCA and/ or its Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 16 Draft, do not cite or quote metabolites under conditions of repeated dosing. End points examined were the same as those assessed in the low dose group.

In the low dose ( 100 µ mol/ kg) group at 4 and 8 hours post dosing, maximum radioactivity was observed in the intestines and the kidney, followed by liver, spleen, testes, lung,

brain, and heart. Radioactivity disappeared from the intestine more rapidly than from any other tissue examined. The radioactivity in the kidney also decreased sharply, but not as fast as in the intestine. The elimination phase in the spleen, testes, and heart followed a similar time pattern but exhibited a much slower elimination rate. In contrast, radiolabel in the liver increased from 4 to 8 hours and then decreased at a slower rate than that observed in the kidney or intestine. In the high dose ( 1000 µ mol/ kg) group, radioactivity in various tissues at 24 hours post dosing were 1.4 to 3.8 fold higher than that observed in the low dose group. Following three daily doses at the high dose, radioactivity was significantly increased in most tissues except for the liver and spleen,

as compared with radioactivity observed after a single dose. The study authors concluded that some accumulation of MCA in the tissues occurred which appeared to be both dose and duration dependent.

Binding of MCA to red blood cells and hemoglobin was low in all treated groups.

However, about 50% of the total radioactivity in the plasma was present in dialyzed plasma in the low dose group, indicating binding of the radiolabel to the plasma proteins. Plasma protein binding was approximately 73% and 79% of total radiolabel in the single and multiple dose high Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 17 Draft, do not cite or quote dose groups, respectively. These values were considered to be significantly higher that those found at the same time points in the low dose group, indicating an effect of dose on the extent of protein binding. Affinity chromatography indicated that approximately 65% of the total plasma radiolabel was associated with albumin in the groups receiving either a single or multiple dose at the high dose level.

In a more recent study, Saghir ( 2001) examined the kinetics of MCA in adult male Sprague Dawley rats exposed to either a subtoxic ( 10 mg/ kg) or a toxic ( 75 mg/ kg) dose via intravenous injection. Doses were selected on the basis of a previous study in which rats ( 6 7 dose group) were treated intravenously with non radiolabeled MCA at doses ranging from 50 to 125 mg/ kg and observed for up to 72 hours following injection for the onset of toxicity ( coma and mortality). For the kinetics study, rats were administered radiolabeled MCA ( carbon position of radiolabel not specified) at either the subtoxic or toxic dose, and 5 animals were scheduled for sacrifice at each of the following post dosing time points: 5, 15, 45 minutes; 2, 4, 8, and 16 hours.

However, some rats treated with the high dose died at the last 4 time points, reducing the number of animals per group to three or four. Blood and urine samples were collected and various organs removed and analyzed for radioactivity. The gastrointestinal tract ( GIT) was separated into stomach ( esophagus and stomach), small intestine, and large intestine, and total radioactivity in each GIT segment was quantified. Average plasma concentration time profiles of total radioactivity and parent MCA measured after intravenous injection were analyzed by a compartmental modeling method using a non linear least square regression program; the best fit Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 18 Draft, do not cite or quote of the data was obtained by using a two compartment model. The time course of MCAassociated radioactivity in selected tissues was also analyzed to determine AUC and mean residence time ( MRT).

MCA was rapidly distributed to tissues. After 5 minutes, only 0.6 and 1.0% of the radioactive dose/ mL remained in the systemic circulation at 10 and 75 mg/ kg, respectively. Less than 0.1% of dose/ mL remained in the plasma at 8 hours post dosing. Most of the radioactivity associated with plasma was parent MCA. The MRT of MCA in plasma was about 4 hours and the apparent volume of distribution at steady state ( V ss) was about 3 times lower at the toxic dose as compared with the subtoxic dose. Disappearance of radioactivity and of parent compound from plasma followed a biexponential pattern. The subtoxic MCA dose was rapidly distributed to tissues whereas the distribution of the toxic dose to tissues was much slower. The AUC of total radioactivity and of parent MCA in plasma was 22 23 times higher at the 75 mg/ kg dose than at the 10 mg/ kg dose, instead of the 7 to 8 fold decrease expected on the basis of dose proportionality. These results indicate that distribution and/ or clearance was significantly slower at the toxic dose. A higher percent of radioactivity was found in the liver and kidney at the subtoxic dose as compared with the toxic dose. At the low dose, the time course of concentrations of MCA in the liver, heart, lungs, brown fat, muscle, and skin paralleled the plasma concentration and peaked at 5 minutes; whereas at the high dose the concentration in the liver peaked more slowly ( at 15 minutes). MCA associated radioactivity entered the brain rapidly and was retained there at almost the same concentrations throughout the study period, with the Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 19 Draft, do not cite or quote percent of accumulation being proportionate to the dose. In the thymus, the distribution of radioactivity occurred in two distinct phases at both doses: an early rapid distribution, followed by a decline within 15 minutes, and a second peak at about 4 hours post injection ( approximately 1%

of the dose/ gram tissue in both dose groups). Dose proportionality in the thymus occurred at all sampled time points. Distribution of both subtoxic and toxic doses was slower to the spleen and testes than to other tissues and was within the expected dose proportionality. A large fraction of both doses was recovered from the GIT within 45 minutes of treatment. At 5 minutes postdosing approximately 10% of each dose was retained in the GIT; at 45 minutes, GIT recovery was 47% and 23% of the subtoxic and toxic doses, respectively. Most of the GIT radioactivity was located in the small intestine and was rapidly reabsorbed; a maximum of only about 5% of dose reached the large intestine. About 2 3% of each dose was found in the colon within 5 minutes of dosing and was attributed by the authors to be due to direct transport from the blood across the gut wall. This study provides useful information about the distribution of MCA, but the time course data should be treated with caution, since the intravenous dosing used in this study bypasses the first pass metabolism relevant to oral exposure.

No new studies were identified on the distribution of MCA following exposure by the dermal or inhalation routes.

C. Metabolism Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 20 Draft, do not cite or quote Trichloroacetic acid Larson and Bull ( 1992) reported the formation of CO 2, glyoxylic acid, oxalic acid, glycolic acid, and dichloroacetic acid ( DCA) following oral administration of 20 or 100 mg/ kg [ 14C] TCA ( position of carbon radiolabel not given) to rats and mice. The authors suggested that TCA was metabolized by a reductive dehalogenation mechanism to form DCA. The formation of lipid peroxidation induced by TCA was given as evidence for this mechanism, which would result in the formation of free radical intermediates capable of binding to cellular lipids. The liver was suggested as the primary site of TCA metabolism, based on decreased TCA metabolism in mongrel dogs following hepatic by pass ( Hobara, 1987b). Figure III 1 summarizes potential pathways for TCA metabolism. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 21 Draft, do not cite or quote C l C C l C l C O OH C l C C l C O OH C C O C l C C O OH H H H C C O OH HO O HO G lycine CO2 Thiod iglyco late G lyo xylic a c id Oxa lic a cid TCA DCA MCA Adapted from B ull, 2 00 0 a nd L ash e t a l. 2000 H O G lycolate Figure III 1. Proposed metabolism of TCA Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 22 Draft, do not cite or quote The formation of both TCA and DCA as metabolites of trichloroethylene or chloral hydrate ( a trichloroethylene metabolite upstream of TCA) suggested that DCA might be formed from TCA. In an attempt to address this possibility, Abbas ( 1996) compared TCA and DCA kinetics following the administration of trichloroethylene or chloral hydrate. Male B6C3F1 mice were administered IV doses of chloral hydrate at 10, 100, or 300 mg/ kg, and blood and urine samples were collected and examined for chloral hydrate ( CH), trichloroethanol ( TCOH),

trichloroethanol glucuronide ( TCOG), TCA, and DCA. The concentration of TCA gradually increased and approached a steady state over a 4 hour period. The blood AUC for TCA was 26.8, 368, and 818 µ mol hour/ L at doses of 10, 100, and 300 mg/ kg, respectively. Significant amounts of DCA were found in mouse blood, although at only a fraction ( 10 20%) of the TCA concentration. DCA remained in systemic circulation over a 4 hour period and mimicked the shape of the TCA concentration time curve. Based on unpublished data that the half life of DCA in mice is only 12 minutes, the authors stated that the continued persistence of DCA in the presence of TCA suggested that DCA formation is dependent on TCA kinetics, implying that DCA is formed from TCA.

Recent evidence calls into question whether DCA is a metabolite of TCA, or at least the degree of conversion. Lash et al. ( 2000) discussed the evidence surrounding this controversy.

According to this review, Larson and Bull ( 1992) may have over reported DCA concentrations in male F344 and B6C3F1 mice, possibly due to the method used for DCA analysis. Ketcha ( 1996,

as cited in Lash, 2000) suggested that the analytical methods used in the earlier studies might have Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 23 Draft, do not cite or quote resulted in conversion of TCA in biological samples to DCA, and led to over estimation of the formation of DCA. Based on these reports, Lash et al ( 2000) concluded that the " true"

concentrations of DCA in biological samples reported in these earlier animal studies are likely to be lower than the reported values, due to analytical artifacts. Thus, the degree ( if any) of TCA metabolism to DCA remains unclear.

The extent of TCA metabolism to DCA may also be species dependent. Volkel ( 1998)

exposed 3 male and 3 female volunteers, along with Wistar rats ( 3/ sex) to 10, 20, or 40 ppm perchloroethene ( tetrachloroethylene) for 6 hours via inhalation and measured metabolites in the urine. TCA was the major metabolite recovered in the urine of both humans and rats, with urinary excretion being more rapid in rats as compared with humans. Blood concentrations of TCA in rats were also consistently much higher than blood concentrations in humans. DCA was found in the rat urine at about one tenth the concentration of TCA, but was not detected in any of the human urine samples. The authors concluded that levels of DCA in rat blood were too high to be solely due to TCA metabolism.

Recent evidence has suggested that rats have a limited capacity for TCA metabolism.

Schultz ( 1999) compared renal and blood clearance of TCA following a single dose of 500 µ mol/ kg administered intravenously administration to male F344 rats and reported blood, renal,

and nonrenal clearance rates of 92.5, 42.1, and 50.4 mL/ hr kg, respectively. Approximately 46%

of the clearance of TCA from the blood was accounted for by renal clearance, and excretion of Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 24 Draft, do not cite or quote TCA in the feces was negligible. Therefore, as much as 54% of the removal of TCA from the blood could be accounted for by either metabolism or tissue sequestration. No data were provided to determine the degree of metabolism in different tissues. The high oral bioavailability of TCA suggests, however, that the liver may have limited ability to metabolize TCA, supporting the idea that tissue sequestration may account for non renal clearance of intravenously, as well as orally, administered TCA.

TCA was poorly metabolized in F344 rats given IV injections of radiolabeled [ 1 14C] TCA at doses of 0, 6.1, 61, or 306 µ mol/ kg ( 0, 1, 10, or 50 mg/ kg) ( Yu, 2000). Although the fraction of the administered radioactivity excreted in the urine at 24 hours post dosing was as much as 84% at the high dose, HPLC analyses of plasma, urine, and liver homogenate were unable to detect any of the reported metabolites of TCA ( oxalate, DCA, glyoxalate or glycolate).

Nevertheless, about 8 12% of the radioactivity was eliminated in exhaled air as CO 2, indicating that some TCA was metabolized. Intravenous administration of TCA also resulted in a significant increase in non extractable [ 1 14C] label in the liver and plasma. The non extractable radiolabel was considered by the authors to represent metabolites biologically incorporated into hepatic macromolecules which were either retained in the liver or secreted into plasma. Alternatively, the radiolabeled carbon may have been covalently bound to macromolecules in liver plasma. The amount of TCA metabolized in 24 hours, including excretion in exhaled air and non extractable binding in the liver and plasma, was estimated to be less than 20% of the total administered dose. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 25 Draft, do not cite or quote Few data are available on enzyme pathways that might play a role in the metabolism of TCA. Pravacek ( 1996) evaluated the hepatotoxicity of DCA and TCA on liver slices from male B6C3F1 mice, as well as the metabolic capacity of the liver for these two compounds. In the studies evaluating cytotoxicity ( as evidenced by potassium content and liver enzyme leakage), the liver slices were exposed for up to 8 hours at concentrations of TCA ranging from 0 to 86 mM ( 14 mg/ mL) TCA. To determine if TCA treatments can alter phase I or phase II biotransformations, the liver slices were exposed to a low or high concentration of DCA or TCA,

and the conversion of 7 ethoxycoumarin to 7 hydroxycoumarin ( a measure of phase I metabolism), and formation of sulfate and glucuronide conjugates of hydroxycoumarin ( a measure of phase II metabolism) were assessed. TCA treatment with 1000 µ g/ mL increased phase I metabolism, but had no effect on phase II metabolism at either 25 or 1000 µ g/ mL. Metabolism of TCA was monitored by the rate of removal of the parent compound. The removal of TCA was not saturable at non cytotoxic concentrations over the range of concentrations tested ( 0 to 5000 µ g/ mL); thus neither the K m ( the concentration at which half maximal metabolic rate is reached)

or V max ( maximum metabolic rate) was estimated. In contrast, DCA metabolism was saturable.

Based on this difference in kinetics, the study authors suggested that TCA and DCA might be metabolized through distinct pathways, a finding consistent with recent data demonstrating that the primary metabolic pathway for DCA is NADPH and GSH dependent ( e. g., Lipscomb et al.,

1995; Cornett et al., 1997, 1999), whereas that of TCA appears to be mediated by cytochrome P

450 pathways. However, the study authors noted that an alternative explanation for these data is that both TCA and DCA share a single metabolic pathway which has a lower capacity for DCA. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 26 Draft, do not cite or quote Ni ( 1996) studied the mechanism of TCA induced hepatic toxicity in an in vitro system.

Incubation of TCA with male B 6C3F1 mouse liver microsomes resulted in free radical generation and lipid peroxidation. Lipid peroxidation products that were observed included acetaldehyde, formaldehyde, malondialdehyde, acetone, and propionaldehyde. Incubation with liver microsomes from mice pretreated with pyrazole, a specific cytochrome P450 2E1 ( CYP2E1)

enzyme inducer, induced about 2 fold higher lipid peroxidation. The authors also reported that in the same experimental system, the same molar concentration of chloral hydrate ( CH) induced lipid peroxidation to the same extent as TCA, and the CH induced lipid peroxidation could be inhibited by 2,4 dichloro 6 phenylphenoxyethylamine, a general cytochrome P450 inhibitor. Thus, the authors suggested that cytochrome P450 is the enzyme system responsible for metabolic activation of TCA, and that CYP2E1 might be the primary isoform responsible for this metabolism.

In order to determine if TCA induced lipid peroxidation ( see study summary in Chapter V) is due to the formation of radical intermediates following dehalogenation of TCA by cytochrome P450 enzymes, Austin ( 1995) evaluated the effects of pretreating mice with TCA.

Male B6C3F1 mice were pretreated with 1000 mg/ L ( estimated to be 228 mg/ kg/ day by the study authors) TCA in drinking water for 14 days, then administered 300 mg/ kg of TCA, DCA, or an equivalent volume of distilled water ( control) by gavage as an acute challenge. Animals were sacrificed 9 hours following the acute challenge, and lipid peroxidation, peroxisome proliferation,

and TCA induced changes in phase I metabolism were measured. Measures of phase I Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 27 Draft, do not cite or quote metabolism included ( 1) changes in 12 hydroxylation of lauric acid ( an assay specific for CYP4A isoform activity, which is believed to be associated with induction of peroxisome proliferation in rats and mice ( Gibson, 1989); ( 2) changes in p nitrophenol hydroxylation ( an assay specific for CYP2E1 activity); ( 3) immunoblot analysis for induction of cytochrome P450 isoforms CYP2E1,

CYP4A, CYP1A1/ 2, CYP2B1/ 2, and CYP3A1; and ( 4) total liver P450. Pretreatment with TCA increased 12 hydroxylation of lauric acid, demonstrating an increase in CYP4A activity ( and apparently reflecting a peroxisome proliferation response), whereas p nitrophenol hydroxylation was unchanged, indicating no effect on CYP2E1 activity. Immunoblot analysis, a measure of the amount of a protein, was consistent with the increase in CYP4A activity. Increased band intensities on the immunoblot appeared to occur at locations corresponding to those which have been identified as the CYP4A2 and CYP4A3 isoform bands. Similarly, immunoblot analysis was consistent with the absence of an effect on CYP2E1 activity, and also showed no changes in CYP1A1/ 2, 2B1/ 2, and 3A1 protein levels. TCA pretreatment did not alter the overall amount of total liver microsomal P450. These data demonstrate that pretreatment of mice with TCA modifies the lipoperoxidative responses ( described in Chapter V) following acute challenge. The authors suggested that this results from activities associated with peroxisome proliferation and might be related to a shift in the expression of P450 isoforms. The increased levels of CYP4A in TCA pretreated mice is consistent with results observed in other studies with other peroxisome proliferators ( Okita, 1992). Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 28 Draft, do not cite or quote TCA may be converted to DCA in situ in the gastrointestinal tract of mice ( Moghaddam,

1996, 1997). Microflora from the cecum of B6C3F1 mice were shown to be able to metabolize TCA under anaerobic conditions. As further evidence for the microbial metabolism of TCA in the gut, mice depleted of microflora by antibiotic treatment and then treated with TCE had higher gut TCA levels and lower gut DCA levels than control mice receiving similar oral doses of TCE without an antibiotic.

Although the metabolism of TCA to DCA has been proposed ( Larson and Bull, 1992), the degree to which this reaction occurs has been debated ( Lash, 2000), and the mechanism of dehalogenation of TCA has not been conclusively determined. The metabolism of both TCA and DCA to similar downstream metabolites, as described in this paragraph, suggests that they may be sequential metabolites in the same pathway. For this reason, a brief summary of DCA metabolism is included here. For a more detailed analysis of data on DCA metabolism the reader is referred to the Drinking Water Criteria Document for DCA ( U. S. EPA, 2001a). DCA is essentially completely eliminated by metabolism, undergoing metabolic conversion via dechlorination and oxygenation to yield glyoxylate, oxalate, carbon dioxide, and several glycine conjugates, including hippuric acid ( Crabb, 1981; Evans and Stacpoole, 1982; Lin, 1993; James, 1998). In vitro experiments have demonstrated that conjugation with glutathione ( GSH) is the primary metabolic conversion pathway for DCA in B6C3F1 mouse, F344 rat, and human liver cytosol ( Lipscomb,

1995; James, 1997). The glutathione dependent oxygenation of DCA to form the initial major Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 29 Draft, do not cite or quote metabolite, glyoxylic acid, is catalyzed by glutathione S transferase Zeta ( GST Zeta) ( Tong,

1998a; Tong, 1998b).

In summary, the metabolism of TCA has not been well characterized. While several studies have suggested that TCA is metabolized to DCA in mice ( Larson and Bull, 1992; Abbas,

1996), concerns regarding potential over estimation of DCA formation reduce confidence in these findings ( Lash, 2000). TCA appears to be metabolized to only a limited extent in rats ( Schultz,

1999; Toxopeus and Frazier, 1998; Yu, 2000). TCA may also be metabolized differently in humans than in rodents ( Lash, 2000). Enzyme systems responsible for TCA metabolism have not been identified in vivo, but in vitro experiments with mouse tissues have provided limited evidence for involvement of a cytochrome P450 mediated pathway ( Ni, 1996; Prevacek, 1996).

Monochloroacetic acid Bhat ( 1990) suggested that MCA can undergo dehalogenation reactions leading to the formation of oxalate and glycine in male Sprague Dawley rats. The mechanism for removal of chlorine from MCA was not described. This study also indicated that MCA can form a glutathione conjugate, which after further degradation is excreted in the urine. In another rat study ( Bhat and Ansari, 1989), MCA was found to react with lipids, based on the appearance of cholesteryl chloroacetate in neutral lipid fractions from hepatic lipid extracts from treated rats.

Figure III 2 presents the proposed metabolism for MCA. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 30 Draft, do not cite or quote Cl Cl GSCH 2 COOH SCH 2 COOH CH 2 CH COOH NH 2 SCH 2 COOH CH 2 COOH ClCH 2 COOH MCA Cholesteryl Chloroacetate COOH COOH Oxalic Acid + COOH CH 2 NH 2 Glycine Glutathione ( GS)

S carboxylmethyl glutathione S carboxylmethyl cysteine Thiodiacetic acid Figure III 2. Proposed metabolic pathway of MCA ( Adapted from Bhat, 1990) Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 31 Draft, do not cite or quote In the Kaphalia ( 1992) study, the distribution of radioactivity in plasma, red blood cells,

plasma proteins, and major organs and tissues was determined 24 hours following a single oral dose of either 100 or 1000 µ mol/ kg [ 1 14C] MCA ( approximately 10 and 100 mg/ kg, respectively)

or following administration of 1000 µ mol/ kg [ 1 14C] MCA ( approximately 100 mg/ kg) daily for 3 days. About 50% of the total radioactivity in the plasma appeared to bind with plasma protein in the single dose, low dose group. Plasma protein binding was significantly higher in both single and repeated dosing high dose groups, with the bound radiolabel constituting approximately 73%

and 79% of total plasma radiolabel for the single and multiple dose groups, respectively.

Affinity chromatography indicated that approximately 65% of the total plasma radiolabel was associated with albumin in the both high dose groups. Although MCA was absorbed and/ or eliminated at a rapid rate, as demonstrated by the distribution of the radiolabel in intestine and kidney ( and its urinary excretion), absorption and elimination phases were increased in other tissues including liver, lung, and heart, as evidenced by retention of significant radiolabel at 48 hours following exposure. The authors hypothesized that these results might be due to MCA conjugation with cholesterol or phospholipids as proposed by Bahat and Ansari, ( 1988) and Bahat et al ( 1990), and incorporation of glycine into proteins. The authors did not rule out the possibility of metabolic incorporation of the   carbon atom into plasma protein via metabolism of MCA to glycine. However, they concluded that the contribution of this pathway to MCA metabolism was minor. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 32 Draft, do not cite or quote In the Saghir ( 2001) kinetics study, blood, urine, and feces samples were collected and assessed for total radioactivity and parent MCA in both the subtoxic ( 10 mg/ kg) and toxic ( 75 mg/ kg) dose groups. The presence of metabolite( s) was determined by assessing the difference between total radioactivity and parent MCA. For bile analysis, additional rats ( number not specified) were treated with 10 mg/ kg MCA, and anesthetized 30 minutes following MCA administration. The bile duct was cannulated, bile was collected for a period of 1 hour and analyzed for metabolite( s). Decreased biotransformation at the toxic dose was noted at early time points ( within 2 hours post dosing), as indicated by significantly lower biliary excretion of the metabolite( s) ( up to 28% less than at the subtoxic dose) and higher urinary excretion of the parent MCA. The time course of decreased biotransformation and increased retention of radioactivity in tissues was associated with the time of onset of severe toxicity and/ or lethality, which occurred between 40 and 70 minutes following dosing with 75 mg/ kg. Rats surviving this time period retained a larger quantity of metabolite( s) in the GIT and lower plasma and tissue radioactivity than rats dying from toxicity. Metabolites were not identified. However, radioactivity found in bile was primarily associated with one major unidentified metabolite that was more polar than the parent compound; the authors suggested that this metabolite was a glutathione conjugate. The authors also concluded that the rate determining step in the toxicity of MCA is its detoxification by the liver, and that the abrupt onset of severe toxicity and/ or death observed in the 75 mg/ kg group was a consequence of the rapid overwhelming of the detoxification capacity of the liver via metabolic saturation. These conclusions indicate that the authors consider the parent MCA to be the active toxic moiety. Alternatively, MCA induced toxicity might be associated with the Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 33 Draft, do not cite or quote formation of lipophilic metabolites resulting from the metabolism of the parent compound. It has been suggested that MCA can alkylate free amino and thiol groups of amino acids, especially under conditions of glutathione depletion ( Hayes et al., 1973).

Several recent studies on MCA metabolism provide conflicting evidence regarding the involvement of glutathione in MCA metabolism. Previous work has demonstrated that the glutathione conjugate, 2 S glutathionyl acetate, was produced from rat liver microsomal incubation and may result from reaction of glutathione with MCA ( Liebler, 1985). To further explore this metabolic pathway, Dowsley et al. ( 1995) studied microsomal incubations of MCA and glutathione. They found that the glutathione conjugate did not form under these conditions.

However, the authors also noted that they could not exclude the possibility of interactions of MCA and glutathione in vivo.

Wijeweera ( 1998) studied the effects of MCA alone or with 1,1 dichloroethylene ( vinylidene chloride) in Sprague Dawley rat liver slices in vitro. Incubation with 100 µ M MCA resulted in a depletion of glutathione levels that was statistically significant for the first 3 hours of exposure ( decreased by 37, 39, and 45% compared to control levels at 1, 2, and 3 hours of treatment, respectively). This concentration of MCA had no effect on potassium ion release from cells ( a measure of cytotoxicity), the appearance of centrilobular necrosis, or heat shock protein expression. In the co treatment experiments, MCA markedly potentiated the hepatotoxicity of 1,1 dichloroethylene, as measured by potassium ion release, centrilobular necrosis, and heat shock Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 34 Draft, do not cite or quote protein expression, suggesting that depletion of cellular glutathione by MCA may enhance 1,1

dichloroethylene toxicity.

Taken together, data from the in vivo studies of Bhat ( 1990), Bhat and Ansari ( 1989),

and Saghir ( 2001) suggest that MCA may form glutathione conjugates. These data are supported to some extent by the results from incubation studies with tissue liver slices ( Wijeweera, 1998) demonstrating MCA associated glutathione depletion. However, glutathione depletion might have been associated with other factors in the Wijeweera ( 1998) experiments,

and direct measurements in in vitro studies ( Dowsley, 1995) did not find evidence for glutathione conjugation. Therefore, the degree to which MCA conjugates with glutathione, and under what conditions, is not known.

D. Excretion Trichloroacetic acid No full toxicokinetics studies were identified for humans. However, TCA in urine is often measured as a biomarker for chlorinated solvent exposure or exposure to disinfectant by products as described previously in the distribution section. In addition, the therapeutic use of chloral hydrate, of which TCA is a metabolite, has also yielded information on the plasma elimination of TCA. Breimer ( 1974) reported that, following oral administration of a single 15 mg/ kg dose of Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 35 Draft, do not cite or quote chloral hydrate to a human volunteer, TCA levels in blood increased rapidly ( consistent with the rapid metabolism of chloral hydrate to TCA) and were maintained for up to 9 hours without decreasing. The authors noted that this observation was consistent with a long half life of TCA in humans of up to 4 5 days. The slow elimination rate in humans was consistent with the plasma half life of approximately 5.5 days observed by these same authors in a dog given a single IV dose of 60 mg/ kg chloral hydrate. Humbert ( 1994) also reported on the plasma clearance of TCA as part of a human pharmacokinetic study of chloral hydrate. Plasma levels of TCA were measured for 3 hours after a single dose of 40 mg/ kg of chloral hydrate in a neonate ( age and sex not given), and for 14 days after a single dose of 6.25 mg/ kg in an adult ( age and sex not given).

Dosing route was not reported for either case. In the neonate, the plasma level of TCA continued to rise throughout the 3 hour sample period; sampling did not continue long enough to estimate an elimination half life. The reported TCA half life in the adult was 4220 minutes ( 2.9 days).

Both of these human studies ( Breimer, 1974; Humbert, 1994) suggest that TCA is long lived in the plasma. However, only limited conclusions regarding TCA elimination can be drawn from these studies. The observed plasma elimination rate of TCA is affected by the rate of its formation from chloral hydrate and from trichloroethanol, another major chloral hydrate metabolite, and by the inherent elimination rate of TCA. Although chloral hydrate is rapidly cleared from plasma, trichloroethanol displays slower removal kinetics. Since TCA is a metabolite of trichloroethanol, the long plasma half life of TCA could in part reflect the rate of trichloroethanol metabolism in the presence of either trichlorethanol or chloral hydrate. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid 1 C. P. Weisel, Robert Wood Johnson Medical School, Piscataway, NJ.

EPA/ OW/ OST/ HECD III 36 Draft, do not cite or quote In contrast to the slow TCA elimination observed secondary to chloral hydrate metabolism, rapid elimination kinetics of TCA were reported in humans following low dose exposure to TCA from swimming pool water. Kim and Weisel ( 1998) reported rapid clearance of TCA following dermal only or dermal plus oral exposures from swimming pool water. In one set of exposures, four subjects ( two/ sex) walked around in the pool ( dermal exposure only) and in another set of exposures, the same four subjects swam ( dermal exposure and presumed oral exposure from incidental ingestion of pool water during swimming). TCA levels in the urine void collected 5 to 10 minutes after the 30 minute exposure in the pool were elevated and generally returned to pre exposure levels within 3 hours. Post exposure urinary excretion of TCA was 1.1

to 3.9 fold higher than background excretion levels, as estimated from TCA levels in urine voided during the 3 hours prior to pool water exposures. Estimated dermal exposure to TCA ( based on the product of exposure duration and TCA concentration in the pool water) was positively correlated with the urinary levels of this compound. The authors suggested ( Weisel, personal communication) 1 that, although other studies have reported a relatively slow elimination rate following oral or inhalation TCA exposures ( Breimer, 1974; Humbert, 1994; Volkel, 1998), the rapid elimination rate observed in the swimming pool study likely resulted from route dependent and dose dependent differences in TCA kinetics. TCA pool water concentrations were low,

ranging from 57 to 871 µ g/ L, with a mean of 420 µ g/ L and a median of 278 µ g/ L. These dermal exposures resulted in doses on the order of 1 µ g, compared with doses on the order of 1 mg/ kg in Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 37 Draft, do not cite or quote the oral studies. This low dermal dose would be rapidly excreted by the kidneys before being available for uptake by the liver as occurs following oral dosing.

Blood elimination half lives of TCA are fairly rapid in rodents. Blood elimination halflives ranged from 5.4 to 6.0 hours for male B6C3F1 mice administered oral gavage doses of 0.03,

0.12, or 0.61 mmol/ kg TCA ( corresponding to 5, 20, and 100 mg/ kg) ( Templin, 1993). Similar results were reported by Schultz ( 1999), who reported that the elimination half life was 8 hours for F344 rats after IV administration of 500 µ mol/ kg ( 82 mg/ kg) of TCA.

A comparative toxicokinetics study by Volkel ( 1998) suggests that there are differences in the elimination rate of TCA between rats and humans. Human volunteers ( 3/ sex) and Wistar rats ( 3/ sex) were exposed to 10, 20, or 40 ppm perchloroethene ( tetrachloroethylene) for a 6 hour inhalation exposure period. The blood and urine levels of TCA, a major metabolite of tetrachloroethylene, were evaluated in this study. Blood levels of TCA in humans were approximately 10 fold lower than in rats immediately after the exposure to 40 ppm tetrachloroethylene, perhaps reflecting species differences in the absorbed dose of the parent compound or differences in the rate of formation of metabolites, including TCA, from tetrachloroethylene. In humans exposed to 40 ppm tetrachloroethylene, blood levels of TCA immediately following the 6 hour exposure period were similar to those measured at 24 hours post exposure, suggesting slow clearance of TCA from the blood. In contrast, TCA blood concentrations in rats at 24 hours post exposure were decreased to 36% of TCA levels measured Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 38 Draft, do not cite or quote immediately following exposure to the 40 ppm tetrachloroethylene for 6 hours. These results suggest that rats clear TCA from the blood more rapidly than do humans. Differences in urinary elimination of TCA in rats and humans were also observed. The cumulative urinary excretion of TCA was dose dependent in both rats and humans, but the cumulative urinary excretion of TCA relative to body weight over the period measured ( 78 hours for humans and 72 hours for rats) was much greater for rats than for humans, more than 10 fold on a body weight basis. The finding that urinary TCA excretion relative to body weight is higher in rats is consistent with the more rapid plasma clearance observed in these rats. However, interpretation of these data in terms of estimating TCA elimination is limited, because the apparent species differences in TCA clearance might reflect differences in the internal dose of the parent compound ( due to differences in systemic absorption of the inhaled dose and/ or differences in the metabolism of tetrachloroethylene to TCA) or true differences in the rate of TCA excretion. Urinary excretion was more rapid in the rat, as indicated by a mean elimination half life in urine of 45.6 hours in humans as compared with 11.0 hours in rats. The authors suggested that the slow rate of TCA excretion in humans reflects the high degree of TCA binding to plasma proteins. The variability in cumulative TCA excretion or excretion rate was low for the 6 human volunteers. For example,

the coefficient of variation for the urinary elimination half life was only 5.4%. The slower urinary and blood clearance of TCA in humans suggests that for a given internal dose of TCA, humans are likely to have a longer systemic exposure to TCA than rats.

The toxicokinetics studies in animals show that the major route of excretion of TCA is in the urine, with a minor amount exhaled as CO 2. Mice and rats given single oral doses of TCA Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 39 Draft, do not cite or quote exhibited similar patterns of excretion over 24 hours ( mice) or 48 hours ( rats) ( Larson and Bull,

1992). Urinary excretion accounted for 57 72% of the administered dose, roughly 90% of which was eliminated as TCA. Other urinary metabolites identified included glyoxylic acid, oxalic acid,

and glycolic acid. Exhalation of CO 2 accounted for 5 11% of the administered compound.

However, concerns about the analytical methods used in this study limit confidence in the results.

Experiments in mongrel dogs revealed that biliary excretion was minimal over periods up to 2 hours after IV administration of TCA ( Hobara, 1986).

More recent studies on the excretion of TCA have resulted in similar findings. Schultz ( 1999) measured parent compound concentrations in the blood, urine, and feces 24 hours after oral or IV dosing of male F344 rats with 500 µ mol/ kg TCA ( 82 mg/ kg). The urine was the major contributor to blood clearance, while feces made a minimal contribution. Apparent renal clearance of the parent compound accounted for only 46% of the total clearance. ( However,

some of the apparent renal clearance was probably attributable to tissue binding.) Putative metabolites of TCA were not measured, and neither was the release of CO 2. Therefore, it is not possible to determine the contribution of each route of excretion to the total administered dose of the parent compound.

Yu ( 2000) also reported that the major route of TCA excretion was the urine following IV injection of radiolabeled [ 1 14C] TCA at doses of 0, 6.1, 61 or 306 µ mol/ kg ( 0, 1, 10, or 50 mg/ kg) in male F344 rats. Within 9 hours post injection, approximately 35 58% of the TCA Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid 2 John M. Frazier, Wright Patterson Air Force Base, Ohio EPA/ OW/ OST/ HECD III 40 Draft, do not cite or quote associated radioactivity was excreted in the urine at all three dose levels; at 24 hours postexposure the cumulative urinary excretion had increased to 47 84% ( as estimated from a figure in the paper and confirmed by the senior study author 2) . Contributions of fecal and respiratory excretion to the total excretion were much lower. Within 24 hours post injection, only 4 7% of the TCA associated radioactivity was excreted in the feces, and about 8 12% was excreted in exhaled air. Urinary excretion was rapid and dose dependent. At the low dose of 6.1 µ mol/ kg ( 1 mg/ kg) TCA, the mean fraction of the initial dose excreted in the urine was 35% at 9 hours postinjection and this percentage had increased to 47% at 24 hours following exposure. At the high dose of 306 µ mol/ kg ( 50 mg/ kg) TCA, the fraction of the initial dose excreted in urine was reported as 58% at 9 hours post injection and 84% at 24 hours. In contrast, the percentage of administered TCA eliminated via the feces and exhaled in the breath decreased with increasing dose. The terminal first order rate constants for TCA disappearance from various tissues after administration of 6.1 µ mol/ kg ( 1 mg/ kg) were determined. As measured by TCA derived radioactivity, elimination from the liver, small intestine, and large intestine was slower than elimination from the plasma, RBC, muscle, and kidney.

Toxopeus and Frazier ( 1998) investigated the kinetics of TCA in isolated perfused rat liver ( IPRL) test system, using livers from male F344 rats. The livers were perfused with either 5 or 50 µ mol of TCA, and TCA concentrations were monitored in perfusion medium and bile for 2 hours.

Uptake of TCA was limited, as discussed above in the Distribution Section. The total TCA Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 41 Draft, do not cite or quote concentration in bile remained relatively constant throughout the exposure period, averaging 44 µ M. Bile excretion was linear over time and cumulative excretion was 0.1% of the total dose by the end of the experiment, suggesting that biliary excretion contributes minimally to overall elimination of TCA.

In summary, the existing data demonstrate that urine is the primary route of excretion of TCA, with exhalation of CO 2 and fecal excretion contributing to a much lesser extent ( Hobara,

1986; Larson and Bull, 1992; Templin et al., 1993; Schultz et al., 1999; Yu, 2000). The urine is also an important route of excretion in humans, although no quantitative data have been identified to estimate the relative contributions of other routes of excretion. The uptake and elimination of TCA from the blood appears to be considerably slower in humans as compared to rodents ( Volkel, 1998; and reviewed in Lash, 2000). However, results from these studies should be treated with caution, since exposure was actually to TCE, a chemical metabolized to TCA. Thus,

apparent species differences in TCA elimination could actually be due to differences in absorption of TCE or conversion of TCE to TCA. Although human data are very limited, they suggest the possibility that human elimination rates might be route and dose dependent. Slow removal kinetics in humans were reported following oral administration of chloral hydrate ( Breimer, 1974;

Humbert, 1994) while rapid elimination human kinetics were reported following low doses resulting from acute dermal absorption of TCA from swimming pool water ( Kim and Weisel,

1998). The plasma elimination rate is of major significance for risk assessment, because slower elimination kinetics, if true, would suggest that humans would receive a higher cumulative internal Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 42 Draft, do not cite or quote dose of TCA ( AUC) than rodents at the same administered dose, if absorption is similar in rodents and humans. However, the data are insufficient to be useful in the assignment of uncertainty factors for animal to human extrapolation.

Monochloroacetic acid Approximately 50% of a subcutaneous dose of radiolabeled MCA was excreted in the urine of male Sprague Dawley rats within 17 hours of dosing ( Hayes, 1973). Kaphalia ( 1992)

reported that urinary excretion of MCA and/ or its metabolites was rapid in male Sprague Dawley rats administered a single oral dose of 100 µ mol/ kg ( approximately 10 mg/ kg); about 90% of the dose was excreted in the urine at 24 hours postdosing.

In the kinetics study by Saghir ( 2001), blood, urine, bile and feces samples were collected and assessed for total radioactivity and parent MCA. The gastrointestinal tract ( GIT) was separated into stomach ( esophagus and stomach), small intestine, and large intestine, and total radioactivity in each GIT segment ( and contents) was quantitated. Anuria was observed at about 4 hours following dosing in rats administered 75 mg/ kg, apparently due to an inability to empty the urinary bladder rather than a decrease in urine production. The rate of urinary excretion of both parent MCA and MCA metabolites was rapid. Within 45 minutes of treatment, 6 8% of both doses was recovered in the urine. At 16 hours, 73% and 58% of the 10 and 75 mg/ kg doses,

respectively, was recovered in the urine, indicating a slower rate of excretion at the higher, toxic Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 43 Draft, do not cite or quote dose. Analysis of urine samples by HPLC indicated that parent MCA comprised 55% and 68%

of the excreted radioactivity at the10 and 75 mg/ kg doses, respectively, and that a greater percentage of urinary radioactivity was associated with metabolite production ( as compared with parent MCA) at the lower dose. Very little radioactivity was excreted in the feces ( approximately 0.6 2%). However, 47% and 23% of the 10 and 75 mg/ kg doses, respectively, was recovered from the GIT at 45 minutes following treatment, with most of the radioactivity being retained in the small intestine. GIT radioactivity was associated primarily with one major ( unidentified)

metabolite. Rats treated with 10 mg/ kg were approximately 4 times more efficient in removing MCA metabolites from the liver into the bile and from the bile into the GIT, as compared with rats treated with 75 mg/ kg. A maximum of 5% of the radioactivity reached the large intestine,

indicating that significant resorption occurred in the small intestine. The time profile of MCA concentrations in the kidney was similar to that in the GIT and was considered by the authors to be consistent with the hypothesis that metabolite( s) excreted with bile were being resorbed in the small intestine and excreted in the urine following further metabolism.

A fatal MCA poisoning after dermal exposure to an 80% solution of MCA was reported by Kulling ( 1992). Measurements of plasma MCA concentrations 4, 6, 8, and 12 hours after exposure found 33, 15, 7.8 and 0.22 mg/ L, respectively. Based on these data, MCA is rapidly cleared from the blood in humans. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 44 Draft, do not cite or quote No new studies were identified that evaluated the excretion of MCA following dosing by the inhalation route.

E. Bioaccumulation and Retention Trichloroacetic acid No new studies were identified that evaluated the bioaccumulation or retention of TCA following longer term dosing by the oral, dermal, or inhalation routes. The plasma elimination half life in humans following oral or inhalation exposure to trichloroethylene, tetrachloroethylene,

or chloral hydrate suggests slow clearance of TCA ( Breimer, 1974; Humbert, 1994; Volkel,

1998). Based on this slow elimination rate, chronic exposure to high oral or inhalation doses might result in an increase in internal doses of TCA. However, these data from dosing with chemicals that are metabolized to TCA, have limited utility in assessing TCA elimination kinetics because the influence of other factors, including other metabolites and alternate metabolic pathways for the parent compounds, is not well characterized. Rapid TCA clearance was observed following low dose exposures by the dermal route ( Kim and Weisel, 1998), suggesting a limited potential for bioaccumulation at doses likely to result from environmental exposures,

and/ or route and dose dependent differences in elimination and/ or clearance.

Monochloroacetic acid Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 45 Draft, do not cite or quote In the study by Kaphalia ( 1992), repeated oral dosing ( once daily for three consecutive days) with 1000 µ mol/ kg ( approximately 100 mg/ kg) [ 1 14C] MCA resulted in an increase in radioactivity in most tissues, except for the liver and the spleen, of male Sprague Dawley rats at 24 hours post dosing as compared with administration of a single dose of 1000 µ mol/ kg [ 1 14C]

MCA . Further, radioactivity in various tissues at 24 hours postdosing were 1.4 to 3.8 fold higher in rats given a single oral dose of 1000 µ mol/ kg [ 1 14C] MCA than that observed in animals treated with a single oral lower dose of 100 µ mol/ kg ( approximately 10 mg/ kg) [ 1 14C] MCA.

The authors concluded that tissue accumulation occurred to some extent and was both dose and exposure duration dependent. The results from the kinetics study by Saghir ( 2001) suggest the possibility of accumulation or retention of MCA and/ or its metabolites in the brain. Male Sprague Dawley rats were treated with a single intravenous injection of 10 or 75 mg/ kg radiolabeled MCA ( subtoxic and toxic dose, respectively). Following treatment, MCA derived radioactivity in the plasma and most other tissues declined over time ( measured at various time points up to 16 hours postdosing). However, MCA associated radioactivity rapidly entered the brain and was retained there at almost the same concentrations throughout the 16 hour study period, with the percent accumulation being proportionate to the dose. MCA radioactivity also remained higher in thymus gland tissue from 2 16 hours following exposure. These data suggest that MCA and/ or its metabolites may have the potential to accumulate in brain tissue and the thymus gland. No new studies were identified that evaluated the bioaccumulation or retention of MCA following longer term dosing by the oral, dermal, or inhalation routes. Based on Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 46 Draft, do not cite or quote metabolism studies, MCA might interact with lipids to form lipid soluble metabolites such as cholesteryl chloroacetate ( Bhat and Ansari, 1989).

F. PBPK models Abbas and Fisher ( 1997) developed PBPK models for TCA and DCA in B6C3F1 mice exposed to trichloroethylene through oral dosing ( by gavage in corn oil), and these models were expanded by Greenberg ( 1999) to include the inhalation route. The main trichloroethylene PBPK model was linked to five TCE metabolite sub models, for chloral hydrate, trichloroethanol,

trichloroethanol glucuronide, DCA, and TCA. Each sub model contained compartments for the liver, lung, kidney, and body. Abbas and Fisher ( 1997) experimentally determined the tissue: blood partition coefficients for all five TCE metabolites. The model was developed using literature values for V max and K m for trichloroethylene, literature values for physiological parameters, and the experimentally determined tissue partition coefficients. Other parameters were fit using data for trichloroethylene and metabolites obtained from male B6C3F1 mice receiving a single gavage dose of 1200 mg/ kg trichloroethylene. The model was validated using the other doses in the same study ( 300, 600, and 2000 mg/ kg). Additional parameters for the inhalation model ( Greenberg, 1999) were developed from male B6C3F1 mice exposed for 4 hours to 600 ppm trichloroethylene; the model was validated with data from a separate inhalation study conducted at 110 748 ppm trichloroethylene. The TCA model adequately described the TCA concentrations in the liver, lungs, kidneys, and blood following trichloroethylene exposure, as well Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 47 Draft, do not cite or quote as urinary excretion of TCA following oral and inhalation exposure to trichloroethylene. But, the DCA models did not fit the experimental data as well as the TCA models. Since the TCA and DCA observed in the model validation studies came either from trichloroethylene metabolism or from conversion of other trichloroethylene metabolites, the results of directly administering TCA or DCA could not be determined. In addition, the first order metabolic rate constants for the conversion of TCA to DCA, and for conversion of DCA to other metabolites were markedly different in the oral and inhalation models, suggesting that there might be route dependency in the metabolism of TCA and DCA. However, since the metabolic rate constants were estimated from TCA and DCA derived from oral versus inhalation exposure to trichloroethylene, the differences in TCA and DCA metabolic rate constants could simply be secondary to metabolic differences upstream of TCA.

Fisher ( 1998) developed a human PBPK model for trichloroethylene. To account for trichloroethylene metabolism and excretion, the model also included two sub models, one for TCA and one for trichloroethanol. The sub model for TCA was constructed to model concentrations of TCA in human blood and urine following inhalation exposure to trichloroethylene. These sub models included compartments for lung, kidney, body, and liver.

The model was optimized using sex specific metabolic rate constants and partition coefficients for humans exposed to 50 or 100 ppm trichloroethylene. Using the model, the authors successfully estimated TCA concentrations in the blood and urine in exposed males and females. However,

since the TCA modeled in this study came from trichloroethylene metabolism, rather than a direct Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 48 Draft, do not cite or quote exposure to TCA, the usefulness of the model for making judgements about the toxicokinetics of TCA following direct exposures is limited.

G. Summary Trichloroacetic acid TCA is readily absorbed by the oral route in rats ( Schultz, 1999) and by the dermal and oral routes in humans ( Kim and Weisel, 1998). Once absorbed, TCA is available for systemic distribution, based on the appearance of TCA in blood after oral exposure in rodents ( Templin,

1993; Schultz, 1999). Tissue distribution of TCA appears to be dependent on the time of measurement following dosing. For example, in one study ( Yu, 2000), radioactivity following administration of [ 1 14C] TCA levels was highest in plasma and well perfused tissues, such as the kidney and liver, for up to 3 hours after exposure. However, after 24 hours, the level of radioactivity in the liver exceeded levels in the plasma. Intermediate levels of radioactivity were measured in other tissues and were much lower in fat. TCA appears to bind plasma proteins ( Templin, 1993; Toxopeus and Frazier; 1998; Schultz, 1999; Lash et al., 2000; Yu, 2000), which is an important determinant of the extent to which TCA partitions from plasma into target tissues.

No studies were identified that investigated the tissue distribution of TCA in humans, but the appearance of TCA in the blood and urine of humans exposed to chlorinated solvents or orally administered chloral hydrate indicates that it is present in the systemic circulation as a downstream Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 49 Draft, do not cite or quote metabolite. No studies investigating the toxicokinetics or degree of maternal to fetus or bloodto breast milk transfer of TCA were located.

TCA is not readily metabolized, as indicated by minimal first pass metabolism in the liver following oral dosing with TCA ( Schultz, 1999) and by limited amounts of radioactivity excreted in exhaled air or present as non extractable radioactivity in plasma and liver following IV administration of [ 1 14C] TCA ( Yu, 2000). Some studies suggest that TCA is metabolized to DCA ( Larson and Bull, 1992; Abbas, 1996). However, confidence in these results is decreased by concerns regarding potential over estimation of DCA levels due to analytical artifacts ( Lash,

2000). The enzymes involved in TCA metabolism have not been determined, but some in vitro studies suggest the involvement of cytochrome P450s ( Ni, 1996; Pravecek ,1996).

The primary route of excretion of TCA is in the urine, with exhalation of CO 2 and fecal excretion contributing to a much lesser extent ( U. S. EPA, 1994; Templin et al., 1993; Schultz et al., 1999; Yu, 2000). The elimination of TCA from the blood appears to be considerably slower in humans than in rodents administered chlorinated solvents of which it is a downstream metabolite ( Volkel, 1998; Lash, 2000). Based on the slower elimination rate in these human studies, chronic exposure to high oral or inhalation TCA doses might result in an increase in internal doses of TCA. On the other hand, rapid clearance was observed following low dose dermal exposures ( Kim and Weisel, 1998), suggesting limited potential for bioaccumulation at Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 50 Draft, do not cite or quote doses likely to result from environmental exposures, and/ or route and dose dependent differences in clearance and/ or elimination.

Monochloroacetic acid The onset of poisoning and the diversity of affected target organs in humans following accidental acute dermal exposure to concentrated MCA solution demonstrates absorption and systemic distribution of this compound following direct skin contact ( Millischer, 1988; Kusch,

1990; Kulling, 1992). Systemic toxicity observed in animal studies following oral dosing also demonstrates MCA absorption and distribution ( U. S. EPA, 1994; DeAngelo, 1997; Saghir,

2001). Saghir ( 2001) reported that radioactivity following an IV dose of [ 14C] MCA ( position of carbon radiolabel not given) distributed rapidly into tissues, resulting in only minimal plasma concentrations at 45 minutes following dosing. Radioactivity in the liver, heart, lungs, and brown fat paralleled the levels detected in plasma. However, levels of radioactivity in the brain displayed different kinetics as compared with plasma, remaining relatively constant throughout the measurement period ( to 16 hours post exposure). Radioactive levels in the thymus gland were higher 2 16 hours post exposure than immediately following dosing, suggesting possible accumulation. No studies investigating the toxicokinetics or degree of maternal to fetus or blood to breast milk transfer of MCA were located. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 51 Draft, do not cite or quote Some studies suggest that MCA can undergo dehalogenation reactions, leading to the formation of oxalate and glycine ( Bhat, 1990). However, the mechanism for removal of chlorine from MCA was not described and the role of glutathione in MCA metabolism remains unclear.

These studies ( Bhat, 1990) also suggest that MCA can form glutathione conjugates. These data are supported to some extent by the results from incubation studies with liver slices ( Wijeweera,

1998), demonstrating MCA associated glutathione depletion. However, glutathione depletion may be associated with other factors in the Wijeweera ( 1998) experiments, and in vitro studies ( Dowsley, 1995) did not find evidence for this reaction. Therefore, the degree to which MCA conjugates with glutathione, and under what conditions, is not known. The data from the Saghir ( 2001) study suggest that MCA associated toxicity occurs when animals are exposed to high doses of MCA that saturate the detoxification capacity of the liver, possibly via glutathione depletion. MCA has also been reported to bind to lipids, as indicated by the appearance of cholesteryl chloroacetate in neutral lipid fractions from hepatic lipid extracts from treated rats ( Bhat and Ansari, 1989). Based on data showing a dose dependent increase in the plasma protein binding of radiolabeled carbon in rats orally treated with [ 1 14C] MCA and slow rates of radiolabel elimination in a number of tissues, Kaphalia et al. ( 1992) have proposed that MCA associated toxicity might be due to the formation of conjugates with proteins and/ or membrane lipids.

Metabolic incorporation of the radiolabeled carbon via metabolism of MCA to glycine is another possible metabolic pathway, although Kaphalia ( 1992) suggest that the contribution of this pathway to MCA metabolism is likely to be minor. Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 52 Draft, do not cite or quote MCA is rapidly cleared from the blood in humans, based on a case report from a dermal poisoning ( Kulling, 1992). In rats, MCA is excreted primarily in urine, with excretion by this route accounting for approximately 50% of a single subcutaneous dose within 16 or 17 hours of dosing ( Hayes, 1973), and approximately 90% within 24 hours of a single, orally administered dose ( Kaphalia, 1992). Saghir ( 2001) reported that excretion of radiolabeled MCA and/ or metabolite( s) was rapid in rats administered either a subtoxic ( 10 mg/ kg) or toxic ( 75 mg/ kg) dose via IV injection; 6 8% of both doses was recovered in the urine within 45 minutes. At 16 hours,

urinary excretion at the toxic dose ( representing 58% of administered compound) was slower than at the subtoxic dose ( representing 73% of administered dose). HPLC analysis of urine samples showed that a total of 55% and 68% of the excreted radioactivity was parent MCA at the 10 and 75 mg/ kg doses, respectively. These values represented approximately 40% of the injected dose at both dosages. Although very little radioactivity ( 0.6 2%) was excreted in the feces, a large fraction of both doses was recovered from the GIT within 45 minutes of treatment. Most of the radioactivity was concentrated in the small intestine, and a maximum of about 5% of dose reached the large intestine, indicating significant reabsorption from the small intestine. The time profile of MCA concentrations in the kidney was similar to that in the GIT, consistent with the authors'

conclusions that metabolite( s) excreted with bile were being reabsorbed in the small intestine and excreted in the urine.

Limited animal data suggest that MCA and/ or its metabolites might accumulate in body tissue, particularly the brain, depending on dose and/ or exposure duration ( Saghir, 2001; Addendum to Drinking Water Criteria Document for Monochloroacetic Acid and Trichloroacetic Acid EPA/ OW/ OST/ HECD III 53 Draft, do not cite or quote Kaphalia, 1992).


[Federal Register Volume 82, Number 204 (Tuesday, October 24, 2017)]
[Notices]
[Pages 49218-49219]
[FR Doc No: 2017-22998]

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DEPARTMENT OF HOMELAND SECURITY

Coast Guard

[Docket No. USCG-2017-0106]

Collection of Information Under Review by Office of Management
and Budget; OMB Control Number: 1625-0073

AGENCY: Coast Guard, DHS.

ACTION: Thirty-day notice requesting comments.

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SUMMARY: In compliance with the Paperwork Reduction Act of 1995 the
U.S. Coast Guard is forwarding an Information Collection Request (ICR),
abstracted below, to the Office of Management and Budget (OMB), Office
of Information and Regulatory Affairs (OIRA), requesting approval for
reinstatement, without change, of the following collection of
information: 1625-0073, Alteration of Unreasonably Obstructive Bridges.
Our ICR describes the information we seek to collect from the public.
Review and comments by OIRA ensure we only impose paperwork burdens
commensurate with our performance of duties.

DATES: Comments must reach the Coast Guard and OIRA on or before
November 24, 2017.

ADDRESSES: You may submit comments identified by Coast Guard docket
number [USCG-2017-0106] to the Coast Guard using the Federal
eRulemaking Portal at http://www.regulations.gov. Alternatively, you
may submit comments to OIRA using one of the following means:
    (1) Email: [email protected].
    (2) Mail: OIRA, 725 17th Street NW., Washington, DC 20503,
attention Desk Officer for the Coast Guard.
    A copy of the ICR is available through the docket on the Internet
at http://www.regulations.gov. Additionally, copies are available from:
Commandant (CG-612), ATTN: Paperwork Reduction Act Manager, U.S. Coast
Guard, 2703 Martin Luther King Jr. Ave. SE., Stop 7710, Washington, DC
20593-7710.

FOR FURTHER INFORMATION CONTACT: Mr. Anthony Smith, Office of
Information Management, telephone<PHONE_NUMBER>, or fax<PHONE_NUMBER>,
for questions on these documents.

SUPPLEMENTARY INFORMATION:

Public Participation and Request for Comments

    This Notice relies on the authority of the Paperwork Reduction Act
of 1995; 44 U.S.C. Chapter 35, as amended. An ICR is an application to
OIRA seeking the approval, extension, or renewal of a Coast Guard
collection of information (Collection). The ICR contains information
describing the Collection's purpose, the Collection's likely burden on
the affected public, an explanation of the necessity of the Collection,
and other important information describing the Collection. There is one
ICR for each Collection. The Coast Guard invites comments on whether
this ICR should

be granted based on the Collection being necessary for the proper
performance of Departmental functions. In particular, the Coast Guard
would appreciate comments addressing: (1) The practical utility of the
Collection; (2) the accuracy of the estimated burden of the Collection;
(3) ways to enhance the quality, utility, and clarity of information
subject to the Collection; and (4) ways to minimize the burden of the
Collection on respondents, including the use of automated collection
techniques or other forms of information technology. These comments
will help OIRA determine whether to approve the ICR referred to in this
Notice.
    We encourage you to respond to this request by submitting comments
and related materials. Comments to Coast Guard or OIRA must contain the
OMB Control Number of the ICR. They must also contain the docket number
of this request, [USCG-2017-0106], and must be received by November 24,
2017.

Submitting Comments

    We encourage you to submit comments through the Federal eRulemaking
Portal at http://www.regulations.gov. If your material cannot be
submitted using http://www.regulations.gov, contact the person in the
FOR FURTHER INFORMATION CONTACT section of this document for alternate
instructions. Documents mentioned in this notice, and all public
comments, are in our online docket at http://www.regulations.gov and
can be viewed by following that Web site's instructions. Additionally,
if you go to the online docket and sign up for email alerts, you will
be notified when comments are posted.
    We accept anonymous comments. All comments received will be posted
without change to http://www.regulations.gov and will include any
personal information you have provided. For more about privacy and the
docket, you may review a Privacy Act notice regarding the Federal
Docket Management System in the March 24, 2005, issue of the Federal
Register (70 FR 15086).
    OIRA posts its decisions on ICRs online at http://www.reginfo.gov/public/do/PRAMain after the comment period for each ICR. An OMB Notice
of Action on each ICR will become available via a hyperlink in the OMB
Control Number: 1625-0073.

Previous Request for Comments

    This request provides a 30-day comment period required by OIRA. The
Coast Guard published the 60-day notice (82 FR 37462, August 10, 2017)
required by 44 U.S.C. 3506(c)(2). That Notice elicited no comments.
Accordingly, no changes have been made to the Collections.

Information Collection Request

    Title: Alterations of Unreasonably Obstructive Bridges.
    OMB Control Number: 1625-0073.
    Summary: The collection of information is a request to determine if
the bridge is unreasonably obstructive.
    Need: 33 U.S.C. 494, 502, 511, 513, 514, 515, 516, 517, 521, 522,
523 and 524 authorize the Coast Guard to remove or alter bridges and
causeways over the navigable waters of the United States and that the
Coast Guard deems to be unreasonably obstructive.
    Forms: None.
    Respondents: Public and private owners of bridges over navigable
waters of the United States.
    Frequency: On occasion.
    Hour Burden Estimate: The estimated burden has decreased from 240
hours to 160 hours a year. There are six additional engineering
projects that have been added to this information collection since
2014. Two of the four previous projects have been completed since 2014;
making a total of eight current projects. The reduction in burden is
based on having previously completed the majority of the review for
each study.

    Authority: The Paperwork Reduction Act of 1995; 44 U.S.C.
Chapter 35, as amended.

    Dated: October 12, 2017.
James D. Roppel,
U.S. Coast Guard, Acting Chief, Office of Information Management.
[FR Doc. 2017-22998 Filed 10-23-17; 8:45 am]
BILLING CODE 9110-04-P