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Disposition of [14C]2,3-dichloropropene in Fischer-344 rats after oral or intraperitoneal administration
Toxicology
Letters,
23 (1984) 119-125
119
Elsevier
TOXLett.
1293
DISPOSITION OF [‘4C]2,3-DICHLOROPROPENE IN FISCHER-344 AFTER ORAL OR INTRAPERITONEAL ADMINISTRATION (Fumigant; urinary excretion; gastro-intestinal
M.A.
MEDINSKY,
Lovelace
J.A.
Inhalation
BOND,
Toxicology
*National
Institute
(Received
April
(Accepted
May 19th, 1984)
J.S. DUTCHER Research
of Environmental
lSth,
absorption)
and L.S. BIRNBAUM*
Institute,
Health
RATS
P.O.
Sciences,
Box 5890, Albuquerque,
Research
NM 87185,
and
Triangle Park, NC 27709 (U.S.A.)
1984)
SUMMARY 2,3-Dichloropropene (2,3-DCP), a component with [‘?]2,3-DCP and given to rats by peroral feces, and expired other routes,
air were collected
over 72 h. Excretion
with 66% to 75010 of the dose excreted
8% of the dose was exhaled carcass
of commercial fumigants (p.0.) or intraperitoneal
as %Oz.
with the highest concentrations
dose was absorbed
from
of radioactivity
in 72 h. Feces contained
and nematocides, was mixed (i.p.) administration. Urine, in urine predominated from
At the end of 72 h, only 2% to 3% of the dose remained of r4C in liver, kidney,
the gastrointestinal
over
13% to 21% of dose.
testes, and lung. Approx.
in the
91% of the p.o.
(GI) tract.
INTRODUCTION
Dichloropropenes are used in combination with dichloropropane in the production of commercially available fumigants and nematocides. The major constituents of these fumigants are c&1,3 and trans-1,3 dichloropropene (55-60’70) with 2,3-DCP occurring in much smaller amounts [l]. 2,3-DCP is also an intermediate in the production of the herbicide 2-chloroallyl diethyldithiocarbamate [2]. Dichloropropenes are substituted ally1 chlorides with a double bond between the first and second carbons. These compounds are liquid at room temperature (bp. approx. 100°C; vapor pressure approx. 28 torr at 25”C), and during agricultural use the fumigant mixtures are injected into the soil. Thus, the greatest potential for exposure of people exists during manufacture of the chemicals or during bulk handling activities. Abbreviations:
0378-4274/84/S
2,3-DCP,
03.00
2,3-dichloropropene;
0 Elsevier
Science
GI, gastrointestinal.
Publishers
B.V.
120
Previous studies had shown that c&1,3, trans-1,3 and 2,3-DCP were directly mutagenic in the Salmonella typhimurium assay [3, 41. One study has reported on the carcinogenicity of cis-1,3-DCP which was sarcomagenic after subcutaneous injection [5]. Work by Hutson et al. [6] and Climie et al. [7] on the retention and pathways for excretion of radiolabeled isomers of 1,3-DCP indicated distinct differences in the pathways for excretion of these isomers, with five times more radioactivity exhaled as i4C02 after oral administration of the trans isomer compared to the cis. The purpose of the present study was to determine the absorption, excretion and tissue distribution of 2,3-DCP 72 h after either p.o. or i.p. administration and to compare these results to those reported for the 1,3-DCP isomers. METHOD
[1,2-‘4C]2,3-Dichloro-l-propene ([14C]2,3-DCP; 3.2 &i/pmol) was obtained from Midwest Research Institute, Kansas City, MO. Unlabeled 2,3-DCP (98% pure) WI, and distilled before was purchased from Aldrich Chemical Co., Milwaukee, use. Radiochemical purity of the [14C]2,3-DCP was found to be 98% using reversephase high-performance liquid chromatography with a 10 pm PBondapak Crs column (Waters Associates, Milford, MA) and a 10 to 100% methanol:water gradient in 30 min at a flow rate of 1 ml/min. All rats used in these studies were male Fischer-344, 13 weeks of age at the time of dosing, born and raised in the barrier-maintained colony of the Lovelace Inhalation Toxicology Research Institute. They were housed two per polycarbonate cage with hardwood chip bedding and filter caps. Rooms were maintained at 68-72°F with a relative humidity of 20-50% and a 12-h 1ight:dark cycle, with light starting at 06.00. Food (Lab Blox, Allied Mills, Chicago IL) and water from bottles with sipper tubes were provided ad lib. throughout the study. [14C]2,3-DCP was dissolved in corn oil, and administered orally in 0.5 ml corn oil to 3 rats at a dose of 32 mg 2,3-DCP/kg body weight. An additional 3 rats were given i.p. injections of 2,3-DCP in corn oil at the same dose. Specific activity of the dosing solution was adjusted by addition of unlabeled 2,3-DCP so that each rat received approx. 5 &i of r4C. Immediately after administration of [‘4C]2,3-DCP, rats were placed in glass metabolism cages (Stanford Glass, Palo Alto, CA) for collection of urine, feces, and expired air. Urine and feces were collected in glass containers on dry ice at 4, 7, 10, 16, 20, 24, 32, 48, 54, and 72 h. Expired air from each cage was pulled at a flow rate of 500 ml/min by a vacuum pump through two gas traps containing 200 ml of 5 M KOH. Samples were removed from the KOH traps every h for the first 4 h, after which time solutions in the traps were sampled and changed at the times of the other excreta collections. At the end of 72 h, rats were killed and blood samples (2 ml) were obtained by
121
heart puncture. Tissues or samples of tissues taken included liver, brain, thyroid, larynx, spleen, thymus, kidneys, adrenals, stomach, small intestine sample, large intestine sample, urinary bladder, testes, fat sample (perirenal and subcutaneous), muscle sample, bone (femur), pelt sample, heart, lung, trachea, remaining GI tract, and carcass. The tissues were weighed, placed in sealed containers, and stored at - 80°C until analysis. Samples of tissues were homogenized in 2 vols of deionized water on ice to prein duplicate samples of vent loss of radiolabeled volatiles. Radiocarbon homogenates was determined by digestion of 300-mg samples of the homogenate with 1 ml of Protosol (New England Nuclear, Boston, MA). Samples were decolorized with 0.5 ml of 30% Hz02 and, after addition of scintillation fluid (Ready-Solv, Beckman, Fullerton, CA), radioactivity in samples was quantitated in a Packard 460C Liquid Scintillation Spectrometer (Packard Instruments, Downers Grove, CA). Quench correction was by automatic external standard, with sufficient counts accumulated to give c 5% error with a 95% confidence interval. Samples of urine and 0.5-ml samples of the 5 M KOH traps were added directly to scintillation fluid for determination of radioactivity. To distinquish 14C02 from exhaled [i4C]2,3-DCP or other volatile metabolites, additional 0.5-ml samples of the KOH traps were acidified with 0.3 ml of concentrated HCl to release CO2 and diluted with 2 ml of deionized water. The residual radioactivity in these samples was considered to be radiolabeled volatiles other than 14C02. Feces were mixed with 5 ml of 1% Triton X-100, vortexed, and 300 mg samples of the homogenate were digested with Protosol and decolorized as described above. Radioactivity in all samples of excreta was determined after addition of scintillation fluid, as described above. Gastrointestinal absorption of 2,3-DCP was calculated by comparing the excretion of radioactivity in urine, feces, and expired 14C02 after p.o. and i.p. administration of [r4C]2,3-DCP as in the following equation: %ABS = 100 - Fp.0, + Fi,p. [(u*.o. +
Ap.o.)/(Ui.p.
+ Ai.p.)],
where U= urinary radioactivity, F= fecal radioactivity, and A = expired 14C02, expressed as % dose excreted during 72 h after either p.o. or i.p. administration of [14C]2,3-DCP. We assumed that the injection of [‘4C]2,3-DCP was equivalent to oral administration with 100% absorption and that &,. was equal to the % radioactivity excreted in feces with 100% absorption of parent compound. Half-times for excretion were determined by least-squares linear regression of the log of the fraction of radioactivity remaining to be excreted as a function of line. T~/z = -0.693/X where X equaled the slope of the regression time. Statistical analyses of data were performed by comparing means using a onetailed Student’s t-test. The level of significance was set at P
3
HOURS
9
AFTER
d
DOSING
HOURS
AFTER
DOSING
c r
0
0
15
30 HOURS
Fig. 1. Cumulative after intraperitoneal sent means
RESULTS
f
45 AFTER
60
75
DOSING
Vo of dose excreted or oral administration
as 14C in urine (a), feces (b), and exhaled as “‘CO2 (c) by rats of 32 mg [‘4C]2,3-DCP/kg body weight. Data points repre-
SEM. n = 3.
AND DISCUSSION
Urine accounted for the majority of 14C excreted after administration of [i4C]2,3-DCP, with 66 and 75% excreted by this route after p.o. or i.p. administration, respectively (Fig. la; Table I). The half-times for excretion were not significantly different after i.p. or p.o. administration being 4.9 +_ 1.3 h and 7.5
123 TABLE I DISTRIBUTION OF 14CAT 72 h AFTER ADMINISTRATION (2,3-DCP) Route of administration
Intraperitoneal injection Oral administration
OF [‘4C]2,3-DICHLOROPROPENE
% Dose found in Urine
Feces
Exhaled CO2
Exhaled 2,3-DCP
Carcass and tissues
75+ ia 66+0.4
13kI 21+ 1
7kO.4 8kI
2kO.2 2+0.1
3kO.6 2kO.I
aData represent means and SEM from 3 rats.
+ 0.6 h (mean f SE), respectively. Radioactivity was also found in the feces (Fig. lb; Table I) and in the expired air as 14C02 (Fig. Ic; Table I). Similar studies of [14C]cis-1,3 and trans-1,3-DCP demonstrated that urine was also the major route of excretion of 14C for these compounds after p.o. administration [6]. Most of the radioactivity administered was eliminated during the first 24 h of the experiment, similar to what was seen with our studies on 2,3-DCP (Fig. 1). Climie et al. [7] identified the major urinary metabolite (92% of radioactivity excreted in urine) of c&l ,3-DCP in rats as N-acetyl-S-[3-chloroprophenyl] cysteine. Metabolism involved formation of a carbon sulfur bond between glutathione and 1,3-DCP, with loss of a Cl atom from the carbon-3 position. The authors proposed that the DCP’s were eliminated in urine because of this efficient glutathionedependent biotransformation. In our study, fecal excretion of 14C was significantly higher (21%) after p.o. administration of [14C]2,3-DCP, compared to i.p. (13%). This difference was probably due to unabsorbed 2,3-DCP. Calculation of GI absorption indicated that 91% of the administered dose was absorbed from the GI tract. Hutson et al. [6] found that only 2-3% of an oral dose of [14C]cis or trans-1,3-DCP was excreted in the feces 4 days after administration, This is less than what we found even after i.p. administration suggesting possible differences in either biliary excretion or enterohepatic circulation between the 1,3-DCPs and 2,3-DCP. In the studies reported here, acidification of samples obtained from 5 M KOH traps decreased the radioactivity remaining in the samples, indicating that most of the 14C in the expired air was due to 14C02 (8% of dose). Less than 2% of the dose was due to acid-stable volatile compounds, which could include unmetabolized 2,3-DCP. Hutson et al. [6] also found little volatile radioactivity other than 14C02 in the expired air of rats given cis- or truns-1,3-DCP. These authors did find a difference between the cis and ~rans isomers in the amount of radioactivity exhaled as r4C02 [6]. Studies with c&1,3- and truns-1,3-DCP showed 5 and 23% of the dose excreted as i4C02, respectively. In studies on 2,3-DCP, we found that approx. 8% of the dose was exhaled as r4C02 after p.o. administration of 2,3-DCP suggesting that in this respect, the pathways for metabolism of 2,3-DCP may be more like the cis-1,3-DCP than trans-1,3-DCP.
124
Similar to what was seen with the cis- and trans-1,3-DCPs [6], from 2-3% of the radioactivity associated with 2,3-DCP remained in the rats at 72 h after dosing (Table I). Major organs for retention of radioactivity associated with 2,3-DCP included the liver, kidney, testes, lung, and brain (Fig. 2). The liver accounted for approximately 20% of the 14C remaining at the end of 72 h. There appeared to be no differences in the retention of 14C, between the two routes of administration studied. When expressed on a per-gram basis, tissues in which concentrations of 14C found were higher than concentrations found in carcass (approx. 8 nmol/g) included the adrenals, spleen, and turbinates (19 to 16 nmol [i4C]DCP equivalents/g tissue) in addition to tissues shown in Fig. 2. In summary, the results of studies reported here indicate that the disposition of 2,3-DCP is similar to that found for the cis-1,3-isomer, with most of the radioactivity excreted within 24 h in the urine and a smaller fraction excreted as COZ. Although most of the ingested 2,3-DCP will be absorbed, any absorbed 2,3-DCP will be rapidly excreted with no apparent accumulation in any tissue.
TISSUE
Fig. 2. nmol of [‘4C]2,3-DCP equivalents present in tissues at 72 h after oral administration (solid bars) or intraperitoneal injection (hatched bars) of 32 mg [?]2,3-DCP/kg body weight. Each bar represents mean + SEM, n=3.
125
ACKNOWLEDGEMENTS
Research conducted as part of the National Toxicology Program under U.S. Department of Energy (DOE) contract number DE-AC04-76V01013 and interagency agreement with the National Institute of Environmental Health Sciences. The facilities used are fully accredited by the American Association for Accreditation of Laboratory Animal Care. The authors gratefully acknowledge the excellent technical assistance of Ms. Frances Straus and Ms. Eileen Cahill and the useful discussions with a number of our colleagues at the Institute, with particular appreciation to Drs. R.O. McClellan and R.F. Henderson. REFERENCES 1 J.P.
Nater and V.H.J.
Gooskens,
Occupational
dermatosis
due to soil fumigant,
Contact
Derm.
2
(1976) 227-229. 2 SRI
International,
573.7007B,
3 F. DeLorenzo, containing
S. Degl’tnnocenti, D. Lutz,
substituted
Biochem.
Economics
A. Ruocco, Cancer
Menlo
Park,
CA,
L. Silengo and R. Cortese,
Res., 37 (1977)
E. Eder and D. Henschler,
ally1 and allylic
Pharmacol.,
Handbook,
1980,
pp.
573.70030-X,
and 573.9003H-K.
1,3-dichloropropene,
4 T. Neudecker, alkyl
Chemical
573.9OOIR-W,
compounds:
Mutagenicity
of pesticides
1915-1917.
Structure-activity
correlation
relationship
of alkylating
in halogen
and mutagenic
and
properties,
29 (1980) 2611-2617.
5 B. Van Duuren, B.M. Goldschmidt, G. Loewengart, A.C. Smith, S. Melchlonne, I. Seldman and D. Roth, Carcinogenicity of halogenated olefinic and aliphatic hydrocarbons in mice, J. Natl. Cancer Inst., 6 D.H.
63, (1979) 1433-1439. Hutson,
fumigant 7 I.J.G. ification
J.A. Moss and B.A. Pickering,
D-D and their metabolites
Climie,
D.H.
Hutson,
B.J. Morrison
of (z)-1,3-dichloropropene
9 (1979) 149-156.
The excretion
in the rat, Food
and G. Stoydin,
(a component
and retention
Cosmet.
Toxicol., Glutathione
of the nematocide
of components
of the soil
9 (1971) 677-680. conjugation
in the detox-
D-D) in the rat, Xenobiotica,
Letters,
23 (1984) 119-125
119
Elsevier
TOXLett.
1293
DISPOSITION OF [‘4C]2,3-DICHLOROPROPENE IN FISCHER-344 AFTER ORAL OR INTRAPERITONEAL ADMINISTRATION (Fumigant; urinary excretion; gastro-intestinal
M.A.
MEDINSKY,
Lovelace
J.A.
Inhalation
BOND,
Toxicology
*National
Institute
(Received
April
(Accepted
May 19th, 1984)
J.S. DUTCHER Research
of Environmental
lSth,
absorption)
and L.S. BIRNBAUM*
Institute,
Health
RATS
P.O.
Sciences,
Box 5890, Albuquerque,
Research
NM 87185,
and
Triangle Park, NC 27709 (U.S.A.)
1984)
SUMMARY 2,3-Dichloropropene (2,3-DCP), a component with [‘?]2,3-DCP and given to rats by peroral feces, and expired other routes,
air were collected
over 72 h. Excretion
with 66% to 75010 of the dose excreted
8% of the dose was exhaled carcass
of commercial fumigants (p.0.) or intraperitoneal
as %Oz.
with the highest concentrations
dose was absorbed
from
of radioactivity
in 72 h. Feces contained
and nematocides, was mixed (i.p.) administration. Urine, in urine predominated from
At the end of 72 h, only 2% to 3% of the dose remained of r4C in liver, kidney,
the gastrointestinal
over
13% to 21% of dose.
testes, and lung. Approx.
in the
91% of the p.o.
(GI) tract.
INTRODUCTION
Dichloropropenes are used in combination with dichloropropane in the production of commercially available fumigants and nematocides. The major constituents of these fumigants are c&1,3 and trans-1,3 dichloropropene (55-60’70) with 2,3-DCP occurring in much smaller amounts [l]. 2,3-DCP is also an intermediate in the production of the herbicide 2-chloroallyl diethyldithiocarbamate [2]. Dichloropropenes are substituted ally1 chlorides with a double bond between the first and second carbons. These compounds are liquid at room temperature (bp. approx. 100°C; vapor pressure approx. 28 torr at 25”C), and during agricultural use the fumigant mixtures are injected into the soil. Thus, the greatest potential for exposure of people exists during manufacture of the chemicals or during bulk handling activities. Abbreviations:
0378-4274/84/S
2,3-DCP,
03.00
2,3-dichloropropene;
0 Elsevier
Science
GI, gastrointestinal.
Publishers
B.V.
120
Previous studies had shown that c&1,3, trans-1,3 and 2,3-DCP were directly mutagenic in the Salmonella typhimurium assay [3, 41. One study has reported on the carcinogenicity of cis-1,3-DCP which was sarcomagenic after subcutaneous injection [5]. Work by Hutson et al. [6] and Climie et al. [7] on the retention and pathways for excretion of radiolabeled isomers of 1,3-DCP indicated distinct differences in the pathways for excretion of these isomers, with five times more radioactivity exhaled as i4C02 after oral administration of the trans isomer compared to the cis. The purpose of the present study was to determine the absorption, excretion and tissue distribution of 2,3-DCP 72 h after either p.o. or i.p. administration and to compare these results to those reported for the 1,3-DCP isomers. METHOD
[1,2-‘4C]2,3-Dichloro-l-propene ([14C]2,3-DCP; 3.2 &i/pmol) was obtained from Midwest Research Institute, Kansas City, MO. Unlabeled 2,3-DCP (98% pure) WI, and distilled before was purchased from Aldrich Chemical Co., Milwaukee, use. Radiochemical purity of the [14C]2,3-DCP was found to be 98% using reversephase high-performance liquid chromatography with a 10 pm PBondapak Crs column (Waters Associates, Milford, MA) and a 10 to 100% methanol:water gradient in 30 min at a flow rate of 1 ml/min. All rats used in these studies were male Fischer-344, 13 weeks of age at the time of dosing, born and raised in the barrier-maintained colony of the Lovelace Inhalation Toxicology Research Institute. They were housed two per polycarbonate cage with hardwood chip bedding and filter caps. Rooms were maintained at 68-72°F with a relative humidity of 20-50% and a 12-h 1ight:dark cycle, with light starting at 06.00. Food (Lab Blox, Allied Mills, Chicago IL) and water from bottles with sipper tubes were provided ad lib. throughout the study. [14C]2,3-DCP was dissolved in corn oil, and administered orally in 0.5 ml corn oil to 3 rats at a dose of 32 mg 2,3-DCP/kg body weight. An additional 3 rats were given i.p. injections of 2,3-DCP in corn oil at the same dose. Specific activity of the dosing solution was adjusted by addition of unlabeled 2,3-DCP so that each rat received approx. 5 &i of r4C. Immediately after administration of [‘4C]2,3-DCP, rats were placed in glass metabolism cages (Stanford Glass, Palo Alto, CA) for collection of urine, feces, and expired air. Urine and feces were collected in glass containers on dry ice at 4, 7, 10, 16, 20, 24, 32, 48, 54, and 72 h. Expired air from each cage was pulled at a flow rate of 500 ml/min by a vacuum pump through two gas traps containing 200 ml of 5 M KOH. Samples were removed from the KOH traps every h for the first 4 h, after which time solutions in the traps were sampled and changed at the times of the other excreta collections. At the end of 72 h, rats were killed and blood samples (2 ml) were obtained by
121
heart puncture. Tissues or samples of tissues taken included liver, brain, thyroid, larynx, spleen, thymus, kidneys, adrenals, stomach, small intestine sample, large intestine sample, urinary bladder, testes, fat sample (perirenal and subcutaneous), muscle sample, bone (femur), pelt sample, heart, lung, trachea, remaining GI tract, and carcass. The tissues were weighed, placed in sealed containers, and stored at - 80°C until analysis. Samples of tissues were homogenized in 2 vols of deionized water on ice to prein duplicate samples of vent loss of radiolabeled volatiles. Radiocarbon homogenates was determined by digestion of 300-mg samples of the homogenate with 1 ml of Protosol (New England Nuclear, Boston, MA). Samples were decolorized with 0.5 ml of 30% Hz02 and, after addition of scintillation fluid (Ready-Solv, Beckman, Fullerton, CA), radioactivity in samples was quantitated in a Packard 460C Liquid Scintillation Spectrometer (Packard Instruments, Downers Grove, CA). Quench correction was by automatic external standard, with sufficient counts accumulated to give c 5% error with a 95% confidence interval. Samples of urine and 0.5-ml samples of the 5 M KOH traps were added directly to scintillation fluid for determination of radioactivity. To distinquish 14C02 from exhaled [i4C]2,3-DCP or other volatile metabolites, additional 0.5-ml samples of the KOH traps were acidified with 0.3 ml of concentrated HCl to release CO2 and diluted with 2 ml of deionized water. The residual radioactivity in these samples was considered to be radiolabeled volatiles other than 14C02. Feces were mixed with 5 ml of 1% Triton X-100, vortexed, and 300 mg samples of the homogenate were digested with Protosol and decolorized as described above. Radioactivity in all samples of excreta was determined after addition of scintillation fluid, as described above. Gastrointestinal absorption of 2,3-DCP was calculated by comparing the excretion of radioactivity in urine, feces, and expired 14C02 after p.o. and i.p. administration of [r4C]2,3-DCP as in the following equation: %ABS = 100 - Fp.0, + Fi,p. [(u*.o. +
Ap.o.)/(Ui.p.
+ Ai.p.)],
where U= urinary radioactivity, F= fecal radioactivity, and A = expired 14C02, expressed as % dose excreted during 72 h after either p.o. or i.p. administration of [14C]2,3-DCP. We assumed that the injection of [‘4C]2,3-DCP was equivalent to oral administration with 100% absorption and that &,. was equal to the % radioactivity excreted in feces with 100% absorption of parent compound. Half-times for excretion were determined by least-squares linear regression of the log of the fraction of radioactivity remaining to be excreted as a function of line. T~/z = -0.693/X where X equaled the slope of the regression time. Statistical analyses of data were performed by comparing means using a onetailed Student’s t-test. The level of significance was set at P
3
HOURS
9
AFTER
d
DOSING
HOURS
AFTER
DOSING
c r
0
0
15
30 HOURS
Fig. 1. Cumulative after intraperitoneal sent means
RESULTS
f
45 AFTER
60
75
DOSING
Vo of dose excreted or oral administration
as 14C in urine (a), feces (b), and exhaled as “‘CO2 (c) by rats of 32 mg [‘4C]2,3-DCP/kg body weight. Data points repre-
SEM. n = 3.
AND DISCUSSION
Urine accounted for the majority of 14C excreted after administration of [i4C]2,3-DCP, with 66 and 75% excreted by this route after p.o. or i.p. administration, respectively (Fig. la; Table I). The half-times for excretion were not significantly different after i.p. or p.o. administration being 4.9 +_ 1.3 h and 7.5
123 TABLE I DISTRIBUTION OF 14CAT 72 h AFTER ADMINISTRATION (2,3-DCP) Route of administration
Intraperitoneal injection Oral administration
OF [‘4C]2,3-DICHLOROPROPENE
% Dose found in Urine
Feces
Exhaled CO2
Exhaled 2,3-DCP
Carcass and tissues
75+ ia 66+0.4
13kI 21+ 1
7kO.4 8kI
2kO.2 2+0.1
3kO.6 2kO.I
aData represent means and SEM from 3 rats.
+ 0.6 h (mean f SE), respectively. Radioactivity was also found in the feces (Fig. lb; Table I) and in the expired air as 14C02 (Fig. Ic; Table I). Similar studies of [14C]cis-1,3 and trans-1,3-DCP demonstrated that urine was also the major route of excretion of 14C for these compounds after p.o. administration [6]. Most of the radioactivity administered was eliminated during the first 24 h of the experiment, similar to what was seen with our studies on 2,3-DCP (Fig. 1). Climie et al. [7] identified the major urinary metabolite (92% of radioactivity excreted in urine) of c&l ,3-DCP in rats as N-acetyl-S-[3-chloroprophenyl] cysteine. Metabolism involved formation of a carbon sulfur bond between glutathione and 1,3-DCP, with loss of a Cl atom from the carbon-3 position. The authors proposed that the DCP’s were eliminated in urine because of this efficient glutathionedependent biotransformation. In our study, fecal excretion of 14C was significantly higher (21%) after p.o. administration of [14C]2,3-DCP, compared to i.p. (13%). This difference was probably due to unabsorbed 2,3-DCP. Calculation of GI absorption indicated that 91% of the administered dose was absorbed from the GI tract. Hutson et al. [6] found that only 2-3% of an oral dose of [14C]cis or trans-1,3-DCP was excreted in the feces 4 days after administration, This is less than what we found even after i.p. administration suggesting possible differences in either biliary excretion or enterohepatic circulation between the 1,3-DCPs and 2,3-DCP. In the studies reported here, acidification of samples obtained from 5 M KOH traps decreased the radioactivity remaining in the samples, indicating that most of the 14C in the expired air was due to 14C02 (8% of dose). Less than 2% of the dose was due to acid-stable volatile compounds, which could include unmetabolized 2,3-DCP. Hutson et al. [6] also found little volatile radioactivity other than 14C02 in the expired air of rats given cis- or truns-1,3-DCP. These authors did find a difference between the cis and ~rans isomers in the amount of radioactivity exhaled as r4C02 [6]. Studies with c&1,3- and truns-1,3-DCP showed 5 and 23% of the dose excreted as i4C02, respectively. In studies on 2,3-DCP, we found that approx. 8% of the dose was exhaled as r4C02 after p.o. administration of 2,3-DCP suggesting that in this respect, the pathways for metabolism of 2,3-DCP may be more like the cis-1,3-DCP than trans-1,3-DCP.
124
Similar to what was seen with the cis- and trans-1,3-DCPs [6], from 2-3% of the radioactivity associated with 2,3-DCP remained in the rats at 72 h after dosing (Table I). Major organs for retention of radioactivity associated with 2,3-DCP included the liver, kidney, testes, lung, and brain (Fig. 2). The liver accounted for approximately 20% of the 14C remaining at the end of 72 h. There appeared to be no differences in the retention of 14C, between the two routes of administration studied. When expressed on a per-gram basis, tissues in which concentrations of 14C found were higher than concentrations found in carcass (approx. 8 nmol/g) included the adrenals, spleen, and turbinates (19 to 16 nmol [i4C]DCP equivalents/g tissue) in addition to tissues shown in Fig. 2. In summary, the results of studies reported here indicate that the disposition of 2,3-DCP is similar to that found for the cis-1,3-isomer, with most of the radioactivity excreted within 24 h in the urine and a smaller fraction excreted as COZ. Although most of the ingested 2,3-DCP will be absorbed, any absorbed 2,3-DCP will be rapidly excreted with no apparent accumulation in any tissue.
TISSUE
Fig. 2. nmol of [‘4C]2,3-DCP equivalents present in tissues at 72 h after oral administration (solid bars) or intraperitoneal injection (hatched bars) of 32 mg [?]2,3-DCP/kg body weight. Each bar represents mean + SEM, n=3.
125
ACKNOWLEDGEMENTS
Research conducted as part of the National Toxicology Program under U.S. Department of Energy (DOE) contract number DE-AC04-76V01013 and interagency agreement with the National Institute of Environmental Health Sciences. The facilities used are fully accredited by the American Association for Accreditation of Laboratory Animal Care. The authors gratefully acknowledge the excellent technical assistance of Ms. Frances Straus and Ms. Eileen Cahill and the useful discussions with a number of our colleagues at the Institute, with particular appreciation to Drs. R.O. McClellan and R.F. Henderson. REFERENCES 1 J.P.
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