Document JNXorg7G1erLZOgaw4KV7mdve

TOXICOLOGY AND aPMJED PHARMACOLOGY 68, 140-151 (1983) The Uptake and Disposition of 1,1-Dichloroethylene in Rats during Inhalation Exposure1,2 C. E. Dallas,* F. W. Weir,* S. Feldman,t L. Putcha4 and J. V. brucrnerJ-3 `Environmental Science Discipline. The University of Texas School of Public Health Houston. Texas tDepartment of Pharmaceutics. University of Houston. College of Pharmacy. Houston. Texas, and tDi vision of Toxicology. Department of Pharmacology. The University of Texas Medical School. Houston, Texas 77025 -.filed October 2. 1982, accepted November 24 1982 The Uptake and Disposition of 1. 1-Dichloroethylene in Rats during Inhalation Exposure. Dallas, C. E., Weir. F vx r. ldman. S., Putcha, L., and Bruckner, J V (1983). Toxicol Appl Pharmaco' 6` J' 'rnc uptake, disposition, and respirators' elimination of 1.1- dichloroethyi ,:.nr,g inhalation exposure were evaluated to gain insight into the pharmacoC 150. or 30c naiocarbon. Anesthetized male Sprague-Dawley rats inhaled 25. 75. for 3 hr from an aluminized Mylar hag through a miniaturized one way breath . , into the trachea Periodic air samples were taken immediately adjacent to the f me separate inhaled air and exhaled breath streams concurrently with blood samples fron. o cannuiated femoral vein and analyzed for 1,1-DCE content by gas chro matography. 1,1-DCE was absorbed very rapidly, in that substantial levels were present in the venous blood at the firs sampling time (i.e.. 2 min). Percentage systemic uptake decreased over time after initiation of exposure until equilibrium was established. Percentage uptake after reach ing equilibrium varied inversely with the exposure concentration. 1,1-DCE venous whole-blood in animals exposed to 25, 75. and 150 ppm l.l-DCE increased rapidly to near steady state - approximately 45 min, as did concentrations of 1,1 -DCE in the exhaled breath and alveolar 4.U calculation of the amount of 1,1-DCE- taken up by the body over the course of the 3-hr exposures revealed that cumulative uptake of the inhaled chemical was statistically linear for the 25-, 75-, and 150-ppm exposures Accumulation plots for 300-ppm exposed animals, how ever, were best fitted to a cubic curv-: Although trends toward the establishment of equi librium were initially seen in the 300-tr :'. exposed animals, levels of 1,1-DCE in the blood and breath rose progressively during the latter hour of the 3-hr exposure period. Thus, despite in creased exhalation of l.l-DCE. these animals could not prevent systemic accumulation of the chemical. Dichloroethylene (1,1-DCE). also known as vmylidene chloride, is used wideb as a monomer in the manufacture of a v ariety of plastic materials. In addition to the potential for exposure in the occupational environ ment, 1,1-DCE and other halocarbons are contaminants of drinking water supplies (U.S. EPA, 1975, 1977), ' Presented in pan at the annual meeting of the Society of Toxicology in Boston, Mass., February 1982. 1 Supported by U.S. EPA Grant R808282 and NIEHS Training Gram ES07090. 1 To whom reprint requests should be sent. The pharmacokinetics and metabolic fate of 1,1-DCE are of considerable interest, in that both toxicity (Andersen and Jenkins, 1977; Andersen et al., 1979a) and carcino genicity (Maltoni el al., 1977) are highly dose dependent. Maltoni et al. (1977), for example, saw no increase over controls in tumor inci dence in male mice exposed to 10 ppm of 1,1-DCE vapor, bui a significant incidence in 25-ppm exposed animals. McKenna et al. (1978a) reported that rats subjected to 10 ppm 1,1 -DCE for 6 hr were capable of metaboliz ing about 98% of the systemically absorbed 0041-008X/83 $3.00 C 1913 by AcWtfnie TTfir Inc Att nghu oi rvirottycuon i *nv form reaerxd 140 dose to nom ola tabolites. Rats s metabolized a s sorbed dose ant as unchanged 1 posed to 200 pp creased hepatic as well as liver; and Jenkins (lc and Jones and ported similar < terns for 1.1 -Df to the hypothe animal's metal detoxification i Investigators question of sati employing ind take during in Bolt (1979) an posed rats to systems and chamber conct sure of svsterr course of upu a number of metabolism. I modeling has; uration kineu pharmacokint ongoing expo limited to the Hepatotoxicn early as 2 hr : 200 ppm 1.1-i would appear .pathway ma; sure, leading icity. Therefo of the curren exposure cor metabolism measuremen and blood sa Since alter of 1,1-DCE elimination SL 066620 UPTAtCE/DlSPOSmON OF 1,1-D1CHLOROETHYLENE 141 dose to nonvolatile, apparently nontoxic me tabolites. Rats subjected to 200 ppm for 6 hr metabolized a smaller percentage of their ab sorbed dose and exhaled a larger percentage as unchanged i, 1-DCE. Nevertheless, rats ex posed to 200 ppm [MC]1.1-DCE exhibited in creased hepatic covalent binding of radiolabel as well as liver and kidney damage. Andersen and Jenkins (1977), McKenna et al. (1978b), and Jones and Hathway (1978) have also re ported similar dose-dependent excretion pat terns for 1,1-DCE. lending additional support to the hypothesis that high doses exceed the animal's metabolic capacity and associated detoxification mechanisms. Investigators have recently dealt with the question of saturable 1,1 -DCE metabolism by employing indirect measures of 1,1-DCE up take during inhalation exposures. Filser and Bolt (1979) and Andersen et al. (1979b) ex posed rats to 1,1-DCE in closed inhalation systems and periodically monitored the chamber concentration of 1,1 -DCE as a mea sure of systemic uptake. The resulung time course of uptake curves were used to derive a number of kinetic constants for 1,1-DCE metabolism. Although this method of kinetic modeling has provided useful estimates of sat uration kinetics, there have been no direct pharmacokinetic studies of 1,1-DCE during ongoing exposures. Direct studies have been limited to the elimination phase of exposure. Hepatotoxictty, however, has been seen as early as 2 hr after fasted rats begin to inhale 200 ppm 1,1-DCE (Reynolds et al., 1975). It would appear that saturation ofdetoxification pathways may occur during ongoing expo sure. leading to altered disposition and tox icity. Therefore, one of the primary objectives of the current investigation was to define the exposure conditions under which saturable metabolism can occur, by direct, repetitive measurement of 1,1-DCE in exhaled breath and blood samples. Since attention in pharmacokinetic studies of 1.1-DCE has been focused primarily on elimination of the chemical, information on its uptake and disposition is lacking. One might predict that uptake and disposition of 1,1-DCE will be dependent upon the com pound's basic physical/chemical properties and upon physiological processes which gov ern the kinetics of anesthetic gases (Goldstein et al., 1974), The onset of cytotoxicity and alterations in metabolism during the course of exposures to 1,1-DCE. however, would be anticipated to after the disposition and ulti mately the uptake of the chemical. Thus, a second major objective of the project was to characterize the uptake and disposition of in haled 1,1-DCE and to determine whether the pharmacodynamics ofthe halocarbon changes during ongoing t sures. METHODS Animats. Adult, male Sprague-Dawley rats were ob tained from Ttmco Breeding Laboratories (Houston. Tex.). The animals were maintained on a constant reverse lightdark cycle, with light from 2200 to 1000 hr and darkness from 1000 io 2200 hr. They were housed in solid-bottom polypropylene cages fined with vented stainless-steel lids. Tap water and Ralston Purina Fonnulab chow were pro vided ad libitum. The rats were used after a 14-day ac climation period, at which time their body weight ranted from 350 to 400 g. Solvent exposures were initiated at the same ume for each animal (at 0100 hr or 3 hr after the beginning of the dark cycle). Test material. 1,1-Dichloroethylene (I. I-DCE) (98% minimum punty) was obtained from Mathcson, Coleman and Bell (Norwood, Ohio.) Animal preparation. The rats were anesthetized by im injection of 0.2 ml of a mixture consisting of ketamine HG:acepromazine maleaten ylazme HG (3:2:1, v:\iv). A tracheostomy was then performed and a minaturized one way breathing valve inserted directly into the trachea. Since such valves were not commercially available, i vahe was designed and fabricated for the study. It consisted of a T-shaped polypropylene tube (6.4 x 0.95 cm i.dL). with a rubber septum set on either side ofa centrally positioned l-cm side arm. A tapered plastic pipet was attached to the side arm. and the pipei tip inserted into the trachea. The septa were positioned so that as the animal inhaled, the inlet septum opened, and the outlet septum dosed. Upon exhalation the flow of air forced the outlet septum open and the inlet septum closed. Thereby, separate air ways were established for the inhaied and exhaled breath streams. The septa were made of flexible latex rubber, im order that they offer minimal resistance to air flow. The valve's dead space (0.75 ml) was comparable to that et SL 066621 142 DALLAS ET Al_ rats of the weight range utilized. The femoral vein of each animal was cannulated so that samples of venous blood could be obtained for 1.1-DCE analysis. A temperature probe was inserted into the rectum so thai core temper ature of the anesthetized animal could be monitored and regulated by means of variable temperature heating pad. The cardiac rate and blood pressure were monitored in four subjects over a 3-hr period and found to be stable. The mean values c SD were: cardiac rate, 390 20; sys tolic pressure, 115 t 8.7; and diastolic pressure, 90 p 6 4. These values are similar to those measured in harnesstrained (Hunt and Kumeldorf, I960) and m unanesthetizcd. unrestrained (Popovic and Kent, 1964) rats. Inhalation system. A known concentration of 1,1-DCE was generated in a 70-ltter aluminized Mylar bag, which was connected in senes by Teflon tubing with a pneu motachograph, the breathing valve, and an empty 70-liler Mylar bag The empty bag served as a reservoir to ac cumulate exhaled gas. Thereby a closed system was cre ated to prevent release of the agent into the laboratory. Samples of the exhaled breath and inhaled air were taken with a gas-tight synnge from sampling ports immediately adjacent to the breathing valve. The animal and the area between the two Mylar bags, including all valve and tub ing connections, were located under a ventilation hood with an exhaust fan to further minimize potential con tamination of the laboratory Experimental procedure. After the breathing valve was inserted into its trachea, the animal was allowed to adjust to breathing through the valve for approximately 10 min (without solvent exposure) until consistent, stable breath ing patterns were established. For individual exposures, a concentration of 25. 75. 150, or 300 ppm of 1,1-DCE was generated in the influent Mylar bag. Once breathing was stabilized, the Mylar bag with the 1,1-DCE was con nected by Teflon tubing to the valve and thus to the animal for a 3-hr exposure period. Blood samples were periodically withdrawn from the femoral vein over the course of the exposure. Samples of exhaled breath were taken at 2- to S-min intervals during this period, while samples of the inhaled air were monitored at 30-rain in tervals. After 3 hr, the bag containing 1,1-DCE was dis connected. and the animal was allowed to breathe room air for 30 min while postexposure blood samples and exhaled breath samples were taken. Respiratory measurements and calculations. The res piration of each animal was continuously monitored. This procedure was accomplished by measuring the airflow, created by respiration of the animal, with a size No. 0 Flash No. 7318 pneumotachograph connected to a Narco No. 7172 strain gauge coupler of a Narco physiograph. The magnitude ofeach respiration was recorded, and each respiratory rate signal was accumulated for a I-min in terval with a Narco GPA-10 integrator unit. The resulting integrated signal was then continuously recorded as vol ume of ruspintton per minute (P*) with a Narco No. 7173 transducer in a second channel of the physiograph. Mean values SD over the course of the 3-hr exposures for the 16 rats utilized in the study were; KE - 221 * 28 ml/min; respirations per minute (/) * 145 21/min; tidal volume ( Kt) * l.Jc 0.2 ml These values are quite similar to val ues for unanesthetized rats (Guyton. 1947) and for tracheosiomized rats anesthetized with thiamylal (Johanson and Pierce, 1971). While exhaled breath levels of 1,1-DCE were measured from air samples taken at the sampling port, alveolar lev els of the solvent could be calculated from the exhaled breath data and the respiratory rate and A certain amount of the inhaled gas containing 1.1-DCE resides in the dead space of the valve and is exhaled without par ticipating m ajveolar gas exchange. By correcting for the contribution of this dead space volume to the exhaled breath 1,1-DCE concentration, an approximation of the ajveolar levels of 1,1-DCE could be calculated (Kelman, 1982). Since the I e and the alveolar concentration of 1,1 -DCE at each sampling point were known, subtraction of the concentration of 1,1-DCE in the alveolar air from the inhaled concentration yielded an approximation of the quantity of compound which remained in the body for each sampling period. Thereby it was possible to monitor percentage systemic uptake and to determine cumulative uptake of 1 .l-DCE over the course of the 3-hr exposure. Analysts of 1,1-DCE in air and blood. The concentra tions of 1,1-DCE in the inhaled and exhaled air were measured with a Perkin-Eimer 3920 gas chromatograph equipped with a flame ionization detector. Air samples of 1.0 ml were taken from the sampling pons with a gastight syringe and injected directly onto a 6-ft X 1/8-in. stainless-steel column packed with 10* FFAP on 80/100 mesh Chromasorb. Operating temperatures were: I60*C, injection pon; 220'C, detector, 109'C, isothermal col umn operation. Chromatographic grade nitrogen was used as the carrier gas with a flow of 30 ml/min. Maximum sensitivity of this detector was maintained with settings of 16 psi for hydrogen and 60 psi for the zero air. 1.1 -DCE levels in the blood were measured by gas chro matographic headspace analysis. A 0.1-ml blood speci men was withdrawn through a stopcock, connected to an indwelling femoral cannula, into a I-ml tuberculin sy nnge. and injected into chilled 1-ml air-tight vials. An equal volume (0.1 mil of heparinized saline was then injected back through the stopcock to replace the lost volume The blood samples were kept under refrigeration overnight, then placed into a 55*C water bath to volatilize the 1,1-DCE (boiling point, 32 C). After allowing 30 mm for equilibrauon of the 1,1-DCE between the blood and head space in the vial, a 0.1-ml sample of the bead space was taken with a gas-tight synnge and injected directly onto a 6-ft x 1 /8-in. stainless steel column packed with Durapak in a Hewlett-Packard gas chromatograph equipped with a **Ni electron capture detector. Operating temperatures were: I50*C, injection port; 250*C, detec tor. 80*C, column oven. The flow rate for nitrogen, the Fig. i. L to 25, 75, measured a for groups carrier gas, was 1,1-DCE in the curve generated known concent Statistical an. analyzed by regr examination of p of animals. By possible to deter cumulative upta no longer linear to describe cun nation was dei assumed straig 1978) of the po level. By use c appropriate tag repealed obser determined. TT SL 066622 UPTAKE/DISPOSITION OF 1,1-DICHLOROETHYLENE 143 Fig. I. Levels of 1,1 -DCE in the exhaled breath during and after inhalation exposures. Rats were exposed to 23, 73, 150, or 300 ppm 1,1-DCE for 3 hr. Concentrations of 1.1-DCE in the exhaled breath were measured at 2- to 8-min intervals during the exposure and for 30 min thereafter. Brackets encase i e SD for groups of four animals. earner gas, was 25 to 28 ml/min. The concentration of 1,1 -DCE in the blood was determined from a standard curve generated daily from blood samples containing i known concentration of 1,1-DCE Statistical analysts of accumulation plots. Data were analyzed by regression analysis and tested for linearity by examination of plots for individual animals and for groups of animals. By examination of individual plots, it was possible to determine the duration of exposure and the cumulative uptake of 1,1-DCE beyond which uptake was no longer linear. The appIbpriateneM of a linear model to describe cumulative uptake at each exposure concen tration was determined by testing for lack of fit of an assumed straight-line model (Kkinbaum and Kuppcr, 1978) of the pooled data of test animals at each exposure level. By use of the residual sum of squares from the appropriate regression analysis, pure error estimates from repeated observations for each concentration could be determined. Tbe level of significance was set at p < 0.05. RESULTS The 1,1-DCE exposure concentrations in the inhalation airway immediately adjacent to the breathing valve were determined at 30min intervals during the exposures. The ac tual values, following each desired concentra tion, represent the mean SE for 24 deter minations per exposure level: 25 ppm, 23.7 1,4 ppm: 75 ppm. 77.1 1.8 ppm; 150 ppm, 145.9 0.6 ppm; and 300 ppm, 295.1 3.8 ppm. Concentrations of 1,1-DCE in the exhaled breath over the course of the 3-hr exposure are shown in Fig. I. The exhaled breath con centrations during the initial min of exposure SL 066623 144 DALLAS ET AL. Fic 2. Alveolar concentrations of l.l-DCE during and after inhalation exposures. Rats were exposed to 25. 75, 150. or 300 ppm l.l-DCE for 3 hr. Concentrations of l.l-DCE in the alveolar air were determined at 2- to S-mtn intervals during the exposure and for 30 min thereafter. Brackets encase i c SD for groups of four animals. were about 60% of the inhaled concentra tions. Levels of 1,1 -DCE in the exhaled breath of animals inhaling 25. 75, and 150 ppm rose until near steady-state levels were achieved within about 30 min. The curves for these exposure groups were asymptotic, in that they continued to gradually increase throughout the 3-hr exposure period. Although exhaled breath levels in rats inhaling 300 ppm sug gested that a near-equilibrium state was es tablished between hours 1 and 2, the levels increased slightly during the last hour of the experiment. Upon cessation of 1,1-DCE in halation, the concentration of 1,1-DCE fell very rapidly m the expired air of all exposure groups. Conversion of the exhaled breath data to alveolar concentrations yielded more defini tive information on the nature of 1,1-DCE uptake. As can be seen in Fig 2. the pattern of 1.1 -DCE uptake was distinctly dose depen dent. At the 25-ppm exposure level, appar ently all the chemical presented to the animal was retained during the first 20 min. There after, the chemical appeared in the alveolar air in increasing amounts until near steady state was reached after about 45 min. Animals presented with 7 5 ppm 1,1 -DCE appeared to Fic 3 P 75. 150. or represents t for sake of absorb all the while the 15C retained all b DCE. The tintwo intermed sembled that At equilibriui these groups sure level. Th; exposed anim this group ro; level over the a trend towar during the sr siantial incre these animal: 45 min. The rapidly in all tinuation of Percentage both dose ar SL 066624 UPTaK/D1SPOSIT)ON OF 1.1-DICHLOROETHYLENE 145 Fig, 3 Percentage systemic upuke or 1.1-DCE dunng inhalauor exposures. Rats were exposed to 35. 75. 150. or 300 ppm 1.1-DCE for 3 hr. Percentage upuke was determined at 8-mm intervals. Each point represents the mean percentage upuke in four animals per group. Sundard deviation brackets are omitted for sake of danty. absorb all the chemical for the first 10 min, while the 150-ppm exposed animals initially retained all but a fraction of the inhaled 1,1DCE. The time course of equilibration at these two intermediate dose levels very closely re sembled that of the 25-ppm exposed animals. At equilibrium the alveolar concentrations in these groups were proportional to the expo sure level. This was not true for the 300-ppm exposed animals. Alveolar concentrations in this group rose to a disproportionately high level over the 3-hr period. Though there was a trend toward establishment of equilibrium during the second hour of exposure, a sub stantial increase in alveolar concentrations in these animals occurred during the final 30 to 45 min. The alveolar concentrations fell very rapidly in all exposure groups upon discon tinuation of 1,1 -DCE inhalation. Percentage systemic uptake of 1,1-DCE was both dose and time dependent (Fig. 3). Per centage uptake in each exposure group de creased markedly over the first 45 min of ex posure and approached near steady state dur ing the following hour. Uptake varied inversely with exposure level, though the magnitude of difference among the 25-, 75-, and 150-ppm groups was quite modest. Average percentage uptake (SD) in these groups over the last 90 min of the 3-hr exposure was as follows: 77,2 0.9%, 25 ppm; 75.7 1.1%, 75 ppm; and 72.5 1.9%, 150 ppm. Approximately 60% of inhaled l, 1 -DCE was retained by the 300ppm exposed animals at 2 hr. Thereafter, pul monary retention in the 300-ppm exposed an imals decreased progressively. Plots of the cumulative uptake of 1,1-DCE during the 3-hr exposures revealed the rates and patterns of accumulation to be concen tration dependent. Figure 4 is a plot of mean cumulative uptake over time for each of the four exposure groups. Cumulative uptake ap peared to be relatively linear during the 3-hr SL 066625 146 Dallas et al. Fig. 4. Cumulative uptake of 1,1-DCE during inhalation exposures. Rats were exposed to 25, 75, 150, or 300 ppm 1,1-DCE for 3 hr. The quantity of 1,1-DCE retained dunng successive 8-min intervals was calculated on the basis of the minute volume and difference between inhaled and alveolar I, I -DCE con centrations. Each point represents the mean value of four animals per group. Standard deviation brackets are omitted for sake of clarity. The pooled data of the test animals at each exposure level were tested for lack of fit to an assumed straight-line model of uptake. exposure in the exposure range of 25 to 150 ppm. Statistical analysis revealed no signifi cant deviation from linearity at the 25- and 75*ppm exposure levels. There appeared to be some departure from linearity, however, during the final hour in the 150-ppm exposed animals. Examination of the cumulative data plots of each of the 4 animals in the group revealed that the rats responded somewhat dif ferently to 150 ppm 1,1-DCE. Plots of two of the animals displayed departure from linear ity after about 2 hr of exposure, one rat, after 2 1 /2 hr and one rat not at all (data not shown). Despite this finding, testing of the pooled data (for the 3 hr of 150-ppm exposure) for lack of fit of a straight-line model showed there to be no significant deviation from the linear model. In contrast, the pooled 300-ppm data did show deviation from the linear model. Indeed, regression analysis determined that a Fic. 5. A exposure to that is direc the lines. Bi cubic model mulauve upta imals. The rate oi tity of 1,1-DC exposure are Fig. 5. It can rate and toia proportional exposure ran true for the 3 the values for would be pr tionality to < SL 066626 UPTAKE/DISPOSITION OF 1,1-DlCHLOROETHYUNE 147 FlG. 5. Average rate of accumulation and total quantity of 1.1-DCE retained over 3 hr of inhalation exposure to 25.15, 150. or 300 ppm 1,1-DCE. The straight lines represent the magnitude of each parameter that is directly proportional to dose. The 25-ppm values are used as the point of ongin for derivation of the lines. Brackets encase it SO for groups of four animals. cubic model was the best description of cu mulative uptake in the 300-ppm exposed an imals. The rate of accumulation and total quan tity of 1,1-DCE retained during the 3 hr of exposure are plotted against exposure level in Fig. S. It can be seen that both accumulation rate and total 1,1-DCE retained were directly proportional to inhaled concentration in the exposure range 25 to 150 ppm. This was not true for the 300-ppm exposed animals, in that the values for both parameters were lower than would be predicted on the basis of propor tionality to dose. The time course of venous whole-blood levels of l,l-DCE in the animals is shown in Fig. 6. Substantial concentrations of 1,1-DCE were found in the blood of all animals at the first sampling time (2 min). Concentrations of 1,1-DCE in the blood of animals inhaling 25, 75, and 150 ppm increased rapidly, reach ing near steady-state levels after about 45 min. The mean (SD) blood levels during the final 30 min of exposure were 180 28 Mg/ml for 25 ppm, 348 45 ug/ml for 75 ppm, and 472 36 fig/nil for 150 ppm. Near steady state or equilibrium was never established in the 300-ppm exposed animals. The concentra- SL 066627 148 Dallas et al. Fig 6. Venous blood levels of I.l-DCE during end ifier inhalation exposures. Rats were exposed to 25, 73, 150, or 300 ppm 1,1-DCE for 3 hr. Concentrations of I.l-DCE in whole venous blood samples were measured at 2- to 30-min intervals during the exposures and for 30 min thereafter. Brackets encase x SD for groups of four animals. tion of 1,1-DCE in the blood of these animals progressively increased over the course of the 3-hr exposure, reaching levels eightfold higher than levels found in the 150-ppm exposed an imals. Although 1,1-DCE blood levels in all groups fell rapidly upon cessation of 1,1-DCE inhalation, the decreases were not so dramatic as the falls in exhaled breath and alveolar con centrations. DISCUSSION Pharmacokinetic data based upon direct measurements of 1,1-DCE and other halo- carbons during exposures are limited. Most pharmacokinetic studies of halocarbons per tain to metabolism and elimination of the chemicals. Investigators typically utilize re labeled halocarbons because of technical dif ficulties in quantifying the volatile parent compounds. In such instances, the investi gator is largely unable to distinguish among parent compound, metabolite(s). and MC which enters the metabolite pool. Astrand (1975) summarizes the results of several stud ies in which levels of parent halocarbons were monitored in human subjects' alveolar air and blood during; ies provide us< elimination c though emph cokmctic inte man studies ; ations, in tha ister levels hig kinetics nor s propriate am ployed in tht used to advan In the abse and dispositio speculate tha' water-insolub current invest Since 1,1-DC soluble molec across membi bed into the s; stantial levels venous circuit (2 min after i other gaseou 1974), the ne alveoli to bio but become [ accumulates pattern was systemic upu age uptake o decreased ov: relatively co proach to eqi lar air was ra low solubility perfusion of t deposition of ited solubilit; demonstrate; alveolar cone exposures. E asymptotic.' librium can tabolism of 1 sues. Deposi SL 066628 UPTAKE/DISPOSITION OF 1,1 -DJCHLOROETHYLENE 149 blood during and after exposures. These stud ies provide useful informauon on uptake and elimination characteristics of halocarbons, though emphasis is Dot placed on pharma cokinetic interpretation of the data. Such hu man studies are limited by ethical consider ations, in that one normally canDOt admin ister levels high enough to produce saturation kinetics nor study toxic halocarbons. An ap propriate animal model, such as that em ployed in the current investigation, can be used to advantage under these circumstances. In the absence of knowledge of the uptake and disposition ofinhaled 1,1-DCE, one might speculate that the chemical will behave as a water-insoluble anesthetic gas. Findings in the current investigation support this hypothesis. Since 1,1-DCE is a small, uncharged, lipidsoluble molecule, it should be absorbed readily across membranes in the pulmonary capillary bed into the systemic circulation. Indeed, sub stantial levels of 1,1-DCE were found in the venous circulation at the initial sampling point (2 min after initiation of exposures). As with other gaseous anesthetics (Goldstein et ai,, 1974), the net transfer rate of 1,1 -DCE from alveoli to blood should initially be very rapid but become progressively slower as the agent accumulates in the blood and tissues. This pattern was reflected by the time course of systemic uptake of 1,1-DCE, where percent age uptake of 25, 75, and 150 ppm 1,1-DCE decreased over time from virtually 100% to a relatively constant steady-state level. Ap proach to equilibrium in blood and in alveo lar air was rapid, indicative of tbe relatively low solubility of 1,1-DCE in blood and poor perfusion of adipose tissue, the major site of deposition of lipophilic compounds. The lim ited solubility of 1,1-DCE in blood was also demonstrated by the dose dependency of the alveolar concentrations at the initiation of the exposures. Equilibration of tbe blood was asymptotic. The delay in attainment of equi librium can be attributed largely to the me tabolism of 1,1-DCE and its deposition in tis sues. Deposition in lipid depots very likely played a more important role during deequili bration. Lipids should release 1,1-DCE more slowly than other tissues, thereby prolonging the presence of the compound in the blood stream. Although we did not monitor blood levels long enough after the exposures to ac curately define the terminal elimination phase, we did observe that 1,1-DCE was present in venous blood longer than in exhaled air. Findings in the current study indicate that the rat's capacity to metabolize and eliminate 1,1-DCE can be exceeded during tbe course of inhalation of 150 ppm or more of 1,1 -DCE. Inhalation of 300 ppm definitely exceeds the animal's capability to assimilate the chemical. Despite evidence after the initial hour of in halation of 300 ppm of the approach of steadystate conditions, 1,1-DCE levels in the blood and alveolar air increased substantially during the final hour of the 3-hr exposures. Signifi cant departures from linearity during the final hour in the plots of cumulative uptake of 1.1DCE also indicate saturation of the animals' elimination mechanisms at 300 ppm. Al though deviation from linearity in three of four 150-ppm exposed animals is not suffi cient to be statistically significant, this finding nevertheless suggests that inhalation of 150 ppm 1,1 -DCE for some 2 hr can also result in saturation kinetics in the rat. This level is in agreement with the 150-ppm saturation point Fiber and Bolt (1979) determined by indirect measurement of 1,1-DCE uptake in rats. Thus, it would be expected that higher vapor concentrations of 1,1-DCE exceed the rat's capacity to metabolize 1,1-DCE. Fiber and Boll (1979) note that 1,1-DCE exhibits a high V__relative to most other halocarbons they tested. On the basis of a similarly de signed study, Andersen et al. (1979b) calcu lated the chamber l7TM, for 1.1 -DCE to be 132 ppm/kg/hr, corresponding to an in vivo of 15.87 mg 1,1 -DCE metabolized/kg/kr. The values are in general agreement with findings in the present study. In plots of cumulative uptake iD the 150-ppm exposed animals, we observed departure from linearity after about SL 066629 150 DALLAS ET AL 2 hr, at which time there was an average total uptake of 1,1-DCE of approximately 23 mg/ kg. As would be anticipated, departure from linearity occurred sooner (after 1 to 1 1/2 hr) in animals inhaling 300 ppm. Average total uptake of 1,1 -DCE at this time was approx imate^ 26 mg/kg. Thus, metabolic saturation seemed to occur during the course of an ex posure when there had been systemic uptake of a finite, limited quantity of 1,1-DCE. it is possible that factors other than met abolic saturation may play a role in the sat uration kinetics observed here. Inhalation of 200 ppm 1.1-DCE for 1 to 2 hr caused a marked reduction in glutathione (GSH) levels in the livers of fasted rats (Reynolds et al., 1980). Although GSH is required in the major 1,1-DCE metabolic pathways (Jones and Hathwav, 1978; Reichert el al., 1979). it is not entirely clear whether a decrease in he patic GSH. per se, will result in a net reduc tion in 1,1-DCE metabolism. Although over night fasting is known to deplete hepatic GSH in rats (Jaeger et al., 1973), Andersen ei al. (1979b) reported no difference in the 1.1-DCE k'rnw between fed and fasted rats. Despite a 60% decrease in GSH content, isolated per fused livers from fasted rats are able to me tabolize 1,1-DCE as rapidly as livers from fed rats (Reichert et al., 1978). These investiga tors did see a modest decrease in 1,1-DCE metabolism in the in vitro system after a diethylmaleate pretreatment which reduced the GSH content by almost 90%. Andersen et al. (1980) also find diethylmaleate and certain other agents which substantially depress he patic GSH to inhibit 1,1-E>CE metabolism by rats. Although these findings suggest that very marked GSH depletion may result in some decrease in net 1,1-DCE metabolism, it ap pears more likely that saturation of the pri mary "activating" enzyme system is respon sible for the altered kinetics seen in high-dose animals in the present study. Hepatotoxicity may also play a role in the saturation kinetics phenomenon. Evidence of cytotoxicity has been seen in livers of lasted rats after 2 hr of inhalation of 200 ppm 1.1DCE (Reynolds et al., 1975, 1980). Frank hemorrhagic centrilobular necrosis was pres ent after 4 hr. Although fasted rats were not utilized in the current study, the reverse lightdark cycle animals we used were exposed to : ,1-DCE during the period when liver GSH levels are lowest (Jaeger et al.. 1973). There fore. it was quite possible that some degree of hepatic injury occurred in the high-dose :nimals during the latter pan of the 3-hr \posures. Damage of the endoplasmic reticu lum of hepatocytes would diminish their ca pacity to metabolize 1.1-DCE. Although Andersen et al. (1979a) saw evidence of this condition within 30 min in immature rats subjected to 200 ppm 1,1-DCE. relatively lit tle effect was seen in mature rats. Reynolds et al. (1980) observed only slight effects on hepatic microsomal parameters in mature rats after 2 hr of inhalation of 200 ppm 1,1-DCE. Not until hepatotoxicity was fully manifest 6 hr after the beginning of the 4-hr, 200-ppm exposure were significant decreases in the mi crosomal parameters manifest. Thus, it would appear that 1,1-DCE-induced deactivation of functional components of the mixed-function oxidase system may not contribute signifi cantly within the ume frame of the .current study to the observed saturation kinetics. Findings in the present study indicate that the rat has a limited capacity to metabolize and eliminate 1,1 -DCE. Since 1,1 -DCE is vol atile and relatively insoluble in blood, in creased amounts of the chemical can be readily eliminated via the lungs upon the onset of metabolic saturation. However, despite in creased exhalation of 1,1-DCE, we observed a progressive rise in blood ievels in the 300ppm exposed animals. The high-dose animals may be at risk from toxic injury, while those inhaling the lower concentrations may tol erate their exposures. Indeed, other investi gators (Jaeger et al., 1974; Andersen et al., 1979a) have reported the threshold for hep atotoxicity in fasted rats to be in this same exposure range. Although the dose-response curves for hepat 1979a) and for It kins. 1977) are in plateau. The abst toxicity despite i uted to saturatioi of 1.1-DCE to tc ent study metabt ing the course c more of 1,1-DC mined whether tl forded protectio they will experie ney, or other or; R ANDERSEN. M. E_ F R. A., AND jENKJ tabolism and the a Toxicol App! Ph Andersen. M. E.. Jenkins, L J.. Jr tuques to assess t ethylene meiabol 395-^09. ANDERSEN, M. E.. toxicity of 1,1 -dt sex, ate, and fasti 163. ANDERSEN, M E.. 7 R. A., and Jenk. of multiple deto ttbolues in the ti col. App! PHorm Astrand, 1. (1975 Ussues of man. Health 1. 199-7 Filser, J. G.. ani ueucs of halogen. 42, 123-136. Goldstein, a., ARt Principles ofDru. 2nd ed.. pp. 338Guyton, a. C. (ic volumes of labor 70-77. Hunt, E. 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