Environ. Sci. Technol. 1988, 22, 1038-1043
Avian Tissues as Biolndicators of Exposure to Alkylleads: Metabolism of Ethyllead Salts by Japanese Quail Kannan Krishnan and Wllllam D. Marshall” Department of Food Science and Agricultural Chemistry, Macdonald College, Ste. Anne de Bellevue, Quebec, Canada H9X I C 0
Japanese quail, in groups of seven birds, were provided drinking water amended with 0.0 or 250 ppm of Pb(N03)2, 25 ppm of Et2PbC12,or 2.5 or 0.25 ppm of Et3PbCl for 8 weeks. Daily egg pools from treated and control groups and soft tissues (liver, kidney, brain, and breast muscle) recovered at the termination of the trials were analyzed for alkyllead salts. For the inorganic lead feeding trial, no evidence for host-mediated methylation was observed in any of the samples. In each of the alkyllead feeding trials, the toxicant was rapidly transferred to the egg. However, the increase in toxicant concentrations with time was not monotonic. Et2Pb2+was metabolized to Et3Pb+,Me2Pb2*, and Me3Pb+in low yield which accumulated in the egg. The major toxicant in soft tissues was Et3Pb+. Metabolic dealkylation of Et3Pb+was a minor process, and Et2Pb2+ accumulated mainly in egg. Only traces of mixed alkyllead cations (Et2MePb+and EtMePb2+)were detected in liver or kidney if either Et3PbC1 or Et2PbC12served as test toxicant. These observations were used to reinterpret burdens of alkylleads observed in earlier avian wildlife vonitoring studies. Introduction
The speciation of alkyllead salts in biological tissues has recently become the focus of intensive study. The myriad forms of alkylleads include the volatile but rather labile tetraalkylleads (R,Pb) which are degraded in the environment either abiotically [photolytically (1,2),thermally, or hydrolytically ( 3 , 4 ) ]or in biological systems (5,6). The products, ionic salts (R3PbX, R2PbX2;R = Me, Et), are more persistent, yet they retain much of the acute mammalian toxicity (7) of their tetraalkyl progenitors. Moreover, they tend to accumulate at least temporarily in lipophilic tissues (7,8), where they cause damage which is distinctly different from classical plumbism. I t has been demonstrated that alkyllead salts and inorganic lead salts may be methylated abiotically (9-11) or “biologically” by lake sediments or bacterial cultures (12-14). However, the biomethylation of Pb2+remains controversial (15, 16). In addition, several chemicals have been reported to catalyze the redistribution of alkyl groups between alkyllead compounds (17,18) or the disproportionation of trialkylleads to result in the tetra- and dialkyllead homologues (9, 13). A recent paper (19) reported concentrations of alkyllead salts (R3Pb+,R,Pb2+; R = Me, Et) in tissues (liver, kidney, and brain) from mature and immature Herring gulls (Larus argentatus) culled from four colonies and in gull eggs culled from 10 colonies within the Great Lakes. An interesting spectrum of alkyllead analytes was present in virtually all tissues. The ethyllead and methyllead tissue burdens were not significantly correlated, indicating independent ethyl- and methyllead sources to the gull populations. The lack of statistically significant differences in burdens between immature and mature gulls suggested a continued exposure of gulls to these toxicants rather than a single catastrophic event. Methyllead salts were ubiquitous even in tissues from sites which were several hundred miles from any known source of these toxicants. 1038
Environ. Sci. Technol., Vol. 22, No. 9, 1988
Moreover, levels of these salts were correlated significantly with lead levels in lake sediment, suggesting methylation but not ethylation of inorganic lead. To our knowledge, only a single study on the toxicity of alkyllead salts to an avian species has been reported (20). In this study starlings, which had received 2.8 or 28 mg/kg of body weight daily for up to 11 days, displayed overt signs of severe toxicity. Residues of alkyllead salts (unspeciated) in kidney, liver, and brain amounted to several percent of the dose administered. The objectives of the current research were to study the metabolism, accumulation in tissues, and transfer into eggs when a suitable avian species received repeated low doses of triethyl-, and diethyl-, or inorganic lead salts. It was considered that the information gained could be used to reevaluate the earlier observations with gulls. Additionally, it was anticipated that the results would provide an estimate by which concentrations of ionic alkylleads in eggs could be related to levels of exposure of the adult to these toxicants. The Japanese quail is considered to be a better model for avian wildlife than the domestic hen (21). On the basis of experimental observations with this species and comparisons with mammalian studies, this species was also considered to be an appropriate model for the study of plumbism (22, 23). Additionally, it represents a species which lays eggs throughout the year. Materials and Methods
Reagents and Standards. Alkyllead chlorides (R3PbC1,R2PbC12;R = Me, Et) and alkyllead butylates (R3BuPb,R2Bu2Pb)were prepared as previously described (24,251. All chemicals were ACS reagent-grade or better, and chromatographic support gases were prepurified-grade. Enzyme preparations were obtained from the Sigma Chemical Co. (St. Louis, MO). The ammoniacal buffer consisted of diammonium citrate (22.6 g), potassium cyanide (4.0 g), and sodium sulfite (24.0 g) made up to 250 mL with distilled water. The pH of the diluted solution was adjusted to 10.0 with concentrated aqueous ammonia. Feeding Trials. Female adult Japanese quail (Coturnix coturnix japonica, a commercial strain purchased from a local breeder), 6-8 months old, were housed, in groups of seven birds, in chick batteries and were provided with feed (Purina turkey chow) and distilled water ad libitum during a 12-14-day acclimatization period. The room temperature was regulated to 25 “C (Et2Pb2+and Pb2+trials) or to 22 “C (Et3Pb+trials). Indirect lighting was set to provide 10 h of darkness and 14 h of light. Weekly weight gains and egg laying patterns were established during this time. Treatments consisted of amending the distilled drinking water with 0 or 250 mg/L Pb(N03)2,with 25 mg/L Et2PbC12,or with 2.50 or 0.25 mg/L Et,PbCl. Fresh amended water was supplied each day after recording the previous day’s consumption. Each trial was conducted for 8 weeks. Daily, eggs from each group were segregated by treatment, removed from their shells, pooled by treatment, homogenized in a Sorvall Omni Mixer (Du Pont Instruments), and frozen at -10 “C to await analysis. At the termination of each trial, birds were sacrificed by deca-
0013-936X/88/0922-1038$01.50/0
0 1988 American Chemical Society
pitation. Soft tissues (liver, kidney, brain, and breast muscle) were excised, rinsed in isotonic saline, dried on tissue paper, weighed and analyzed separately (liver, muscle) or pooled (brain, kidney), and analyzed. Enzymatic Hydrolysis. Samples of tissue or egg homogenate (2.5 g) were incubated in 50-mL Nalgene screw-cap centrifuge tubes at 37 "C for 24 h in 20 mL of 5% ethanol/0.5 M sodium dihydrogen phosphate buffer (pH 7.5) containing 40 mg each of lipase (type VII, Sigma Chemical Co., St. Louis, MO) and protease (type XIV). Extraction. Ammoniacal buffer (5 mL) was added to the hydrolysate. The diluted hydrolysate was then extracted 3 times with 0.01% (w/v) dithizone in hexane (10 mL). The pooled dithizone extracts were centrifuged at 2350g for 10 min (5 "C) to hasten phase separation. The organic phase was concentrated to 1.0 mL in precalibrated tubes (equipped with screw-cap tops and Teflon liners) under a gentle stream of nitrogen at 30 "C. Derivatization. n-Butylmagnesium chloride (0.5 mL, 2.27 M in tetrahydrofuran, Alfa Products, Ventron Corp., Danvers, MA) was added to the concentrated organolead dithizonates. The tubes were capped, vortexed 10 s, magnetically stirred for 10 min at ambient temperature, and cooled in an ice bath. Excess Grignard reagent was destroyed by the dropwise addition of 1M FINO3. The reaction mixture was diluted to 10 mL with water, shaken for 30 s, and centrifuged for 5 min at 310g (room T). The hexane layer was removed, and the aqueous layer was reextracted with 5 mL of fresh hexane. The organic extracts were combined, dried over sodium sulfate, reduced to 1 mL under a gentle stream of nitrogen, placed in a sample vial, and capped for immediate analysis. Sample Analysis. A gas chromatograph (GO-quartz tube-atomic absorption spectrometer (QT-AAS) as previously described (25) was used for the quantitation of samples. Three replicate determinations were performed on each sample homogenate. In addition, each butylated extract was quantified 2 or 3 times by comparison with external standards containing Me,BuPb, Me2Bu2Pb, Et3BuPb, and Et2Bu2Pb. Methylethyllead compounds were identified by prediction of retention times with Kovat's retention index and from retention times of alkylbutyllead standards (24). Actual retention times of methylethylleads were confirmed from transalkylation mixtures, and quantitation of these compounds was achieved by comparison with a similar analyte for which standards were available. Thus, quantitation of MeEt2Pb+ and MeEtPb2+was based on the instrumental response to and recoveries for Me3Pb+ and Et2Pb2+,respectively. Recovery Experiments. Three samples of each quail tissue (liver, kidney, brain, and breast muscle) were spiked at a level of 5-6 ng/g wet weight (as Pb) with a mixture of Me3PbC1, Me2PbC12, Et,PbCl, and Et2PbC12. The percentage recovery of each analyte was determined by dividing the mean peak area of the recovered butylate by the mean peak area of a butylated spike solution diluted to the expected (assuming 100% recovery) concentration. Recoveries, the average of three replicate determinations, are recorded in Table I. No alkylleads were detected in control tissues or eggs which had not been intentionally spiked. Statistical Analyses. The data were analyzed according to the method of covariance taking into account the dependent variable associated with each independent variable. If the F ratio indicated a significant timetreatment interaction, the data set was decomposed so that only treatment means for the same time interval were compared. Duncan's New Multiple Range Test was used
Table I. Mean Recoveries of Ionic Alkyllead Compounds from Separate Quail Tissues" meanb analyte percent recovery i 1 SD tissue
Me3Pb+
MezPb2+
Et3Pb+
EtzPb"
egg liver kidney brainC
92f3 83f7 84 f 6 72 f 2
75f6 18k6 22 f 7 17 f 4
78f8 71i7 74 f 5 61 f 2
89f7 66i9 74f 6 50 f 2
Spiked at 5-6 ng/g wet weight (as Pb) into control tissue which did not contain detectable quantities of analytes. * N = three replicate determinations. Calculated from three replicate injections of a single determination. 100
,
I
40
T
1 0
2
6
4
8
WEEKS
Figure 1. Mean water consumed (mL/bird, daily average for each week) by groups of Japanese quail which were provided drinking water 25.0 mg/L EtzPbCIz(O),or 250.0 mg/L Pb(NO& amended with 0.0 (O), (+). Means for the same time interval bearing different letters are significantly (p < 0.05) different.
to identify significant ( p treatment means.
0.05) from the water consumption by birds treated with 250 mg/L of Pb(N0J2 (Figure 1). Water consumption for the various trials was compared on a weekly basis (representing the average of the seven previous daily consumptions). On the basis of the water consumed, the mean level of Et2PbClzingested/bird per day was calculated to be 5.66 f 0.90 mg/kg of body weight. Although water consumption by the treated groups was reduced significantly, mean body weights (recorded weekly) and daily egg laying patterns were unaffected by these treatments. Exposure of the birds to the amended water resulted in a prompt transfer of the toxicant into egg: 9.8 f 0.8 mg/kg wet weight of Et2Pb2+was detected in the pooled egg sample 1 day after the commencement of the trial. The level of this toxicant in egg increased, but not monotonically, during the first 33 days of the trial (Figure 2), reaching a maximum of 1.04 f 0.11 mg/kg wet weight. On day 27 and in subsequent days of the trial, Et,Pb+ was also detected in the egg pools (Figure 3). Although the level of this toxicant increased with time, it remained at less than 3.2% of the Et2Pb2+concentration in the same egg pool. On day 50, traces of Me2Pb2+were detected, and on subsequent days of the trial Me,Pb+ was also detected. The Environ. Sci. Technol., Vol. 22,
No. 9, 1988 1039
i
T
3OY
Figure 2. Concentrations of Et2Pb2+in daily egg pools from quail which had received drinking water amended with 25.0 mg/L Et2PbC12. Error bars represent 1 SD based on three replicate determinations.
Level
Day Figure 3. Concentrations of Et,Pb+, Me,Pb2+, and Me,Pb+ in daily egg pools from quail which had received drinking water amended with 25.0 mg/L Et2PbC12. Error bars represent 1 SD based on three replicate determinations.
levels of Me2Pb2+and Me3Pb+ increased from day 50 to day 56; although Me3Pb+remained a trace contaminant (6.5 f 1.6 ng/g wet weight on day 56), levels of Me2Pb2+ became appreciable (82.1 f 6.6 ng/g on day 56). During the EtzPb2+trial, no steady-state concentration of ionic alkylleads in eggs was achieved (Figure 2); the levels of the toxicants continued to increase in the egg pool throughout the trial. No explanation for the abrupt decrease in EhPb2+concentration in the day 33 egg pool was evident although this event corresponded roughly to the appearance of increased concentrations of Et,Pb+ in the egg samples. There were no significant differences between the control and treated group in mean body weight, mean egg volume, mean egg mass, or number of eggs laid during this period. Although no fatalities occurred which could be attributed to the toxicant (one control bird died of unknown causes), the trial was terminated after 8 weeks, in part because of a decreased egg production by both treated and control birds (a result of old age). At sacrifice there were no significant differences in mean body weights between the treated and control groups. In terms of organ weight to body weight ratios (Table 11) only the kidney was significantly elevated 0, < 0.01); liver and 1040
Environ. Sci. Technol., Vol. 22, No. 9, 1988
Table 11. Means of Organ Weight to Body Weight Ratios (X103) for Treated and Control Birds
treatment control Pb(NOJ2, 250 pg/g EtzPbClz, 25 pg/g control Et3PbC1, 2.5 hg/g Et3PbC1, 0.25 pg/g
kidney liver weight/body weightlbody weight weight 5.0 f 0.9 7.9 f 1.0" 8.3 f 3.8" 7.3 f 0.9 7.0 f 0.7
8.3 f 1.2
36.1 f 8.5 37.6 f 9.5 32.9 f 4.3 32.1 f 4.8 29.3 f 4.8 30.7 f 3.3
brain weight/ body weight 3.1 f 0.4 2.6 f 0.3 2.8 & 0.1 4.0 f 0.3 3.7 f 0.4 3.9 i 0.2
" Sienificantlv different from mean control value (D < 0.01). brain weightlbody weight ratios were unchanged. Virtually all tissues contained appreciably more Et3Pb+ than Et2Pb2+(Table 111). Clearly a toxification mechanism is operative in quail which converts ingested Et2PbC12to Et3Pb+. Whereas metabolic dealkylation has been observed in several mammalian species (8), a metabolic conversion of dialkyllead to trialkyllead has not been reported previously to our knowledge. Surprisingly, Et3Pb+
Table 111. Mean Organolead Cation Concentrations (ng/g Wet Weight) in Soft Tissues from the Diethyllead Dichloride Feeding Trial to Japanese Quail analyte mean' f 1 SD Et,Pb' Et2Pb2+
analyte range Et,Pb+ Et,Pb2+
brain poolb 16.2 f 0.05 5.1 f 0.01 kidney poolc 161.6 f 32.3 52.6 f 14.6 liverd 49.6 f 44.9 22.0 f 7.8 14.9-150.1 muscled 15.3 f 19.8 18.2 f 4.8 2.3-55.7
3o
analvte meana f 1 SD EtzMePb+ EtMePbZt
I
1
11.9-31.9 9.2-27.1
livere kidneye
0.6 i 0.3 1.5 f 0.3
1.4 f 0.4 1.9 f 0.6
Corrected for recoveries. *Three replicate injections of one pooled sample. Two replicate injections of three separate sample pools. d T ~ replicate o injections of three separate samples from each bird. eThree replicate injections from a single pooled sample.
intake was calculated to be 61.2 f 11.5 mg/kg body weight. As with the Et2Pb2+trial, there were no significant differences (treated vs control group) in mean body weights, number of eggs laid, or mean egg mass. At sacrifice, after 8 weeks of daily dosing, the kidney weight to body weight ratio was significantly elevated (p < 0.01, Table 11). However, no other significant differences were observed. No alkyllead salts were detected in any of the daily egg pools or in any of the soft tissues at sacrifice. Thus, in this species, metabolic alkylation of Pb2+was not observed at any time during the 2-month trial. Two feeding trials were also conducted in which EbPbC1 was added at 2.5 or 0.25 ppm to distilled drinking water. Water consumption, relative to controls, was significantly depressed for the high treatment group only (Figure 4). Mean Et3PbC1ingestedlbird per day was 61 f 5 and 425 f 36 pg/kg of body weight for the low and the high treatments, respectively. As with the Et2PbClz feeding trial, the toxicant was rapidly transferred to egg (Figures 5 and 6); pooled egg samples contained 0.1 f 0.07 and 1.1 f 0.1 ng/g wet weight of Et3Pb+ (low and high treatment, respectively) 1 day after the commencementof these trials. Et2Pb2+was ubiquitous in the egg pool samples, indicating that metabolic dealkylation, which had been reported
"i
I4
Level
Et3Pb+ (
3 Et2Pb2+ 6
4 2
o,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,
123456789111111111122222222223333333333444444444445555555 01234567890123456789012345678~012345678~0123456
Figure 5. Concentrations of Et3Pb+ and Et2Pb2+cations in daily egg pools from Japanese quail which had received drinking water amended with 0.25 mg/L Et3PbCI. Error bars represent 1 SD based on three replicate determinations. Environ. Sci. Technol., Vol. 22, No. 9, 1988
1041
T
60 T
I
i
T
50
L w e i (ppb)
30+
BOY
Figure 6. Concentrations of Et3Pb+and Et2Pb2+cations in the daily egg pools from quail which had received drinking water amended with 2.5 mg/L Et,PbCI. Error bars represent 1 SD based on three replicate determinations. Table IV. Mean Organolead Cation Concentrations in Soft Tissues from the Triethyllead Chloride Feeding Trials to Japanese Quail
Table V. Statistical Correlations" between Mean Ionic Alkyllead Concentrations in Liver and Muscle from Quail Which Had Received Water Amended with 2.5 or 0.25 mg/L Et,PbCl
analyte range analyte mean f 1 SD tissue
EtsPb', pg/g
,
EtzPb2+,ng/g
pg/g
ng/g
brain" 250 rgikg 2.5 mg/ kg
kidneyb 250 r d k g 2.5 mg/kg liver 250 &kgc 2.5 mg/kgd musc1ee 250 a / k g 2.5 mg/kg
0.06 i 0.007 0.45 f 0.009
2.4 f 0.4 23.1 f 1.1
0.30 f 0.1 1.83 0.6
*
31.3 f 14.3 239.8 129.5
*
0.42-0.53 1.06-2.96
11.0-48.0 107.2-418.2
0.29 i 0.1 1.11 f 0.03
21.3 f 17.9 70.5 f 34.9
0.20-0.34 0.66-2.33
11.0-48.0 48.8-153.8
0.08 f 0.03 0.34 0.1
6.1 i 2.9 21.0 f 7.4
0.03-0.13 0.27-0.61
2.3-9.7 15.0-38.6
*
a Three replicate injections of a single pooled sample. Two replicate injections of three separate pooled samples. Individual livers analyzed separately. dSingle or duplicate sample from each bird. eThree samples from each bird.
previously in mammals (8, 26), was active in this avian species as well. Both trials were characterized by a relatively constant concentration of Et,Pb+ in egg particularly during the latter stages of these studies. The mean concentration of Et3Pb* in egg was 5.2 f 2.4 and 31.6 f 16.2 ng/g (low and high dose trials, respectively), and the mean level of EbPb2+was somewhat more variable, 4.6 f 2.3 and 20.5 f 17.5 ng/g wet weight. These differences in burdens reflected the differences in the mean levels of toxicant ingested between the two trials. At sacrifice, no significant differences in organ weight to body weight ratios were detected in either of these trials (Table 11). Although ethylleads were detected in all tissues (Table IV), methylleads were not detected in any tissue or in egg. The ratio of EhPb2+metabolite to Et,Pb+ cation concentrations, in separate tissues, was constant in both trials (0.074 0.03%) and was considerably higher in soft tissues than in egg. Triethyllead cation accounted for 89-97% of the total ionic alkyllead burden in soft tissues. In addition, the concentrations of EgPb+ in liver or muscle were correlated significantly wtih Et2Pb2+metabolite concentration in either of these tissues (Table V). Similarly, the concentrations of these analytes in kidney were correlated significantly (Table VI). These observations are consistent with an equilibration of toxicant burdens among the soft tissues and an active transport of the more
*
1042
Environ. Sci. Technol., Vol. 22, No. 9, 1988
tissue and analyte
muscle liver Et2Pb2+ Et,Pb+
muscleb Et3Pb' 0.9350 prob > lrl 0.0003** muscleb EtzPb" prob > Irl liverb Et.Pb+ prob > Irl liverb EtsPb2+ prob > Irl liverb EtzMePb+ prob > Irl musclec Et3Pb' prob > Irl musclee Et2Pb2+ prob > Irl liverc EtsPb' prob > Irl
liver liver liver Et2Pb2+ EbMePb+ EtMePbZ+
0.9796 0.0001**
0.9777 0.0001**
0.5326 0.1741
0.8328 0.0103*
0.9054 0.0002**
0.9885 0.0001**
0.5165 0.1900
0.8073 0.0154*
0.9481 0.003**
0.6049 0.1121
0.8443 0.0084'
0.5180 0.1885
0.8191 0.0129* 0.8645 0.0056**
0.8703 0.0023**
0.8669 0.0025**
0.7945 0.0105'
0.8596 0.0030**
0.8720 0.0022** 0.9396 0.0002**
'(**) significant at < 0.01 level; (*) significant at 2.5 mg/L trial. OFrom the 0.25 mg/L trial.
< 0.05 level. bFrom the
Table VI. Statistical Correlationsa between Ionic Alkyllead Concentrations in Kidney from Quail Which Had Received Water Amended with 2.5 mg/L Et3PbC1 analyte Et,Pb+ prob > Irl EtzPb2+ prob > Irl Et2MePb+ prob > Irl "(**) significant a t
EtzPb2+ 0.8585 0.0064**
EtzMePb+ 0.9086 0.0018** 0.8924 0.0029**
EtMePb2+ 0.9123 0.0016** 0.7640 0.0273* 0.8902
0.0030**
< 0.01 level;
(*) significant a t
< 0.05 level.
polar metabolite into egg. Et,Pb+ toxicant and Et2Pb2+ metabolite were accumulated in soft tissues to approximately the same proportion in both trials. The concentration of either analyte, in any tissue, from the high-level trial may be compared with the concentration in the
Table VII. Mean Mixed Alkyllead Cation Concentrations in Soft Tissues" from Quail Which Had Received 2.5 mg/L Et3PbC1 in Drinking Water for 8 Weeks
tissue brain' liverd kidnef
analyte meanb f 1 SD EtzMePb+, EtMePb*+, ng/g ng/g
* *
3.2 0.04 4.5 f 2.7 5.8 1.8
6.3 1.2
* 5.6 * 5.5
analyte range EtzMePbt, ng/g
EtMePb2+, ng/g
ndf-8.2 3.8-8.5
nd-18.5 2.9-19.2
None detected in muscle. Corrected for recoveries. Three replicate injections of one pooled sample. d T ~ replicate o injections from three separate samples from each individual. 'Two replicate injections from one sample from each individual. 'nd = not detected.
correspondingtissue from the low-level trial (average ratio: 5.6 & 0.3, EhPb+; 6.3 f 0.5, EhPb2+). These ratios are very similar to the corresponding ratio for the average daily intake (425/61 = 7.0) between the two trials, indicating that the metabolic response was quantitatively similar in the two trials. Mixed ethylmethylleads (EbMePb+ and EtMePb2+but not EtMe2Pb+),although present only at trace levels, were ubiquitous in liver, kidney, and pooled brain samples from the 2.5 ppm of Et3PbC1feeding trial (Table VII). The concentrations of these analytes in kidney were correlated significantly with the concentrations of individual ethyllead cations in kidney (Table VI) and, similarly, in liver concentrations of EtMePb2+ (but not Et2MePb+)were correlated with concentrations of ethyllead salts (Table V). These results are consistent with a metabolic methylation of ethyllead salts in this species. However, relative to the concentration and toxicity of triethyllead cation, levels of mixed alkylleads are probably not toxicologically important. It is interesting to note that the mean burdens of these mixed alkyllead cations were approximately 4-fold higher (3.8-5.3) in the Et3PbC1feeding trial (425 pg/kg of body weight per day) than in the Et2PbC12trial (5.66 mg/kg of body weight per day) despite the 7-fold lower daily dosage. To the extent that quail represents a suitable model for other avian wildlife species, it is considered that the mixed alkyllead cations which were detected in Herring gulls and Mallard ducks probably resulted from ingestion and not from host-mediated metabolism of ethylleads. The current, feeding trials also indicate that metabolic methylation of' ingested Pb2+ is not a detectable process in quail and suggest that methyllead cation burdens in avian wildlife result from ingestion of the preformed toxicants. Finally, the ratios of mean Et2Pb2+to mean Et3Pb+concentrations in kidney and liver of Herring gulls from the Great Lakes (kidney, 0.35; liver, 0.42) more closely resembled the ratios in Table I11 (kidney, 0.35; liver, 0.44) than the corresponding ratios in Table IV (kidney, 0.11; liver, 0.07). This would suggest that the major toxicant ingested by gulls w a EbPb2+and that Et3Pb+burdens resulted, at least in part, from metabolic conversion of diethyllead. In toto, the results indicate that ethyllead salts are rather poorly accumulated in eggs or soft tissues. Further, they
suggest that egg is a suitable indicator tissue for exposure of avian species to Et2Pb2+and that soft tissues (liver, kidney, and brain) are the preferred indicator tissues for Et3Pb+ exposure. Registry No. Et2PbC12,13231-90-8;Et3PbC1, 1067-14-7;Pb, 7439-92-1; Me3Pb+, 14570-16-2; Me2Pb2+,21774-13-0; Et3Pb+, 14570-15-1; Et2Pb2+,24952-65-6; Et2MePb+, 105956-70-5; EtMePb2+, 106673-67-0.
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I
Received for review June 8,1987. Revised manuscript received December 28, 1987. Accepted February 29, 1988. This study was financially supported by the Natural Science and Engineering Research Council of Canada (GO316 and A6687).
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