Environ. Sci. Technol. 1999, 33, 2395-2402
Transfer of Cadmium and Mercury to Sheep Tissues N I C H O L A S A . B E R E S F O R D , * ,† ROBERT W. MAYES,‡ NEIL M. J. CROUT,§ PATRICIA J. MACEACHERN,‡ BEVERLEY A. DODD,† CATHERINE L. BARNETT,† AND C. STUART LAMB‡ Institute of Terrestrial Ecology, Merlewood Research Station, Grange-over-Sands, Cumbria, LA11 6JU, U.K., Macaulay Land Use Research Institute, Craigiebuckler, Aberdeen, Scotland, AB15 8QH, U.K., and School of Biology, University of Nottingham, Sutton Bonington, Loughborough, Leicestershire, LE12 5RD, U.K.
Toxic heavy metals such as cadmium and mercury can enter the diet of farm animals by a variety of environmental exposure routes and, hence, contaminate food products derived from those animals. Therefore, there is a need to be able to predict the likely levels of contamination in animal tissues if exposed to a contaminated diet and also to estimate how rapidly an animal will decontaminate once the source of contamination is removed from the diet. Data on the transfer and excretion rates of Cd and Hg from tissues have previously been inadequate to allow the development of dynamic models to predict changes in the degree of contamination of different tissues of ruminants. A study is described during which a group of sheep were given a single oral administration of 109Cd and 203Hg. Measurements of the concentrations of the radioisotopes in tissue samples were subsequently made over a period of 1 year. The resultant data were used to develop compartment models to describe the behavior of the two metals in sheep tissues. To our knowledge the models developed are the first to allow the time-dependent prediction of the potential Cd and Hg contamination of animal-derived food products. Previously only advised transfer coefficients were available; we demonstrate that these are of little value for cadmium and mercury due to their slow rates of accumulation and excretion.
Introduction There are a number of processes such as smelting of nonferrous ores, waste incineration (1), and the disposal of sewage sludge onto land (2) that release toxic metals into the environment resulting in locally enhanced levels. Contamination of surface soils by heavy metals can be especially high in areas of historical or current mining (e.g., ref 3). These activities have resulted in the requirement for assessments of the likely concentrations in foodstuffs produced in the areas around such industrial or mining sites (e.g., refs 4-6). A more recent, potential source of heavy metals releases into * Corresponding author tel: 441539532264; fax: 441539535941; e-mail:
[email protected]. † Institute of Terrestrial Ecology. ‡ Macaulay Land Use Research Institute. § University of Nottingham. 10.1021/es9811041 CCC: $18.00 Published on Web 05/29/1999
1999 American Chemical Society
the environment is large-scale plastics fires (7). Many plastics contain heavy metals, most notably cadmium, that can be incorporated into plastics at the rate of 41 kg of Cd (t of plastic)-1 (8). There is also the potential for incidents such as that which occurred in Europe during 1989-1990 when imported animal feed contaminated with heavy metals was accidentally allowed to enter the diet of farm animals (9, 10). Although Pb was the prime contaminant of concern in this instance, levels of Cd and Hg within the feed were also above national guideline levels. There is a clear requirement to be able to predict both the levels of heavy metal contaminants in the tissues of animals exposed to contaminated diets and also the rates of decontamination of those tissues once the source of dietary contamination has been removed. Although there is wealth of information on the mechanisms of toxic effects of heavy metals in mammals (11, 12) and some data on the transfer of heavy metals from diet to animal tissues measured during feeding trials (13-17), there is inadequate data currently available to develop and/or validate food chain models describing the time-dependent transfer of these contaminants to the milk and meat of farm animals. Within the United Kingdom, sheep are generally the farm livestock most likely to graze contaminated former mine workings. The purpose of the work described in this paper was to develop dynamic models describing the transfer of Cd and Hg to the tissues of sheep.
Experimental Section Protocol. Thirty-three Scottish Blackface lambs aged 6-7 months were individually housed and fed a diet of dried grass pellets (900 g of dry matter (DM)/day) and alkali-treated straw pellets (196 g of DM/day) for a period of approximately 4 weeks prior to the start of the study. The animals were all obtained from two flocks maintained by the Macaulay Land Use Research Institute and were of a similar live-weight and body condition. The lambs were placed in metabolism cages and allocated to 11 groups of three on the basis of anticipated sacrifice dates of between 0.5 and 365 days after radioisotope administration. Care was taken to ensure that animals originally obtained from each of the two flocks were distributed evenly throughout the intended sacrifice dates. Twelve of the animals (those selected for sacrifice 45-365 days after radioisotope administration) were housed in cages fitted with collection chutes and separators to allow total collections of feces and urine. On a single occasion (day 0) solutions containing 109CdCl2 (2.10 ( 0.023 MBq; mean ( SE) and 203HgCl2 (0.75 ( 0.020 MBq) were administered to each lamb directly into the rumen via a tube passed down the esophagus. Because of its short radioactive half-life (46.8 days), the administered activity of 203Hg was doubled for each subsequent anticipated sacrifice date after day 24. The specific activities of the two radioisotope solutions as supplied were 0.038 mg of Hg/MBq of 203Hg and 0.001 mg of Cd/MBq of 109Cd. Therefore, each animal was administered 0.021 ( 0.0002 mg of stable cadmium with the isotopic solution. Animals to be slaughtered before day 45 were administered 0.028 ( 0.0013 mg of stable mercury; thereafter, this was approximately double for each subsequent slaughter date in line with the increased administration of 203Hg. Total feces and urine outputs were collected and weighed from the animals in metabolism cages equipped with separation chutes for 24 days after radioisotope administraVOL. 33, NO. 14, 1999 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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tion. Subsamples of excreta were weighed into 700-mL or 2.3-L plastic containers and stored at -20 °C to await analysis. Immediately prior to sacrifice by captive bolt, the liveweight of each animal was recorded. After collection of a sample of blood (>2 L, mixed with heparin), the liver, kidneys, lungs (day 0.5-10 only), forestomach contents (rumen; day 0.5-24 only), and combined intestinal and cecal contents (hind gut; day 0.5-24 only) were removed together with samples of muscle (hind leg), bone (femur), mesenteric fat (day 0.5-10 only), and skin (day 0.5-10 only). Only two animals were sacrificed 365 days after radioisotope administration; the third animal died of an unknown illness. To determine the proportion of live-weight attributable to muscle, bone, and fat, total dissections on a half of the carcass (divided down the spine) were performed from one sheep sacrificed on each of 1.5, 6, 16, 90, and 181 days after radioisotope administration. On three occasions during the study, the total shaved skin was also removed and weighed. Blood samples were centrifuged (2200g for 20 min) to separate plasma and red blood cells (RBC); the RBC were washed by resuspension in 0.15 M NaCl solution followed by centrifugation (2200g for 20 min). Soft tissues samples were sliced into approximately 1 cm3 pieces and well mixed before being weighed into plastic containers (130 mL-2.3 L). All samples were stored at -20 °C. Approximately 400 g of fresh bone, free from soft tissue, was dissolved in 400 mL of concentrated HNO3 and diluted to 700 mL with deionized water. The amounts of feed offered were increased to 1213 g of DM/day after day 90 and 1320 g of DM/day after day 181, maintaining the same grass pellet:straw pellet ratio. Samples of feed were retained throughout the study. Sample Analyses. The activity concentrations of radioisotopes in tissue and excreta samples were determined using hyper-pure germanium detectors. The detectors were calibrated using mixed γ standards of appropriate matrix and volume. Analyses times ranged from 1000 to 240 000 s depending upon the individual sample activity, where possible a counting error of less than 10% of the result for each isotope was achieved. The date of isotope administration was used as the reference date to correct for radioactive decay. Plasma and RBC samples were made up to a standard volume with deionized water prior to analysis. After freeze-drying, subsamples of the dietary components were digested in concentrated nitric acid (BDH Aristar) under reflux. The Cd concentrations in diluted aliquots of the digests were determined by inductively coupled plasma mass spectrometry. Mercury concentrations were estimated by atomic fluorescence spectrometry. Limits of detection were taken to be three times the standard deviation of determinations conducted on 18 blank samples prepared at the same time as the sample digests (18). To ensure adequate quality control, a certified ryegrass reference sample was analyzed (Community Bureau of Reference CRM281). Modeling Methods. For both cadmium and mercury, a similar model structure was used, and this is shown in Figure 1. This structure was chosen on the basis that this represents a simplified understanding of the transfer of pollutants from the diet to the circulatory system and hence to excreta and tissues. The modeling methods used were similar to those employed to develop models to predict the dynamic behavior of a range of pollutant radionuclides in the tissues of farm livestock, including radioisotopes of cesium (19), cerium, ruthenium, and silver (20). For input into the model parametrization, the experimental data was expressed as the proportion of administered activity determined in a given pool (unitless) (see Tables 1 and 2). Excretion of Cd and Hg from the circulatory system was modeled as one pathway (see Figure 1) rather than as separate routes to represent endogenous fecal excretion and urine. 2396
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This decision was made on the basis of the experimental results, which are discussed later. The kidney compartment represents Cd or Hg within the kidney tissue rather than the metals being transferred from the extracellular fluid compartment (ECF) to urine via the kidney. The model comprised a set of simultaneous first-order differential equations, and the symbols for the models’ rate coefficients (day-1) are defined in Figure 1:
dR ) -argR dt dG ) -argR + aegECF - (agex + age)G dt dECF ) -ageG + aleL + ameM + akeK + alueLu + dt aboeBo + aseS + arbceRBC + afateFat - (aeg + ael + aek + aem + aelu + bebo + aerbc + aes + aefat + aeex)ECF dL ) aelECF - aleL dt dK ) aekECF - akeK dt dM ) aemECF - ameM dt dLu ) aeluECF - alueLu dt dBo ) aeboECF - aboeBo dt dRBC ) aerbcECF - arbceRBC dt dS ) aesECF - aseS dt dFat ) aefatECF - afateFat dt dEx ) agexG + aeexECF dt The RBC compartment was not included in the Cd model since 109Cd was not detectable in this compartment (see Results). The models were implemented using the modeling software ModelMaker 3.0 (21). Within the software, the differential equations were solved using the fourth-order Runge-Kutta algorithm, and model fitting was undertaken by the Marquardt method (22). To account for the different magnitude of the data for the various compartments, the fitting procedure minimized the weighted sum of square residuals (χ2). For each data point, the weighting factor was the reciprocal of the variance of the mean estimated from sample replicates. Confidence intervals ∆yn,t for the nth compartment at time t were calculated by estimating the vector of the derivatives [dyn,t /daj] where aj is the jth parameter. These were then combined by summing over i ) 1,n and j ) 1,n where n is the number of estimated parameters in the model:
(∆yn,t)2 )
dy dy cov(i,j) ∑da da i
j
TABLE 1. Percentage of Administered muscle liver kidney ECF RBC bone lung fat skin rumen intestine sum in tissuesc
mean SE mean SE mean SE mean SE mean SE mean SE mean SE mean SE mean SE mean SE mean SE mean SE
Cd in Sampled Tissues throughout the Studya
109
day 0.5
day 1.5
day 3
day 6
day 10
day 16
day 24b
day 45
day 90
day 181
day 365b
1.43 × 10-3 5.25 × 10-4 2.31 × 10-2 7.87 × 10-3 2.31 × 10-3 7.42 × 10-4
