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A total of 128 primary mammary tumors (66 of them malignant) occurred in 35 female beagles injected with {sup 226}Ra at eight dose levels ranging from 0.2 to 440 kBq/kg body mass as young adults, while a total of 156 mammary tumors (57 of them malignant) were seen in 46 female control beagles not given any radioactivity. Sixty-three of 65 control dogs and 59 of 61 dogs given {sup 226}Ra survived the minimum age for diagnosis of mammary tumors of 3.75 years. Based on the observed age-dependent tumor incidence rates in the controls and on the corresponding number of dog-years at risk, the total number of observed malignant tumors in the radium group was statistically greater than the number of expected malignant tumors (66 observed vs 34 expected, P < 0.005). There was no such difference for the benign tumors. Cox regression analysis indicated no increased risk for the first tumor occurrence in irradiated dogs. Cox regression analysis of the multivariate risk sets showed no significantly increased risk for the occurrence of benign tumors but a statistically higher risk of 1.66 with a confidence interval of 1.15-2.40 for the occurrence of malignant tumors. The increased risk was dependent on dose,more » but a dependence on the frequency of previous occurrence of mammary tumors could not be confirmed. Censoring ovariectomized dogs at time of surgery decreased the relative risks slightly but did not alter the significance. Exposure to diagnostic X rays with cumulative exposures below 0.2 Gy had no effect on tumor formation. It is unknown whether the increased risk for malignant mammary tumors was due to some initial deposition of radium in sensitive tissue, a possible irradiation of fatty mammary tissue from transient radon {yields} polonium deposition, or a general effect of the overall radium deposition on the immune system of the dogs that lowered their resistance to formation of mammary tumors. 27 refs., 5 figs., 4 tabs.« less

The natural survival, relative to properly chosen controls, of 26 beagle dogs injected once intravenously with an average of 0.58 ? 0.04 kBq 239pu/kg, 23 dogs injected with 2.31 ? 0.43 kBq 226Ra/kg, 13 dogs injected with 1.84 ? 0.26 kBq 228Ra/kg, 12 dogs injected with 0.56 ? 0.030 kBq 228Th/kg, and 12 dogs injected with 21.13 ? 1.74 kBq 9oSr/kg was evaluated statistically. The amounts of these radionuclides are related directly to the estimated maximum permissible body burdens for humans suggested in ICRP II (1959). They constitute a level of exposure that initially was assumed to cause no deleterious effects in dogs. This study had two objectives: (1) identification of homogeneous control groups against which to evaluate the survival of the irradiated groups and (2) comparison of the survival characteristics and estimation of mortality or hazard rate ratios for control dogs vs dogs injected with the baseline dosages given above. It was shown, by goodness-of-fit plots, that the Cox proportional hazards model was an appropriate method of analysis. Therefore, covariates that possibly could influence survival were tested for significance. Only the effects of grand mal seizure, which is caused in epileptic dogs by an external stimulus and can be fatal if untreated, were significant (P < 0.0001). Consequently, in the final model, death from grand mal seizure was considered as accidental. After censoring the dogs dying from grand mal seizure, it was established that the data for the control groups from previous and contemporary experiments could be pooled. The change in hazard rates relative to controls resulting from exposure to the baseline radionuclide level was modest, 1.6 times for 239Pu (P = 0.033), 1.0(4) for 226Ra (P = 0.86), 1.9 for 228Ra (P = 0.035), 2.5 for 228Th (P < 0.001), and 0.52 for 9oSr (P = 0.041). Bone tumor induction was clearly elevated in dogs injected with 239Pu and 228Th. When the effect of these bone tumors on survival was removed by censoring, the dogs injected with 239Pu were indistinguishable from the controls. In contrast, the effects of bone tumor on group survival of the 228Ra and 228Th dogs were not significant. Thus, no additional life-shortening effects beyond those attributable to bone tumor were suggested by these data for 239pu, but other, as yet unspecified, confounders are suggested for 228Ra and 228Th. ? 1991 Academic Press, Inc.

The influence of age at injection of226 Ra or239 Pu on skeletal deposition and local distribution, the pattern of bone tumor formation, and postinjection survival was assessed in parallel short-term studies of mechanisms and lifetime toxicity. Beagles received a single intravenous injection of226 Ra or239 Pu at 3 months (juveniles), 17-19 months (young adults) or 60 months (mature). Data from short-term studies of mechanisms and dosimetry and from one dosage level (41 kBq ${}^{226}{\rm Ra}/{\rm kg}$ or 11 kBq ${}^{239}{\rm Pu}/{\rm kg}$ body mass) of each of the toxicity experiments were compared. Skeletal growth and turnover produced differential initial deposition and distribution patterns typical for each age group. At 1 week after injection, skeletal retention of226 Ra or239 Pu was 68 and 68%, respectively, in the juveniles, 32 and 46% in the young adults, and 31 and 43% in the mature dogs. Comparing individual bones in the juveniles, gradients in the concentration of239 Pu were small since all bones ...

An unusually high incidence of kidney disease (tubular degeneration and necrosis with fibrous replacement) was observed among 24 beagles injected at about 5 years of age with 116 or 329 kBq 226Ra kg-1 but not among an additional 10 beagles given about 39 kBq 226Ra kg-1. This 226Ra solution also contained 210Pb, 210Bi, and 210Po. To determine whether the kidney disease was related to the radiation from 226Ra and its short-lived progeny or to the alpha radiation from 210Po, 2 beagles about 7 years of age were injected with 451 kBq 226Ra kg-1 of 210Po citrate. Measurements of polonium retention in the kidneys of 4 additional beagles given 210Bi citrate enabled us to model the retention of these emitters in the dog kidney and to estimate the kidney dose from the alpha radiation of 210Po following injection of either 226Ra + 210Bi + 210Po or 210Po only. Autoradiography revealed that almost equal concentrations of 210Po were in the tubular epithelium and/or its basement membrane and in the glomeruli, but very little of the 210Bi deposited in kidney tissue was present in the glomeruli. Radiation damage to the kidneys similar to that observed previously in beagles given 226Ra solutions that also contained 210Bi and 210Po was seen among the beagles given 210Po but not in the dogs given purified 226Ra. The analysis of these data indicated that the relatively high incidence of kidney disease among the mature beagles injected with 226Ra and its accompanying 210Bi and 210Po resulted from alpha irradiation of the kidneys by the substantial amount of 210Po that was in the injection solution.

Fourteen young adult beagles were given an intravenous injection of either ${}^{241}{\rm Am}({\rm III})$ citrate or ${}^{239}{\rm Pu}({\rm IV})$ citrate (labeled with239 Pu) followed by a single intravenous injection of Ca-DTPA after an interval of 1 min, 6 min, 30 min, 150 min, 8 hr, 1 day, or 3 days. Animals were sacrificed 7 days after DTPA injection. Retention of241 Am at death was influenced strongly by early treatment. Dogs given Ca-DTPA at 1, 6, and 30 min retained about 3, 10, and 29%, respectively, of the injected241 Am, while the animal treated at 150 min retained 45%. Beagles given241 Am and then DTPA at 8 hr, 1 day, and 3 days, retained 58, 73, and 72%, respectively. For241 Am contamination, the first DTPA treatment should be given as soon as possible. Total-body retention of ${}^{239+237}{\rm Pu}$ was affected less dramatically by early treatment. The dogs given DTPA at 1, 6, and 30 min retained 31, 28, and 34% at death, while the...

The anatomical distribution of /sup 239/Pu in the proximal humerus was studied in four beagles injected at 90 days of age and sacrificed 7, 28, 56, and 128 days after injection, respectively. Initial deposition in each humerus was 3.1% of the injected Pu; 41% of that was retained with a net halftime of 248 days; for the remaining 59% no decrease was observed with time. Sixty-one percent of the initial deposit was concentrated in the proximal epiphysis and metaphysis. Average radiation dose rates to trabecular bone of the epiphysis, to those parts of the metaphysis that were formed before and/or after injection, and to the area corresponding to the primary and secondary spongiosa were determined by fission track autoradiography. Local dose rates within the range of 10 ..mu..m of the endosteal surface, to the marrow and to osseous tissue were calculated. The possible effects of local radiation doses were correlated with areal densities of cells and types of cells at risk. Comparisons were made with corresponding sites from the humerus of beagles injected as young adults.

Twelve juvenile beagles 90 days old were injected intravenously with 2.8 ..mu..Ci /sup 239/Pu/kg. All attained normal skeletal size and normal weight. However, all died from osteosarcoma with an average survival time of 1306 +- 163 days after injection. At death the liver contained 12.7 +- 2% of the injected /sup 239/Pu, and less than 1% was in the total of kidneys, lung, and spleen. In a liver homogenate, free of nuclei and debris, most of the Pu was bound to ferritin in the cytosol or found with microsomes. No overt evidence of radiation-induced liver damage was discernible. At death, the juvenile skeletons contained 56.0 +- 3.1% compared to 44% in beagles injected as young adults. Highly active bone growth and turnover produced a skeletal redistribution pattern of Pu which differed from that observed in young adults. The cumulative average dose to the anatomical skeleton was 2850 +- 443 rad. Seventeen bone tumors were produced, located predominantly in areas with high concentrations of Pu and rapid bone turnover rates. Skeletal radiographs taken at death revealed 33 fractures, all of which had healed.

Retention of administered226 Ra was substantially greater in beagles injected as 3-month-old juveniles than as 1.4-year-old young adults, but the measured ${}^{222}{\rm Rn}/{}^{226}{\rm Ra}$ ratio ...

When /sup 239/Pu(IV) is injected intravenously, its fractional retention in the skeleton of the beagle decreases and then approaches an approximately constant value. The initial fractional deposition of /sup 239/Pu is independent of dose level, but the long term fractional retention is less at the lower dose levels (0.0055 to 0.095 ..mu..Ci/kg) than at the high dose levels (0.30 to 2.9 ..mu..Ci/kg). These findings are in contrast with results on the retention of /sup 239/Pu(IV) in the liver of the beagle. The rate of decrease in hepatic retention of /sup 239/Pu diminishes with decreasing dose level. These opposing effects result from cellular necrosis which is an effect of the ..cap alpha.. irradiation from /sup 239/Pu. Skeletal retention equations are presented for each of the three higher dose levels of /sup 239/Pu and for the group of lower dose levels. These equations and those previously reported for the liver are used to calculate cumulative average radiation doses to the skeleton including marrow, and to the liver throughout the observed lives of the beagles, which were injected in young adulthood at six dose levels of /sup 239/Pu(IV) ranging from 0.016 to 2.9 ..mu..Ci/kg. In the skeletal dose calculations an experimentally determined valuemore » of (7.5 +- 1.0) percent for skeletal weight relative to body weight at injection is used. The concentration of plutonium in plasma has been measured from 1 min to 12.5 years following injection. A sum of four exponentials has been fit to the data. Samples were obtained from dogs at six dose levels, and from well, ill, and terminal dogs.« less

Retention of226 Ra was substantially lower in mature beagles injected at 5 years of age compared to corresponding values for young adult beagles injected at 17 months of age. As with young adults, ...

The retention and distribution of /sup 241/Am and /sup 65/Zn, given as the DTPA (diethylenetriaminepentaacetic acid) chelates, were studied in rats for 48 hr following intravenous injection. The retention and distribution of (/sup 14/C)DTPA, injected as the zinc chelate, were determined in rats and two beagles. At 24 hr postinjection (P.I.) rats retained 5% of the /sup 241/Am, 35% of the /sup 65/Zn, and 4% of the (/sup 14/C)DTPA. The loss of these nuclides from blood plasma could be described by the sum of two exponentials, and in rats the respective biological half-lives (t/sub 1/2/) for early times P.I. are t/sub 1/2/ /sup 65/Zn = 1.4 hr, t/sub 1/2/ /sup 241/Am = 0.75 hr, and t/sub 1/2/ (/sup 14/C)DTPA = 0.64 hr; the second component half-times for /sup 241/Am, /sup 65/Zn, and (/sup 14/C)DTPA are 0.2, 6, and 1.6 days, respectively. In beagles the initial t/sub 1/2/ for loss of (/sup 14/C)DTPA from blood was 0.77 hr and the second component t/sub 1/2/ was 4.4 days. The urine was the primary route of excretion in both species. In both rats and dogs, the kidney and liver contained the highest concentration of /sup 241/Am, /sup 65/Zn, and (/sup 14/C)DTPA. The concentrationmore » of /sup 65/Zn in the liver reached a plateau after approximately 4 hr. Significant amounts of /sup 65/Zn, however, were found in the lung, spleen, and femur. In rats the tissue : plasma concentration ratios of (/sup 14/C)DTPA in most organs studied peaked at about 4 hr P.I. In dogs the concentration of (/sup 14/C)DTPA in all soft tissues, except for the brain, exceeded that of plasma at 52 hr P.I. However, in the skeletal parts the concentration was less than in plasma.« less

Equations have been derived, from the results of total-body and partial-body counting and γ-ray counting of individual bones and soft tissues, which describe the retention of injected241 Am in the liver, in the nonliver tissue (including skeleton), and in the skeleton of young adult beagles. Retention was found to be dependent upon injection level, and different sets of equations were developed for dogs given about (a) 2.8 μCi/kg, (b) 0.9 μCi/kg, (c) 0.3 μCi/kg, and (d) 0.1 μCi/kg and less. Liver retention, R1, was characterized by a single exponential equation of the form $R_{{\rm L}}=ce^{-\beta t}$ , with c = 0.49 ± 0.04 and β = a function of injection level. Nonliver tissue was assigned a retention equation of the form $R_{{\rm NL}}=d+\alpha +J(1-e^{-ml})$ , with $d=0.102\pm 0.024\ e^{-2.22t}$ , α = 0.41 ± 0.04, and both J and m as a function of injection level. Skeletal retention was found to be about 0.885 ± 0.037 of nonliver retention with ...

The subcellular distribution of curium (/sup 243/ /sup 244/Cm) was studied in canine liver from 2 hr to 47 days after injection of 3 ..mu..Ci/sup 243/ /sup 244/Cm/kg of body weight. The pattern of distribution for Cm was similar to other trivalent actinide elements studied previously (Am, Cf). Initially (2 hr), most of the nuclide was found in the cytosol and at least 90 percent was protein bound. About 70 percent of the Cm was bound to ferritin, approximately 5 percent was associated with a protein of MW approximately 200,000, and approximately 25 percent was found in the low-molecular-weight region (approximately 5000). The decrease in the Cm content of cytosol, nuclei, and microsomes coincided with an increase in the amount associated with mitochondria and lysosomes. The concentration of the Cm in the mitochondrial fraction was higher than it was in the lysosomal fraction at each time studied. In the mitochondrial fraction approximately 30 percent of the Cm was bound to membranous or granular material, and 70 percent was found in the soluble fraction. The Cm concentration initially associated with cell nuclei was high but had diminished to 20 percent of the 2 hr concentration by 20 days post injection (PI).more » The subcellular distribution of Cm in the liver of a dog which had received the same dose and was terminated because of severe liver damage was studied at 384 days PI. The liver weighed 130 g and contained approximately 30 percent of the injected Cm. In contrast, a normal liver weighs 280 g and at 2 hr PI contains approximately 40 percent of the injected dose. The subcellular distribution of Cm in this severely damaged liver differed from the pattern observed at earlier times after injection. The relative concentration of Cm in the cytosol was doubled; it was higher in the nuclei-debris fraction; and it was lower in the mitochondrial and lysosomal fractions when compared to earlier times.« less

The distribution, retention, and excretion of 233U were measured in seven beagles injected intravenously with 2.8 /zCi 233U(VI)/kg (-3 mg/dog) in citrate buffer and sacrificed at times ranging from 1 to 726 days postinjection (PI). The concentration of U in plasma decreased rapidly, and for the first 24 hr PI could be described by a triphasic exponential equation with half-times of 0.92, 46, and 315 min. The uranium in the blood at 5 min PI was present as either a protein (transferrin-albumin) or a carbonate/bicarbonate complex. Up to 60% of the injected U was excreted during the first day, most of it in the urine. Whole-body retention was 17% at 7 days, 10% at 94 days, 7.6% at 1 year, and less than 5% at 2 years PI. Except for the kidney, only a small fraction of the injected U was retained in soft tissues. At 1 day PI, the liver contained 1.1% of the injected U and its subcellular distribution was similar to that seen with Pu(IV). At 1 day PI, 7.7% of the U was found in the skeleton; this amount decreased with a biological half-life of 883 days. Initially, U was deposited nonuniformly on bone surfaces with areas of intense hot spots adjacent to areas with intermediate and low concentrations. At later times redistribution and/or diffusion of U into the bone volume produced a deposition pattern intermediate to that of 239Pu (surface) and 226Ra (volume). The heterogeneity of surface deposits persisted, and isolated segments of the trabecular bone showed intense surface deposits at times greater than 1 year PI. Twenty-two percent of the injected U was found in the kidneys at 1 day PI with high concentrations localized in the proximal tubules. The initially high concentration of U in this organ was correlated with extensive diuresis and a persistent elevation in blood urea nitrogen. Damage to proximal tubules was attributed primarily to chemical toxicity but was aggravated by simultaneous a irradiation. It is postulated that the deposition of U on trabecular endosteal surfaces will induce osteosarcoma(s) following prolonged exposure to 233U or 232U.

The retention, distribution, and dosimetry of injected /sup 224/Ra were studied in six young adult beagles killed 0.04 to 7 days after /sup 224/Ra administration. Biological retention of /sup 224/Ra in the skeleton averaged 48% (37 to 60%), and soft-tissue retention decreased from 61% at 0.04 days to 6% at 7 days. At death there was more /sup 212/Pb in red blood cells and liver than the amount in transient equilibrium with the /sup 224/Ra in these tissues. There was an excess of 2/sup 12/Bi over /sup 224/Ra in kidneys, and a deficiency of both 2/sup 12/Pb and /sup 212/Bi relative to /sup 224/Ra in eyes and bone. The retention equations derived from the data suggest that the fractional escape of /sup 220/Rn produced in bone by /sup 224/Ra decay was roughly 0.08. From the retention of /sup 224/Ra, /sup 212/Pb, and /sup 212/Bi in the skeleton and selected soft tissues, it was calculated that the injection of 1 ..mu..Ci /sup 224/Ra/kg into an adult beagle having 75 g skeleton/kg body results in cumulative average doses to the blood-filled organs of 40 rad to the skeleton, 13 rad to the kidneys, 8 rad to the liver, 11 rad to themore » eyes, and 0.8 rad to the testes and ovaries.« less

Decorporation of Pu and Am by tetrameric catechoylamide (CAM) ligands has been investigated in beagles and mice. Eight dogs were injected intravenously (iv) with 237 + 239Pu(IV) + 241Am(III) citrate, and 30 min later, pairs of dogs were injected iv with 30 mumole/kg of 3,4,3-LICAM(C) [N1,N5,N10,N14-tetrakis(2,3-dihydroxy-5-sulfobenzoyl)tetr aazatetradecane, tetrasodium salt], 3,4,3-LICAM(S) [N1,N5,N10,N14-tetrakis(2,3-dihydroxy-4-carboxybenzoyl)te traazatetradecane, tetrasodium salt], CaNa3-DTPA, or each of the latter two ligands. Blood was sampled, and excreta were collected for 7 days, at which time the dogs were sacrificed and nuclide retention in liver and nonliver tissue was measured. Groups of five mice were each given 238Pu(IV) or 241Am(III) citrate iv; 3 min later 30 mumole/kg of a CAM ligand was injected intraperitoneally, mice were killed at 24 hr, and separated excreta and tissues were analyzed. In the dogs, average retention at 7 days of the injected Pu and Am, respectively, was as follows: 12 and 70% after treatment with a CAM ligand alone; 30 and 20% after DTPA; 12 and 20% after LICAM(S) plus DTPA; 90 and 89% without a ligand. In the mice, mean retention of the injected Pu and Am, respectively, was as follows: 14 and 66% after treatment with LICAM(C); 21 and 54% after LICAM(S); 91 and 87% without a ligand. In both species, about 99% of net Pu excretion (excretion with ligand - excretion without ligand) promoted in 24 hr by DTPA or LICAM(S) was in the urine, whereas about 10% of net Pu excretion promoted by the less hydrophilic LICAM(C) was in feces. Delayed excretion of both Am and Pu was significant in all ligand-treated dogs. Comparison of the nuclide content of tissues of ligand-treated mice with those of mice killed 3 min after nuclide injection indicated that the CAM ligands chelated circulating Pu and Am and prevented further deposition. In addition, the CAM ligands removed much of the presumably loosely bound Pu present in liver and skeleton at the time of ligand injection. LICAM(C) was more effective in removing Pu from liver and LICAM(S) was more effective in the skeleton. Moderate to severe uremia and histological evidence of cell killing in the distal tubules of the kidney were observed in the four dogs injected once with 30 mumole/kg of LICAM(S).(ABSTRACT TRUNCATED AT 400 WORDS)

Total-body and skeletal retention of 228Th were determined in a group of 104 young adult male and female beagles for about the first 7 years after the injection of 0.00159 to 2.76 muCi/kg. Ratios of 224Ra / 228Th , 212Pb / 228Th , and 212Bi / 228Th in the skeleton and in soft tissues of 20 beagles were measured as a function of time after injection. A humerus, femur, and ulna from 20 dogs dying 7 to 554 days after injection were sectioned, and the 228Th concentration was obtained for each piece. Percentage biological retention in the skeleton of 228Th at t days after injection could be described as 68.1 e-0. 000180t . Ratios of daughter-to-parent activity in soft tissue showed no definite trend with dose level or time and averaged Ra/Th = 0.56, Pb/Th = 0.83, and Bi/Th = 0.91, whereas the ratios for the skeleton varied with both dose level and time. Ratios of activity in the skeleton from lowest to highest dose level after 2 years following injection ranged between Ra/Th = 0.88 to 0.95, Pb/Th = 0.78 to 0.92, and Bi/Th = 0.77 to 0.90. Retained 228Th was deposited most heavily in parts of the skeleton with much trabecular bone, much bone surface area, and high bone remodeling rates. No changes in this deposition pattern could be discerned during the 554 days over which the measurements of sectioned long bones were made.

The distribution of241 Am in the thyroid glands of three dogs after intravenous injection was studied. The glands were removed, frozen immediately in liquid N2, and counted. Later the thyroid tissue was homogenized and separated into fractions by differential centrifugation. Eighty-five to ninety per cent of the241 Am was found with the crude fibrous residue. Combining this fibrous material with cell nuclei and membranes accounted for more than 90% of the total activity found in the thyroid. Only very small quantities were found with microsomes, mitochondria, and in the 105,000 × g supernatant. It is concluded that241 Am is selectively bound by the connective tissue elements of the thyroid. The concentration of241 Am in thyroid connective tissue was found to be 337 times that of a sample of subcutaneous loose connective tissue and 4 times that of connective tissue derived from the liver.

This paper is another in the series of studies of the excretion, retention, and distribution of Th228 and its decay products in adult beagles (1-4). These metabolic studies are in turn a part of the long-term studies of the biological effects of Th228 and other radionuclides in beagles (5). This communication reports measurements of the activities of Ra224 and Pb212 in one humerus, the third lumbar vertebra, the liver, the spleen, and the kidneys through 3.5 years after injection. The activity of Bi212 in kidneys is also presented. These results, together with those for Th228 (3), are then used to estimate average radiation dose rates for bone, liver, and kidneys.

In the canine liver ${}^{241}{\rm Am}({\rm III})$ is distributed ubiquitously at the subcellular level. Significant concentrations were measured in the hepatic connective tissue, the nuclear fraction, the cytosol, the mitochondrial fraction, and the microsomal fraction, and increase in the order given. In the cytosol most of the americium is bound by ferritin. It was also bound by lipofuscin, which was identified histochemically, and by an unidentified material with a molecular weight of 1500. ${}^{241}{\rm Am}\text{-ferritin}$ was isolated in pure form by gel electrophoresis. (Since ferritin that is heavily loaded with iron is found in the microsomal fraction, the high concentration of americium observed in the microsomal fraction could be an anomaly.) The transfer of americium from transferrin to ferritin was shown to occur in vitro as is the case with ${}^{239}{\rm Pu}({\rm IV})$. Partial identification of ${}^{239}{\rm Pu}\text{-ferritin}$ in hepatic cytosol was also achieved. Two concurrent mechanism...

When human blood serum that had been tagged in vitro with monomeric PuIV was subjected to gel filtration, most of the PuIV was found in two molecular-weight ranges, that of the low-molecular-weight serum proteins and that of small molecules and ions. The distribution of PuIV between the two peaks was variable. In addition, a small amount was eluted in the highest-molecular-weight fraction. By a combination of gel filtration, ion exchange, and electrophoresis the PuIV protein complex was isolated, and the protein was identified as transferrin, the protein that transports iron. PuIV appeared to be bound at the iron-binding sites, and the reaction between PuIV and transferrin was shown to be reversible. The stability constant of the complex is high but less than that of the very stable FeIII transferrin complex. The variation in distribution of PuIV between transferrin and the low-molecular-weight region thus is related to the variation in the amount of iron bound by the transferrin.

BRUENGER, F. W., STEVENS, W., AND STOVER, BETSY J. Americium-241 in the Blood: In Vivo and in Vitro Observations. Radiation Res. 37,349-360 (1969). The concentration in canine blood of trivalent 241Am that had been injected intravenously decreased at a rate comparable with those of divalent alkaline earths. A small fraction of the 241Am was associated with blood cells, but the major part was in the serum. Complexes with serum proteins were demonstrated in sera obtained at 1 and 5 minutes after injection. Americium-241 complexes with both transferrin and albumin were identified and isolated in in vitro experiments with human sera. The AmIII complex with transferrin is less stable under physiological conditions than either the FeIII- or PuIV-transferrin complex. Evidence was obtained for a third AmIII-protein complex.

Small amounts of a number of metal ions have been found in melanins obtained from natural sources, and ${}^{140}{\rm Ba}$ and226 Ra have been found to deposit in tissues containing melanin. In the studies reported here, some aspects of the chemical nature of the incorporation were investigated with radionuclides of high specific activity. The uptake of metal ions by synthetic melanin from a physiological buffer increased with decreasing hydrated ionic radius, decreased with decreasing pH, and decreased in the presence of chelating agents. Replacement of melanin-bound ${\rm Ra}^{++}$ with H+ demonstrated the reversibility of the reaction. Comparative studies with synthetic melanin and natural melanin plus its protein showed that the role of the protein in the reactions is small. Both the reaction characteristics and the chemical structures of melanin and of weak acid cation exchangers are similar, which suggests that metal ions and melanin interact by ion exchange.

The interaction between Pu239 and the constituents of the blood is a subject that has been pursued by a number of workers (1-4). It has been established that Pu239 is bound to plasma proteins (1-4). The identities of the particular blood proteins that bind plutonium are only tentatively known (1, 2), and they are thought to be A-globulins. The separation and identification of the proteins to which plutonium is bound is difficult, since neither the chemistry of proteins nor that of plutonium is simple. Plutonium can exist in four oxidation states in aqueous solution. The studies reported herein are limited to one oxidation state, PuIV, whereas earlier studies used PuVI (1, 2). This small highly charged ion (PuIV) essentially does not exist as such in systems of biological interest. Instead it binds with many anions, including OH(hydrolysis), to form chemical entities of lower net charge. Blood serum or plasma offers an enormous number of ionic binding sites. Thus, PuIV in blood is undoubtedly involved in an unknown number of unidentified competitive equilibria. These characteristics exclude the use of ion exchange in these studies, for the ion exchange system would introduce new competitive equilibria. Electrophoretic separation of the proteins is also excluded, since PuIV could be released from the protein and move independently under the appropriate (unknown) voltage gradient (5; also F. W. Bruenger, unpublished observations). The advent of gel-filtration as described by Flodin and Killander (6) has provided a new approach to this problem. This method of separating proteins is primarily mechanical in nature, in contrast with the chemical and electrical methods mentioned above, and hence it is much less likely to perturb the PuIV-protein system. The work reported here is a preliminary study of the separation of the proteins that bind PuIV, using the method of gel filtration.

The distribution of PuIV in blood plasma of beagles was determined as a function of time after injection of PuIV in a citrate buffer solution. The plasma was subjected to gel filtration on G100-G200 columns. Significant amounts of PuIV were found in the region of low-molecular-weight proteins as well as in the molecular weight range of small molecules or ions. The amount of PuIV bound to protein increased during the first few hours after injection, reached a maximum at about 7 hours, and then decreased steadily. Concomitantly the PuIV found in the region of low-molecular-weight compounds decreased continuously from 5 minutes to 30 days. The decrease in the amount of PuIV in the region of low-molecular-weight compounds occurred more rapidly than did the decrease in PuIV associated with low-molecular-weight proteins. The proteins that bound PuIV were identified as transferrin and albumin. These protein-PuIV complexes were separated by ion-exchange chromatography. Their identity was confirmed by electrophore...