ARCHIVES
OF
BIOCHEMISTRY
AND
BIOPHYSICS
79,
200-205
(1969)
Absorption and Distribution of Cr51in the Albino Rat] R. D. MacKenzie,2’ 3 R. A. Anwar, R U. Byerrum and C. A. Hoppert From the Kedzie Chemical Laboratory, Michigan State University, East Lansing, Michigan
ReceivedJune 11,
1958
INTRODUCTION
Extensive industrial use of chromium and disposal of hexavalent chromium wastes have resulted in contamination of drinking water in the United States. Well waters have been reported to contain up to 25 parts per million (p.p.m.) chromium (l), whereas a survey of 24 municipal water supplies showed concentrations from 0.001 to 0.04 p.p.m. (2). Although the complete nature of the chromium action in living organisms has not as yet been elucidated, small quantities of this element will activate several enzyme systems (3-5). On the other hand, chromate administered at 134 p.p.m. in drinking water for 2-3 months produced kidney lesions and liver damage (6). Since chromate consumption by animals and man is occurring in varying quantities in water, a knowledge of its absorption, distribution, and excretion is essential. Visek and co-workers (7) fed trivalent CrK1to rats and found that most was excreted in the feces within 4 days and only about 0.5% was absorbed. Since their investigation did not include orally administered hexavalent chromium, high specific activity chromium-51 was used to study the absorption, distribution, and excretion of chromate after administration of nontoxic quantities. EXPERIMENTAL Three weighing
experiments were begun using albino rats of the Sprague-Dawley strain about 300 g. Half the animals in each experiment were fasted for 24 hr. be-
1 Presented in part at the annual meeting of the American Institute of Nutrition, Chicago, Illinois, April l&1957. The work was supported by a grant from the National Institutes of Health, U. S. Public Health Service. 2 Part of a thesis presented to the School for Advanced Graduate Studies in partial fulfillment of requirements for the degree of Doctor of Philosophy, Department of Chemistry, Michigan State University, East Lansing, Michigan. a Present address: Department of Biochemistry, Wm. S. Merrell Co., Cincinnati 15, Ohio. 4 Stock diet consists of the following (in ‘% by weight) : ground yellow corn meal, 32.5; ground whole wheat, 25.0; powdered whole milk, 22.5; linseed meal, 10.0; alfalfa, 6.0; brewer’s yeast, 3.0; sodium chloride, 1.0. 200
CHROMIUM-51
fore administration
IN
RBTS
201
of chromium, whereas the other half were fed a stock diet4 ad
libitum.
The radioactive sodium chromate solutions to be administered were prepared by oxidizing Cr61C13with Hz02 in 6 N NaOH. The chromate solutions were adjusted to pH 7.5 with 6 N HCI, and the trivalent solutions to pH 5 before administration. After administration of radioactive chromium, animals u-ere killed and selected tissues were oxidized with concentrated nitric acid and 300/, HEOZ . A 2-ml. portion, represent’& all or a known amount of each sample, was evaporated on a glass planchet and counted in a well-type scintillation tube containing a thallium-activated sodium iodide crystal. Only counts greater than 1.5 times the background count were considered to have a significant amount of radioactivity.
I&xperiment I. Absorption and Distribution
of Orally Adm,inistered Chromate
Sodium chromate solution, 1.5 ml., containing 57 pg. chromium and having a radioactivity of 22.8 PC., was administered by stomach tube to each of the rats. The quantity of chromium administered was similar to the amount a rat, would receive per day while drinking water containing 2 p.p.m. chromium. Rat,s drinking water containing this level of chromium showed no symptoms of toxicity during a year (8). After the rats were administered the radioactive chromat,e, they were placed in metabolism cages. The fasted animals did not receive food until 2 hr. aft,er t,he chromate was fed. Feces and urine were collected separately. This was facilitated by spraying the met,abolism cages with an acyrlic plastic and placing a similarly treated small-mesh screen at, the bottom of the funnel-type metabolism cage to reduce fecal contamimtion of the urine. The feces were removed from t,he cage daily to minimize this VOW tamination. The rats were killed at intervals of 1, 7, and 14 days following administration of the radioactive chromium. Liver, kidney, stomach (plus contents), intestine (plus contents), blood, spleen, brain, lung, and submaxillary gland as well as the urine and feces were selected for analysis.
h’xperiment II. Red Cell Absorption of Chrom,iu,m Administered by Stomach Tube There is no satisfactory way known to differentiate between trivalent and hexavalent chromium in most tissues. Gray and Sterling (9), however, have shown that eryt,hrocytses will absorb chromate ions and will contain nearly all the hexavalent, chromium in the blood, whereas trivalent chromium remains almost exclusively in blood plasma. Therefore, this difference in absorption by red cells was used as a means of distinguishing between the two forms of chromium at the time chromiunl was absorbed into the blood stream. Sixteen rats received by stomach tube 125 PC. of either hexavalent or trivalent chromium in 1 ml. solution containing 131 pg. chromium. Four hours after the rats were administered the chromium, they were anesthetized with ether and blood was removed from the heart. The syringes were rinsed with heparin solution to prevent clotting. The blood was centrifuged, and the red cells and plasma were prepared for counting as previously described.
Experiment III.
Red Cell Absorption of Chromium Injected into the Intcstinc
To determine the role of the stomach wall in chromium absorption, the stomach was bypassed by injecting chromium directly into the lumen of the intestine. Three
202
MACKENZIE,
ANWAR,
BYERRUM
AND
HOPPERT
groups of eight animals were given (a) Na&rOI , (b) CrC13 , or (c) 50:50 mixture of hexavalent and trivalent chromium. Half the animals in each group were fasted, the other half fed ad libitum. The rats were anesthetized with ether, and an incision of about 2 cm. was made just below the lower rib in the center of the abdomen. The chromium solutions were injected into the lumen of the intestine about 4 cm. below the stomach. The radioactivity of the solutions was the same as in Expt. II and, therefore, the data of the two experiments are directly comparable. Four hours after injection, the animals were sacrificed and the blood was measured for radioactivity as in Expt. II. RESULTS
Absorption and Distribution
of Orally Administered Chromate
The average radioactivity of several tissues at selected intervals after feeding radioactive chromium are found in Table I. The values are percentages of the administered dose, contained in each tissue, and each number is the average of four analyses. Little or no radioactivity could be detected in brain, salivary gland, or lung. Liver showed a maximal uptake of about 1% of the administered dose, whereas kidney, blood, and spleen had a maximal content of 0.1-0.2 %. Spleen initially had a low content of radioactive chromium, but this level persisted unchanged for 2 weeks. Generally the tissues of the fasted animals contained more chromium than the nonfasted. If the quantity of chromium excreted in the urine is a measure of the quantity of chromium absorbed, it can be seen that about 6% of the administered chromium was absorbed by the fasted animals and about 3 % by the nonfasted. The greatest part of the administered dose was excreted in the feces.
The Distribution
TABLE I and Retention of Chromate in the Rat after a Single Oral Dose Per cent of the administered
Time, days Condition
Tissue Stomach Intestine Blood Liver Kidney Spleen Urine Feces Total, y.
1
dose/total tissue 14
7
Fasted
Nonfasted
Fasted
Nonfasted
1.964 26.78 .17 1.03 0.14 0.02 2.9 64.1 97.1
2.22 18.2 0.03 0.21 0.03
