Document oDq0dwg09EmvB2OJDLa8vMDKE

Reprinted from Tk e Jo t h u s t a i. os* Nu t r it io n Vol, 100, No. 1, January 1970 The American Institute of Nutrition 1970 Zirconium, Niobium, Antimony, Vanadium and Lead in Rats: Life term studies*1 2 HENRY A. SCHROEDER2 MARIAN MITCHENER a n d ALEXIS P. NASON Department of Physiology, Dartmouth Medical School, Hanover, New Hampshire, and Brattleboro Memorial Hospital, Brattleboro, Vermont 05301 ABSTRACT To evaluate innate effects of the trace dements zirconium, niobium, antimony and vanadium, and to reevaluate those of lead, 603 rats of the Long-Evans strain were fed a diet containing relatively small amounts of these elements in an environment reasonably free of trace contaminants. Groups of 100 or more divided as to sex' were given S ppm (as metal) either zirconium,, niobate, anthnonite, or vanadyl ions, and ^5 ppm lead (males only) in drinking water from the time of weaning until natural death, and compared With an equal number of controls. Chromium 1 ppm was in the water. These doses were tolerable for growth which was enhanced in the male niobium group. Innate toxicity in terms of life span and longevity occurred in the antimony groups. In rats given antimony, nonfasting serum glucose levels were lower than fasting, an unusual finding. Increased incidences of glycosuria occurred in the zirconium, niobium and lead groups. Serum cholesterol was abnormal in the antimony and vanadium groups. Antimony and lead accumu lated in soft tissues, the former with age. No element was tumqrigenic. Lead-fed males lost weight from 24 to 30 months of age and their coats were poor, the only signs of toxicity. A previous series of chromium-deficient rats given the same dose of lead showed early mortality, shortened life span and decreased longevity. Chromium may be antagonistic to lead toxicity. Innate biological effects of small doses of trace elements in drinking water, given to mice and rats from the time of weaning until death are being studied in an en vironment relatively free of contaminating metals. The purposes of these time-consum ing experiments are to ascertain whether an orally given element -- usually a metal -- has favorable or adverse effects on growth and survival or is inert biologically, and whether or not a common human chronic disease is reproduced in these ani mals. We have reported effects of cad mium, lead, chromium and its deficiency (1-5), arsenic, germanium and tin (6) in rats. Under study are selenium, tellu rium, nickel and molybdenum. The present report concerns zirconium, niobium, anti mony and vanadium. The previous study on lead was made in chromium-deficient rats, a fact of which we were not aware at that time (2); therefore it was repeated in males supplemented with chromium. All of these trace elements are found in human tissues, and have been given to mice (7, 8). METHODS The diet of seed rye flour (60% ), dry skim milk (30% ), com oil (9% ) and io dized sodium chloride (1% ) with added vitamins and ferrous sulfate, the basal drinking water containing soluble salts of zinc (50 ppm), manganese (10 ppm), cop per (5 ppm), trivalent chromium (1 ppm), cobalt (1 ppm) and molybdenum (1 ppm), and the environmental conditions of the laboratory have been described previously (8). To the water was added one of the following as trace metal concentration: zirconium sulfate 5 ppm, sodium niobate 5 ppm, potassium antimony tartrate 5 ppm, vanadyl sulfate 5 ppm or lead nitrate 25 ppm.s The diet contained 2.66 ng/g zir- Received for publication June 20, 2969. 1 Supported by Public Health Service Research Grant no. HE-05076 from the National Heart Institute, Con tract DA 2595 from the U.S. Army, Cooper Laborator ies, Inc., and the CIBA Pharmaceutical Products, Inc. 2 Address for request of reprints: 9 Belmont Avenue, Brattleboro, Vermont 05301. s Previous reports (1, 2) have stated that the dose of lead was 5 ppm. The dose was incorrect. A typo graphical error in our laboratory instructions resulted in 5 times the desired dose being used, both in the experiments begun in 1960 and in those here reported which were begun in 1965. When the error was dis covered, the larger amount was continued. J. NuxainoN, 200: 59- .i-Ht N36918 60 HENRY A. SCHKOEDER, MARIAN MITCHENER AND ALEXIS P. NASON conium, 1.62 ug/g niobium, 3.2 ug/g va Numbers of rats dying during die 3 weeks nadium and 0.2 ug/g lead. Antimony was of the epidemic were as follows: Males, not detected. All values are in terms of dry control 19, zirconium 5, niobium 12, anti weight. mony 9, lead 22, vanadium 17; females, Random-bred pregnant female rats of control 12, zirconium 4, niobium 6, anti the Long-Evans strain were purchased4 mony 3, vanadium 17. These animals were and their offspring bom and weaned in our removed from the series and survival laboratory. Groups of 50 or more of each curves corrected from that time, using the sex, 4 in a cage, were given the basal water smaller numbers. and one of the metals. Lead was given only to males. Controls received the basal water. The total number of rats used was 603. Animals were weighed at weekly inter vals from weaning time to six weeks of age, then at monthly intervals. They were disturbed for measurements of blood pres sure which required anesthesia, for sam pling of blood which required wanning, and for cleaning their cages at weighing time. Animals dying a natural death were weighed and dissected; grossly visible tu mors, other lesions, and heart, lung, kid ney, liver and spleen were described and fixed in Bouin's solution. Portions of the same tissues were frozen in polyethylene bottles and later ashed and analyzed for the elements given. A low-temperature asher6 was used for antimony; for other elements tissues were ashed at 450 in muffle furnaces. Analytical methods and their sensitivities have been reported in the study on mice (7); for lead, both colori metric methods (2) and atomic absorption spectrophotometry were used. Extraction of antimony in 2% ammonium pyrrolidine dithiocarbamate and analysis by atomic absorption spectrophotometry improved sensitivity considerably above that previ ously found (7). The sensitivity of the method for vanadium was suspect; there fore, analyses for this element were not done. Methods for serum constituents, uri nalysis and blood pressure have been re ported in other similar studies (6). Fasting serum glucose levels were measured in animals deprived of food for 18 hours; nonfasting levels came from animals al lowed free access to food. During the 4 years of these experiments, an epidemic of virulent pneumonia struck the rat colony, killing a sizable number of animals before it was controlled by oral RESULTS Growth rq.tes. None of the four metals affected growth or mature weights of fe males consistently (table 1). The oldest fe males in all but the vanadium group were significantly heavier than their controls. Males fed zirconium were heavier than the controls at three intervals and lighter at two. Males fed niobium were heavier at six intervals. Antimony, vanadium and lead had negligible effects on growth and ma ture weight. Therefore, niobium appeared to enhance the growth of male rats; the opposite effect was observed in mice of both sexes (7). Survival and longevity. In table 2 are given mean and median ages and 75% and 90% life spans of the various groups. There were no significant differences from controls in the zirconium, niobium, lead and vanadium groups. Antimony, however, was innately toxic, males surviving 106 days and females 107 days less than the controls at median life spans, and 70 and 165 days less when 90% were dead. Sur vival curves are shown in figures 1 through 4. This toxicity of antimony was not seen in mice (7). Innate toxicity was also demonstrated when longevities, defined as the mean age of the last surviving 10%, were calculated. In the antimony groups these ages were significantly reduced compared to control ages. The oldest antimony-fed male lived 202 days and the oldest female 152 days less than their controls. Males of the nio bium group also had decreased longevity, and the last survivor was 171 days younger than his control. The opposite effect was seen in male mice (7). Blood and urinary findings. Fasting se rum glucose levels are given in table 3. penicillin. Fortunately, enough rats in each 4BLU: (LE) strain, Blue Spruce Farms, Inc., Alta- group survived to continue the experiment. mont. New York. s Tracerlab 500-A, Richmond, Calif. 9 DUP050312629 TRACE METALS IN RATS 61 TABLE X Weights of rats given zirconium, niobium, antimony, vanadium and lead Age Control days g Males 30 72.1* 4.2* 60 189.5* 6.0 90 270.0* 8.9 120 312 * 9.3 ISO 341.5* 8.9 180 364.7* 8.7 360 443.8*14.9 540 507.4*:16.4 Zirconium S 88.5*2.39 204.0*4.7 285,7* 6.2 313.7*8.7 377.5* 6.3 9 392.0* 6.5 * 405.2* 8.4* 469.0*8.19 Niobium 0 91.9* 1.4 9 193.0* 3.7 309,8* 4.39 362.6 * 4.2* 385.3* 5.9* 392.8 * 5.5* 473.1* 6.9* 497.1*11.3 Antimony 9 89.3* 1.5* 207.7* 4.3* 257.7* 3.8 312.0* 7.2 334.4* 9.4 350.0* 8.7 453.0* 9.0 475.0*10.2 Vanadium 9 76.4* 3.9 184.2* 6.3 253.6* 9.5 306.7* 8.7 331.4* 2.7 370.0 * 9.3 427.7*11.7 472.8*17.3 Lead 9 55.7* 3.1s 171.2* 3.3 4 265.8* 4.5 314.0* 6.8 353.1* 8.1 369,7* S3, 443.7* 8.1 461.5* 12.0 Females 30 64.7* 2.1 60 154.2* 6.0 90 197.1* 5.4 120 225.2* 5.3 150 233.8* 4.0 180 250.5* 4.9 360 262.6* 5.9 540 262.4* 9.8 82.1*2.1 * 159.3* 1.9 204.4* ID 232.0* 2.7 250.0* 2.5* 263.7*4.3 267.0*4.2 299.25.32 89.7* 0.9* 162.3* 3.3 216.4* 3.3 239.7* 3.3 247.0* 3.9 262.4* 3.1 271.5* 7.0 282.8* 4.3 82.9* 1.63 163.5* 2.4 199.8* 2.6 217.6* 4.4 238.2* 2.9 246.2* 3.2 283.1* 3.9* 295.2* 4.0 63.2* 2.7 162.1* 3.7 193.4* 5.9 223.0* 6.1 235.7* 2.7 246.0* 9.3 274.1*11.7 282.9*10.0 i Mean SEM. Differences from controls have been analyzed by Student's t test, giving the P values of foot notes 2,3.4 and 5. if < 0.005. Survived and longevity of rats, days Metal No. Mean rats age 50% dead 75% dead 90% dead Controls 52 819 872 974 1057 5 54 910 912 1050 1157 Zirconium <3 56 870 881 1019 1077 9 58 935 947 1099 1187 Niobium d 52 853 892 959 1035 9 56 994 998 1P87 1207 Antimony<J 51 746 766 955 987 9 59 797 805* 900 992 Leadc? 52 877 883 931 951 Vanadium<? 52 813 860 918 1091 9 61 922 961 1051 1170 i Mean + s e m of last 10% of animals surviving, t Differs from controls, P < 0.001, by Student's t test, s Differs from controls, P < 0.025, by chi-square analysis. Last 1232 1347 1189 1291 1061 1342 1030 1195 1262 1218 1313 Longevity1 1160* 27.8 1304*36.0 1127* 23.0 1247*17.4 1045 * 4.r* 1247* 21.3 999 7.8 2 1092*30.0 1071*66.0 1147*35.5 1269*34.5 Females fed zirconium and vanadium showed somewhat elevated levels compared to their controls. The lowest male value was in the lead group. Nonfasting glucose levels were higher than fasting ones in all groups but the antimony, where they were lower. In males, differences were signifi cant in all but the vanadium group. In the female zirconium and antimony groups the two glucose levels were not signifi cantly different. This unusual lack of re sponse of serum glucose to food in anti mony-fed rats of both sexes has not been duplicated in groups of rats fed seven other elements, and occurred only in males given selenate and females given tellurite. Glycosuria8 was found in 23% of 90 controls, 43% of 23 in the antimony group, 52% of 56 in the zirconium group, 63% of 16 in the lead group, 71% of 24 in the niobium group and 12% of 17 in the va nadium group. By chi-square analysis the differences from the controls were signifi cant in the zirconium (P < 0.01), the lead (P < 0.005) and the niobium groups (P 0 Glucose in the urine was measured "by Combiatix (Ames Co., Elkhart, Ind.) on nonfasting mature rats about 18 months of age. DUP050312630 Fig. 1 Survival curves of male rats. Solid line, controls; dotted line, zirconium-fed; dot- dashed line, niobium-fed; dashed line, antimony-fed. The lessened survival and longevity of the group fed antimony is apparent, although there was only one significant difference by chi-square analysis at the 33-month interval (P < 0.0005). There were 52 to 56 rats in each group. MONTHS Fig. 2 Survival curves of female rats. Solid line, controls; dotted line, zirconium-fed; dot-dashed line, niobinm-fed; dashed line, antimony-fed. The decreased survival of the group fed antimony is obvious; it was significant by chi-square analysis at 30 and 33 months of age. The zirconium group had a significantly lower mortality at 24 and 27 months of { age, as did the niobium group at 24 months of age. There were 56 to 64 rats in each group. <0.0001). No significant differences in proteinuria were found between the sev eral groups, although niobium-fed animals of both sexes and vanadium-fed females showed no samples with 3- or 4-plus pro tein, whereas 17 to 31% of the others had these amounts. There was half as much protein in the urine of niobium-fed fe males as in the controls. Fasting serum cholesterol levels, previ ously published (5), are shown in table 4 for comparison. In both sexes significant i l DU P050312631 TRACE METALS IN RATS 63 Fig. 3 Survival curves of vanadium-fed rats. Solid liner male controls; dotted line, males fed vanadium; dash-dotted line, female controls; dashed line, females fed vanadium. By chi-square analysis, the curve of the vanadium-fed females differed significantly from that of their controls at 24 months (P < 0.01) and at 27 months (P<0.05), mortality being less. There were 52 to 61 rats in each group. Fig. 4 Survival curves of lead-fed male rats. Solid line, controls; dotted line, present series of rats fed 25 ppm lead, with 1 ppm chromium in water of both groups. The hatched line is the curve of the previous series of lead-fed rats (2) without chromium in water, and includes the nearly 20% early mortality. This curve was significantly lower (P < 0.05 to < 0.0005) than that of their controls (not shown) from 18 to 30 months of age when the early mortality was excluded (2). There were 52 to 62 rats in each group. differences from the controls occurred in the vanadium and antimony groups and in the male zirconium and female niobium groups, values being higher in males and lower in females. The high level in females was probably the result of insufficient chro mium, for we have shown that 5 ppm is required for normal levels of serum cho- DUP050312632 64 HENRY A. SCHROEDER, MARIAN MIXCHENER AND ALEXIS P. NASON TABLE 3 Serum glucose and cholesterol levels in rats fed zirconium, niobium, antimony, lead and vanadium1 Glucose Fasting Nonfastmg tuoiesteroi Males Controls Zirconium Niobium Antimony Lead Vanadium days 718 921 889 852 737 697 mg/100 ml 106.53:3.61 106.13:9.9 112.33=5.7 114.93:6.8 81.83=3.6 8 107.8 8.9 134.45.1 133.34.7 137.59.9 94.56.24 147.3 5.7 121.3 5.4 mg/100 ml 77.52.1 89.75.6* 75.7 2.7 97.6 4.9 * 86.6 6.5 91.65.1* Females Controls Zirconium Niobium Antimony Vanadium 698 921 907 859 711 79.6 8.2 111.45.64 94.3 5.3 86.2 5.0 96.22.5* 114.2 5.4 120.5 3.3 107.5 8.0 82.5 7.0 8 116.1 5.9 116.0 d; 6.0 100.7 9.0 78.6 4.8 97.0 5.6 8 67.9 9.2* 1 Differences between fasting end nonfasting levels of glucose were significant in all groups of mates CP < 0.03 to P < 0.001) out the vanadium, and in the control CP < 0.001), niobium (p < 0.0S) and vanadium groups (P < 0.00S) of females. Note decline in nonfasting level in antimony group in both sexes. Data on cholesterol previously published < 5). Twelve animals in each group. 8 Mean s e m. Differences from comparable controls have been analyzed by Student's t test, giving the P values of footnotes 3,4 and 3, P < 0.01. *P < 0.005. P< 0.025. TABLE 4 Mean heart and body weights of rats and gross tumors Metal No. rats Weight autopsded at death Heart weight Controld $ Zirconium <f 9 Niobium<J 9 Antimonyc? 9 Leade? Vanadium cf 9 Normal,1 billed age 315 days c? 9 fir mg 50 334 1498 39 234 949 46 324 1280 53 244 1019 46 346 1315 52 234 992 50 340 1215 47 243 982 43 290 1204 31 320 1223 39 232 988 5 517 1311 5 300 904 > Data from Blue Spruce Farms, Inc., Altamont, N. Y. Ratio x 1000 HW/BW 4.49 4.06 3.95 4.18 3.80 4.24 3.57 4.04 4.15 3.83 4.26 2.54 3.01 Tumors No. % 10 20.0 14 35.9 7 15.2 20 37.7 7 15.2 16 30.8 6 12.0 18 38.3 7 16.3 13 42.0 15 38.4 lesterol in females, whereas 1 ppm appears to be enough for males (5). Elevated systolic blood pressure was not found in 10 animals of each sex and each group. Heart weights. Mean heart and body weight at death and their ratios axe shown in table 4, There were no obvious differ ences in body weights among the various groups. The hearts of males fed zirconium, niobium, antimony and vanadium weighed 14.6%, 12.2%, 18.9% and 18.3% less, respectively, than those of the controls, whereas the hearts of females weighed 3.5 to 7.4% more. Tumors. None of these 5 metals was tumorigenic (table 4), as evidenced by vis ible tumors at necropsy. There were more than twice as many tumors in females as in males (P < 0.0001). No metal signifi- =. <: Si ; ii* Vv: ' : M i-M DUP050312633 TRACE METALS DM RATS 65 cantly suppressed the incidence of tumors. Accumidation of elements in tissues. Evidence for accumulation of zirconium in rat tissues was poor (table 5). The same phenomena were observed in mice (7). Deposition of antimony in tissues was clear (table 6) as it was in mice (7), none being detected in control samples and in the diet. Furthermore, antimony accumulated with age in rats. From ages 279 to 1070 days, or about 9 to 35 months, pooled samples showed a tendency to increase in concen tration, with a correlation coefficient (r) of 0.525 (P < 0.05). The mean concentra tion of antimony in five tissues of all rats analyzed was 13.1 ug/gram. No obvious accumulation of niobium occurred in rats fed this element (table 7), with the pos sible exception of spleen, which was not controlled. Niobium accumulated in spleens of mice (7). There was a definite, but surprisingly small, accumulation of lead in soft tissues (table 8), considering the dose used. Blanching of the incisor teeth, which has occurred in a number of older rats, Organ, sex TABLE 5 Zirconium in rat tissues, wet weight1 Controls 2 Fed zirconium 8 No. rats Mean No. N.D. No. rats Mean Difference Males Kidney Liver Heart Lung Spleen Tumors Mean* mis 42 10.5 4 35 10.5 3 39 7.8 26 * 35 6.4 3 33 3.1 3 3 2.7 1 7.8 -- M/9 10 9.7 9 11.2 10 17.2 3 7.4 10 35.1 5 ---- -- 17.7 M/9 -0.8 + 0.7 *4- 9.4 + 1.0 432.0 -- + 9.9 Females Kidney Liver Heart Lung Spleen Tumors Mean7 38 5.4 9 38 3.4 0 31 7.7 5* 38 9.6 0 31 22.3 4 3 4.5 1 -- 9.3 -- 37 12.5 35 6.7 37 9.9 37 10.5 35 19.9 -- -- -- 11.9 + 7.1 +6.3 +2.2 + 0.9 --2.4 +2.6 * Tissues were pooled in groups of 2 to 16, usually 4 to 7. 2 Control rats were 144 to 900 days old. * Zirconium-fed rats were 427 to 1172 tissues. Limit of detection of the method days old. AH zircomum-fed rats had was 0.01 to 0.017 ag/gram. wet weight. zirconium N.D., not in their detected. Differences between controls and zirconium-fed rats have been treated by chi-square analysis, resulting in the P values of footnotes 4, 5 and 6. 4 P < 0.001. s P < 0.005. P< 0.05. ? Excluding tumors. TABLE 6 Antimony in rat tissues. dry weight, all ages1 Organ Controls No. rats Mean Fed antimony No. rats Mean Range Kidney Liver Heart Lung Spleen M/9 9 N.D.S 9 N.D. 9 N.D. 9 N.D. 9 N.D. M/9 58 10.14 4.6-34.4 40 11.57 1.7-60.1 66 12.10 4.0-28.0 34 17.67 4.0-30.0 62 15.97 4.1-53.5 * Tissues were pooled in lots of 2 to 8. .. * Not detected. Limit of detection of antimony by the method used was 0.9 jig/gram dry weight, or 0.25 jug/xnilliliter MIBK. DU P050312634 665;c HENRY A. SCHROEDER, MARIAN MITCHENER AND ALEXIS P. NASON TABLE 7 Niobium, in rats, wet weight1 1 'Organ Controls 2 No. rats No. N.D.4 Mean Fed .niobium No. No. rats N.D.4 m/s Kidney 35 0 1.71 `liver 26 15 0.51 Heart 28 15 2.27 Lung 30 0 1.50 Spleen ---- -- Mean excluding spleen 119 30 1.53 29 S 29 7 29 4 29 8 29 4 116 27 - * Tissues were pooled in groups of 2 to 15, usually 4 to 7, / Controls were 300 to 911 days old. s Niobium-fed rats were 276 to 780 days old. ' not detected. Limit of detection by the method was 0.1 jug/gxazu wet weight. TABLE 8 Lead in rat tissues,1 wet weight Mean MS/9 2.07 0.52 1.73 1.01 7.23 1.33 , Organ Controls No. rats Mean Fed lead No. rats Mean Kidney Liver Heart Lung Spleen Mean s/a 21 0.50" 22 0.64 12 0.82 26 1.24 15 2.31* 96 1.06 s.a/a 40 2.65 9 30 2.06 25 1.28* 33 1.04 23 1.04* 151 1.71 * Tissues were pooled in lots of 2 to 7, s In control rats, lead was not detected in 2 kidneys and 2 spleens. In lead-fed rats, it was not detected in 5 hearts and 4 spleens. Limit of detection by the method was 0.05 mb/gram, s Differs from controls by Student's t test, P < 0.01. was found in the various groups with the older animals inconsistently. Nor was there following frequencies: zirconium 7, nio good analytical evidence that zirconium bium 5, antimony 5, lead 9, vanadium 9 was consistently absorbed by -rats. A logi and controls 12. It was somewhat more cal explanation was suggested by Blumen- prevalent in females than in males (9.796 thal.7 Zirconium sulfate in water solution and 6.0%, respectively). dates to hydrous zirconia, ZrCVHaO, in a DISCUSSION long chain, just as do chromic salts; this complex is insoluble and probably non In studies such as these we are attempt absorbable. Animals do absorb zirconium ing to evaluate some of the trace dements found in the human environment, human food and human tissues which have no known physiological roles and thus may be considered "abnormal," pending fur ther investigation. We may obtain clues from food where it probably exists as a soluble chelate; it is probably broken down to insoluble and inert hydrous zirconia in tissues, in particles with diameters of 20 to 50 A. to biological activity, favorable or adverse, Niobium as niobate was probably ab by examining growth, survival, longevity, sorbed and excreted readily in the urine, as tumorigenesis and certain serum constitu occurs in man (9). Its effect on body ents; microscopic sections may reveal path weight of male rats is unexplained. In ological changes. terms of longevity, it appeared to be toxic The present study and the previous one also to males. on mice (7) reveal no evidence that zir Antimony was more toxic to rats than conium as fed has any biological activity, to mice (7) in terms of survival and Ion- except possibly to affect body weight of s' Blumenthai, F. N,, personal communication. DUP050312635 TRACE METALS IN RATS 67 gevity. This element was tolerable for (13); in our chronic ones depressant ef growth and its innate toxicity did not ap fects on circulating cholesterol were con pear until later life, indicating that growth fined to female rats. Vanadium pentoxide is not always a valid index of the adverse in drinking water was highly toxic to rats effects of an element. There appeared to at 49 (lg/milliliter (14): the valence state be disturbances in glucose and cholesterol of this metal may influence its toxicity. metabolism associated with its ingestion, Effects of antimony potassium tartrate but no signs of injury to the heart, as have been extensively studied (10); in rats might be expected with larger doses (10). oral doses up to 100 mg/kilogram body No toxicity was demonstrated by tetravalent vanadium, according to any of the measurements made in this study. Growth, body weights and survival were almost identical to or slightly above the controls. In these chromium-supplemented rats, lead was not toxic to males in terms of~ growth and survival. Older rats, nowever. appeared to have lost weight; their coats showed considerable loss of hair and they were sluggish in their movements. At 2 years of age, they weighed 421 11.7 g (40 g less than at 1.5 years), whereas controls weighed 508 13.1 g (P < 0.005); at 30 months of age their weight was 378 32.0 g compared to the weight of the controls 444 15.8 g (P < 0.05). At 3 years of age mean weights did not differ significantly. A similar difference in weight was observed in the former series of chro mium-deficient lead-fed male rats (2) at 2 weight did not influence growth but con sistently injured the heart (15). Accord ing to our experiences, doses calculated at 350 ug/kilogram were innately toxic; larger ones have shortened life spans (15). To compare innate toxicides of these five elements with others up to half life spans, median ages at death of the six groups previously reported (2, 6) and the nine included here and to be reported later are shown in table 9. The amounts of chro mium supplemented in the water are also indicated. In both sexes, the elements showing this early type of toxicity (early compared to the whole lives of the ani mals) were selenite, as reported (16); lead, reported in the first series (2); germanium (6); antimony and cadmium (2). Similar toxicities were observed in mice. No tox icity relative to the controls occurred with chromium, selenate, niobium, the second years of age (P < 0.005). There was a re lead series (the present paper), zirconium, markable resistance to accumulation of Tead in the sott tissue_or-thesp rats given- 25 ppm in water; although bone was not analyzed, this tissue probably stored lead, as 61% of the lead in the human body is in bone. Whether or not the first series of lead-fed rats can be compared with the vanadium, tellurite, arsenite, or nickel. Tin was toxic to female rats and mice, but not to males. These data concern oral ingestion of the elements, and do not apply to ele ments absorbed into (he lungs from pol luted air or injected parenterally. Median ages of male control rats in the second done 5 years later is problematical; however, the diet and regimen were iden tical, the rats were of the same type, the environment had not altered and controls for both series had similar life spans and three series differed by 106 and 18 days, and in the females by 33 and 34 days (table 9). The rats for the first series and the first lead, chromium and cadmium groups were obtained from a different sup no early deaths. If the profound differences _ plier 8; the remainder came from the pres in mortality between the two serjes-caa-be- ent one.8 Possibly males of the first strain accepted, it appears that chromium pm-- were naturally long lived. tects agamsT~fiinate lead toxicity. If any of the five trace elements in this According to Browning ( iu), zirconium study are essential for rats and mice, they has a very low order of toxicity, whereas are required at concentrations less than niobium is chronically toxic in doses more those present in our diet. There is no evi than 100 times those given here (11), in dence, however, that antimony and lead hibiting hepatic succinic dehydrogenase are essential elements. Unfortunately, rel-* 9 (12). In acute experiments vanadyl ions suppressed hepatic cholesterol synthesis a Rockland Farms, New City, N. Y. 9 See footnote 4. DUP050312636 68 HENRY A. SCHROEDER, MARIAN MITCHENER AND ALEXIS F. NASON TABLE 9 Median ages of rats fed various trace elements 1 Element Control I * Selenate Chromium Niobium Lead H 2 Zirconium Tin Control IX 2 Vanadium Control III2 Tellurite Arsenite Cadmium Nickel Antimony Germanium Lead I2 Selenite Total Chromium in water ppm 0 5 5 1 1 1 1 1 1 S 5 1 0 5 1 1 0 5 Male No. Median rats age days 52 978 49 964 54 922 48 892 52 883 56 881 55 876 53 872 52 860 55 854 52 844 s 53 825 s 69 822 s 52 822 54 766 55 738 s 62 729 s 48 60 s 971 Female No. Median rats age days 52 945 55 1002 54 950 50 998 s ---- 57 947 56 330 s 80 912 61 961 46 878 52 894 s 55 912 s 58 805 51 928 61 805 52 833 60 727 53 342 s 953 i All elements were given in water at 5 ppm, except for selenite and selenate at 3 ppm, tellurite at 2 ppm, and lead at 25 ppm. __ , . 8 Meaning of Roman numerals! I refers to (2), II to the present paper, in to results yet to be published. e Toxic to mice in terms of survival and longevity. atively large amounts of zirconium, nio bium and vanadium occurred naturally in tbe diet, the last two largely in com oil; therefore, we have no evidence for or against essentiality of these three elements. LITERATURE CITED 1. Schroeder, H. A., W. H. Vinton, Jr. and J. J. Balassa 1963 Effects of chromium, cad mium and lead on the growth and survival of rats. J. Nutr. SO; 48. 2. Schroeder, H. A., J. J. Balassa and W. H. Vinton, Jr. 1965 Chromium, cadmium and lead in rats: Effects on life span, tumors and tissue levels. J. Nutr. 86: 51. 3. Mertz, W., E. E. Roginski and H. A. Schroeder 1965 Some aspects of glucose metabolism of chromium-deficient rats raised in a strict ly controlled environment. J. Nutr. 86:107. 4. Schroeder, H. A. 1966 Chromium defi ciency in rats: A syndrome simulating dia betes mellitus with retarded growth. J. Nutr. 88: 439. 5. 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