Effects of Various Aluminium Compounds Given Orally to Mice on Al Tissue Distribution and Tissue Concentrations of Essential Elements

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1 C Pharmacology & Toxicology 2000, 86, Printed in Denmark. All rights reserved Copyright C ISSN Effects of Various Aluminium Compounds Given Orally to Mice on Al Tissue Distribution and Tissue Concentrations of Essential Elements Maria Długaszek 1, Maria A. Fiejka 2, Alfreda Graczyk 1, Janina Cz. Aleksandrowicz 2 and Maria Słowikowska 2 1 Institute of Optoelectronics, Military University of Technology, Kaliskiego 2, Warsaw, and 2 Department of Sera & Vaccines Evaluation, National Institute of Hygiene, Chocimska 24, Warsaw, Poland (Received May 4, 1999; Accepted November 2, 1999) Abstract: To evaluate the risk of gastrointestinal long-term aluminium (Al) exposure, aluminium distribution and the levels of the following essential elements: Ca, Mg, Zn, Cu, and Fe in tissue were studied. Aluminium was administered in drinking water as aluminium chloride, dihydroxyaluminium sodium carbonate or aluminium hydroxide. Mice (strain Pzh:SFIS) were exposed to a total dose of 700 mg Al in long-term treatment (for each Al compound nω15). Concentrations of Al, Ca, Mg, Zn, Cu, and Fe in stomach, kidneys, bone and liver were analyzed by atomic absorption spectrometry. After AlCl 3 treatment, aluminium was found to accumulate in all tested tissues. A significant decrease in Fe concentration in liver and Zn in kidneys was observed in comparison to concentrations of these elements in the control group. In the Al(OH) 3 -treated group, accumulation of aluminium was observed in bone only and decline of Fe concentration in stomach and Cu in liver and kidney. In the NaAl(OH) 2 CO 3 -treated group the increase in Al concentration was significant in bone; there was no change in concentration of essential elements in the examined tissues. The observed aluminium accumulation was not accompanied by changes in Ca and Mg concentration except for bone. This study showed that oral administration as a route of Al exposure can result in diverging accumulation of aluminium in tissues, the concentration depending on the chemical form. Aluminium is a chemically active element. As a rule, Al produces stable compounds in biological systems, more stable than Ca and Mg. The small ionic radius of Al, close in size to the ionic radii of Fe and Mg, makes it possible to substitute these elements in biologically active compounds. Aluminium combines with many compounds, important with respect to their biological function, e.g. proteins (transferrin, chromatin, proteins G, calmodulin, enzymes), ATP, GTP, DNA, RNA, phosphates, fluorides, thus affecting their biological functions (Trapp 1986; Macdonald & Martin 1988). Aluminium induces disturbances of function of the nervous, osseous and erythropoietic systems. The syndrome of dialysis encephalopathy, dialysis osteomalacia and microcytic anaemia has been reported in patients dialysed due to renal insufficiency. Observation of the same symptoms in patients not dialysed but treated with aluminium containing drugs pointed to verification supported explanations of mechanisms of absorption of Al from orally administered drugs. Aluminium is absorbed from the alimentary tract in the small intestine and, because of favourable ph, in the stomach and duodenum, too. Aluminium intestinal absorption is determined by a number of factors, for instance solubility of Al compound, the presence in the alimentary tract of organic acids (citric, ascorbic), diet deficient in Fe and Ca, Author for correspondence: Maria Długaszek, Military University of Technology, Institute of Optoelectronics, Kaliskiego 2, Warsaw, Poland (fax π ). renal insufficiency, immaturity of the alimentary tract, gastric hyperacidity, severe systemic diseases (Ittel 1993; Powell & Thompson 1993). Balance studies have shown that 125 mg Al can be deposited in the human organism. Patients treated with antacid drugs, which contain Al(OH) 3 or NaAl(OH) 2 CO 3 as the principal agent, on an average receive a daily dose of 1 g Al, which, may be further increased toeven5galperday(greger & Baier 1983; Wilhelm et al. 1990). Such drugs uptill now considered to have no side effects, are commonly applied, about 30% of the population use antacids. Although Al absorption in the alimentary tract is estimated to be below 1%, there are reports on aluminium accumulation in the tissues of patients treated with medicines containing Al compounds (Joff et al. 1989; Umbreit 1993; Jeffrey et al. 1996). The accumulation is followed by a number of disturbances of the biochemical function of the cell, thus leading to gradual degeneration of neurones, osteomalacia and bone fragility, and changes in the erythropoietic system. In the present study we have attempted to evaluate Al accumulation in tissues of mice after long-term oral exposure to aluminium chloride, dihydroxyaluminium sodium carbonate or aluminium hydroxide, and Al effect on the level of essential elements. Materials and Methods Chemicals. Aluminium hydroxide (3%), was obtained from the Serum and Vaccine Production Plant (Cracov, Poland). Aluminium chloride (AlCl 3 ) was purchased from Matthey Chemicals

2 136 MARIA DŁUGASZEK ET AL. Table 1. Al, Ca, Mg, Zn, Cu, Fe concentrations in stomach tissue of mice (mg/g wet weight), exposed orally to 700 mg Al in long-term treatment. Values represent X S.D. (nω15). To examine the significance of the differences between the animal groups (experimental and control), as concerns a specific characteristics, Student s t-test was used. The significance of the differences was verified at PΩ0.05. Al(OH) Control group NaAl(OH) 2 CO Control group AlCl P Control group Limited. Alugastrin (NaAl(OH) 2 CO 3 ) was produced by Polfa, Poland. Animal treatment. Young female mice (21 days of age, initial body weight 16 g strain Pzh:SFIS) were divided into three experimental groups of 15 animals each. The first group was given Al(OH) 3 suspended in drinking water at doses of 0.5 mg Al/ml. The second and the third group of animals was given NaAl(OH) 2 CO 3 and AlCl 3, respectively, also dissolved in drinking water at doses of 0.5 mg Al/ ml. Each experimental group of animals had its own control group of 5 animals each. Control animals received tap water ad libitum. Mice were fed commercial mouse chow. The diet was analysed to contain average amounts of Al 64.5 mg/g in chow and 14.5 ng/ml in tap water. The daily dose of Al estimated from water and diet was mg in the control groups. Fluid, food consumption and body weight were recorded once a week. The treatment was continued until the total dose of Al given to each experimental group of mice reached the value of 700 mg Al; it lasted 159 days for Al(OH) 3, for NaAl(OH) 2 CO days, and for AlCl days. The animals were killed by cervical dislocation 48 hr after the last treatment. Tissues (whole stomach, liver, bone tibia, both kidneys) from treated and control mice were weighed and stored in aluminium free plastic containers at 18æ for analysis of Al and essential elements. Analyses of Al and of essential elements. Aluminium was measured both in tissue and chow samples by electrothermal atomic absorption spectrometry GFAAS (Perkin Elmer Model 2100 atomic absorption spectrophotometer and HGA-700, graphite furnace with the AS-70 autosampler). The details of the procedure used have been previously reported by Fiejka et al. (1996). The analysis of Al in diet and water was performed by the same method as described previously (Fiejka et al. 1996). For verification of the alu- minium determination method, the NBS Standard Reference Material 1577a was used. The samples from all tissues and mixed chow were wet-ashed in a mixture of perchloric acid and nitric acid at boiling temperature. In the drinking water samples, the Al concentration was determined directly by electrothermal atomic absorption spectrometry. All steps during the sample preparation were carried out under dust-free conditions. Determination of Ca, Mg, Zn, Fe, and Cu was performed by flame atomic absorption spectrometry under standard conditions (The Perkin-Elmer Corporation 1982). Statistical analysis. The concentrations of elements were expressed as arithmetic mean S.D. Student s t-test was used for analysis of the experimental data. The significance of the differences was verified at PΩ0.05. Age related differences in Al levels in tissues of control animals (c. Al(OH) 3 experiment time 159 days versus c. AlCl 3 - experiment time 239 days) were tested by one way ANOVA. Results Accumulation of aluminium in tissues of experimental animals after long term oral administration of Al(OH) 3, NaAl (OH) 2 CO 3 or AlCl 3 is presented in tables 1 4. The time needed for consumption of 700 mg Al resulted from varied daily intake (mice of control group drank 8.9 ml water, mice of treated groups accordingly drank Al(OH) ml, NaAl(OH) 2 CO ml, and AlCl ml). Table 2. Al, Ca, Mg, Zn, Cu, Fe concentrations in kidney tissue of mice (mg/g wet weight), exposed orally to 700 mg Al in long-term treatment. Values represent X S.D. (nω15). To examine the significance of the differences between the animal groups (experimental and control), as concerns a specific characteristics, Student s t-test was used. The significance of the differences was verified at PΩ0.05. Al(OH) P Control group NaAl(OH) 2 CO Control group AlCl Control group

3 BIOAVAILABILITY AND DISTRIBUTION OF ALUMINIUM COMPOUNDS 137 Table 3. Al, Zn, Cu, Fe (mg/g wet weight), Ca and Mg (mg/g wet weight) concentrations in tibia of mice exposed orally to 700 mg Al in long-term treatment. Values represent X S.D. (nω15). To examine the significance of the differences between the animal groups (experimental and control) as concerns specific characteristics, Student s t-test was used. The significance of the differences was verified at PΩ0.05. mg/g mg/g mg/g mg/g mg/g mg/g Al(OH) Control group NaAl(OH) 2 CO Control group AlCl Control group Stomach Al concentration in the three control groups was within the range mg/g. Significantly high Al accumulation in the stomach was noted in the group of NaAl(OH) 2 CO 3 -treated animals (range mg/g) and in the AlCl 3 group (range mg/g). In comparison to this experimental group the concentration of Al in the stomachs of mice after Al(OH) 3 treatment was twice lower (range mg/g) and not significantly different from control mice. Aluminium concentration in kidneys of Al(OH) 3 -treated mice (range mg/g) and in NaAl(OH) 2 CO 3 -treated animals (range mg/g) were generally comparable to the respective control groups of animals (range mg/g). Significant Al accumulation in kidneys was found only in the group of mice after AlCl 3 treatment (range , in comparison to the range mg/g of control group). Aluminium accumulation in bone was found in two experimental groups; mean Al concentration was significantly elevated in Al(OH) 3 -(range mg/g) and AlCl 3 - treated mice (range mg/g), as compared to those in the control groups ( ; mg/g). A significant increase in Al concentration in liver was noted only in AlCl 3 -treated mice (range mg/g), when compared to the respective control (range mg/g). Unexpected findings were some changes in the concentrations of Al in tested tissues of control mice in regard to the age of animals. A tendency to decreased Al level in tested tissues of control groups was observed in relation to the length of life of the animals. In control mice stomach (AlCl 3 ; time of experiment 239 days) the concentration of Al was by 48% lower than in other control mice (Al(OH) 3 ; time of experiment 159 days, ). The decline of Al concentration in these control groups was also noted in kidneys, 58% (), in bone 42% (), and in liver 71% (). Concentrations of Ca, Mg, Zn, Cu, and Fe in tissues of mice exposed to Al(OH) 3, NaAl(OH) 2 CO 3 or AlCl 3 are presented in table 1 4. Only in the animal group exposed to Al(OH) 3 there was an increase in Mg concentration in bones, and a decrease in Fe concentration in stomach and Cu in kidneys and liver. Animal exposed to AlCl 3 showed a decline of Fe concentration in the liver and of Zn in the kidneys. As in the case of aluminium, the organs of control mice showed an age-related tendency to declined concentration of some essential elements. In the stomach the content of Ca, Mg, and Fe declined by 23%, 15%, 10% respectively. In kidneys, the Zn concentration decreased by 10% and Cu, Fe by 18%, 6% respectively, but the Ca content increased by 52%. In bones, the level of Ca and Zn concentration Table 4. Al, Ca, Mg, Zn, Cu, Fe concentrations in liver tissue of mice (mg/g wet weight), exposed orally to 700 mg Al in long-term treatment. Values represent X S.D. (nω15). To examine the significance of the differences between the animals groups (experimental and control), as concerns specific characteristics, Student s t-test was used. The significance of the differences was verified at PΩ0.05 Al(OH) Control group NaAl(OH) 2 CO Control group AlCl 3 0, Control group P 0.001

4 138 MARIA DŁUGASZEK ET AL. decreased by 6% and 12% respectively. The liver showed a decrease in Zn by 41% and Cu by 28% while Ca and Fe concentration increased by 15% and 20% respectively. Discussion Aluminium absorption via food and medications from the gastrointestinal tract is one of the systemic routes of human exposure to aluminium. Although aluminium is poorly absorbed from the gastrointestinal tract in persons with normal renal function, further experimental studies are still necessary to evaluate aluminium distribution at the organ and tissue level and its potential influence on mineral metabolism. In this study, aluminium accumulation was evaluated in the tissues of mice following long-term exposure to aluminium, administered as chloride, dihydroxyaluminium sodium carbonate, and hydroxide, as well as aluminium effects on the level of bioelements in tissues. In our animal model 700 mg of Al is comparable to the amounts which may be taken up by patients treated with aluminium containing drugs. The chemical form of aluminium is one of the essential factors determining its bioavailability in the digestive tract. Therefore, compounds, including aluminium hydroxide and carbonate, which are principal constituents of drugs commonly applied for gastric hyperacidity were selected with regard to solubility. The degree of aluminium absorption in the digestive tract may also be influenced by dietary ligands and minerals, and by factors such as age of the animal, as well as age-related changing features such as body weight, function of the digestive tract and excretion, the dynamics of metabolic processes and sensitivity to Al toxicity. Aluminium accumulation in tissues is also genetically conditioned (Fosmire et al. 1993; Greger & Radzanowski 1995). Our experimental model showed the highest elevation of aluminium in the tissues tested after exposure to AlCl 3, lower values being detected after the administration of NaAl(OH) 2 CO 3 and Al(OH) 3 except for bone. Exposure to AlCl 3 induced an increase in aluminium concentration in all tissues selected for testing, 3.5 times in stomach, and comparable in liver (3.6 times), in relation to controls. Lower increase was found in bone tissue (1.7 times) and kidneys (twice). Administration of NaAl(OH) 2 CO 3 induced aluminium accumulation in the stomach (concentration doubled) and in the liver (increased 1.6 times), while slight increase only was seen in bone tissue (statistically insignificant). The lowest effects were seen after Al(OH) 3 administration, which was followed by only 1.5 times increase in aluminium concentration in the bone tissue. Depending on the exposure used, aluminium concentration in the bone tissue was higher after all the administered chemical forms, though in the case of NaAl(OH) 2 CO 3, the increase was statistically insignificant. After long term oral administration, aluminium accumulation in bone and liver of mice was comparable with findings in other experimental models (Fosmire et al. 1993; Oteiza et al. 1993). However, much higher aluminium concentrations than in these experimental models were observed after exposure to aluminium lactate (Anghileri et al. 1994), which might be associated with higher bioavailability of aluminium from lactate than from hydroxide or chloride (Yokel & McNamara 1988). In our experimental model, age-related differences in aluminium concentration in the tissues of control groups were observed. As the animals grew older, the aluminium level declined in the stomach from 2.97 to 1.54 mg/g (48%), in the kidneys from 0.43 to 0.18 mg/g (58%), in the bones from 2.77 to 1.61 mg/g (42%), and in the liver from 0.24 to 0.07 mg/g (71%). The dynamics of the changes were most intense in the liver. Similar age-related observations of changes in aluminium concentration in animal tissues have been reported (Fosmire et al. 1993; Golub et al. 1993; Anghileri et al. 1994; Greger & Radzanowski 1995; Domingo et al. 1996). These findings confirm that a high rate of metabolic processes in young organisms and lack of effective protection mechanisms are favourable for accumulation of toxic elements, including aluminium. Another aim of this study was to examine the impact of dietary aluminium on mineral metabolism in various tissues of mice. The interaction of aluminium with essential elements was previously reported. The concentration of Al and Ca in the tissues of mice treated orally with Al was determined by Anghileri et al. (1994). Calcium concentrations in the brains of Al-treated animals were higher than in controls but the differences were not statistically significant. The highest concentration of 45 Ca 2π ions was observed in the liver (Anghileri 1992). Muller et al. (1993) observed a significant rise of Ca in serum and spleen of adult rats, as well as in newborns whose mothers received oral aluminium lactate. This is in line with data by Severson et al. (1992), who reported much lower levels of Ca in the osseous tissue of aluminium-intoxicated animals. Aluminium increase in spleen and kidneys of animals given water with low Ca contents was observed by Wills et al. (1993). Calcium-deficient diet causes increase in Al contents in serum and bones of rats, not given this element, probably due to bones of such animals being more fragile (Konishi et al. 1996). Brown & Schwartz (1992) assumed that aluminium binds with ligands at the same sites as Fe 3π, and they observed significant Al increase in the liver and spleen of rats fed iron-deficient diet. Golub et al. (1996) did not to observe the effect of dietary aluminium excess (doses about 100 mg) on Fe level in the brain and liver, nor did Oteiza et al. (1993) observe any change in Fe concentration in the cerebrospinal fluid, brain, liver and bones, in mice given about 300 mg Al. After administration to mice of 900 mg Al, Golub et al. (1995) found lower Fe levels in the brain and cerebrospinal fluid. Konishi et al. (1996), in addition to aluminium also detected Fe in the frontal region of bone mineralization. A negative effect of aluminium on Mg levels in serum, bones, and kidneys was observed in sheep after oral exposure. Lower Mg concentration in bones is probably not

5 BIOAVAILABILITY AND DISTRIBUTION OF ALUMINIUM COMPOUNDS 139 only a result of its lower concentration in blood, but also points to disturbances of Mg metabolism (Allen 1985; Muller et al. 1993). It was also noted that Mg level is lower in carcasses of newborn offspring of rats fed aluminium compounds, as these rats also showed lower Mg level in blood (Muller et al. 1993). No significant differences in zinc concentration in the brain, cerebrospinal fluid, liver, and bones were found by Oteiza et al. (1993) in mice that received aluminium with the diet. A decline in the contents of this element was observed by Muller et al. (1993) in the kidneys of rats after administration of aluminium compounds. In complex studies of long-term impact of Al (as Al(NO 3 ) 3 in citrate solution) on accumulation of elements (Ca, Mg, Zn, Cu, Fe, and Mn) in tissues (liver, bones, testicles, spleen, kidneys, brain) Sanchez et al. (1997), observed interactions between Al and the elements tested. In some cases the changes were dose- and age-related. In general, no significant differences were observed in the concentration of elements in our study. The lack of significant differences between the contents of mainly Ca and Mg is probably a result of the different concentrations of these elements and aluminium in the tissues tested, the organism might have at disposal adequate homeostatic properties, which permit under conditions of the aluminium stress of the experiment to preserve status of the elements in analyzed tissues. To conclude, aluminium is absorbed from the digestive tract of animals administered orally as a chloride, hydroxide or carbonate as demonstrated by higher contents in all tissues (stomach, kidneys, bones, liver). The degree of accumulation depends, however, on the tissue type and the chemical form of aluminium, as aluminium is assimilated from compounds contained in antacid drugs. References Allen, V. G.: Influence of aluminum on magnesium metabolism, magnesium in cellular processes and medicine. 4th Int. Symp. on Magnesium, Blacksburg 1985, Analytical Methods for Atomic Absorption Spectrophotometry. The Perkin Elmer Corporation, Norwalk, Connecticut, USA, Anghileri, L. J.: Effects of complexed iron and aluminum on brain calcium. NeuroToxicology 1992, 13, Anghileri, L. J., P. Maincent & P. Thouvenot: Long-term oral administration of aluminum in mice. Aluminum distribution in tissues and effects on calcium metabolism. Ann. Clin. Lab. Scie. 1994, 24, Brown, T. S. & R. Schwartz: Aluminum accumulation in serum, liver and spleen of Fe-depleted and Fe-adequate rats. Biol. Trace Elem. Res. 1992, 34, Domingo, J. L., J. Liorens, D. J. Sanchez, M. Gomez, J. M. Liobet & J. Corbella: Age-related effects of aluminum ingestion on brain aluminum accumulation and behaviour in rats. Life Sci. 1996, 58, Fiejka, M., E. Fiejka & M. Długaszek: Effect of aluminium hydroxide administration on normal mice: tissue distribution and ultrastructual localization of aluminium in liver. Pharmacology & Toxicology 1996,78, Fosmire, G. J., S. J. Focht & G. E. McCleran: Genetics influences on deposition of aluminium in mice. Biol. Trace Res. 1993, 37, Golub, M. S., B. Han & C. L. Keen: Aluminum alters iron and manganese uptake and regulation of surface transferrin receptors in primary rat oligodendrocyte cultures. Brain Res. 1996, 719, Golub, M. S., B. Han, C. L. Keen & M. E. Gershwin: Developmental patterns of aluminum excess on manganese deficiency. Toxicol. 1993, 81, Golub, M. S., B. Han, C. L. Keen, M. E. Gershwin & R. P. Tarara: Behavioural performance of Swiss Webster mice exposed to excess dietary aluminum during development and as adults. Toxicol. Appl. Pharmacol. 1995, 133, Greger, J. L. & M. J. Baier: Excretion and retention of low moderate levels of aluminium by humans subject. Fd. Chem. Toxic. 1983, 21, Greger, J. L. & G. M. Radzanowski: Tissue aluminium distribution in growing, mature and ageing rats: relationship to changes in gut, kidney and bone metabolism. Fd. Chem. Toxic. 1995, 33, Ittel, T. H: Determinants of gastrointestinal absorption and distribution of aluminium in health and uremia. Nephrol. Dial. Transplant. 1993, Suppl., 1, Jeffrey, E. H., K. Abreo, E. Burgess, J. Cannata & J. L. Greger: Systemic aluminum toxicity: effects on bone, hematopoietic tissue, and kidney. J. Toxicol. Environ. Health 1996, 48, Joff, P., E. Olsan, J. G. Heaf, B. Gammelgaard & J. Podenphant: Aluminum concentrations in serum, dialysate, urine, and bone among patients undergoing continuous ambulatory peritoneal dialysis (CAPD). Clin. Nephrol. 1989, 32, Konishi, Y., K. Yagyu, H. Kinebuchi, N. Saito, T. Yamaguchi & Y. Ohtsuki: Chronic effect of aluminium ingestion on bone in calcium deficient rats. Pharmacology & Toxicology 1996, 78, Macdonald, T. L. & B. Martin: Aluminum ion in biological systems. TIBS 1988, 13, Muller, G., D. Burnel, A. Gery & P. R. Lehr: Elements variations in pregnant and non pregnant female rats orally intoxicated by aluminum lactate. Biol. Trace Elem. Res. 1993, 39, Oteiza, P., C. L. Keen, B. Han & M. S. Golub: Aluminum accumulation and neurotoxicity in Swiss-Webster mice after long-term dietary exposure to aluminum and citrate. Metabolism 1993, 42, Powell, J. J. & R. P. H. Thompson: The chemistry of aluminium in the gastrointestinal lumen and its uptake and absorption. Proc. Nutr. Soc. 1993, 52, Sanchez, D. J., M. Gomez, J. M. Llobet, J. Corbella & J. L. Domingo: Effects of aluminium on the mineral metabolism of rats in relation to age. Pharmacology & Toxicology 1997, 80, Severson, A. R., C. F. Haunt, C. E. Friling & T. E. Huntley: Influence of short-term aluminum exposure on demineralized bone matrix induced bone formation. Arch. Toxicol. 1992, 66, Trapp, G. A.: Interaction of aluminum with cofactors, enzymes, and other proteins. Kidney Int., 1986, Suppl., 29, Umbreit, M. H.: Nowe spojrzenie na właściwości biologiczne jonów glinu. Farmacja Polska 1993, 49, 1 5. Wilhelm, M., D. E. Jager & F. K. Ohnesorge: Aluminium toxicokinetics. Pharmacology & Toxicology 1990, 66, Wills, M. R., C. D. Hewitt, J. Savory & M. M. Herman: Long-term oral aluminum administration in rabbits. II. Brain and other organs. Ann. Clin. Lab. Sci. 1993, 23, Yokel, R. A. & P. J. McNamara: Influence of renal impairment, chemical form, and serum protein binding on intravenous and oral aluminum kinetics in rabbit. Toxicol. Appl. Pharmacol. 1988, 95,