Biol Trace Elem Res (2014) 161:32–37 DOI 10.1007/s12011-014-0061-1
The Study of Barium Concentration in Deciduous Teeth, Impacted Teeth, and Facial Bones of Polish Residents Agnieszka Fischer & Piotr Malara & Danuta Wiechuła
Received: 14 May 2014 / Accepted: 29 June 2014 / Published online: 1 August 2014 # Springer Science+Business Media New York 2014
Abstract The study determines the concentration of Ba in mineralized tissues of deciduous teeth, permanent impacted teeth, and facial bones. The study covers the population of children and adults (aged 6–78) living in an industrial area of Poland. Teeth were analyzed in whole, with no division into dentine and enamel. Facial bones and teeth were subjected to the following preparation: washing, drying, grinding in a porcelain mortar, sample weighing (about 0.2 g), and microwave mineralization with spectrally pure nitric acid. The aim of the study was to determinate the concentration of Ba in deciduous teeth, impacted permanent teeth, and facial bones. The concentration of barium in samples was determined over the ICP OES method. The Ba concentration in the tested bone tissues amounted to 2.2-15.5 μg/g (6.6 μg/g± 3.9). The highest concentration of Ba was present in deciduous teeth (10.5 μg/g), followed by facial bones (5.2 μg/g), and impacted teeth (4.3 μg/g) (ANOVA Kruskal-Wallis rank test, p=0.0002). In bone tissue and impacted teeth, Ba concentration increased with age. In deciduous teeth, the level of Ba decreased with children’s age.
Keywords Barium . Deciduous teeth . Impacted teeth . Facial bones
A. Fischer (*) : D. Wiechuła Department of Toxicology, School of Pharmacy with the Division of Laboratory Medicine, Medical University of Silesia, Katowice, Poland e-mail: [email protected] P. Malara Institute of Engineering Materials and Biomaterials, Silesian University of Technology, Gliwice, Poland
Introduction Barium (Ba) is a common element in the environment. Its share in lithosphere, mostly in the form of barite (BaSO4), amounts to approximately 0.04 %. In Poland, barite motherlodes occur in the southern part of the country (Sudeten, the Świętokrzyskie Mountains, Tatra Mountains) [1]. Barium can be released into the environment during geological processes, but its main source in the environment is the anthropogenic activity. Barium release accompanies mining operations and coal and fuel combustion processes. Its use is very broad and diverse (e.g., chemical, plastic, metallurgical, pharmaceutical, cosmetic, and medical industries). Barium present in the soil is relatively stable; however, acidification of the substrate leads to a formation of watersoluble compounds. The content of Ba in drinking water is highly variable and depends, among other things, on its hardness. Higher contents of Ba occur in acidic water when compared with alkaline [2, 3]. Plants absorb Ba directly from the environment, whereas animals absorb it from food. The concentration of barium in tissues of living organisms is diverse and depends on a position in the trophic chain [3]. The human body absorbs it mainly by ingestion and, in a negligible quantity, by inhalation. The average concentration in food does not exceed 0.1 mg Ba/kg [3]. A daily dose of Ba supplied with food to the human body amounts to 0.3–1.8 mg, and its donor is mainly milk and dairy products (25 % of dose) [3, 4]. Ba absorption from food is also subject to individual variability as it is greater among younger people [3]. During pregnancy, Ba transport from mother to fetus is limited by the placenta, which is indicated by lower concentrations of Ba in the umbilical cord sera [5, 6]. The literature studies also showed that the Ba concentration in mothers’ milk correlates with its concentration in hard tissues of deciduous teeth [6]. In addition, a clear increase in the quotient of Ba/Ca
Barium Concentration in Teeth and Facial Bones
is observed after cessation of breast-feeding and introduction of modified milk into the diet of infants [6]. The physiological role of barium in the body remains unexplained. It is known that this element is not involved in metabolism and does not participate in the process of maintaining homeostasis [7]. Barium is present in the body in various tissues (lung, fat tissue, skin, eye tissue), but 91 % of the concentration of this element is accumulated by the bone tissue [3, 4]. The main component of the mineralized tissues is hydroxyapatite, which has the ability to exchange elements. The inclusion of Ba in the hydroxyapatite structure occurs at the expense of the concentration of physiologically essential elements, including calcium [8, 9]. Literature research indicates a possibility of making an elemental analysis of teeth taking into account the type of calcified tissues and the mineralization period [6, 10–12]. Teeth being formed in utero, record birth as the neonatal line, define the pre- and postnatal developmental periods [6, 10, 11]. Calcified tissues accumulated different concentration of metals. Differences in the concentration of elements in enamel and dentine are observed. For example, the levels of Pb and Mn were higher in the dentine than in enamel [10, 11]. The highest concentration of Pb was found in postnatal dentine [10]. Barium is an element of which occurrence in the environment is connected mainly with anthropogenic activity. In an organism, it accumulates itself in bone tissue. In the available literature, there are no current research results on barium content in teeth of inhabitants of Poland. The objective of the study was to determine the concentration of Ba in teeth and bones of people living in the industrial areas of southern Poland (Silesian Region). The Silesian Region is one of the most industrialized and urbanized regions in Poland, characterized by high traffic and intensive emissions. In this area, there are acidic soils (pH 4.0–6.7), containing 100– 400 mg Ba/kg [13]. Research, including the authors’ own research, onto the determination of concentration of other elements (e.g., Cu, Cr, Pb, Ca, Fe) in teeth was conducted among inhabitants of the Silesian agglomeration. The obtained results indicate a higher metal concentration in teeth of inhabitants of the Silesian Region than in other regions of Poland [14–16]. In the study, the levels of Ba in three kinds of calcified tissues (deciduous teeth, permanent impacted teeth, and facial bones) were determined. The teeth were analyzed as a whole without making a histological division into calcified tissues. The differences in the concentration of Ba in the tissues covered by the study were analyzed depending on gender and age.
Materials and Methods The subjects of the research were samples of mineralized tissues from persons residing in the south of Poland. People
33
included in the research lived in the towns of the province of Silesia (Katowice, Tychy, Chorzow, Sosnowiec, BielskoBiala) since birth. The study tissues (n=35) included samples of facial bones (n=14), permanent impacted teeth (n=10), and deciduous teeth (n=11). All collected tissue samples came from healthy individuals that did not indicate a presence of chronic diseases of the body requiring treatment in the interview. The study covered healthy teeth that did not have signs of decay and fillings. The weight of the bone tissue given to research was at least 0.15 g. The research did not include tissues samples from people suffering from chronic diseases requiring pharmacological treatment. The research did not include teeth with decay and fillings. Among the mineralized tissues of the body, the teeth are a relatively easily accessible tissue. They can be collected at the time of medically reasoned tooth extractions and as the postoperative residual forwarded for further research. Deciduous teeth can be taken for testing over a completely noninvasive method to the patient, during physiological replacement of teeth. The deciduous teeth were collected over a noninvasive method during a physiological replacement. Permanent teeth were removed during a planned medical, surgical extraction of teeth. During the procedure of a surgical extraction, especially for impacted teeth, the formation of bone chips is a common complication. During the procedure, bone chips are removed to accelerate the process of healing of postextraction wounds. The investigated teeth and bone chips were postoperative residue. Studied deciduous teeth came from children (n=11, aged 6–12 years, mean 8.5±2.2) and were collected during natural replacement of teeth to permanent ones. Permanent teeth (n=10) came from people aged 16-41 (mean 29.0±10.5) and were removed during a planned medical, surgical extraction of tooth. The teeth were completely impacted, remaining in the upper or lower jaw bone. Completely impacted teeth are fully formed teeth, surrounded by bone tissue from all sides. Molar teeth were assigned for the research. Of all the types of teeth (incisors, canines, premolars, molars), third molars are the kind of teeth which is usually subject to irregularities in the process of eruption. During surgical tooth extraction procedures, the postoperative residue can also include bone fragments. Mandibular and maxillary bones (n=14) of people aged 16–78 (mean 44.4±21.5) were submitted for the research. All collected tissues samples came from women (n=16, aged 6–77 years, mean 30.9±24.9) and men (n=19, aged 7–78 years, mean 26.3±18.1). Bone tissues obtained for the research were subjected to the following preparation: washing, rinsing (tap water/distilled water), drying to constant weight (85 °C), grinding in a porcelain mortar, drying (85 °C/3 h), sample weighing (about 0.2 g), and mineralization (microwave digestion system
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Fischer et al.
MAGNUM II (prod. ERTEC-Poland), spectrally pure nitric acid; Merck, Germany). The concentration of barium in bone samples was determined over the ICP OES method (Perkin Elmer Optima 5300 DVTM). For calibration, the working solutions were prepared by a precisely measured dilution of standard solutions (Merck, Germany). The verification of determination accuracy was established by means of the standard addition method using the model of mass concentration of given metals. Blank samples were used after every ten samples. The procedure was validated by NIST 1486 (Bone Meal). Statistical Analysis Statistical results were calculated in Statistica 10 for Windows. Nonparametric tests were used to obtain the results of the research (Mann-Whitney U test for two groups and ANOVA Kruskal-Wallis rank for more than two groups). The main criterion for selection of nonparametric tests, apart from the abnormal distribution of results, was the small group size. The correlation analysis was based on Spearman’s nonparametric rank tests. To evaluate the statistical significance of the data, the probability of ≤0.05 was considered significant.
Results Table 1 shows the results of statistical analysis of the concentration of Ba in the studied tissues. The concentration of Ba in mineralized tissue samples amounted to 2.20–15.47 mg/g (arithmetic mean=6.61 μg/g). The highest concentration of Ba was present in deciduous teeth (10.47 μg/g), followed by facial bones (5.21 μg/g), and the lowest was in the impacted permanent teeth (4.33 μg/g) (Table 1). Ba concentration differences within the bone tissue groups were statistically significant (ANOVA Kruskal-Wallis rank test, p=0.0002). The change in the Ba concentration in the studied tissues occurring with age of the subjects is shown in Fig. 1. The graph illustrating the variation of the concentration of Ba in the function Table 1 Concentration of Ba (in μg/g) in deciduous teeth, impacted teeth, and bone tissues Number Mean ± SD GM Deciduous teeth 11 Impacted teeth 10 Bone tissues 14
Median Min. Max.
10.47±3.24 9.96 11.11* 4.33±2.22 3.90 3.33* 5.21±2.06 4.84 4.92*
5.55 2.20 2.50
15.47 9.24 9.29
GM geometric mean, Min. minimum values, Max. maximum values *p=0.0002 (significant differences, ANOVA Kruskal-Wallis rank) *p=0.0002 (significant differences, ANOVA Kruskal-Wallis rank)
Fig. 1 The relationship between the concentration of Ba in deciduous teeth, impacted teeth, and bone tissues and age. Spearman’s rank correlation
of age has proven an increase of the concentration of this element with increasing age in the case of permanent teeth and facial bones. In the case of deciduous teeth, the level of Ba decreased with increasing age of the subjects (Fig. 1). The lowest concentration of Ba in deciduous teeth was observed in the oldest person in the study (5.55 μg/g; age 12 years). For facial bones and permanent teeth, the lowest levels of Ba occurred among persons of 25 years of age. The correlation
Barium Concentration in Teeth and Facial Bones
coefficient specifying the change of the concentration of Ba in all studied tissues with increasing age of the patients was −0.43 and was statistically significant (Spearman’s rank correlation, p=0.01). An analysis of co-occurrence of Ba concentration changes with age of the subjects in different types of mineralized tissues (deciduous teeth, permanent impacted teeth, facial bones) showed that the statistically significant correlation coefficient had defined only the changes in the concentration of Ba and age in the impacted teeth (Spearman’s rank correlation, r=0.64, p=0.05). Studies of Ba concentration changes in mineralized tissues of men and women have shown that higher levels of Ba concentration characterized the samples taken from women (Fig. 2). Higher average levels of Ba in samples from women compared to men concerned all studied bone tissues (deciduous teeth, permanent teeth, facial bones). The greatest diversity of the concentration of Ba, depending on gender of the subjects, was characteristic for facial bone samples (median value 4.65 μg Ba/g in samples from men, 6.64 μg Ba/g in samples from women) (Fig. 2). The difference in the concentration of Ba in the bone tissues from men and women was not statistically significant (Mann-Whitney U test, p=0.14).
Discussion As revealed by the study, the concentration of Ba in mineralized tissues amounted to an average of 6.61 μg/g (range 2.20– 15.47) (Table 1). The specified level of Ba was in the range of Ba concentration provided by other authors [7, 9, 16]. However, the concentration of Ba in the studied bone tissues was Fig. 2 Concentration of Ba (in μg/g) in deciduous teeth, impacted teeth, and bone tissues of male and female
35
higher than the concentration of this element in the biopsied rib bones of Japanese people, amounting to 1.3 μg/g [17]. Similarly, the concentration of Ba in impacted teeth surpassed the content of Ba in third molars of Taiwanese population twice [18]. A different level of concentration of Ba in mineralized tissues of people of Europe and Asia [17, 18] may be the result of a diverse contamination associated with the influence of different environmental (e.g., pH of the soil, pollution) and cultural factors (e.g., dietary preferences) of the studied populations. Places where surveyed people lived since birth belong to the most industrialized and urbanized areas of Poland [19], which increase the amount of bioavailable forms of Ba in the environment [2, 3]. Eating habits of Polish citizens also contribute to increasing the level of Ba in the body (a diet rich in animal protein, milk, and dairy products). Arearelated, statistically significant differences in the concentration of Ba in deciduous teeth have also been observed in the literature research including a population of people in Africa and Europe [20]. The study of Ba concentration was conducted in three types of bone tissues: deciduous teeth, permanent teeth, and facial bones. The analysis of the results showed that the concentration of Ba in particular types of mineralized tissues was significantly different (p≤0.05). The average level of Ba in samples of facial bones and permanent teeth was similar (amounting to 5.21 and 4.33 μg Ba/g) (Table 1). Permanent teeth covered by the survey were completely impacted molar teeth. These teeth remained in the bones of the upper or lower jaw, from all sides surrounded by bone tissue, and had no contact with the environment of the mouth. Among the studied mineralized tissues, the largest Ba concentration appeared
36
in deciduous teeth (10.47±3.24 μg/g) (Table 1). When compared to permanent teeth, a larger concentration of metals in the mineralized tissues of deciduous teeth has already been observed in previous own studies [21]. In the light of literature reports [3], the increase of the levels of Ba in deciduous teeth is affected by an increased absorption of environmental Ba by younger people. Among the study tissues (completely impacted teeth, facial bones, deciduous teeth), only deciduous teeth have direct contact with food and inhaled air in the mouth. Thus, there is potential for absorption of elements into the body in this way as well. Therefore, higher levels of Ba in deciduous teeth may be the result of transportation of this element to mineralized tooth tissue with blood, as well as direct incorporation from the environment. The content of barium in mineralized tissues of teeth may be affected by divergent histologic structure of permanent and milk teeth and by different mineralization state of individual tissues [18, 22]. The study showed that the level of Ba increased with age of the respondents in the case of impacted teeth and facial bones (Fig. 1). Changes in the concentration of Ba occurring in mineralized tissues of impacted teeth and facial bones have been defined by positive correlation coefficient. For samples of permanent teeth, the coexistence of Ba concentration changes in a function of age was described as significantly important correlation coefficient (0.64). Ba contents >6 μg/g in facial bone tissues and impacted teeth occurred mainly among people over the age of 40. In the light of literature research, the concentration of Ba among people aged 31–40 was higher than in the case of subjects up to 30 years of age [18]. The obtained results reflect the Ba accumulation mechanism occurring in the bone tissue of the body [3]. In case of deciduous teeth, the level of Ba decreased with increasing age of the subjects (Fig. 1). The differences in the Ba concentration in mineralized tissues of deciduous teeth have been defined by a statistically insignificant correlation coefficient (r=−0.39). The lowering of the level of elements’ concentration, mainly trace elements (Cr, Cu, Pb, Cd) in deciduous teeth with increasing age of the surveyed children, was already observed in previous own studies [23] and other authors’ study [24]. The results of literature research also show that the level of Ba in the body among children increases with the introduction of nutrients that complement breast-feeding [7]. The confirmation of this phenomenon may be the fact that in the conducted studies, the greatest level of Ba (>10 μg Ba/g) was found in samples taken from deciduous teeth of the youngest children (up to 10 years of age) (Fig. 1). The analysis of changes in the concentration of Ba in the examined tissues of men and women, at a significance level of p≤0.05, did not show statistically significant differences (Fig. 2). Statistically insignificant differences in the concentration of Ba in permanent teeth of men and women were found by Szostak et al. in the studies of an early medieval
Fischer et al.
coastal population of Gdańsk (Poland) [9]. Similarly to the studies of Szostek et al. [9], samples of all types of bone tissues taken from women (permanent teeth, deciduous teeth, facial bones) had higher levels of the Ba concentration as compared to samples from men (Fig. 2). The largest differences in the concentration of Ba in the samples from women and men concerned facial bones and then impacted teeth. The smallest differences in the concentration of Ba, depending on the gender of the subjects, were found in deciduous teeth. The level of Ba in deciduous teeth of boys and girls was similar and amounted to 11 and 10 μg Ba/g.
Summary The study showed a statistically significant difference in the concentration of Ba in mineralized tissues of deciduous teeth, permanent impacted teeth, and facial bones of people living in the industrial areas of southern Poland. The highest level of Ba was found in deciduous teeth, then in permanent impacted teeth and facial bones. The examined teeth and bone tissues obtained from women were characterized by a higher concentration of Ba when compared to samples from men, but the resulting differences were not statistically significant. In the mineralized tissues of permanent teeth and facial bones, unlike hard tissues of deciduous teeth, the concentration of Ba increased with age of the subjects. A statistically significant correlation coefficient (R=0.64) described only the changes of Ba concentration and the age of the subjects in mineralized tissues of impacted teeth. Acknowledgments This work was financed by the Medical University of Silesia in Katowice (contract no. KNW-1-124/K/4/0).
References 1. Polish Geological Institute, National Research Institute, http://www. pgi.gov.pl/attachments/article/434/war1_50zal.jpg. Accessed April 2014 2. Rosborg I, Nihlgard B, Gerhardsson L (2003) Ambio 32:440–446 3. Oskarsoon A, Reeves AL (2007) Barium. In: Nordberg GF et al (eds)Handbook on the Toxicology of metals. Elsevier, Amsterdam, pp 407–414 4. WHO (1990) Environ Health Criteria 107 5. Krachler M, Rossipal E, Micetic-Truk D (1999) Concentrations of trace elements in sera of newborns, young infants, and adults. Biol Trace Elem Res 68:121–135 6. Austin C, Smith TM, Bradman A, Hinde K, Joannes-Boyau R, Bishop D, Hare DJ, Doble P, Eskenazi B, Arora M (2013) Barium distributions in teeth reveal early-life dietary transition in primates. Nature 13(498):216–220 7. Tomczyk J, Szostek K, Komarnitki, Mańkowska-Pliszka H, Zalewska M (2013) Dental caries and chemical analyses in
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reconstruction of diet, health and hygienic behaviour in the Middle Euphrates (Syria). Arch Oral Biol 58:740–751 Rosenthal HL (1981) Content of stable strontium in man and animal biota. In: Skloryna SC (ed) Handbook of stable Strontium. Plenum, New York Szostek K, Gołąb H, Pudło A (2009) The use of strontium and barium analyses for the reconstruction of the diet of the early medieval coastal population of Gdańsk (Poland): a preliminary study. HOMO-J Comp Hum Biol 60:359–372 Arora M, Kennedy JB, Elhlou S, Pearson NJ, Walker DM, Bayl P, Chan SWY (2006) Spatial distribution of lead in human primary teeth as a biomarker of pre- and neonatal lead exposure. Sci Total Environ 371:55–62 Arora M, Hare D, Austin C, Smith DR, Doble P (2011) Spatial distribution of manganese in enamel and coronal dentine of human primary teeth. Sci Total Environ 409:1315–1319 De Souza-Guerra C, Barroso RC, de Almeida AP, Pexioto AIT, Moreira S, de Sousa FB, Gerlach RF (2014) Anatomical variations in primary teeth microelements with known differences in lead content by micro-synchrotron radiation X-Ray fluorescence (μSRXRF)—a preliminary study. J Trace Elem Med Biol 28:186–193 Lis J, Pasieczna A (1995) Geochemical atlas of Upper Silesia. Państwowy Instytut Wydawniczy, Warszawa Kwapuliński J, Malara P, Fischer A, Wiechuła D (2004) The teeth as the bioindicators of environmental exposure to chromium. Pol J Env Stud 13:43–48 Fischer A, Wiechuła D, Kwapuliński J (2004) The lead content of teeth in inhabitants of the Silesian region. Czas Stomatol 57(3):177–182
37 16. Manea-Krichten M, Patterson C, Miller G, Settle D, Erel Y (1991) Comparative increases of lead and barium with age in human tooth enamel, rib and ulna. Sci Total Environ 107: 179–203 17. Yoshinaga J, Suzuki T, Morita M, Hayakawa M (1995) Trace elements in ribs of elderly people and elemental variation in the presence of chronic diseases. Sci Total Environ 162:239–252 18. Liu HY, Chao JH, Chuang CY, Chiu HL, Yang CW, Sun YC (2013) Study of P, Ca, Sr, Ba and Pb levels in enamel and dentine of human third molars for environmental and archaeological research. Adv Archeol 3(2):71–77 19. Inspection of Environmental Protection (2010) Wyniki pięcioletniej ocent jakości powietrza w strefach w Polsce. IEP, Warszawa 20. Brown CJ, Chenery SR, Smith B, Tomkins A, Roberts GJ, Sserunjogi L, Thompson M (2002) A sampling and analytical methodology for dental trace element analysis. Analyst 127(2):319–323 21. Fischer A, Wiechuła D, Postek-Stefańska L, Kwapuliński J (2009) Concentrations of metals in maxilla and mandible deciduous and permanent teeth. Biol Trace Elem Res 132:19–26 22. Szpringer-Nodzak M, Wochna-Sobańska M (2005) Stomatologia wieku rozwojowego. PZWL, Warszawa 23. Fischer A, Wiechuła D, Przybyła-Misztela C (2013) Changes of concentrations of elements in deciduous teeth with age. Biol Trace Elem Res 154:427–432 24. He B, Huang S, Zhang C, Jing J, Hao Y, Xiao L, Zhou X (2011) Mineral densities and elemental content in different layers of healthy human enamel with varying teeth age. Arch Oral Biol 56:997–1004
The Study of Barium Concentration in Deciduous Teeth, Impacted Teeth, and Facial Bones of Polish Residents Agnieszka Fischer & Piotr Malara & Danuta Wiechuła
Received: 14 May 2014 / Accepted: 29 June 2014 / Published online: 1 August 2014 # Springer Science+Business Media New York 2014
Abstract The study determines the concentration of Ba in mineralized tissues of deciduous teeth, permanent impacted teeth, and facial bones. The study covers the population of children and adults (aged 6–78) living in an industrial area of Poland. Teeth were analyzed in whole, with no division into dentine and enamel. Facial bones and teeth were subjected to the following preparation: washing, drying, grinding in a porcelain mortar, sample weighing (about 0.2 g), and microwave mineralization with spectrally pure nitric acid. The aim of the study was to determinate the concentration of Ba in deciduous teeth, impacted permanent teeth, and facial bones. The concentration of barium in samples was determined over the ICP OES method. The Ba concentration in the tested bone tissues amounted to 2.2-15.5 μg/g (6.6 μg/g± 3.9). The highest concentration of Ba was present in deciduous teeth (10.5 μg/g), followed by facial bones (5.2 μg/g), and impacted teeth (4.3 μg/g) (ANOVA Kruskal-Wallis rank test, p=0.0002). In bone tissue and impacted teeth, Ba concentration increased with age. In deciduous teeth, the level of Ba decreased with children’s age.
Keywords Barium . Deciduous teeth . Impacted teeth . Facial bones
A. Fischer (*) : D. Wiechuła Department of Toxicology, School of Pharmacy with the Division of Laboratory Medicine, Medical University of Silesia, Katowice, Poland e-mail: [email protected] P. Malara Institute of Engineering Materials and Biomaterials, Silesian University of Technology, Gliwice, Poland
Introduction Barium (Ba) is a common element in the environment. Its share in lithosphere, mostly in the form of barite (BaSO4), amounts to approximately 0.04 %. In Poland, barite motherlodes occur in the southern part of the country (Sudeten, the Świętokrzyskie Mountains, Tatra Mountains) [1]. Barium can be released into the environment during geological processes, but its main source in the environment is the anthropogenic activity. Barium release accompanies mining operations and coal and fuel combustion processes. Its use is very broad and diverse (e.g., chemical, plastic, metallurgical, pharmaceutical, cosmetic, and medical industries). Barium present in the soil is relatively stable; however, acidification of the substrate leads to a formation of watersoluble compounds. The content of Ba in drinking water is highly variable and depends, among other things, on its hardness. Higher contents of Ba occur in acidic water when compared with alkaline [2, 3]. Plants absorb Ba directly from the environment, whereas animals absorb it from food. The concentration of barium in tissues of living organisms is diverse and depends on a position in the trophic chain [3]. The human body absorbs it mainly by ingestion and, in a negligible quantity, by inhalation. The average concentration in food does not exceed 0.1 mg Ba/kg [3]. A daily dose of Ba supplied with food to the human body amounts to 0.3–1.8 mg, and its donor is mainly milk and dairy products (25 % of dose) [3, 4]. Ba absorption from food is also subject to individual variability as it is greater among younger people [3]. During pregnancy, Ba transport from mother to fetus is limited by the placenta, which is indicated by lower concentrations of Ba in the umbilical cord sera [5, 6]. The literature studies also showed that the Ba concentration in mothers’ milk correlates with its concentration in hard tissues of deciduous teeth [6]. In addition, a clear increase in the quotient of Ba/Ca
Barium Concentration in Teeth and Facial Bones
is observed after cessation of breast-feeding and introduction of modified milk into the diet of infants [6]. The physiological role of barium in the body remains unexplained. It is known that this element is not involved in metabolism and does not participate in the process of maintaining homeostasis [7]. Barium is present in the body in various tissues (lung, fat tissue, skin, eye tissue), but 91 % of the concentration of this element is accumulated by the bone tissue [3, 4]. The main component of the mineralized tissues is hydroxyapatite, which has the ability to exchange elements. The inclusion of Ba in the hydroxyapatite structure occurs at the expense of the concentration of physiologically essential elements, including calcium [8, 9]. Literature research indicates a possibility of making an elemental analysis of teeth taking into account the type of calcified tissues and the mineralization period [6, 10–12]. Teeth being formed in utero, record birth as the neonatal line, define the pre- and postnatal developmental periods [6, 10, 11]. Calcified tissues accumulated different concentration of metals. Differences in the concentration of elements in enamel and dentine are observed. For example, the levels of Pb and Mn were higher in the dentine than in enamel [10, 11]. The highest concentration of Pb was found in postnatal dentine [10]. Barium is an element of which occurrence in the environment is connected mainly with anthropogenic activity. In an organism, it accumulates itself in bone tissue. In the available literature, there are no current research results on barium content in teeth of inhabitants of Poland. The objective of the study was to determine the concentration of Ba in teeth and bones of people living in the industrial areas of southern Poland (Silesian Region). The Silesian Region is one of the most industrialized and urbanized regions in Poland, characterized by high traffic and intensive emissions. In this area, there are acidic soils (pH 4.0–6.7), containing 100– 400 mg Ba/kg [13]. Research, including the authors’ own research, onto the determination of concentration of other elements (e.g., Cu, Cr, Pb, Ca, Fe) in teeth was conducted among inhabitants of the Silesian agglomeration. The obtained results indicate a higher metal concentration in teeth of inhabitants of the Silesian Region than in other regions of Poland [14–16]. In the study, the levels of Ba in three kinds of calcified tissues (deciduous teeth, permanent impacted teeth, and facial bones) were determined. The teeth were analyzed as a whole without making a histological division into calcified tissues. The differences in the concentration of Ba in the tissues covered by the study were analyzed depending on gender and age.
Materials and Methods The subjects of the research were samples of mineralized tissues from persons residing in the south of Poland. People
33
included in the research lived in the towns of the province of Silesia (Katowice, Tychy, Chorzow, Sosnowiec, BielskoBiala) since birth. The study tissues (n=35) included samples of facial bones (n=14), permanent impacted teeth (n=10), and deciduous teeth (n=11). All collected tissue samples came from healthy individuals that did not indicate a presence of chronic diseases of the body requiring treatment in the interview. The study covered healthy teeth that did not have signs of decay and fillings. The weight of the bone tissue given to research was at least 0.15 g. The research did not include tissues samples from people suffering from chronic diseases requiring pharmacological treatment. The research did not include teeth with decay and fillings. Among the mineralized tissues of the body, the teeth are a relatively easily accessible tissue. They can be collected at the time of medically reasoned tooth extractions and as the postoperative residual forwarded for further research. Deciduous teeth can be taken for testing over a completely noninvasive method to the patient, during physiological replacement of teeth. The deciduous teeth were collected over a noninvasive method during a physiological replacement. Permanent teeth were removed during a planned medical, surgical extraction of teeth. During the procedure of a surgical extraction, especially for impacted teeth, the formation of bone chips is a common complication. During the procedure, bone chips are removed to accelerate the process of healing of postextraction wounds. The investigated teeth and bone chips were postoperative residue. Studied deciduous teeth came from children (n=11, aged 6–12 years, mean 8.5±2.2) and were collected during natural replacement of teeth to permanent ones. Permanent teeth (n=10) came from people aged 16-41 (mean 29.0±10.5) and were removed during a planned medical, surgical extraction of tooth. The teeth were completely impacted, remaining in the upper or lower jaw bone. Completely impacted teeth are fully formed teeth, surrounded by bone tissue from all sides. Molar teeth were assigned for the research. Of all the types of teeth (incisors, canines, premolars, molars), third molars are the kind of teeth which is usually subject to irregularities in the process of eruption. During surgical tooth extraction procedures, the postoperative residue can also include bone fragments. Mandibular and maxillary bones (n=14) of people aged 16–78 (mean 44.4±21.5) were submitted for the research. All collected tissues samples came from women (n=16, aged 6–77 years, mean 30.9±24.9) and men (n=19, aged 7–78 years, mean 26.3±18.1). Bone tissues obtained for the research were subjected to the following preparation: washing, rinsing (tap water/distilled water), drying to constant weight (85 °C), grinding in a porcelain mortar, drying (85 °C/3 h), sample weighing (about 0.2 g), and mineralization (microwave digestion system
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MAGNUM II (prod. ERTEC-Poland), spectrally pure nitric acid; Merck, Germany). The concentration of barium in bone samples was determined over the ICP OES method (Perkin Elmer Optima 5300 DVTM). For calibration, the working solutions were prepared by a precisely measured dilution of standard solutions (Merck, Germany). The verification of determination accuracy was established by means of the standard addition method using the model of mass concentration of given metals. Blank samples were used after every ten samples. The procedure was validated by NIST 1486 (Bone Meal). Statistical Analysis Statistical results were calculated in Statistica 10 for Windows. Nonparametric tests were used to obtain the results of the research (Mann-Whitney U test for two groups and ANOVA Kruskal-Wallis rank for more than two groups). The main criterion for selection of nonparametric tests, apart from the abnormal distribution of results, was the small group size. The correlation analysis was based on Spearman’s nonparametric rank tests. To evaluate the statistical significance of the data, the probability of ≤0.05 was considered significant.
Results Table 1 shows the results of statistical analysis of the concentration of Ba in the studied tissues. The concentration of Ba in mineralized tissue samples amounted to 2.20–15.47 mg/g (arithmetic mean=6.61 μg/g). The highest concentration of Ba was present in deciduous teeth (10.47 μg/g), followed by facial bones (5.21 μg/g), and the lowest was in the impacted permanent teeth (4.33 μg/g) (Table 1). Ba concentration differences within the bone tissue groups were statistically significant (ANOVA Kruskal-Wallis rank test, p=0.0002). The change in the Ba concentration in the studied tissues occurring with age of the subjects is shown in Fig. 1. The graph illustrating the variation of the concentration of Ba in the function Table 1 Concentration of Ba (in μg/g) in deciduous teeth, impacted teeth, and bone tissues Number Mean ± SD GM Deciduous teeth 11 Impacted teeth 10 Bone tissues 14
Median Min. Max.
10.47±3.24 9.96 11.11* 4.33±2.22 3.90 3.33* 5.21±2.06 4.84 4.92*
5.55 2.20 2.50
15.47 9.24 9.29
GM geometric mean, Min. minimum values, Max. maximum values *p=0.0002 (significant differences, ANOVA Kruskal-Wallis rank) *p=0.0002 (significant differences, ANOVA Kruskal-Wallis rank)
Fig. 1 The relationship between the concentration of Ba in deciduous teeth, impacted teeth, and bone tissues and age. Spearman’s rank correlation
of age has proven an increase of the concentration of this element with increasing age in the case of permanent teeth and facial bones. In the case of deciduous teeth, the level of Ba decreased with increasing age of the subjects (Fig. 1). The lowest concentration of Ba in deciduous teeth was observed in the oldest person in the study (5.55 μg/g; age 12 years). For facial bones and permanent teeth, the lowest levels of Ba occurred among persons of 25 years of age. The correlation
Barium Concentration in Teeth and Facial Bones
coefficient specifying the change of the concentration of Ba in all studied tissues with increasing age of the patients was −0.43 and was statistically significant (Spearman’s rank correlation, p=0.01). An analysis of co-occurrence of Ba concentration changes with age of the subjects in different types of mineralized tissues (deciduous teeth, permanent impacted teeth, facial bones) showed that the statistically significant correlation coefficient had defined only the changes in the concentration of Ba and age in the impacted teeth (Spearman’s rank correlation, r=0.64, p=0.05). Studies of Ba concentration changes in mineralized tissues of men and women have shown that higher levels of Ba concentration characterized the samples taken from women (Fig. 2). Higher average levels of Ba in samples from women compared to men concerned all studied bone tissues (deciduous teeth, permanent teeth, facial bones). The greatest diversity of the concentration of Ba, depending on gender of the subjects, was characteristic for facial bone samples (median value 4.65 μg Ba/g in samples from men, 6.64 μg Ba/g in samples from women) (Fig. 2). The difference in the concentration of Ba in the bone tissues from men and women was not statistically significant (Mann-Whitney U test, p=0.14).
Discussion As revealed by the study, the concentration of Ba in mineralized tissues amounted to an average of 6.61 μg/g (range 2.20– 15.47) (Table 1). The specified level of Ba was in the range of Ba concentration provided by other authors [7, 9, 16]. However, the concentration of Ba in the studied bone tissues was Fig. 2 Concentration of Ba (in μg/g) in deciduous teeth, impacted teeth, and bone tissues of male and female
35
higher than the concentration of this element in the biopsied rib bones of Japanese people, amounting to 1.3 μg/g [17]. Similarly, the concentration of Ba in impacted teeth surpassed the content of Ba in third molars of Taiwanese population twice [18]. A different level of concentration of Ba in mineralized tissues of people of Europe and Asia [17, 18] may be the result of a diverse contamination associated with the influence of different environmental (e.g., pH of the soil, pollution) and cultural factors (e.g., dietary preferences) of the studied populations. Places where surveyed people lived since birth belong to the most industrialized and urbanized areas of Poland [19], which increase the amount of bioavailable forms of Ba in the environment [2, 3]. Eating habits of Polish citizens also contribute to increasing the level of Ba in the body (a diet rich in animal protein, milk, and dairy products). Arearelated, statistically significant differences in the concentration of Ba in deciduous teeth have also been observed in the literature research including a population of people in Africa and Europe [20]. The study of Ba concentration was conducted in three types of bone tissues: deciduous teeth, permanent teeth, and facial bones. The analysis of the results showed that the concentration of Ba in particular types of mineralized tissues was significantly different (p≤0.05). The average level of Ba in samples of facial bones and permanent teeth was similar (amounting to 5.21 and 4.33 μg Ba/g) (Table 1). Permanent teeth covered by the survey were completely impacted molar teeth. These teeth remained in the bones of the upper or lower jaw, from all sides surrounded by bone tissue, and had no contact with the environment of the mouth. Among the studied mineralized tissues, the largest Ba concentration appeared
36
in deciduous teeth (10.47±3.24 μg/g) (Table 1). When compared to permanent teeth, a larger concentration of metals in the mineralized tissues of deciduous teeth has already been observed in previous own studies [21]. In the light of literature reports [3], the increase of the levels of Ba in deciduous teeth is affected by an increased absorption of environmental Ba by younger people. Among the study tissues (completely impacted teeth, facial bones, deciduous teeth), only deciduous teeth have direct contact with food and inhaled air in the mouth. Thus, there is potential for absorption of elements into the body in this way as well. Therefore, higher levels of Ba in deciduous teeth may be the result of transportation of this element to mineralized tooth tissue with blood, as well as direct incorporation from the environment. The content of barium in mineralized tissues of teeth may be affected by divergent histologic structure of permanent and milk teeth and by different mineralization state of individual tissues [18, 22]. The study showed that the level of Ba increased with age of the respondents in the case of impacted teeth and facial bones (Fig. 1). Changes in the concentration of Ba occurring in mineralized tissues of impacted teeth and facial bones have been defined by positive correlation coefficient. For samples of permanent teeth, the coexistence of Ba concentration changes in a function of age was described as significantly important correlation coefficient (0.64). Ba contents >6 μg/g in facial bone tissues and impacted teeth occurred mainly among people over the age of 40. In the light of literature research, the concentration of Ba among people aged 31–40 was higher than in the case of subjects up to 30 years of age [18]. The obtained results reflect the Ba accumulation mechanism occurring in the bone tissue of the body [3]. In case of deciduous teeth, the level of Ba decreased with increasing age of the subjects (Fig. 1). The differences in the Ba concentration in mineralized tissues of deciduous teeth have been defined by a statistically insignificant correlation coefficient (r=−0.39). The lowering of the level of elements’ concentration, mainly trace elements (Cr, Cu, Pb, Cd) in deciduous teeth with increasing age of the surveyed children, was already observed in previous own studies [23] and other authors’ study [24]. The results of literature research also show that the level of Ba in the body among children increases with the introduction of nutrients that complement breast-feeding [7]. The confirmation of this phenomenon may be the fact that in the conducted studies, the greatest level of Ba (>10 μg Ba/g) was found in samples taken from deciduous teeth of the youngest children (up to 10 years of age) (Fig. 1). The analysis of changes in the concentration of Ba in the examined tissues of men and women, at a significance level of p≤0.05, did not show statistically significant differences (Fig. 2). Statistically insignificant differences in the concentration of Ba in permanent teeth of men and women were found by Szostak et al. in the studies of an early medieval
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coastal population of Gdańsk (Poland) [9]. Similarly to the studies of Szostek et al. [9], samples of all types of bone tissues taken from women (permanent teeth, deciduous teeth, facial bones) had higher levels of the Ba concentration as compared to samples from men (Fig. 2). The largest differences in the concentration of Ba in the samples from women and men concerned facial bones and then impacted teeth. The smallest differences in the concentration of Ba, depending on the gender of the subjects, were found in deciduous teeth. The level of Ba in deciduous teeth of boys and girls was similar and amounted to 11 and 10 μg Ba/g.
Summary The study showed a statistically significant difference in the concentration of Ba in mineralized tissues of deciduous teeth, permanent impacted teeth, and facial bones of people living in the industrial areas of southern Poland. The highest level of Ba was found in deciduous teeth, then in permanent impacted teeth and facial bones. The examined teeth and bone tissues obtained from women were characterized by a higher concentration of Ba when compared to samples from men, but the resulting differences were not statistically significant. In the mineralized tissues of permanent teeth and facial bones, unlike hard tissues of deciduous teeth, the concentration of Ba increased with age of the subjects. A statistically significant correlation coefficient (R=0.64) described only the changes of Ba concentration and the age of the subjects in mineralized tissues of impacted teeth. Acknowledgments This work was financed by the Medical University of Silesia in Katowice (contract no. KNW-1-124/K/4/0).
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