Accepted Manuscript Title: Reference values of hair toxic trace elements content in occupationally non-exposed Russian population Author: Anatoly V. Skalny Margarita G. Skalnaya Alexey A. Tinkov Eugeny P. Serebryansky Vasily A. Demidov Yulia N. Lobanova Andrei R. Grabeklis Elena S. Berezkina Irina V. Gryazeva Andrey A. Skalny Alexandr A. Nikonorov PII: DOI: Reference:
S1382-6689(15)00119-2 http://dx.doi.org/doi:10.1016/j.etap.2015.05.004 ENVTOX 2260
To appear in:
Environmental Toxicology and Pharmacology
Received date: Revised date: Accepted date:
21-3-2015 8-5-2015 10-5-2015
Please cite this article as: Skalny, A.V., Skalnaya, M.G., Tinkov, A.A., Serebryansky, E.P., Demidov, V.A., Lobanova, Y.N., Grabeklis, A.R., Berezkina, E.S., Gryazeva, I.V., Skalny, A.A., Nikonorov, A.A.,Reference values of hair toxic trace elements content in occupationally non-exposed Russian population, Environmental Toxicology and Pharmacology (2015), http://dx.doi.org/10.1016/j.etap.2015.05.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
1
Reference values of hair toxic trace elements content in occupationally non-exposed
2
Russian population
3 4
Anatoly V. Skalny 1,2,3, Margarita G. Skalnaya 2,3, Alexey A. Tinkov 1,4, Eugeny P. Serebryansky
5
3
6
Irina V. Gryazeva 3, Andrey A. Skalny 3,5, Alexandr A. Nikonorov 4
, Elena S. Berezkina
8
1
9
Sovetskaya st., 14, Yaroslavl, 150000, Russia
,
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Laboratory of biotechnology and applied bioelementology, Yaroslavl State University,
10
2
11
UNESCO), Orenburg State University, Pobedy Ave. 13, Orenburg 460352, Russia
12
3
13
Zemlyanoy Val St. 46, Moscow 105064, Russia
14
4
15
460000, Russia
16
5
17
Agency, Bekhtereva str. 1, St. Petersburg 192019, Russia
an
Institute of Bioelementology (Russian Satellite Centre of Trace Element – Institute for
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Russian Society of Trace Elements in Medicine, ANO “Centre for Biotic Medicine”,
te
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Department of Biochemistry, Orenburg State Medical University, Sovetskaya St., 6, Orenburg,
Federal State Scientific Institution “Institute of Toxicology”, Federal Medico-Biological
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, Vasily A. Demidov 3, Yulia N. Lobanova 3, Andrei R. Grabeklis
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Corresponding author
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Alexey A. Tinkov, MD, PhD
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Laboratory of biotechnology and applied bioelementology, Yaroslavl State University,
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Sovetskaya st., 14, Yaroslavl, 150000, Russia
23
Tel: +7-961-937-81-98
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E-mail: [email protected]
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1 Page 1 of 11
Abstract
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A total of 5908 occupationally non-exposed adults (4384 women and 1524 men) living in
3
Moscow and Moscow region were involved in the current investigation. Hair Al, As, Be, Bi, Cd,
4
Hg, Li, Ni, Pb, Sn, Sr content was estimated by inductively-coupled plasma mass spectrometry
5
using NexION 300D. Men are characterized by significantly higher hair Al, As, Cd, Hg, Li, and
6
Pb content. At the same time, hair levels of Bi, Ni, Sn, and Sr were significantly higher in
7
women. Consequently, the reference ranges were estimated for male, female, and general cohort
8
as coverage intervals in accordance with IUPAC recommendations.
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Key words: inductively coupled plasma mass spectrometry; trace elements; hair; reference
11
ranges; coverage intervals; IUPAC recommendations
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1. Introduction
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Heavy metal exposure has been shown to be associated with a number of diseases (Järup, 2003)
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and its monitoring is an essential part of environmental and health care (Wolterbeek, 2002). Hair
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has been widely used as a bioindicator of human exposure to heavy metals (Bencko, 1995). It
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has been shown that hair may successfully reflect environmental and occupational exposure to
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lead (Sen, 1996), mercury (Kruzikova et al., 2009), aluminium (Yokel, 1982) and combination of
19
multiple metals (Wang and Fu, 2009). At the same time, a number of side factors like
20
demography, lifestyle and geography (Christensen, 1995) may affect hair trace element content.
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Despite the presence of a number of studies indicating reference ranges of hair metal content, the
22
existing data are inconsistent (Mikulewicz et al., 2013). Therefore, the primary aim of the current
23
study is estimation of reference ranges of hair toxic trace elements content in adult Russian
24
population.
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2. Materials and methods 2 Page 2 of 11
A total of 5908 occupationally non-exposed adults (4384 women and 1524 men) aged from 20 to
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60 years and living in Moscow and Moscow region were involved in current investigation. The
3
investigation has been carried out in accordance with the principles of the Declaration of
4
Helsinki for studies involving humans and was approved by the Local Ethics Committee. All
5
examinees gave their informed consent prior to the inclusion in the study.
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The participants were selected for the study with the use of the following exclusion criteria: i)
7
occupational exposure to toxic metals; ii) smoking (both present and former smokers); iii) acute
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inflammatory diseases; iv) endocrine disorders; v) metallic implants; vi) pregnancy and lactation;
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vii) vegetarian diet, viii) alcohol abuse.
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Proximal parts of occipital scalp hair (0.1 gram) were collected. Briefly, hair samples were
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washed with acetone and then rinsed thrice with deionized water (Zhao et al., 2012). After
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washing hair samples were dried at 60°C on air with subsequent microwave degradation. Briefly,
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0.05 g of hair samples were introduced into Teflon tubes and added with concentrated HNO3.
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Digestion was performed in a Berghof speedwave four system during 20 minutes at 170-180°C.
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The obtained solutions were added with distilled deionized water to a final volume of 15 ml.
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Hair toxic trace element content (Al, As, Be, Bi, Cd, Hg, Li, Ni, Pb, Sn, Sr) was estimated by
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inductively-coupled plasma mass spectrometry with NexION 300D (PerkinElmer Inc., Shelton,
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CT 06484, USA) using Dynamic Reaction Cell technology removing the majority of
19
interferences with minimal loss of analyte sensitivity and equipped with ESI SC-2 DX4
20
autosampler (Elemental Scientific Inc., Omaha, NE 68122, USA).
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Manufacturer’s specifications were used for ICP-MS system preparation. Calibration was
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performed using standards containing 0.5, 5, 10, and 50 μg/l ultra-trace elements prepared from
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Universal Data Acquisition Standards Kit (PerkinElmer Inc., Shelton, CT 06484, USA) by
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dilution with distilled deionized water acidified with 1% HNO3. An internal online
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standardization using yttrium-89 isotope was also performed. Internal standard containing 10
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μg/l yttrium was prepared from Yttrium (Y) Pure Single-Element Standard (PerkinElmer Inc.,
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Shelton, CT 06484, USA) on a matrix containing 8% 1-butanol (Merck KGaA), 0.8% Triton X-
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100 (Sigma-Aldrich, Co.), 0.02% tetramethylammonium hydroxide (Alfa-Aesar, Ward Hill, MA
3
01835 USA), and 0.02% ethylenediaminetetraacetic acid (Sigma-Aldrich, Co). For laboratory
4
quality control the certified reference material of human hair GBW09101 (Shanghai Institute of
5
Nuclear Research, Shanghai, China) was used.
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All analyses were performed in accredited clinic-diagnostic laboratory of autonomous non-profit
7
organization “Centre for Biotic Medicine” (Moscow, Russia), International Union of Pure and
8
Applied Chemistry (IUPAC) company associate.
9
The obtained data were treated with Statistica 11 software (Statsoft, USA). Data distribution was
10
evaluated by the Shapiro-Wilk test. As the distribution was not Gaussian, all variables were log
11
transformed to approximate a normal distribution. Natural logarithms were used for further
12
statistical analyses. One-way ANOVA with Fisher’s LSD post hoc test were used for group
13
comparisons. The values were considered significantly different at p < 0.05.
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The reference values of hair toxic trace elements content were based on calculation of the 0.95
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coverage intervals with 0.95 confidence intervals for the upper and lower limits in accordance
16
with IUPAC recommendations (Poulsen et al., 1997).
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3. Results and discussion
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The descriptive statistics of the male and female cohorts (Table 1) indicates that age of the
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examinees was not statistically different between the gender groups (p = 0.245). Hair trace
21
elements analysis in men and women (Table 1) indicated that the hair level of certain trace
22
elements is sex-related. In particular, men are characterized by significantly higher hair
23
aluminium (p < 0.001), arsenic (p < 0.001), cadmium (p < 0.001), lithium (p = 0.001), mercury
24
(p = 0.002), and lead (p < 0.001) content. At the same time, hair levels of bismuth (p < 0.001),
25
nickel (p < 0.001), tin (p < 0.001), and strontium (p < 0.001) were significantly higher in women
26
in comparison to the respective values in men. No significant difference was found between men 4 Page 4 of 11
and women in hair beryllium content (p = 0.222). Hypothetically, significantly increased hair
2
mercury, lead, arsenic, and aluminium content in males may result from increased outdoor
3
activity in men. The obtained data is in agreement with earlier studies indicating sex-related
4
difference in hair cadmium, lead (DiPietro et al., 1989), arsenic (Wolfsperger et al., 1994),
5
mercury (Shimomura et al., 1980), nickel (Michalak et al., 2012), aluminium (Chojnacka et al.,
6
2010), and strontium (Dongarra et al., 2011). At the same time, our research data on hair tin
7
content are in contrast to the study of Chojnacka et al. (2010), who have failed to detect a
8
significant gender difference in hair Sn levels. Consequently, sex-specific reference ranges
9
should be used while using hair as an indicator of environmental exposure.
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Reference ranges calculated as coverage intervals for hair toxic element content in a males,
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females and general cohort are provided in Table 2. The obtained data on adults’ hair metal
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content reference ranges are generally comparable with earlier published studies performed in
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Sweden (Rodushkin and Axelsson, 2000), France (Goullé et al., 2005), and Poland (Chojnacka et
14
al., 2010). At the same time, the most significant difference between our reference ranges and the
15
recently published ones were observed in the case of hair cadmium, lead, mercury, and strontium
16
content. It is notable that in the studied cohort hair cadmium and lead content was significantly
17
lower than the respective values postulated in all three previous studies (Rodushkin and
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Axelsson, 2000; Goullé et al., 2005; Chojnacka et al., 2010). The opposite situation was
19
observed in the case of hair mercury and strontium. In particular, our reference values were
20
higher than the reference values obtained for hair levels of these metals by Rodushkin and
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Axelsson, 2000, Goullé et al., 2005, and Chojnacka et al., 2010. It should be noted that our
22
recent data is generally comparable with our earlier published reference values obtained by
23
means of inductively coupled plasma atomic emission spectrometry (Skalny, 2003). The
24
observed difference in reference ranges of hair trace element content may be a consequence of a
25
number of factors, like geographical location, climate, and the overall development of industry in
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the selected location (Christensen, 1995). Moreover, all four studies involved examinees of
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different ages that may also have a significant influence on hair trace elements content
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(Ambeskovic et al., 2013). Finally, various statistical approaches to estimation of the reference
3
ranges may also result in some contradictions.
4
Generally, the reference ranges for hair toxic trace element in male, female, and general samples
5
have been estimated. Consistency of the obtained data with previously published studies, a large
6
number of samples examined, and the use of Dynamic Reaction Cell technology during analysis
7
indicate high quality of the data obtained. The estimated reference ranges may be used in
8
environmental risk assessment (Tamburo et al., 2011).
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Conclusions
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The present study has demonstrated that men are characterized by significantly higher hair Al,
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As, Cd, Hg, Li, and Pb content. Hair levels of Bi, Ni, Sn, and Sr were significantly higher in
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women. Consequently, the reference ranges calculated as coverage intervals in accordance with
14
IUPAC recommendations are different for males, females, and the general cohort.
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Acknowledgements
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The current research is supported by Russian Ministry of Education and Science within project
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No. 2014/258-544. The authors would like to thank the anonymous reviewers for their helpful
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comments and suggestions that significantly improved the manuscript quality.
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Conflict of interest
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The authors declare that there are no conflicts of interest
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1. Ambeskovic, M., Fuchs, E., Beaumier, P., Gerken, M., Metz, G.A., 2013. Hair trace
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elementary profiles in aging rodents and primates: links to altered cell homeodynamics
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and disease. Biogerontology 14, 557-567.
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pollutants in occupational and environmental settings. Toxicology 101, 29-39.
3. Chojnacka, K., Zielińska, A., Górecka, H., Dobrzański, Z., Górecki, H., 2010. Reference values for hair minerals of Polish students. Environ. Toxicol. Pharmacol. 29, 314-9.
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2. Bencko, V., 1995. Use of human hair as a biomarker in the assessment of exposure to
4. Christensen, J.M., 1995 Human exposure to toxic metals: factors influencing
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interpretation of biomonitoring results. Sci. Total Environ. 166, 89-135. 5. DiPietro, E.S., Phillips, D.L., Paschal, D.C., Neese, J.W., 1989. Determination of trace
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elements in human hair. Reference intervals for 28 elements in nonoccupationally
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exposed adults in the US and effects of hair treatments. Biol. Trace Elem. Res. 22, 83-
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100.
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6. Dongarrà, G., Lombardo, M., Tamburo, E., Varrica, D., Cibella, F., Cuttitta, G., 2011.
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Concentration and reference interval of trace elements in human hair from students living
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in Palermo, Sicily (Italy). Environ. Toxicol. Pharmacol. 32, 27-34.
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7. Goullé, J.P., Mahieu, L., Castermant, J., Neveu, N., Bonneau, L., Lainé, G., Bouige, D., Lacroix, C., 2005. Metal and metalloid multi-elementary ICP-MS validation in whole blood, plasma, urine and hair. Reference values. Forensic Sci. Int. 153, 39-44.
8. Järup, L., 2003. Hazards of heavy metal contamination. Br. Med. Bull. 68, 167-82. 9. Kruzikova, K., Kensova, R., Blahova, J., Harustiakova, D., Svobodova, Z., 2009. Using human hair as an indicator for exposure to mercury. Neuro Endocrinol. Lett. 30, 177-81.
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10. Michalak, I., Mikulewicz, M., Chojnacka, K., Wołowiec, P., Saeid, A., Górecki, H.,
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2012. Exposure to nickel by hair mineral analysis. Environ. Toxicol. Pharmacol. 34,
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727-34.
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11. Mikulewicz, M., Chojnacka, K., Gedrange, T., Górecki, H., 2013. Reference values of elements in human hair: a systematic review. Environ. Toxicol. Pharmacol. 36, 1077-86. 12. Poulsen, O.M., Holst, E., Christensen, J.M., 1997. Calculation and application of
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coverage intervals for biological reference values. Pure Appl. Chem. 69, 1601-1611.
5
13. Rodushkin, I., Axelsson, M.D., 2000. Application of double focusing sector field ICP-MS
6
for multielemental characterization of human hair and nails. Part II. A study of the
7
inhabitants of northern Sweden. Sci. Total. Environ. 262, 21-36.
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Lead Exposure. J. Hum. Ecol. 7, 133-141.
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14. Sen, J., 1996. Human Scalp Hair As An Indicator of Environmental Lead Pollution and
15. Shimomura, S., Kimura, A., Nakagawa, H., Takao, M.., 1980. Mercury levels in human
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hair and sex factors. Environ. Res. 22, 22-30.
16. Skalny, A. V., 2003. Reference values of chemical elements concentration in hair,
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obtained by means of ICP-AES method in ANO Centre for Biotic Medicine. Trace.
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Elem. Med. 4, 55-56.
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17. Tamburo, E., Dongarrà, G., Varrica, D., D’Andrea, D., 2011. Trace elements in hair of
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urban schoolboys: a diagnostic tool in environmental risk assessment. Geophysical
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Research Abstracts 13, 1157
18. Wang, T., Fu, J., Wang, Y., Liao, C., Tao, Y., Jiang, G., 2009. Use of scalp hair as indicator of human exposure to heavy metals in an electronic waste recycling area. Environ. Pollut. 157, 2445-51.
19. Wolfsperger, M., Hauser, G., Gössler, W., Schlagenhaufen, C., 1994. Heavy metals in
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human hair samples from Austria and Italy: influence of sex and smoking habits. Sci.
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Total Environ. 156, 235-42.
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20. Wolterbeek, B., 2002. Biomonitoring of trace element air pollution: principles, possibilities and perspectives. Environ. Pollut. 120, 11-21.
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1 2
21. Yokel, R.A., 1982. Hair as an indicator of excessive aluminum exposure. Clin. Chem. 28, 662-5. 22. Zhao, L.J., Ren, T., Zhong, R.G., 2012. Determination of lead in human hair by high
4
resolution continuum source graphite furnace atomic absorption spectrometry with
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microwave digestion and solid sampling. Analyt. Lett. 45, 2467-81.
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Table 1
Men (n = 1524)
Women (n = 4384)
Parameter Mean
SD
Median P5
P95
an
P95
Mean
General sample (n = 5908) SD
Median P5
P95
Mean
SD
37.026
22.575 56.306 37.914 10.306
36.314
22.969 55.894 37.560 10.198
36.506
22.844 56.008 37.651 8.646
Al, μg/g
5.704
1.926
21.436 7.876
8.439
4.876
1.742
16.020 6.609
8.694
6.936
1.785
16.958 6.936
0.110
As, μg/g
0.046
0.019
0.213
0.074
0.149
0.021
0.006
0.097
0.035
0.091
0.045
0.007
0.135
0.045
0.026
Be, μg/g
0.002
0.000
0.014
0.003
0.008
0.002
0.000
0.010
0.003
0.030
0.003
0.000
0.011
0.003
1.931
Bi, μg/g
0.036
0.007
0.342
0.187
1.917
0.043
0.007
0.735
0.251
1.937
0.234
0.007
0.674
0.234
0.309
Cd, μg/g
0.015
0.004
0.168
0.063
0.594
0.011
0.003
0.070
0.024
0.074
0.034
0.003
0.090
0.034
1.003
Hg, μg/g
0.566
0.076
2.744
0.891
1.086
0.498
0.112
2.002
0.734
0.969
0.775
0.106
2.212
0.775
0.115
Li, μg/g
0.013
0.005
0.070
0.026
0.167
0.010
0.005
0.063
0.023
0.091
0.024
0.005
0.065
0.024
1.057
Ni, μg/g
0.198
0.078
0.809
0.303
0.504
0.265
0.091
1.167
0.451
1.188
0.413
0.087
1.057
0.413
14.943
Pb, μg/g
0.501
0.121
4.689
2.528
29.289
0.295
0.083
1.462
0.532
1.408
1.046
0.088
2.142
1.046
2.568
Sn, μg/g
0.088
0.028
0.402
0.212
2.938
0.133
0.025
2.760
0.647
2.416
0.535
0.026
2.151
0.535
18.468
Sr, μg/g
1.119
0.313
6.624
2.102
4.097
3.990
0.683
25.370 8.696
21.157
6.935
0.455
20.319 6.935
ep te
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Age, years
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Median P5
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Table 1. Descriptive statistics of age and hair toxic trace elements content in adult men and women
8.646
P5, P95 – 5 and 95 percentile boundaries; SD – standard deviation;
Page 10 of 11
Table 2
Table 2. Coverage intervals for hair trace elements content in adult Russian population calculated in accordance with IUPAC recommendations. Men (n = 1524)
Women (n = 4384)
General sample (n = 5908)
Al, μg/g
3.355-15.279
2.778-10.545
2.913-11.627
As, μg/g
0.036-0.117
0.008-0.062
0.010-0.078
Be, μg/g
0.000-0.007
0.000-0.005
0.000-0.005
Bi, μg/g
0.017-0.234
0.017-0.336
0.014-0.342
Cd, μg/g
0.009-0.088
0.005-0.042
0.006-0.056
Hg, μg/g
0.130-1.365
0.185-1.094
0.168-1.189
Li, μg/g
0.009-0.039
0.009-0.040
Ni, μg/g
0.132-0.540
0.168-0.779
0.159-0.704
Pb, μg/g
0.291-2.358
0.160-0.917
0.187-1.389
Sn, μg/g
0.051-0.265
0.082-1.158
0.076-1.009
Sr, μg/g
0.650-4.173
1.570-15.181
1.148-10.929
δ
0.008
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ed
0.005
0.009-0.040
0.004
ce pt
δ – coverage uncertainty;
ip t
Metal content
β level of expectation in coverage interval - 0.95;
Ac
γ level of confidence of the coverage uncertainty - 0.95.
Page 11 of 11
S1382-6689(15)00119-2 http://dx.doi.org/doi:10.1016/j.etap.2015.05.004 ENVTOX 2260
To appear in:
Environmental Toxicology and Pharmacology
Received date: Revised date: Accepted date:
21-3-2015 8-5-2015 10-5-2015
Please cite this article as: Skalny, A.V., Skalnaya, M.G., Tinkov, A.A., Serebryansky, E.P., Demidov, V.A., Lobanova, Y.N., Grabeklis, A.R., Berezkina, E.S., Gryazeva, I.V., Skalny, A.A., Nikonorov, A.A.,Reference values of hair toxic trace elements content in occupationally non-exposed Russian population, Environmental Toxicology and Pharmacology (2015), http://dx.doi.org/10.1016/j.etap.2015.05.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
1
Reference values of hair toxic trace elements content in occupationally non-exposed
2
Russian population
3 4
Anatoly V. Skalny 1,2,3, Margarita G. Skalnaya 2,3, Alexey A. Tinkov 1,4, Eugeny P. Serebryansky
5
3
6
Irina V. Gryazeva 3, Andrey A. Skalny 3,5, Alexandr A. Nikonorov 4
, Elena S. Berezkina
8
1
9
Sovetskaya st., 14, Yaroslavl, 150000, Russia
,
us
Laboratory of biotechnology and applied bioelementology, Yaroslavl State University,
10
2
11
UNESCO), Orenburg State University, Pobedy Ave. 13, Orenburg 460352, Russia
12
3
13
Zemlyanoy Val St. 46, Moscow 105064, Russia
14
4
15
460000, Russia
16
5
17
Agency, Bekhtereva str. 1, St. Petersburg 192019, Russia
an
Institute of Bioelementology (Russian Satellite Centre of Trace Element – Institute for
M
Russian Society of Trace Elements in Medicine, ANO “Centre for Biotic Medicine”,
te
d
Department of Biochemistry, Orenburg State Medical University, Sovetskaya St., 6, Orenburg,
Federal State Scientific Institution “Institute of Toxicology”, Federal Medico-Biological
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18
3,5
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7
3,5
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, Vasily A. Demidov 3, Yulia N. Lobanova 3, Andrei R. Grabeklis
19
Corresponding author
20
Alexey A. Tinkov, MD, PhD
21
Laboratory of biotechnology and applied bioelementology, Yaroslavl State University,
22
Sovetskaya st., 14, Yaroslavl, 150000, Russia
23
Tel: +7-961-937-81-98
24
E-mail: [email protected]
25 26
1 Page 1 of 11
Abstract
2
A total of 5908 occupationally non-exposed adults (4384 women and 1524 men) living in
3
Moscow and Moscow region were involved in the current investigation. Hair Al, As, Be, Bi, Cd,
4
Hg, Li, Ni, Pb, Sn, Sr content was estimated by inductively-coupled plasma mass spectrometry
5
using NexION 300D. Men are characterized by significantly higher hair Al, As, Cd, Hg, Li, and
6
Pb content. At the same time, hair levels of Bi, Ni, Sn, and Sr were significantly higher in
7
women. Consequently, the reference ranges were estimated for male, female, and general cohort
8
as coverage intervals in accordance with IUPAC recommendations.
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Key words: inductively coupled plasma mass spectrometry; trace elements; hair; reference
11
ranges; coverage intervals; IUPAC recommendations
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1. Introduction
14
Heavy metal exposure has been shown to be associated with a number of diseases (Järup, 2003)
15
and its monitoring is an essential part of environmental and health care (Wolterbeek, 2002). Hair
16
has been widely used as a bioindicator of human exposure to heavy metals (Bencko, 1995). It
17
has been shown that hair may successfully reflect environmental and occupational exposure to
18
lead (Sen, 1996), mercury (Kruzikova et al., 2009), aluminium (Yokel, 1982) and combination of
19
multiple metals (Wang and Fu, 2009). At the same time, a number of side factors like
20
demography, lifestyle and geography (Christensen, 1995) may affect hair trace element content.
21
Despite the presence of a number of studies indicating reference ranges of hair metal content, the
22
existing data are inconsistent (Mikulewicz et al., 2013). Therefore, the primary aim of the current
23
study is estimation of reference ranges of hair toxic trace elements content in adult Russian
24
population.
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2. Materials and methods 2 Page 2 of 11
A total of 5908 occupationally non-exposed adults (4384 women and 1524 men) aged from 20 to
2
60 years and living in Moscow and Moscow region were involved in current investigation. The
3
investigation has been carried out in accordance with the principles of the Declaration of
4
Helsinki for studies involving humans and was approved by the Local Ethics Committee. All
5
examinees gave their informed consent prior to the inclusion in the study.
6
The participants were selected for the study with the use of the following exclusion criteria: i)
7
occupational exposure to toxic metals; ii) smoking (both present and former smokers); iii) acute
8
inflammatory diseases; iv) endocrine disorders; v) metallic implants; vi) pregnancy and lactation;
9
vii) vegetarian diet, viii) alcohol abuse.
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Proximal parts of occipital scalp hair (0.1 gram) were collected. Briefly, hair samples were
11
washed with acetone and then rinsed thrice with deionized water (Zhao et al., 2012). After
12
washing hair samples were dried at 60°C on air with subsequent microwave degradation. Briefly,
13
0.05 g of hair samples were introduced into Teflon tubes and added with concentrated HNO3.
14
Digestion was performed in a Berghof speedwave four system during 20 minutes at 170-180°C.
15
The obtained solutions were added with distilled deionized water to a final volume of 15 ml.
16
Hair toxic trace element content (Al, As, Be, Bi, Cd, Hg, Li, Ni, Pb, Sn, Sr) was estimated by
17
inductively-coupled plasma mass spectrometry with NexION 300D (PerkinElmer Inc., Shelton,
18
CT 06484, USA) using Dynamic Reaction Cell technology removing the majority of
19
interferences with minimal loss of analyte sensitivity and equipped with ESI SC-2 DX4
20
autosampler (Elemental Scientific Inc., Omaha, NE 68122, USA).
21
Manufacturer’s specifications were used for ICP-MS system preparation. Calibration was
22
performed using standards containing 0.5, 5, 10, and 50 μg/l ultra-trace elements prepared from
23
Universal Data Acquisition Standards Kit (PerkinElmer Inc., Shelton, CT 06484, USA) by
24
dilution with distilled deionized water acidified with 1% HNO3. An internal online
25
standardization using yttrium-89 isotope was also performed. Internal standard containing 10
26
μg/l yttrium was prepared from Yttrium (Y) Pure Single-Element Standard (PerkinElmer Inc.,
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Shelton, CT 06484, USA) on a matrix containing 8% 1-butanol (Merck KGaA), 0.8% Triton X-
2
100 (Sigma-Aldrich, Co.), 0.02% tetramethylammonium hydroxide (Alfa-Aesar, Ward Hill, MA
3
01835 USA), and 0.02% ethylenediaminetetraacetic acid (Sigma-Aldrich, Co). For laboratory
4
quality control the certified reference material of human hair GBW09101 (Shanghai Institute of
5
Nuclear Research, Shanghai, China) was used.
6
All analyses were performed in accredited clinic-diagnostic laboratory of autonomous non-profit
7
organization “Centre for Biotic Medicine” (Moscow, Russia), International Union of Pure and
8
Applied Chemistry (IUPAC) company associate.
9
The obtained data were treated with Statistica 11 software (Statsoft, USA). Data distribution was
10
evaluated by the Shapiro-Wilk test. As the distribution was not Gaussian, all variables were log
11
transformed to approximate a normal distribution. Natural logarithms were used for further
12
statistical analyses. One-way ANOVA with Fisher’s LSD post hoc test were used for group
13
comparisons. The values were considered significantly different at p < 0.05.
14
The reference values of hair toxic trace elements content were based on calculation of the 0.95
15
coverage intervals with 0.95 confidence intervals for the upper and lower limits in accordance
16
with IUPAC recommendations (Poulsen et al., 1997).
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3. Results and discussion
19
The descriptive statistics of the male and female cohorts (Table 1) indicates that age of the
20
examinees was not statistically different between the gender groups (p = 0.245). Hair trace
21
elements analysis in men and women (Table 1) indicated that the hair level of certain trace
22
elements is sex-related. In particular, men are characterized by significantly higher hair
23
aluminium (p < 0.001), arsenic (p < 0.001), cadmium (p < 0.001), lithium (p = 0.001), mercury
24
(p = 0.002), and lead (p < 0.001) content. At the same time, hair levels of bismuth (p < 0.001),
25
nickel (p < 0.001), tin (p < 0.001), and strontium (p < 0.001) were significantly higher in women
26
in comparison to the respective values in men. No significant difference was found between men 4 Page 4 of 11
and women in hair beryllium content (p = 0.222). Hypothetically, significantly increased hair
2
mercury, lead, arsenic, and aluminium content in males may result from increased outdoor
3
activity in men. The obtained data is in agreement with earlier studies indicating sex-related
4
difference in hair cadmium, lead (DiPietro et al., 1989), arsenic (Wolfsperger et al., 1994),
5
mercury (Shimomura et al., 1980), nickel (Michalak et al., 2012), aluminium (Chojnacka et al.,
6
2010), and strontium (Dongarra et al., 2011). At the same time, our research data on hair tin
7
content are in contrast to the study of Chojnacka et al. (2010), who have failed to detect a
8
significant gender difference in hair Sn levels. Consequently, sex-specific reference ranges
9
should be used while using hair as an indicator of environmental exposure.
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Reference ranges calculated as coverage intervals for hair toxic element content in a males,
11
females and general cohort are provided in Table 2. The obtained data on adults’ hair metal
12
content reference ranges are generally comparable with earlier published studies performed in
13
Sweden (Rodushkin and Axelsson, 2000), France (Goullé et al., 2005), and Poland (Chojnacka et
14
al., 2010). At the same time, the most significant difference between our reference ranges and the
15
recently published ones were observed in the case of hair cadmium, lead, mercury, and strontium
16
content. It is notable that in the studied cohort hair cadmium and lead content was significantly
17
lower than the respective values postulated in all three previous studies (Rodushkin and
18
Axelsson, 2000; Goullé et al., 2005; Chojnacka et al., 2010). The opposite situation was
19
observed in the case of hair mercury and strontium. In particular, our reference values were
20
higher than the reference values obtained for hair levels of these metals by Rodushkin and
21
Axelsson, 2000, Goullé et al., 2005, and Chojnacka et al., 2010. It should be noted that our
22
recent data is generally comparable with our earlier published reference values obtained by
23
means of inductively coupled plasma atomic emission spectrometry (Skalny, 2003). The
24
observed difference in reference ranges of hair trace element content may be a consequence of a
25
number of factors, like geographical location, climate, and the overall development of industry in
26
the selected location (Christensen, 1995). Moreover, all four studies involved examinees of
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5 Page 5 of 11
different ages that may also have a significant influence on hair trace elements content
2
(Ambeskovic et al., 2013). Finally, various statistical approaches to estimation of the reference
3
ranges may also result in some contradictions.
4
Generally, the reference ranges for hair toxic trace element in male, female, and general samples
5
have been estimated. Consistency of the obtained data with previously published studies, a large
6
number of samples examined, and the use of Dynamic Reaction Cell technology during analysis
7
indicate high quality of the data obtained. The estimated reference ranges may be used in
8
environmental risk assessment (Tamburo et al., 2011).
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Conclusions
11
The present study has demonstrated that men are characterized by significantly higher hair Al,
12
As, Cd, Hg, Li, and Pb content. Hair levels of Bi, Ni, Sn, and Sr were significantly higher in
13
women. Consequently, the reference ranges calculated as coverage intervals in accordance with
14
IUPAC recommendations are different for males, females, and the general cohort.
15
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Acknowledgements
17
The current research is supported by Russian Ministry of Education and Science within project
18
No. 2014/258-544. The authors would like to thank the anonymous reviewers for their helpful
19
comments and suggestions that significantly improved the manuscript quality.
20
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Conflict of interest
22
The authors declare that there are no conflicts of interest
23 24
References
6 Page 6 of 11
1
1. Ambeskovic, M., Fuchs, E., Beaumier, P., Gerken, M., Metz, G.A., 2013. Hair trace
2
elementary profiles in aging rodents and primates: links to altered cell homeodynamics
3
and disease. Biogerontology 14, 557-567.
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pollutants in occupational and environmental settings. Toxicology 101, 29-39.
3. Chojnacka, K., Zielińska, A., Górecka, H., Dobrzański, Z., Górecki, H., 2010. Reference values for hair minerals of Polish students. Environ. Toxicol. Pharmacol. 29, 314-9.
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2. Bencko, V., 1995. Use of human hair as a biomarker in the assessment of exposure to
4. Christensen, J.M., 1995 Human exposure to toxic metals: factors influencing
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4
interpretation of biomonitoring results. Sci. Total Environ. 166, 89-135. 5. DiPietro, E.S., Phillips, D.L., Paschal, D.C., Neese, J.W., 1989. Determination of trace
11
elements in human hair. Reference intervals for 28 elements in nonoccupationally
12
exposed adults in the US and effects of hair treatments. Biol. Trace Elem. Res. 22, 83-
13
100.
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6. Dongarrà, G., Lombardo, M., Tamburo, E., Varrica, D., Cibella, F., Cuttitta, G., 2011.
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Concentration and reference interval of trace elements in human hair from students living
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in Palermo, Sicily (Italy). Environ. Toxicol. Pharmacol. 32, 27-34.
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7. Goullé, J.P., Mahieu, L., Castermant, J., Neveu, N., Bonneau, L., Lainé, G., Bouige, D., Lacroix, C., 2005. Metal and metalloid multi-elementary ICP-MS validation in whole blood, plasma, urine and hair. Reference values. Forensic Sci. Int. 153, 39-44.
8. Järup, L., 2003. Hazards of heavy metal contamination. Br. Med. Bull. 68, 167-82. 9. Kruzikova, K., Kensova, R., Blahova, J., Harustiakova, D., Svobodova, Z., 2009. Using human hair as an indicator for exposure to mercury. Neuro Endocrinol. Lett. 30, 177-81.
23
10. Michalak, I., Mikulewicz, M., Chojnacka, K., Wołowiec, P., Saeid, A., Górecki, H.,
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2012. Exposure to nickel by hair mineral analysis. Environ. Toxicol. Pharmacol. 34,
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727-34.
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11. Mikulewicz, M., Chojnacka, K., Gedrange, T., Górecki, H., 2013. Reference values of elements in human hair: a systematic review. Environ. Toxicol. Pharmacol. 36, 1077-86. 12. Poulsen, O.M., Holst, E., Christensen, J.M., 1997. Calculation and application of
4
coverage intervals for biological reference values. Pure Appl. Chem. 69, 1601-1611.
5
13. Rodushkin, I., Axelsson, M.D., 2000. Application of double focusing sector field ICP-MS
6
for multielemental characterization of human hair and nails. Part II. A study of the
7
inhabitants of northern Sweden. Sci. Total. Environ. 262, 21-36.
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Lead Exposure. J. Hum. Ecol. 7, 133-141.
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14. Sen, J., 1996. Human Scalp Hair As An Indicator of Environmental Lead Pollution and
15. Shimomura, S., Kimura, A., Nakagawa, H., Takao, M.., 1980. Mercury levels in human
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hair and sex factors. Environ. Res. 22, 22-30.
16. Skalny, A. V., 2003. Reference values of chemical elements concentration in hair,
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obtained by means of ICP-AES method in ANO Centre for Biotic Medicine. Trace.
14
Elem. Med. 4, 55-56.
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17. Tamburo, E., Dongarrà, G., Varrica, D., D’Andrea, D., 2011. Trace elements in hair of
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urban schoolboys: a diagnostic tool in environmental risk assessment. Geophysical
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Research Abstracts 13, 1157
18. Wang, T., Fu, J., Wang, Y., Liao, C., Tao, Y., Jiang, G., 2009. Use of scalp hair as indicator of human exposure to heavy metals in an electronic waste recycling area. Environ. Pollut. 157, 2445-51.
19. Wolfsperger, M., Hauser, G., Gössler, W., Schlagenhaufen, C., 1994. Heavy metals in
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human hair samples from Austria and Italy: influence of sex and smoking habits. Sci.
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Total Environ. 156, 235-42.
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20. Wolterbeek, B., 2002. Biomonitoring of trace element air pollution: principles, possibilities and perspectives. Environ. Pollut. 120, 11-21.
8 Page 8 of 11
1 2
21. Yokel, R.A., 1982. Hair as an indicator of excessive aluminum exposure. Clin. Chem. 28, 662-5. 22. Zhao, L.J., Ren, T., Zhong, R.G., 2012. Determination of lead in human hair by high
4
resolution continuum source graphite furnace atomic absorption spectrometry with
5
microwave digestion and solid sampling. Analyt. Lett. 45, 2467-81.
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Table 1
Men (n = 1524)
Women (n = 4384)
Parameter Mean
SD
Median P5
P95
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P95
Mean
General sample (n = 5908) SD
Median P5
P95
Mean
SD
37.026
22.575 56.306 37.914 10.306
36.314
22.969 55.894 37.560 10.198
36.506
22.844 56.008 37.651 8.646
Al, μg/g
5.704
1.926
21.436 7.876
8.439
4.876
1.742
16.020 6.609
8.694
6.936
1.785
16.958 6.936
0.110
As, μg/g
0.046
0.019
0.213
0.074
0.149
0.021
0.006
0.097
0.035
0.091
0.045
0.007
0.135
0.045
0.026
Be, μg/g
0.002
0.000
0.014
0.003
0.008
0.002
0.000
0.010
0.003
0.030
0.003
0.000
0.011
0.003
1.931
Bi, μg/g
0.036
0.007
0.342
0.187
1.917
0.043
0.007
0.735
0.251
1.937
0.234
0.007
0.674
0.234
0.309
Cd, μg/g
0.015
0.004
0.168
0.063
0.594
0.011
0.003
0.070
0.024
0.074
0.034
0.003
0.090
0.034
1.003
Hg, μg/g
0.566
0.076
2.744
0.891
1.086
0.498
0.112
2.002
0.734
0.969
0.775
0.106
2.212
0.775
0.115
Li, μg/g
0.013
0.005
0.070
0.026
0.167
0.010
0.005
0.063
0.023
0.091
0.024
0.005
0.065
0.024
1.057
Ni, μg/g
0.198
0.078
0.809
0.303
0.504
0.265
0.091
1.167
0.451
1.188
0.413
0.087
1.057
0.413
14.943
Pb, μg/g
0.501
0.121
4.689
2.528
29.289
0.295
0.083
1.462
0.532
1.408
1.046
0.088
2.142
1.046
2.568
Sn, μg/g
0.088
0.028
0.402
0.212
2.938
0.133
0.025
2.760
0.647
2.416
0.535
0.026
2.151
0.535
18.468
Sr, μg/g
1.119
0.313
6.624
2.102
4.097
3.990
0.683
25.370 8.696
21.157
6.935
0.455
20.319 6.935
ep te
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Age, years
Ac c
Median P5
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Table 1. Descriptive statistics of age and hair toxic trace elements content in adult men and women
8.646
P5, P95 – 5 and 95 percentile boundaries; SD – standard deviation;
Page 10 of 11
Table 2
Table 2. Coverage intervals for hair trace elements content in adult Russian population calculated in accordance with IUPAC recommendations. Men (n = 1524)
Women (n = 4384)
General sample (n = 5908)
Al, μg/g
3.355-15.279
2.778-10.545
2.913-11.627
As, μg/g
0.036-0.117
0.008-0.062
0.010-0.078
Be, μg/g
0.000-0.007
0.000-0.005
0.000-0.005
Bi, μg/g
0.017-0.234
0.017-0.336
0.014-0.342
Cd, μg/g
0.009-0.088
0.005-0.042
0.006-0.056
Hg, μg/g
0.130-1.365
0.185-1.094
0.168-1.189
Li, μg/g
0.009-0.039
0.009-0.040
Ni, μg/g
0.132-0.540
0.168-0.779
0.159-0.704
Pb, μg/g
0.291-2.358
0.160-0.917
0.187-1.389
Sn, μg/g
0.051-0.265
0.082-1.158
0.076-1.009
Sr, μg/g
0.650-4.173
1.570-15.181
1.148-10.929
δ
0.008
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0.005
0.009-0.040
0.004
ce pt
δ – coverage uncertainty;
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Metal content
β level of expectation in coverage interval - 0.95;
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γ level of confidence of the coverage uncertainty - 0.95.
Page 11 of 11
