Mineral Composition of Human fascia lata

Transcription

1 Biol Trace Elem Res DOI /s y Mineral Composition of Human fascia lata Anna Leśniewicz & Milena Furtak & Wiesław Żyrnicki & Joanna Dawidowicz & Krzysztof Maksymowicz & Sylwia Szotek Received: 21 March 2014 /Accepted: 17 April 2014 # Springer Science+Business Media New York 2014 Abstract The mineral composition of pathologically unchanged human fascia lata was examined here using inductively coupled plasma optical emission spectrometry (ICP- OES) method for the first time. The total concentrations of Ag,Al,B,Ba,Ca,Cd,Co,Cr,Cu,Fe,Mg,Mn,Mo,Ni,P,Pb, Sr, Ti, V and Zn were simultaneously measured in the tissue secured during autopsy. The age-related changes and between-gender differences in mineral composition of the examined tissue were investigated and discussed. Keywords Human fascia lata. Mineral composition. Trace elements. ICP-OES Introduction The human body consists of approximately 60 elements present in different organic and inorganic compounds [1]. It is known that both excess and deficiency of main and trace A. Leśniewicz (*): M. Furtak : W. Żyrnicki Analytical Chemistry Division, Faculty of Chemistry, Wrocław University of Technology, Wybrzeże Wyspiańskiego 27, Wrocław, Poland anna.lesniewicz@pwr.wroc.pl J. Dawidowicz Veterinary Clinic Brynów, Brynowska 25c, Katowice, Poland K. Maksymowicz Faculty of Medicine, Department of Forensic Medicine, Wrocław Medical University, J. Mikulicza-Radeckiego 4, Wrocław, Poland S. Szotek Department of Biomedical Engineering, Mechatronics and Theory of Mechanisms, Faculty of Mechanical Engineering, Wrocław University of Technology, Wybrzeże Łukasiewicza 7/9, Wrocław, Poland elements is influencing proper functioning of the human organism. Therefore, all the investigations concerning elemental composition of human body are important for assessment of element concentration range in healthy organisms (i.e. reference levels) as well as quantity characteristic for pathological stages and metal and non-metal poisonings. Under examinations, carried out since nearly decades, varieties of human body fluids and tissues have been taken. For example, blood [2 4] and serum [5], urine [2, 3], bile [3], brain [3, 6, 7], breast tissue [8], kidney [3], liver [3], lung [3, 9], stomach [3], bones [10], ligaments [11], tendon [12, 13], ureter [14], hair [2], nails [15] andskin[16] have been analysed for determination of the total concentration of elements, recognized as reference level, as well as their variability related to age, environment and state of the organism health. The most of the reported investigations implicate only few elements, usually recognized as essential such as Ca, Cu, Fe, Mg, Se and Zn [4, 5, 11, 14, 16]. Only few research papers have been devoted to multi-elemental analysis of body fluids and tissues [2, 6, 8]. Measurement based on quantification of major and trace elements have been performed using spectroscopic methods mostly, as for example: FAAS [5], GF-AAS [5], HG-AAS [8], inductively coupled plasma optical emission spectrometry (ICP-OES) [4, 6, 11, 14], ICP-MS [2] and INNA [8, 9, 16]. According to our best knowledge, mineral composition of the human fascia lata was not examined with the use of spectroscopic methods allowing to determine simultaneously so many main and trace elements. The human fascia lata the deep fascia of the thigh is a strong and fibrous connective tissue, composed of collagen mainly, surrounding and isolating the muscles, internal organs and other soft structures of the body and providing structural support and protection. Because of its merits, the fascia lata is considered as a unique and extraordinary biomaterial which can be used in

2 Leśniewicz et al. medicine and tissue engineering as an autologous or allogenic dura matter substitute or as a scaffolding material applied in soft tissues repairing [17] orspinesurgery[18, 19]. Therefore, as a part of multidisciplinary investigations leading to extension of our knowledge about chemical, biological and mechanical properties of the human fascia lata, the study of its mineral composition were undertaken. Main and trace elements were determined in the pathologically unchanged human (men and women) fascia lata tissues. Material and Methods Samples The pathologically unchanged human fascia lata samples, the deep fascia of the thigh, were collected during autopsy. This study was approved by the Bioethics Committee of Wrocław Medical University (KB-262/2010). The standard reference materials were used for verification of measurements accuracy and all the applied analytical procedures repeatability. Because of lack of certified reference material of fascia, TORT 2 Lobster Hepatopancreas Standard Reference Material was analysed. Sample Preparation Accurately weighed portion of soft tissue was transferred into a Teflon digestion vessel. Next, 4.5-mL of concentrated nitric acid (Merck KGaA, Darmstad, Germany) was added. Decomposition of samples was carried out in microwave digestion system (Milestone, MLS-1200, MEGA). Six-step programme, with maximum power 650 W, was simultaneously applied to the six samples. After cooling the sample, solutions were quantitatively transferred into a 10-mL volumetric flask and filled up with deionised water to the mark. To ensure high quality of trace metal determination, with each set of minimum three parallel samples, a blank sample was conducted through the complete analytical procedure and used for correction of analytical signals measured by ICP- OES. All reagents used in this study were of analytical grade at least or better. Before the use, all the glassware and plastic bottles were washed with distilled water, cleaned with diluted nitric acid in ultrasonic bath and finally rinsed several times with deionised. All dissolutions and dilutions were performed with deionized water obtained in our laboratory from EASYpure (Barnstead, Thermolyne Corporation, USA) water purification system. All chemicals used in this study were at least of analytical grade and were tested for possible contamination. Measurements The element determinations were performed with the use of atomic emission spectrometry with inductively coupled argon plasma as the excitation source. Multi-elemental measurements were performed for the analysed samples. A Jobin- Yvon 38S spectrometer was equipped with a concentric nebuliser (Burgener Teflon nebuliser) and cyclonic spray chamber. The operating parameters and analytical line wavelengths chosen for the measurements were typical for this kind of determinations. The calibration curve method was used for calibration of instrument. Aqueous standard solutions were prepared by dilution of the ICP multi-element standard (Merck KGaA, Darmstad, Germany) solution. Results Total concentrations of Ag, Al, B, Ba, Ca, Cd, Co, Cr, Cu, Fe, Mg, Mn, Mo, Ni, P, Pb, Sr, Ti, V and Zn were measured in healthy fascial tissue samples collected by pathologist from persons in the age ranging from 19 to 72. Comparison of the results of the elements determinations in soft tissue fascia samples for women and men and taking into account age of the persons is presented in Table 1. Total concentrations of the elements determined in analysed samples are shown as an arithmetic mean and its standard deviation in relation to sample donors sex and age. In all the examined samples, as it could be expected in the case of biological tissues, the highest concentration was observed for P and Ca. The determined P and Ca concentrations varied from 216 to 349 and 233 to 367 μg g 1,respectively. Relatively high content was also noticed for Mg and Zn. For those elements, concentrations were found in the ranges: and μg g 1. Lower concentrations were determined for Al, Cu and Fe. In the case of those metals, contents ranged as follows: , and μgg 1. Contents ranging between 1.0 and 0.5 μgg 1 was noticed for B and Sr. Less than 0.4 μg g 1 of Ag, Ba, Cd, Cr, Mn, Ni Pb and Ti was quantified in the examined samples. None of the examined samples contain detectable amount of Co, Mo and V. The highest barium, calcium, chromium, strontium and zinc contents were observed in fascia lata samples taken from women donors. However, the highest concentration of iron, phosphorus and magnesium was determined in tissues obtained from men, especially young, i.e. at age ranging from 19 to 38, than for woman. The comparison of the element concentrations in samples obtained from women and man donors as well as in samples from people at different age ranges were made using t test at the 95 % level of significance. According to the t test, statistically significant difference between samples taken from men and women occurred only between the

3 Mineral Composition of Human fascia lata Table 1 Concentration of the elements in human fascia (mean value ± standard deviation [μg/g]) and its relation to donors sex and age Element Women Men t calculated ;S a At the age of At the age of t calculated ;S a Ag 0.005± ± ± ± Al 3.67± ± ± ± ; S B 0.18± ± ± ± Ba 0.21± ± ± ± Ca 367±30 233± ; S 264±16 235± ; S Cd 0.011± ± ± ± Co <0.016 <0.008 <0.014 <0.008 Cr 0.32± ± ± ± Cu 0.98± ± ± ± Fe 5.68± ± ± ± Mg 44.9± ± ± ± Mn 0.15± ± ± ± Mo <0.015 <0.015 <0.016 <0.014 Ni 0.37± ± ± ± P 216±10 342± ; S 349±12 308± ; S Pb 0.31± ± ± ± Sr 0.69± ± ; S 0.52± ± Ti 0.035± ± ± ± V <0.009 <0.008 <0.007 <0.009 Zn 13.4± ± ± ± t critical =4.303 (n=3; α=0.05) S significant a Indication of the difference statistical significance means estimated for Ca, P and Sr. Nevertheless, statistically significant difference in elements content in samples achieved from people at the age ranging between 19 and 38 and donors at the age of appeared for Al, Ca and P. For those elements calculated values of t parameter (t calculated ) were higher than the critical value of this test (t critical =4.303). This was specified by character S following t calculated value in Table 1. Content of Al, Ca and P in human fascia lata samples decreases with ageing, similar decrease of Mg, and P content in ureters, ligaments and tendons was also observed [12 14]. However, definite increase of calcium content with the aging was noticed in those samples [12 14]. Relationships between Al, Ca, P and Mg content in fascia samples and the donor s age were estimated by calculation of Pearson s linear correlation coefficient (r). The correlation coefficient values (r) assessed for the investigated pairs of elements, i.e. Al-Ca, Al-Mg, Al-P, Ca-Mg, Ca-P and Mg-P as well as for their concentration and the donors age assessed were found to be lower than 0.2 or close to 0.5 in the case of Ca and P concentration (r= 0.474), what indicates negligible or moderate relationship between those elements concentration in fascia samples taken from people at different age. The smallest variation in mineral concentrations measured in the examined samples from different donors was observed for Mg. Magnesium content varied only by few percent. Not too large differences were noted in Cu, Fe, Ni, P, Pb Ti and Zn contents. The greatest discrepancies in metal concentrations among the examined groups of the samples were noticed for Ag, Ba, Cd, Cr, Mn and Sr. In the case of those elements, concentration differences were larger than 80 % and for Sr differed by nearly three orders of magnitude. Analysis of Standard Reference Material lobster tissue (TORT 2; Lobster Hepatopancreas) was performed for verification of the applied analytical procedures and accuracy. The concentration of all the studied elements was measured in reference material samples mineralised by mixture of nitric acid in closed-vessel system with the aid of microwave energy. The t test was used to discover a statistical significance of the reference occurring between the experimental and certified values at 95 % confidence level. The results of this test with the indication of difference statistical significance (S or NS) and the comparison of experimentally obtained and certified concentration values of are shown in Table 2. For most of the examined elements measured in analysed reference material, a very good agreement between experimental data, i.e. determined total concentrations of the elements analysed here, and certified values was observed, indicating good accuracy of the experimental results. The best

4 Leśniewicz et al. Table 2 Measurement accuracy verification analysis of the standard reference material: Lobster Hepatopancreas (TORT 2) Element Concentrations in μg/g t calculated ;SorNS a Experimental Certified Ag 0.15±0.01 Lack of data Al 31.6±3.7 Lack of data B 4.53±0.12 Lack of data Ba 1.93±0.08 Lack of data Ca 4,650±140 Lack of data Cd 24.9± ± ; NS Co 0.50± ± ; NS Cr 0.76± ± ; NS Cu 98.0± ± ; NS Fe 105± ±13.0 0; NS Mg 1,310±80 Lack of data Mn 12.1± ± ; NS Mo 0.92± ± ; NS Ni 2.26± ± ; NS P 8,140±570 Lack of data Pb 0.33± ± ; NS Sr 43.5± ± ; NS Ti 0.80±0.01 Lack of data V 1.65± ± ; NS Zn 167± ± ; NS t critical =4.303 (n=3; α=0.05) S significant, NS not significant a Indication of the difference statistical significance agreement between determined and certified concentration values was observed for Fe and V, where the recovery equals 99.9 and %, respectively. Very high accuracies were also observed for Co, Cr, Mo and Sr our results differed from the certified values by only a few percent. For Cu, Ni, Pb, and Zn, the differences in concentrations (measured and certified) were found to be satisfactory if one takes into account standard deviation uncertainties of the measurements. Only for Cd and Mn significant differences between values were noticed for those elements determinations recovery was lower than 90 %. The t test indicated that no statistically significant difference occurred between the measured and certified concentration values for Cd, Co, Cr, Cu, Fe, Mn, Mo, Ni, Pb, Sr, V and Zn. For those elements, calculated values of t parameter (t calculated ) were lower than the critical value of this test (t critical ) equal to This was signified by NS in the Table 2. The precision of the measurements of the standard reference material samples was investigated by analysis of three subsamples and for the most of the examined elements the relative standard deviations were found to be lower than 5 %. In the cases of Al and Cr measurements, the relative standard deviation values exceeded 10 %. Final remarks For the first time, the total concentrations of 20 minerals were measured in human fascia lata samples, which seem to be essential for investigation of mineral tissue homeostasis indispensable for proper functioning of all human body cells and tissues. The used microwave digestion technique in connection with ICP-OES is a good method for determination of macroelements as well as microelements important for human organism especially such as Co, Cr, Cu, Fe, Mo, Ni, Pb, Sr, V and Zn. Acknowledgments The work was financed by a statutory activity subsidy from the Polish Ministry of Science and Higher Education for the Faculty of Chemistry of Wrocław University of Technology and by National Science Centre grant no. N N Conflict of Interest interest. References The authors declare that they have no conflict of 1. Nelson DL, Cox MM (2000) Lehninger principles of biochemistry, 3rd edn. W. H. Freeman, New York. ISBN Goullé JP, 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(1): Lech T, Sadlik JK (2011) Zinc in postmortem body tissues and fluids. Biol Trace Elem Res 142: Massadeh A, Gharibeh A, Omari K, Al-Momani I, Alomari A, Tumah H, Hayajneh W (2010) Simultaneous determination of Cd, Pb, Cu, Zn, and Se in human blood of Jordanian smokers by ICP- OES. Biol Trace Elem Res 133:1 11. doi: /s y 5. Kalkan A, Bulut V, Avci S, Celik I, Bingol NK (2002) Trace elements in vital hepatitis. J Trace Elem Med Biol 16: Rahil-Khazen R, Bolann BJ, Ulvik RJ (2002) Correlations of trace element levels within and between different normal autopsy tissues analyzed by inductively coupled plasma atomic emission spectrometry (ICP-AES). BioMetals 15: Krebsa N, Langkammer C, Goessler W, Ropele S, Fazekas F, Yen K, Scheurer E (2014) Assessment of trace elements in human brain using inductively coupled plasma mass spectrometry. J Trace Elem Med Biol 28: Ng KH, Looi LM, Bradley DA (1997) The elemental composition of breast tissue: can this be related to breast particle deposition? J Radioanal Nucl Chem 217(2): Rogero SO, Saiki M, Saldiva PH, Daliberto ML (1994) Determination of trace elements in human lung samples. Biol Trace Elem Res 43 45: Zaichick V, Zaichick S (2009) Instrumental neutron activation analysis of trace element contents in the rib bone of healthy men. J Radioanal Nucl Chem 281:47 52

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