- Email: [email protected]
Determination of trace elements in three Chrysomela (Coleoptera, Chrysomelidae) species by EDXRF analyses
ARTICLE IN PRESS
Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378 www.elsevier.com/locate/jqsrt
Determination of trace elements in three Chrysomela (Coleoptera, Chrysomelidae) species by EDXRF analyses A. Karabuluta, I˙. Aslanb, R. Dumlupınarc, E. Tiras- og˘lud, G. Budaka, a Physics Department, Faculty of Art and Science, Atatu¨rk University, 25240 Erzurum, Turkey Plant Protection Department, Faculty of Agriculture, Atatu¨rk University, 25240 Erzurum, Turkey c Biology Department, Faculty of Art and Science, Atatu¨rk University, 25240 Erzurum, Turkey d Physics Department, Faculty of Arts and Sciences, Karadeniz Technical University, Trabzon, Turkey b
Received 9 April 2004; received in revised form 3 September 2004; accepted 4 September 2004
Abstract In this investigation, the concentration levels of potassium, calcium, iron and nickel in the three Chrysomela species were measured in the region of Erzurum (Turkey). These concentrations measured by energy dispersive X-ray fluorescence (EDXRF) spectrometry were analyzed. From these results, the difference between K, Ca, and Fe concentrations is not significant, but is significant for Ni concentrations. Therefore, the presence of an association between the determination of species and Ni concentrations levels was observed in the present study. These results are presented and discussed in this paper. r 2004 Elsevier Ltd. All rights reserved. Keywords: Trace element analysis; EDXRF analysis; Chrysomela
1. Introduction Chrysomela collaris Linnaeus, C. populi Linnaeus, and C. saliceti (Weise) are very common in European countries and Turkey and very important pest species on poplar [1,2]. C. populi, a poplar leaf beetle, is an important insect that has emerged as a predominant insect pest of poplars in the Europe, Asia and Turkey and is spreading everywhere more so in the Palearctic region Corresponding author. Tel.:+90 4422314161; fax: +90 4422360948.
E-mail address: [email protected] (G. Budak). 0022-4073/$ - see front matter r 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.jqsrt.2004.09.038
ARTICLE IN PRESS 374
A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
[1–3]. The major pests, C. collaris and C. populi feed on Populus and rarely feed on Salix species, often completely defoliating the cultivated poplar area in the Palearctic region. Generally, C. saliceti feeds on willow than poplar. The widespread occurrence of insecticide resistance has made the poplar leaf beetle rely heavily on conventional broad spectrum insecticides in the past, which are difficult and costly to manage in the poplar production areas. The result has been increased economic losses, due to increased yield loss or insecticide costs. To tackle this problem, extensive effort has been directed toward the development of economical and nonchemical approaches to manage this pest. Elytra of C. populi and C. saliceti are red or rusty. Generally, these species have very similar morphologic characters, but the body of C. populi is bigger than C. saliceti. Therefore, some entomologists confuse these species. The elytra of C. collaris are bronze or dark blue. Element analyses of zooplankton are important for investigating the uptake and transfer of substances along the trophic chain in order to define the role of these organisms in biochemical pathways [4]. There are three important functions (osmotic, structural, and biochemical) of elements in the insect life [1]. (1) The elements contribute in the formation of osmotic balance in tissues and cells. (2) They have a role in the formation of important biomolecules as protein, nucleic acid, and lipid. (3) They participate in the formation of several enzymes by binding to specific proteins as prosthetic group (cofactor) and moreover also in the formation of NADP and ATP, which have important roles in energy metabolism and redox reactions [5]. The advent of commercially available energy dispersive spectrometers for X-ray fluorescence (XRF) measurements has provided an economical and powerful tool for environmental, clinical, chemical, geological and industrial analysis. XRF is a non-destructive, fast, multi-element technique for analyzing the surface layer and determining major, as well minor as some trace elements in thin and thick samples of all sizes and forms. Although X-ray spectrometry measurement is simple for a quantitative study, accurate quantitative measurements often depend on matrix correction procedures which require a large number of standards. One of the major problems posed by geological materials is the sample preparation. Since the beginning of the application of EDXRF technique as an analytical tool, a great deal of research has been done in mineralogical and biological materials by different workers [6–9]. The aim of the present study was the investigation of the metal contents of elytra of C. collaris, C. populi, and C. saliceti in Turkey by using EDXRF analyses. The experimental method and the discussion of the analytical results are given below.
2. Experimental 2.1. Apparatus A block diagram of the counting system is given in Fig. 1. The experimental setup consists of an EDXRF spectrometer, a Si(Li) detector, a sample changer, a high-voltage source, a preamplifier, a linear amplifier, a radioactive source, a sample, an oscilloscope, and a computer-controlled multi-channel analyzer system including the analog-to-digital converter (ADC).
ARTICLE IN PRESS A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
375
Fig. 1. Geometry of experimental setup.
2.2. Sample preparation Samples were irradiated by 59.5 keV photons emitted by an annular 50 mCi 241Am annular radioactive source for Fe and Ni determination and irradiated by 5.9 keV photons emitted by an annular 50 mCi 55Fe radioactive source for K and Ca. The insect samples, adults of C. collaris, C. populi, and C. saliceti, were collected throughout the growing seasons from the poplar trees of Erzurum (Turkey) using a hand sweep net and mouth aspirator. After collection, these adult beetles were extracted from their elytra and these elytra were washed three times in triple distilled water in the laboratory. After drying at 60 1C at a constant weight, the samples were digested with HNO3 and H2O2 in a microwave oven. To make the samples as homogenous as possible, the dry material was ground by hand in a ceramic mortar. The samples were then transferred onto a polyethylene myler film. A circular sample radius of approximately 3 cm was used. The matrix effects and their inaccurate correction are usually the main sources of error in X-ray fluorescence analysis. In the thin samples or samples of intermediate thickness that are studied here, enhancement effects are usually absent. The background count rate, which is mainly due to scatters in the sample matrix, increases with sample thickness, and therefore the detection limit which is extrapolated for thick samples in similar counting conditions is less favorable [10], particularly for elements with low atomic numbers. A sample thickness of less than 0.01 g/cm2 has been chosen as a reasonable compromise between saturation thickness and optimal peak-to-background ratios. 2.3. Measurements The choice of a suitable radioisotope and geometry for a radiation source involves several compromises. Our experience indicates that the best performance is obtained with an annular source configuration, as reported in the earlier papers [11,12]. The spectra were analyzed by using the method developed and described in a previous work [11]. Figs. 2a and b present typically spectra analyzed by using the EDXRF technique.
ARTICLE IN PRESS 376
A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378 500
400
Counts
Peaks of primary radiation 300
Coherent peak (59.5 keV)
FeKα NiKα
200
Compton peak 100
0 0
500
1000
(a)
1500
2000
Channel number 500
Counts
400
300 KKα
200
Ca Kα
100
MnKα
MnKβ
0 0
500
1000 Channel number
(b)
Fig. 2. (a) EDXRF spectra of
241
1500
Am source; (b) EDXRF spectra of
2000 55
Fe source.
Experimental analysis in the present paper was performed by using standard addition methods for the simultaneous determination of K, Ca, Fe, and Ni in C. collaris, C. populi, and C. saliceti. The method involves the addition of known quantities of the analyte to the specimen. If the analyte is presented at low levels and no suitable standards are available, standard addition may prove to be an alternative, especially if the analyst is interested in only one analyte element. The principle is the following: adding a known amount of analyte i ðDW i Þ to the unknown sample gives an increased intensity Ii+DIi. Assuming a linear calibration, the following equations apply: I i ¼ MiW i
(1)
for the original samples and I i þ DI i ¼ M i ðW i þ DW i Þ
(2)
for the sample with the addition. Thus, the method assumes that linear calibration is adequate throughout the range of addition because it assumes that an increase in the concentration of
ARTICLE IN PRESS A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
377
analyte by amount DW i will increase the intensity by M i DW i : These equations can be solved for the weight fraction of element i (Wi). To check the linearity of the calibration, the process can be repeated by adding different amounts of the analyte to the sample and plotting the intensity measured versus the concentrations added. The intercept of the line on the concentration axis equals Wi. The intensities used for the calibration must be corrected for background and line overlap. This method is mainly suitable for the determination of trace and minor concentration levels because the amount of DW i added to the sample must be in proportion to the amount Wi in the sample itself. The extrapolation error can be quite large if the slope of the line is not known accurately. Adding significant amount of additives to the sample, however, may lead to non-linearity, because it alters the matrix effect [13].
3. Results and discussion Fig. 2a and b show the spectra obtained within 1 h. The average concentrations found for different elements in the samples are listed in Table 1. The purpose of the research reported here was to investigate the content of trace elements in the elytra of species of C. collaris, C. populi, and C. saliceti in the same genus. We determined concentrations of K, Ca, Fe, and Ni in the C. collaris, C. populi, and C. saliceti. From these results, we showed that the difference between K, Ca, and Fe concentrations is not significant, but the difference between Ni concentrations is significant. Individuals belonging to the same species and living in different geographical regions can possess some small morphological and biochemical content differences that originate from adaptation to ecological conditions. These differences can be clearly determined in individuals of the same species that belong to the same genus. But, the morphological and biochemical differences between different species belonging to the same genus living in the same habitat are of genetic origin and the food they feed on may have an effect on their biochemical content. Chrysomela species used in this study were collected from a small habitat. Thus, the effect of geographical conditions on biochemical content was highly eliminated. In this case, we can evaluate our findings related to the concentrations of K, Ca, Fe, and Ni elements (Table 1) in two ways. First, it may be due to the reflection of genetic differences on their biochemical contents. Second, it may be due to feeding of these species with different foods. However, in order to understand which idea is true, elemental analysis of many species must be carried out. If it is proved that the first idea is true, EDXRF analysis may be helpful in the Table 1 Concentrations of elements obtained from all species elytra Material
C. collaris C. populi C. saliceti a
Not determined.
Concentration (%) K
Ca
Fe
Ni
0.632370.0316 0.569170.0285 0.596370.0298
0.061870.0031 0.049270.0025 0.071070.0036
0.022970.0012 0.033370.0017 0.032270.0016
0.030270.0015 —a 0.008370.0004
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identification of species which have been doubted. As shown in Table 1, the Ni values are 0.030270.0015, not determined, 0.08370.0004 in the elytra of C. collaris, C. populi, and C. saliceti, respectively. Ni concentration was significantly lower in C. populi compared to C. collaris. Ni concentrations in C. saliceti were not examined because the concentrations were too low and the line intensities were close to the background levels. Therefore, our data confirm that the determination of species may participate in differences between Ni concentrations.
References [1] Aslan I˙, O¨zbek H. Faunistic and systematic studies on the subfamily Chrysomelinae (Coleoptera, Chrysomelidae) in Artvin, Erzincan and Erzurum provinces. Turk J Zool 1999;23:751–67 [in Turkish, with English summary]. [2] Warchalowski A, Chrysomelidae, the leaf-beetles of Europe and the Mediterranean area. Natura optima dux Foundation, Warszawa; 2003. p. 600. [3] Jolivet P, Petitpierre E, Hasiao TH. Biology of Chrysomelidae, Series entomologicia, vol. 42. Dordrecht, MA: Kluwer Academic Publishers; 2003. p. 606. [4] Mages M, Woelfl S, Tumpling W. A method for trace element determination of single Dahpnia specimens using total reflection X-ray fluorescence spectrometry. Spectrochim Acta B 2001;56:2209–17. [5] Kadıog˘lu A, Bitki Fizyolojisi. Trabzon, Turkiye; 1999. p. 35–40 [in Turkish]. [6] Karabulut A, Budak G. Radiosotope X-ray fluorescence analysis of some elements in Colemanite ore. Spectrochim Acta B: At Spectrosc 2000;55:91–5. [7] Karabulut A, Budak G, Polat R, Gu¨rol A, Levent M, S- ahin Y. EDXRF analysis of murgul pyrite ore concentrates. JQSRT 2002;72:741–6. [8] C - evik U, Ergen E, Budak G, Karabulut A, Tiras-og˘lu E, Apaydin G, Kopya AI. Elemental analysis of Ak,caabat tobacco and its ash by EDXRF spectrometry. JQSRT 2003;78:409–15. [9] Gu¨rol A, Ergen E, Karabulut A, Polat R, Altinkaynak K, Budak G. Determinattion of P, S, Cl, K, and Ca in human urinary stones by EDXRF. Instrum Sci Technol 2004;32:69–76. [10] Nguyen TH, Huynh VH. Application of EDXRF to the determination of lead and other trace elements in the body fluids of industrial workers in Vietnam. X-ray Spectrometry 1996;25:3–14. [11] S- ahin Y, Karabulut A, Budak G. A practical method for the analysis of overlapped peaks in energy dispersive X-ray spectra. Appl Spectrosc Rev 1996;31:333–45. [12] Budak G, Karabulut A, Demir L, S- ahin Y. Monte Carlo calculations of XRF intensities in annular arrangements of radioactive source, sample, cylindrical collimator and detector. Appl Spectrosc Rev 1996;31:321–32. [13] Van Grieken RE, Markowicz AA. Handbook of X-ray Spectrometry. Bassel, Hong Kong, New York: Marcel Dekker Inc.; 1993. p. 314 [Chapter 5].
Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378 www.elsevier.com/locate/jqsrt
Determination of trace elements in three Chrysomela (Coleoptera, Chrysomelidae) species by EDXRF analyses A. Karabuluta, I˙. Aslanb, R. Dumlupınarc, E. Tiras- og˘lud, G. Budaka, a Physics Department, Faculty of Art and Science, Atatu¨rk University, 25240 Erzurum, Turkey Plant Protection Department, Faculty of Agriculture, Atatu¨rk University, 25240 Erzurum, Turkey c Biology Department, Faculty of Art and Science, Atatu¨rk University, 25240 Erzurum, Turkey d Physics Department, Faculty of Arts and Sciences, Karadeniz Technical University, Trabzon, Turkey b
Received 9 April 2004; received in revised form 3 September 2004; accepted 4 September 2004
Abstract In this investigation, the concentration levels of potassium, calcium, iron and nickel in the three Chrysomela species were measured in the region of Erzurum (Turkey). These concentrations measured by energy dispersive X-ray fluorescence (EDXRF) spectrometry were analyzed. From these results, the difference between K, Ca, and Fe concentrations is not significant, but is significant for Ni concentrations. Therefore, the presence of an association between the determination of species and Ni concentrations levels was observed in the present study. These results are presented and discussed in this paper. r 2004 Elsevier Ltd. All rights reserved. Keywords: Trace element analysis; EDXRF analysis; Chrysomela
1. Introduction Chrysomela collaris Linnaeus, C. populi Linnaeus, and C. saliceti (Weise) are very common in European countries and Turkey and very important pest species on poplar [1,2]. C. populi, a poplar leaf beetle, is an important insect that has emerged as a predominant insect pest of poplars in the Europe, Asia and Turkey and is spreading everywhere more so in the Palearctic region Corresponding author. Tel.:+90 4422314161; fax: +90 4422360948.
E-mail address: [email protected] (G. Budak). 0022-4073/$ - see front matter r 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.jqsrt.2004.09.038
ARTICLE IN PRESS 374
A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
[1–3]. The major pests, C. collaris and C. populi feed on Populus and rarely feed on Salix species, often completely defoliating the cultivated poplar area in the Palearctic region. Generally, C. saliceti feeds on willow than poplar. The widespread occurrence of insecticide resistance has made the poplar leaf beetle rely heavily on conventional broad spectrum insecticides in the past, which are difficult and costly to manage in the poplar production areas. The result has been increased economic losses, due to increased yield loss or insecticide costs. To tackle this problem, extensive effort has been directed toward the development of economical and nonchemical approaches to manage this pest. Elytra of C. populi and C. saliceti are red or rusty. Generally, these species have very similar morphologic characters, but the body of C. populi is bigger than C. saliceti. Therefore, some entomologists confuse these species. The elytra of C. collaris are bronze or dark blue. Element analyses of zooplankton are important for investigating the uptake and transfer of substances along the trophic chain in order to define the role of these organisms in biochemical pathways [4]. There are three important functions (osmotic, structural, and biochemical) of elements in the insect life [1]. (1) The elements contribute in the formation of osmotic balance in tissues and cells. (2) They have a role in the formation of important biomolecules as protein, nucleic acid, and lipid. (3) They participate in the formation of several enzymes by binding to specific proteins as prosthetic group (cofactor) and moreover also in the formation of NADP and ATP, which have important roles in energy metabolism and redox reactions [5]. The advent of commercially available energy dispersive spectrometers for X-ray fluorescence (XRF) measurements has provided an economical and powerful tool for environmental, clinical, chemical, geological and industrial analysis. XRF is a non-destructive, fast, multi-element technique for analyzing the surface layer and determining major, as well minor as some trace elements in thin and thick samples of all sizes and forms. Although X-ray spectrometry measurement is simple for a quantitative study, accurate quantitative measurements often depend on matrix correction procedures which require a large number of standards. One of the major problems posed by geological materials is the sample preparation. Since the beginning of the application of EDXRF technique as an analytical tool, a great deal of research has been done in mineralogical and biological materials by different workers [6–9]. The aim of the present study was the investigation of the metal contents of elytra of C. collaris, C. populi, and C. saliceti in Turkey by using EDXRF analyses. The experimental method and the discussion of the analytical results are given below.
2. Experimental 2.1. Apparatus A block diagram of the counting system is given in Fig. 1. The experimental setup consists of an EDXRF spectrometer, a Si(Li) detector, a sample changer, a high-voltage source, a preamplifier, a linear amplifier, a radioactive source, a sample, an oscilloscope, and a computer-controlled multi-channel analyzer system including the analog-to-digital converter (ADC).
ARTICLE IN PRESS A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
375
Fig. 1. Geometry of experimental setup.
2.2. Sample preparation Samples were irradiated by 59.5 keV photons emitted by an annular 50 mCi 241Am annular radioactive source for Fe and Ni determination and irradiated by 5.9 keV photons emitted by an annular 50 mCi 55Fe radioactive source for K and Ca. The insect samples, adults of C. collaris, C. populi, and C. saliceti, were collected throughout the growing seasons from the poplar trees of Erzurum (Turkey) using a hand sweep net and mouth aspirator. After collection, these adult beetles were extracted from their elytra and these elytra were washed three times in triple distilled water in the laboratory. After drying at 60 1C at a constant weight, the samples were digested with HNO3 and H2O2 in a microwave oven. To make the samples as homogenous as possible, the dry material was ground by hand in a ceramic mortar. The samples were then transferred onto a polyethylene myler film. A circular sample radius of approximately 3 cm was used. The matrix effects and their inaccurate correction are usually the main sources of error in X-ray fluorescence analysis. In the thin samples or samples of intermediate thickness that are studied here, enhancement effects are usually absent. The background count rate, which is mainly due to scatters in the sample matrix, increases with sample thickness, and therefore the detection limit which is extrapolated for thick samples in similar counting conditions is less favorable [10], particularly for elements with low atomic numbers. A sample thickness of less than 0.01 g/cm2 has been chosen as a reasonable compromise between saturation thickness and optimal peak-to-background ratios. 2.3. Measurements The choice of a suitable radioisotope and geometry for a radiation source involves several compromises. Our experience indicates that the best performance is obtained with an annular source configuration, as reported in the earlier papers [11,12]. The spectra were analyzed by using the method developed and described in a previous work [11]. Figs. 2a and b present typically spectra analyzed by using the EDXRF technique.
ARTICLE IN PRESS 376
A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378 500
400
Counts
Peaks of primary radiation 300
Coherent peak (59.5 keV)
FeKα NiKα
200
Compton peak 100
0 0
500
1000
(a)
1500
2000
Channel number 500
Counts
400
300 KKα
200
Ca Kα
100
MnKα
MnKβ
0 0
500
1000 Channel number
(b)
Fig. 2. (a) EDXRF spectra of
241
1500
Am source; (b) EDXRF spectra of
2000 55
Fe source.
Experimental analysis in the present paper was performed by using standard addition methods for the simultaneous determination of K, Ca, Fe, and Ni in C. collaris, C. populi, and C. saliceti. The method involves the addition of known quantities of the analyte to the specimen. If the analyte is presented at low levels and no suitable standards are available, standard addition may prove to be an alternative, especially if the analyst is interested in only one analyte element. The principle is the following: adding a known amount of analyte i ðDW i Þ to the unknown sample gives an increased intensity Ii+DIi. Assuming a linear calibration, the following equations apply: I i ¼ MiW i
(1)
for the original samples and I i þ DI i ¼ M i ðW i þ DW i Þ
(2)
for the sample with the addition. Thus, the method assumes that linear calibration is adequate throughout the range of addition because it assumes that an increase in the concentration of
ARTICLE IN PRESS A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
377
analyte by amount DW i will increase the intensity by M i DW i : These equations can be solved for the weight fraction of element i (Wi). To check the linearity of the calibration, the process can be repeated by adding different amounts of the analyte to the sample and plotting the intensity measured versus the concentrations added. The intercept of the line on the concentration axis equals Wi. The intensities used for the calibration must be corrected for background and line overlap. This method is mainly suitable for the determination of trace and minor concentration levels because the amount of DW i added to the sample must be in proportion to the amount Wi in the sample itself. The extrapolation error can be quite large if the slope of the line is not known accurately. Adding significant amount of additives to the sample, however, may lead to non-linearity, because it alters the matrix effect [13].
3. Results and discussion Fig. 2a and b show the spectra obtained within 1 h. The average concentrations found for different elements in the samples are listed in Table 1. The purpose of the research reported here was to investigate the content of trace elements in the elytra of species of C. collaris, C. populi, and C. saliceti in the same genus. We determined concentrations of K, Ca, Fe, and Ni in the C. collaris, C. populi, and C. saliceti. From these results, we showed that the difference between K, Ca, and Fe concentrations is not significant, but the difference between Ni concentrations is significant. Individuals belonging to the same species and living in different geographical regions can possess some small morphological and biochemical content differences that originate from adaptation to ecological conditions. These differences can be clearly determined in individuals of the same species that belong to the same genus. But, the morphological and biochemical differences between different species belonging to the same genus living in the same habitat are of genetic origin and the food they feed on may have an effect on their biochemical content. Chrysomela species used in this study were collected from a small habitat. Thus, the effect of geographical conditions on biochemical content was highly eliminated. In this case, we can evaluate our findings related to the concentrations of K, Ca, Fe, and Ni elements (Table 1) in two ways. First, it may be due to the reflection of genetic differences on their biochemical contents. Second, it may be due to feeding of these species with different foods. However, in order to understand which idea is true, elemental analysis of many species must be carried out. If it is proved that the first idea is true, EDXRF analysis may be helpful in the Table 1 Concentrations of elements obtained from all species elytra Material
C. collaris C. populi C. saliceti a
Not determined.
Concentration (%) K
Ca
Fe
Ni
0.632370.0316 0.569170.0285 0.596370.0298
0.061870.0031 0.049270.0025 0.071070.0036
0.022970.0012 0.033370.0017 0.032270.0016
0.030270.0015 —a 0.008370.0004
ARTICLE IN PRESS 378
A. Karabulut et al. / Journal of Quantitative Spectroscopy & Radiative Transfer 94 (2005) 373–378
identification of species which have been doubted. As shown in Table 1, the Ni values are 0.030270.0015, not determined, 0.08370.0004 in the elytra of C. collaris, C. populi, and C. saliceti, respectively. Ni concentration was significantly lower in C. populi compared to C. collaris. Ni concentrations in C. saliceti were not examined because the concentrations were too low and the line intensities were close to the background levels. Therefore, our data confirm that the determination of species may participate in differences between Ni concentrations.
References [1] Aslan I˙, O¨zbek H. Faunistic and systematic studies on the subfamily Chrysomelinae (Coleoptera, Chrysomelidae) in Artvin, Erzincan and Erzurum provinces. Turk J Zool 1999;23:751–67 [in Turkish, with English summary]. [2] Warchalowski A, Chrysomelidae, the leaf-beetles of Europe and the Mediterranean area. Natura optima dux Foundation, Warszawa; 2003. p. 600. [3] Jolivet P, Petitpierre E, Hasiao TH. Biology of Chrysomelidae, Series entomologicia, vol. 42. Dordrecht, MA: Kluwer Academic Publishers; 2003. p. 606. [4] Mages M, Woelfl S, Tumpling W. A method for trace element determination of single Dahpnia specimens using total reflection X-ray fluorescence spectrometry. Spectrochim Acta B 2001;56:2209–17. [5] Kadıog˘lu A, Bitki Fizyolojisi. Trabzon, Turkiye; 1999. p. 35–40 [in Turkish]. [6] Karabulut A, Budak G. Radiosotope X-ray fluorescence analysis of some elements in Colemanite ore. Spectrochim Acta B: At Spectrosc 2000;55:91–5. [7] Karabulut A, Budak G, Polat R, Gu¨rol A, Levent M, S- ahin Y. EDXRF analysis of murgul pyrite ore concentrates. JQSRT 2002;72:741–6. [8] C - evik U, Ergen E, Budak G, Karabulut A, Tiras-og˘lu E, Apaydin G, Kopya AI. Elemental analysis of Ak,caabat tobacco and its ash by EDXRF spectrometry. JQSRT 2003;78:409–15. [9] Gu¨rol A, Ergen E, Karabulut A, Polat R, Altinkaynak K, Budak G. Determinattion of P, S, Cl, K, and Ca in human urinary stones by EDXRF. Instrum Sci Technol 2004;32:69–76. [10] Nguyen TH, Huynh VH. Application of EDXRF to the determination of lead and other trace elements in the body fluids of industrial workers in Vietnam. X-ray Spectrometry 1996;25:3–14. [11] S- ahin Y, Karabulut A, Budak G. A practical method for the analysis of overlapped peaks in energy dispersive X-ray spectra. Appl Spectrosc Rev 1996;31:333–45. [12] Budak G, Karabulut A, Demir L, S- ahin Y. Monte Carlo calculations of XRF intensities in annular arrangements of radioactive source, sample, cylindrical collimator and detector. Appl Spectrosc Rev 1996;31:321–32. [13] Van Grieken RE, Markowicz AA. Handbook of X-ray Spectrometry. Bassel, Hong Kong, New York: Marcel Dekker Inc.; 1993. p. 314 [Chapter 5].