- Email: [email protected]
Ion microprobe analyses of ancient human bone
Beam Intemctiens with Materials&Atoms
ELSEVIER
Nuclear
Instruments and Methods in Physics Research
B 136 138 ( 1998) 3% 333
Ion microprobe analyses of ancient human bone
’
St. Jankuhn a3*,T. Butz a, R.-H. Flagmeyer “, T. Reinert ‘, J. Vogt ‘, B. Barckhausen b, J. Hammer1 b, R. Protsch von Zieten b, D. Grambole ‘, F. Herrmann ‘, K. Bethge d
Abstract
At the Rossendorf nuclear microprobe facility, a beam of protons with MeV energy was used to analyse ancient human bones of the Merowingian period (63th century AD). Emitted X-rays were detected to determine the elemental composition of the bones and to estimate the influence of the burial environment on the elemental content of the skeletons. In cross sections of human femora. a different behaviour of the radial distributions of the main and trace elements like P, Ca. Mn. Fe, Zn. Br. and Sr was observed using lateral-resolved FPIXE. This result indicates post 0 1998 Elsevier mortem mineral exchange processes and diagenetic alteration during burial of bone tissue in soil. Science B.V. Kr~~ordu:
Microprobe;
PIXE; Ancient
human
bone; Elemental
1. Introduction In previous studies of ancient human bone, we have determined bone mineral density (BMD) [I] and chemical composition [2]. For a quantitative interpretation of the data, however, one must consider post mortem mechanisms of diagenesis originating from soil environment. Pate et al. already
*Corresponding author. Fax: +49 341 97 32497; e-mail: jankuhn@,rz.uni-leipzigde: WWW: http://www.uni-leipzig.de/ -nfp/. ’ Supported by the Bundesministerium fiir Bildung. Wissenschaft. Forschung und Technologie. Germany. under grant 03-BU9LEl. 0168-583X/98/$19.00 0 1998 Elsevier Science B.V. All rights reserved. PIISOl68-583X(97)00751-9
profiles
emphasize this problem in [3]: Wlzut happens to horn NS it lw~es the biosphere rid enters the grosphere? Thus, e.g., Lambert et al. have exposed excavated human bone to aqueous solutions containing high concentrations of a single added metal ion in order to examine the extent of introduction of contaminating materials during burial. These authors have established that the major contamination is relatively close to the surface [4]. In agreement with that work, we found out using proton-induced X-ray emission spectroscopy (PIXE) that striking alterations of the elemental distributions in the bone take place within a surface (periosteal) region of about 1 mm thickness [5]. In order to characterize this behaviour
more precisely, we carried out PIXE at a nuclear microprobe (uPIXE) to enhance the lateral resolution.
2. Samples and experimental The skeletons investigated are findings from former Merowingian burial places used between the 6th and 8th century AD. The locations of the excavation places near Bockenheim, Edesheim. and Essingen in Rheinland-Pfalz, Germany, are indicated in the map in Fig. 1. For the determination of the elemental distributions, the sample preparation was performed as follows: From femoral shafts, cross sections of about 1 mm thickness were cut with a diamond saw at about 12 cm, i.e., l/3 the distance apart from the proximal epiphysis (Fig. 2). Altogether, 18 cross-section samples were prepared, six from each excavation place. The uPIXE measurements were performed at the Rossendorf nuclear microprobe at the 3 MV Tandetron accelerator [6]. All experiments were carried out in vacuum with a 3 MeV proton beam of (3 x 3) urn? area. The beam current at the target was 200 pA. The cross sections were scanned
Monnheim
Fig. 1, Map of archaeological excavation rowingian cemeteries near Bockenheim. singen.
places of former MeEdesheim. and Es-
Fig. 2. Cross section of the left femoral shaft of a male Merowingian individuum (skeleton no. 18/l of Essingen) of the age group Adultus I (:I- 30 yr at the time of death). The arrow indicates the scan line on which the elemental distributions shown in Fig. 4 were recorded.
radially from the outer edge (Periost) in the direction to the middle of the bone (Endost) over a distance of 1.55 mm at maximum, with 128 points for each line scan. These scans were repeated successively 120 times in order to reduce the sample heating caused by the ion beam within the irradiation time of 3 h per sample. In this scan mode, the preset charge per step was 100 pC so that a total charge of 12 nC was collected per point. In order to detect the characteristic X-rays emitted from the target. a Si(Li) detector made by EC&G 80 mm’ active area, AEx = 180 eV energy resolution at EY = 5.9 keV -- was placed 46 mm from the samples at 0 = 120” backward angle. In front of the detector. a 150 urn Mylar foil was used as a filter to limit the count rate to about 2 kHz in order to suppress the intensity of the lines of the most abundant matrix elements (P and Ca) as well as to avoid pile-up effects. Fig. 3 shows a typical PIXE spectrum of the dense tissue (Substantia corticalis) of a bone cross section recorded on a fixed spot (point mode) but otherwise under the same experimental conditions described above. The uPIXE spectra were analysed by means of a standard peak-fit program-code which extracts the peak areas of the element regions of interest after removing the background.
__._---. --_ _. ~~. _
St. Junk&
331
et ul. I Nucl. Insir. and Mrrh. in Phys. Rex B 136-138 (1998) 329-333
t”““““““’
P
6x10'
4x10' 2x101.. 6~10~
. .
, , ,
, ,
: Ca
6x10' 4x10' 2x16-
*Qwv@w--*
6oooEnergy E,(keV)
Fig. 3. PIXE spectrum recorded section shown in Fig. 2.
on a single point of the cross
: : : ! : : : : : : : : : : : Mn
6000. 4000
I
r
3. Results and discussion
Fig. 4 shows a series of line-scan profiles representing selected elemental distributions of a femoral cross section. Different concentration profiles with the following characteristic features were observed. (i) The X-ray yields of phosphorus, stron(not shown) have an tium, and bromine approximately constant level over the whole scanned range. (ii) In the direction from periost to endost, the concentrations of calcium and iron decrease rapidly within the first 100 urn to constant levels. (iii) Manganese is enriched at certain areas in the bone. (iv) The zinc concentration decreases less rapidly than those of Ca and Fe, extends more deeply into the bone, and local Zn accumulations are more numerous and broader but less intense than those of Mn. There are several biochemical (A) and geochemical (B) processes which may explain the observed elemental profiles. In the case of the transition-metal zinc, such processes are: (A.i) Bone mineralization. Zinc plays an important role in the growth of new skeletal tissue and is localized in areas of bone formation [7]. (A.ii) Meraholisnz. Zinc is also involved in the general metabolism of humans so that relations may exist between dietary intake [S], diseases [9], or occupational exposure [lo] and the bone mineral composition. (B.i) LISfSusion. Zinc penetrates into the periosteal surface of the dead bone from the surrounding burial matrix and causes metal-ion exchange comparable to Mg, Sr, and Pb [l 1,4]. (B.ii) Microbial activity. Microorganisms carry
0.2
0.4
0.6
0.6
1.0
1.2
14
16
Distance from the surface (mm)
Fig. 4. Elemental distributions of P, Ca. Mn. Fe, Zn, and Sr obtained by scanning the sample seen in Fig. 7.
zinc into the skeleton as a product of their metabolism [ 121. The distances between points of higher Zn concentration observed as local maxima in the distributions were determined to be in the range of 80-140 urn (cf. Fig. 5). This is the typical radius of the osteons which form the bone tissue [lo]. An association with lines of arrested growth can not be excluded, although, this is a phenomenon mainly observed in infantile bone [13]. Obviously, biochemical processes (A) are mainly responsible for the local enrichments.
332
St. Junkuhn cf al. I Nucl. Instr. and Meth. in Phys. Rex B 136-138 (199X) 329-333
500 400
and associated soil will help to clarify this behaviour of the concentration profiles mentioned above. Relationships of Zn with other elements were not observed. Thus, the same situation exists for manganese: its concentration profile is not correlated with other elements in the whole scanned area. Mn is known to cause less exchange because only voids and defects in the bone tissue are available to it [4].
Bo351 MaleIRighUAdultusll
MaleIRighUAdultusll
4. Conclusions
Es675 Female/RightfAdultusI
0
0.2
0.4
0.6
0.6
1.0
1.2
1.4
I .6
Distance from the surface (mm)
Fig. 5. Zinc Bockenheim,
distribution patterns of selected Edesheim, and Essingen.
samples
from
The total length scale for the decrease in Zn concentration is in the range of 400-800 pm. This could be related to a diffusion depth of the element which penetrates from the burial environment into the skeleton. It is believed that this diffusion-like decrease has geochemical (B) rather than biochemical reasons. At first sight, this depth varies over the total number of samples which were analysed, and it seems relatively independent on the burial place. In general, Zn is strongly bound to the bone tissue, and it is not possible to remove Zn by applying different solvents [7]. Further investigations on bone
and outlook
We used the PPIXE technique to get radial profiles of major and trace elements in a variety of ancient human bones. Among the investigated element distributions, the Zn profiles are the most interesting ones. A possible explanation for these characteristic profiles is that Zn enrichment in the centres of bone formation is superimposed onto a diffusion profile of Zn from the environment into the bone. Further experiments should allow to distinguish between the biochemical and the geochemical origin of the elemental alteration. Therefore, we will continue with the analysis of associated soil found aside the excavated bones to reveal a relationship between the elemental content inside and outside the bone by comparing skeletons from different burial sites. For this purpose, a nuclear microprobe is a powerful technique which could provide better understanding of bones at three stages: the initial calcification stage (mineralization, bone formation); during life (bone remodelling, disease, accident, increasing age, diet, occupational and environmental exposure); and finally the post mortem diagenesis (ionic substitution, secondary mineralization, fossilization).
References 111 J. Hammerl. Ph.D. Thesis. Univ. Frankfurt a.M.. 1990. VI St. Jankuhn. T. Butz. R.-H. Flagmeyer, T. Reinert. J. Vogt, J. Hammerl. R. Protsch von Zieten. M. Wolf. H. Baumann. K. Bethge. I. Symietz. in: J.L. Duggdn. I.L. Morgan (Eds.), CP392. Appl. of Accelerators in Res. and Ind.. AIP Press, New York, 1997. p. 575. [31 F.D. Pate, J.T. Hutton, K. Norrish, Appl. Geochem. 4 (1989) 303.
St. Jankuhn et ul. I Ntd.
Instr. und Meth. in Phys. Res. B 136-138 (IY9X) 329-333
[4] J.B. Lambert. S.V. Simpson, S.G. Weiner, J.E. Buikstra, J.
Archaeol. Sci. 12 (1985) 85. [S] St. Jankuhn. H. Baumann, K. Bethge. T. Butz, R.-H. Flagmeyer, J. Hammerl. R. Protsch von Zieten, T. Reinert. 1. Symietz. J. Vogt. M. Wolf. in: D. Dirksen. G.v. Bally (Eds.). Series of the Int. Sot. on Optics within Life Sciences, Vol. iv, Springer, Berlin, 1997, p. 137. [6] F. Herrmann. D. Grambole. Nucl. Instr. and Meth. B 104 (1995) 26. [7] S.B. Doty. K.W. Jones, H.W. Krdner. R.E. Shroy. A.L. Hanson, Nucl. Instr. and Meth. 181 (1981) 159.
333
[8] L.L. Klepinger, Ann. Rev. Anthropol. 13 (1984) 75. [9] J.D. Robertson. D.L. Samudralwar, W.R. Markesbery. Nucl. Instr. and Meth. B 64 (1992) 553. [lo] U. Lindh. D. Brune, G. Nordberg, P.-O. Wester, Sci. Tot. Environ. 16 (1980) 109. [I l] T.A. Elliott. G.W. Grime. Nucl. Instr. and Meth. B 77 (1993) 537. [12] G. Grupe. H. Piepenbrink. Appl. Geochem. 4 (1989) 293. [13] D. Grambole. F. Herrmann, B. Herrmann. Nucl. Instr. and Meth. B 109/110 (1996) 667.
ELSEVIER
Nuclear
Instruments and Methods in Physics Research
B 136 138 ( 1998) 3% 333
Ion microprobe analyses of ancient human bone
’
St. Jankuhn a3*,T. Butz a, R.-H. Flagmeyer “, T. Reinert ‘, J. Vogt ‘, B. Barckhausen b, J. Hammer1 b, R. Protsch von Zieten b, D. Grambole ‘, F. Herrmann ‘, K. Bethge d
Abstract
At the Rossendorf nuclear microprobe facility, a beam of protons with MeV energy was used to analyse ancient human bones of the Merowingian period (63th century AD). Emitted X-rays were detected to determine the elemental composition of the bones and to estimate the influence of the burial environment on the elemental content of the skeletons. In cross sections of human femora. a different behaviour of the radial distributions of the main and trace elements like P, Ca. Mn. Fe, Zn. Br. and Sr was observed using lateral-resolved FPIXE. This result indicates post 0 1998 Elsevier mortem mineral exchange processes and diagenetic alteration during burial of bone tissue in soil. Science B.V. Kr~~ordu:
Microprobe;
PIXE; Ancient
human
bone; Elemental
1. Introduction In previous studies of ancient human bone, we have determined bone mineral density (BMD) [I] and chemical composition [2]. For a quantitative interpretation of the data, however, one must consider post mortem mechanisms of diagenesis originating from soil environment. Pate et al. already
*Corresponding author. Fax: +49 341 97 32497; e-mail: jankuhn@,rz.uni-leipzigde: WWW: http://www.uni-leipzig.de/ -nfp/. ’ Supported by the Bundesministerium fiir Bildung. Wissenschaft. Forschung und Technologie. Germany. under grant 03-BU9LEl. 0168-583X/98/$19.00 0 1998 Elsevier Science B.V. All rights reserved. PIISOl68-583X(97)00751-9
profiles
emphasize this problem in [3]: Wlzut happens to horn NS it lw~es the biosphere rid enters the grosphere? Thus, e.g., Lambert et al. have exposed excavated human bone to aqueous solutions containing high concentrations of a single added metal ion in order to examine the extent of introduction of contaminating materials during burial. These authors have established that the major contamination is relatively close to the surface [4]. In agreement with that work, we found out using proton-induced X-ray emission spectroscopy (PIXE) that striking alterations of the elemental distributions in the bone take place within a surface (periosteal) region of about 1 mm thickness [5]. In order to characterize this behaviour
more precisely, we carried out PIXE at a nuclear microprobe (uPIXE) to enhance the lateral resolution.
2. Samples and experimental The skeletons investigated are findings from former Merowingian burial places used between the 6th and 8th century AD. The locations of the excavation places near Bockenheim, Edesheim. and Essingen in Rheinland-Pfalz, Germany, are indicated in the map in Fig. 1. For the determination of the elemental distributions, the sample preparation was performed as follows: From femoral shafts, cross sections of about 1 mm thickness were cut with a diamond saw at about 12 cm, i.e., l/3 the distance apart from the proximal epiphysis (Fig. 2). Altogether, 18 cross-section samples were prepared, six from each excavation place. The uPIXE measurements were performed at the Rossendorf nuclear microprobe at the 3 MV Tandetron accelerator [6]. All experiments were carried out in vacuum with a 3 MeV proton beam of (3 x 3) urn? area. The beam current at the target was 200 pA. The cross sections were scanned
Monnheim
Fig. 1, Map of archaeological excavation rowingian cemeteries near Bockenheim. singen.
places of former MeEdesheim. and Es-
Fig. 2. Cross section of the left femoral shaft of a male Merowingian individuum (skeleton no. 18/l of Essingen) of the age group Adultus I (:I- 30 yr at the time of death). The arrow indicates the scan line on which the elemental distributions shown in Fig. 4 were recorded.
radially from the outer edge (Periost) in the direction to the middle of the bone (Endost) over a distance of 1.55 mm at maximum, with 128 points for each line scan. These scans were repeated successively 120 times in order to reduce the sample heating caused by the ion beam within the irradiation time of 3 h per sample. In this scan mode, the preset charge per step was 100 pC so that a total charge of 12 nC was collected per point. In order to detect the characteristic X-rays emitted from the target. a Si(Li) detector made by EC&G 80 mm’ active area, AEx = 180 eV energy resolution at EY = 5.9 keV -- was placed 46 mm from the samples at 0 = 120” backward angle. In front of the detector. a 150 urn Mylar foil was used as a filter to limit the count rate to about 2 kHz in order to suppress the intensity of the lines of the most abundant matrix elements (P and Ca) as well as to avoid pile-up effects. Fig. 3 shows a typical PIXE spectrum of the dense tissue (Substantia corticalis) of a bone cross section recorded on a fixed spot (point mode) but otherwise under the same experimental conditions described above. The uPIXE spectra were analysed by means of a standard peak-fit program-code which extracts the peak areas of the element regions of interest after removing the background.
__._---. --_ _. ~~. _
St. Junk&
331
et ul. I Nucl. Insir. and Mrrh. in Phys. Rex B 136-138 (1998) 329-333
t”““““““’
P
6x10'
4x10' 2x101.. 6~10~
. .
, , ,
, ,
: Ca
6x10' 4x10' 2x16-
*Qwv@w--*
6oooEnergy E,(keV)
Fig. 3. PIXE spectrum recorded section shown in Fig. 2.
on a single point of the cross
: : : ! : : : : : : : : : : : Mn
6000. 4000
I
r
3. Results and discussion
Fig. 4 shows a series of line-scan profiles representing selected elemental distributions of a femoral cross section. Different concentration profiles with the following characteristic features were observed. (i) The X-ray yields of phosphorus, stron(not shown) have an tium, and bromine approximately constant level over the whole scanned range. (ii) In the direction from periost to endost, the concentrations of calcium and iron decrease rapidly within the first 100 urn to constant levels. (iii) Manganese is enriched at certain areas in the bone. (iv) The zinc concentration decreases less rapidly than those of Ca and Fe, extends more deeply into the bone, and local Zn accumulations are more numerous and broader but less intense than those of Mn. There are several biochemical (A) and geochemical (B) processes which may explain the observed elemental profiles. In the case of the transition-metal zinc, such processes are: (A.i) Bone mineralization. Zinc plays an important role in the growth of new skeletal tissue and is localized in areas of bone formation [7]. (A.ii) Meraholisnz. Zinc is also involved in the general metabolism of humans so that relations may exist between dietary intake [S], diseases [9], or occupational exposure [lo] and the bone mineral composition. (B.i) LISfSusion. Zinc penetrates into the periosteal surface of the dead bone from the surrounding burial matrix and causes metal-ion exchange comparable to Mg, Sr, and Pb [l 1,4]. (B.ii) Microbial activity. Microorganisms carry
0.2
0.4
0.6
0.6
1.0
1.2
14
16
Distance from the surface (mm)
Fig. 4. Elemental distributions of P, Ca. Mn. Fe, Zn, and Sr obtained by scanning the sample seen in Fig. 7.
zinc into the skeleton as a product of their metabolism [ 121. The distances between points of higher Zn concentration observed as local maxima in the distributions were determined to be in the range of 80-140 urn (cf. Fig. 5). This is the typical radius of the osteons which form the bone tissue [lo]. An association with lines of arrested growth can not be excluded, although, this is a phenomenon mainly observed in infantile bone [13]. Obviously, biochemical processes (A) are mainly responsible for the local enrichments.
332
St. Junkuhn cf al. I Nucl. Instr. and Meth. in Phys. Rex B 136-138 (199X) 329-333
500 400
and associated soil will help to clarify this behaviour of the concentration profiles mentioned above. Relationships of Zn with other elements were not observed. Thus, the same situation exists for manganese: its concentration profile is not correlated with other elements in the whole scanned area. Mn is known to cause less exchange because only voids and defects in the bone tissue are available to it [4].
Bo351 MaleIRighUAdultusll
MaleIRighUAdultusll
4. Conclusions
Es675 Female/RightfAdultusI
0
0.2
0.4
0.6
0.6
1.0
1.2
1.4
I .6
Distance from the surface (mm)
Fig. 5. Zinc Bockenheim,
distribution patterns of selected Edesheim, and Essingen.
samples
from
The total length scale for the decrease in Zn concentration is in the range of 400-800 pm. This could be related to a diffusion depth of the element which penetrates from the burial environment into the skeleton. It is believed that this diffusion-like decrease has geochemical (B) rather than biochemical reasons. At first sight, this depth varies over the total number of samples which were analysed, and it seems relatively independent on the burial place. In general, Zn is strongly bound to the bone tissue, and it is not possible to remove Zn by applying different solvents [7]. Further investigations on bone
and outlook
We used the PPIXE technique to get radial profiles of major and trace elements in a variety of ancient human bones. Among the investigated element distributions, the Zn profiles are the most interesting ones. A possible explanation for these characteristic profiles is that Zn enrichment in the centres of bone formation is superimposed onto a diffusion profile of Zn from the environment into the bone. Further experiments should allow to distinguish between the biochemical and the geochemical origin of the elemental alteration. Therefore, we will continue with the analysis of associated soil found aside the excavated bones to reveal a relationship between the elemental content inside and outside the bone by comparing skeletons from different burial sites. For this purpose, a nuclear microprobe is a powerful technique which could provide better understanding of bones at three stages: the initial calcification stage (mineralization, bone formation); during life (bone remodelling, disease, accident, increasing age, diet, occupational and environmental exposure); and finally the post mortem diagenesis (ionic substitution, secondary mineralization, fossilization).
References 111 J. Hammerl. Ph.D. Thesis. Univ. Frankfurt a.M.. 1990. VI St. Jankuhn. T. Butz. R.-H. Flagmeyer, T. Reinert. J. Vogt, J. Hammerl. R. Protsch von Zieten. M. Wolf. H. Baumann. K. Bethge. I. Symietz. in: J.L. Duggdn. I.L. Morgan (Eds.), CP392. Appl. of Accelerators in Res. and Ind.. AIP Press, New York, 1997. p. 575. [31 F.D. Pate, J.T. Hutton, K. Norrish, Appl. Geochem. 4 (1989) 303.
St. Jankuhn et ul. I Ntd.
Instr. und Meth. in Phys. Res. B 136-138 (IY9X) 329-333
[4] J.B. Lambert. S.V. Simpson, S.G. Weiner, J.E. Buikstra, J.
Archaeol. Sci. 12 (1985) 85. [S] St. Jankuhn. H. Baumann, K. Bethge. T. Butz, R.-H. Flagmeyer, J. Hammerl. R. Protsch von Zieten, T. Reinert. 1. Symietz. J. Vogt. M. Wolf. in: D. Dirksen. G.v. Bally (Eds.). Series of the Int. Sot. on Optics within Life Sciences, Vol. iv, Springer, Berlin, 1997, p. 137. [6] F. Herrmann. D. Grambole. Nucl. Instr. and Meth. B 104 (1995) 26. [7] S.B. Doty. K.W. Jones, H.W. Krdner. R.E. Shroy. A.L. Hanson, Nucl. Instr. and Meth. 181 (1981) 159.
333
[8] L.L. Klepinger, Ann. Rev. Anthropol. 13 (1984) 75. [9] J.D. Robertson. D.L. Samudralwar, W.R. Markesbery. Nucl. Instr. and Meth. B 64 (1992) 553. [lo] U. Lindh. D. Brune, G. Nordberg, P.-O. Wester, Sci. Tot. Environ. 16 (1980) 109. [I l] T.A. Elliott. G.W. Grime. Nucl. Instr. and Meth. B 77 (1993) 537. [12] G. Grupe. H. Piepenbrink. Appl. Geochem. 4 (1989) 293. [13] D. Grambole. F. Herrmann, B. Herrmann. Nucl. Instr. and Meth. B 109/110 (1996) 667.