SHORT COMMUNICATION

Journal of Radioanalytical and Nuclear Chemistry, Vol. 247, No. 1 (2001) 175�178

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W. M. Kwiatek,1 B. Kubica,1 R. Grybos′′ ,2 M. Kros′′niak,3 E. M. Dutkiewicz,1 R. Hajduk1

1 Institute of Nuclear Physics, Cracow, Poland2Department of Chemistry, Jagiellonian University, Poland

3Department of Molecular Biology, Jagiellonian University, Poland

(Received February 21, 2000)

Proton induced X-ray emission (PIXE) and atomic absorption spectroscopy (AAS) were used for vanadium determination in animal tissues. The

vanadium concentration levels were determined in blood, kidneys and livers taken from rats. Two groups of the animals were treated with different

diets. The diet for the first group was supplemented with vanadium compounds while the diet for the second one was assumed to be a �normal�

diet. The second group was treated as control. In order to achieve the best minimum detectable limit (MDL)1 the samples were subject to a special

sample preparation procedure. Blood and kidneys were mineralized with an APDC compound. The mineralization process was performed

according to the procedure described previously.2 The application of PIXE3 is very useful for different types of samples. PIXE measurements were

performed with a proton beam at the Institute of Nuclear Physics in Cracow, Poland while the AAS measurements were done at the Institute of

Molecular Biology, Jagiellonian University, Poland. The concentration levels of vanadium in blood and kidneys are compared and discussed.

There were no significant statistical differences between results of vanadium concentration levels determined by the abovementioned techniques.

The PIXE technique had the advantage over the AAS technique of giving a broad spectrum of trace elements analyzed in a single measurement.

Therefore with the help of sample preparation procedure the application of the PIXE method seems to be suitable for such analyzes.

Introduction

Vanadium is a trace element widely distributed in the

environment. It is a contaminant of many ores, petroleum

oils, coals, and used in the steel and chemical

industries.4 It is present at trace level in various sources

for nutrition such as meat, fish, vegetables, milk, etc.5,6

from which it is taken up by animals and humans.

Vanadium has been shown to produce several important

biological effects in living organisms.7�9 The insulin-

mimetic action of vanadium both in vitro and in vivo has

been demonstrated in numerous studies.8,10�12 Its

therapeutic potential has been shown in clinical studies

with diabetic patients.13,14

It has been estimated that no more than 1% of

vanadium normally taken up with the diet is absorbed.15

The concentration of vanadium (ng/g dry tissue weight)

in tissues of nonsupplemented rats depends on the organ

in the order: brain (2 ng/g) < fat < blood < heart <

muscle < lung < liver < testes < spleen < kidney

(133 ng/g).16 All organs accumulate vanadium in a

dose16 and type of compound manner.17

Vanadium in body fluids and tissues can be determined

by various methods such as: atomic absorption spectrometry

(AAS),18,19 voltametry,20 inductively coupled plasma

atomic emission spectrometry (ICPAES),21 inductively

coupled plasma mass spectrometry (ICPMS),22 neutron

activation analysis (NAA),23 and 48V-radiolabeled tracer.17

Comprehensive overviews on vanadium determination in

biological materials have been published24 and it seems that

PIXE has not been applied yet for this purpose. Therefore,

the authors have applied that technique since it has been

recognized as a multi-elemental analytical method.

Experimental

The experiment was carried out on male Wistar rats,

weighting 150±29 g. Animals were housed in wire cages

in a room at 20±2 °C with natural light-dark cycles. The

animals were allowed free access to commercial pelleted

food and tap water. The rats were divided into control

and vanadium treated (VT) groups. The animals in group

VT received bis (1,10-phenanthroline) oxovanadium

(IV) sulphate trihydrate (VO(phen)2SO4.3H2O)

suspended in 1% methylcellulose by one-time oral dose

of 200 mg/kg of rat. Bis (1,10-phenanthroline)

oxovanadium (IV) sulphate trihydrate was prepared by

the modified method described in the literature.25 Two

days after the administration of vanadyl complex the rats

were exanguinated under light ether anesthesia before

sacrificing. Then the samples of blood, kidney and liver

were taken out from the animals frozen at �20 °C and

dried at 55 °C to constant mass.

Sample preparation procedure

Mineralization procedure: High purity compounds as

HNO3, H2O2, and APDC (ammonium pyrrolidine-

dithiocarbomate) were used. Nitric acid was additionally

purified by means of distillation. Samples were treated

with HNO3, H2O2 for 20 minutes at 1200 °C. When they

were fully mineralized the solution was divided into two

parts. One was analyzed by atomic absorption

spectroscopy (AAS) and APDC compound was added to

the second one in order to form complexes with metals

that exist in the samples. The solution was filtered

0236�5731/2001/USD 17.00 Akadémiai Kiadó, Budapest

© 2001 Akadémiai Kiadó, Budapest Kluwer Academic Publishers, Dordrecht

W. M. KWIATEK et al.: DETERMINATIONOFVANADIUM INANIMALTISSUESBY PIXE ANDAAS

through a Millipore filter GVWP 22. The undiluted

complexes were left on the filter and then the filter was

dried at 50 °C for 1 hour. The prepared filters were the

targets in PIXE analysis for the determination of trace

element concentration.

Normal preparation procedure: Samples were dried

for 24 hours at 50 °C and then powdered in an agate

mortar. Then the samples were pressed into pellets of

10 mm in diameter and 1 mm thick. Such a pellet was

placed on Scotch tape and fixed to an aluminum frame.

Then the frames were put into the chamber and analyzed

by PIXE.

Methods and apparatus

The PIXE method was applied in the Institute of

Nuclear Physics in Cracow. A proton beam of 2.5 MeV

from the Van de Graaff accelerator was used for these

analyses. Both types of samples (filters and pellets) were

analyzed for 15 minutes in order to get good statistics in

the X-ray characteristic spectra. The spectra were

detected with a Si(Li) detector with an energy resolution

of 160 eV for 5.9 keV X-rays. For normalization

backscattered protons were used. All the spectra

(characteristic X-rays and particle spectra) were

registered with an ADCAM system.

The AAS measurements were performed at the

Institute of Molecular Biology, Jagiellonian University.

A Perkin Elmer 5100 ZL apparatus with Zeeman

background correction device was used for vanadium

content determination. The measurements were done by

non-flame absorption spectrometry. The samples were

injected into a graphite tube.

Results and discussion

The results obtained from both techniques are

presented in Tables 1 and 2. Table 1 presents vanadium

concentration levels in samples taken from both rat

groups. The concentrations are given in µg/g dry mass.

In these tables the results from PIXE and AAS are

compared.

As seen in Table 1 there is no statistical difference

between the concentration levels determined with the

two techniques. But the application of PIXE enabled the

multi-elemental determination shown in Table 2. For

determination of trace element concentration levels the

IAEA standards were used. Those standards include:

H-4 - animal muscle, H-8 horse kidney, and A-13 animal

blood.

Table 1. Vanadium concentration levels determined by PIXE and AAS in rat samples (µg/g dry mass)

Sample Vanadium concentration Vanadium concentration

determined by AAS determined by PIXE

Bt 0.195 ± 0.029 0.078 ± 0.019

Vt 1.167 ± 0.187 2.133 ± 0.457

Bn 0.548 ± 0.093 0.457 ± 0.192

Vn 2.787 ± 0.371 2.237 ± 0.095

Bk 0.344 ± 0.043 0.444 ± 0.067

Vk 1.537 ± 0.199 1.490 ± 0.568

Bt � liver from control group of rats,Vt � liver from vanadium treated rats,Bn � kidney from control group of rats,Vn � kidney from vanadium treated rats,Bk � blood from control group of rat,Vk � blood from vanadium treated rats.Estimated errors for AAS are about 15%.

Table 2. Trace element concentration levels determined by PIXE

Sample V Mn Fe Cu Zn Pb Se Br

Bt 0.078 ± 0.019 0.622 ± 0.071 267 ± 45 61.6 ± 6.9 231 ± 30 17.9 ± 2.5 2.0 ± 0.5 0.089 ± 0.010

Vt 2.133 ± 0.457 4.438 ± 0.567 75 ± 15 68.0 ± 8.0 60 ± 15 2.8 ± 3.5 1.9 ± 0.5 0.209 ± 0.040

Bn 0.457 ± 0.192 0.906 ± 0.112 1180 ± 360 324 ± 46 538 ± 55 22.4 ± 2.8 8.4 ± 1.5 0.163 ± 0.035

Vn 2.239 ± 0.095 4.744 ± 0.596 5090 ± 820 80 ± 25 51.5 ± 6.5 69 ± 9.5 4.4 ± 0.9 2.316 ± 0.350

Bk 0.444 ± 0.067 2.723 ± 0.382 2930 ± 330 53 ± 15 16.1 ± 2.2 24.0 ± 4.0 2.6 ± 0.7 0.533 ± 0.200

Vk 1.490 ± 0.568 0.785 ± 0.142 482 ± 95 150 ± 25 36.7 ± 4.8 6.3 ± 0.9 9.8 ± 3.5 0.171 ± 0.035

Bt � liver from control group of rats,Vt � liver from vanadium treated rats,Bn � kidney from control group of rats,Vn � kidney from vanadium treated rats,Bk � blood from control group of rat,Vk � blood from vanadium treated rats.

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W. M. KWIATEK et al.: DETERMINATIONOFVANADIUM INANIMALTISSUESBY PIXE ANDAAS

Figures 1 and 2 show PIXE spectra of kidney and

blood samples. Spectra (a) correspond to samples with

no vanadium treatment while spectra (b) relate to the

samples treated with vanadium compounds. The spectra

do not differ in elemental composition. There is a

significant difference in vanadium peak intensity. Figure

3 shows two spectra of the same blood sample. Spectrum

(a) corresponds to the thick sample while spectrum (b)

corresponds to the sample prepared by the

mineralization procedure. As seen in the case of

spectrum (b), the calcium and potassium peaks are

reduced in comparison to those in spectrum (a). This

reduction is due to the application of APDC.

Conclusions

PIXE analysis is very sensitive to multi-trace element

determinations. The advantage of this technique is its

multi-elemental character at the time while the AAS

technique gives more precise results but each element

has to be determined separately. In order to increase the

sensitivity of PIXE, the mineralization procedure of the

sample is very useful and helps to achieve a better

determination at lower levels.

Fig. 1. The PIXE spectra of kidney sample: a � thick target,

b � thin target that followed mineralization procedure

Fig. 2. The PIXE spectra of blood sample: a � thick target,

b � thin target that followed mineralization procedure

Fig. 3. The PIXE spectra of blood sample: a � thick target,

b � thin target that followed mineralization procedure

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W. M. KWIATEK et al.: DETERMINATIONOFVANADIUM INANIMALTISSUESBY PIXE ANDAAS

*

This work was supported by KBN Grant No. 8 T11E 017 16.

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