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Trace element contamination in blood-collecting devices
-1 Inlrm~rrr~mri Jmrnd
4 Nuduur Mrdrt me md Biology, Vol 6, pp Pergamon Press Ltd 1979 Prmted m Breat Bntam
X7
case of zinc. Use of disposable syringes to collect blood samples seems to avoid this contamination.
to 21 I
Introduction
Trace Element Contamination in Blood-collecting Devices
THE past few years have witnessed an ever increasing use of the proton-induced X-ray emission (PIXE) method for quantitative multi-elemental determinations in biological samples.“) Many of the important trace elements are found in these specimens at very low concentrations (nanograms or even less). It is clear that a quantitative analysis of such small quantities could be greatly affected by the ever present risk of contamination during the sample handling procedures. Thus it is important to evaluate as precisely as possible all eventual elemental contaminations coming from the accessories used in the sample collection, handling, and storage procedures.“’ In this work we have determined by PIXE techniques the elements present in these accessories and their influence in the trace element analysis of plasma.
R. LECOMTE,’P. PARADIS,’S. MONARO,’ M. BARRETTE,~ G. LAM~UREUX’and H. A. MBNARD~ ‘Laboratoire de Physique Nucleaire, Universite de Montreal, Montreal, Quebec, Canada, 2Departement de Medecine Nucleaire et Radiobiologie, Faculte de Medecine, Universite de Sherbrooke, Sherbrooke, Quebec, Canada, and ‘Unite de Maladie Rhumatismale, Departement Medecine, Faculte de Medecine. Universite de Sherbrooke, Sherbrooke. Quebec, Canada (Rrceiced
_____-
in ProgressIF
Work
de
de
15 April 1979) Experimental
We have used proton-induced X-ray emission (PIXE) methods to determine the trace-element contamination from devices commonly used for blood collection. After a qualitative evaluation of the elemental composition of the devices, we have measured the contamination in de-ionized water and plasma samples following various experimental procedures. We have found that the Vacutainer tubes are the main source of contamination, but their effect in plasma elemental analysis is significant only in the
Procedures and Results
The blood-collecting devices acting as possible agents of contamination are needles, disposable syringes, collecting tubes (Vacutainers), their stoppers, and infusion sets.* A list of tested Vacutainer tubes which, together with their stoppers, are recognized as the main source of contaminationt3 ” is presented in Table 1. First, we have obtained a complete profile of the elements present in the materials mentioned above. This was effected by bombarding fragments of these devices with a 3 MeV proton beam from the Van de Graaff accelerator of the Universite de Montreal. No quantitative results could be obtained employing this procedure since our analysis method relies on the use of thin targetst doped with an internal standard.@.‘) However, elemental concentrations were determined relatively to Zn which is always present (in different amounts) in these devices. The results on the elemental composition of the Vacutainer stoppers are presented in Table 2. The same procedure was employed in the case ot the stainless steel needles and plastic materials form-
* Vacutainer tubes from Becton, Dickinson & Co. of Canada Ltd, Mississauga, Ontario. Disposable syringes from Jelco Laboratories, Rareton, New Jersey 08869. Minicath-21 infusion sets from Deseret Pharmaceutical Co., Sandy, Utah 84070. i The word “target” is employed in the technical jargon of the nuclear physicists or accelerator people. It describes here the sample (as blood serum or plasma) in its final form and ready to be bombarded by a proton beam. Details on the plasma sample preparation can be found in Ref. (6).
TARLE 1. List of tested “Vacutainer”
tubes
Tube No.
Tube vol. (ml)
3200KA 3200NT 3200 LS3200 3273PS
10 10 10 10 3
32oOQS
10
Stopper lubrication
Interior
Green Pink Red Yellow Gray
Silicone Glycerine Silicone Silicone Glycerine
None None Silicone Silicone None
Lavender
Silicone
None
Stopper color
coating
Additive
-
-. 207
Sodium heparin None None None Potassium oxalate and sodium fluoride. EDTA (K,) and sorbate
Work
208 TABLE
2.
Relative elemental composition tainer stoppers
in progress
of Vacu-
Stopper color
Ti
Cr
Fe
Zn
Green Pink Red Yellow Gray Lavender
0.11 0.94 0.25 0.88 * 0.30
1.6 0.004 ~
0.03 0.17 0.29 0.03
--
0.04
1.0 1.0 1.0 1.0 1.0 1.0
*The concentration of this element could not be measured in the gray stopper since the Ti K-ray peak is masked by the very intense L-ray peak of barium which is observed only in this stopper.
ing the walls of the disposable syringes and infusion sets.t The X-ray spectra showed the presence of Fe, Cr and Ni in the needles and of Zn and Ti in the plastics. tA small amount of iron is present in some plastics. A special case is the black plastic of disposable syringes which, in addition to zinc, contains barium and traces of iron and strontium. On the contrary, no trace element heavier than potassium is detected in the disposable material used to prepare the targets (test tubes and pipets from Becton, Dickinson & Co., Falcon plastics, Oxnord, California 93030).
7000 t
After the presence of possible trace element contaminants in the collecting devices was established, their eventual contaminating effects on the plasma specimens were looked for and determined quantitatively. This was carried out by using the following procedure (the “standard” manipulation): 1 ml of plasma or de-ionized water was introduced into pre-opened Vacutainer tubes. The tubes were sealed again, attached to a multi-purpose rotor and shaken for 1 min. The liquid was then left at rest in contact with the stopper for 30 min and subsequently shaken again for another 1 min. The purpose of carrying out some tests with the de-ionized water is simply to determine the ultimate detectability limits of the elemental contamination of the specimen. In fact, the de-ionized water does not contain any heavy elements. Furthermore, in comparison with biological fluids, the absence of an organic matrix in the de-ionized water targets produces a much less significant background in the low-energy end of the X-ray spectra. These two factors increase the detection sensitivity by 1 order of magnitude for elements lighter than copper. An X-ray spectrum from a plasma target (treated with the standard manipulation described above) is presented in Fig. 1. The results extracted from different Vacutainer tubes (see Table 1) are presented in Table 3 (for de-ionized water) and Table 4 (for plasma). The results shown in Table 3 are representative of the sensitivity which can be achieved with our experimental techniques and analysis methods.@!‘) The
(a)
L
CHANNEL
NUMBER
FIG. 1. Characteristic X-ray spectrum from plasma after the standard test (see text) with the green-stoppered tube (a) and reference spectrum of the plasma pool (b). The zinc contribution of the Vacutainer tube is evident in (a).
+ * + + * k
0.1 1.3 0.1 0.3 950 100
Z
2.3 1.2 1.2 1.7 18.1 9.6
+ * & * * +
Ca 0.3 0.3 0.1 0.1 0.3$ 1.5$
0.11 0.03 0.02 0.03 0.23 0.23 + 0.01 f 0.01 _t 0.01 _+ 0.03 + 0.03
f 0.04
Ti
contamination
Cr
of de-ionized
0.06 0.11 0.05 0.07 0.26 0.33
31 36 63 29 37 475 200
72 63 71 68 69 73 76
k + _t + _t + f
Ca 7 7 12 5 8 14t lit
< 0.05 < 0.08 < 0.08 <0.06 < 0.06
Cr
Ti
of plasma
concentration
1.12 1.16 1.26 1.21 1.27 1.8 1.26
f + ) f k k +
Fe
contained
<0.004 0.012 * 0.009 0.005 f 0.003 <0.006 0.06 &- 0.04 0.06 + 0.04
0.09 0.23 0.18 0.14 0.08 0.5 0.11
0.04 0.05 0.02 0.02 < 0.3
+ 0.01 * 0.07 + 0.01 * 0.02 0.02 + 0.24
cut
1 SD)*
Ni
f + + + + ) _t tubes have to be compared
1.83 1.77 1.76 1.83 1.81 1.79 2.06
cu
tubes (mg/l. f 1 SD)*
0.04 * 0.02 0.04 + 0.02 <0.04 0.06 t_ 0.03 <0.04 0.09 2 0.03 0.09 * 0.03
in Vacutainer
* The concentration values are determined from a series of 5 tested tubes. The results for different obtained from 10 measures on the pool of plasma used to perform all tests. t The Ca analysis could be perturbed by the very intense potassium peaks $ The Cu analysis could be perturbed by the very intense zinc peaks.
+ + + + + & +
Pool Green Pink Red Yellow Gray Lavender
301 253 302 285 280 2430 1630
K
TABLE 4. Trace element
0.01 0.10 0.04 0.06 0.03 0.06
Nit
tubes (/lg per tube f
1.3 1.6 2.2 3.4 1.2 53
k 0.6 _+ 0.8 + 0.3 _t 0.8 * 1.2 * 4
Zn
0.87 4.3 5.2 5.2 7.2 3.4 51
2.72 2.61 2.72 2.70 2.70 2.65 2.69
+ k + k k + +
Br 0.10 0.15 0.13 0.09 0.08 0.03 0.16
< 0.008 to.01 <0.006 <0.005 <0.04 < 0.05
Br
values (first line)
&- 0.03 * 0.3 + 0.8 & 0.8 + 0.6 + 0.6 * 11 with the reference
0.05 0.06 0.09 0.02 0.02 0.10 0.181
Zn
and lot-to-lot variations limit the precision of such results. are non-negligible for the very thin de-ionized water targets
+ + + + + +
Fet
water from Vacutainer
contamination values are determined from a series of 3 tested tubes. Tube-to-tube Ni and Cu are present in the formvar backing of the targets and their contributions Ca analysis could be perturbed by the very intense potassium peaks. Cu analysis could be perturbed by the very intense zinc peaks.
2.1 1.7 0.5 0.7 4100 1600
Tube type
* The i Fe, $ The $ The
Green Pink Red Yellow Gray Lavender
Tube type
TARLF 3.Trace element
210
Work
detection limits (expressed in pg per tube) are: Ca (0.02) Cr (OOOS), Fe (0.003), Cu and Zn (0.002), Br (0.005) and MO (0.02). In the case of Pb, we could detect its L X-rays corresponding to an amount of 0.01 pg per tube. These results demonstrate the contamination of water by K, Ca, Ti and Zn. The minute quantities of Fe, Ni and Cu are due to the formvar used as backing for the water targets. The use of tubes with gray and lavender stoppers is certainly not advised since they contain a large quantity of additives with chelate properties which strongly contaminate the samples. From an inspection of the data shown in Table 4, it can be inferred immediately that only the concentration of zinc in the plasma samples is affected significantly by contamination. The situation is not as clear for Ni and Ti, since more sensitive techniques than those employed here would be needed to obtain conclusive results for these trace elements. It should be remarked that the so-called standard manipulation does not really reproduce the actual blood-taking situation. Indeed, this standard manipulation was devised with the purpose of emphasizing the possibility of trace element contamination in the plasma samples. The introduction of a number of precautions could probably reduce the trace element contamination. To check this important point, a series of tests have been carried out following different procedures. These tests (numbered 1, 2, 3 and 4) and the results on the Zn contamination are presented in Table 5 (only zinc is included in this table, since no significant contamination from the other trace elements could be detected). One can notice that there is a decrease in the Zn contamination with the tests 1, 2 and 3. Such a decrease however, is not sufficient to allow a quantitative measurement of this element in plasma. Finally, tests on the other blood-collecting devices such as disposable syringes and infusion sets were performed. The tests on the disposable syringes were carried out by piping in and out, several times, 1 ml of de-ionized water through the needle of the syringe. No significant element contamination through leachTARLE 5. Complementary
Test number
ing could be detected in the water samples with the exception of zinc whose maximum concentration was measured at 0.3~cg per syringe. When plasma was used, the contamination from zinc increased its plasma level by approximately 20”;,. This indicates that using a full syringe (10 ml) of plasma in the normal drawing situation will lead to a very small zinc contamination in the tinal samples, The tests carried out with the infusion sets did not yield very clear results. Their contamination effects are smaller than those due to the Vacutainer tubes with which they are always employed in the clinical blood-collecting procedures.
Conclusion
The results obtained in this work indicate that the Vacutainer tubes are generally safe when used for trace element determination in plasma or serum samples. Only in the case of blood collected for zinc determination do these tubes fail. since the specimens show large contaminations even when the blooddrawing procedures are carried out with great care. Probably the zinc contamination may be avoided with the use of a disposable syringe of Jelco type. Furthermore, it has been proven that the other accessories used with the Vacutainer tubes in the bloodcollecting procedures can be considered as chemically inert in trace element studies. As a conclusion of this work, we wish to compare the present results with those obtained from previous similar studies. Our calcium data from the de-ionized water compare very well with those of PRAGAY rt d(3) (These workers have investigated four different types of tubes, including those with the red and pink stoppers.) The agreement with the values obtained by for calcium is more haphazard. Some FOSTER et ~1.‘~’ of their results compare well with ours whereas some other data indicate a much larger contamination (up to 14pg per tube). This is due, most probably, to the different manipulations (which are not described) employed by these workers and to the use of different extracting agents. HFLMAN et ~1.‘~’have investigated
tests on zinc contamination
Filling method Via a needle through stopper of unopened tubes Pipets, opened tubes Pipets, opened tubes
Table 4f Poo&
in progress
Plasma volume (ml)
1 10
I
Pipets, opened tubes
1
Pipets, opened
1
tubes
by green-stoppered
Vacutainer
Manipulation
tubes
Results* (mgil. i: 1 SD)
Standard+
1.80 f 0.15 (4)
Standard Only five inversions of the tubes Standard followed by a storage of 3 weeks Standard
1.28 k 0.13(2) 1.77 & 0.14(4) 16 + 5(2)
4.3 f 0.3 (5) 0.87 & 0.03 (10)
* From series of tested tubes whose number is given in parenthesis. Only the results for the tubes with green stoppers are shown here since they are representative for the other tubes (see Table 4). t The standard manipulation is described in the text. $ Excerpted from Table 4. 4 Obtained from the plasma pool used as reference.
711
Work in progress the contamination for a group of 9 elements and found a positive contamination only for Mg, Ca, Zn and Pb. No comparison can be done for Mg since this element is not detectable using the present experimental techniques. 16.‘) At first sight, the agreement between our results and theirs on the contamination from Ca and Zn (2 and 4 pg per tube, respectively) is fairly good. However, these authors claim to have inverted the tubes only five times and this type of manipulation should give smaller contamination values (see Table 51. Furthermore, the amount of lead (0.6 pg per tube) measured as contaminant by these workers was never detected in our X-ray spectra in spite of our technlcally attainable 0.01 pg per tube sensitivity limit. .4clino~~led!~rments~The financial support of the National Research Council and Medical Research Council of Canada is gratefully acknowledged.
References 1,
JOHANSSONS. A. E. and JOHANSXIN T. B. !vucl. Instrum. Meth. 137, 473 (1976). 2, ANAND V. D., WHITE J. M. and NINE H. V. C/in. Chem. 21, 595 (1975). 3, PRAGAY D. A., HOWARD S. F. and CHII.UK~~ M. E. Clin. Chem. 17, 350 (1971). 4, FOSTERL. B., FRINGS C. S., DUNN R. T.. BOWERS G. N., PYRUS J. and DOUMAS B. Clin. (‘hem. 16. 546 (1970). 5, HELMAN E. Z., WALLICK D. K. and R~:IN(X)LII 1. M. C/in. Chem. 17, 61 (1971). 6, BARRETTE M., LAMOUREUX G.. LIBEL. E.. LECOMTER., PARADIS P. and MONARO S. .Yuc/. Instrum. Meth. 134, 189 (1976). 7. LECOMTER., PARADIS P., MONARO S.. B.~RRET~F M., LAMOUREUX G. and M~NARD H. A. Nucl. Instrum. Meth. 150, 289 (1978).
4 Nuduur Mrdrt me md Biology, Vol 6, pp Pergamon Press Ltd 1979 Prmted m Breat Bntam
X7
case of zinc. Use of disposable syringes to collect blood samples seems to avoid this contamination.
to 21 I
Introduction
Trace Element Contamination in Blood-collecting Devices
THE past few years have witnessed an ever increasing use of the proton-induced X-ray emission (PIXE) method for quantitative multi-elemental determinations in biological samples.“) Many of the important trace elements are found in these specimens at very low concentrations (nanograms or even less). It is clear that a quantitative analysis of such small quantities could be greatly affected by the ever present risk of contamination during the sample handling procedures. Thus it is important to evaluate as precisely as possible all eventual elemental contaminations coming from the accessories used in the sample collection, handling, and storage procedures.“’ In this work we have determined by PIXE techniques the elements present in these accessories and their influence in the trace element analysis of plasma.
R. LECOMTE,’P. PARADIS,’S. MONARO,’ M. BARRETTE,~ G. LAM~UREUX’and H. A. MBNARD~ ‘Laboratoire de Physique Nucleaire, Universite de Montreal, Montreal, Quebec, Canada, 2Departement de Medecine Nucleaire et Radiobiologie, Faculte de Medecine, Universite de Sherbrooke, Sherbrooke, Quebec, Canada, and ‘Unite de Maladie Rhumatismale, Departement Medecine, Faculte de Medecine. Universite de Sherbrooke, Sherbrooke. Quebec, Canada (Rrceiced
_____-
in ProgressIF
Work
de
de
15 April 1979) Experimental
We have used proton-induced X-ray emission (PIXE) methods to determine the trace-element contamination from devices commonly used for blood collection. After a qualitative evaluation of the elemental composition of the devices, we have measured the contamination in de-ionized water and plasma samples following various experimental procedures. We have found that the Vacutainer tubes are the main source of contamination, but their effect in plasma elemental analysis is significant only in the
Procedures and Results
The blood-collecting devices acting as possible agents of contamination are needles, disposable syringes, collecting tubes (Vacutainers), their stoppers, and infusion sets.* A list of tested Vacutainer tubes which, together with their stoppers, are recognized as the main source of contaminationt3 ” is presented in Table 1. First, we have obtained a complete profile of the elements present in the materials mentioned above. This was effected by bombarding fragments of these devices with a 3 MeV proton beam from the Van de Graaff accelerator of the Universite de Montreal. No quantitative results could be obtained employing this procedure since our analysis method relies on the use of thin targetst doped with an internal standard.@.‘) However, elemental concentrations were determined relatively to Zn which is always present (in different amounts) in these devices. The results on the elemental composition of the Vacutainer stoppers are presented in Table 2. The same procedure was employed in the case ot the stainless steel needles and plastic materials form-
* Vacutainer tubes from Becton, Dickinson & Co. of Canada Ltd, Mississauga, Ontario. Disposable syringes from Jelco Laboratories, Rareton, New Jersey 08869. Minicath-21 infusion sets from Deseret Pharmaceutical Co., Sandy, Utah 84070. i The word “target” is employed in the technical jargon of the nuclear physicists or accelerator people. It describes here the sample (as blood serum or plasma) in its final form and ready to be bombarded by a proton beam. Details on the plasma sample preparation can be found in Ref. (6).
TARLE 1. List of tested “Vacutainer”
tubes
Tube No.
Tube vol. (ml)
3200KA 3200NT 3200 LS3200 3273PS
10 10 10 10 3
32oOQS
10
Stopper lubrication
Interior
Green Pink Red Yellow Gray
Silicone Glycerine Silicone Silicone Glycerine
None None Silicone Silicone None
Lavender
Silicone
None
Stopper color
coating
Additive
-
-. 207
Sodium heparin None None None Potassium oxalate and sodium fluoride. EDTA (K,) and sorbate
Work
208 TABLE
2.
Relative elemental composition tainer stoppers
in progress
of Vacu-
Stopper color
Ti
Cr
Fe
Zn
Green Pink Red Yellow Gray Lavender
0.11 0.94 0.25 0.88 * 0.30
1.6 0.004 ~
0.03 0.17 0.29 0.03
--
0.04
1.0 1.0 1.0 1.0 1.0 1.0
*The concentration of this element could not be measured in the gray stopper since the Ti K-ray peak is masked by the very intense L-ray peak of barium which is observed only in this stopper.
ing the walls of the disposable syringes and infusion sets.t The X-ray spectra showed the presence of Fe, Cr and Ni in the needles and of Zn and Ti in the plastics. tA small amount of iron is present in some plastics. A special case is the black plastic of disposable syringes which, in addition to zinc, contains barium and traces of iron and strontium. On the contrary, no trace element heavier than potassium is detected in the disposable material used to prepare the targets (test tubes and pipets from Becton, Dickinson & Co., Falcon plastics, Oxnord, California 93030).
7000 t
After the presence of possible trace element contaminants in the collecting devices was established, their eventual contaminating effects on the plasma specimens were looked for and determined quantitatively. This was carried out by using the following procedure (the “standard” manipulation): 1 ml of plasma or de-ionized water was introduced into pre-opened Vacutainer tubes. The tubes were sealed again, attached to a multi-purpose rotor and shaken for 1 min. The liquid was then left at rest in contact with the stopper for 30 min and subsequently shaken again for another 1 min. The purpose of carrying out some tests with the de-ionized water is simply to determine the ultimate detectability limits of the elemental contamination of the specimen. In fact, the de-ionized water does not contain any heavy elements. Furthermore, in comparison with biological fluids, the absence of an organic matrix in the de-ionized water targets produces a much less significant background in the low-energy end of the X-ray spectra. These two factors increase the detection sensitivity by 1 order of magnitude for elements lighter than copper. An X-ray spectrum from a plasma target (treated with the standard manipulation described above) is presented in Fig. 1. The results extracted from different Vacutainer tubes (see Table 1) are presented in Table 3 (for de-ionized water) and Table 4 (for plasma). The results shown in Table 3 are representative of the sensitivity which can be achieved with our experimental techniques and analysis methods.@!‘) The
(a)
L
CHANNEL
NUMBER
FIG. 1. Characteristic X-ray spectrum from plasma after the standard test (see text) with the green-stoppered tube (a) and reference spectrum of the plasma pool (b). The zinc contribution of the Vacutainer tube is evident in (a).
+ * + + * k
0.1 1.3 0.1 0.3 950 100
Z
2.3 1.2 1.2 1.7 18.1 9.6
+ * & * * +
Ca 0.3 0.3 0.1 0.1 0.3$ 1.5$
0.11 0.03 0.02 0.03 0.23 0.23 + 0.01 f 0.01 _t 0.01 _+ 0.03 + 0.03
f 0.04
Ti
contamination
Cr
of de-ionized
0.06 0.11 0.05 0.07 0.26 0.33
31 36 63 29 37 475 200
72 63 71 68 69 73 76
k + _t + _t + f
Ca 7 7 12 5 8 14t lit
< 0.05 < 0.08 < 0.08 <0.06 < 0.06
Cr
Ti
of plasma
concentration
1.12 1.16 1.26 1.21 1.27 1.8 1.26
f + ) f k k +
Fe
contained
<0.004 0.012 * 0.009 0.005 f 0.003 <0.006 0.06 &- 0.04 0.06 + 0.04
0.09 0.23 0.18 0.14 0.08 0.5 0.11
0.04 0.05 0.02 0.02 < 0.3
+ 0.01 * 0.07 + 0.01 * 0.02 0.02 + 0.24
cut
1 SD)*
Ni
f + + + + ) _t tubes have to be compared
1.83 1.77 1.76 1.83 1.81 1.79 2.06
cu
tubes (mg/l. f 1 SD)*
0.04 * 0.02 0.04 + 0.02 <0.04 0.06 t_ 0.03 <0.04 0.09 2 0.03 0.09 * 0.03
in Vacutainer
* The concentration values are determined from a series of 5 tested tubes. The results for different obtained from 10 measures on the pool of plasma used to perform all tests. t The Ca analysis could be perturbed by the very intense potassium peaks $ The Cu analysis could be perturbed by the very intense zinc peaks.
+ + + + + & +
Pool Green Pink Red Yellow Gray Lavender
301 253 302 285 280 2430 1630
K
TABLE 4. Trace element
0.01 0.10 0.04 0.06 0.03 0.06
Nit
tubes (/lg per tube f
1.3 1.6 2.2 3.4 1.2 53
k 0.6 _+ 0.8 + 0.3 _t 0.8 * 1.2 * 4
Zn
0.87 4.3 5.2 5.2 7.2 3.4 51
2.72 2.61 2.72 2.70 2.70 2.65 2.69
+ k + k k + +
Br 0.10 0.15 0.13 0.09 0.08 0.03 0.16
< 0.008 to.01 <0.006 <0.005 <0.04 < 0.05
Br
values (first line)
&- 0.03 * 0.3 + 0.8 & 0.8 + 0.6 + 0.6 * 11 with the reference
0.05 0.06 0.09 0.02 0.02 0.10 0.181
Zn
and lot-to-lot variations limit the precision of such results. are non-negligible for the very thin de-ionized water targets
+ + + + + +
Fet
water from Vacutainer
contamination values are determined from a series of 3 tested tubes. Tube-to-tube Ni and Cu are present in the formvar backing of the targets and their contributions Ca analysis could be perturbed by the very intense potassium peaks. Cu analysis could be perturbed by the very intense zinc peaks.
2.1 1.7 0.5 0.7 4100 1600
Tube type
* The i Fe, $ The $ The
Green Pink Red Yellow Gray Lavender
Tube type
TARLF 3.Trace element
210
Work
detection limits (expressed in pg per tube) are: Ca (0.02) Cr (OOOS), Fe (0.003), Cu and Zn (0.002), Br (0.005) and MO (0.02). In the case of Pb, we could detect its L X-rays corresponding to an amount of 0.01 pg per tube. These results demonstrate the contamination of water by K, Ca, Ti and Zn. The minute quantities of Fe, Ni and Cu are due to the formvar used as backing for the water targets. The use of tubes with gray and lavender stoppers is certainly not advised since they contain a large quantity of additives with chelate properties which strongly contaminate the samples. From an inspection of the data shown in Table 4, it can be inferred immediately that only the concentration of zinc in the plasma samples is affected significantly by contamination. The situation is not as clear for Ni and Ti, since more sensitive techniques than those employed here would be needed to obtain conclusive results for these trace elements. It should be remarked that the so-called standard manipulation does not really reproduce the actual blood-taking situation. Indeed, this standard manipulation was devised with the purpose of emphasizing the possibility of trace element contamination in the plasma samples. The introduction of a number of precautions could probably reduce the trace element contamination. To check this important point, a series of tests have been carried out following different procedures. These tests (numbered 1, 2, 3 and 4) and the results on the Zn contamination are presented in Table 5 (only zinc is included in this table, since no significant contamination from the other trace elements could be detected). One can notice that there is a decrease in the Zn contamination with the tests 1, 2 and 3. Such a decrease however, is not sufficient to allow a quantitative measurement of this element in plasma. Finally, tests on the other blood-collecting devices such as disposable syringes and infusion sets were performed. The tests on the disposable syringes were carried out by piping in and out, several times, 1 ml of de-ionized water through the needle of the syringe. No significant element contamination through leachTARLE 5. Complementary
Test number
ing could be detected in the water samples with the exception of zinc whose maximum concentration was measured at 0.3~cg per syringe. When plasma was used, the contamination from zinc increased its plasma level by approximately 20”;,. This indicates that using a full syringe (10 ml) of plasma in the normal drawing situation will lead to a very small zinc contamination in the tinal samples, The tests carried out with the infusion sets did not yield very clear results. Their contamination effects are smaller than those due to the Vacutainer tubes with which they are always employed in the clinical blood-collecting procedures.
Conclusion
The results obtained in this work indicate that the Vacutainer tubes are generally safe when used for trace element determination in plasma or serum samples. Only in the case of blood collected for zinc determination do these tubes fail. since the specimens show large contaminations even when the blooddrawing procedures are carried out with great care. Probably the zinc contamination may be avoided with the use of a disposable syringe of Jelco type. Furthermore, it has been proven that the other accessories used with the Vacutainer tubes in the bloodcollecting procedures can be considered as chemically inert in trace element studies. As a conclusion of this work, we wish to compare the present results with those obtained from previous similar studies. Our calcium data from the de-ionized water compare very well with those of PRAGAY rt d(3) (These workers have investigated four different types of tubes, including those with the red and pink stoppers.) The agreement with the values obtained by for calcium is more haphazard. Some FOSTER et ~1.‘~’ of their results compare well with ours whereas some other data indicate a much larger contamination (up to 14pg per tube). This is due, most probably, to the different manipulations (which are not described) employed by these workers and to the use of different extracting agents. HFLMAN et ~1.‘~’have investigated
tests on zinc contamination
Filling method Via a needle through stopper of unopened tubes Pipets, opened tubes Pipets, opened tubes
Table 4f Poo&
in progress
Plasma volume (ml)
1 10
I
Pipets, opened tubes
1
Pipets, opened
1
tubes
by green-stoppered
Vacutainer
Manipulation
tubes
Results* (mgil. i: 1 SD)
Standard+
1.80 f 0.15 (4)
Standard Only five inversions of the tubes Standard followed by a storage of 3 weeks Standard
1.28 k 0.13(2) 1.77 & 0.14(4) 16 + 5(2)
4.3 f 0.3 (5) 0.87 & 0.03 (10)
* From series of tested tubes whose number is given in parenthesis. Only the results for the tubes with green stoppers are shown here since they are representative for the other tubes (see Table 4). t The standard manipulation is described in the text. $ Excerpted from Table 4. 4 Obtained from the plasma pool used as reference.
711
Work in progress the contamination for a group of 9 elements and found a positive contamination only for Mg, Ca, Zn and Pb. No comparison can be done for Mg since this element is not detectable using the present experimental techniques. 16.‘) At first sight, the agreement between our results and theirs on the contamination from Ca and Zn (2 and 4 pg per tube, respectively) is fairly good. However, these authors claim to have inverted the tubes only five times and this type of manipulation should give smaller contamination values (see Table 51. Furthermore, the amount of lead (0.6 pg per tube) measured as contaminant by these workers was never detected in our X-ray spectra in spite of our technlcally attainable 0.01 pg per tube sensitivity limit. .4clino~~led!~rments~The financial support of the National Research Council and Medical Research Council of Canada is gratefully acknowledged.
References 1,
JOHANSSONS. A. E. and JOHANSXIN T. B. !vucl. Instrum. Meth. 137, 473 (1976). 2, ANAND V. D., WHITE J. M. and NINE H. V. C/in. Chem. 21, 595 (1975). 3, PRAGAY D. A., HOWARD S. F. and CHII.UK~~ M. E. Clin. Chem. 17, 350 (1971). 4, FOSTERL. B., FRINGS C. S., DUNN R. T.. BOWERS G. N., PYRUS J. and DOUMAS B. Clin. (‘hem. 16. 546 (1970). 5, HELMAN E. Z., WALLICK D. K. and R~:IN(X)LII 1. M. C/in. Chem. 17, 61 (1971). 6, BARRETTE M., LAMOUREUX G.. LIBEL. E.. LECOMTER., PARADIS P. and MONARO S. .Yuc/. Instrum. Meth. 134, 189 (1976). 7. LECOMTER., PARADIS P., MONARO S.. B.~RRET~F M., LAMOUREUX G. and M~NARD H. A. Nucl. Instrum. Meth. 150, 289 (1978).