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Atomic fluorescence spectrometry combined with reduction aeration for the determination of mercury in biological samples
Analytica Chimica Acta, 242 (1991) 203-208 Elsevier Science Publishers B.V.. Amsterdam
203
Atomic fluorescence spectrometry combined with reduction aeration for the determination of mercury in biological samples G. Vermeir
*, C. Vandecasteele
’ and R. Dams
Laboratory of Analytical Chemistry University of Gent, Institute for Nuclear Sciences, Proeftuinstraat 86, B-9000 Gent (Bel@n) (Received
31st May 1990)
Abstract A sensitive atomic fluorescence system for the determination of mercury was optimized. The system consists of a continuous-flow mercury vapour generator coupled to a fluorescence spectrometer specific for mercury. A new gas-liquid separator was developed. A detection limit of 0.9 ng Hg 1-l was obtained. The system was combined with a microwave oven for dissolving samples in closed Teflon PFA vessels. Accurate results were obtained for certified reference materials, and biological samples such as urine and hair were analysed. Keywords; Atomic
fluorescence
spectrometry;
Mercury;
Biological
samples
scribed a flow-injection system for the determination of mercury by CVAFS. Bloom and Fitzgerald [9] used Carbotrap preconcentration followed by cryogenic gas chromatography with CVAFS detection for the determination of organomercurial species in gaseous samples. However, the application of AFS was hampered because no commercial system was available until a few years ago. This paper describes the optimization of a commercial system for the analysis of biological samples for mercury by AFS. Moreover, and in contrast to previous work [5,6], the accuracy of the method was assessed by analysing several certified reference materials. Additionally hair and urine samples were analysed and the results were compared with literature values.
In previous papers [1,2] the determination of mercury in biological samples by cold vapour (CV) atomic absorption spectrometry (AAS) was addressed. This required preconcentration on a gold absorber in order to achieve sufficiently low detection limits. Atomic fluorescence spectrometry (AFS) allows a more sensitive determination of mercury than AAS. This method has frequently been applied to the analysis of water samples [3,4]. For several interesting biological samples, direct analysis is possible without any preconcentration [5,6]. Caupeil et al. [5] applied AFS spectrometry to the determination of mercury in fish samples at the mg kg-’ level. Thompson and Godden [6] tested their system on urine and blood samples. Ferrara et al. [7] improved the AFS technique by using a mercury radiofrequency electrodeless discharge lamp as excitation source. Morito et al. [S] de-
EXPERIMENTAL
Reagents ’ Research Director tific Research.
0003-2670/91/$03.50
of the Belgian
National
0 1991 - Elsevier
Fund
All chemicals and water from
for Scien-
Science Publishers
B.V.
were of analytical-reagent grade a Millipore Mini-Q purification
G. VERMEIR
204
system was used throughout. A stock standard solution of mercury (1.000 g 1-l) was prepared by dissolving mercury(I1) oxide in 14 M nitric acid and diluting to, 1 1. Working standard solutions (ng 1-i range) were prepared daily by dilution immediately before use. All standards, blank and sample solutions were stabilized for up to 10 h with 2% (v/v) nitric acid, Tin(I1) chloride solution was prepared by dissolving 20 g of the dihydrate (highest purity, UCB) in SO ml of boiling 12 M hydrochloric acid followed by dilution with 200 ml of water and 600 ml of 1.5 M sulphuric acid. This solution was cooled and purged with nitrogen for 30 min. Dissolution procedure
A lOO-mg amount of solid sample or 10 ml of liquid sample was weighed directly into a Teflon PFA digestion vessel and sub-boiled concentrated nitric acid (1 ml for the solid samples and 5 ml for the liquid samples) was added. Six sealed vessels and a beaker filled with 50 ml of water, to absorb excess of microwave energy, were placed inside a plastic box which was closed and placed in a microwave oven (Amana, Model R.S. 560A). The dissolution programme consisted of three steps: 20% power for 8 rnin, 40% power for 8 min and 60% power for 4 min. After cooling, the digest was
Fig. 1. Schematic
flow diagram
of the system.
transferred to a volumetric 100 ml with water. Mercury
flask
and
ET AL.
diluted
to
determination
Mercury was determined using an AFS system (PS Analytical, Sevenoaks), consisting of a mercury vapour generator, a fluorescence spectrometer and a computer. This system copes with normal laboratory conditions, but it was optimized to lower the detection limits. The continuous-flow mercury vapour generator (Model PSA 10003 hydride generator; PS Analytical) consists (Fig. 1) of a carrier gas supply, a pumping system delivering a continuous flow of tin(I1) chloride solution, blank and sample, a Tpiece where the tin(I1) chloride solution and the blank or the sample are mixed and a gas-liquid separator. The sample and blank streams are switched using a combination of two three-port, two-way miniature Teflon valves as indicated in Fig. 1. Blank and samples are pumped at a flowrate of 7 ml min-’ and tin(I1) chloride solution at a flow-rate of 3 ml mm’. The atomic fluorescence spectrometer (Merlin Model PSA 10023 fluorescence detector; PS Analytical) consists of a high-intensity mercury lamp, an open chimney designed by Thompson and Godden [6] into which the mercury vapour is fed by means of the argon
DETERMINATION
OF MERCURY
IN BIOLOGICAL
205
SAMPLES
carrier gas, an interference filter to isolate the 254-nm resonance line and a photometric detector. The response is displayed digitally as a reading from 0 to ,199 and as a 0-lo-mV analogue output that can be interfaced to a PC or chart recorder. The analogue output is converted to a digital signal by an analogue-to-digital convertor. A Tulip PC records the time-resolved fluorescence signal (one measurement per second during 300 s). Integration is carried out over ca. 260 s (peak-area mode). Correction of the peak area for baseline drift is possible using time windows. The system can also be used in the peak-height mode.
RESULTS
AND
The sensitivity obtained with argon carrier gas was approximately ten times greater than that with nitrogen. This is caused by the quenching of excited mercury atoms by nitrogen molecules. Argon was used in all subsequent measurements. The flow-rate was varied from 200 to 600 ml mm’. In this range 200 ml rnin- ’ was optimum for both peak height and peak area (Fig. 2). When the central tube of the chimney was sheathed with argon, the signal decreased. Therefore, the use of the sheath gas was discontinued. The tin(I1) chloride concentration has a small influence on the signal, but a concentration of ca. 21 g 1-l seems optimum. Peak :
200
1
area
(x100)
Peak height .__~_~
600
200
1
01 0
I_____--_-
100
area
(x1000)
-~~~
Fig. 3. Effect of the sample uptake rate on the signal (mercury standard solution, 10 pg 1-l; calibration range 1).
DISCUSSION
Optimization
600
Peak s”r
200
300
400
L.
500
600
-0 700
Ar flow (ml/mid Fig. 2. Effect of argon flow-rate on the fluorescence signal (10 solution; calibration range 10). 0, Peak pg Hg 1-l standard area; +, peak height.
The signal obtained over 1 h (twelve measurements of 5 min) was stable to within 2%. Longterm variations were also within reasonable limits. The effect of the sample uptake rate on the fluorescence signal is shown in Fig. 3. The line bends from a sample uptake rate of 5 ml mm’. This means that above this sample uptake rate the mercury is not completely removed from the solution. The graph shows that at a sample uptake rate of 7 ml min-‘, which is the rate at which the mercury vapour generator operates, ca. 90% of the mercury present was liberated. To increase the efficiency of volatilization of mercury from the solution, the gas-liquid separator shown in Fig. 4 was devised. A glass sinter was sealed across the end of the gas inlet, causing fine bubbles to be carried through the solution. This increased the solution-gas interface area appreciably. This system increased the signal by 20%, and improved the baseline stability, compared with the original separator shown in Fig. 1. The signal rises to a maximum level once the valves have switched between the blank stream and the sample stream. When the sampling period is finished, the signal returns to the baseline level. It is clear that because of short-term fluctuations of the signal, calculation of the peak area will yield more reproducible results than just using the peak height after a given time. Therefore, the signal is integrated from 0 to 300 s, whereas the sample is introduced from 5 to 95 s.
206
G. VERMEIR
r--
DETECTOR
Ar CARRIER GAS I
r-
\
I
>
Sn Cl2 *SAMPLE I
O2 GLASS
Fig. 4. New gas-liquid
SINTER
separator.
As the recovery of mercury from the standard and sample may differ slightly, because of matrix differences, it is desirable that the recovery of mercury from both the standard and the sample are almost 100%. This was verified in two ways. First, the fraction going to waste was collected for a standard and for a spiked sample and was found to contain less than 5% of the total mercury concentration. Second, the signal obtained was compared with that obtained in a batch system [1,2] after quantitative aeration. It appeared that the present system yields an integrated signal equal to 96.7% with a standard deviation of 2.7% (five determinations) of that obtained with the batch system. The new gas-liquid separator had a better efficiency, but a larger amount of water vapour was produced, which on condensation may produce stray light, resulting in baseline drift. Therefore, a water trap consisting of a quartz tube filled with shredded paper [lo] was placed between the gasliquid separator and the detector. An effect of water vapour was noted only when measuring in the most sensitive range. Mercury has a pronounced tendency to absorb on surfaces, resulting in a memory effect. On
ET AL.
changing from a low-mercury sample (0.4 pg 1-l) to a high-mercury sample (4.0 pg l-i), or vice versa, a representative signal is obtained only after 4-5 analyses. This effect can be reduced by decreasing the surface in contact with the mercury vapour and by adopting a proper measuring sequence. In addition, decreasing the tube length between the mercury vapour generator and the detector from 500 to 160 mm increased the sensitivity by 5%. Effect of organic solvents The presence of 0.01,0.05 or 0.1% (v/v) acetone in the sample did not affect the baseline stability and with a 5 PI-181-i sample gave no reduction in peak area. The presence of 0.01, 0.05 or 0.1% (v/v) benzene in the sample did not affect the baseline, but the peak area for a 5 pg 1-l standard was reduced by 0, 20 and 40%, respectively. The large decrease with benzene is thought to be caused by relatively strong absorption of the 253.7-nm radiation. According to Thompson and Reynolds [3], the effect is decreased when air is used as the carrier gas, but this would reduce the sensitivity considerably. Alternatively, the method of standard additions can be used. Moreover, it is highly unlikely that the final sample will contain such high concentrations of organic compounds. Detection limit and precision The detection limit, defined as the mercury concentration corresponding to three times the standard deviation of the blank, corresponds to 0.9 ng 1-l. This detection limit can be compared with that obtained with AAS. Only when mercury is preconcentrated on a gold-coated absorber from a l-l sample during 1 h is this detection limit decreased to 1 ng ll’. Therefore, comparison of the absolute amount detectable by the two techniques gives a better illustration of the superior properties of AFS for mercury (2 pg by AFS, compares with 100 pg by AAS). The precision is generally better than 1% (r.s.d.) in the pg 1-l concentration range and better than 5% in the ng 1-i concentration range. Analysis of reference materials In order to verify the accuracy of the method, the optimized procedure was used to analyse the
DETERMINATION
OF MERCURY
TABLE
1
Analysis
of biological
Sample
IN BIOLOGICAL
reference Mercury
materials
207
SAMPLES
and real samples
content
Found
x
Certified s
n
BCR CRM 151 Milk Powder (spiked) (ng g-‘) BCR CRM 186 Pig
4.8
3
0.003
3
55.5
3.9
3
57*15
11.55
0.24
5
12.3kO.9
102.6
Kidney (pg g-‘) NBS SRM 1566 Oyster Tissue (ng g-‘) BCR RM 397 Human Hair (pg g-‘)
1.987
101*10 1.97+0.04
Literature 1630 4.87
Hair (ng g-‘) Urine (pg I-‘) a Provisional
certified
60 0.02
3 3
a [13]
30-4300 0.8-6.4
value.
certified reference materials BCR CRM 151 Milk Powder (spiked), BCR CRM 186 Pig Kidney and NBS SRM 1566 Oyster Tissue. The results obtained (Table 1) agreed well with the certified values, indicating the accuracy of the method. In addition, this laboratory participated in a certification programme for the determination of trace elements in human hair, organized by the Community Bureau of Reference (BCR; Commission of the European Communities). The hair samples were taken from subjects living in an Italian rural environment. The samples were cleaned with water and ethanol, ground under liquid nitrogen to a final size down to 80 pm and homogenized in a drum. Stability and homogeneity were investigated and found to be acceptable for the production of a reference material. The results obtained for the candidate reference material are given in Table 1. The present results agreed well with the inter-laboratory mean and with the proposed certified value [ll]. Determination
of mercury
in human
hair
and
urine
Head hair readily incorporates methylmercury at the time the hair is formed [12]. The concentration in newly formed hair is directly proportional
to the blood concentration. Once incorporated into hair, the concentration remains stable for many years. Provided that steps are taken to avoid external contamination, hair forms a record of previous exposure. The optimized method was used to analyse a hair sample, taken without special precautions to avoid contamination (Table 1). The mercury concentration in the hair was 1630 ng g-i with a standard deviation of 60 ng gg’. This value falls within the range given in the literature [13] for unexposed subjects, i.e., 30-4300 ng g-i. The mercury concentration in the reference material (see above) is much higher and falls in the range given by Baron and Schweinsberg [13] for exposed subjects, i.e., 1400-15 300 ng g-‘. Urine is the chief medium of excretion of inorganic divalent mercury and is the most commonly used for biological monitoring. Concentrations in urine probably indicate kidney levels. The optimized method was used to analyse a urine sample immediately after collection (Table 1). The urine sample was analysed both by calibration against aqueous standards and by using the standard addition method. The concentrations obtained with the two methods were 4.87 + 0.02 and 4.57 + 0.27 pg l-i, respectively. The blank for the entire analytical procedure was 13.9 & 1.0 ng 1-i and the detection limit was 30 ng 1-i assuming that a lo-ml sample of urine was digested and diluted to 100 ml and 10 ml of this dilution was analysed. The concentration found falls within the range given by Baron and Schweinsberg [13] (0.86.4 pg I-‘) for unexposed subjects.
Conclusion
The determination of mercury in solutions by AFS combined with mercury vapour generation is a simple, sensitive and specific method for determining low levels of mercury. The main advantage over CVAAS is that ng 1-i concentration levels can be measured without preconcentration on gold. The accuracy is shown by the good agreement between the results obtained and the certified values. The determinations are precise (better than 5% r.s.d.) and sensitive (detection limit 0.9 ng 1-l).
208 REFERENCES 1 G. Vermeir, C. Vandecasteele and R. Dams, Microchim. Acta, Part III, (1988) 305. 2 C. Vermeir, C. ,Vandecasteele and R. Dams, Anal. Chim. Acta, 220 (1989) 257. 3 K.C. Thompson and G.D. Reynolds, Analyst, 96 (1971) 771. 4 V.I. Muscat and T.J. Vickers, Anal. Chim. Acta, 57 (1971) 23. 5 J.E. Caupeil, P.W. Hendrikx and J.S. Bongres, Anal. Chim. Acta. 81 (1976) 53. 6 K.C. Thompson and R.G. Godden, Analyst, 100 (1975) 544.
G. VERMEIR
ET AL,
7 R. Ferrara, A. Scritti, C. Barghigiani and A. Petrorino, Anal. Chim. Acta, 117 (1980) 391. 8 H. Morita, T. Kimoko and S. Shimomura, Anal. Lett. 16 (1983) 1187. 9 N. Bloom and W.F. Fitzgerald, Anal. Chim. Acta, 208 (1988) 151. 10 D.C. Stuart, Anal. Chim. Acta 101 (1978) 421. 11 B. Griepink, Ph. Quevauviller, E.A. Maier and H. Munkau, The certification of the contents (mass fraction) of Cd, Hg, Pb, Se and Zn in human hair, CRM 397, Commission of the European Communities, to be published. 12 T.W. Clarkson, J. Am. COIL Toxicol., 8, No. 7 (1989) 1291. 13 P. Baron and F. Schweinsberg, Zentralbl. Hyg. Umweltmed., 188 (1989) 84.
203
Atomic fluorescence spectrometry combined with reduction aeration for the determination of mercury in biological samples G. Vermeir
*, C. Vandecasteele
’ and R. Dams
Laboratory of Analytical Chemistry University of Gent, Institute for Nuclear Sciences, Proeftuinstraat 86, B-9000 Gent (Bel@n) (Received
31st May 1990)
Abstract A sensitive atomic fluorescence system for the determination of mercury was optimized. The system consists of a continuous-flow mercury vapour generator coupled to a fluorescence spectrometer specific for mercury. A new gas-liquid separator was developed. A detection limit of 0.9 ng Hg 1-l was obtained. The system was combined with a microwave oven for dissolving samples in closed Teflon PFA vessels. Accurate results were obtained for certified reference materials, and biological samples such as urine and hair were analysed. Keywords; Atomic
fluorescence
spectrometry;
Mercury;
Biological
samples
scribed a flow-injection system for the determination of mercury by CVAFS. Bloom and Fitzgerald [9] used Carbotrap preconcentration followed by cryogenic gas chromatography with CVAFS detection for the determination of organomercurial species in gaseous samples. However, the application of AFS was hampered because no commercial system was available until a few years ago. This paper describes the optimization of a commercial system for the analysis of biological samples for mercury by AFS. Moreover, and in contrast to previous work [5,6], the accuracy of the method was assessed by analysing several certified reference materials. Additionally hair and urine samples were analysed and the results were compared with literature values.
In previous papers [1,2] the determination of mercury in biological samples by cold vapour (CV) atomic absorption spectrometry (AAS) was addressed. This required preconcentration on a gold absorber in order to achieve sufficiently low detection limits. Atomic fluorescence spectrometry (AFS) allows a more sensitive determination of mercury than AAS. This method has frequently been applied to the analysis of water samples [3,4]. For several interesting biological samples, direct analysis is possible without any preconcentration [5,6]. Caupeil et al. [5] applied AFS spectrometry to the determination of mercury in fish samples at the mg kg-’ level. Thompson and Godden [6] tested their system on urine and blood samples. Ferrara et al. [7] improved the AFS technique by using a mercury radiofrequency electrodeless discharge lamp as excitation source. Morito et al. [S] de-
EXPERIMENTAL
Reagents ’ Research Director tific Research.
0003-2670/91/$03.50
of the Belgian
National
0 1991 - Elsevier
Fund
All chemicals and water from
for Scien-
Science Publishers
B.V.
were of analytical-reagent grade a Millipore Mini-Q purification
G. VERMEIR
204
system was used throughout. A stock standard solution of mercury (1.000 g 1-l) was prepared by dissolving mercury(I1) oxide in 14 M nitric acid and diluting to, 1 1. Working standard solutions (ng 1-i range) were prepared daily by dilution immediately before use. All standards, blank and sample solutions were stabilized for up to 10 h with 2% (v/v) nitric acid, Tin(I1) chloride solution was prepared by dissolving 20 g of the dihydrate (highest purity, UCB) in SO ml of boiling 12 M hydrochloric acid followed by dilution with 200 ml of water and 600 ml of 1.5 M sulphuric acid. This solution was cooled and purged with nitrogen for 30 min. Dissolution procedure
A lOO-mg amount of solid sample or 10 ml of liquid sample was weighed directly into a Teflon PFA digestion vessel and sub-boiled concentrated nitric acid (1 ml for the solid samples and 5 ml for the liquid samples) was added. Six sealed vessels and a beaker filled with 50 ml of water, to absorb excess of microwave energy, were placed inside a plastic box which was closed and placed in a microwave oven (Amana, Model R.S. 560A). The dissolution programme consisted of three steps: 20% power for 8 rnin, 40% power for 8 min and 60% power for 4 min. After cooling, the digest was
Fig. 1. Schematic
flow diagram
of the system.
transferred to a volumetric 100 ml with water. Mercury
flask
and
ET AL.
diluted
to
determination
Mercury was determined using an AFS system (PS Analytical, Sevenoaks), consisting of a mercury vapour generator, a fluorescence spectrometer and a computer. This system copes with normal laboratory conditions, but it was optimized to lower the detection limits. The continuous-flow mercury vapour generator (Model PSA 10003 hydride generator; PS Analytical) consists (Fig. 1) of a carrier gas supply, a pumping system delivering a continuous flow of tin(I1) chloride solution, blank and sample, a Tpiece where the tin(I1) chloride solution and the blank or the sample are mixed and a gas-liquid separator. The sample and blank streams are switched using a combination of two three-port, two-way miniature Teflon valves as indicated in Fig. 1. Blank and samples are pumped at a flowrate of 7 ml min-’ and tin(I1) chloride solution at a flow-rate of 3 ml mm’. The atomic fluorescence spectrometer (Merlin Model PSA 10023 fluorescence detector; PS Analytical) consists of a high-intensity mercury lamp, an open chimney designed by Thompson and Godden [6] into which the mercury vapour is fed by means of the argon
DETERMINATION
OF MERCURY
IN BIOLOGICAL
205
SAMPLES
carrier gas, an interference filter to isolate the 254-nm resonance line and a photometric detector. The response is displayed digitally as a reading from 0 to ,199 and as a 0-lo-mV analogue output that can be interfaced to a PC or chart recorder. The analogue output is converted to a digital signal by an analogue-to-digital convertor. A Tulip PC records the time-resolved fluorescence signal (one measurement per second during 300 s). Integration is carried out over ca. 260 s (peak-area mode). Correction of the peak area for baseline drift is possible using time windows. The system can also be used in the peak-height mode.
RESULTS
AND
The sensitivity obtained with argon carrier gas was approximately ten times greater than that with nitrogen. This is caused by the quenching of excited mercury atoms by nitrogen molecules. Argon was used in all subsequent measurements. The flow-rate was varied from 200 to 600 ml mm’. In this range 200 ml rnin- ’ was optimum for both peak height and peak area (Fig. 2). When the central tube of the chimney was sheathed with argon, the signal decreased. Therefore, the use of the sheath gas was discontinued. The tin(I1) chloride concentration has a small influence on the signal, but a concentration of ca. 21 g 1-l seems optimum. Peak :
200
1
area
(x100)
Peak height .__~_~
600
200
1
01 0
I_____--_-
100
area
(x1000)
-~~~
Fig. 3. Effect of the sample uptake rate on the signal (mercury standard solution, 10 pg 1-l; calibration range 1).
DISCUSSION
Optimization
600
Peak s”r
200
300
400
L.
500
600
-0 700
Ar flow (ml/mid Fig. 2. Effect of argon flow-rate on the fluorescence signal (10 solution; calibration range 10). 0, Peak pg Hg 1-l standard area; +, peak height.
The signal obtained over 1 h (twelve measurements of 5 min) was stable to within 2%. Longterm variations were also within reasonable limits. The effect of the sample uptake rate on the fluorescence signal is shown in Fig. 3. The line bends from a sample uptake rate of 5 ml mm’. This means that above this sample uptake rate the mercury is not completely removed from the solution. The graph shows that at a sample uptake rate of 7 ml min-‘, which is the rate at which the mercury vapour generator operates, ca. 90% of the mercury present was liberated. To increase the efficiency of volatilization of mercury from the solution, the gas-liquid separator shown in Fig. 4 was devised. A glass sinter was sealed across the end of the gas inlet, causing fine bubbles to be carried through the solution. This increased the solution-gas interface area appreciably. This system increased the signal by 20%, and improved the baseline stability, compared with the original separator shown in Fig. 1. The signal rises to a maximum level once the valves have switched between the blank stream and the sample stream. When the sampling period is finished, the signal returns to the baseline level. It is clear that because of short-term fluctuations of the signal, calculation of the peak area will yield more reproducible results than just using the peak height after a given time. Therefore, the signal is integrated from 0 to 300 s, whereas the sample is introduced from 5 to 95 s.
206
G. VERMEIR
r--
DETECTOR
Ar CARRIER GAS I
r-
\
I
>
Sn Cl2 *SAMPLE I
O2 GLASS
Fig. 4. New gas-liquid
SINTER
separator.
As the recovery of mercury from the standard and sample may differ slightly, because of matrix differences, it is desirable that the recovery of mercury from both the standard and the sample are almost 100%. This was verified in two ways. First, the fraction going to waste was collected for a standard and for a spiked sample and was found to contain less than 5% of the total mercury concentration. Second, the signal obtained was compared with that obtained in a batch system [1,2] after quantitative aeration. It appeared that the present system yields an integrated signal equal to 96.7% with a standard deviation of 2.7% (five determinations) of that obtained with the batch system. The new gas-liquid separator had a better efficiency, but a larger amount of water vapour was produced, which on condensation may produce stray light, resulting in baseline drift. Therefore, a water trap consisting of a quartz tube filled with shredded paper [lo] was placed between the gasliquid separator and the detector. An effect of water vapour was noted only when measuring in the most sensitive range. Mercury has a pronounced tendency to absorb on surfaces, resulting in a memory effect. On
ET AL.
changing from a low-mercury sample (0.4 pg 1-l) to a high-mercury sample (4.0 pg l-i), or vice versa, a representative signal is obtained only after 4-5 analyses. This effect can be reduced by decreasing the surface in contact with the mercury vapour and by adopting a proper measuring sequence. In addition, decreasing the tube length between the mercury vapour generator and the detector from 500 to 160 mm increased the sensitivity by 5%. Effect of organic solvents The presence of 0.01,0.05 or 0.1% (v/v) acetone in the sample did not affect the baseline stability and with a 5 PI-181-i sample gave no reduction in peak area. The presence of 0.01, 0.05 or 0.1% (v/v) benzene in the sample did not affect the baseline, but the peak area for a 5 pg 1-l standard was reduced by 0, 20 and 40%, respectively. The large decrease with benzene is thought to be caused by relatively strong absorption of the 253.7-nm radiation. According to Thompson and Reynolds [3], the effect is decreased when air is used as the carrier gas, but this would reduce the sensitivity considerably. Alternatively, the method of standard additions can be used. Moreover, it is highly unlikely that the final sample will contain such high concentrations of organic compounds. Detection limit and precision The detection limit, defined as the mercury concentration corresponding to three times the standard deviation of the blank, corresponds to 0.9 ng 1-l. This detection limit can be compared with that obtained with AAS. Only when mercury is preconcentrated on a gold-coated absorber from a l-l sample during 1 h is this detection limit decreased to 1 ng ll’. Therefore, comparison of the absolute amount detectable by the two techniques gives a better illustration of the superior properties of AFS for mercury (2 pg by AFS, compares with 100 pg by AAS). The precision is generally better than 1% (r.s.d.) in the pg 1-l concentration range and better than 5% in the ng 1-i concentration range. Analysis of reference materials In order to verify the accuracy of the method, the optimized procedure was used to analyse the
DETERMINATION
OF MERCURY
TABLE
1
Analysis
of biological
Sample
IN BIOLOGICAL
reference Mercury
materials
207
SAMPLES
and real samples
content
Found
x
Certified s
n
BCR CRM 151 Milk Powder (spiked) (ng g-‘) BCR CRM 186 Pig
4.8
3
0.003
3
55.5
3.9
3
57*15
11.55
0.24
5
12.3kO.9
102.6
Kidney (pg g-‘) NBS SRM 1566 Oyster Tissue (ng g-‘) BCR RM 397 Human Hair (pg g-‘)
1.987
101*10 1.97+0.04
Literature 1630 4.87
Hair (ng g-‘) Urine (pg I-‘) a Provisional
certified
60 0.02
3 3
a [13]
30-4300 0.8-6.4
value.
certified reference materials BCR CRM 151 Milk Powder (spiked), BCR CRM 186 Pig Kidney and NBS SRM 1566 Oyster Tissue. The results obtained (Table 1) agreed well with the certified values, indicating the accuracy of the method. In addition, this laboratory participated in a certification programme for the determination of trace elements in human hair, organized by the Community Bureau of Reference (BCR; Commission of the European Communities). The hair samples were taken from subjects living in an Italian rural environment. The samples were cleaned with water and ethanol, ground under liquid nitrogen to a final size down to 80 pm and homogenized in a drum. Stability and homogeneity were investigated and found to be acceptable for the production of a reference material. The results obtained for the candidate reference material are given in Table 1. The present results agreed well with the inter-laboratory mean and with the proposed certified value [ll]. Determination
of mercury
in human
hair
and
urine
Head hair readily incorporates methylmercury at the time the hair is formed [12]. The concentration in newly formed hair is directly proportional
to the blood concentration. Once incorporated into hair, the concentration remains stable for many years. Provided that steps are taken to avoid external contamination, hair forms a record of previous exposure. The optimized method was used to analyse a hair sample, taken without special precautions to avoid contamination (Table 1). The mercury concentration in the hair was 1630 ng g-i with a standard deviation of 60 ng gg’. This value falls within the range given in the literature [13] for unexposed subjects, i.e., 30-4300 ng g-i. The mercury concentration in the reference material (see above) is much higher and falls in the range given by Baron and Schweinsberg [13] for exposed subjects, i.e., 1400-15 300 ng g-‘. Urine is the chief medium of excretion of inorganic divalent mercury and is the most commonly used for biological monitoring. Concentrations in urine probably indicate kidney levels. The optimized method was used to analyse a urine sample immediately after collection (Table 1). The urine sample was analysed both by calibration against aqueous standards and by using the standard addition method. The concentrations obtained with the two methods were 4.87 + 0.02 and 4.57 + 0.27 pg l-i, respectively. The blank for the entire analytical procedure was 13.9 & 1.0 ng 1-i and the detection limit was 30 ng 1-i assuming that a lo-ml sample of urine was digested and diluted to 100 ml and 10 ml of this dilution was analysed. The concentration found falls within the range given by Baron and Schweinsberg [13] (0.86.4 pg I-‘) for unexposed subjects.
Conclusion
The determination of mercury in solutions by AFS combined with mercury vapour generation is a simple, sensitive and specific method for determining low levels of mercury. The main advantage over CVAAS is that ng 1-i concentration levels can be measured without preconcentration on gold. The accuracy is shown by the good agreement between the results obtained and the certified values. The determinations are precise (better than 5% r.s.d.) and sensitive (detection limit 0.9 ng 1-l).
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