Ultratrace determination of mercury in water following EN and EPA standards using atomic fluorescence spectrometry
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- Labatzke, T. & Schlemmer, G. Anal Bioanal Chem (2004) 378: 1075. doi:10.1007/s00216-003-2416-x
Chemical vapour generation has been used in combination with atomic fluorescence spectrometry to determine mercury at ultratrace concentrations down to 0.1 ng L−1. A time-based injection of 1 mL of solution for measurement was sufficient to generate a steady-state detector response in the direct mode of measurement. The detection limit calculated from a ten-point calibration curve according to DIN 32645 was 0.26 ng L−1. Instrument noise is limited by reflected radiation from the light source rather than by the dark current of the photomultiplier. The detection limit is directly influenced by the reagent blank which was 2 ng L−1 in the experiments described. Focusing by amalgamation and subsequent thermal desorption generates a detector response which is about eight times higher in peak intensity and about twice as large in integrated intensity. The detection limit under these conditions is 0.09 ng L−1 which can be further improved by preconcentration of larger volumes of solution for measurement. The cycle time for one individual reading is about 40 s without amalgamation and 125 s with amalgamation. The linear dynamic range of the system is five orders of magnitude with a single photomultiplier gain setting. The carry-over is less than 0.3% in direct measurement mode. Reference water samples and a surface water containing approximately 5 ng L−1 were used to prove the validity of the method for real samples. Good accuracy and recoveries of 103% were calculated using the fast direct determination technique.
KeywordsAtomic fluorescence spectrometryChemical vapour generationWaterMercury
For many years mercury has been one of the elements monitored at concentrations below 1 µg L−1 in environmental samples such as drinking water or surface water . Despite rigorous control in many countries the global mercury pollution is increasing according to the United Nations Environment Program (UNEP)  and global world-wide pollution is higher than previously estimated. Mercury is one of the most critical heavy metals in the environment and in the biological organism. It is considered to be an endocrine-disrupting chemical and it has become obvious that even very low concentrations of total mercury, e.g. in lake water, can be accumulated by a factor of more than 10,000 in fish . Most recent investigations by Frech and co-workers  prove that, depending on the kind of ecological system, even extremely low concentrations of mercury can be accumulated in biological organisms to levels which are no longer below threshold values for food. It is obvious that determinations of mercury at 1 ng L−1 levels and below will have to become routine in laboratories in the near future .
The classical method for total mercury determination at low concentrations is the so-called “chemical vapour generation” technique which was first described by Hatch and Ott . Mercury, present in the sample as a variety of species is oxidized, so that the solution for measurement contains only the doubly charged cation Hg2+. In a second chemical pre-treatment step it is reduced to its metallic form by use of either stannous chloride (SnCl2), a rather mild reducing agent, or NaBH4, which is much more reactive but much less selective . The neutral mercury is driven from the solution by a carrier gas (usually argon) and transported to the measurement cell. Alternatively it can be trapped on a gold/platinum gauze and, after complete stripping of all matrix components, rapidly desorbed thermally from the gauze . Trapping results in focusing of mercury in the measurement cell by a factor of at least five compared with the direct mode of measurement. The increase in sensitivity can usually be transformed into five times better detection limits . The time for an individual determination will be about three times longer with the amalgamation technique, because two more operating cycles, preconcentration and desorption, are required. The advantages of amalgamation are more complete analyte–matrix separation and hence an even better selectivity. The disadvantages are higher risk of contamination and carry over and longer cycle times. A further improvement in detection limit with the amalgamation technique is achievable if sample volumes beyond the optimum in direct measurement are preconcentrated and released. At the cost of larger sample volumes and longer cycle times the detection limits can theoretically be reduced in a manner which depends linearly on sample volume. However, in real life, very high preconcentration volumes require correspondingly high masses and volumes of reagents which will inevitably increase the contamination level relative to the targeted detection limit. There will be a limit of the preconcentrated volume at which the detection limit is no longer defined by the standard deviation of the spectrophotometer but by the standard deviation of the blank level.
Traditionally the analyte vapour was measured by atomic absorption spectrometry, either in a standard AA spectrometer using mercury hollow-cathode lamps or electrodeless discharge lamps or with a dedicated mercury analyser with special low-pressure mercury lamp and photomultiplier tube or photodiode or a charge-coupled device-type detector . Whereas the first type of instrument results in limits of determination (LOD) of about 100 ng L−1 without amalgamation, a specific mercury AAS analyser might afford an LOD of lower than 10 ng L−1 without amalgamation. Atomic fluorescence spectrometry (AFS) is an attractive alternative to AAS. Its fundamentals were investigated and described  only shortly after AAS has been commercially introduced to instrumental analysis, but its application to routine element analysis in flame atomizers was limited. If measurements are performed in argon-flushed cells after analyte–matrix separation, however, AFS has distinct advantages over AAS . These are predominantly spectrophotometric detection limits, which are at least an order of magnitude superior to those of AAS with specific light source and detector, and a dynamic spectrometric working range of 4 to 5 orders of magnitude. The theoretically predicted benefits were later shown experimentally  and today the most sensitive mercury analysers are based on atomic fluorescence. Both the US EPA and the European Community recently issued standards which reflect the required limits of determination and the new instrumental developments for the determination of mercury. In the new standards for ambient waters in US EPA method 1631  the required limits of detection and determination are 0.2 ng L−1 and 0.5 ng L−1 respectively. In European Community method EN 13506  the stated working range is between 1 ng L−1 and 100 ng L−1. The method detection limit in the latter method was found to be below 1 ng L−1.
In the work discussed in this paper a new specific mercury analyser based on the chemical vapour generation technique with atomic fluorescence detection was used to study fundamental factors influencing the working range of mercury determinations. Using EPA method number 1631, the factors which define analytical quality were investigated.
The fluorescence obtained from a line source is linearly dependent on the intensity of the incident radiation and the concentration of fluorescing atoms. From theoretical considerations the linear dynamic range should be greater than four orders of magnitude. To match the best signal-to-noise ratio with the required dynamics of photomultiplier and amplifiers, the system is equipped with an automatic gain setting which is based on measurement on the actual line source intensity and combined with sensitivity information obtained from instrument performance measurements during first installation of the system.
Generation and control of mercury vapour
sample injection—reaction—separation—drying—determination in the fluorescence cell;
sample injection—reaction—separation—drying—collection 1—thermal desorption—determination in the fluorescence cell; or
sample injection—reaction—separation—drying—collection 1—thermal desorption 1—collection 2—thermal desorption 2—determination in the fluorescence cell.
Sample or carrier can be directly passed into waste without contact with the reactor unit or the gas–liquid separator. This way the sampling part can be cleaned selectively.
A second valve behind the gas–liquid separator can be switched to “waste” to make sure that the whole reaction and separation unit can be flushed and cleaned without contaminating the gas drying unit, collectors, or measurement cell. This valve is also active if liquid bubbles are detected in the reaction gas stream coming from the gas liquid separator.
Finally a high stream of about 830 mL min−1 can be activated to flush the measurement cuvette rapidly, to minimize the time for one analysis. This procedure is called “fast baseline return” (FBR).
Function–time procedure for determination of mercury by AFS by direct introduction of generated mercury vapour into the fluorescence cell
Time interval (s from start)
Flow S/C/R (mL min−1)
Flow Ar (mL min−1)
Function–time procedure for determination of mercury by AFS with amalgamation–desorption of the generated mercury vapour
Time interval (s from start)
Flow S/C/R (mL min−1)
Flow Ar (mL min−1)
reflected light from the excitation source;
mercury fluorescence as a result of contamination of argon, tubing, and reservoirs;
mercury fluorescence as a result of contamination of carrier, reductant, or reagents; and
mercury fluorescence from the measurement solution.
Reflected light is quantitated and a correction is made shortly before each read cycle. It is also used to control the intensity output of the mercury radiation source and correct for minimal possible drifts in source intensity. At the same time argon, cleaned by activated carbon, which is passing directly through the fluorescence cell might add infinitesimally small Hg-specific fluorescence to the baseline reading. This also is corrected in the baseline auto-zero cycle. This reading should be very small and more or less constant. It will not add to the blank reading but might influence the baseline noise level.
In a second and independent mode of operation, the argon can be passed over the reaction unit while carrier and reduction are active. This way a “constant” blank level of carrier and reagent is automatically corrected for. If the blank level is elevated, however, it will increase the baseline noise level also.
The generated analyte gas is flushed through the absorption cell with a fairly low argon flow. Although variable between 85 and 250 mL min−1 it is usually optimised for highest mercury density and optimum peak shape in the range 85 mL min−1 without amalgamation and 170mL min−1 with amalgamation. In these circumstances mercury can be removed from the measurement volume completely after about a minute only. With FBR transient Hg can be efficiently removed within less than 5 s.
Reagents and standards
Ultrapure deionised water (resistivity at least 18 MΩ) was used for all operations. All reagents were at least “analytical grade” or higher. Borosilicate containers were used to prepare and store all standards and all solutions for measurement. For preparation of blanks and standards the containers were cleaned with bromine-containing blank solutions for at least 24 h.
In all experiments we followed closely the recommendations given by EPA method 1631 .
The stock solution for oxidizing and stabilizing standards and samples consists of 10.8% KBr (w/v) and 15.2% KBrO3 (w/v) in concentrated HCl; 10 mL per litre of sample or standard is used to stabilise and acidify the solutions for measurement.
The carrier consists of 1.8% (w/v) dilute hydrochloric acid.
The reduction solution contains 2% (w/v) SnCl2 in 3.6% (w/v) hydrochloric acid. It is purged for at least 2 h by bubbling purified argon through the container.
The hydroxylamine hydrochloride stock is a 12% (w/v) solution in ultrapure water. The reductant is purged from Hg by bubbling purified argon through the solution after addition of 1 mL SnCl2 solution.
Standards were prepared from a 1 mg L−1 Hg stock solution and stabilized by addition of the bromine-containing solution described above. The pre-reduction solution was added less than 1 h before measurement of the sample. Prereduced samples at concentrations below 100 ng L−1 are stable for a few hours only.
A river water certified reference material CRM-ORMS-2 (National Research Council Canada, Ottawa, Canada) was used for validation of the data. The CRM was diluted 1:5. Its original mercury concentration is 30.6 ng L−1 (±2.3 ng L−1). The CRM was handled and pretreated in the same way as blanks and samples. Unfiltered surface water from the river Saale was used for recovery experiments. The water was stabilized with the bromine-containing solution described above immediately after collection.
Results and discussion
Photometric noise and detection limits
With amalgamation on one gold net the limits of detection and determination were 0.090 and 0.325 ng L−1, respectively, using the 10-point calibration plot and 1 mL of sample per injection. After preconcentration of 3 mL of sample, the respective values were 0.065 ng L−1 for detection limit and 0.234 ng L−1 for quantitative determination limit.
Linear dynamic range, working range, carry-over, and relative standard deviation
Carry-over from 1 µg L−1 mercury on consecutive blank readings
Solution for measurement
Peak intensity (counts)
Peak intensity, reagent blank corrected
Concentration (µg L−1)
Concentration reagent blank corrected (µg L−1)
The relative standard deviation at elevated concentrations of approximately ten times the limit of determination is mainly a result of the reproducibility of sample introduction and chemical reaction . The RSD (n=11) for a 10 ng L−1 standard was 0.7% without amalgamation and 0.7% with amalgamation.
Determination of mercury in water samples
Determination of CRM-ORMS-2 and water from the river Saale. Sets 1 to 9 are within-batch determinations with three replicates per samples. The average values and standard deviations stated in the text are averages from individual runs (between-batch average) and standard deviations
Determined (ng L−1)
s.d. (ng L−1)
Saale+5 ng L−1
Avg Saale+5 ng L−1
Although blank control of mercury in containers, tubing, gases, and all reagents and solutions used is of central importance if the element has to be determined at concentrations below 10 ng L−1, these conditions can be obtained in a routine laboratory without excessive equipment or operations. It has been found that method detection limits below 0.5 ng L−1 are possible if the blank level can be limited to about 2 ng L−1 Hg. The spectrometric detection limits of optimised atomic fluorescence spectrometers can reach the currently most stringent detection limit requirements even without amalgamation. Compared with atomic absorption spectrometry they are lower by about one order of magnitude. They can be improved by another order of magnitude if the amalgamation technique is applied. A second inherent advantage of atomic fluorescence over atomic absorption spectrometry is the linear dynamic working range of five orders of magnitude. The wide dynamic range can be used only if carry-over from sample to sample is minimised. Values of less than 0.5% carry-over are possible even for the critical element mercury by optimisation of liquid and gas velocities and pathways. Quenching has to be avoided or minimised by careful gas–liquid separation, optimised drying of the mercury cold vapour, and, in complex samples, by the amalgamation technique.