Simple and Versatile Atomic Fluorescence System for Determination of Nanogram Quantities of Mercury V. I. Muscat,l T. J. Vickers,lI2and Anders Andrena Florida State University, Tallahassee, Fla. 32303 A flameless atomic fluorescence system for mercury which makes use of either reduction-aeration or combustion techniques for the generation of mercury vapor and a silver amalgamator for collection of mercury prior to the final measurement i s described. The system is capable of quantitative determination of mercury in samples containing as little as 0.6 ng of Hg. Results are reported for application of the system t6 a variety of samples, including water, rock, wheat flour, and natural sediments. CLOSED SYSTEM reduction-aeration atomic absorption and atomic fluorescence techniques for the determination of nanogram quantities of mercury have been compared in a recent report ( I ) . The reduction-aeration method of mercury atomization, which has achieved wide use (2) in the atomic absorption determination of mercury, has been shown to be applicable to atomic fluorescence measurements, and the atomic fluorescence technique has been found to have better sensitivity and lesser requirements for instrumentation than the atomic absorption technique. The present report describes a system which overcomes certain limitations of previous atomic fluorescence methods for the determination of mercury and reports results obtained with this system for a variety of sample types. The present atomic fluorescence system makes use of a silver amalgamator to collect the mercury prior to the fluorescence measurement. Mercury vapor in a carrier gas stream is brought into close contact with silver wire at ambient temperatures and is effectively removed from the gas stream by amalgam formation. The mercury is subsequently released for the final measurement step by heating. Such noble metal traps for separation and concentration of mercury have been in use for many years (3) and have recently been used with atomic absorption spectrophotometry (4-8). EXPERIMENTAL
Apparatus. A summary of the experimental facilities is given in Table I. The fluoresence vapor cell and measurement system are identical to those described previously ( I ) , and the gas train is as shown schematically in Figure 1. Two types of mercury generators have been employed in this study-a reduction-aeration system for solution samples, and a furnace system for solid samples. In the reduction-aeration system mercury present in solution as Hg(I1) was reduced by addition of Sn(II), and the resulting mercury vapor was 1 2
Department of Chemistry. Author to whom correspondence should be directed. Department of Oceanography.
(1) V. I. Muscat and T. J. Vickers, Anal. Chim. Acta, in press. (2) D. C. Manning, At. Absorption Newslett., 9,97 (1970). (3) Schumacher and W. L. Jung, Z . Anal. Chem., 39,12 (1900). (4) W. W. Vaughn and J. H. McCarthy, US.,Geol. Sur., Pi rof. PUP.,501-D D123 (1964). (5) U. Ulfvarson, Acta Chem. Scand., 21,641 (1967). (6) V. Lidums and U. Ulfvarson, ibid., 22,2150 (1968). (7) G. Thilliez, Chim. Anal., 50, 226 (1968). (8) G. W. Kalb, At. Absorption Newslett., 9,84 (1970). 218
Table I. Experimental Facilities Hg pen-lamp, Ultra Violet Products, Inc. Excitation source 30-mm diameter, 50-mm focal length External optics fused silica lens Heath EU-700 scanning monochromator, Spectrometer 350-mm Czerny-Turner mounting, 1180 grooves/emmgrating blazed for 2500 A, f/7, 20 A/mm reciprocal linear dispersion at exit slit in the 1st order, ganged entrance and exit bilaterally adjustable straight slits R166 solar blind multiplier phobotube, Detector Hamamatsu TV, 1690-3200 A spectral response Detector power supply Heath EU-701,300-1500 V dc, 0-1.5 mA Heath Model EU-703-31, 10-'-lO-l A Amplifier full scale deflection Hewlett-Packard Model 680 potentioRecorder metric recorder Square borosilicate glass 2-cm tubing with Fluorescence cell 2 x 2.5 cm Vycor windows on two adjacent sides Variable transformer Allied Radio Corp., Type 64-940. Input 120 V, 50-60 Hz Sargent, Cat. No. S-36400 Tube furnace
swept into the amalgamator by bubbling the selected carrier gas through the solution. The reduction-aeration vessel employed was similar to that described by Kalb (8). In the furnace system, solid samples were heated in a tube furnace, and a suitable carrier gas was used to sweep the released mercury vapor into the amalgamator. The furnace tube was a 450 mm long, 13-mm i.d. fused silica tube with ground glass joints at both ends for connection to the gas train. In use, the central 200 mm of the tube was placed in the furnace. A trap containing lead acetate was placed between the furnace tube and the amalgamator to remove volatile sulfides from the gas stream. The silver amalgamator consisted of a 180 mm long, 9-mm i.d. fused silica tube which was closely packed with 36 gauge silver wire. Facility for heating the amalgamator was obtained by wrapping the tube with approximately 5 feet of standard resistance wire (3 ohms/foot), the ends of which were connected to a variable transformer. A preliminary study indicated that mercury could best be freed from the amalgamator by applying approximately 70 V across the wire for 35 sec. Higher power settings decreased the life of the heater without significantly improving the sensitivity of the determination. Reagents. The solutions used were : stannous chloride, 10% w/v solution in 1N "OB; hydroxylamine hydrochloride, 25 % w/v solution; potassium permanganate, 6 75 w/v solution; and potassium peroxydisulfate, 6% w/v solution. All standard mercury solutions were prepared from mercuric chloride. A stock solution of 100 pg/ml was prepared by dissolving 0.1354 gram of mercuric chloride and diluting
ANALYTICAL CHEMISTRY, VOL. 44, NO. 2, FEBRUARY 1972
Carrier Gas
Flow Meter
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Fluorescence Cell
Mercury Generator
4 Figure 1. Schematic diagram of gas train
A small quantity of oxygen was added to the argon stream through a T-joint to facilitate combustion. With the sample in place and the flow of gas started, the tube was placed in the furnace, which had previously been heated to approximately 800 “C. Combustion was complete within 120 seconds. The tube was disconnected from the gas train, the flow of argon through the amalgamator was started, and the remainder of the measurement was carried out in the same way as described for the reduction-aeration method. Both methods required periodic cleaning of the amalgamator to maintain a low blank. This was accomplished by heating the amalgamator for several minutes at a 85-V heater setting and flushing with argon or air. Blank
RESULTS AND DISCUSSION
Figure 2. Typical recorder tracing of fluorescence signals to 1 liter with I N “03. Two drops of K M n 0 4 were added to each mercury solution. All solutions were stored in polyethylene containers. Reagents were checked regularly for Hg content. Stannous chloride was found to have significant Hg contamination, but this was easily eliminated by aerating the stannous chloride solution for 5-10 minutes before use, thereby removing the elemental Hg. One bottle of “03, obtained for use in the digestion procedure, gave a high blank reading and was rejected. All other reagents were used as received. Glassware. All glassware used in preparing samples for the reduction-aeration procedure was washed with detergent, rinsed with 1 M ” 0 3 and a chloroform solution of dithizone, and heated for several hours in an oven at 125 “C. Procedure. Samples to be analyzed by the reductionaeration method may require prior treatment to convert all mercury to Hg(I1). In this study samples were digested by treatment with 6 ml of 1:4 HpS04-HN03and 2 ml each of potassium permanganate and potassium peroxydisulfate solutions. All such digestions were carried out overnight. Prior to analysis, the samples were transferred to the reaction flask, excess oxidant was removed with hydroxylamine hydrochloride solution, and the total volume was adjusted to 50 ml. Three milliliters of stannous chloride was added through a rubber septum at the top of the flask to reduce the mercury to its elemental state. An argon flow of 1.5 l./min was started and allowed to continue for 2 minutes, at which time the flow was altered to bypass the reduction-aeration flask and flush the system of foreign gases. The gas flow was then turned off and the amalgamator resistance heater turned on. After 35 seconds, the argon flow was again started, and the heater was turned off as mercury was flushed from the amalgamator and into the fluorescence cell. The resulting fluorescence signal was recorded and the peak height was taken as a measure of the intensity of fluorescence. The amalgamator was allowed to cool before additional samples were run, and the cycle time for the entire process was approximately 9 minutes. Solid samples were analyzed by the furnace technique. Samples were weighed into an 11-mm o.d., 1-mm wall thickness fused silica tube, approximately 6 cm in length, and placed in the center of the furnace tube. The furnace tube was placed in the gas train, and the flow of gas started.
Effect of Carrier Gas. Argon has been used as the carrier gas for fluorescence measurements to take advantage of its low cross section for quenching of fluorescence. Measurements were also made using air as the carrier gas, and it was found that the enhancement in the fluorescence signal obtained by use of argon is approximately a factor of 100. The sensitivity obtained with air was acceptable for many purposes. However, with the present system it is no more difficult to use argon than it is air, and the very substantial increase in signal which is obtained with argon warrants its use. Analytical Curves. Figure 2 shows a typical recorder tracing of the atomic fluorescence signals. The peak height was used as a measure of the fluorescence intensity. Blank and scatter signals were negligible under the conditions of the experiment, and it is apparent from Figure 2 that the noise level was also very low. Figure 2 points out one of the advantages of the open amalgamator system compared to the closed system previously employed ( I ) : with the open system the mercury vapor enters the fluorescence cell as a “plug” and a larger signal is obtained than with the closed system, in which the mercury vapor is evenly distributed throughout the system. With the same carrier gas (air) and sample concentration, the open system reduction-aeration method gives a signal approximately 10 times that obtained with the closed system. Different methods must be used in preparing analytical curves for the reduction-aeration and furnace methods. Solution standards of mercuric chloride were used with the reduction-aeration method. When standard solutions were run, the digestion step was omitted; but otherwise the procedure was identical to that employed for running samples by the reduction-aeration method. Mercury vapor standards were used to prepare analytical curves for the furnace method. Air in a polyethylene container was saturated with mercury vapor at room temperature. Known volumes of air were drawn from the container by syringe and injected, through a rubber septum, into an argon stream flowing into the amalgamator. Knowledge of the container temperature allowed calculation of the nanograms of mercury per cubic centimeter of air from tabulated values of the vapor pressure of mercury. The vapor pressure of mercury at room temperature is such that the volumes to be transferred by syringe are
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Table 11. Comparison of Results for Two Rock Samples Hg found, ppm This Sample study AF AA NA" w-1 0.18 0.18 ( I ) 0.18 (9) 0.17 (IO) GSP-1 0.015 0.01s ( I I ) 0.021 (11) a Neutron activation analysis.
Nanograms Hg
Figure 3. Comparison of analytical curves obtained by the reduction-aeration and vapor pressure methods A . vapor pressure method B . reduction-aeration method
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Figure 4. Analytical curve for furnace method Error bars are for f l standard deviation as computed from three replicates in a convenient range. For example, at 20 "C, the conversion factor is 13.1 ng of Hg per cma air. Analytical curves obtained by the two methods are shown in Figure 3. Despite the differences in the calibration techniques for the reduction-aeration and furnace methods, there is good agreement between the two methods. Direct comparison of the two methods was made for samples of U. S. Geological Survey Rock Standard W-1. Both the furnace and reduction-aeration method gave a mean value of 0.18 ppm of Hg with similar precision. The greater sensitivity of the furnace technique makes possible the use of considerably smaller samples. For the determination of Hg in the W-1 rock standard, 15 to 30 mg of sample was used for the furnace method and 300 to 600 mg for the reduction-aeration method. This study has been directed primarily toward examination of the furnace technique. Figure 4 shows in greater detail the low concentration range of applicability of the vapor pressure calibration method. The standard deviation of individual measurements is indicated for three replicates at each concentration. From the measurements shown in Figure 4, 220
it was estimated that as little as 0.6 ng of Hg could be quantitatively determined. This amount gave a signal twice the blank value, and the standard deviation for three measurements was in the range of 10 to 20% of the amount present. The minimum determinable amount can probably be further lowered by careful attention to the reduction of blank values, but it seemed unrealistic to pursue this aspect further, since in this range of mercury concentration the minimum determinable amount in a sample will reflect uncertainties in standards and sample preparation more than limitations of the measurement method. Determination of Mercury in Reference Materials and Sediments. The methods developed in this study were applied to the determination of mercury in samples of water, rock, and wheat for which values of mercury content have been reported elsewhere. The water sample was a Federal Water Quality Administration (Analytical Quality Control Laboratory, Cincinnati, Ohio 45202) reference material with a "true" value of 4.2 ng/ml of Hg. Using the reduction-aeration procedure, a measured value of 4.2 f 0.4 ng/ml was obtained. The mercury content of two rock samples, U. S. Geological Survey W-1 and GSP-1, was determined using both the reduction-aeration and furnace techniques. Pooled results for ten determinations on W-1 and four on GSP-1 are compared with values reported elsewhere in Table 11. Mercury treated wheat flour samples (International Atomic Energy Agency, Code 66/10) offered an opportunity to test the furnace technique for recovery of mercury present in an organic matrix. Addition of oxygen to the argon stream during the combustion and inclusion of the lead acetate filter between the furnace and amalgamator were essential to successful application of the furnace technique to the wheat samples. Three replicates, with sample sizes ranging from 18 to 30 mg, gave a mean value of 5.1 ppm with a standard deviation of 0.5 ppm. Data accompanying the wheat flour sample report the mercury content to be 4.59 + 1.32 ppm. This value is a pooled mean of neutron activation analysis data from 15 laboratories. A value of 4.92 + 0.45 ppm has been reported (12) for an atomic absorption determination. In addition to the reference materials, a large number of mercury determinations have been performed for natural sediment samples of high organic content. Initial attempts to apply the furnace technique to these samples proved unsuccessful, and the difficulty was ascribed to the chloride content of the samples. The interference was eliminated by placing a second amalgamator in the gas train. The upstream amalgamator (Le., the one nearer the furnace) was (9) W. R. Hatch and W. L. Ott, ANAL,CHEM., 40,2085 (1968). (10) M. Fleisher, Geochim. Cosmochim. Acta, 29, 1271 (1965). ( 1 1 ) F. J. Flanagan, ibid., 33, 81 (1969). (12) J. F. Uthe, F. A. J. Armstrong, and M. P. Stainton, J . Fish. Res. Bd. Can., 21, 805 (1970).
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operated at approximately 400 "C during the combustion process and removed the interfering species from the gas stream without hindering the progress of mercury vapor into the downstream amalgamator. A similar system for removal of the chloride interference has been described by Joensuu (13). For a number of sediment samples, mercury determinations have been carried out by three methods: furnace technique AF, wet digestion (reduction-aeration) AF, and wet digestion (reduction-aeration) AA. Within the precision of the methods, identical Hg concentrations were found by all three methods. For example, duplicate determinations by each of the methods for a representative sediment sample gave a mean Hg concentration of 0.28 ppm with a standard deviation for the six determinations of 0.03 ppm. Sample sizes were approximately 1 gram for the AA method, 0.7 gram for the wet digestion AF method, and 0.3 gram for the furnace A F method. Nondispersive System. Although a monochromator was employed for the preliminary studies and the measurements so far reported in this study, in the optimized system the monochromator serves no useful function, since all the fluorescence occurs at a single wavelength, 2537 A, and the intensity of scattered radiation is very small at all wavelengths within the spectral response of the detector. In order to test the utility of a nondispersive system the monochromator was removed, and the fluorescence cell was placed directly in front of the aperture of the multiplier phototube module (Heath EU-701). The field of view of the detector was limited by placing a disk with a I-mm opening in the detector module aperture. The performance of this system was in all respects equal to that of the dispersive system, and the non______ (13) 0.Joensuu, Appl. Specfrosc., 25, 526 (1971).
dispersive system is now in routine use for the determination of mercury in natural sediment samples with the furnace technique. CONCLUSIONS
The present measurement system has several attractive features which make its use advantageous when compared to the closed system atomic fluorescence approach ( I ) . With the amalgamator nearly all the mercury vapor is presented in the fluorescence cell at the same time. The closed system inevitably distributes the mercury vapor over a relatively large volume, and thus only a fraction of the mercury vapor is present in the fluorescence cell at any one time. A factor of ten increase in signal is obtained due to the increased concentration of mercury in the fluorescence cell when the amalgamator is used. With the closed system, it is difficult to arrange to use a carrier gas other than air, but with the amalgamator argon, or other gases, can conveniently be used. Replacement of air by argon results in approximately a 100-fold increase in the fluorescence signal. The amalgamator serves to separate mercury from potential interferants. The use of the furnace technique would be much more difficult if the amalgamator were not used. With the amalgamator the amount of sample required for a determination is reduced. With the furnace technique, little or no sample preparation is required for solid samples. Thus, compared to the reduction-aeration technique, the time required for an analysis is reduced and the likelihood of contamination is lessened. RECEIVED for review July 6, 1971. Accepted September 29, 1971.
An Automated Stopped-Flow Spectrophotometer with Digital Sequencing for Millisecond Analyses P. M. Beckwith and S. R. Crouch Department of Chemistry, Michigan State Uniuersity, East Lansing, Mich. 48823
A com pletely automatic stopped-f low spectrophotometer is described which features a vertical flow system to minimize problems with air bubbles, pneumatically actuated valves for directing the liquid flow and dispelling waste solutions, and a spring-loaded stopping syringe. The entire operating cycle of the system is controlled by a digital sequencing system or by manual switches. The dead time of the flow system was determined to be 5 + 1 msec. Mixing was also found to be complete -5 msec after stopping the flow. Kinetic determinations of Fe(lll) using the Fe(lll)-SCNreaction and of phosphate using the formation reaction of 12-molybdophosphoric acid are presented. Approximately 1000 phosphate samples can be analyzed per hour using the stopped-flow system. STOPPED-FLOW SPECTROPHOTOMETRY has become an increasingly popular technique for fundamental studies of rapid reactions. In addition, the stopped-flow technique has been recently shown to have considerable potential for quantitative analytical determinations based upon rapid
reaction rate measurements (1-3). One of the more attractive features of the stopped-flow technique is the extremely short analysis time which can be achieved using moderately rapid reactions. To take full advantage of the potential of stopped-flow methods for rapid analyses, it is highly desirable to automate the sample handling and mixing operations as well as the data acquisition and evaluation steps. A noteworthy achievement in automating the latter steps has been reported by Willis et al. ( 4 ) and Desa and Gibson (9,who have described on-line computer systems for the processing of stopped-flow data. (1) A. C. Javier, S. R. Crouch, and H. V. Malmstadt, ANAL. CHEM., 41, 239 (1969). (2) J. B. Pausch and D. W. Margerum, ibid., p 226. (3) D. W. Margerum, J. B. Pausch, G. A. Nyssen, and G. F. Smith, ibid., p 233. (4) B. G. Willis, J. A. Bittikofer, H. L. Pardue, and D. W. Margerum, ibid.,42, 1340 (1970). ( 5 ) R. J. Desa and Q. H. Gibson, Comput. Biomed. Res., 2, 494
(1969).
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