A necessary prerequisite for this approach is the availability of retention indices of the compounds to be searched for. Compilations like those discussed by Schupp and Lewis (25), already in a computer-compatible format, should be particularly suited. In our laboratory, a collection of retention indices of drugs and metabolites has been accumulated by the routine use of the described computer program GCRET during the past year, which is now used, in combination with a collection of mass spectra, for the automatic identification of drugs in body fluids of comatose patients (26). Detailed examples of this technique will be discussed in a separate publication (27). Because of the predictability of the retention indices of polyamino
alcohols (see above), this approach should also be readily applicable to the characterization of complex mixtures of these oligopeptide derivatives for the purpose of automation of protein sequencing. Although this paper has described the application of a computer program to rather specific areas in biochemical and biomedical analysis, this program is flexible enough to be of general use for the identification of any compounds which are amenable to gas chromatography.
(25) 0. E. Schupp I l l and J . E. Lewis, Res./Deve/op., 21 (5),24 (1970). (26) H. Nau and K. Biemann, submitted to Anal. Lett. (27) C. E. Costello, H. S. Hertz, T.Sakai, and K. Biemann, submitted to Clin. Chem.
Received August 6, 1973. Accepted October 23, 1973. Financial support from the U S . Public Health Service, National Institutes of Health (Grant No. GM 05472 and RR 00317) is gratefully acknowledged.
ACKNOWLEDGMENT The authors are indebted to Catherine E. Costello for accumulating the data presented in Figure 3.
Mercury Determinations in Natural Waters by Persulfate Oxidation James J. Alberts, James E. Schindler, and Richard W. Miller Department of Zoology, University of Georgia, Athens, Ga, 30602
Peter W. Carr Department of Chemistry, University of Georgia, Athens, Ga, 30602
Glooshenko ( I ) , and Coyne and Collins (2) have shown that aqueous solutions of mercury do not store well in glass or plastic containers. Since most field sampling involves sample storage and transportation times which are significant with respect to the loss rate, the conventionally accepted Hatch and Ott (3) method for sample digestion and analysis may exhibit negative errors due to the loss of mercury(I1). This loss is not inherent in the method of Hatch and Ott, but is an integral part of any analysis method which requires storage of the mercury sample and subsequent transfer of an aliquot to an analysis vessel. Additionally, analytical techniques involving the basic digestion procedures of the Hatch and Ott method suffer from inability to store large numbers of digested samples for long periods of time prior to analysis and require considerable time per analysis due to the large volume of the analysis chambers. Omang ( 4 ) employs an open system t o shorten analysis time; however, the requirements of a 1liter sample and a 24-hour digestion with acid-permanganate limits the usefulness of this method in the field. This report describes a procedure for determining mercury a t the parts-per-billion (ppb) level in aqueous solution which utilizes an open analysis system, requires small ( 1 ) W. A . Giooshenko. J . Phycol., 5,224 (1969). (2) R. V. Coyne and J. A . Collins, Anal. Chem.. 44 1093 (1972) (3) W. R. Hatch and W. L. Ott. Anal. Chem., 40, 2085 (1968). (4) S. H. Omang, Ana/. Chim. Acta, 53, 415 (1971).
434
sample volume, is easily handled in the field and during transportation, has little loss of mercury during storage, allows oxidation at elevated temperatures, has improved analysis time, and ensures that the sample is sealed until opened in the analysis system, thereby releasing all of the mercury present as the metallic vapor. The method reported is an adaption of the Menzel and Vaccaro (5) ampoule method for determination of dissolved organic matter in natural waters, involving the oxidation of organic matter by persulfate in an ampoule sealed under nitrogen and heated to 120 "C. Persulfate has been used as an oxidant for organomercurials (6) and in conjunction with strong mineral acids-ie., " 0 3 , HzSOa-for the determination of mercury in soils and sediments (7, 8). However, these procedures do not expose the samples to temperatures in excess of 60 "C, and are subject to adsorption losses since the mercury is associated with solids rather than in aqueous solution. EXPERIMENTAL Materials. A l l chemicals used in t h i s work were reagent grade a n d a l l solutions were prepared in glass d i s t i l l e d water. Glassware
(5) D. W. Menzel and R. F. Vaccaro. Limnol. Oceanogr., 9, 138 (1964). (6) Environmental Protection Agency, Cincinnati, Ohio, No. 16020-07/71 (1971) . (7) I. K. Iskandar, J, K . Syers. L. W. Jacobs, D. R. Keeney, and J. T. Gilmour. Analyst (London), 97, 388 (1972). (8)J. R. Melton, W. L. Hoover, and P. A. Howard, Soil Sci. SOC.Amer. R o c . 35, 850 (1971).
A N A L Y T I C A L C H E M I S T R Y , VOL. 46, NO. 3, M A R C H 1974
T
I /
I-1
Figure 1. Analysis system
Tygon tubing leading from an air pump to a 9-inch, movable cannula; B, leading to absorption cell was washed with concentrated nitric acid, rinsed with distilled water, dried a t 120 "C and stored in the drying oven. The samples were contained in Kimax glass ampoules (Kimble Glass, 12012L-10) of 10-ml volume. Prior to filling, the ampoules were fired at 500 "C for 3 to 4 hours. During this period and until filled and sealed, the mouths of the ampoules were covered with aluminum foil. Ampoules prepared in this fashion show very low blanks for a period of a t least several months. Equipment. All measurements were carried out using a PerkinElmer Atomic Absorption Spectrophotometer (Model 303) with a Perkin-Elmer Flameless Mercury System modified as shown in Figure 1. The per cent absorption-time curves were recorded on a Perkin-Elmer strip chart recorder (Model 56) set on the 10-mV range and chart speed of 20 mm/min. The recorder was interfaced to the spectrophotometer via a Perkin-Elmer Automatic Null Recorder Readout Unit (303-0103). The spectrophotometer was operated a t the manufacturer's suggested settings for mercury. Procedure. Approximately 170 mg of potassium persulfate (KzSzOs) was placed in the ampoule followed by 5 ml of sample, which was introduced by a long tip, 10-ml graduated Mohr pipet, and 0.2 ml of 3% (v/v) phosphoric acid. The ampoule was sealed under air within 1 hour of filling and autoclaved for 45 minutes a t 120 'C within 1.5 hours to 3 days after sealing (Table 11). The actual analysis was initiated by placing the ampoule in the Tygon tubing sleeve shown in the insert for Figure 1. The glass neck was crushed with a pliers. Immediately after the seal was broken, 0.5 ml of 10% (w/v) stannous chloride was introduced uia a syringe and the purge cannula from the Flameless Mercury System air pump inserted into the solution. The mercury vapor was swept into a 10-cm absorption cell equipped with plastic end windows and was vented directly to a fume hood. The purge air was supplied at a flow rate of 3 l./min and the volume of the system from sample to absorption cell was approximately 56 cm3.
A,
RESULTS AND DISCUSSION Figure 2 represents the recorder response (per cent absorption us. time) for the present technique and the meth-
Ampoule B r e a k e r ( 5 4 4 insert1
I-ml disposable syringe with 4-inch movable cannula; C. Tygon tubing exit
od of Hatch and Ott. This figure illustrates that peak height is quite reproducible from sample to sample and that the sample transfer time is much shorter than in the method of Hatch and Ott which employs a large volume sample container. If mercury is eliminated from the sample a t a rate controlled by convective diffusion, then the shortened analysis time is due, a t comparable flow rates, to the decreased volume of the sample, and the decrease in the volume of the entire gas system. The decrease in analysis time decreases gas phase broadening of the sample and provides larger peak signals than the Hatch and Ott procedure without recycling. Calibration curves prepared with mercuric chloride are shown in Figure 3. The absorbance values on Figure 3A were obtained at a noise filter setting of 2 and attenuation of X1 on the recorder interface. At this setting 100 ppb mercury will give -0.730 absorbance unit. Similarly, the absorbance values on Figure 3B were obtained at a noise filter setting of 2 and attenuation X3. Table I represents the precision of the ampoule technique. It should be noted that subsequent standard solutions of 10 ppb mercury concentration prepared with plastic, disposable, 10-ml graduated Serological pipets (Falcon Plastics, No. 7530) gave a per cent relative standard deviation of 2.6 for five replicate determinations. Therefore, plastic pipets may be used for sample preparation in the field with little concern for loss of precision. Table I1 represents the recoveries of several forms of mercury from solutions of varying composition. Samples which had been autoclaved and stored for a period of 4 weeks show absorbance values significantly less than those of freshly prepared standards. Since this deviation did not manifest itself until after the 3rd week of storage, it appears that this method of storage is effective for a period
A N A L Y T I C A L C H E M I S T R Y , VOL. 46, NO. 3, M A R C H 1974
435
Figure 2. Recorder response in per cent absorption ws. time A, Response of Hatch-Ott method for 10 ng Hg and 50 ng Hg; B. Response of ampoule technique for 5 ng Hg to 50 ng Hg
(Y
0
0 c n L
0 0 0
a
0
10
20
30 4 0
50
60
70
BO
90
ppb
He
100
0
2
1
6
B
10
Figure 3. Standard curves for ampoule mercury technique in absorbance units ws. concentration in ppb H g A , Curve for 0 to 100 ppb Hg at recorder attenuation X 1 and noise suppression 2; 8. Curve for 0 to 10 ppb Hg at recorder attenuation X3 and noise suppression 2
Table I. Effect of Concentration on Precision Absorbance
70 o b
1 ppb Hg ( 5 ng) 2 ppb Hg (10 ng) 5 ppb Hg (25 ng) 10 ppb Hg ( 5 0 ng)
0.019c 0 . 036c 0.090c 0 . 21gC
4.3 2.3 2.7 2.5
10 ppb Hg ( 5 0 ng) 20 ppb Hg (100ng) 5 0 ppb Hg (250 ng) 100 ppb Hg (500 ng)
0 .055d 0 . 116d 0 .2 m d 0 . 614d
2.2 6.0 6.0 3.6
Concentrationa
a Sample is mercuric chloride, concentration in solution (absolute quantity). Relative standard deviation based on a minimum of four measurements per concentration. Recorder interface at attenuation X3. d Recorder interface at attenuation X1.
*
of at least 2 weeks without significant loss of sample. The reason for this extension of the storage period is not fully known. It is possible that the oxidation procedure with persulfate affects the hydrated layer of the glass and prevents penetration of the mercury into this layer. Also, the 436
A N A L Y T I C A L CHEMISTRY, VOL. 46,
NO. 3,
fact that the ampoules are sealed until introduction of reducing agent in a closed system may limit the amount of mercury which is lost through volatilization. Studies were conducted to investigate the effect of other dissolved species upon the analysis, Table 11. Sodium salts of chloride, nitrate, and sulfate were added to 10-ppb mercury standard solutions to give various concentrations of interfering ion up to 35 parts per thousand (ppt). Similarly, dextrose to a concentration of 50 parts per million (ppm) carbon was added to 10-ppb mercury standards to determine the effect of oxidizable organic matter on the method, and mercury standards were made using water from a farm pond near the laboratory. Nitrate, sulfate, dextrose, and natural pond water did not affect the determination even at the maximum concentrations used in these experiments. However, chloride above 20 ppm showed a reduction of the peak heights, with complete elimination of the signal at approximately 200 ppm C1. This interference was eliminated in the case of sea water which had been refluxed in an open system with excess persulfate for a period of 4 hours prior to the addition of mercury salts. During the reflux period, the presence of
M A R C H 1974
Table 11. Recovery of Mercury from Various Sources
Mercury source
1. HgClz 2. Methyl mercuric
Concentration, ppb H g
10 5 10
Media
HzO H?O
Relative responsea
Hz0
1.00 1.00 0.97
HZO
1.06
HzO; samples stored sealed for 3 days before autoclaving HzO; samples stored sealed for 3 days before autoclaving HeO; samples stored for 3 weeks after autoclaving H,O; samples stored for 3 weeks after autoclaving H,O; samples stored for 4 weeks after autoclaving H 2 0 ; samples stored for 4 weeks after autoclaving HzO; 50 ppm carbon as dextrose HZO; 35 ppt Nosas NaN03 H,O; 35 ppt S01’as Na2SOi H?O; 20 ppm C1as NaCl Natural pond water
1.00
acetate 3. HgClz
5 10 5
4. HgClz
10 5
5. HgClz
10 5
6 . HgClz
10
7. HgC12
10
8. HgClz
10
9. HgClz
10
10. HgClz
10
0.98
1.02 0.95 0.91 0.84 1.00
0.94 0.97 0.98 0.98
Defined as relative to mercuric chloride in distilled water.
chlorine gas was noticed escaping from the open end of the reflux condenser. It appears that the persulfate oxidation causes chloride to be converted to chlorine gas which is trapped in the ampoules, This gas occupies the head space of the ampoules and is released into the flow system when the ampoule is broken for analysis. The entrained chlorine then oxidizes the elemental mercury which is swept out of the ampoule after addition of stannous chloride. The oxidized mercury is no longer volatile and settles out on the walls of the tubing. Any mercury in higher oxidation states
which may reach the absorption cell will not be read since the instrument is set to detect absorption of the 253.7-nm elemental mercury line. To see if the head gas could be eliminated, samples of 10 ppb mercury with concentrations of chloride >50 ppm were placed in the ampoule breaker and shaken vigorously just prior to opening in an attempt to supersaturate the liquid phase with chlorine. This treatment restored the mercury peak height to the expected level for concentrations of chloride to 200 ppm. A further attempt to reduce the gas before reducing the oxidized mercury in the sample involved injection of 0.5 ml, 10% SnCl2 above the unbroken ampoule in the ampoule breaker. This treatment, followed by vigorous shaking, allowed extension of the determination to 500 ppm C1. However, above this level, further attempts a t eliminating the excess chlorine by means of SnCl2 saturated glass, wool, and SnC12 liquid traps placed downstream of the ampoule failed to prevent the reduction in peak height with increase in concentration of dissolved chloride. The mercury signal was eventually totally eliminated at 5 ppt C1. Since fresh water chloride concentrations are usually below 12 ppm C1 ( 9 ) , the method will have significant interferences in estuarine and oceanic waters. The procedure described represents a simple and rapid (between 30 and 40 samples at the 10-ppb level may be run in an hour) technique of determining mercury in natural waters. It also alleviates the problems of storage and sample handling attendant with other methods of mercury analysis. It should be emphasized that the procedure has been investigated only in aqueous samples. Unpublished data from these laboratories indicate that samples containing solid-liquid phase boundaries-i. e., sediment and tissue-undergo only partial oxidation of the organic carbon under these experimental conditions. Therefore, these latter samples may be more profitably analyzed with other existing methods. Received for review April 11, 1973. Accepted October 4, 1973. This project has been financed in part with Federal funds from the Environmental Protection Agency under grant No. R-800427. The contents do not necessarily reflect the views and policies of the Environmental Protection Agency, nor does mention of trade names or commercial products constitute endorsement or recommendation for use. (9) D. A. Livingston, U.S.Geol. Sur. Prof. Paper 440-G (1963).
Direct X-Ray Spectrometric Determination of Bromine in Water Yoetz Deutsch Geochemistry Department, Geological Survey of Israel, 30 Malkhei lsrael St., Jerusalem, lsrael
A rapid X-ray spectrometric method for the routine determination of bromine in water was investigated. Once calibrated, the procedure developed requires no standards and is simple, rapid. precise, and at least as accurate as the conventional “wet chemical” method (
