'I
The accuracy of the column RIA method was also tested by analyses of 179 clinical specimens. These samples were assayed previously by a dextran-coated charcoal technique. A comparison of the results from the two assays is given in Figure 3. The correspondence bitween the two sets of data is very satisfactory, the correlation coefficient was 0.99. These results demonstrate that the column RIA procedure is a valid analytical method for the quantification of digitoxin in clinical specimens. It is rapid and possesses acceptable precision, sensitivity, and accuracy. In addition, the method is convenient because both the incubation and separation functions are incorporated into a single device, thus minimizing transfers.
A/' : r = 0.99 SAMPLES
In
ACKNOWLEDGMENT
I
1
I
I
n
I
1
1
10
20
30
40
50
60
70
80
DI GI TOXl N (ng/ml) DEXTRAN-CQ9TED CHARCOAL PROCEDURE Correspondence of results obtained with the column procedure and the dextran-coated charcoal method Figure 3.
The authors are grateful to Mrs. Betty Byers of the South Bend Medical Foundation for her kind assistance in sudplying the clinical specimens and reference values.
LITERATURE CITED RIA
patients on digitoxin therapy is immunologically indistinguishable from the digitoxin in the standards. In other words, the antibody recognizes the digitoxin in clinical specimens and standards as identical. As shown in Table 11, acceptable results were obtained in a parallelism study over a sixfold range of dilution. These data indicate that various components in the serum, including potentially cross reacting materials, do not interfere in the test. Accuracy. The accuracy of the assay was evaluated by determining the recovery of exogenous digitoxin added to clinical specimens. Assays were performed on specimens before and after the addition of aliquots of a normal plasma containing known amounts of added digitoxin. The amount of digitoxin recovered from each specimen was then calculated. The results of this analysis given in Table I11 indicate acceptable recovery.
(1)S. Shapiro, D. Slone, G. P. Lewis, and H. Jick, J. Chronic DIs., 22, 361 (1969). (2)I. J. Giuffra and H. L. Tseng, N. Y. State J. Med., 52, 581 (1952). (3)P. L. Rodensky and F. Wasserman, Arch. htern. Med.. 108,171 (1961). (4)A. Schott, Postgrad. Med. J., 40, 628 (1964). (5) M. S.Gotsman and V. Schrire, S.Afr. Med. J., 40,590 (1966). (6)D. W. Duhme, D. J. Greenblatt. and J. Koch-Weser, Ann. Intern. Med., 80,516 (1974). (7)T. W. Smith and E. Haber, Pharmacol. Rev., 25, 219 (1973). (8)J. E. Doherty, W. H. Perkins, and W. J. Flanigan Ann. Intern. Med., 66, 116 (1967). (9)G. C. Oliver, Jr.. B. M. Parker, D. L. Brasfield, and C. W. Parker, J. CIin. Invest., 47, 1035 (1968). (IO) T. W. Smith, J. PharmacoI. Exp. Ther., 175, 352 (1970). (11) S. J. Updike, J. D. Simmons. D. H. Grant, J. A. Magnuson, and T. L. Goodfriend, CIin. Chem., 19, 1339 (1973). (12)T. W. Smith, V. P. Butler. Jr., and E. Haber, Biochem., 9, 331 (1970). (13)V. P. Butler, Jr., and J. P. Chen, Proc. Nat. Acad. Sci. USA, 57, 71 (1967). (14)J. Feldman. and D. Rodbard, "Principles of Competitive Protein-Binding Assays", W. D. Odell and W. H. Daughaday, Ed., Llppincott Co., Philadelphia, Pa., 1971,p 158. (15) C. D. Hawker, Anal. Chem., 45, 878 A (1973).
RECEIVEDfor review February 28,1975. Accepted April 21, 1975.
Determination of Arsenic by Anodic Stripping Voltammetry and Differential Pulse Anodic Stripping Voltammetry Gustaf Forsberg, Jerome W. O'Laughlin, and Robert G. Megargle' Department of Chemistry, University of Missouri, Columbia, Mo. 6520 7
S. R. Koirtyohann Environmental Trace Substances Center, University of Missouri, Columbia, Mo. 6520 7
The determination of arsenlc by anodic stripplng voltammetry (ASV) and differential pulse anodic strlpplng voltammetry (DPASV) was Investigated. Factors affectlng sensitivity and precislon included pH, deposition potential, supporting electrolyte, and the nature of the working electrode. The detection limit of both DPASV and ASV was 0.02 ng/ml. The Present address, Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115. 1586
ANALYTICAL CHEMISTRY, VOL. 47, NO. 9, AUGUST 1975
sensitivity increased with Increasing acid concentration and one molar solutlons of either hydrochloric or perchloric acids were most suitable as supporting electrolytes. Gold was found to be superlor to platlnum as a worklng electrode material. The most satisfactory procedure for reduclng arsenlc(V) to arsenic( Ill), whlch Is necessary because arsenic( V) Is electrolnactlve, Involved heating arsenlc( V) wlth NaZS03 In concentrated acid solutions.
Many classical methods for the determination of arsenic such as the Marsh ( I ) and Gutzeit ( 2 ) methods require relatively large amounts of arsenic and are totally unsuitable for the detection of arsenic in biological samples in the nanogram t o subnanogram range, a subject of great interest in monitoring arsenic in the environment. Spectrophotometric methods such as t h e molybdehum blue method ( 3 ) and t h e silver diethyldithiocarbamate ( 4 ) procedure are subject t o numerous interferences and d o not have t h e required sensitivity. Normal flame atomic absorption is relatively insensitive t o arsenic ( 5 ) .Procedures have also been developed which involve reducing arsenic t o arsine, then determining t h e arsine by atomic absorption (6-8) or emission (9). Arsenic can be determined at trace levels by neutron activation analysis (10-12), but this requires access t o a nuclear reactor and sophisticated counting equipment. Various electrochemical techniques have been investigated for the determination of arsenic at trace levels. Electrochemical methods have great sensitivity, in some cases approaching t h a t of mass spectroscopy and neutron activation analysis. T h e instrumentation required is relatively simple and generally costs far less than t h a t required for spectrochemical techniques. Another advantage of electrochemical techniques is their ability t o distinguish between t h e different oxidation states of arsenic. Arsenic determinations have been made using classical d c polarography (13,14), oscillopolarography (15), ac polarography (15), square wave polarography (16) and differential pulse polarography (17). Myers and Osteryoung (1 7) report the detection limit for arsenic by the latter method was approximately 0.3 ng/ml. A number of ions, notably those of lead, tin, and thallium were serious interferences, and methods of eliminating these interferences increased the detection limit to 20 ng/ml. Anodic stripping voltammetry (ASV) has seen little use in arsenic determinations, but it is potentially one of t h e most sensitive of all methods and seemed a promising technique t o investigate for trace arsenic determinations. T h e use of ASV t o determine concentrations of arsenic as low as 2 x 10-'M has been reported (18). Recent advances in instrumentation and the development of such techniques as differential pulse anodic stripping voltammetry (19) (DPASV) suggest, however, that considerable improvements in sensitivity and accuracy could be easily achieved. T h e present paper is a report on the determination of arsenic by ASV and DPASV. Parameters likely to affect sensitivity and accuracy were investigated including pH, deposition potential, supporting electrolyte, the nature of the working electrode, and t h e reduction of arsenic(V) t o arsenic(II1). T h e effect of a number of ions which might interfere with the determination of arsenic is also reported.
EXPERIMENTAL Reagents. Stock solutions of arsenic(II1) and arsenic(V) were prepared by dissolving reagent grade As203 and NasAsOd in deionized water with the required amount of NaOH then acidifying with HC1. Subsequent dilutions of these solutions were made as required. When reagent grade hydrochloric and nitric acids were used as supporting electrolytes for analyzing for arsenic in concentration of 1@M or less, they were further purified by electrodeposition or distillation. Triply distilled perchloric acid available from the G. Frederick Smith Co. proved to be the purest electrolyte available. In 1M concentration, it was used as the supporting electrolyte in analyzing for arsenic in concentration as low as 0.02 ng/ ml. All other chemicals were reagent grade and used without further purification. Apparatus. Two different stripping analysis instruments were used in the research. A PAR (Princeton Applied Research Inc.) Model 174 Polarographic Analyzer was used for the experiments involving DPASV and some of the later experiments involving ASV. A semi-automatic stripping analysis instrument was used for
most of the experimental work involving ASV. Its design and operation have been described earlier (20). A variety of cells were used. In each type, the electrodes were mounted in a Teflon cell cover constructed to fit the cell in use. The initial design used a cell of 100-ml capacity with 70 ml being the normal volume of solution used. The solution was stirred by a motor-driven glass impeller. For subsequent work, 10-ml cells were made from glass or polyethylene. The normal cell solution volume was 5 ml. Agitation was accomplished by bubbling nitrogen through the solution. These smaller cells were used in all research involving DPASV studies. The working electrodes were gold or platinum wires fused into glass tubes. The platinum electrodes were 0.64 mm in diameter and approximately 1 cm in length. The gold electrodes were 0.25 mm in diameter and approximately 1 cm in length. The counter electrodes were platinum and were similar in size and construction to the platinum working electrodes. A Ag/AgCl electrode filled with 1M HC1 was used as the reference electrode. The body of this electrode was a glass tube fitted with a porous Vycor plug. Procedure. This solution to be analyzed was introduced into the cell and deaerated with nitrogen for 10 minutes. The arsenic was then deposited at the selected potential for a predetermined time. When the motor driven glass impeller was used to provide stirring, it was stopped 20 sec before the end of the deposition period. The cell voltage was scanned from the deposition potential to a preselected potential, usually about 0.4 to 0.5 volt more positive than the peak potential of the arsenic oxidation peak. Voltammograms were recorded using an x-y recorder. For ASV experiments, voltage scan rates of 50 and 100 mV/sec were used. Lower scan rates afforded greater peak resolution but smaller recorded areas. Apparent sensitivity (or peak area) was directly proportional to the voltage scan rate. Scan rates of about 200 mV/sec or greater, require the presentation of the voltammogram on an oscilloscope and were not used. For DPASV experiments, a voltage scan rate of 10 mV/sec, a pulse amplitude of 50 mV, and a measurement frequency of 0.5 sec were used. A thorough discussion of the effects of varying DPASV instrumental parameters is available in the literature (21). The area of the arsenic oxidation peak was directly proportional to the concentration of arsenic. The area of the oxidation peak was usually determined by triangulation. In instances where the shape of the oxidation peak made area determination by triangulation difficult, the peak area was determined by cutting out the chart paper under the peak and comparing its weight with the weight of a paper of known area. The concentration of arsenic was determined by the use of either calibration curves or the standard addition technique where known amounts of a standard solution of arsenic(II1) were added and the increase in peak area was observed. The work of Trushina and Kaplan (18) appears to indicate that the height of the arsenic oxidation peak is directly proportional to the concentration of arsenic, but this was not pursued in this research. Additions to the cell solution were made with Eppendorf micropipets. RESULTS AND DISCUSSION Working Electrodes. An attempt was made t o use a hanging mercury drop working electrode. This proved unsuccessful. T h e arsenic oxidation peak appeared as a shoulder on the mercury oxidation wave and was of little analytical utility. Silver working electrodes were unsuitable for similar reasons. Platinum was a suitable electrode material and was employed in t h e initial studies. Subsequently, gold working electrodes were found to have some advantages over platinum and were substituted for platinum. Gold has a higher hydrogen overvoltage than platinum. This reduces the problem of simultaneous evolution of hydrogen while depositing arsenic. With gold working electrodes, it was possible t o use a deposition potential for arsenic a t which adsorbed films of hydrogen did not form on the electrode as evidenced by the lack of a n anodic wave from the oxidation of adsorbed hydrogen. This was usually not the case with platinum working electrodes. T h e large anodic waves from t h e oxidation of the adsorbed hydrogen films from the surface of the platinum working electrode seriously interfered with the analysis of very dilute arsenic solutions. With more concentrated arsenic solutions, enough arsenic was deposited on the working electrode ANALYTICAL CHEMISTRY, VOL. 47, NO. 9, AUGUST 1975
1587
T
1
25 ua
I 1
t1 -
A
I tl
13
I
15
i 7
E I vOlti v s AglAgCI electrode)
Figure 1. ASV voltammograms ( A ) 1M HCI and (B) 1 fig/ml As(ll1) in 1M HCI. pt electrode, I-min plating time, 50 mV/sec scan rate, 0.8 pA/mm
during the deposition step to largely prevent the buildup of a film of adsorbed hydrogen. Another advantage of gold electrodes is that they yield higher and sharper oxidation peaks. This was attributed to enhanced reversibility of the electrode reaction in both the plating and stripping step. Cyclic voltammetry studies of gold and platinum electrodes supported this assumption. A substantially larger separation of the anodic and cathodic peaks in the cyclic voltammograms of the arsenic(II1)-arsenic(0) couple was observed when platinum working electrodes were used in place of gold working electrodes. There are serious difficulties, however, associated with the use of either gold or platinum working electrodes. Their response is very strongly dependent upon past history, pretreatment, and the formation of oxide films. Oxide films have been shown to greatly alter the kinetics of electrode reactions ( 2 2 ) .Precise, carefully controlled pretreatment of the electrodes is necessary to assure reproducible electrode response. A number of pretreatment procedures have been developed and are discussed at length in the literature (23). Several of these pretreatment procedures were evaluated in this work. The most effective approach involved storing the electrodes in 6M "03 until just before use to ensure a thoroughly oxidized electrode surface. The electrodes were then thoroughly rinsed in distilled water. They were then allowed to stand in air-free 1M HC104 a t 0.0 volts until the current decayed to a small fraction of its initial value. This procedure will remove most of the oxide film from the electrode surface and help assure reproducible response. T o avoid the buildup of oxide films while using the electrodes, the cell potential should not be scanned to potentials where gold or platinum oxidize. In general this means not letting the cell potential exceed +0.7 volt relative to the Ag/AgCl reference electrode. Reduction of Arsenic(V) to Arsenic(II1). In samples which have undergone the customary wet ashing procedures with strong oxidizing agents such as nitric and perchloric acids, any arsenic present will be in the pentavalent oxidation state. Because arsenic(V) is not electrolytically reducible in most supporting electrolytes (24, 25), the reduction of arsenic(V) to arsenic(II1) has been extensively studied (25, 26). Some of the reductants which have been used include potassium iodide, cuprous chloride, hydrazine sulfate, and hydroxylamine hydrochloride. For the purpose of determining arsenic by ASV, the reduction problem is A quantitative reduction of arsetwofold. nic(V) to arsenic(II1) is desired, and excess reductant or the byproducts of the reduction reaction should not inter1588
ANALYTICAL CHEMISTRY, VOL. 47, NO. 9. AUGUST 1975
I
::i '0
1
11
I 2
I
3
I
1
I
I
I
I
i
4
5
6
7
8
9
10
I
PH
Figure 2. Variation of peak area with pH Pt electrode, 1 fig/ml As(lll), -0.7-V deposition potential, 1-min plating time, 50 mV/sec scan rate, 0.2 NA/mm
fere with the determination of arsenic(II1). When such reductants as cuprous ion, iodide ion, or hydrazinium salts were used, their oxidation waves obscured that portion of the current-voltage curve where the arsenic oxidation peak occurred. Attempts to eliminate this problem by extracting the arsenic(II1) into benzene and then back-extracting into a clean supporting electrolyte were unsuccessful. The medium exchange technique, which involved plating arsenic from one solution and stripping it into another, also proved unsuccessful. The most successful reduction procedure involved the use of Na2S03 as the reductant. A solution of arsenic(V) was heated in concentrated acid solutions with Na2S03 for 20 to 30 min at 80 to 100 "C. The acid concentration should be at least 1M and preferrably higher. Heating helps drive the reduction reaction to completion and removes the excess SOz. Blanks were low and the procedure proved quite suitable for the determination of arsenic a t the submicrogram level. Recoveries were determined by comparing peak areas derived from reduced solutions of arsenic(V) with those from standard solutions of. arsenic(II1). For ten different solutions of 1 wg/ml arsenic(V) in 2M perchloric acid, the average percent recovery was 97.2% with a relative standard deviation of 6.9%. S u p p o r t i n g Electrolyte. A variety of supporting electrolytes including sulfuric, nitric, perchloric, phosphoric, and hydrochloric acids were investigated. These preliminary studies showed that 1M solutions of the other acids all afforded approximately the same sensitivity for determining arsenic. The arsenic oxidation peak potential was approximately 0.3 volt and varied depending upon the choice of electrode, electrolyte, and voltage scan rate. Voltammograms of a solution 1M in HCl ( A ) and a solution 1M in HC1 and also containing 1 wg/ml of arsenic(II1) ( B ) are shown in Figure 1. A study was made of the dependence of peak area (sensitivity) on pH. Buffer solutions were used to provide supporting electrolytes of the desired pH and enough arsenic(II1) was added to give an arsenic(II1) concentration of 1 wg/ml. Arsenic was deposited on a platinum electrode for 1 minute then stripped. The results obtained are given in Figure 2. The solutions used for the first four pH values (pH = 1, 2 , 2.5, 3) were prepared from 0.2M KCl and 0.2M HC1 with chloride ion concentration and ionic strength constant. Sensitivity remained nearly constant up to pH 3.5 and decreased markedly at higher pH although the ionic strength remained approximately constant at 0.1M for solutions a t higher pH values. It was concluded that the ionic strength was not a major factor in the variation of
1
-.I
-.3
..5
-.I E lvolts
-.9 VI
-1.1
-1.3
-1.5-1.7
-19
AqlAgCl electrodel
Figure 4. Variation of peak area with deposition potential in HC104 Figure 3. Variation of peak area with concentration of HC104 Au electrode, 0.1 pg/ml As(lll), -0.2-V deposition potential, 4-min plating time, 100 mV/sec scan rate, 0.04 pA/mm
electrode response with pH. The deposition potential of maximum response would also shift in the same direction, and this could also be a factor in the observed decrease in sensitivity. Studies of the variation in sensitivity with deposition potential in electrolytes of different pH showed, however, that in the case of platinum working electrodes using a deposition potential of -0.7 volt minimized the effect this had on sensitivity. These studies will be discussed in a subsequent section. The variation in peak area with the concentration of hydrochloric and perchloric acids was also studied. A series of hydrochloric and perchloric acid solutions with molarities ranging from to 6M were used; no other electrolyte was added. The concentration of arsenic(II1) was 0.1 pg/ml. Arsenic was deposited for four minutes on a gold working electrode a t a potential of -0.2 volt and then stripped. The results of the studies performed in HC104 are presented in Figure 3. A marked decrease in sensitivity with decreasing acid concentration is observed. Similar results were observed for the studies performed in HC1. The polarographic half wave potential for the reduction of arsenic(II1) to arsenic(0) is reported to be a complex function of the acid concentration of the supporting electrolyte (24) tending to shift to more cathodic values with increasing pH. The deposition potential of maximum response would also be expected to shift to more negative values. T h e resultant decrease in plating effiency could be one of the major reasons for the decrease in sensitivity with increasing pH. This was confirmed by studies of the variation in peak area with deposition potential in supporting electrolytes with different acid concentrations. These studies are discussed below. Another factor to be considered is the decrease in ionic strength which has been shown to decrease sensitivity (27). The magnitude of this effect would likely be minor compared to the effect of the shift in the deposition potential of maximum response. Deposition Potential. T h e variation in sensitivity with deposition potential was investigated. Both platinum and gold working electrodes were used as well as a variety of supporting electrolytes. The results obtained using a platinum working electrode and 0.01, 0.1, and 1.OM perchloric acid as the supporting electrolyte are presented in Figure 4. The results obtained with hydrochloric acid of the same molarities were very similar. The concentration of arsenic(II1) was 1 pg/ml. Arsenic was deposited for 1 minute then stripped. The decrease in sensitivity with deposition potentials greater than about -0.7 volt is presumably due to the reduction of some of the deposited arsenic metal to arsine gas. The data obtained for 1M hydrochloric acid cor-
Pt electrode, 1 pg/ml As(lll). 1-min plating time, 50 mV/sec scan rate, 0.4 pA/mm. ( 0 )1M "301. (M)0.1 M HCIO4, (A)0.01 M HCIOl
2. 5
2. 0
I 5 10
5
0 E 'volts
vi
Ag AgCl e'ectridei
Figure 5. Variation of peak area with deposition potential and pH. Pt electrode, 1 pg/ml As(lll), 1-min plating time, 50 mV/sec scan rate, 0.2 wA/mm. ( 0 )pH 4. (4 PH 7,(WpH 10
responded well with the reported half-wave potentials for the reduction of arsenic(II1) to arsenic(0) (-0.43 volt vs. the SCE) and the reduction of arsenic(0) to arsine gas (-0.6 volt vs. the SCE) in 1M HC1 ( 2 4 ) . Using platinum working electrodes in 1M HCl, Trushina and Kaplan ( 1 8 ) reported a curve similar in shape to the curve for 1M HC1 obtained in the present work but with the maximum peak height a t slightly more negative potentials. It can be seen in Figure 4 that the deposition potential of maximum response is shifted to slightly more negative potentials as the acid concentration is decreased. This was also true when hydrochloric acid was used as the supporting electrolyte. This parallels the negative shift of the polarographic half wave potential for the reduction of arsenic(II1) to arsenic(0). The most suitable deposition potential appeared to be -0.5 volt when using a platinum working electrode in these electrolytes. A study was also made of the variation of sensitivity with deposition potential in buffer solutions with pH values of 4, 7, and 10. Again, a platinum working electrode was used, the arsenic(II1) concentration was 1 pglml, and the deposition time was 1 minute. The results are presented in Figure 5 . It can be seen that the deposition potential of maximum response shifts to more negative values with increasing pH. This also parallels the negative shift of the polarographic half wave potential. An examination of Figures 4 and 5 shows that using a deposition potential of -0.7 volt minimizes the effect of the shift of the deposition potential of maximum response when studying the variation in sensitivity in electrolytes of various pH. ANALYTICAL CHEMISTRY, VOL. 47, NO. 9, AUGUST 1975
1589
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---
5
5
4
5 = Y
5
3
2
I O
3
1
- I E ivolls
. I VI
- 5
- 1
Aq'AgCl electrodel
Figure 6. Variation of peak area with deposition potential in HCIO4 Au electrode, 0.1 @g/mlAs(lll), 4-min plating time, 100 mV/sec scan rate, 0.04 kA/mm (0)1M HCIO4, ( m ) 0.1M HCI04, (V)0.01 M HCIO4
Studies of the variation in sensitivity with deposition potential were conducted using a gold electrode and hydrochloric and perchloric acid solutions with molarities of 0.01, 0.1, and 1. The concentration of arsenic(II1) was 0.1 Mg/ml and the deposition time was 4 minutes. The results are presented in Figure 6 for perchloric acid. The results when hydrochloric acid was used as the supporting electrolyte were very similar. I t can be seen that there is a pronounced shift of the deposition potential of maximum response to more negative values as the concentration of acid decreases. The studies of the relationship between sensitivity and concen-, tration of perchloric acid which are shown in Figure 3 as well as similar studies performed in HC1 were performed keeping the deposition potential a t -0.2 volt. I t can be seen from Figure 6 that the shift of the deposition potential of maximum response to more negative values is largely responsible for the decrease in sensitivity with decreasing acid concentration. A sharp decrease in sensitivity with deposition potentials greater than the deposition potential of maximum response can be observed in Figure 6. This is probably due to the formation of a film of adsorbed hydrogen on the surface of the electrode. This film greatly retards the deposition of arsenic. In 1M HC1 and HC104, hydrogen evolution commenced a t a potential of about -0.3 volt a t a gold working electrode compared to a potential of about -0.1 volt a t a platinum working electrode. While deposition potentials greater than -0.2 volt afforded the most sensitivity with gold working electrodes, a rapid decrease in sensitivity apparently due to the formation of a film of adsorbed hydrogen was observed if a number of voltammograms were obtained using the same electrode a t this potential. It was decided to use a deposition potential of -0.2 volt to avoid this problem. The hydrogen overvoltage of gold is higher than that of platinum. Because of this, it is possible to reduce arsenic a t the gold electrode a t potentials where the evolution of hydrogen is much less than a t a platinum electrode a t comparable potentials. This could account for the greater sensitivity of the gold working electrode. The curves of deposition potential vs. sensitivity obtained using a platinum working electrode do not exhibit as sharp a decrease in sensitivity after the deposition potential of maximum response as those sensitivity vs. deposition curves obtained using a gold working electrode. A possible explanation of this might be that because hydrogen evolution occurs a t less negative potentials on a platinum electrode it has, therefore, suppressed the deposition of arsenic for the entire 1590
ANALYTICAL CHEMISTRY, VOL. 47, NO. 9, AUGUST 1975
.I
*I
E ivolts
t 5
VI
i 7
t B
Ag'AgCl electrodel
Figure 7. Enhancement of peak area by the addition of Au (A) 20 pg/ml As(lll), (B)20 kg/ml As(lll) and 2 bg/mi Au(lll). 1M HC104, Pt electrode, 1-min plating time, -0.5-V deposition potential, 50 mV/sec scan rate, 1.6 kA/mm
portion of the sensitivity vs. deposition curve and not just for part of the curve as would be the case when a gold working electrode is used. Sensitivity and Precision. ASV proved to be an exceedingly sensitive technique for determining arsenic. Gold working electrodes were found to be more sensitive than platinum working electrodes for reasons discussed above and were used for the bulk of the sensitivity and precision studies except where specifically mentioned. Using gold working electrodes, the detection limit of ASV for arsenic was 0.020 ng/ml or IO0 pg of arsenic(II1) in 5 ml of solution. The electrolyte used was 1M HC104. A deposition time of only 20 minutes was required. In Table I, data are presented relating peak area to concentration of different concentration levels of arsenic(II1). The relative standard deviation of the method was calculated on the basis of dividing the area of the arsenic oxidation peak by the concentration of arsenic(II1). The average relative standard deviation was 15%or less for the concentration levels of arsenic studied. Using platinum working electrodes, the detection limit for determining arsenic by ASV was 0.14 ng/ml or 10 ng of arsenic in 70 ml of solution. Thirty-minute deposition times were required for this concentration range. With the use of smaller cells, it proved possible to detect 5 ng of arsenic(II1) in 5 ml of solution using a deposition time of twenty-five minutes. The use of DPASV greatly shortened deposition times. With the use of DPASV and platinum working electrodes, it proved possible to detect 0.5 ng of arsenic(II1) in 10 ml of solution with a deposition time of only 10 minutes. With a gold working electrode, the detection limit for DPASV was the same as that for ASV, 0.02 ng/ml or 100 pg of arsenic(II1) in 5 ml of 1M perchloric acid. The deposition time was ten minutes. Data are presented in Table I1 relating the area of the arsenic oxidation peak to concentration of arsenic(II1). These data were obtained using DPASV. A comparison with the data in Table I shows that the deposition times required for DPASV are significantly shorter than those required for ASV. The ultimate detection limit for both methods was, however, the same since this was determined by the purity of the supporting electrolyte. The average relative standard deviation of the results obtained by DPASV was 10% or less, which was slightly less than that obtained by ASV. At higher concentration levels of arsenic(III), it was observed that the area of the arsenic oxidation peak no longer
Table I. ASV Sensitivity Studies peak area, mm2
CAS(III)I,~n g / m l
Peak area, m m 2
peak area, m m 2
CAs(III)], ng/ml
[ A s ( I I I ) I , ~n g / m l
5170 4200 4150 3789 3750 3562 3289 3409 3374
10 20 30 40 50 60 70 80 90 100
Peak area, m m 2
CAs(III)I, n g / m l
1162 116.2 2684 134.2 3342 111.4 3960 99.0 5560 111.2 7872 131.2 8330 119.0 8928 111.5 10026 111.1 12580 125.8 0 Relative standard deviation, 15.3%; deposition time, 20 min; deposition potential, -0.2 V: scan rate. 100 mV/sec; current sensitivity, 0.004 wA/mm; supporting electrolyte, 1M HC104. * Relative standard deviation, 9.1%; deposition time, 5 min; deposition potential, -0.2 V; scan rate, 100 mV/sec; current sensitivity, 0.04 fiA/mm; supporting electrolyte, 1M HC104. 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80 0.90
517 840 1245 1515 1875 2137 2302 2737 3037
Table 11. DPASV Sensitivity Studies peak area, mm2
peak area, m m 2
CAs(1IIU , ' n g / m l
Peak area, m m 2
CAs(III)lng/ml
[ A s ( ~ I I ) I ,n~g / m l
Peak area, m m 2
LAs(IIIp, n g / m l
0.1 442 4420 10 54 0 54.0 0.2 816 4080 20 1050 52.5 0.3 1156 3853 30 1725 57.5 0.4 1462 3655 40 2475 61.9 0.5 1768 3536 50 2929 58.5 0.6 2040 3400 60 3375 56.3 0.7 2482 3546 70 4050 57.8 0.8 2788 3485 80 4500 56.3 0.9 3230 3584 90 4800 53.3 1.0 3502 3502 100 5250 52.5 3493 1.1 3842 3485 1.2 4182 a Relative standard deviation, 8.4%; deposition time, 5 min; deposition potential, -0.2 V; scan rate, 10 mV/sec; pulse amplitude, 50 mV; measurement frequency. 0.5 sec; current sensitivity, 0.01 p A / m m ; supporting electrolyte, 1M HC104. Relative standard deviation, 5.4%; deposition time, 1 min; deposition potential. -0.2 V; scan rate. 10 mV/sec; pulse amplitude, 50 mV; measurement frequency, 0.5 sec; current sensitivity. 0.20 fiA/mm; supporting electrolyte. 1M HC104.
increased linearly with respect to the concentration of arsenic(II1). The concentration levels a t which this occurred varied with the surface area of the electrode, the length of the deposition period, and stirring efficiency but were generally greater than 1 kg/ml of arsenic(II1). This phenomenon was attributed to inhibition of the deposition of arsenic by the formation of a monolayer of adsorbed arsenic, a poor electrical conductor, at the electrode surface. This same phenomenon has been observed by other workers (24, 28). Interferences. A number of elements were examined to determine if they interfered with the determination of arsenic by ASV. The concentration of arsenic(II1) in these studies was 0.1 Mg/ml. As expected, 100-fold excesses of sodium, potassium, calcium, iron, cobalt, and nickel did not interfere. Lead and cadmium in 10-fold excesses also did not interfere. Silver, selenium, antimony, and bismuth present in amounts approximately equal to that of arsenic also did not interfere. Mercury, however, did interfere, its oxidation peak overlapping that of arsenic. Another problem stems from the fact that mercury and gold form an amalgam and the removal of mercury from a gold electrode can be difficult. Copper presented the most serious interference problem. In addition to having overlapping oxidation peaks, copper and arsenic have been shown to combine strongly (291, and the presence of even a slight excess of copper greatly reduces electrode response to arsenic. Trushina and Kaplan (18) reported that, when using platinum working electrodes to determine arsenic by ASV,
the addition of a gold salt to the cell greatly increased electrode response to arsenic. This effect was confirmed in this research. Figure 7 shows voltammograms of a 1M HC104 solution containing 20 pg/ml of arsenic(II1) ( A ) and that same solution after sufficient Au3+ had been added to make the concentration of gold 2 wg/ml ( B ) .A possible explanation of this enhancement effect is that arsenic would be plating out, a t least in part, onto a gold surface and the enhanced reversibility of the arsenic(II1)-arsenic(0) couple resulted in more arsenic being deposited onto the working electrode in the same amount of time. Another possible explanation of this enhancement effect would be that the gold being plated out on the platinum electrode greatly increased the surface area of the electrode and deposition of arsenic was much less hindered by the arsenic already deposited on the electrode surface. The possibility that the enhancement effect was due to arsenic impurities in the gold solution being added to the cell solution was investigated and this was found not to be a significant factor. Added gold salts were shown to have no effect on the response of gold working electrodes to arsenic.
LITERATURE CITED (1) "Standard Methods of Chemical Analysis", N. H. Furman, Ed., D. Van Nostrand. Princeton, N.J., 1966, p 124. (2) N. J. Goldstone, lnd. Eng. Chem., Anal. Ed., 18, 797 (1946). (3) D. F. Boltz and M. G. Meiion, Anal. Chem., 19, 873 (1947). (4) E. Gastinger, Mikrochh. Acta, 4, 526 (1972). (5)G. F. Kirkbright and L. Ransom, Anal. Chem.. 43, 1238 (1971). (6) W. Holak, Anal. Chem., 41, 1712 (1969). (7) F. J. Fernandez and D. C. Manning, At. Absorpt. Newsl., I O , 86 (1971). ANALYTICAL CHEMISTRY, VOL. 47, NO. 9, AUGUST 1975
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(8) (9) (10) (11) (12) (13) (14) (15) (16) (17) (18) (19) (20) (21)
E. F. Dalton and A. J. Malonski. At. Absorpt. News/., 10, 92 (1971). F. E. Lichte and R. K. Skogerboe, Anal. Chem., 44, 1480 (1972). R. E. Milkinson and W. S. Hardcastle: WeedSci., 17 (4), 536-7 (1969). A. Golanski, J. Radioanal. Chem., 3 (3-4), 161-73 (1969). S. Gohda, Bull. Chem. SOC.Jpn., 45 (6), 1704-8 (1972). A. Hiroshi. Hitosubashi, JArts Sci., 9, 35 (1968). L. Rozanski. Chem. Anal. (Warsaw), 16, 793 (1971). L. S. Naderhino, E. L. Grizaid. and E. G. Novakovaskaya. Tr. Leningr. Politekh. lnst., N304, 141 (1970). K. Hagiwara and T. Murase, BunsekiKagaku, 14, 757 (1960). D. J. Meyers and J. Osteryoung, Anal. Chem., 45, 267 (1973). L. F. Trushina and A. A. Kaplan, Zh. Anal. Khim., 25, 1616 (1970). H. Siergerman and G. O'Dom; Amer. Lab., 4 (6), 59 (1972). R. G. Megargle, Rev. Sci. hstrum., 43, 43 (1972). T. R. Copeland, J. H. Christie, R. A. Osteryoung, and R. K. Skogerboe. Anal. Chem., 45, 2171 (1973).
(22) D. G. Davis, Talanta, 3, 335-345 (1960). (23) L. Meites, "Polarographic Techniques", 2nd ed., Interscience. New York, 1965, pp 428-39. (24) L. Meites, J. Am. Chem. SOC.,76, 5927 (1954). (25) J. P. Arnold and R. M. Johnson, Talanta, 16, 1191 (1969). (26) R. K. Simon, G. D. Christian, and W. C. Purdy, Am. J. Clin. Pathol., 49, 207 (1968). (27) T. R. Copeland, J. H. Christie, R. K . Skogerboe, and R. A. Osteryoung, Anal. Chem.. 45, 995-96 (1973). (28) I. M. Kolthoff and J. J. Lingane, "Polarography", Interscience. New York, 1941, pp 261-262. (29) T. Kuwabra and S. Suzuki, Bull. Chem. SOC.Jpn, 44, 1690-4 (1973).
RECEIVEDfor review October 2, 1974. Accepted May 16, 1975.
New Easy Method for Obtaining Approximate Redox Potentials of Radicals, Produced by 6oCo-T-Radiolysis,Using Heteropoly Electrolytes of Molybdenum and Tungsten as Electron Acceptors. The Redox Potential of Some Alcohol and Organic 'Acid Radicals Elias Papaconstantinou Chemistry Division, Nuclear Research Center "Demokritos ", Aghia Paraskevi Attikis, Athens, Greece
The reducing ability of short lived radicals can be studied by reacting them with electron acceptors whose redox potential is known. Heteropoly electrolytes of molybdenum and tungsten are used for this purpose. The reduction of these species proceeds in distinct reduction steps producing the corresponding heteropoly blues until the potential of the next reduction step is more negative than the potential of the radical. The apparent redox potentials of some alcohols and formic acid radicals have been determined.
Reactions are generally divided into two main categories: substitution and redox reactions. Whereas bond making and bond breaking is the main concern in substitution reactions, the redox potentials of the reactants are, from a thermodynamic point of view, the important parameters in redox reactions. I t has been the purpose of this project to find an easy way to study the redox properties of organic radicals. I t is obvious that the classical methods ( I ) , that is electrochemical, cannot be used for species whose life time is of the order of microseconds, and redox potentials cannot be calculated from thermodynamic parameters simply because the data are quite limited. Organic radicals are produced extensively in radiation chemistry and photochemistry participating in all kinds of chemical and biochemical redox reactions such as enzymatic reactions, phosphorylation, etc. (2, 3 ) . Their life time, as mentioned earlier, is of the order of psec, thus making conventional experimentation out of question. With this in mind, we undertook the task of finding a rather easy way to obtain their redox potentials. For this reason, we have selected a series of heteropoly anions of molybdenum and tungsten as electron acceptors, namely the 18-molybdodiphosphate ["4]6[PzMo18062], the 18-tungstodiphosphate ["4]6[P2W18062], and the 12tungstophosphate [Nas][PW12040],henceforth designated, for simplicity, as Mala, Wls, and Wl2, respectively. The se1592
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lection of the system was made for the following reasons (4-8):
a) These compounds are reduced in distinct reduction steps by addition of 1 to 6 electrons without decomposition. b) Reduction can be easily followed spectrophotometrically. c) Reduction becomes more difficult as more electrons are added to the anions; Table I. d ) The reversibility of polarographic waves allows comparison with the corresponding reduction steps produced with chemical reducing reagents and polarographically. e) Although the mechanism of the electron transfer is not known even with the reducing reagents (Cr2+, Eu2+, V2+)for which the reduction stoichiometry has been established, one expects, because of the bulkiness of the structure of these anions and the apparent distribution of charge, a rather simple electron transfer. (To a fairly good approximation Wl2, and related complexes are spherical with diameter of about 12 8, (9) whereas the other two and related complexes are ellipsoidal with axes of 12 and 17 8, ( I O ) . ) That is, since the structure of the anions remains intact after the reduction (7, 11, 1 2 ) , no bond breaking or bond formation is likely to take place during reduction. What, then, is anticipated is electron acceptors whose abilit y to accept electrons depends on the reducing power of the reagent and not on the specific redox mechanism that takes place a t the time. While this work was in progress, E. Hayon et al. reported massive data on this subject ( 1 3 ) .Their method uses pulse radiolysis to produce the radicals, titrating them, subsequently, with various organic electron acceptors with different redox potentials. The 50% point in the titration curve is the redox potential of the radical (14,15). A referee has kindly pointed out that, Hayon et al.'s results were recently reinterpreted by them (as presented a t recent conferences and in manuscripts "in press") to show that equilibrium conditions did not prevail for most of the radicals
