8411
Anal. Chem. 1993, 65, 3411-3416
Determination of Ammonia in Seawater Using Catalytic Cathodic Stripping Voltammetry Anne-Marie Harbin and Constant M. G. van den Berg' Oceanography Laboratory, Liverpool University, Liverpool L69 3BX, United Kingdom
A procedure is presented for the determinationof ammonia in seawater using adsorptive cathodic stripping voltammetry (CSV). The ammonia is reacted with formaldehyde (13% w/v) at pH 3.8 to form an imino compound (methylenimine) which is adsorbedon the hangingmercury drop electrode (HMDE). Subsequently, a differential pulse scan is made toward more negative potentials and the reduction current is measured. Optimized conditions include an adsorption potential of -0.85 V and an adsorption time of 60 a; the voltammetric scan was initiated from-0.6 V. The peak potential was at -0.91 V. The CSV response was linearly related to the ammonia concentration between 10 and 3000 nM. The limit of detection for ammonia was 4 nM. Comparative measurements of the concentration of ammonia in samples from the estuary of the River Mersey and in lake waters using the CSV method and conventional spectrophotometric technique showed good agreement betweenthe two methods. The methodwas applied successfully to the determinationof low ammonia levels in seawater from the water column of the northwest Mediterranean. INTRODUCTION Ammonia occurs in seawater as an intermediate of the nitrogen cycle, being produced by bacterial reduction, assimilation, and cellular breakdown (ammonia here implies the sumof ammoniaand ammonium). Typical concentrations found in seawater vary from 0-25 pM in coastal waters to C0.5 p M in surface oceanic waters.' Existing methods for ammonia analysis include spectrophotometry, fluorometry, and potentiometry. Usually ammonia in seawater is determined by a spectrophotometric method based on the Berthelot reaction between ammonia, phenol, and hypochlorite leading to the formation of an indophenol dye.2.3 The reaction time is 1h, and the method is subjected to interferences from dissolved organic nitrogen compounds such as amino acids, urea, and nucleic acids.4 The sensitivity has been much improved by solvent extraction preconcentration, achieving a limit of detection of 3.5 nM.5 Ammonia in seawater can be detected potentiometrically by ion-selective electrode with a limit of detection of 0.1 pM.697
-
* Author to whom correspondence should be addressed.
(1) Sharp, J. H. In Nitrogen in the Marine Environment; Carpenter, E. J., Capone, E. D., Us.; Academic: London, 1983; p 2. (2) Strickland, J. D. H.;Parsons, T. R. A Practical Handbook of Seawater Analysis, 2nd ed.;Bull. Fish. Re8. Board Can. 1976, No. 167, pp 1-310. (3) Graeshoff,K.; Ehrhardt, M. E.; Kremling, K.Methods of Seawater Analy8k; Verlag Chemie: Weinheim, Germany, 1983; p 419. (4) Patton, C. J.; Crouch, S. R. Anal. Chem. 1977,49,464. ( 6 ) Bmzineki, M. A. Mar. Chem. 1987,20, 277. (6) Gilbert, T. R.; Clay, A. M. Anal. Chem. 1973,45,1757. 0003-2700/93/0365-3411$04.00/0
Flow injection methods include the detection of a change in pH upon diffusion of ammonia through a membranes or the use of fluorimetry (detection limit 20 nM) after derivatization with o-phthalaldehyde? which suffers from interference by volatile alkylamines. The latter method has been improved by preceding the reaction by diffusion of ammonia across a microporous Teflon membrane.10 This also forms the basis of a fiber-optic sensor with a reaction time of 5 min." Comparatively high levels (10 pM) of ammonia can be determined by polarography.12 Lower levels (50 nM) of ammonia can be detected by linear-sweeppolarography after reaction with 1-naphthol and hypochloritein strongly alkaline solution, pH >lo." This high pH could not be used for seawater as it leads to precipitation of calcium and magnesium hydroxides. Ammonia concentrations of 3-30 rM in natural water can be determined by differential-pulse polarography after reacting with formaldehyde at elevated temperature.l* Preliminary experiments to evaluate the use of indophenol blue dye and o-phthalaldehyde for the determination of ammonia using adsorptive cathodic stripping voltammetry (CSV) were unsuccessful as several mutually interfering reductive peaks were formed which were not qualitative for ammonia. In the present report, we describe a procedure to determine low levels (4 nM) of ammonia by adsorptive CSV after reaction with formaldehyde. The method is simple and requires no sample pretreatment involving extraction, and alkylamines do not interfere. It can form the basis for a sensitive FIA method with voltammetric detection.
EXPERIMENTAL SECTION Reagents and Equipment. Voltammetric analyses were carried out using either a PAR 174Apolarograph with a PAR 303 static mercury drop electrode (dropsurface area 2.94 mm2) or an Autolab polarograph connected to a Metrohm 663 hanging mercury drop electrode (HMDE surface area 0.38 mm2) controlled by an AT 80286IBM-compatiblepersonal computer. The referenceelectrode was an Ag/AgC13M KC1whereas the counter electrode was a platinum wire. The PAR 174A polarograph had been altered (by shortening the time interval between pulse) to increase the pulse rate to 10 s-l, and scans were recorded on a Kipp en Zonen X-Yrecorder. Solutions were stirred using a PTFE-coated, star-shapedmagnetic stirring bar, propelled by a magnetic stirrer. The voltammetric cell was glass and had a samplevolume of 10mL. Cyclic voltammetry,scan rate, reaction time, adsorption time, and adsorption potential experimentswere carried out using the Autolab polarograph connected to a Metrohm 663 hanging mercury drop electrode. Solutionswere purged with water-saturatednitrogen prior to voltammetric analyses. The pH meter (Metrohm Model 625) (7) Garside, C.; Hull,I. G.; Murray, S. Limnol. Oceanogr. 1978,23, 1073. (8) Willason, S. W.; Johnson, K.S. Mar. Biol. 1986,91, 286. (9) Genfa, Z.; Dasgupta, P. K. Anal. Chem. 1989,61,404. (10) Jones, R. D. Limnol. Oceanorg. 1991,30,814. (11) Karr,S.; Arnold, M. A. Anal. Chem. 1992,64, 2438. (12) Nyman, G. J.; Ragle, J. L.; Lunde, P. F. Anal. Chem. 1960,32,362. (13) Zhao, Z.; Mo, J. Electroanalysis 1990,2, 139. (14) McLean, J. D.; Stenger, V. W.; Reim, R. E.; Long, M. W.; Hiller, T. A. Anal. Chem. 1978,50, 1309. 0 1993 American Chemical Society
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* ANALYTICAL CHEMISTRY, VOL.
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was calibrated usingNBS buffers. A Philips VM 705 microwave was ueed for heating samples when required. Special w e was taken to exclude paaaible solution contaminationwith ammonia from the laboratoryatmosphere. Solutions containing high concentrations of ammonia were stored in a fumehood, and solutions containing low levels of ammonia and samples were prepared on a laminar-flow bench supplied with fiitered air. Disposable plastic gloves were worn during sample handling. Even so,it was found impossible to store samples for an extendedperiodof time in the laboratoryenvironment without considerable contamination. Analyses were therefore carried out as Boon as possible upon sampling, whereas samples from the Mediterranean were stored frozen and wrapped in two plastic baga to minimize interaction with the atmosphere. Reagents were BD$I AnalaR grade unleea otherwise stated. Water (MQ produced by an Milli-Q deionizer system (Millipore) was ueed to prepare reagents. Formaldehyde (3795,w/v; Aldrich) was purified by paeeing through a column (250 X 10 mm) of cation-exchaugereah (DowexX8-SO, 100mesh, in the H+ and was stored in a glaes bottle. A stock solution containing 1 mMammonia waa prepared from ammonium chloridewhich had been dried at 110 "C for 1 h. An aqueous stock solution of the pH buffer contained 1.3 M d u m acetate trihydrate in 10 M acetic wid (BDHGPR). Aqueous stoek solutions of surfactanta, dodeeylbenzenesulfonicacid, cetylpyridiniumbromide (Sigma), and Triton X-100 were prepared by dissolution of the solids in water. Seawater ueed to optimize the method was collected from the Menai Straits and had a ealinity of 33 psu (practical salinity units). Samples from the Meraey Estuary were collected on March 20,1992,using a v-I (jet foil) from the Natural River Authority (NorthWest),by manually filling glass d i t y bottles usingbucket (polypropylene)collected water. The entire salinity region was coveredwithin 4 h. Samples were stored in the dark andanalyleduponretunatothelaboratory(within4h). Samples from the MediterraneanSea were collected during a cruise of the Valdivia as part of theEROS 2000program. The station location was 38" N, 0.6" E (station 81,February 1991. Collection took place at depths ranging from 21 to 2701 m using Teflon-coated Go-Flo bottles (General Oceanics). Samples were fiitered (0.4 pm, Nuclepore) and frozenimmediately after collection. Analysis took place on return to the laboratory. Procedure for Determining Ammonia in Seawater. A 5.5-mL sample aliquot was pipeted into the voltammetric cell; 1.0mL of buffer solution and 3.5mL of formaldehyde were added to give fiial concenetrationsof 1.1 M acetate pH buffer and 13% (w/v)formaldehyde. The pH was -3.8. Thereaction was allowed to proceed for 20 min. The cell was deaerated for 4 min, whereafter the cell potential was set to -0.85 V, a new mercury drop was extruded, and the timer was started. The adsorption time was 60 s (with stirring) followed by a 10-s quiescence time. The scan was initiated at -0.6 V and the conditions were as follows: negative scan diredion, differential-pulse mode, pulse height 50 mV, pulse rate 10 e-*, and scan rate 10mV/s. The peak potential for the ammonia-formaldehyde compound was at -0.91 V. The procedure was repeated with a separate sample aliquot to which an addition of ammonia standard was added sufficient to at leaat dauble the original peak current.
RESULTS AND DISCUSSION Cyclic Voltemmetry. A reduction peak is obtained a t 4.91 V when a potential scanis made from 4.6 V in a negative direction in a solution containing the ammonia-reagent mixture according to the procedure outlined above (Figure 1). The fmt cyclic scan was preceded by 609 adsorption and immediately followed by a second scan (Figure 1,dotted l i e ) . The reverse scan exhibits a stepwise decrease in the reduction current instead of an oxidation peak, indicative of a catalytic current similar to that produced upon the electrochemical reduction of the formazone complex of platinum.16J6 The second cyclic scan produced a smaller reductive peak than the first because this scan had not been preceded by
-0.6
-0.7
-0.8
-0.9
-1.0
-1.2
-1 1
potential (V)
Flgwr 1. Cyclic voltammetry of methyknimhre formed by readon of 500 nM of ammonla wlth farmaldehyde (l3%, w/v) in seawater at pH 3.8(0.13Msodlumacetateand 1.0Maoeakacld). Thereactkntlme was 20 mh. The scan rate wao 10 mVls and adsorptbn (e0e) at -0.8 V. A subsequent8econdscanonthe8amemerwydrop(notpreceded by adsorptbn) is portrayed by a dotted line.
adsorption. The formaldehyde and the ammonia are known to form an imino-type compound in acidic solution." The production of this compound (methylenimine) is as follow: H,C4
+ NH;
-
[HsN+CH20-- H,N==CH,OI
-
Ha+"
(1)
This reaction is similar to that preceding the determination of platinum, where formaldehyde reads with hydrazine to form the imino compound formazone (CHpNHNH2)+.'6 Compounds of the imiio type (>C-N-) are readily reduced at a high overpotential electrode such as mercury18 in acidic media. The carbon-nitrogen bond of the imine is reduced via a two-electron reduction to an amine.18 In acidic media imines usually exhibit two one-electron-reduction waves;18 however, a single two-electron wave is observed if the two reduction potentials are comparable,m aa may be the case here. The occurrence of a catalytic current indicates that the reduction triggers a catalytic generation of hydrogen on the shoulder of the hydrogen wave, analogous to that occurring at the reduction of the formazone complex of platinum.16J6 The more positive potential of the catalytic hydrogen wave than that for the normal hydrogen wave is not in disagreement with this scenario as the hydrogen wave is shifted to this negative potential by irreversibilityat the mercury electrode.
On the baais of the preliminary experiments into the reaction mechanism and the above discussion, the following reduction mechanism is tentatively proposed for the methyleaimine: In acidic medium protonation occurs H,C=NH
+ H+
-
H,C=NH2+
(2) The protonated methylenimine adsorbs on the HMDE during the preconcentration step. One of the protons on the protonated imine group is reduced to hydrogen a t -4.9 V 2H2C=NH2+ +2e-
-
2H2C=NH
+ H,
(3) The product is recycled by the protonation reaction 2. The peak shape of the reductive voltammetric scan (scan 1 in Figure 1)indicates that the catalytic current is terminated (16)van den Berg, C. M. G.;Jacinto, G.S. A d . Chim.Acta 1988,211, 129. (16)zhao, Z.;hsiaer, H. AM^. Chem. l9M,68,1498. (17)Morrison, R. T.;Boyd, R. N. Organic Chemistry, 3rd ed.;Allyn, Bacon: New York, 1974. (18)Rifi, M.R.Introduction to Organic Electrochemistry; Dekker: New York, 1974. (19)Lund, H.Talonta 1964,12,1085. (20) Zuman, P.InprosreaSinPhyuical Organic Chemktru;Wtweiwr, T., Taft, T.,I%.; Wdey Interscience: New York, 1967;V d 6, p 120.
ANALYTICAL CHEMISTRY, VOL. 65, NO. 23, DECEMBER 1, 1993
a
A
3413
A
pdanlial
Y
-0.92 Y
I
Y . W V
02004006008001MH)
scan rata (mV/s)
I
,
0.0 -0.45 -0.70 -0.95 -1.20 -1.45 -1.70 Potential (V)
6
n n -0.5
-1.0
adsorption potential
-1.5
'0
N)
I
t
0.0 -0.45 -0.70 -0.95 -1.20 -1.45 -1.70 Potential (V)
Flgurr 2. ac vottammetry of the HMDE In seawater wlth 0.1 M acetate buffer and formaldehyde. Scans after 60-s adsorption: (A) seawater wlthaddltlonsof formaldehyde and buffer: (B) seawater reagent mixture plus ammonia. (A) a, seawater; b, seawater 0.1 M acetate buffer: c, (b) 2% (wlv) formaldehyde; d, (b) 7 % (w/v) formaldehyde; e, (b) 4- 13% (w/v) fmldehyde. (B) a, no ammonia; b, 1 pM ammonia; c, 5 pM ammonia; d, 5 pM ammonia, wlthout adsorption.
+
+
+
by reduction of the protonated methylenimine to methylamine H2C=NH2++ 2 6 + H+
-
H,CNH,
(4)
ac Voltammetry. ac voltammetry was used to verify the adsorptive electrode process during the accumulation step. The ac frequency was 75 Hz, the modulation amplitude 10 mV, and the phase angle 90° to sample the capacitance specifically. Preliminary scans (initiated at -0.2 V preceded by 60-8 adsorption at -0.8 V) for seawater with additions of acetate buffer and increasing amounts of formaldehyde (without added ammonia) showed (Figure 2A) that the capacitance current at potentials negative of -0.2 V was suppressed by the addition of acetate buffer, presumably as a result of displacement of the major ions of seawater from the electrode surface by the acetate. Additions of formaldehyde caused increased suppression of the capacitance current at potentials between -0.2 and -1.0 V and produced two cathodic peaks at -1.15 and -1.5 V in the region of the hydrogen wave. The hydrogen wave itself is not apparent from the ac scans due to the electrochemical irreversibility of the hydrogen production. The shapes of the peaka suggest that their origin is not due to capacitanceeffects as this would produce sharp peaks, it is more likely that they reflect the faradaic current resulting from a two-step reduction of the formaldehyde. Additions of ammonia to seawater containing the formaldehyde-acetate buffer reagent mixture showed (Figure 2B) that the capacitance current between-0.2 and -1.0 Vis further suppressed, indicating that methylenimine adsorbs on the HMDE even when present at comparatively low concentrations in the presence of a large excess of formaldehyde. The capacitance current was not suppressed when a scan was carried out in the reagent mixture without stirred adsorption immediately after extrusion of a new mercury drop (Figure 2B,scan d) confirming the adsorptive process. Effect of Varying the Scan'Rate, Adsorption Potential, and Adsorption Time. Variation of the scan rate between 2 and 200 mV/s showed that the cathodic current increased
ob
50
100
150
adsorption time (s)
Flgurr 3. Effect of varying the scan rate (A), the adsorption potentlai
(a), and the adsorption time (C) on the CSV response for ammonia In
seawater: (A) 500 nM ammonia, 60-8 adsorption at -0.8 V, ilnear sweep scans: (B) 25 and 250 nM ammonia, 60-sadsorptkm, differentlalpulse mode; (C) 25 and 250 nM ammonia, adsorption at -0.8 V, differential-pulse mode.
nonlinearly with the scan rate (Figure 3A). A linear increase would be expected for a reversible reduction of an adsorbed compound. The nonlinear increase is in accordance with the proposed mechanism where the current depends on a catalytic effect in addition to the reduction of the adsorbed compound. The peak potential was found to shift to more negative potentials with increasing scan rate (Figure 3A), suggesting slow kinetics of the reduction step. Comparativeexperimenta using linear-sweep voltammetry and voltammetry using a differential-pulse modulation of 10 pulses/s showed that the sensitivity was much improved by the modulatedwave form,presumably because of the repeated measurement of the reduction step within a slowly scanned potential interval. Comparative experiments using squarewave voltammetry (step height 2.5 mV, pulse height 25 mV, using the Autolab polarograph) gave poorer sensitivity at frequencies above 10 Hz, presumably due to the irreversible nature of the catalytic reduction wave coupled with the shortened period of current sampling sooner after the beginning of each pulse at high frequencies. The differentialpulse wave form was therefore used in subsequentexperiments and for analytical purposes. Variation of the adsorption potential showed that the reduction current increased with the adsorption potential between -0.5 and -0.8 V, giving greatest adsorption at potentials between -0.75 and -0.85 V (Figure 2B); the reduction current diminished strongly at adsorptionpotentials more negative than -0.9 V due to reduction of the deposited material. The increased adsorption at potentials more negative than -0.5 V suggests that the adsorbing compound is positively charged as the charge on a mercury electrode is negative at potentials more negative than -0.5 V (zero point of charge in chloride solutions),2l in agreement with the proposed adsorption of protonated methylenimine, which has a positive charge (eq 2).
ANALYTICAL CHEMISTRY, VOL. 65, NO. 23, DECEMBER 1, 1993
3414
5
20
-
- - A
15
-current---
B -
1
i
., oo-s----10
-/r---
15
1
I
-
,,
-.1 _/---
-3
+3.5 30
20 acetate (M)
formaldehyde (% wiv)
D
E
$20,
l
2vMNH3
,
6 5 f
.c
m
'
1
10 ' ,,-
,
g
L
0
4 g - 3 p
I
200nMNH3
-
8
:I
10 20 30 40 50 60 reaction time (min)
PH
Figure 4. Effects of varying the solution composition on the DPCSV peak height for ammonia in seawater using an adsorption time of 30 s: (A) variation of the formaldehyde concentration; 200 nM ammonia in seawater of pH 3.8 and containing 1.0 M acetic acid and 0.13 M sodium acetate. (6) Variation of the acetate concentration at pH 3.8; 400 nMammonia in seawater. (C)Variatlonof the pH; 200 nM ammonia in seawater. (D) Variation of the reaction time on the CSV peak height for low (200 nM)andhigh (2pM)concentrationsof ammonia in seawater.
Variation of the deposition time at two levels of ammonia showed that the peak current increased -4-fold at the lower level of ammonia (25 nM) when the deposition time was increased from 0 to 60 s, whereas the increase was much less (-30 7% ) at the higher level (50 nM) of ammonia (see Figure 3C). The peak height diminished gradually at longer adsorption times, indicative of either drop saturation or competitive adsorption by other solution components such as unreacted formaldehyde. A similar apparent saturation was obtained at the lower concentration of 25 nM, indicating that this effect is due to competitive adsorption rather than to surface saturation by adsorption of the methylenimine. Optimizationof the Concentrations of Formaldehyde, and Buffer, the Solution pH, and the Reaction Time. The methylenimine was found to be stable in weakly acidic solution for several hours and could be formed conveniently by addition of the formaldehyde to seawater sample in the voltammetric cell. The effect of varying the reagent concentration was studied at a buffer concentration of 0.13 M sodium acetate and 1.0 M acetic acid (Figure 4A). The peak height rose sharply with the formaldehyde concentration up to 13% (w/v) formaldehyde, showing a working range between 13% and 17% ,whereafter it decreased markedly, presumably due to competitive adsorption on the mercury drop. The buffer concentration was optimized by varying the acetate concentration between 0.1 and 1.5 M (concentrations less than 0.1 M gave no peaks probably because the pH was much lowered by the added formaldehyde, which was acidic as a result of the cleaning process, which involved treatment with acidic ion-exchange resin). The peak height was found to increase with the acetate concentration up to an optimum concentration of 1 M acetate whereafter the increase was negligible. The same relationship between the acetate concentration and the CSV sensitivity was seen when the experiment was carried out in MQ water. An attempt was made to increase the catalytic effect by using stronger acids (sulfuric acid and hydrochloric acid) as background electrolyte in the seawater. The DPCSV peak height increased with the sulfuric acid concentration up to
-
(21) Heyrovsky, J.; Kuta, J. Principles of Polarography; Academic Press: New York, 1966; p 21.
0.2 M, whereafter it decreased. The effect of increasing the hydrochloric acid concentration of the peak height was minor, the sensitivity increasing by -20% from 0.01 to 1 M hydrochloric acid. The DPCSV sensitivity for ammonia using acetate buffer was greater than using either of these acids, so the acetate was selected for analytical purposes. Variation of the pH between 2.0 and 5.5 showed that the DPCSV peak height increased with the pH until pH -4, whereafter it decreased strongly (Figure 4C). The decrease at higher pH values may be due to a change in the formation efficiency of the methylenimine, or to deprotonation, of the methylenimine (eq 2): A lowering of the catalytic effect as a result of deprotonation of the methylenimine would be in accordance with the proposed reaction mechanism. Several voltammetric peaks were observed in more alkaline solutions (not apparently related to the ammonia concentration), suggesting that instability of the methylenimine was responsible for this effect at high pH; further evidence for this explanation is that the peak height could not restored by lowering of the pH again to near ita optimal value, showing that the effect was irreversible. It is not clear why the sensitivity diminishes rapidly at pH values below 3.5, but it is likely that variations in adsorption efficiencyof protonated and unprotonated substances may play a role. The peak potential was found to shift in a negative direction as the pH was increased as a result of increased stability of the methylenimine. The shift is especiallystrong at pH values near 3.4, suggesting that the acidity constant (pK) for protonation of methylenimine has a value near 3.4 in seawater. Variations in the protonation of the acetate were not responsible for this effect as the value for the acidity constant is 4.5 in seawater (calculated using constants22 valid for an ionic strength of 0.1 and corrected for major ion competition). The time required for the formation of the methylenimine from formaldehyde and ammonia was investigated at ammonia concentrations of 200 nM and 2 pM. At the lower concentration of ammonia the peak height appeared to increase with time up to 15 min, whereafter it leveled off. However, a longer reaction time of -30 min was required for completion at the higher level of ammonia (Figure 4D). We attempted to accelerate the slow reaction kinetics and decrease the measuring time by heating the seawater-reagent mixture. Several types of bacteria, algae, and fungi are known to not be affected significantly by short periods (up to 3 min) of heating of seawater using a microwave oven.23 Hence, the use of microwaves for a short period should have a negligible effect on the phytoplanktonic content of seawater samples and ammonia from cellular breakdown should not be released. For this reason, use of a microwave oven to heat the sample solution was investigated. Heating of 10mL of the seawaterreagent mixture in the voltammetric cell (covered by a watch glass) by microwave oven for 5 s at 125 W caused a 10 "C temperature rise and caused the reaction to approach completion, as indicated by CSV measurementa after different reaction times subsequent to the microwave heating (samples were found to cool to room temperature during the deaeration step prior to the CSV scan). However, the sensitivity was decreased (by -25% ) when the microwave was used, possibly due to partial breakdown of the organic reagents. Ammonia concentrations could still be accurately determined as the same effect was present after the standard addition, so the microwave oven can be used to accelerate the analysis time at high ammonia concentrations. Interferences. The determination of the ammoniaformaldehyde complex by adsorptive CSV suffers potentially
-
-
(22) Smith, R. M.; Martell, A. E. Critical Stability Constants; 2nd Supplement; Plenum Press: New York, 1989; Vol. 6, p 300. (23) Keller, M. D.;Bellows, W. K.; Guillard,R. R. L.J. Exp. Mar. B i d . Ecol. 1988, 117,279.
ANALYTICAL CHEMISTRY, VOL. 65, NO. 23, DECEMBER 1, 1993 IL
I *
DBS
O ' -0.7 0
3411
0.5
1.0
1.5
-0.8
-0.9
-1
'
potential (V)
2.0
surfactant concentration (mgll)
Flgm 1. Effect of surfactants on the CSV determination of 250 nM ammonia in seawater. The adsorption time was 60 8.
from interferences as a result of competitive adsorption of electroactive and surface-active compounds. Various aliphatic and aromatic amines could also react with formaldehyde to form surface-active compounds. The possibility of such interferenceswas investigated by determining ammonia by CSV using the optimized conditions in the presence of amino acids and amines at concentrations much higher than those occurring in seawater. The CSV peak for 200 nM ammonia was not affected by the addition of 500 nM of the followingamino acids: cysteine,leucine, asparagine,histidine, and glycine. The successive addition of 2 pM hydroxylamine and methylamine yielded peaks at the same potential as that of ammonia. However, the apparent CSV sensitivities for these compounds were very low, 0.002 nA/nM per 60 s for hydroxylamine and 0.0005nA/nM per 60 s for methylamine, compared to 0.5 nA/nM per 60 s for ammonia, suggesting that the response was due to traces of ammonia in the hydroxylamine and methylamine. Possible interferenceby metals was investigatedby addition of metals known to give peaks in voltammetry to concentrations far above those normally occurring in seawater. Addition of 1pM zinc, lead, cadmium,nickel, iron(III),copper, and cobalt caused no effect on the CSV response of ammonia. Comparison showed that the CSV sensitivity for ammonia in seawater slightly (-25%) lower than in distilled water, illustrating that interference by the major ions in seawater is minor. However, the difference necessitates recalibration of the CSV sensitivity for each sample individually if they originate from varying salinities. The possible interference by surfactants in seawater was investigated by addition of anionic (dodecyl benzenesulfonic acid, DBS), cationic (cetylpyridinium bromide, CPB), and nonionic (Triton X-100)Surfactants. The Triton X-100and CPB were found to decreasethe CSV peak height for ammonia (Figure 5) whereas an increase in peak height was caused by the addition of DBS. More work would be requiredto explain the improvement from DBS, but a possible explanation is related to an apparent decrease in the slope of the hydrogen wave; possibly adsorption of substances which increase the hydrogen wave was hindered whereas the adsorption of methylenimine was unhindered. The reduction peak of the ammonia compound is a shoulder on the reduction wave of hydrogen, and at low concentrations (5-50 nM NHs), it can be difficult to evaluate the peak height especially in the presence of organics. A low concentration of DBS (0.35mg/ L)can be used to improve the shape of the background of the CSV scan and thus the sensitivity for low ammonia concentrations. Higher concentrations of DBS caused a significant "foaming" effect during stirred adsorption, which was found to be unsuitable for routine analysis. Limit of Detection and Dynamic Range. The limit of detection was calculated from repeated determinations of ammonia in a seawater sample (surface seawater from the Mediterranean) containing a low level of ammonia; a concentration of 25 i 1.2 nM ammonia (n = 9) was found. The
O
1 / 0
1
2
3
4
ammonia (pM)
Figwe6. (A) Voltammetric scans for ammonia inseawater by DPCSV. The scans were determined under optimized condlMon.8 and show seawater wlth successive standard addltkns of 25,60, and 100 nM ammonia. (B)Peak hdght for ammonia in seawater as a functkm of the ammonia concentration. Adsorptkn time 60 s.
3u limit of detection using these data was 4 nM with an adsorption time of 60 8. CSV scans illustrating the response for a low level of ammonia in seawater with subsequent additions of 25,50,and 100nM ammonia are shown in Figure 6A. The ammonia content of the formaldehyde after several purification steps was typically 5-10 nM, contributing -2-3 nM to each sample; the contribution of thii source hae to be taken into account at low ammonia concentrations (by subtraction of the reagent contribution to the detected ammonia concentration) unless a more effective method of reagent purification is developed. Contribution of ammonia contamination from the buffer solution was found to be negligible. Thelinearityof response of the CSV method was evaluated from scans at enhanced concentrations of ammonia in the seawater. The peak current was found to increase linearly with the ammonia concentration between 10 nM and 3 pM at a 60-8 adsorption time; the increase leveled off a t higher ammonia concentrations, probably as a result of saturation of the electrode surface (Figure 6B). . Ammonia in Natural Water Samples. The method was tested on samples from freshwater, estuarine and oceanic origin. Samples from the estuary of the River Mersey (England) were collected at high tide on March 20, 1992. Comparativedeterminationsof the concentration of ammonia were carried out by the conventional spectrophotometric method and by CSV. It can be seen that the ammonia concentrations decreased from a very high level of 150 pM at the freshwater end (at salinities 1-4)to lower levels of 10 pM at the seawater end (salinities 30-35) probably due to ammonia input from sewage in the freshwater end of the estuary (Figure 7A). The samples at the freshwater end had been diluted to remain within the linear range for CSV. The CSV sensitivity was calibrated by ammonia addition to each sample, whereas the spectrophotometric measurements had been calibrated using a single calibration graph. Comparison of the results of the CSV and spectrophotometric methods (Figure 7A,B) shows good agreement for ammonia concentrations below -70 pM whereas the voltammetric method gave slightly lower results than the spectrophotometric method at higher concentrations (low salinities). The plot of voltammetric versus spectrophotometric results (Figure 7B)
- -
ANALYTICAL CHEMISTRY, VOL. 65, NO. 23, DECEMBER 1, 1993
8410
a slope indistinguishable of unity (0.99 i 0.006)and a Y-axis intercept indistinguishable from 0 (0.09 f 0.006).
200 A
i / '0
salinity ammonia (pM) 0 2 4 6 8 1012
50 100 156200 ammonia (uM) CSV ammonia (pM)
Medfierranean
10
Results of ammonia determinations by CSV in samples from the northwestMediterraneanW(38ON,0 . 6 O E)are shown in Figure 7D. The ammonia concentration can be seen to increase from very low levels (66 nM) in the surface waters to a subsurface maximum of 800 nM at -4OO-m depth, presumably caused by breakdown of algal material. Below this the ammonia concentration drops to lower levels in the deeper waters whereas it increases again in the bottom waters, perhaps due to releasee from the sedimenta. This profile is consistent with the known behavior of ammonia in oceanic conditions and illustrlltes the applicability of this method to samples containing low concentrations of thb important nutrient.
CONCLUSIONS
! \ I !
The optimized conditionsfor the determinationof ammonia in seawaterusing the reduction of theammonkformaldehyde - 51 ; 3 complex include a depaaition potential of -0.85 V, a formFIgIm 7. Apphtlom of the csv meulod to Mmanh In natuel aldehyde concentration of 13% (w/v), and a pH of 3.8. A watem: (A)amnmtn-byCSVand-ln limit of detection of 4 nM usinga 60-8deposition time, coupled ~ ~ ~ o c o m p r r b o n - - with the rehtive simplicity of the analytical procedure, detectd by and VoltMWneblc meihocb. The facilitates the determination of low levels of ammonia in ntraiohth hdcatesa 1:l rdutbrwtrlp: (C)UrmaJpcmmntratbns unpolluted natural waters and illustrates the advantages of by CSV and in Lake Esthwalte water: @) vertlcal prollk of ammonla In the nazhwbst hkdhrranean (38°30.0' N, voltammetry over the more traditional spectrophotometric 0.6°00.0' E). methods. The methad hae beentested by comparativenwasurementa hadaslopecloeetounity(1.06~O0.012),andaX-~intercept of ammonia in estuarine water and freshwater samples by claw to zero (-0.8 f 1.3 pM). Deviation of the spedaophospectrophotometryand~l~,theresulteshowinggood tometic Bensitivlty from the calibration graph graph at low agreement. The CSV method hae furthermore been sucslllinitiesmay be the cause for the deviation in thispart (high cessfully applied to the determination of ammonia in s a m p b ammonia) of the concentration range. from the Mediterranean a t concentrations between 66 and The concentratione of ammonia in samples from Lake 800 nM. Esthwaite water (England) obtained by the CSV method and ACKNOWLEDGMENT by epectrophotometry can be compared in Figare 7C. The ammonia concentrations were between 0.06 and 12 PM, A.M.H. wan financially supported by an award from the showing enrichment in the bottom waters due to deasea NERC. histance with sample collection from the Memy from the sedimenta, and very good agreement between the estuary by Peter Janes (NRA North West) ia gratefully two methods ia apparent. Linear regression of the spectroacknowledged. photometric data as a function of the voltammetric data gives
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(2a) Achtmberg, E.P.;Zhang, H.;Gledhill, M.;van den Berg, C. M. G.In Water Pollution Reaearch Report of the Environmental Research Program ofthe CEC Martin, J.-M., Bart$, H.,Eds.; CEC Bruae&, in
proceee.
RECEIVEDfor review May 10, 1993. Accepted September 1, 1993.0 Abstract publiehed in Advance ACS Aktracta, October 16,1993.
