7/27/2019 10.1365_s10337-006-0145-y

1/4

Determination of Low-Level

Ethylenediaminetetraacetic Acid in Water

Samples by Ion Chromatography with

Ultraviolet Detection

T. Kemmei1,&, S. Kodama1, A. Yamamoto2, Y. Inoue3, K. Hayakawa4

1 Toyama Institute of Health, 17-1 Nakataikoyama, Imizu, Toyama 939-0363, Japan;E-Mail: tomoko.kenmei@pref.toyama.lg.jp

2 Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, Matsumoto-cho 1200,Kasugai 487-8501, Japan

3 Department of Preventive Medicine and Environmental Health, Osaka City University Medical School,Asahi-machi, Abeno, Osaka 545-8585, Japan

4 Graduate School of Natural Science and Technology, Kanazawa University, Kakuma-machi,Kanazawa 920-1192, Japan

Received: 25 August 2006 / Revised: 17 November 2006/ Accepted: 21 November 2006Online publication: 14 December 2006

Abstract

A convenient and sensitive ion chromatographic (IC) method for the analysis of ethylenedi-aminetetraacetic acid (EDTA) in water samples was proposed. Using a fast reversible reaction

of free EDTA and metalEDTA complexes into Fe(III)EDTA complex in the presence of Fe(III)ions, sample solutions were applied to an ion-exchange column using a mobile phase (pH 2.3),which was composed of 100 lM Fe(III) chloride and 5 mM methanesulfonic acid. The additionof Fe(III) solution (100 lL) containing 10 mM Fe(III) chloride and 0.5 M methanesulfonic acid tothe sample solution (10 mL) permitted the injection of a large volume (400 lL) of sample, whichallowed for greater sensitivity. The proposed IC method gave a highly linear (r2 > 0.999)calibration curve ranging 0.0051.0 lM EDTA and had a limit of detection of 1.5 nM. Highrepeatability (RSD < 2.1%) and recoveries (88108%) were also obtained. With this method,total EDTA level in raw and drinking waters were analyzed successfully.

Keywords

Ion chromatographyDrinking waterEthylenediaminetetraacetic acid

Introduction

Ethylenediaminetetraacetic acid (EDTA),

the most widely employed aminopoly-

carboxylic acid, is a synthetic complex-

ing agent that is used in industrial and

commercial processes. The hexadentate

ligand of EDTA is a very powerful che-

lating agent that forms a stable complex

with polyvalent metal ions in a 1/1 pro-

portion [13].

EDTA is provided 4,5005,500 ton -

year)1 in Japan. Huge amounts of EDTA

from many sources are discharged to the

aquatic environment. EDTA is not

removed during wastewater treatment

[46] and is hard to biologically degrade

[5, 7, 8]. It has been reported that only

the Fe(III)EDTA complex was quickly

degraded by photolysis and other metal

EDTA complexes were very slowlytransformed [6]. While EDTA may not

pose a significant risk to human health or

ecological systems [9], it increases the

levels of certain heavy metals dissolved

even at very low concentrations [10]

by both releasing adsorbed metals from

sediments and preventing their removal

through precipitation. Thus, EDTA could

be considered as a critical environmental

chemical with regard to contamination of

surface water and groundwater [5]. Due

to the potential effects of EDTA on metal

fate during treatment and after discharge,sensitive analytical techniques are needed

for determining metalEDTA complexes

in environmental samples as well as in

drinking water.

A variety of chromatographic meth-

ods have been developed for the analysis

of EDTA. Gas chromatographic meth-

ods using a nitrogen phosphorus-specific

detector [6, 11, 12] or mass spectrometer

(MS) [13] have been used. These meth-

ods, however, require a preliminary

derivatization step, which is tedious and

time-consuming. The complexation con-

2007, 65, 229232

DOI: 10.1365/s10337-006-0145-y0009-5893/07/02 2006 Friedr. Vieweg & Sohn/GWV Fachverlage GmbH

Full Short Communication Chromatographia 2007, 65, February (No. 3/4) 229

7/27/2019 10.1365_s10337-006-0145-y

2/4

stant of EDTA with Fe(III) ion is par-

ticularly high (pKc 25.1) [3], and Fe(III)

EDTA complex has a UV absorption

peak. So, UV detection of Fe(III)

EDTA complex has been used in high

performance liquid chromatography

(HPLC) [1419], ion chromatography

(IC) [20] and capillary electrophoresis

[21]. However, these methods require a

concentration step for EDTA concen-

trations less than 10 lg L)1. Recently, a

low level of EDTA has been analyzed by

HPLCMS [22], ICMS [23, 24] and IC-

inductively coupled plasma MS [25].

However, these apparatus are not used

widely.

In this paper, we propose a conven-

tional IC method using UV detection for

the analysis of EDTA at 1.5 nM con-

centration (about 0.5 lg L)1).

Experimental

Chemicals

Disodium EDTA dihydrate (Na2H2EDTA2H2O), ferric monosodium EDTA

tri-hydrate (FeNaEDTA3H2O), calcium

disodium EDTA dihydrate (CaNa2EDTA

2H2O) and other metal EDTA hydrates

were obtained from Dojindo (Kumamoto,

Japan). Water was purified with a Milli-Q

SP.TOC. (Millipore, Tokyo, Japan).

Other chemicals (analytical grade) were

purchased from Wako (Osaka, Japan).

Apparatus for IC

The IC system consisted of a Toyo Soda

CCPD pump (Tokyo, Japan), a Rheo-

dyne 7125 injector with a 400 lL sample

loop, a Shimadzu SPD-10AV UV detec-

tor, a Shimadzu CTO-10AC column oven

(Kyoto, Japan), a Shodex DEGAS deg-

asser (Tokyo, Japan).

Standard and SamplePreparation

A stock standard of EDTA (1 mM) pre-

pared by dissolving disodium EDTA

dihydrate in purified water. The stock

solution was stored at 4 C and diluted

daily.

Environmental water samples (15 river

waters, 8 riverbed waters, 37 ground

waters, and 12 spring waters) were col-

lected in Toyama prefecture, placed in

polyethylene bottles and stored in the

dark at 4 C until analysis. Nine mineral

water samples were purchased from a

local market.

Fe(III) solution containing 10 mM

Fe(III) chloride and 0.5 M methanesulf-

onic acid was prepared by dissolving

135 mg of Fe(III) chloride hexahydrate

and 2.4 g of methanesulfonic acid in

50 mL purified water. Unless stated

otherwise, 100 lL of the Fe(III) solution

was added to 10 mL of the diluted stan-

dard solutions or samples before they

were applied to IC.

Chromatographic Conditions

Separations by IC were attained with a

4.6 mm i.d. 150 mm Hitachi gel IC

column (#2740; Hitachi, Tokyo, Japan)

thermostated at 40 C. This column was

made of polymethacryrate bonded with

alkyl quarternary ammonium and packed

with 10 lm particles. The mobile phase

(pH 2.3), unless stated otherwise, was

composed of 100 lM Fe(III)Cl3 and

5 mM methanesulfonic acid, and the flow

rate was 1 mL min)1. Analytes were

detected at 260 nm. Calculations of

EDTA concentrations were based on

peak heights.

Primary investigations with a 4.6 mm

i.d. 150 mm SUPELCOGEL TPR-100

column (Sigma-Aldrich Japan, Tokyo,

Japan) were done usuing 300 mM

methanesulfonate as a mobile phase

(pH 0.7). This column was made of

poly(divinylbenzene/methacryrate) and

packed with 5 lm particles and could be

used a below pH 2.

Results and Discussion

Conversion of EDTAand MetalEDTA Complexesinto Fe(III)EDTA ComplexThrough an IC Column

In the first experiment, it was tried to

analyze EDTA as free EDTA by reversed

phase HPLC with a mobile phase at very

low pH. That is, 1 mM of EDTA solution

was applied to a SUPELCOGEL TPR-

100 column with 300 mM methanesulfo-

nate (pH 0.7) as a mobile phase. Two

peaks were detected at 260 nm (Fig. 1a).

When 0.2 mM Fe(III)EDTA complex

was applied to the column, only one peak

was detected (Fig. 1b), whose retention

time was the same as that of the first peak

detected by applying EDTA. Therefore,

the first and second peaks in Fig. 1a

appeared to correspond to Fe(III)EDTA

complex and free EDTA, respectively.

Free EDTA or metalEDTA complexes

flowing through an HPLC column were

previously found to react with Fe(III) in

the column, forming Fe(III)EDTA

complex, and free EDTA and many

metalEDTA complexes could be easily

converted into Fe(III)EDTA complex by

addition of Fe(III) ions [15]. Moreover,

the peak height of Fe(III)EDTA at

260 nm was significantly higher than that

of free EDTA. Thus, on the basis of the

reaction of free EDTA and metalEDTA

complexes into Fe(III)EDTA complex in

the presence of Fe(III) ions, 0.1 mM

EDTA solution was analyzed as Fe(III)

EDTA complex using a Hitachi gel #2740

IC column and a mobile phase containing

5 mM methanesulfonate (pH 2.3). The

conversion rate of free EDTA into

Fe(III)EDTA complex increased with

increasing Fe(III) chloride concentra-

tion in the mobile phase, reaching 95% at

50100 lM Fe(III) chloride (Fig. 2).

Similarly, the conversion rates of

metalEDTA complexes into Fe(III)

EDTA complex were analyzed. Metal

EDTA complexes such as Ca(II), Cu(II),

Mg(II), Mn(II), Pb(II), Zn(II), Co(II),

and Ni(II)EDTA were commercially

Retention time (min)

4mAU

(a)

(b)

0 2 4

4mAU

Fig. 1. Chromatograms of (a) 1 mM freeEDTA and (b) 0.2 mM Fe(III)EDTA com-plex applied to a SUPELCOGEL TPR)100column with 300 mM methanesulfonate (pH0.7) as the mobile phase

230 Chromatographia 2007, 65, February (No. 3/4) Full Short Communication

7/27/2019 10.1365_s10337-006-0145-y

3/4

available as their hydrates and were

dissolved to 0.1 mM solution. Al(III)

and Cr(III)EDTA complexes could not

be purchased, so we prepared Al(III)

and Cr(III)EDTA solutions(0.1 mM) by

mixing Aluminium(III) nitrate solution

or Chromium chloride solution with

EDTA solution and boiled for 1 h. As

a result, Ca(II), Cu(II), Mg(II),

Mn(II), Pb(II), Zn(II) and Al(III)

complexes were converted into Fe(III)

EDTA complex at rates more than

90% (Fig. 3). However, Co(II)EDTA

complex showed only 71% conversion,

and Cr(III) and Ni(II)EDTA com-

plexes were not almost converted to

Fe(III)EDTA complex. Nowack et al.

[15] also reported that Cr(III) and

Ni(II)EDTA complexes were hard to

react with Fe(III) ions. EDTA exists

mainly in the form of Ca(II)EDTA or

Zn(II)EDTA complexes in natural

waters [11]. So, it seems that, in the case

of drinking waters, the low conversion

rates of Co(II), Cr(III) and Ni(II)

EDTA complexes into Fe(III)EDTA

complex have little effect on measure-

ments of the total EDTA concentrations.

Analysis of EDTA

In order to analyze with high sensitivity,

the effect of sample volume injected onthe analysis of EDTA was investigated.

When a stock standard solution of

EDTA was diluted in water, an increase

in injection volume resulted in broader

system peaks. In IC, system peaks are

generally derived from differences of the

composition between injection solution

and mobile phase. When water was

applied to the IC system, system peaks

were appeared before EDTA peak and a

larger volume injection of water broad-

ened the width of them. As a result, with

an injection volume of 100 lL, systempeaks overlapped with the peak of

EDTA. On the other hand, when a mix-

ture of 10 mL of EDTA solution and

100 lL of Fe(III) solution(in which the

concentrations of Fe(III) chloride and

methanesulfonic acid were the same as

those in the mobile phase) was applied to

the column, EDTA was successfully

analyzed with injection volume ranging

from 10 to 400 lL. Therefore, a 400 lL

injection volume composed of 100 lM

Fe(III)Cl3 and 5 mM methanesulfonic

acid was used for further experiments.

Linearity (r2 > 0.999) was demon-

strated in the range 0.0051.0 lM b y a

standard curve for EDTA. The detection

limit, defined as a signal-to-noise ratio of

3, was 1.5 nM (approximately 0.5 lg L)1).

The large volume injection allowed low

detection limit without a separate pre-

concentration step before injection. The

precision of five consecutive determina-

tions was evaluated at 0.1 lM of EDTA.

High repeatability of peak height

(RSD 2.1%) was obtained. To measure

0

20

40

60

80

100

Ca Cu Mg Mn Pb Zn Al Co Cr Ni

Conversionr

ates(%)

Metal species of metal-EDTA complexes

Fig. 3. Conversion rates of metalEDTA complexes into Fe(III)EDTA complex in a Hitachi gel#2740 column using 100 lM Fe(III)Cl3 and 5 mM methanesulfonate (pH 2.3) as the mobile phase

0

50

100

0 50 100

FeCl3(M)

Conversionrates(%)

Fig. 2. Effect of the concentration of Fe(III) chloride in the mobile phase on the conversion rate offree EDTA into Fe(III)EDTA complex. A Hitachi gel #2740 column with 5 mM methanesulf-onate (pH 2.3) as a mobile phase was used

0 2 4 6 8

Retention time (min)

0.1mAU

(b)

(a)

Fig. 4. Chromatograms of (a) steel-canned mineral water sample and (b) 0.1 lM EDTA standardsolution. Chromatograms were obtained with a Hitachi gel #2740 column with 100 lM Fe(III)Cl3and 5 mM methanesulfonate (pH 2.3) as the mobile phase

Full Short Communication Chromatographia 2007, 65, February (No. 3/4) 231

7/27/2019 10.1365_s10337-006-0145-y

4/4

recoveries, a standard solution of EDTA

was added to 15 water samples to a final

concentration of 0.1 lM. Good recoveries

(88108%) were obtained.

Using the proposed IC method,

EDTA in 75 water samples containing 15

river waters, 8 riverbed waters, 37 ground

waters, 12 spring waters, and 9 mineral

waters were analyzed. EDTA was

detected in only one mineral water sample

(Fig. 4a). The peak corresponded to a

concentration of 0.01 lM (3.0 lg L)1).

Conclusion

Since free EDTA and its metal com-

plexes, except Co(II), Cr(III) and

Ni(II)EDTA, were easily converted to

Fe(III)EDTA in the presence of Fe(III)

ions, EDTA was detected as Fe(III)

EDTA at 260 nm. Total EDTA in raw

and drinking water samples, which exists

mainly in the form of Ca(II)EDTA or

Zn(II)EDTA, was successfully analyzed

at low level (1.5 nM) by using the large

volume injection without a tedious pre-

concentration step.

Acknowledgements

This work was supported by a Grant-in-

Aid for Scientific Research (C)18580337

from the Japan Society for the Promotion

of Science.

References

1. Schwarzenbach G, Gut R, Anderegg G(1954) Helv Chim Acta 37:937957

2. Schwarzenbach G, Anderegg G (1957)Helv Chim Acta 40:17731792

3. Skochdopole R, Chaberek S (1959) J InorgNucl Chem 11:222233

4. Gardiner J (1976) Water Res 10:5075145. Alder AC, Siegrist H, Gujer W, Giger W

(1990) Water Res 24:7337426. Kari FG, Giger W (1996) Water Res

30:1221347. Cech JS, Chudoba J (1988) Acta Hydro-

chim Hydrobiol 16:3133238. Bolton H, Li SW, Workman DJ, Girvin

DC (1993) J Environ Qual 22:1251329. Wolf K, Gilbert PA (1992) In: Huntzinger

O (ed) The handbook of environmentalchemistry, vol 3. Springer, Berlin Heidel-berg New York

10. Erei Y, Morgan JJ (1992) GeochimCosmochim Acta 56:41574167

11. Xue H, Sigg L, Kari FG (1995) EnvironSci Technol 29:5968

12. Sillanpaa M, Sorvari J, Sihvonen ML(1996) Chromatographia 42:578582

13. Nishikawa Y, Okumura T (1995) J Chro-matogr A 690:109118

14. Bergers PJM, Groot AC (1994) Water Res28:639642

15. Nowack B, Kari FG, Hilger SU, Sigg L(1996) Anal Chem 68:561566

16. Geschke R, Zehringer M (1997) FreseniusJ Anal Chem 357:773776

17. Nirel PM, Pardo PE, Landry JC, Revaclier R(1998) Water Res 12:36153620

18. Bedsworth WW, Sedlak DL (2001)J Chromatogr A 905:157162

19. Sillanpaa M, Kokkonen R, Sihvonen ML(1995) Anal Chim Acta 303:187192

20. Voloschik IN, Litvina ML, Rudenko BA(1994) J Chromatogr A 671:249252

21. Pozdniakova S, Ragauskas R, Dikcius A,Padarauskas A (1999) Fresenius J AnalChem 363:124125

22. Dodi A, Monnier V (2004) J ChromatogrA 1032:8792

23. Bauer KH, Knepper TP, Maes A, SchatzV, Voihsel M (1999) J Chromatogr A837:117128

24. Collins RN, Onikso BC, Mclaughlin MJ,Merrington G (2001) Environ Sci Technol35:25892593

25. Ammann AA (2002) J Chromatogr A947:205216

232 Chromatographia 2007, 65, February (No. 3/4) Full Short Communication