ORIGINAL PAPER

A miniaturised isotachophoresis methodfor magnesium determination

Jeff E. Prest & Sara J. Baldock & Peter R. Fielden &

Nicholas J. Goddard & Bernard J. Treves Brown

Received: 31 October 2008 /Revised: 22 December 2008 /Accepted: 7 January 2009 /Published online: 10 February 2009# Springer-Verlag 2009

Abstract The use of malonic acid as a complexing agenthas enabled a new method to be devised to allow thedetermination of magnesium to be made using miniaturisedisotachophoresis. Using a leading electrolyte of 10 mmolL−1 caesium hydroxide and 2 mmol L−1 malonic acid atpH 5.1 gave the method a high specificity towardsmagnesium. Investigations using a poly(methyl methacry-late) chip device with an integrated conductivity detectorshowed that no interference from calcium, strontium,barium and sodium should occur. The method was foundto be linear over the range of magnesium concentrationsfrom 0.625 to 75 mg L−1 and the limit of detection wascalculated to be 0.45 mg L−1. Separations were demon-strated with water samples but the procedure should also beapplicable to more complex sample matrices such asinorganic explosive residues, blood or urine.

Keywords Isotachophoresis .Miniaturisation .

Microdevice .Magnesium .Metal analysis

Introduction

There is a requirement to carry out the analysis ofmagnesium in a large number of applications. Suchapplications include the analysis of inorganic explosiveresidues, water analysis and biomedical applications. Thereare a variety of analytical methods that can be used for thedetermination of magnesium. Traditionally, ion-selective

electrodes [1] and atomic absorption spectroscopy [2] haveseen widespread use. More recently, methods have beendeveloped for other techniques such as ion chromatography(IC) [3]. The use of IC for the analysis of small inorganiccations has proven to be successful and it is the maintechnique currently used for the analysis of inorganicexplosive residues [4]. The use of capillary zone electro-phoresis (CZE) has also been advocated as a lower-costalternative to IC for performing magnesium analysis. Thus,CZE has been applied to the analysis of magnesium in arange of samples such as blood serum and urine [5],seawater [6], gunshot residues [7] and inorganic explosiveresidues [8].

Isotachophoresis (ITP) is one of the families of electro-phoretic separation techniques. Although less commonlyemployed than CZE, it however contains a number ofuseful features which are not found in CZE. These featuresinclude the ability to preconcentrate dilute samples andhandle sample components with significantly differentconcentrations. ITP can be used for the analysis of manytypes of substances but is particularly suited to the analysisof small ions such as organic acids [9] and lanthanide metalcations [10]. It is therefore another method that could beapplied to the determination of magnesium. Numerousreports of the use of capillary-scale ITP for separations ofmagnesium-containing samples have been presented. How-ever, in many of these reports, the analysis of magnesiumwas not the primary target of the investigation. Therefore,in such situations, the devised methods are not necessarilyrobust for the determination of magnesium in the presenceof other common cations. A number of methods usingcapillary ITP have been proposed that include magnesiumas a target analyte in samples of surface and well waters[11], seawater [12], rainwater [13], silage [14] and plantextracts [15].

Anal Bioanal Chem (2009) 394:1299–1305DOI 10.1007/s00216-009-2603-5

J. E. Prest (*) : S. J. Baldock : P. R. Fielden :N. J. Goddard :B. J. Treves BrownSchool of Chemical Engineering and Analytical Science,The University of Manchester,P.O. Box 88, Manchester, UK M60 1QDe-mail: j.prest@manchester.ac.uk

Miniaturisation of chemical analysis systems offersnumerous potential advantages such as the ability toachieve faster separations and produce low-cost devicesand portable instruments. ITP is a technique that is highlysuitable for operation in a miniaturised format. This isbecause the preconcentration effect is useful for dilutesamples in systems with small injection volumes; thetechnique only requires simple instrumentation and con-ductivity detection, which is easily scalable, is an appro-priate detection system. Therefore, numerous miniaturisedITP devices have been proposed. These devices have beenused for a number of applications including the analysis ofsmall anions in wine [16] and anions in inorganic explosiveresidues [17]. To date, little work involving magnesium hasbeen suggested. However, magnesium is present in modelmixtures in a number of papers evaluating device perfor-mance [18, 19]. CZE which may also be applied easily in aminiaturised format, due to the similar requirement ofsimple instrumentation, has seen some use in this area.However, despite one of the earliest applications usingmagnesium to test a device [20], there has been only limitedfurther application. Methods have been suggested foranalysing water [21–23] and other beverages [24, 25] usingminiaturised CZE.

This paper describes the development of a new methodthat allowed for the determination of magnesium to bemade using miniaturised isotachophoresis. The aim of thework was to produce a method that can be applied to theanalysis of magnesium in samples such as inorganicexplosive residues or potable water. Thus, the methodneeded to be free from interference from sodium andcalcium, in particular, but also from species such aspotassium, strontium and barium.

Materials and methods

Instrumentation

The separations carried out as part of this work wereperformed on a miniaturised poly(methyl methacrylate)(PMMA) chip device. The device incorporated two mainseparation channels, the layout of which is shown in theschematic diagram depicted in Fig. 1. The channel linkingthe junction point to well B that incorporated an on-columnconductivity detector (with 75-μm-diameter platinum wire(Aldrich, Gillingham, UK) electrodes positioned in anopposed parallel arrangement) was 200 μm wide by300 μm deep, whereas all other channels were 300 μmwide by 300 μm deep. This device was fabricated in-houseand has been described previously by the authors [17].

The constant currents required to perform the separationswere provided by a PS350 high-voltage power supply

(Stanford Research Systems, Sunnyvale, CA, USA), con-figured to supply negative voltages of up to 5 kV.Conductivity detection was performed using a system builtin-house, which used capacitive coupling to ensure isola-tion of the low-voltage detection circuitry from the highvoltages used to drive the separations. Transport of electro-lytes and samples was achieved by means of a hydrody-namic system that incorporated gravity feed reservoirs forsamples and electrolytes, formed of the barrels of dispos-able plastic syringes and a series of two-way solenoidactuated valves (LFAA1201718H, The Lee Company,Westbrook, CT, USA) located at the access points to thechip, as shown in Fig. 1.

Control of the instrumentation was carried out using astandard PC with LabVIEW software (version 7.1, NationalInstruments, Austin, TX, USA). Hardware interfacing wasachieved using three National Instruments cards, a PCI-GPIB card for the power supply, a PCI-6601 timing anddigital input/output board for the detector and a PCI-6503digital input/output card for the valves. The interfacing wascontrolled using the NIDAQ driver (National Instruments),programmed using LabVIEW.

Separation conditions

A seven-step programme, shown in Table 1, was used toperform the isotachophoretic separations. Steps 1 and 2 inthis programme were used to load leading electrolyte intothe separation channels. These steps also flushed out anysolutions present from preceding analyses. In step 3, theterminating electrolyte was loaded into the device. Loadingand injection of the sample took place in steps 4 and 5, withthe channel between the cross and the junction point used

LE

TE

Waste

Waste

Sample

Conductivity detector

JunctionPointCross

A

B

C

D

E

57 mm

44 m

m

Fig. 1 Schematic diagram of the miniaturised PMMA separationdevice. The channel between the junction point and well B is 200 μmwide and 300 μm deep, whereas all other channels are 300 μm wideand 300 μm deep. Letters A, B, C, D and E refer to the wells throughwhich solutions can enter and leave the device with control of flowsmade using solenoid actuated valves. LE and TE refer to leading andterminating electrolytes, respectively

1300 J.E. Prest et al.

as a 5.1-μL fixed-volume injector. In steps 6 and 7, the actualisotachophoretic separation was performed. This was doneby applying a constant current between wells B (ground) andC (high voltage). In step 6, a current of 35 μAwas applied.This current was subsequently reduced to 20 μA in step 7 toassist in the detection of short zones. When the isotachopho-retic separation was taking place, all of the valves wereclosed, giving a hydrodynamically sealed system.

In isotachophoresis, quantitative information is containedin the step lengths and qualitative information in the stepheights. This information was extracted from the dataproduced by the conductivity detector using a LabVIEWprogramme written in-house. In this work, relative stepheight (RSH) was used as a qualitative measure. This valuewas calculated using the following expression:

RSH ¼ fS � fLEfTE � fLE

where:fLE = frequency of the response produced by the leading

electrolyte step (Hz); fS = frequency of the responseproduced by the sample step (Hz); fTE = frequency of theresponse produced by the terminating electrolyte step (Hz).

This expression includes frequency terms due to thedetector used. The design is based on an astable oscillatorwhich results in the output of a frequency rather than aresistance or voltage as is more commonly seen withconductivity detectors.

Chemicals

Electrolytes were produced using caesium hydroxidemonohydrate, hydroxyethyl cellulose (HEC; molecularweight of approximately 250,000), pivalic acid and Tris(hydroxymethyl) aminoethane (Tris; all Aldrich, Gilling-ham, UK). The following chemicals were added to theleading electrolyte to investigate potential complexingagents: N-(2-acetamido)iminodiacetic acid (ADA; Acros),N-(2-hydroxyethyl)iminodiacetic acid (HIDA) and succinic

acid (both Fluka) and malonic acid and oxalic acid (bothAldrich). Stock sample solutions, 1,000 mg L−1, wereproduced using the following chemicals: magnesium chlo-ride hexahydrate, manganese(II) chloride tetrahydrate,potassium carbonate, sodium chloride, strontium chloridehexahydrate and zinc chloride (all Aldrich), calcium chloridedihydrate (Fluka, Gillingham, UK), barium perchlorate(Acros, Loughborough, UK) and ammonium nitrate (BDH,Poole, UK). All chemicals used had purities of at least 99%with the exception of manganese(II) chloride tetrahydratewhich had a purity of 98+%. Electrolytes and samples wereprepared using >18-MΩ water (Elga Maxima Ultra Pure,Vivendi Water Systems, High Wycombe, UK).

Results and discussion

ITP electrolyte system

The initial part of this study involved finding a suitableelectrolyte system to allow the determination of magnesiumto be made using isotachophoresis. An important require-ment of the electrolyte system was that it must have littlepossibility of interference from other common cationicspecies. In particular, likely sources of interference werethought to be other alkaline earth metals and sodium. Thealkaline earth metals calcium, barium and strontium cancause problems in the determination of magnesium due tothese species having many similar chemical properties.Sodium can cause problems as it has an absolute electro-phoretic mobility of 51.9×10−9 m2 V−1 s−1 [26] which issimilar to that of magnesium which is 54.9×10−9 m2 V−1

s−1 [26]. Of these potential interferences, calcium andsodium are likely to be present, at potentially highconcentrations, in samples for many applications. There-fore, it was particularly important that the devised methodgave good resolution between magnesium and these twospecies. Barium and strontium are less likely to beencountered in samples but can be present in residues fromimprovised explosive devices that have been producedusing pyrotechnics.

In isotachophoresis, there are a number of mechanisms,such as changing the pH at which the separation is carriedout, changing the leading ion concentration or usingcomplexing agents, that can be used to assist in achievinga specific separation. For this work, changing the pH wouldhave little effect on improving the separation as all of thespecies mentioned above are strong bases. Changing theleading ion concentration could assist the separation ofmagnesium from monovalent sodium but would not helpthe separation from the other alkaline earth metals.Therefore, it was thought that the most useful method ofachieving a good separation of magnesium from other

Table 1 Separation programme used for performing miniaturised ITPon a PMMA chip

Step Time, s Current, μA Valve status

A B C D E

1 20 0 ■ □ ■ ■ □2 20 0 ■ □ ■ □ ■3 1 0 ■ ■ □ □ ■4 0.5 0 □ ■ ■ □ ■5 0.3 0 □ ■ ■ ■ □6 200 35 ■ ■ ■ ■ ■7 1,000 20 ■ ■ ■ ■ ■

■ closed, □ open

A miniaturised isotachophoresis method for magnesium determination 1301

species was to incorporate a complexing agent as a co-counter ion in the leading electrolyte. There was a widevariety of substances that could potentially be used for thispurpose as magnesium forms complexes with manyligands. A number of previous studies using capillaryisotachophoresis have suggested some complexing agentsthat could be used to change the effective mobility ofmagnesium. Successful isotachophoretic separations ofmagnesium, calcium, strontium, barium and sodium weremade through the use of HIDA [27]. The incorporation of1,2-cyclohexanediamine-N,N,N′,N′-tetraacetic acid into theleading electrolyte was also found to permit a similarseparation, albeit with only small differences in mobilitiesbetween some of the species, notably calcium and magne-sium [28]. Other methods that have been applied tomanipulate the effective mobilities of calcium and mag-nesium in capillary isotachophoretic separations haveinvolved the use of ethylenediaminetetraacetic acid (EDTA)[12], a combination of 18-crown-6 ether and α-hydroxyi-sobutyric acid [29], ADA [15], nitrilotrismethylenephos-phonium acid in an unbuffered system [11] and acombination of 18-crown-6 ether and sulphuric acid in anunbuffered system [14]. The use of EDTA involvescarrying out an anionic separation and involves a methodthat is complicated by the fact that a zone of non-complexed ligand also migrates isotachophoretically. How-ever, these studies do not include information of thebehaviour of barium and/or strontium with the devisedelectrolyte systems. Therefore, it is not certain whetherthese methods would allow the required magnesiumseparation to be achieved. Another approach that enabledthe separation of magnesium, calcium, strontium, bariumand sodium using capillary isotachophoresis was to usewater–polyethylene glycol (PEG) mixtures as the leadingelectrolyte solvent [30]. However, the relatively high PEGcontent, 45% v/v, required for the separation to be achievedmeans this method may not be suitable for use in aminiaturised device with hydrodynamic fluid transport.

In this work, five electrolyte systems were tried to assesswhether they were suitable for allowing the isotachophoreticdetermination of magnesium to be made. These systems usedthe following species as complexing agents: ADA, HIDA,malonic acid, oxalic acid and succinic acid. The HIDA systemwas selected on the basis of its previous successful use incapillary-scale isotachophoresis [27]. Despite the lack ofinformation regarding the behaviour of barium andstrontium, the ADA system was deemed worthy of furtherconsideration due to it previously being shown to offer anexcellent separation of calcium and magnesium [15]. The useof malonic acid, oxalic acid and succinic acid was selectedon the basis of the stability constants of the complexesformed between alkaline earth metals and these species [31].The devised systems were essentially identical in all aspectswith the exception of the complexing agent incorporated intothe leading electrolyte. Full details of the compositions of theelectrolyte systems are given in Table 2.

The systems were evaluated by performing a similar setof experiments with each of the different leading electro-lytes. For comparative purposes, separations were alsocarried out using an electrolyte system with similarconditions but containing no complexing agent (full detailsof this system, denoted as pivalic, are also given in Table 2).These experiments involved performing a series of separa-tions of single-component samples. The samples used forthese comparisons were 30 mg L−1 barium, 20 mg L−1

calcium, 10 mg L−1 magnesium, 10 mg L−1 sodium and25 mg L−1 strontium. For each of the samples, the RSHswere calculated and the results obtained are displayed inFig. 2.

The experiments showed that the different complexingagents had different effects on the mobilities of the differentcations. This effect can be seen in Fig. 2 in the changes inRSH values observed. In ITP, the step height, and henceRSH, is related to the effective mobility of a particular ion.In three of the systems, those incorporating ADA, HIDAand succinic acid, the complexing agent was found to retard

Table 2 Composition of electrolyte systems that were investigated to identify a suitable system to allow the determination of magnesium

Electrolyte system ADA Oxalic Succinic HIDA Malonic Pivalic

Leading electrolyte 10 mmol L−1

CsOH10 mmol L−1

CsOH10 mmol L−1

CsOH10 mmol L−1

CsOH10 mmol L−1

CsOH10 mmol L−1

CsOHComplexing agent 2 mmol L−1

ADA1 mmol L−1

oxalic acid2 mmol L−1

succinic acid2 mmol L−1

HIDA2 mmol L−1

malonic acid–

pH buffer Pivalic acid Pivalic acid Pivalic acid Pivalic acid Pivalic acid Pivalic acidpH 5.1 5.1 5.1 5.1 5.1 5.1Additive 1 mg mL−1

HEC1 mg mL−1

HEC1 mg mL−1

HEC1 mg mL−1

HEC1 mg mL−1

HEC1 mg mL−1

HECTerminatingelectrolyte

10 mmol L−1

Tris10 mmol L−1

Tris10 mmol L−1

Tris10 mmol L−1

Tris10 mmol L−1

Tris10 mmol L−1

Tris

ADA N-(2-acetamido)iminodiacetic acid, HEC hydroxyethyl cellulose, HIDA N-(2-hydroxyethyl)iminodiacetic acid

1302 J.E. Prest et al.

the effective mobility of calcium more strongly than theother cations. This result is not ideal as it could lead to anincrease in the possibility of co-migration between calciumand magnesium. This is because in the system without anyadditional complexing agent calcium has a higher effectivemobility, exhibiting an RSH±standard deviation (SD) of0.446±0.024 (n=3), than that of magnesium, whichexhibits an RSH±SD of 0.569±0.008 (n=3). The resultsobtained indicated that with the systems containing HIDAor succinic acid it would be unlikely that a magnesium stepwould be obtained that was clearly separated from calcium.It was thought that in this work the smaller difference incalcium to magnesium resolution with HIDA compared tothat achieved with capillary-scale ITP [27] was due to thepH used. In the previous report using capillary ITP, pHlevels of 6.2 or 6.8 were used whereas in this work the pHwas 5.1. With ADA, it should be possible to separatemagnesium from both calcium and sodium. However, toallow a robust determination of magnesium to be made, itwas thought that a system offering a better resolution fromsodium would be preferable.

The two systems that result in the largest change in thebehaviour of magnesium are those containing malonic acidand oxalic acid as complexing agents. Of these, the lattercomplexing agent has the most significant effect on themobility of magnesium. This fits in with the stabilitycomplexes of the magnesium complexes. These are 3.43[31] and 2.85 [31] for the oxalate and malonate complexes,respectively (as a comparison, those of succinate complexesare 2.15 [31]). Both of these systems would seem to besuitable for the analysis of magnesium. However, it wasthought that if oxalic acid was used problems could beencountered with certain samples due to the low solubility

of oxalate complexes [26]. Indeed, some unusual behaviourwas noted with the oxalate system. In particular, shorterthan expected zones were noted with calcium, a problemthat may have been related to precipitation effects. Aprevious study noted that multiple steps were observed foroxalate in a system incorporating calcium as a complexingagent, which the authors attributed to precipitation prob-lems [32]. Therefore, it was decided to investigate furtherthe use of the malonic acid system.

Additional separations were carried out to see whetherother cations may cause interference in the determination ofmagnesium using the malonic acid system. Two furtherspecies that could be found in many samples, particularlythose from an agricultural environment or explosiveresidues, were potassium and ammonium. However, inves-tigations using single-component samples of these cationsshowed that no interference would occur. This is becausethe species have a high mobility and both were found toeither co-migrate with the leading ion or form poorlyresolved low steps. Checks were also carried out to seewhether any problems would occur due to the presence oftransition metals. To do this, experiments were carried outusing 20 mg L−1 manganese(II) as a sample. This specieswas found to yield a step with an RSH±SD of 0.939±0.040. This is somewhat higher than that of magnesium soit does not cause any interference problems. As manganese(II) generally forms weaker complexes than the other first-row transition metals, iron(II), cobalt(II), nickel(II) andcopper(II), no interference is likely from these otherspecies. Zinc(II) often behaves in a similar manner tomagnesium so the behaviour of this species was alsoinvestigated. When a sample of 20 mg L−1 zinc(II) wasanalysed, no obvious step was seen suggesting that theeffective mobility of this species was lower than that ofTris, the terminating ion. This result fits with a previousstudy that found that zinc(II) formed stronger complexeswith malonic acid than magnesium did [33].

Analytical performance

As the miniaturised ITP method that employed malonicacid as a complexing agent was found to offer a means ofanalysing magnesium without interference from commoncations, it was subsequently used to carry out a range ofseparations involving mixtures. Initially, a series of modelsamples was investigated. An example of an isotachophero-gram obtained with such a system is shown in Fig. 3. Theexample shown is for a mixture of barium and magnesium.The result clearly shows that both of these species andsodium that was present as background contamination gaveclear steps, which were well resolved from one another. Theseparation shown in Fig. 3 indicated that the method wouldalso be suitable for performing barium determinations. If

Rel

ativ

e S

tep

Hei

ght

Electrolyte System

ADA Oxalic Succinic HIDA Malonic

Ca2+

Na+

Mg2+

Sr2+

Ba2+

0.0

0.2

0.4

0.6

0.8

1.0

Pivalic

Fig. 2 Graph showing the effect of the addition of complexing agentsto the leading electrolyte on the relative step heights of sodium andalkaline earth cations. Full details of the compositions of theelectrolyte systems used are detailed in Table 2. The error barsshown represent two standard deviations based on three replicate runs

A miniaturised isotachophoresis method for magnesium determination 1303

strontium or calcium were present in mixtures, thesespecies were found to form mixed zones with sodium.

To evaluate the analytical performance of the devisedmethod, a calibration curve was produced for magnesium.This was done by performing a series of separations with11 samples containing magnesium concentrations in therange 0.625 to 75 mg L−1. Good results were obtained withthese experiments which involved analysing each sample infour repeat runs. In these experiments, the step height,which in ITP contains the qualitative information in theseparation, was found to be repeatable. Across the wholeset of samples analysed, the RSH±SD of the magnesiumstep was found to be 0.673±0.030 (n=44). Good repro-ducibility in the quantitative information contained from anisotachophoretic separation, the step length, was alsoobserved. The relative standard deviations (RSDs) of thesteps noted with the different samples were all found to fallin the range of 1.4% to 6.5%. A calibration curve wasproduced from the results using weighted linear regression.The equation of the curve was calculated to have anintercept of −0.2 s and a slope of 3.3 s L mg−1, with SDs ofthe slope and intercept of 0.42 and 0.13, respectively. Thecurve exhibited good linearity with a correlation coefficientof 0.992 calculated. The calibration curve was used toestimate a limit of detection (LOD) for magnesium usingthe miniaturised ITP method. The method used to calculatethe LOD was based on a blank result plus three times thestandard deviation of that value. In ITP, you do not get ameaningful blank result, unless background contaminationis present, so the LOD was calculated using an estimatedblank. In most instances, the intercept of the calibration

curve can be used to calculate an estimated blank.However, in this work, the intercept had a small negativevalue, which could lead to the possibility of a negativeLOD. Thus, the LOD was calculated as zero plus threetimes the standard deviation of the intercept and was foundto be 0.45 mg L−1. This figure for the miniaturised ITPmethod is of a similar order of magnitude, albeit higher, tothat achieved using miniaturised CZE [23] whilst beingapplicable for a wider range of sample concentrations. TheLOD is lower than the level that would be required in manyapplications, such as using magnesium in blood as a markerfor pulmonary disease [34].

The method was then tested on the analysis of somewater samples. The samples used were three still mineralwaters commercially available in the UK, Brecon Carrog,Fairbourne Spring and Highland Spring, and tap water. Inall cases, a step for magnesium was identified. An exampleof such a separation is shown in Fig. 4. This result, whichwas for a sample of Highland Spring mineral water, showsa clear magnesium step that is well separated from themixed zone arising from calcium and sodium. Goodreproducibility was noted for the magnesium steps in thewater samples. The RSDs of the RSHs were between 0.6%and 3.6% and the zone lengths were between 2.0% and5.5%. Using the calibration curve, the amount of magne-sium present in the samples was found to be 2.8 mg L−1 inthe tap water, 5.5 mg L−1 in Fairbourne Spring water,6.9 mg L−1 in Highland Spring water and 13.3 mg L−1 inBrecon Carrog water. For the mineral waters, this representsa reasonable agreement with the magnesium concentrationsstated on the bottles which were 4.4 mg L−1, 8.0 mg L−1

and 16.5 mg L−1, respectively. All of the mineral waters

Time/s

Fre

quen

cy R

espo

nse

/ kH

z

344

346

348

350

250 290 330 370

Ld

Tm

Ba2+Na+

Mg2+

Fig. 3 Separation of cations using the malonic acid containingelectrolyte system. The sample illustrated contained 7.5 mg L−1

magnesium and 30 mg L−1 barium achieved together with backgroundsodium contamination. Leading electrolyte 10 mmol L−1 caesiumhydroxide, 2 mmol L−1 malonic acid, pH 5.1 (pivalic acid), with 1 mgmL−1 HEC added. Terminating electrolyte 10 mmol L−1 Tris. Ld =leading ion; Tm = terminating ion

346

348

350

300 350 400 450 500

352

Ld

Tm

Ca & Na2+ +

Mg2+

Time/s

Fre

quen

cy R

espo

nse

/ kH

z

Fig. 4 Isotachopherogram of the analysis of a sample of HighlandSpring mineral water. Leading electrolyte 10 mmol L−1 caesiumhydroxide, 2 mmol L−1 malonic acid, pH 5.1 (pivalic acid), with 1 mgmL−1 HEC added. Terminating electrolyte 10 mmol L−1 Tris. Ld =leading ion; Tm = terminating ion

1304 J.E. Prest et al.

contain higher concentrations of calcium than magnesium.In particular, Fairbourne Spring contains high concentra-tions of both calcium, 55 mg L−1, and sodium, 31.2 mg L−1.The analysis of this sample shows the ability of the methodto allow the determination of magnesium to be made in thepresence of significant amounts of other species. With theFairbourne spring water sample, the mixed zone resultingfrom the calcium and sodium had a length±SD of 187.6±7.0 s (n=3) that was considerably longer than that ofmagnesium of 18.2±1.0 s (n=3).

Conclusions

A range of potential complexing agents were tried to find asuitable electrolyte system to allow the determination ofmagnesium to be made using miniaturised isotachophoresis.These studies showed that the use of malonic acid as acomplexing agent in isotachophoresis was effective inproducing a method that was highly selective towards mag-nesium. Using a leading electrolyte of 10 mmol L−1 caesiumhydroxide at pH 5.1 incorporating 2 mmol L−1 of malonicacid, the effective mobility of magnesium was retardedsignificantly, allowing this species to be comprehensivelyseparated from sodium, calcium, strontium and barium.

The new procedure was designed to allow the determi-nation to be made without interference from cations thatcould be found in inorganic explosive residues so it shouldbe suitable for use in applications requiring the analysis ofsuch samples. The selectivity of the method towardsmagnesium means that it should also be suitable for wateranalysis and may be applicable to blood analysis and urineanalysis. The method was found to offer good linearity,with a correlation coefficient of 0.992, over the concentra-tion range from 0.625 to 75 mg L−1 and a limit of detectionof 0.45 mg L−1.

Acknowledgements The authors would like to thank the UK HomeOffice and the Engineering and Physical Sciences Research Council(UK) for funding this research programme and the ForensicExplosives Laboratory of the Defence Science and TechnologyLaboratory, Fort Halstead, for their support.

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