Magnetic techniques for the detection and … Magnetic techniques for the detection and...

REVIEW

Magnetic techniques for the detection and determinationof xenobiotics and cells in water

Ivo Safarik & Katerina Horska & Kristyna Pospiskova &

Mirka Safarikova

Received: 15 February 2012 /Revised: 15 April 2012 /Accepted: 16 April 2012 /Published online: 12 May 2012# Springer-Verlag 2012

Abstract Magnetic techniques based on the application ofmagnetic nanoparticles and microparticles and films havebeen successfully used for the determination and detectionof different types of xenobiotics (e.g. herbicides, insecti-cides, fungicides, aromatic and polyaromatic hydrocarbons,pentachlorophenol and heavy metal ions) as well as viruses,microbial pathogens and protozoan parasites in water sam-ples. Preconcentration of xenobiotics from large volumes ofsamples can be performed using magnetic solid-phaseextraction, stir-bar sorptive extraction and related proce-dures. This review provides basic information about thesetechniques. Published examples of successful applicationsdocument the importance of these simple and efficient pro-cedures employing magnetic materials.

Keywords Bioanalytical methods . Enzymes .

Immunoassays/ELISA . Organic compounds . Water .

Preconcentration

Introduction

Human activity is accompanied by the production of bothorganic and inorganic xenobiotics (contaminants). These

contaminants are usually analysed using highly sophisticat-ed analytical techniques. High-performance liquid chro-matography and gas chromatography systems equippedwith different types of detectors are routinely used for assaysof organic xenobiotics, whereas atomic absorption spectro-photometry and inductively coupled plasma mass spectrosco-py are used for assays of inorganic xenobiotics. Thesemethods usually require extensive analyte preconcentration,experienced technical staff and expensive equipment andreagents.

As a consequence, attention has also been focused on thedevelopment of new methods enabling relatively simple,rapid and sensitive assays. Immunoassays seem to be methodsof choice, both for screening and for analytical purposes; theyhave been used for many years in clinical chemistry as reli-able, sensitive and selective methods to determine low con-centrations of organic compounds in, for example, blood,urine and tissue extracts [1].

The possibility to use immunoassays for environmentalstudies was recognized in the early 1970s. Immunoassayshave been developed for a broad range of pesticides andcontaminants of industrial origin. This interesting topichas been reviewed in several articles and book chapters[1–5].

Immunoassays can benefit from the application of mag-netic nanoparticles or microparticles. In general, magneti-cally responsive particles can be used in an enormousnumber of applications, ranging from molecular biology towastewater treatment [6–8]. Magnetic particles exhibit thefollowing important properties [8, 9]:

1. Selective separation and removal of magnetically respon-sive nanoparticles and microparticles and other relevantmaterials from complex samples using an external mag-netic field (e.g. an appropriate magnetic separator, perma-nent magnet, or electromagnet).

2. Targeting and localization of magnetic particles to thedesired place using an external magnetic field (e.g.

I. Safarik (*) :K. Horska :M. SafarikovaDepartment of Nanobiotechnology,Institute of Nanobiology and Structural Biology of GCRC,Na Sadkach 7, 370 05 Ceske Budejovice, Czech Republice-mail: [email protected]

I. SafarikRegional Centre of Advanced Technologies and Materials,Palacky University, Slechtitelu 11, 783 71 Olomouc,Czech Republic

K. PospiskovaDepartment of Biochemistry, Faculty of Science,Palacky University, Slechtitelu 11, 783 71 Olomouc,Czech Republic

Anal Bioanal Chem (2012) 404:1257–1273DOI 10.1007/s00216-012-6056-x

sealing the rotating objects using magnetic fluids ormagnetic drug targeting).

3. Heat production caused by magnetic particles subjectedto a high-frequency alternating magnetic field. Thisphenomenon is employed especially during magneticfluid hyperthermia, e.g., for cancer treatment.

4. Increase of a negative T2 contrast by magnetic iron oxidenanoparticles during magnetic resonance imaging.

5. Great increase of apparent viscosity of magnetorheolog-ical fluids when they are subjected to a magnetic field.

6. Magnetic nanoparticles and microparticles can be usedfor magnetic modification of diamagnetic biologicalmaterials (e.g. cells) and magnetic labelling of biologi-cally active compounds (e.g. antibodies and enzymes).This property can be efficiently employed in the devel-opment of magnetic assays for determination and detec-tion of xenobiotics and cells.

The conversion of classic immunoassays (performed, e.g.,in microtitration plates) to the magnetic version (performedmainly in test tubes with the help of magnetic separators)opened new possibilities for their applications, such as theiruse during field surveys. Two types of magnetic-particle-based immunoassays can be distinguished:

1. Immunomagnetic assays, where an appropriate antibody(or antibodies) is immobilized on a magnetic carrier(instead of in the wells of the microtitration plates)

2. Magnetoimmunoassays, where magnetic particles serveas a detectable label (instead of the enzymes, radio-nuclides or luminescent molecules used in standardimmunoassays)

Immunomagnetic assays are very similar to standardmicrotitration plate assays; the main difference is that spe-cific antibodies are bound to the magnetically responsiveparticles, which can be separated from the suspension usingappropriate magnetic separators based on strong rare-earthpermanent magnets.

Immunomagnetic techniques have also been successfullyused for the detection of pathogenic bacteria and viruses infood and clinical microbiology [10, 11]. This technique hasalso been employed in environmental microbiology for thedetection of target viruses, bacteria and protozoan parasitesin water samples of different origin.

Also enzymes (e.g. acetylcholinesterase) immobilized onmagnetic carriers can be used for the detection of xeno-biotics acting as enzyme inhibitors, such as carbofuran,paraoxon, malaoxon, and paraoxon-methyl [12].

In many cases the target analytes present in water sam-ples have to be preconcentrated before appropriate analyti-cal procedures are performed. Currently, preconcentration isusually performed using liquid–liquid extraction or solid-phase extraction (SPE) processes. Recently, a new

extraction technique based on the application of magnetical-ly responsive adsorbents has been developed; this techniqueis known as magnetic SPE (MSPE) [13]. Another technique,called stir-bar sorptive extraction (SBSE), is based on theuse of a magnetic stir bar covered with a layer of an appro-priate adsorbent [14].

The synthesis of magnetic ferrites has been used to removemetal pollutants from wastewater. The method involves theincorporation of metal ions in the ferrite structure during itssynthesis by a coprecipitation method [15–17].

It can be clearly seen that magnetically responsive mate-rials can be widely used in detection and analytical systemsbecause of the performance advantages they offer comparedwith similar solids that lack magnetic properties [15]. All theabove-mentioned procedures will be briefly summarized inthe following sections.

Immunomagnetic techniques for detectionand determination of xenobiotics, viruses and cells

Different types of magnetic nanoparticles and microparticlesare available commercially. The magnetic particles for theassays have to enable both efficient immobilization of anti-bodies and other biomolecules (e.g. enzymes) and theirrapid separation using a simple permanent magnet or mag-netic separator. For commercially available RaPID Assaykits, intended for determination of xenobiotics, silanizedmagnetic iron oxide microparticles are used as a carrier.Immunomagnetic separations of cells are usually based onthe use of Dynabeads.

Currently, two commercially available assay systems fordetermination of xenobiotics based on the application ofmagnetic particles for antibody immobilization are used,namely, RaPID Assay (SDIX, formerly Strategic Diagnostics,USA) and Abraxis (Abraxis, USA) kits (see Fig. 1).

A typical assay can be performed as follows. In thisexample, the volumes and times for both SDIX and Abraxisatrazine assays will be reviewed. The protocols for otheranalytes are similar. First, 200 μL of water sample is directlycombined with 250 μL of the atrazine enzyme conjugateand 500 μL of antibody-coupled magnetic particle suspen-sion in a test tube. After vortexing, this mixture is incubatedfor 15 min, and then the magnetic particles are pulled to theside of the test tube by placing it in a special magnetic rackfor 2 min. The particles are rinsed twice with washingsolution and then 500 μL of colour reagent (containinghydrogen peroxide and 3,3′,5,5′-tetramethylbenzidine in anorganic base) is added. After 20 min, this reaction is stoppedwith 500 μL of sulfuric acid solution and the absorbance ofthe tube contents is measured (after magnetic separation) ina spectrophotometer at 450 nm. Because a competitiveimmunoassay is used, where the antigen (analyte) in the

1258 I. Safarik et al.

analysed sample competes with enzyme (horseradish perox-idase) labelled antigen to bind with immobilized antibodies,colour development is inversely proportional to the analyteconcentration; a darker colour corresponds to lower analyteconcentration and vice versa. The analyte concentration inan unknown sample is calculated using the standard calibra-tion curve generated from kit standards after performing thereaction and reading with a spectrophotometer.

The kits currently available commercially, together withthe applications described in the area of water contaminantstudies and the sensitivities of the kits, are presented inTable 1.

The spectrophotometric detection of the reaction productduring the immunoassays is very simple and is the basis ofportable systems enabling field analyses. However, manystudies have been performed recently where immunomag-netic particles were coupled with electrochemical sensors[18, 19]. The main advantage of such a combination is thesubstantial decrease of the detection limit (e.g. 50 timeslower for PCBs) in comparison with standard magneticimmunoassays with spectrophotometric detection [20, 21].

Immunomagnetic assays for determination of pesticideswere compared with “standard” analytical procedures, suchas GC-MS. In the case of atrazine determination, the RaPIDAssay kit was internally consistent with a variation of lessthan 3 % between 156 duplicate standards that were run toproduce standard curves. For the 217 samples examined,atrazine concentrations measured by the two analytical tech-niques were highly correlated (r00.96). Compared with theconcentrations measured by GC-MS, the magnetic assay pro-duced no false negatives and few false positives (5.53 %). Themagnetic assay tended to overestimate atrazine concentrationsslightly, probably the result of cross-reaction in the assay byatrazine metabolites and structurally related triazine herbi-cides. Even so, the immunomagnetic assays proved to be areliable, relatively simple and cost-effective screening tech-nique for atrazine contamination [22].

Not only the individual chemical compounds, but alsoviruses and whole cells can be successfully separated using

magnetic particles. Magnetic separation of viruses and cellshas several advantages in comparison with other techniquesused for the same purpose. It permits the target cells to beisolated directly from crude materials such as environmentalwater samples, sediments, soil, blood, bone marrow, tissuehomogenates, stool, fermentation media and food. Com-pared with other conventional methods of cell separation,magnetic separation is relatively simple and fast. The largedifferences between the magnetic permeabilities of the mag-netic and non-magnetic cells enable highly selective sepa-rations. Cells isolated by magnetic separation processes areusually pure, viable and unaltered. The whole separationprocess can be performed in the same tube running multiplesamples simultaneously in a fraction of the usual time.Separated microbial cells are usually grown on selectiveagars [10].

Immunomagnetic separation employs magnetic particleswith bound antibodies specific against the target (micro)organism or virus to be separated. Recently, immunomag-netic separation has been successfully used in water virolo-gy, microbiology and parasitology to detect important viral,microbial and parasitic contamination. Table 2 shows sev-eral typical applications of immunomagnetic separation fordetection of viruses, whereas Table 3 presents examples ofsuccessful separation of important microbial pathogens andTable 4 examples of successful separation of protozoanparasites. Several immunomagnetic beads are availablecommercially, especially from Invitrogen; namely Dyna-beads anti-Salmonella, Dynabeads anti-E. coli O157, Dyna-beads EPEC/VTEC O26, Dynabeads EPEC/VTEC O103,Dynabeads EPEC/VTEC O111, Dynabeads EPEC/VTECO145, Dynabeads anti-Legionella and Dynabeads anti-Lis-teria. The same company also produces immunomagneticparticles for the separation of parasitic protozoa, e.g. Cryp-tosporidium oocysts (Dynabeads anti-Cryptosporidium) andsimultaneous separation of Cryptosporidium oocysts andGiardia cysts (Dynabeads GC-Combo). The diameter ofthe magnetic particles used for immobilization of antibodies(Dynabeads) is 2.8 μm. Analogous immunomagnetic beadscan also be obtained from Lab M (UK), namely CaptivateO26, Captivate O103, Captivate O111, Captivate O145,Captivate O157 and Captivate Salmonella. However, immu-nomagnetic particles can also be prepared in the laboratoryusing appropriate magnetic beads and an appropriate anti-body [23] (see Fig. 2).

Immunomagnetic separation has been implemented intostandard procedures for detection of parasites in water—USEPA method 1622 (detection of Cryptosporidium) [24] andUS EPA method 1623 (detection of Cryptosporidium andGiardia) [25]. Both methods employ a relatively simpleprocedure. A water sample is filtered and the oocysts, cystsand extraneous materials are retained on the filter. Althoughthe EPA has only validated the method using laboratory

Fig. 1 Abraxis kit for the determination of atrazine. (Reproduced withthe permission of Abraxis, USA)

Magnetic techniques for the detection of xenobiotics in water 1259

Tab

le1

Examples

ofxeno

biotic

assays

inwater

samples

usingcommercially

availableim

mun

omagnetic

kitsandph

otom

eter

measurement

Xenobiotics

Characteristics

Kitused

Assay

range(ppb)

Water

system

Com

ments

Reference

Acetochlor

Herbicide,pesticide

Abraxis

0.10–2.5

Wastewater

treatm

entplanteffluents

Acetochloras

themajor

toxicant

intank

truck

cleaning

wastewater

effluent

[55]

Atrazine

Herbicide,pesticide

RaPID

Assay

0.4-5.0

Estuarine

waters

Com

parisonwith

GC

[56]

Ebroriverwater

Annualmonito

ring

(from

March

1995

toJune

1996)

[57]

Realestuarine,

coastalandspiked

water

samples

Evaluationof

2differentim

munoassays;solid

-phase

extractio

nfollo

wed

byLC-D

AD

[58]

Surface

water

inOntario,Canada

Spatialandseasonal

variations

measurement

[59]

Abraxis

0.1–5.0

Water

from

drinking

water

treatm

entplants

Study

ofatrazine

occurrence

intheUSA

[60]

Water

from

atrazine-spikedmicrocosm

sStudy

ofatrazine

asachem

ical

stressor

[61]

Water

samples

from

industrial

effluents

Com

parisonwith

SPEfollo

wed

byLC-A

PCI-MS

[62]

Carbofuran

Insecticide

RaPID

Assay

0.06-5.0

Water

samples

from

Daphnia

magna

experiments

Study

ofsimultaneousexposure

ofDaphnia

magna

tosuspendedsolid

sandcarbofuran

[63,

64]

Fish,

bluegills

(Lepom

ismacrochirus)

Com

parisonof

2commercial

immunoassays

[65]

2,4-Dichloro-

phenoxyacetic

acid

Herbicide

RaPID

Assay

0.7-50.0

12surfacewatersin

thePiedm

ont,USA

12%

ofsamples

tested

positiv

e[66]

17β-Estradiol

Estrogen

Abraxis

1.5-25.0

Realandspiked

water

samples

Com

parisonwith

chromatographytechniques

[67]

Glyphosate

Herbicide

Abraxis

0.075-4.0

Surface

water

Evaluationof

theanalytical

procedure

[68]

Metolachlor

Herbicide

RaPID

Assay

0.05-5.0

Surface

runoff,tiledrainage

Com

parisonwith

SPE-G

CandSPME-G

C[69]

12surfacewatersin

thePiedm

ont,USA

66%

ofsamples

tested

positiv

e[66]

Surface

water

samples

Com

parisonof

ELISA

andGC/M

S[70]

Surface

water

inOntario,Canada

Spatialandseasonal

variations

measurement

[59]

PAHs

PAHs

RaPID

Assay

0.7-50.0

River

water

samples

Com

parisonwith

GC-M

Stechniques

[71]

CarcinogenicPA

Hs

PAHswith

4or

morerings

RaPID

Assay

0.2-10.0

Environmentalsedimentsamples

from

thePatos

Lagoonestuary,

southern

Brazil

Com

parisonwith

GC-FID

[72]

Water

samples

from

industrial

effluents

Com

parisonwith

SPEfollo

wed

byLC-A

PCI-MS

[62]

PBDEs

PDBEs

Abraxis

0.025-1.0

Water

from

amunicipal

water

source,areservoir,alake

andapond

BDE-47andBDE-99congenersmostreadily

recognized

[73]

Brackishwatersin

theHaw

aiianIslands

Analysisof

PBDEsin

Haw

aiianeuryhalin

efish

andcrabs

[74]

PCP

Pesticide,woodpreservativ

eRaPID

Assay

0.06-10.0

Water

samples

from

industrial

effluents

Com

parisonwith

SPEfollo

wed

byLC-APCI-MS

[62]

Pyrethroids

Insecticide

Abraxis

1.0-15.0

Surface

waters

Monito

ring

inSan

Juan

County,WA,USA

[75]

Spinosad

Insecticide

RaPID

Assay

0.02-1.0

Spikedwater

samples

Methodology

developm

ent

[76]

Sulfamethazine

Antibacterial

agent

Abraxis

0.05-5.0

Wastewater

from

variousstages

inwastewater

treatm

entplants

SPEandLC-M

S/M

Sused

forcomparison

[77]

Triclopyr

Herbicide

RaPID

Assay

0.03-3.0

Leachatefrom

soilcolumns

Study

ofpesticidetransport

[78]

Triclosan

Antibacterial

agent

Abraxis

0.02-1.0

Wastewater

treatm

entplantinfluentsandeffluents

Allwastewater

samples

tested

show

edthepresence

oftriclosan

[79]

Wastewater

treatm

entplantinfluentsandeffluents

Triclosan

toxicity

onbiofilm

algaeandbacteria

exam

ined

[80]

The

kits

mentio

nedwereob

tained

from

SDIX

(formerly

Strategic

Diagn

ostics;

RaPID

Assay)andAbraxis

during

2011.Alsokits

fortheassays

ofalachlor,metolachlor

(Abraxis),beno

myl/

carbendazim

(RaPID

Assay)andPCBs(bothAbraxisandRaPID

Assay)wereavailable.

PAHpo

lycyclicarom

atichy

drocarbo

n,PDBEpo

lybrom

inated

diph

enyl

ether,PCPpentachlorop

heno

l,GCgaschromatog

raph

y,LCliq

uidchromatog

raph

y,DADdiod

e-arraydetection,

SPEsolid

-ph

aseextractio

n,ACPIatmosph

eric

pressure

chem

ical

ionizatio

n,MSmassspectrom

etry,SP

MEsolid

-phase

microextractio

n,FID

flam

eionizatio

ndetection


filtration of bulk water samples shipped from the field, fieldfiltration may also be used. Then, materials on the filter areeluted and the eluate is centrifuged to pelletize the oocystsand cysts, and the supernatant fluid is aspirated. The oocystsand cysts are magnetized by attachment of magnetic beadsconjugated to anti-Cryptosporidium antibodies (method1622) or magnetic beads conjugated to anti-Cryptosporidiumand anti-Giardia antibodies (method 1623). The magnetizedoocysts and cysts are separated from the extraneous materialsusing a magnet, and the extraneous materials are discarded.The magnetic bead complex is then detached from the oocystsand cysts. The oocysts and cysts are stained on well slideswith fluorescently labelled monoclonal antibodies and 4′,6-diamidino-2-phenylindole. The stained sample is examinedusing fluorescence and differential interference contrast mi-croscopy. Qualitative analysis is performed by scanning eachslide well for objects that meet the size, shape and fluores-cence characteristics of Cryptosporidium oocysts or Giardiacysts. Quantitative analysis is performed by counting the totalnumber of objects on the slide confirmed as oocysts or cysts.

Pathogenic microorganisms obtained using immunomag-netic separation can also be detected using molecular biol-ogy methods, especially polymerase chain reaction (PCR).Detection limits using molecular methods such as PCR maybe lower when compared with conventional growth-basedassays, and also have the advantage of increased specificity.Achieving low detection limits in any environmental path-ogen assay is of paramount importance, especially in water

samples, where the presence of a single organism may resultin human illness. However, successful PCR requires nucleicacid that is free from inhibitors and interfering compounds.Nucleic acids have to be released from the concentrated micro-organisms after chemical or mechanical disintegration. Mag-netic particles coated with streptavidin can be used to bindbiotin-labelled oligonucleotide capture probes. The strong af-finity between biotin and streptavidin (KD010

-15 M) per-mits the separation of hybrid from non-target nucleic acid,interfering compounds and chemical species. This techniqueof combining magnetic capture hybridization with PCR hasbeen applied to pathogen detection in a wide variety of samplematrices, including water samples containing Salmonella[26].

Magnetic techniques with immobilized enzymes

The mechanism of action of many pesticides and otherxenobiotics is designed to impact on a specific enzymefound within the target organism or even an unintendedenzyme. Organophosphate and carbamate pesticides aredesigned to inhibit acetylcholinesterase, and this enzymehas been used most often in enzymatic detection of thesepesticides. Also, other enzymes can be inhibited by thepesticides and other xenobiotics and the extent of inhi-bition is correlated to the concentration of the analyte[27].

Table 2 Examples of (immuno)magnetic separation of viruses from water samples

Virus Water sample Magnetic system Comments Reference

Adenoviruses Water from theTamagawa river, Japan

Dynabeads M-280 sheep anti-mouseIgG with bound mouse antiadenovirusmonoclonal antibodies

Combination of IMS with directquantitative real-time PCR

[81]

Enteroviruses Seeded environmentalwater samples

Dynabeads M-280 sheep anti-mouseIgG coated with mouse antienterovirusmonoclonal antibody

Comparison with reportercell system responding to viralreplication based on fluorescentresonance energy transfer

[82]

Hepatitis A virus Spiked surface river waterand seawater samples

Dynabeads M-280 with bound murinemonoclonal antibodies

Combination of IMS with reversetranscription/nested PCR

[83]

Spiked environmentalwater samples

Streptavidin MagneSphereparamagnetic particles (Promega)with bound biotinylatedrabbit anti-HAV IgG

Combination of IMS with reversetranscription PCR

[84]

Noroviruses Seeded environmentalwater samples

MyOne streptavidin-coated magneticbeads with bound synthetic biotinylatedhisto-blood group antigens

Histo-blood group antigensused as binding moleculesinstead of antibodies

[85]

Norwalk-like virus Environmentalwater samples

Rabbit polyclonal antibodies bound toDynabeads M-280 with sheepanti-rabbit IgG

Virus capture was not influencedby the content of humic acidsin the samples

[86]

Rotavirus Seeded environmentalwater samples

Dynabeads protein A with boundantibodies against group A rotavirus

IMS combined with quantitativereverse transcription-PCR

[87]

Spiked water samples Dynabeads M-280, sheep anti-mouseIgG with bound murinemonoclonal antibodies

IMS combined with reversetranscription PCR

[88]

IMS immunomagnetic separation, PCR polymerase chain reaction


Not only free enzymes, but also enzymes immobilized onmagnetic carriers can be used for the detection of xenobioticsacting as their inhibitors. Acetylcholinesterase immobilizedon magnetic particles and integrated in a flow-injection

system via a magnetic reactor was used for amperometricdetermination of carbofuran, paraoxon, malaoxon andparaoxon-methyl [12]. Later, an ultrasensitive method fordetermination of pesticides in a glass lab-on-a-chip by means

Table 3 Examples of immunomagnetic separation of pathogenic microorganisms from water samples

Microorganism Water sample Magnetic system Comments Reference

Desulfovibriovulgaris

Subsurface water samples Streptavidin-coupled Dynabeadswith bound biotin-labelled antibodies

Samples taken frombioremediation areas

[89]

Escherichia coli Beach waters Magnetic particles (Bangs) withbound anti-E. coli antibodies

IMS and ATP bioluminescenceused for selective captureand quantification

[90]

Real and spikedwater samples

Dynabeads M-280 streptavidinwith bound biotin-labelled polyclonalrabbit antibody to E. coli

IMS coupled with quantumdot labelling

[91]

Untreated wastewatersamples from California,North Carolina, and Ohio

Magnetic particles with boundrabbit polyclonal anti-E. coli antibody

IMS/ATP method used [92]

Escherichia coliO157:H7

Wastewater from municipalsewage treatment plantand slaughterhouses

Dynabeads anti-E. coli O157 E. coli O157 is commonlypresent in animal andhuman wastewaters

[93]

Spiked sewage andriver waters

Dynabeads anti-E. coli O157 IMS coupled with selectivemedia, immunoassaysand biochemical tests

[94]

Spiked water samples Dynabeads anti-E. coli O157and Dynabeads streptavidinwith bound biotinylatedmonoclonal antibodyto O157 LPS

Immunomagneticelectrochemiluminescence

[95]

Drinking water fromprivate water suppliesin the Netherlands

Dynabeads anti-E. coli O157 E. coli O157:H7 was isolatedfrom 2.7 % of the samplesthat otherwise met thedrinking water standards

[96]

Helicobacterpylori

Spiked water fromthe Fyris river

Dynabeads M-280 withbound polyclonal rabbitantibody to H. pylori

Combination ofIMS and PCR

[97]

Legionellapneumophila

Spiked water samples andwater from potablehot water systems

Dynabeads M-280 tosylactivatedwith bound polyclonalantibody to L. pneumophila

Combination of IMSand real-time PCR

[98]

Spiked water samplesand water fromenvironmental sources

Dynabeads M-280 tosylactivatedwith bound monoclonalantibody to L. pneumophila

IMS–culture assay inenvironmental sampleswith high levels ofinterfering microflora

[99]

Spiked water samples Dynabeads M-280 precoatedwith sheep anti-mouse IgGand coated with mouseanti-L. pneumophila IgG3k

Combination of IMS and PCR [100]

Dynabeads MyOne streptavidinwith bound biotinylatedpolyclonal anti-Legionella antibody

IMS and detection by sandwichELISA and PCR amplificationof the ompS gene

[23]

Mycobacteriumavium subsp.paratuberculosis

Water from watertreatment works.Spiked water samples

Dynabeads M-280 withbound polyclonal rabbitantibody to M. aviumsubsp. paratuberculosis

4.7 % of samples testedpositive. Combination ofIMS with PCR studied

[101] [102]

Mycobacteriumulcerans

Spiked water samples Dynabeads coated withsheep anti-mouse IgG

Magnetic bead sequencecapture PCR

[103]

Dynabeads precoated withsheep anti-mouse IgG andcoated with mouse polyclonal IgG

Combination of IMS and PCR [104]

ATP adenosine triphosphate


of enzymatic inhibition of acetylcholinesterase immobi-lized on magnetic beads was developed. The reproduc-ible insertion of a controlled amount of enzyme-coupledmagnetic beads inside the chip channel and their immo-bilization in a capture region with the aid of a magnetic

field enables the easy renewal of the biosensing materialafter each determination in a highly reproducible man-ner. The detection of carbofuran (one of the most toxiccarbamate pesticides) has been achieved down to thenanomolar level [28].

Table 4 Examples of immunomagnetic separation of protozoan parasites from water samples

Parasite Water sample Magnetic system Comments Reference

Cryptosporidiumparvum

Spiked turbid water Dynabeads GC-Combo Nested PCR detection [105]

Karst water samples Dynabeads M-280streptavidin with boundbiotin-labelled antibodyto C. parvum

Comparison withimmunofluorescence assay

[106]

Spiked water samples Dynabeadanti-Cryptosporidium

Effects of pH and magneticmaterial on IMS

[107]

Spiked water samples Dynabeadanti-Cryptosporidium

High-quality DNA preparation [108]

Potomac river watershed Dybaneadsanti- Cryptosporidium

Separation for DNA extraction [109]

Wastewater concentrates Dynabeadanti-Cryptosporidium

Modified US EPA method 1622 [110]

Cryptosporidiumparvum andGiardia lamblia

Spiked water samples Dynabeads GC-Combo Fresh, aged, viable andheat inactivated oocystsdo not influence IMS

[111]

Evaluation of cyst lossin standard procedure

[112]

Cryptosporidiumparvum and Giardiaintestinalis

Spiked water samples Dynabeads GC-Combo Portable continuous flowcentrifugation as an alternativeconcentration method

[113]

Cryptosporidiumand Giardia

Filtered concentratedtap water, secondaryeffluent waterand purified water

Dynabeads GC-Combo Flow-through IMS system [114]

Wastewater samples fromthe effluents of secondarysedimentation and tertiarytreatment

Dynabeads GC-Combo Improvements of membranefiltration/elution and IMSsteps of modified USEPA method 1623

[115]

Raw and treated wastewaterand sludge samples

Dynabeads GC-Combo IMS followed byimmunofluorescentassay microscopy and PCR

[116]

Raw and treated wastewater,water from the Seineriver in Paris

Dynabeads GC-Combo Faecal bacterial indicators,enteroviruses and oocysts ofToxoplasma gondii also assessed

[117]

Water samples from 12wastewater treatment plants

Dynabeads GC-Combo PCR-RFLP analysis [118]

Enterocytozoonbieneusi

Spiked water samples Immunotech beads orDynabeads with boundspecific antibodies

PCR used for detection [119]

Giardia lamblia Spiked water samples Dynabeads GC-Combo Optimization of separationparameters

[120]

Dynabeads M-280streptavidin

Indirect IMS followedby PCR detection

[121]

Primary- and tertiary-treatedwastewater effluents

Anti-Giardia Dynabeads Infectivity of G. lamblia cystsassessed in Meriones unguiculatus

[122]

Real and spikedwater samples

Sheep anti-mouse IgGantibody-coatedmagnetite particles

Labelling of cysts with mouseIgG anti-Giardia antibody;82 % of seeded cysts recovered

[123]

Toxoplasmagondii

Unspiked and spikeddrinking and surface water

Goat anti-mouse IgM-coatedmagnetic beads withbound specific antibody

No positive environmentalsamples found

[124]

RFLP restriction fragment length polymorphism


A highly specific and sensitive novel electrochemicalimmunosensing platform for screening exposure to organo-phosphorus agents consists of magnetic beads with immo-bilized butyrylcholinesterase (BChE) and beads with anti-BChE antibody. The system is based on simultaneousimmunodetection of enzyme activity and immunoassay ofthe total amount of enzyme in the samples. The differencebetween the total amount of enzyme and amount of activeenzyme is determined as a phosphorylated enzyme adduct.This method can detect 2 % BChE inhibition. Human plas-ma was used as a matrix; however, this procedure could alsohave interesting applications in water analyses [29].

A phenol biosensor based on the immobilization oftyrosinase on the surface of modified magnetic MgFe2O4

nanoparticles was developed. The tyrosinase was first cova-lently immobilized on core–shell (MgFe2O4–SiO2) magneticnanoparticles, which were modified with an amino group onthe surface. The resulting magnetic bionanoparticles wereattached to the surface of a carbon paste electrode with thehelp of a permanent magnet. The immobilization matrix pro-vided a goodmicroenvironment for retention of the bioactivityof tyrosinase. Phenol was determined by the direct reductionof biocatalytically generated quinone species at −150 mVversus the saturated calomel electrode. The linear range fordetermination of phenol concentration was from 1×10-6 to2.5×10-4 M, with a detection limit of 6.0×10-7 M [30].Alternatively, a Fe3O4 nanoparticle–chitosan nanocompositewas used for entrapment of tyrosinase for the detection ofphenolic compounds such as catechol. This biosensor was arapid, simple and cost-effective way of analysis with a linearrange of 8.3×10−8–7.0×10−5 M [31].

Different magnetic particles can be used for immobiliza-tion enzymes, such as in the case of an amperometricscreen-printing biosensor for the detection of bisphenol Awhere tyrosinase immobilized on amino-functionalizednickel nanoparticles were compared with nickel

nanoparticles immobilized on iron oxide and gold nano-particles. The biosensor based on magnetic nickel nanopar-ticles had a response time of less than 30 s, a linear rangefrom 9.1×10-7 to 4.8×10-5 M and a detection limit of 7.1×10-9 M, was stable for more than 100 assays and hadcharacteristics comparable to or better than those of theother two types of biosensors [32]. In another example,tyrosinase was immobilized on streptavidin magnetic par-ticles via glutaraldehyde activation and transferred to thesurface of a carbon paste electrode; this biosensor wasdesigned for the screening of the inhibitory potency ofdifferent skin-whitening agents [33].

Enzymes inhibited by the presence of heavy metal ionshave also been tested after their immobilization on magneticparticles; the procedure is useful especially when colouredsamples or samples containing suspended solid impuritiesare to be assayed. Selected heavy metal ions such as Ag+

and Pb2+ inhibited the activity of trypsin immobilized onmagnetic nanoparticles in the form of ferrofluid. The per-centage of inhibition was simply evaluated from the differ-ence in the activities of the non-inhibited and inhibitedtrypsin samples using spectrophotometric measurementwith an artificial substrate [34].

In some cases the enzyme can be substituted by alterna-tive materials. Magnetic magnetite nanoparticles can mimicperoxidase activity and thus they have been used for thedetermination of hydrogen peroxide in rainwater, based ontheir catalytic effect on the oxidation of N,N-diethyl-p-phe-nylenediamine sulfate to a coloured product with a strongabsorption maximum at 550 nm [35]. Alternatively, haemo-globin can be used for the same purpose [36].

Examples of enzymes and relevant proteins immobilizedon magnetic carriers and used for determination of xeno-biotics in water are given in Table 5.

Magnetic techniques for preconcentration of xenobiotics

Analysis of both organic and inorganic xenobiotics (as wellas of biologically active compounds) in water systems oftenrequires preconcentration of the target analyte(s) from largevolumes of solutions and/or suspensions. This process isoften accompanied by partial purification of the analyte(s).The sample preparation is often the most time-consumingstep in chemical analysis, accounting on average for 61 % ofthe time typically required to perform the analytical tasks[37]. The sample preparation is also the source of much ofthe imprecision and inaccuracy of the overall analysis [38].

Presently, considerable attention is being paid to SPE as away to isolate and preconcentrate desired components froma sample matrix. SPE offers an excellent alternative to theconventional sample preparation methods, such as liquid–liquid extraction. The separation and preconcentration of an

Fig. 2 Electron microscope image of Legionella pneumophila boundto immunomagnetic beads—Dynabeads MyOne streptavidin (Invitro-gen) with bound biotinylated polyclonal anti-Legionella antibody.(Reproduced with permission from [23])


Tab

le5

Examples

ofenzymes

andrelatedproteins

immob

ilizedon

magnetic

carriers

andtheirapplicationin

water

analysis

Enzym

e/ECnu

mber

Xenob

iotics

Water

system

Magnetic

carrier/activ

ation(immob

ilizatio

n)Com

ments

References

Acetylcho

linesterase

/3.1.1.7

Carbo

furanand

malaoxo

n(pesticides)

Spikedwater

samples

Amino-term

inated

magnetic

particles

(BioMag

4100

)/glutaraldehy

deFlow

injectionanalysisof

pesticides

indrinking

water

[12,

125]

Dichlorvo

s(pesticide)

Water

samples

Au-do

pedmagnetic

Fe 3O4nano

particles

Electrochem

ical

sensor

forthe

detectionof

released

thiocholine

[126]

Carbo

furan(pesticide)

Spikedwater

samples

Dyn

abeads

M-270

epox

yLab-on-a-chip

assaywith

ultrasensitiv

ecarbofuran

detection

[28]

Acetylcho

linesterase

(genetically

engineered)

Chlorpy

riph

os-oxo

nand

chlorfenvinp

hos(pesticides)

Spikedwater

samples

Metal-chelate-fun

ctionalized

magnetic

microbeads/affinity

interaction

with

6-Histail

Amperometricmeasurements

ofreleased

thiocholine

[127]

Perox

idase/1.11.1.7

Hyd

rogenperoxide

Spikedsamples,

disinfectants

Magnetite–chito

sanmicrospheres

Amperometricbiosensorbasedon

amod

ifiedglassy

carbon

electrod

e[128]

Trypsin/3.4.21.4

Ag+,Pb2

+,4-am

inob

enzamidine,

bacitracin,safranin,thionin

Water

solutio

nsMagnetiteparticles(ferrofluid)/

water

solublecarbod

iimide

Spectroph

otom

etricdeterm

ination

ofdecrease

ofim

mob

ilized

tryp

sinactiv

ity

[34]

Tyrosinase/1.14

.18.1

Skin-whitening

agents

(tyrosinaseinhibitors—

kojic,

benzoicandazelaicacids)

Mod

elsolutio

nsStreptavidinMasterbeads

(magnetic

particles50

0nm

),Ademtech,

France/glutaraldehy

de

Carbo

npasteelectrod

e,am

perometricassay

[33]

Pheno

lIndu

strial

wastewater

Core–shell(M

gFe 2O4–SiO

2)magnetic

nano

particlessurfacemod

ifiedwith

aminegrou

ps/glutaraldehyd

e

Carbo

npasteelectrod

e,cyclic

voltammetricand

amperometricmetho

d

[30]

Pheno

liccompo

unds

Mod

elsolutio

nsFe 3O4nano

particle–chito

san

nano

compo

site/entrapm

ent

Glassycarbon

electrod

e,am

perometricdetection

[31]

Bisph

enol

AAqu

eous

solutio

nsAmino-functio

nalized

magnetic

Nior

Fe 3O4nano

particles,Aunano

particles/

glutaraldehy

de

Amperometricbiosensor,

screen-printed

graphite

electrod

e[32]

Haemog

lobin

Hyd

rogenperoxide

Spikedsamples,

disinfectants

Magnetite–chito

sanmicrospheres

Amperometricbiosensor,

glassy

carbon

electrod

e[36]


analyte from large volumes of solution can take a lot of timeusing a standard column SPE. That is why a new procedurefor SPE based on the use of magnetic or magnetizableadsorbents called magnetic solid-phase extraction (MSPE)has been developed recently [13]. Magnetic particles havealso been implemented in other extraction and preconcen-tration procedures [39]. Other procedures, such as SBSE[14], rotating-disk sorptive extraction [40] and stir-rodsorptive extraction [151], employ a magnetic stir bar orother mixing element covered with an adsorbent layer.Stir membrane extraction uses an iron wire to enablemagnetic stirring of the membrane during the extractionprocess [152].

Magnetic solid-phase extraction

In this procedure, a magnetic adsorbent (either a generalone, such as magnetic charcoal, or a magnetic affinityadsorbent) is added to a solution or suspension containingthe target analyte(s). The analyte is adsorbed on to themagnetic adsorbent and then the whole complex is recov-ered from the suspension using an appropriate magneticseparator. The analyte is consequently eluted from therecovered adsorbent and analysed [13] (see Fig. 3).

Different types of magnetic adsorbents, such as magneticcharcoal [13, 41], reactive copper phthalocyanine dyeimmobilized on silanized magnetite particles (blue magne-tite) [13, 42] and magnetic derivatives of Chromosorb,Tenax TA, Tenax GR, Porapak, Chezacarb, polyamide andpolyphenyleneoxide [41, 43, 44], have been used for thepreconcentration of various organic xenobiotics, includingdyes, tensides, nonylphenol derivatives and alkylphenols,from water systems.

MSPE has been efficiently used for the detection ofcompounds illegally used as fish drugs in aquacultures.Recently, malachite green (basic green 4, CI 42000) hasfound extensive use all over the world in the fish farmingindustry as a fungicide, ectoparasiticide and disinfectant; a

similar situation is valid for crystal violet (basic violet 3, CI42555). Both dyes are linked to an increased risk of cancerand that is why the use of malachite green has been bannedin aquacultures intended to produce fish for human con-sumption [42]. Malachite green and crystal violet in thechromatic form specifically interact with copper phthalocy-anine dye immobilized on magnetic particles [45]. Thisfeature enables specific separation and concentration of thetarget dyes (and other specifically binding compounds, suchas polyaromatic hydrocarbons) not only from solutions, butalso from suspension systems. After MSPE, the target dyesare eluted with a small amount of methanol and the spectraare recorded. Concentrations of both dyes in the range 0.5–1.0 μg L-1 water could be reproducibly detected. The dyescan be detected not only in potable water, but also in riverwater [42].

Recently, new progressive materials such as graphenehave been suggested for MSPE. Two-dimensional planargraphene sheets were immobilized onto silica-coated mag-netic microspheres by simple adsorption. The graphenesheets were used as adsorbent material to extract six sulfon-amide antibiotics from water samples. Under the optimalconditions, a rapid and effective determination of targetcompounds in environmental water samples was achieved;the limits of detection ranged from 0.09 to 0.16 ng mL-1

[46].Silica-coated magnetic nanoparticles modified with γ-

mercaptopropyltrimethoxysilane were used for fast and selec-tive SPE of trace amounts of cadmium, copper, mercury andlead ions in environmental and biological samples (includingseawater) prior to their determination by inductively coupledplasma MS [47]. In other approaches, silica-coated magnetitenanoparticles were functionalized with salicylic acid and usedfor SPE and preconcentration of copper, cadmium, nickel andchromium ions from water and food samples [48], and mes-oporous silica–magnetite microspheres with immobilized imi-nodiacetic acid were used for preconcentration of cadmium,manganese and lead ions [49].

Fig. 3 Magnetic solid-phase extraction procedure. (Reproduced with permission from [150])


Tab

le6

Examples

ofxeno

bioticsprecon

centratedby

magnetic

solid

-phase

extractio

n

Xenob

iotics

Characteristics

Water

system

Magnetic

system

Com

ments

Reference

Crystal

violet,safranin

ODyes

Spikedwater

samples

Reactivecopp

erph

thalocyanine

dye

attached

tosilanizedmagnetite

240-fold

enrichmentof

crystalviolet

from

800mLof

sample

[13]

Crystal

violet,safranin

ODyes

Spikedwater

samples

Magnetic

charcoal

460-fold

enrichmentof

crystalviolet

from

800mLof

sample

[13]

Malachite

green,

crystalviolet

Dyes

Spikedwater

samples

Magnetitewith

immob

ilized

copp

erph

thalocyanine

dye

0.5-1.0μg

ofdy

ein

1Lof

water

detected

[42]

Non

ionicsurfactants

Surfactants

Spikedwater

samples

Magnetic

derivativ

esof

charcoal,

poly(oxy

-2,6-dim

ethy

l-1,4-ph

enylene)

andpo

lyam

ide

Hyd

roph

obic

adsorbentsexhibitedthe

bestextractio

ncharacteristics

[41]

Alkylph

enolsand

oxyethylated

alky

lpheno

lsSurfactantsprecursors,

plasticizers

Spikedwater

samples,

riverandlake

samples

Magnetic

derivativ

esof

Chrom

osorb,

Tenax,Porapak,Chezacarb

and

polyph

enyleneoxide

Detectio

nlim

itswerebetween

0.7and1μg

L-1.

[43,

44,12

9]

Polyaromatic

hydrocarbo

nsHyd

rocarbon

sSpikedwater

samples

n-Octadecylph

osph

onic

acid

mod

ified

mesop

orou

smagnetic

nano

particles

Adsorbent

with

extrem

ely

hydrop

hobicprop

erties

[130]

Sulfonamides

Antim

icrobial

drug

Spikedandenvironm

ental

water

samples

Octadecyltrim

ethy

lammon

ium

brom

ide

adsorbed

onto

magnetitenano

particles

Aconcentrationfactor

of1,00

0was

achieved

(extractionof

500mLof

water

samples)

[131]

UV

filters

from

cosm

etic

prod

ucts

Organic

compo

unds

containing

benzenering

Water

samples

ofdifferent

origin

(tap,riverandsea).

Oleic

acid

coated

magnetic

nano

particles

GC-M

Sdeterm

ination

[132]

Polyaromatic

hydrocarbo

nsHyd

rocarbon

sLakewater

Carbo

n–ferrom

agnetic

nano

compo

site

GC-M

Sdeterm

ination

[133]

PAHs,ph

thalateesters

Organic

compo

unds

Sno

w,tapandriverwater

Barium

alginate

cagedFe 3O4–C-18

magnetic

nano

particles

HPLCassaywith

fluo

rescence

detection

[134]

Diethylstilb

estrol,

oestrone,oestriol

Estrogens

Tap,mineral

and

Pearlriverwater

Magnetic

silicaparticlescoated

with

hydrox

y-term

inated

multiw

alled

carbon

nano

tubes

The

extractio

nefficiencies

were

95.9,93

.9and52

.4%,respectiv

ely

[135]

Sarin,soman,tabu

n,cyclosarin

Nerve

agents

Spikedwater

samples

Magnetic

multiw

alledcarbon

nano

tubes

Elutio

nwith

chloroform

,recoveries

60–96

%[136]

Methy

lmercury

Organom

etallic

derivativ

eSeawater

Fe 3O4/polyanilin

enano

particles

GC-M

Sdeterm

ination

[137]

Cd,

Cu,

Hgand

Pbions

Heavy

metal

ions

Spikedandreal

water

samples

Silica-coatedmagnetic

nano

particlesmod

ified

with

γ-mercaptop

ropy

ltrim

etho

xysilane

Indu

ctivelycoup

ledplasmaMS

used

forthestud

y[47]

Ag,

Cd,

Cuand

Znions

Heavy

metal

ions

Tap

andmineral

water

samples

Magnetic

nano

particlescoated

with

3-(trimetho

xysilyl)-1-propanetio

land

mod

ifiedwith

2-am

ino-5-mercapto-

1,3,4-thiadiazole

Indu

ctivelycoup

ledplasmaop

tical

emission

spectroscopy

determ

ination

[138]

Fluoride

Anion

Spikedwater

samples

Magnetic

iron

oxides

nano

particles

Spectroph

otom

etricdeterm

ination

[139]

HPLChigh

-perform

ance

liquidchromatog

raph

y


Inspiration can also be taken from food analysis, whereMSPE using a magnetic, phenyl-functionalized silica adsor-bent has been used for the determination of tetracyclines inmilk samples [50]; other magnetic adsorbents—e.g. magne-tite/silica/poly(methacrylic acid-co-ethylene glycol dimetha-crylate) composite microspheres and magnetic carbonnanotubes—have been used for sulfonamide preconcentra-tion [51] and oestrogen determination [52] from the samematrix.

Typical examples of MSPE for water analysis are shownin Table 6.

Incorporation of magnetic particles in other extractionprocedures

A new two-step microextraction technique, combining dis-persive liquid–liquid microextraction (DLLME) and disper-sive solid-phase microextraction (SPME), was developedfor the fast GC-MS determination of polycyclic aromatichydrocarbons in environmental samples. Any organic sol-vent immiscible with water can be used as an extractant inDLLME. In the approach presented, hydrophobic magneticnanoparticles were used to retrieve the extractant (1-octanol)together with the analyte. Because of the rapid mass transferassociated with the DLLME and the dispersive SPME steps,fast extraction could be achieved. The desorption of analyteswas performed by sonication with acetonitrile as the solvent.

Enrichment factors ranging from 110- to 186-fold wereobtained for the analytes. This two-step extraction methodwas successfully used for the fast determination of polycy-clic aromatic hydrocarbons in river water samples [39]. Thescheme of the procedure is shown in Fig. 4.

Stir-bar sorptive extraction and related procedures

SBSE is a new solventless sample preparation method forthe extraction and enrichment of organic compounds fromaqueous matrices. The method is based on the same princi-ples as SPME; a magnetic stir bar covered with an appro-priate adsorbent is used as the adsorbing element. Comparedwith SPME, a relatively large amount of extracting phase iscoated on a magnetic stir bar. Solutes are extracted into thecoating, based upon their octanol–water partitioning coeffi-cient and upon the sample–extraction medium phase ratio.The technique has been applied successfully to trace analy-sis in environmental, biomedical and food applications.Extremely low detection limits can be obtained [14].

Polydimethylsiloxane-coated stir bars are currently avail-able commercially (Gerstel, Müllheim an der Ruhr, Ger-many). These stir bars have three essential parts (Fig. 5).The first one is a magnetic stirring rod, which is necessaryfor transferring the rotating movement of a stirring plate tothe sample liquid. The second part of the stir bar is a thinglass jacket that covers the magnetic stirring rod. The third

Fig. 4 Dispersive liquid–liquidmicroextraction (DLLME)coupled with dispersivesolid-phase microextraction(D-μ-SPE). (Reproducedwith permission from [39])

Fig. 5 Magnetic stir bar covered with polydimethylsiloxane (PDMS)


and outermost part is the layer of polydimethylsiloxanesorbent into which the analytes are extracted. The extractionprocedure is quite simple. SBSE of a liquid sample isperformed by placing a suitable amount of sample in aheadspace vial or other container. A polydimethylsiloxane-coated stir bar is added and the sample is stirred for 30–240 min. The extraction time is controlled kinetically and isdetermined by the sample volume, stirring speed and stir bardimensions; it has to be optimized for a given application.Optimization is normally accomplished by measuring theanalyte recovery as a function of the extraction time [14].

After extraction, the stir bar is removed, dipped on aclean paper tissue to remove water droplets and introducedto the thermal desorption unit, connected to a gas chromato-graph. In some cases, rinsing the stir bar slightly withdistilled water to remove adsorbed sugars, proteins or othersample components is recommended; this step will preventthe formation of non-volatile material during the thermal-desorption step. Rinsing does not cause solute loss, becausethe sorbed solutes are present in the polydimethylsiloxane

phase. Finally, the solutes are thermally desorbed. The de-sorption temperatures are application-dependent, primarilydetermined by the volatility of the solutes, and are typicallybetween 150 and 300 °C. Desorption can be accomplishedin 5–15 min under a 10–50 mL min-1 helium flow. As analternative to thermal desorption, organic solvents can beused for desorption [14].

Preconcentration of numerous analytes has been de-scribed in the literature; typical examples connected withwater analysis are shown in Table 7. SBSE also enables thedevelopment of multiresidue methods for simultaneous pre-concentration and subsequent determination of several doz-en of xenobiotics [53]. Recently, a new ethylene glycol/silicone twister, based on polydimethylsiloxane/ethyleneglycol copolymer, has become available from Gerstel.

An alternative procedure for the preconcentration of tar-get analytes, called rotating-disk sorptive extraction, isbased on the extraction onto a small rotating disk made ofTeflon containing a sorbent phase of polydimethylsiloxaneon one of its surfaces. In this way, the disk, which has a high

Table 7 Examples of xenobiotics extracted by the stir-bar sorptive extraction procedure (using magnetic stir bars coated with a polydimethylsi-loxane layer) from water samples

Xenobiotics Characteristics Water system Comments Reference

Bayrepel Insect repellent Spiked and real water samples GC-MS detection [140]

Benzylbutylphthalate Endocrine disruptor Real water samples GC-MS detection [141]

Bis(2-ethylhexyl) adipate Endocrine disruptor Real water samples GC-MS detection [141]

Buprofezin Insecticide Spiked water samples LC-MS/MS detection [142]

Chlorfenvinphos Insecticide Spiked water samples LC-MS/MS detection [142]

Chlorpyriphos-ethyl Insecticide Spiked water samples LC-MS/MS detection [142]

Diazinon Insecticide Spiked and real water samples LC-MS/MS detection [142]

Fenthion Insecticide, avicide, acaricide Spiked water samples LC-MS/MS detection [142]

Hexythiazox Acaricide Spiked water samples LC-MS/MS detection [142]

Imazalil Fungicide Spiked and real water samples LC-MS/MS detection [142]

4-tert-Octylphenol Endocrine disruptor Real water samples GC-MS detection [141]

Prochloraz Fungicide Spiked water samples LC-MS/MS detection [142]

Pyrethroids Insecticides Spiked water samples Capillary GC-MS detection [143]

Tolclofos-methyl Bactericide Spiked water samples LC-MS/MS detection [142]

Sex steroid hormones,phthalates, alkylphenols, tamoxifen

Endocrine disruptingcompounds

Real water samples GC-MS detection [144]

Acetylsalicylic acid, ibuprofen,diclofenac sodium, naproxen,mefenamic acid, gemfibrozil

Drugs Spiked water samples HPLC-DAD [145]

11 PBDEs Endocrine disruptingcompounds

Spiked water samples Injection and capillaryGG-MS detection

[146]

Permethrin, icaridin, DMP,DEET, butopyronoxyl, R-326,MGK-264, piperonyl butoxide

Insect repellents Real water samples ThermodesorptionGC-MS detection

[147]

46 acidic and polarorganic pollutants

Phenols, acidic herbicides,pharmaceuticals and others

Real water samples GC-MS detection [148]

15 PAHs Hydrocarbons Real water samples Column LC andfluorescence detection

[149]

DMP dimethyl phthalate, DEET N,N-diethyl-m-toluamide


surface area, contacts only the liquid sample, which can bestirred at higher velocity than with the stir bar used in SBSE,without damaging the phase while at the same time facili-tating analyte mass transfer to the polydimethylsiloxanesurface. With increasing rotational velocity, the amount ofextracted analyte significantly increases because the stag-nant layer concomitantly decreases. On the other hand, theextracted amount concomitantly increases with extractiontime, reaching equilibrium at approximately 20 min. Theprocedure was used for the determination of nonylphenol inreal water samples after elution of the adsorbed analyte withmethanol [40]. In the case of determination of colouredcompounds (e.g. malachite green), another approach hasbeen used. After extraction, the sorbent phase with theconcentrated analyte was separated from the Teflon diskand used directly for malachite green determination bysolid-phase spectrophotometry at 624 nm, without thenecessity of a desorption step [54]. The scheme of the diskand the separation process is shown in Fig. 6.

Stir-rod sorptive extraction was proposed as a new pro-cedure to avoid the friction loss of adsorbent coatings duringthe stirring processes. As shown in Fig. 7, a monolithic-polymer-coated stir rod is used. The polymer has octadecylgroups (hydrophobic) and sulfonic acid groups (ionexchange). A small magnet enables efficient movement ofthe stir rod in an analysed solution using a magnetic stirrer.This system was efficiently used for the separation of fluo-roquinolones [151]. The preconcentration of polycyclicaromatic hydrocarbons from water samples was performed

using poly(ethylene glycol dimethacrylate)/graphene com-posite as a stir bar coating [153].

Because extraction capabilities depend on the extractantgeometry, polymeric membranes are promising materialsthanks to their high sample/adsorbent contact surface. Anovel approach which combines the extraction capabilitiesof polymeric membranes with the advantages of their mag-netic stirring in the analysed solution has been developed;this system was used for monitoring of polycyclic aromatichydrocarbons in water samples [152].

Conclusions

The need for rapid, cost-effective and high-throughput ana-lytical methods for environmental monitoring is increasing.Magnetic nanoparticles and microparticles are very versa-tile, and the examples presented clearly document that mag-netically responsive materials can significantly simplify andimprove determination and detection of both organic andinorganic xenobiotics, nucleic acids and cellular pathogens.The sensitivity and selectivity of these methods depend onthe appropriate functionalization of magnetic materials andmolecules immobilized on their surfaces. In many cases,magnetic techniques can be performed more rapidly andwith lower consumption of chemicals than standard analyt-ical procedures; in addition, magnetic techniques are usuallymore environmentally friendly. Methods using magneticmaterials can be easily integrated into existing analyticaland microbiology laboratory protocols because only a fewadditional items, such as a magnetic separation rack andcommercially available kits, are needed. Water researchand technology can benefit greatly from the application ofsmart magnetic materials and the corresponding analyticalprocedures in laboratory practice.

Fig. 7 Stir-rod sorptive extraction. (Reproduced with permission,from[151])

Fig. 6 The rotating disk and the rotating-disk sorptive extractionprocedure. (Reproduced with permission from [40])


Acknowledgment This research was supported by the Grant Agencyof the Czech Republic (project no. P503/11/2263).

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