Journal of Saudi Chemical Society (2016) 20, S624–S631

King Saud University

Journal of Saudi Chemical Society

www.ksu.edu.sawww.sciencedirect.com

ORIGINAL ARTICLE

A novel hydrogel based microextraction of analytes

* Corresponding author. Tel.: +98 441 2972143; fax: +98 441

2776707.

E-mail address: m.bahram@urmia.ac.ir (M. Bahram).

Peer review under responsibility of King Saud University.

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http://dx.doi.org/10.1016/j.jscs.2013.05.002

1319-6103 ª 2013 Production and hosting by Elsevier B.V. on behalf of King Saud University.

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Morteza Bahram *, Foroogh Keshvari, Naimeh Mohseni

Department of Chemistry, Faculty of Science, Urmia University, Urmia, Iran

Received 22 November 2012; accepted 6 May 2013Available online 15 May 2013

KEYWORDS

Dispersive extraction;

Malachite green;

Crystal violet;

pH-Sensitive hydrogel

Abstract We report on a new method for the preconcentration and microextraction of analytes

from aqueous samples using a pH-sensitive hydrogel. It is referred to as semisolid–liquid dispersive

microextraction (SSLDM) and has the advantages of both dispersive liquid–liquid microextraction

(DLLME) and solid-phase extraction (SPE). The surface area of the droplets is large, so the equi-

librium state is quickly reached. The high density of the gel facilitates phase separation, and disad-

vantages of existing methods are overcome in that there is no need for extraction and dispersive

solvents (as in conventional DLLME) and no need for desorption (as in SPE). The SSLDMmethod

integrates sampling, extraction and concentration into one single and solvent-free step. The method

was exemplarily applied to the preconcentration of malachite green (MG) and crystal violet (CV)

which then were quantified by spectrophotometry. Specifically, poly (styrene-alt-maleic acid), a

pH-sensitive hydrogel, was used to extract MG and CV from water samples. The linear analytical

range is from 0.01 to 1 lmol L�1 for MG, and from 0.05 to 1 lmol L�1 for CV. The correlation

coefficient for MG and CV is 0.999 and 0.993, respectively. The limit of detection for MG and

CV is 0.011 and 0.014 lmol L�1, respectively.ª 2013 Production and hosting by Elsevier B.V. on behalf of King Saud University. This is an open access

article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

1. Introduction

Sample pre-treatment is often the substantial and rate deter-mining step in a measurement process. One of the classical

sample preparation methods is liquid–liquid extraction(LLE). This technique requires extensive amounts of hazard-ous solvent which results in the generation of large amounts

of pollutants (Barcelo et al., 1990). Although, solid-phase

microextraction (SPME) is a solvent-free sample preparationtechnique, reduced volumes (few microliters) of solvents areused only if liquid desorption is carried out (Prosen and

Zupancic-Kralj, 1999). Endeavors in the exploration of novelgreen and miniaturized sample pretreatment techniques haveled to the development of various microextractions methods

namely single drop microextraction (SDME), solid-phase mic-roextraction (SPME), hollow fiber liquid phase microextrac-tion (HF-LPME), dispersive liquid–liquid microextraction(DLLME) and solidified floating organic drop microextraction

(SFODME).In SDME a drop of immiscible extracting solvent (about 1–

10 lL) is suspended from a syringe into the liquid or gaseous

sample medium. After extracting of analyte(s) of intrest for aperiod of time, the organic drop is retracted back into themicrosyringe and is injected to a suitable detector for

(a)

(b)

(c)

Figure 1 Structure of alt-PSMA (a), malachite green (b) and

crystal violet (c).

A novel hydrogel based microextraction of analytes S625

quantification of analytes. During extraction, the target ana-lytes are extracted from aqueous sample into the hanging dropbased on passive diffusion, and extraction recoveries are essen-

tially determined by the organic solvent to water partitioncoefficients. Direct immersion (DI)-SDME, headspace (HS)-SDME, three phases SDME and continuous flow microextrac-

tion are common modes of SDME (Jeannot and Cantwell,1996; Dadfarnia and Haji Shabani, 2010). In the HF-LPMEmethod the extracting phase is placed inside the lumen of a

porous hydrophobic hollow fiber which forms a supported li-quid membrane (SLM). The extraction is done in two-phaseand three-phase sampling modes. In the two-phase samplingmode, the analytes are extracted from an aqueous sample into

the organic solvent immobilized in the pores of the hollow fi-ber. In the three-phase sampling mode, the analytes are ex-tracted from an aqueous sample into the organic solvent

immobilized as a supported liquid membrane, and then intothe acceptor solution placed inside the lumen of the hollow fi-ber (Pedersen-Bjergaard and Rasmussen, 1999). In the

SFODME method a droplet of an immiscible solvent withmelting point near room temperature (10–30 �C) is floatedon the surface of the agitated aqueous sample in order to max-

imize the contact between the two solutions. The sample vial isthen placed in an ice bath to solidify the droplet which is easilyremoved and is allowed to melt for determination (KhaliliZanjani et al., 2007).

In 2006, Assadi and co-workers (Rezaee et al., 2006) intro-duced dispersive liquid–liquid microextraction (DLLME)which is based on a ternary component solvent system. In this

microextraction method, the appropriate mixture of extractingsolvent and disperser solvent is rapidly injected into the aque-ous sample containing the analytes of interest. Thereby, a clou-

dy solution is formed and due to the large surface area betweenextraction solvent and aqueous sample, the extraction is al-most independent of time. Simplicity of operation, rapidity,

low cost, high recovery and enrichment factors are the mainadvantages of DLLME. But this technique requires relativelylarge amounts of the disperser solvent, at most times, recover-ies decrease proportionately for less hydrophobic species and

lower extraction efficiencies will be resulted (Gharehbaghiet al., 2009; Molaakbari et al., 2011), applying a higher densityof extraction solvent creates some problems with some instru-

ments such as ICP-OES and reverse-phase HPLC (Rezaeeet al., 2010), and it is not a selective extraction technique (Fon-tana et al., 2010). Dispersive solid-phase extraction (DSPE),

often referred to as the ‘‘QuEChERS’’ (Quick, Easy, Cheap,Effective, Rugged, and Safe) method, was first introduced by(Anastassiades et al. (2003) and has become increasingly pop-ular for multi-residue analysis of multiclass pesticides in a wide

variety of agricultural products (Wang et al., 2007; Lambro-poulou and Albanis, 2007; Guo et al., 2010). As a clean-upstep, the crude extract is cleaned up by the addition of a small

amount of SPE sorbent material to an aliquot of the extract toremove the matrix co-extractives. The clean-up is easily carriedout by just shaking and centrifugation (Regueiro et al., 2011;

Dıez et al., 2006; Wu et al., 2009). The main disadvantage ofthe QuEChERS is its lower extracted concentration (nearly2–5 times) compared to other most traditional methods (Leho-

tay et al., 2005).In 1992, Igarashi and Yotsuyanagi (Igarashi and Yottuyan-

agi, 1992; Ghiasvand et al., 2005; Igarashi et al., 2000), re-ported an alternative homogeneous liquid–liquid extraction

method based on pH-dependent phase separation. Neutraliza-tion of perfluorooctanoate ion (PFOA–) in water-miscible or-ganic solvent solution (such as acetone, 1,4-dioxane and

tetrahydrofuran) induces phase separation, providing a smallvolume (down to 0.1 mL, depending on the amount ofPFOA� which has been added) of an oily and transparent

water-immiscible liquid phase, from a large volume of theaqueous solution. This phase separation was reversible withthe pH change in the solution, corresponding to below and

above the pKa value of perfluorooctanoic acid (HPFOA).The above method has been applied for organic and inorganicanalytes. Our proposed method bears a strong resemblance tothis work.

The aim of this work was to develop a simple, efficient andlow-cost procedure, which involves the semisolid–liquid dis-persive microextraction instead of classical dispersive liquid–li-

quid and dispersive solid–liquid microextraction methods. ApH-sensitive hydrogel was employed which consists of Poly-styrene-alt-maleic acid (Fig. 1a) for the preconcentration and

determination of malachite green (Fig. 1b) and crystal violet(Fig. 1c) in water samples. Malachite green (MG) and crystalviolet (CV) are triphenylmethane dyes. Malachite green acts

as a liver tumor-enhancing agent (Rao, 1995). It has been re-ported to cause carcinogenesis, mutagenesis, chromosomalfractures, teratogenicity and respiratory toxicity (Srivastavaet al., 2004). Since today some techniques for the determina-

tion of trace amounts of MG have been reported (Pourrezaand Elhami, 2007; Afkhami et al., 2010; Liu et al., 2011; Bah-ram et al., 2011; Tao et al., 2011). Some of these methods are

laborious and require sophisticated instruments like LC–MS(Tao et al., 2011; Dowling et al., 2007; Valle et al., 2005).CV is effective in the treatment of fungal infection and is uti-

lized in coloring paper, dyeing cottons and woods. At present,the use of CV in aquaculture has raised serious concerns be-

S626 M. Bahram et al.

cause it has been reported that CV may cause human carcino-genesis and mutagenesis (Littlefield et al., 1985). It has beenbanned for use in many countries. Several methods have been

proposed for determining and removing CV (Saeed et al.,2010; Jana et al., 2010; Madrakian et al., 2011; Nezamzadeh-Ejhieh and Banan, 2012). In most cases, these methods require

complicated pretreatment procedures.Recently we presented a new preconcentration method

using pH-sensitive hydrogel as a cloud point extraction meth-

od (Bahram et al., 2011). In this work a novel method usingpH-sensitive hydrogel is represented entitled ‘‘Hydrogel basednovel semi solid–liquid dispersive microextraction’’. The earlypresented work (Bahram et al., 2011) is classified as a cloud

point extraction method while this work is a new version ofdispersive extraction methods. In the presented method hydro-gel (HG) was dissolved in water and hydrogel solution was ob-

tained. The extractant phase (dilute hydrogel) was rapidlyinjected into the aqueous sample by a syringe. Because thepH of the aqueous sample prior to injection has been fixed

on the pH lower than that required by the hydrogel to beclouded, instantly, a cloudy solution was formed in the testtube. By centrifugation, the hydrogel-rich phase (which con-

tains the analyte of interest) is separated and detected by asuitable detector. The newly introduced procedure has impor-tant advantages over conventional extraction techniques, sinceit is fast, easy to operate, efficient and avoids using highly toxic

chlorinated solvents. The effect of experimental parameterssuch as extraction time, concentrations of HCl and HG onthe extraction efficiency were investigated and optimized.

2. Materials and methods

2.1. Reagents and solutions

All chemicals used in the experiments were of analytical grade

and used without further purification. All solutions were pre-pared with distilled water. HCl, acetonitrile and malachitegreen, crystal violet, styrene, maleic anhydride, benzoyl perox-

ide, tetrahydrofuran, diethyl ether, and NaOH were fromMerck (Darmstadt, Germany, www.merck.de). Hydrogel(0.4 g) was dissolved in 100 mL distilled water to obtain a con-centration of (0.4% w/v).

Poly (styrene-alt-maleic acid) (Fig. 1a) as an alternatingcopolymer (alt-PSMA) is a readily synthesized copolymer ofstyrene and maleic acid that incorporates two carboxylic

Table 1 The variables and values used for central composite design

Coded factor levels for MG

Variable name �1.428 (low) �F1 HCl (mol L�1) 0.017

F2 HG (%w/v) 0.038

F3 Time (min) 3

Coded factor levels for CV

Variable name �1.607(low) �F1 HCl (mol L�1) 0.0016

F2 HG (%w/v) 0.0048

F3 NaNO3 (mol L�1) 0.022

F4 Time (min) 1.8

groups and a phenyl group in each repeating unit. The molec-ular weight and polydispersity of the PSMA copolymers canbe controlled by varying the molar ratio of free-radical initia-

tor and volume of the added solvent. In alt-PSMA (120,000average MW), the hydrophobic phenyl side group, contributedby the styrene unit, is directly attached to the hydrocarbon

backbone. This compound derives its anionic charge fromtwo free carboxyl groups (pKa 1.9 and 6.0) of maleic acid,which are also directly attached to the hydrocarbon backbone

instead of the aromatic ring (Pirrone et al., 2010) and this com-pound is commercially available (Sigma–Aldrich, Product no.662631, www.sigmaaldrich.com).

2.2. Preparation of poly (styrene-alt-maleic acid)

Firstly poly (styrene-alt-maleic anhydride) (PSMA) was pre-pared through a thermally initiated free-radical polymerization

of styrene and maleic anhydride according to the literaturemethod (Henry et al., 2006).

2.2.1. Instruments

The spectrophotometric measurements were done with a PGmode T80 UV–Vis double-beam spectrophotometer (Japan)utilizing a 1-cm especial quartz cell (volume 0.5 mL). For

acceleration phase separation the Urum-Tadjihiz centrifuge(Urmia, Iran) was used.

2.2.2. Statistical software

Essential Regression and Experimental Design for chemistsand Engineers (EREGRESS), as a MS Excel Add-in software(DD, 1998–2001; Bulacov et al., 2006) was used to design the

experiments and to model and analyze the results.

2.2.3. Extraction procedure for malachite green

10 mL of water sample containing appropriate amount of MG

was placed in a 15 mL falcon tube and 0.75 mL HCl (0.5 mol L�1) was added and rapidly 2 mL of hydrogel dissolved in water(0.4% w/v) injected into the aqueous sample by a syringe. Sep-

aration of two phases was achieved by centrifugation for 3 minat 1800 rpm. The aqueous phase was easily decanted by simplyinverting the tube. The sedimented phase was diluted with

0.5 mL of acetonitrile and transferred into a 1 mL quartz cellto measure the absorbance of the solution at 617 nm againsta reagent blank. It should be noted that, instead of acetonitrilefor dilution of final extract, a moderate alkaline solution could

(CCD) for malachite green and crystal violet.

0.849 0 0.849 +1.428 (high)

0.024 0.035 0.046 0.053

0.052 0.072 0.092 1.06

5 8 11 13

1 0 1 +1.607(high)

0.004 0.008 0.012 0.014

0.012 0.024 0.036 0.044

0.020 0.050 0.080 0.098

3 5 7 8.2

Table 2 List of experiments in the CCD (coded values) and the response of each run for MG and CV.

Design points Factors for crystal violet Response

F1 F2 F3 F4

1 �1 1 �1 1 0.358

2 0 0 1.607 0 0.814

3 1 1 �1 1 0.702

4 0 �1.607 0 0 1.22

5 1 1 1 �1 0.492

6 �1 1 1 �1 0.408

7 0 0 0 �1.607 0.622

8(cp) 0 0 0 0 0.686

9 1.607 0 0 0 0.442

10 1 1 1 1 0.68

11 0 1.607 0 0 0.446

12 �1 1 1 1 0.45

13(cp) 0 0 0 0 0.446

14 1 �1 �1 �1 0.778

15 �1 �1 �1 1 0.654

16 �1 �1 �1 �1 0.632

17 1 �1 1 1 0.75

18 1 �1 �1 1 1.04

19 �1 1 �1 �1 0.192

20(cp) 0 0 0 0 0.346

21 0 0 0 1.607 0.924

22 �1 �1 1 �1 0.724

23 0 0 �1.607 0 0.734

24(cp) 0 0 0 0 0.826

25 �1 �1 1 1 1.072

26 1 �1 1 �1 0.782

27 �1.607 0 0 0 0

28 1 1 �1 �1 0.366

Design points Factors for malachite green Response

F1 F2 F3

1 0.849 �0.849 �0.849 0.851

2 �0.849 �0.849 �0.849 0.776

3 0.849 0.849 �0.849 0.924

4 �0.849 0.849 0.849 0.81

5 1.428 0 0 0.751

6 0 0 1.428 0.957

7 0 0 �1.428 0.981

8 0 �1.428 0 0.801

9 0.849 �0.849 0.849 0.838

10(cp) 0 0 0 0.927

11(cp) 0 0 0 0.925

12 �0.849 �0.849 0.849 0.74

13(cp) 0 0 0 0.95

14 0.849 0.849 0.849 0.94

15 �0.849 0.849 �0.849 0.898

16(cp) 0 0 0 0.904

17 0 1.428 0 1.12

18 �1.428 0 0 0.513

cp: Indicates central points.

A novel hydrogel based microextraction of analytes S627

be used. In this case, this method can be classified as a new dis-persive extraction method free of any organic solvent.

2.2.4. Extraction procedure for crystal violet

10 mL of water sample containing appropriate amount of CVwas placed into a 15 mL falcon tube. 0.5 mL of HCl (0.2 mol L�1) and 1 mL of NaNO3 (1 M) were added and rapidly 0.1 mL

of hydrogel was dissolved in water (0.4% w/v) was injected

into the aqueous sample by a syringe. Separation of two phaseswas achieved by centrifugation for 3 min at 2000 rpm. Theaqueous phase was easily decanted by simply inverting the

tube. The hydrogel extract of this procedure was diluted withacetonitrile (0.3 mL) and transferred into a 0.5 mL quartz cellto measure the absorbance of the solution at 597 nm against a

reagent blank.

Table 3 Some characteristics of the constructed models.

MG CV

Regression equation Coefficient Value Regression equation Coefficient Value

Response = b0 +

b1 · HCl + b2 · HCl ·HCl + b3 · HG · HG

b0 �0.520 Response = b0 + b1 · HCl +

b2 · HG+ b3 · HCl · HCl +

b4 · HCl · NaNO3 + b5 · HG · HG+

b6 · NaNO3 · NaNO3 + b7 · time · time

b0 0.291

|R| 0.964 b1 71.84 |R| 0.913 b1 204.35

R2 0.930 b2 �954.43 R2 0.833 b2 �43.08R2 adjusted 0.915 b3 22.51 R2 adjusted 0.774 b3 �9707.2Standard error 0.03748 Standard error 0.129 b4 �533.04Points no. 18 Points no. 28 b5 567.14

PRESS 0.04 PRESS 0.59 b6 56.77

R2 for prediction 0.862 R2 for prediction 0.702 b7 0.00455

S628 M. Bahram et al.

2.2.5. Central composite design

There are different factors that affect the extraction process. In

order to obtain the optimum conditions, the effect of differentparameters such as concentration of HCl, concentration ofhydrogel, and extraction time was investigated and the opti-

mum conditions were obtained. Therefore, three independentvariables, namely the concentration of HCl (F1), HG (F2),and time of extraction (F3) were studied at five levels with four

repeats at the central point. Preliminary studies showed thepoly (styrene-alt-maleic acid) solution becomes cloudy atacidic pH (pH < 4), therefore instead of pH optimization;the effect of concentration of HCl on the preconcentration

procedure was studied. Circumscribed CCD was used to de-sign the experiments. For each of the four studied variables,high (coded value: +1.428) and low (coded value: �1.428)set points were selected to construct an orthogonal design assummarized in Table 1.

Effect of different parameters concentration of HCl, con-

centration of hydrogel (HG), salting effect (NaNO3) andextraction time were investigated for crystal violet as anothercomponent and the optimum conditions were obtained. For

this case, four independent variables, namely the concentrationof HCl (F1), HG (F2), salting effect (F3) and time of extraction(F4) were studied at five levels with four repeats at the centralpoint. Also for crystal violet preconcentration and extraction

high (coded value: +1.607) and low (coded value: �1.607)set points were selected to construct an orthogonal design (Ta-ble 1).

Table 2 lists the coded values of designed experiments basedon CCD for both analytes.

Polynomial equations, response surface, and central plots

for a particular response were produced using EREGRESS.For an experimental design with three factors, the modelincluding linear, quadratic, and cross terms can be expressed

as Eq. (1).

Response ¼ b0 þ b1 � F1 þ b2 � F2 þ b3 � F3 þ b4 � F1

� F1 þ b5 � F2 � F2 þ b6 � F3 � F3 þ b7

� F1 � F2 þ b8 � F1 � F3 þ b9 � F2 � F3 ð1Þ

F1 to F3 are the variable parameters, and b0 to b9 are the coef-ficient values obtained through multiple linear regressions. The

response surface plots were obtained through a statistical pro-cess that describes the design and the modeled CCD data. Re-

sponse surface methodologies graphically illustrate the

relationships between parameters and responses and are theway to obtain an exact optimum.

For experimental design with four factors (for crystal violet

case) the same model was designed which includes 14 coeffi-cients and one intercept from b0 to b14 (Eq. (2)).

Response ¼ b0 þ b1 � F1 þ b2 � F2 þ b3 � F3 þ b4 � F4

þ b5 � F1 � F1 þ b6 � F1 � F2 þ b7 � F1

� F3 þ b8 � F1 � F4 þ b9 � F2 � F2 þ b10

� F2 � F3 þ b11 � F2 � F4 þ b12 � F3 � F3

þ b13 � F3 � F4 þ b14 � F4 � F4 ð2Þ

3. Result and discussion

3.1. Experimental design

In order to find the important factors and build a model tooptimize the procedure, a full quadratic model including allterms of Eq. (1) for malachite green and Eq. (2) for crystal vio-

let was used in the first step. Then in order to obtain a simpleand yet realistic model, the insignificant terms were eliminatedfrom the model through a ‘backward elimination’ process. For

Figure 2 Response surface plot for malachite green.

Table 4 Optimum conditions obtained by response surface modeling.

MG CV

Variable name Optimum values Selected values Variable name Optimum values Selected values

F1 HCl (mol L�1) 0.029–0.037 0.037 F1 HCl (mol L�1) 0.008–0.012 0.01

F2 HG (%w/v) 0.077–0.098 0.08 F2 HG (%w/v) 0.004–0.008 0.004

F3 Time (min) Insignificant Instantaneously F3 NaNO3 (mol L�1) 0.09–0.1 0.1

F4 Time (min) Insignificant Instantaneously

Figure 3 Response surface plots for crystal violet.

Table 5 Analytical characteristics of the proposed method.

Analytical characteristics Malachite green Crystal violet

Regression equationa Abs = 1 · 106 C � 0.016 Abs = 1 · 106C � 0.059

R2 b 0.999 0.993

Linear range (lmol L�1) 0.01–1 0.05–1

LOD (lmol L�1)d 0.011 0.014

Repeatability (RSD) c f 4.42 2.72

Regression equation before extractiona Abs = 37495C � 0.007 y = 39199C+ 0.015

Concentration factor 20 20

Improvement factor e 26.67 25.5

a Concentration of malachite green in mol L�1.b Squared regression coefficient.c Relative standard deviation for five replicate determinations of 10�7 mol L�1 malachite green.d LOD, limit of detection for S/N= 3.e The ratio of the slope of the calibration graph for the pH-sensitized hydrogel extraction method to that of the slope of the calibration graph

without preconcentration.f Relative standard deviation for seven replicate determinations of 10�7 mol L�1 crystal violet.

A novel hydrogel based microextraction of analytes S629

Table 6 Preconcentration and spectrophotometric determination of malachite green in the spiked water samples by the proposed

method.

Sample Analyte Added (mol L�1) Found (mol L�1) Recovery (%)

Tap water 1 (Nazlou, Urmia-Iran) MG 2 · 10�7 1.9 · 10�7 95

CV 4 · 10�7 3.9 · 10�7 97%

Tap water 2 (Urmia, Iran) MG 2 · 10�7 1.87 · 10�7 93.5

CV 4 · 10�7 3.85 · 10�7 96%

Figure 4 Spectra of hydrogel-rich phase containing malachite

green (8 · 10�7mol L�1) and crystal violet (1 · 10�6mol L�1)

diluted with acetonitrile against blank.

S630 M. Bahram et al.

optimization of malachite green extraction by the eliminationof insignificant terms from the constructed model, calibration

of R2 decreased to 0.930 but adjusted R2 (Radj), and R for pre-diction (R2

pred) increased to 0.915 and 0.862, respectively. Thereduced model using significant linear, quadratic and interac-

tion parameters are shown in Table 3. The results for crystalviolet case are also shown in Table 3.

3.2. Response surface method and selection of the optimumconditions

In order to gain insight about the effect of each variable, thethree dimensional (3D) plots for the predicted responses were

formed based on the model function. One of the response sur-face plots is represented in Fig. 2, which shows the 3D plots ofabsorbance of samples (617 nm) versus pairs of variables while

the other variable was kept at the center level. As shown inFig. 2, there was a non-linear relation between the responseand the variables, because the surface plots of the response

are curved. Using the response surfaces the optimum condi-tions are reached and represented in Table 4. Also Fig. 3 rep-resents the response surface plots for crystal violet analysis.

3.3. Analytical characteristics

Table 5 summarizes the analytical characteristics of the opti-mized method, including regression equation, linear range,

limit of detection, reproducibility, preconcentration andimprovement factors. Because the amount of malachite greenor crystal violet in 10 mL of sample solution is measured after

preconcentration by pH-sensitive hydrogel extraction in a finalvolume of 0.5 mL (0.2 mL Hydrogel rich-phase + 0.3 mL ace-

tonitrile), the solution is concentrated by a factor of 20. Theimprovement factor, defined as the ratio of the slope of the cal-ibration graph for the pH-sensitive hydrogel extraction meth-

od to that of the calibration graph in the presence ofhydrogel without preconcentration (pH > 4). The analyticalcharacteristics for crystal violet and malachite green analysis

are also represented in Table 5.

3.4. Application of the method

This newly introduced method was applied to the determina-

tion of malachite green/crystal violet in several spiked watersamples. The results are presented in Table 6. The spectraare also shown in Fig. 4. The recoveries are close to 100%

and indicate that this method was helpful for the preconcentra-tion and determination purposes.

4. Conclusion

In this paper, we focused on the investigation of a semisolid–liquid dispersive microextraction based on pH-sensitive hydro-

gel. The method has high preconcentration capabilities in avery short time. By using hydrogel, the presented dispersiveextraction method would be free of the organic solvent method

which can be classified as a green analytical method. Highlytoxic, chlorinated solvents, which are usually employed asextractants in DLLME are overcome. In comparison to tradi-tional cloud point extraction methods (using surfactants) and

temperature controlled ionic liquid dispersive liquid-phasemicroextraction, this process avoids the heating and coolingstep, which may lead to the degradation of some thermal

unstable compound, and significantly reduces the extractiontime.

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