American Journal of Analyt ical Chemistry, 2011, 2, 1-8
doi:10.4236/ajac.2011.21001 Published Online February 2011 (http://www.SciRP.org/journal/ajac)
Extraction and Determination of Three Chlorophenols by
Hollow Fiber Liquid Phase Micr oextraction -
Spectrophotometric Analysis, and Evaluation
Procedures Using Mean Centering of Ratio
Spectra Method
Zarrin Es’haghi
Department of C hemi st ry , Faculty of Sciences, Payame Noor University, Mashhad, Iran
Received October 16, 2010; revised November 25, 2010; accepted November 27, 2010
Abstract
A method termed hollow fiber liquid phase microextraction (HF-LPME) was utilized to extract three chlo-
rophenols, 2-chlorophenol (2-CP), 2,4-dichlorophenol (2,4-DCP) and 2,4,6-trichlorophenol (2,4,6-TCP),
separately from water. The extracted chlorophenols were then separated, identified, and quantified by
UV-Vis spectrophotometry with photodiode array detection (UV-Vis/DAD). In the study, experimental con-
ditions such as organic phase identity, acceptor phase volume, sample agitation, extraction time, acceptor
phase NaOH concentration, donor phase HCl concentration, salt addition, and UV absorption wavelength
were optimized. The statistical parameters of the proposed method were investigated under the selected con-
ditions. The analytical characteristics of the method such as detection limit, accuracy, precision, relative
standard deviation (R.S.D.) and relative standard error (R.S.E.) were calculated. The results showed that the
proposed method is simple, rapid, accurate and precise for the analysis of ternary mixtures.
Keywords: Chlorophenol, Hollow Fiber Liquid Phase Microextraction (HF-LPME), UV-Vis
Spectrophotometry, Photodiode Array Detector
1. Introduction
Cholorophenols (CPs) are chemical species known to be
highly toxic and a potential threat to public health. CPs
are used extensively as preservatives, fungicides, pesti-
cides, disinfectants, and intermediates in many industries
[1]. CPs are generated from phenols during the treatment
of tap water with chlorine and are considered to be one
of the most obnoxious contaminants [2,3] because they
deteriorate taste and produce an unfavorable smell.
Moreover, they are thought to be serious health hazards
because they accumulate in moderate amounts and show
high toxicity [4,5]. These hazardous materials are usually
detected in human urine because of the intake of food
and water containing CPs and other chlorinated sub-
stances as metabolites present in the environment [6,7].
In order to assess human exposure to CPs, a reliable and
sensitive analytical method is required.
Many analytical methods, including capillary electro-
phoresis [7], high-performance liquid chromatography
(HPLC) [8,9] and gas chromatography (GC) [10,11], are
available for the determination of CPs in human urine
samples. Methods such as ultraviolet-visible spectropho-
tometry may be profitable for some applications instead
of expensive methods such as HPLC and CEC.
Despite highly selective separation and sensitive in-
strumentation for quantification, the direct introduction
of analytes to the instruments is not usually compatible
with environmental determinations.
The sample preparation step in an analytical process
typically consists of an extraction procedure that results
in the isolation and enrich ment of components of interest
from a sample matrix.
Classical extraction procedures consume large amounts
of solvents, thus creating environmental and occupa-
tional hazards, and often provide very little selectivity.

2 Z. ES’HAGHI
These results emphasize the need for reduction of solvent
use, automation, and miniaturization.
Microextraction techniques are the result of looking
for the miniaturization of classical extraction techniques
in order to expend minimum analysis time and chemicals
[12]. In recent years, efforts have been made towards the
miniaturization of the traditional liqu id–liquid extraction.
These techniques, based on the contact between two im-
miscible liquids, fall into two main categories, namely
single-drop microextraction (SDME) [13,14] and hollow
fiber liquid phase microextraction (HF-LPME) [15,16].
Hollow fiber liquid-phase microextraction (HF-LPME)
was introduced by Pedersen-Bjergaard and Rasmussen to
improve the stability and reliability of single drop liq-
uid-phase microextraction (LPME). In this microextrac-
tion technique, a water-immiscible organic solvent was
immobilized in the pores of a porous hollow fiber, and
formed a supported liquid membrane.
The lumen of the hollow fiber was filled with mi-
cro-liter amounts of an acceptor phase. The analytes
were extracted from the aqueous donor phase into the
organic solvent, and then captured into the acceptor
phase. After extraction, the extracted phase was directly
introduced to the analytical instruments [15]. Three
phase HF has been applied to the determinatio n of CPs in
this research.
2. Experimental Section
2.1. Reagents and Samples
2-Chlorophenol (2-CP), 2,4-dichlorophenol (2,4-DCP)
and 2,4,6-trichlorophenol (2,4,6-TCP) were purchased
from Sigma-Aldrich GmbH (Steinheim, Germany). Me-
thanol and acetone and the other solvents used for this
study were of HPLC grade. 1-Octanol was purchased
from Merck, Darmestadth, Germany.
Stock solutions o f the chloroph enols were prepared by
dissolving each chlorophenol in methanol to obtain 100
mg·mL-1 solutions. Aliquots of these stock solutions
were diluted with water to prepare standard working so-
lutions at a concentration of 100 ng/mL. All the other
reagents and solvents used were of analytical reagent
grade.
A Q3/2 Accurel polypropylene hollow fiber mem-
brane (600 µm i.d., 200 µm wall thickness, 0.2 µm pore
size) was purchased from Membrana GmbH (Wuppertal,
Germany). The hollow fiber was cut into 4.0 cm seg-
ments, cleaned with acetone, and dried before use.
De-ionized water was purified in a Milli-Q water puri-
fication system (Millipore, Bedford, MA, USA). Field
samples, a reservoir and a tap water sample, were col-
lected from Mashhad, Iran.
2.2. Extraction Process and Initial Conditions
A 14-mL aqueous solution (0.1 M HCl) containing all
analytes at concentrations of 100 ng/mL each and a 1 cm
stir bar were placed in a 16-mL sample vial previously
loaded with 1 g of NaCl. The solution was stirred at 1000
rpm for 2 min to dissolve the NaCl. Acceptor solution
(12.5 µL, 0.1 M NaOH) was withdrawn into the micro-
syringe. The needle tip of the microsyringe was then
inserted into the 5.0 cm hollow fiber, and the fiber was
filled with the acceptor phase. Before that, the fiber pores
were impregnated with organic solvent. After impregna-
tion, excess organic solvent was carefully removed from
the lumen of the fiber. Then, the fiber was moved to the
aqueous sample solution. The magnetic stirrer was set at
a speed of 500 rpm. The setup arrangement is illustrated
in Figure 1. After extracting for 40 min, the stirrer was
stopped. All acceptor phase (about 12 µL) was with-
drawn back into the microsyringe. Finally, the acceptor
phase was injected into a UV-Vis cell and then was di-
luted to 2.0 mL with acceptor phase solution and intro-
duced into the spectrophotometer for further analysis.
3. Results and Discussion
In this experiment, the chlorophenols were extracted
from an acidified donor phase (sample solution) into an
organic phase, which impregnated the pores of a hollow
fiber. The chlorophenols were then converted into chlo-
rophenolate anion by extracting the chlorophenols from
the organic phase into an alkaline acceptor phase (en-
riched extract). The extracted analytes were then quanti-
fied by UV-Vi s spectrophotometer/DAD.
3.1. Optimization of Variables
The microextraction conditions for the three chlorophe-
Figure 1 . Hollow fiber liquid phase microextraction appa-
ratus.

Z. ES’HAGHI
3
nols must be optimized. In this purpose the univariant
method was conducted to pursue the optimal experimen-
tal conditions for the microextraction procedure. Ana-
lytical parameters, including organic phase identity, ac-
ceptor phase volume, stirring rate, extraction time, ac-
ceptor phase NaOH concen tration, donor ph ase HCl con-
centration, and salt addition were investigated and iden-
tified to improve efficiency. The conditions for LLLME
are as follows: 14 mL of 6.0 M HCl, 1.5 g of NaCl(s) as
the donor phase, 15 µL of 0.1 M NaOH as the acceptor
phase, 1,2,4-trichlorobenzene/1-octanol (70/30) as the
organic phase, 900 rpm stirring speed, and 45 min ex-
traction time.
3.2. Selection of Organic Solvent
The selection principles for a suitable organic solvent are
as follows: First, the solvent should be easily immobi-
lized in the pores of the polypropylene hollow fiber.
Even more important is that the solvent must be of low
volatility and immiscible with water. Most important of
all, the solubility’s of the analytes in the organic solvent
must be higher than in the donor phase. Some proper
organic solvents were examined for extracting chloro-
phenols. Accordingly, o-xylene (OX), p-xylene (PX),
octanol (OC), and 1,2,4-trichlorobenzene (TCB) were
tested. The results shown in Figure 2 indicate how ex-
traction efficiencies were achieved with these aromatic
solvents.
Two xylenes with non-polar functional groups showed
acceptable extraction ability, presumably by forming
induced dipoles by ring-electrons. Trichlorobenzene also
has high extraction efficiency because the molecular
structure of tri-chlorobenzene is very similar to those of
the chlorophenols, except that the low polarity of tri-
chloro benzene is such as to cause low extraction effi-
ciencies for monochlorophenols. To improve extraction
for all the chlorophenols, extractions utilizing various
solvent mixtures (v/v = 1/1) were tested. As trial results
indicated, the extractions of toxic polychlorophenols
were more effective with trichlorobenzene, but 1-octanol
was completely compatible with the polypropylene hol-
low fiber and thus use of solvent mixtures achieves better
reproducibility and precision. Therefore 1,2,4-trichloro-
benzene/octanol (70/30) was selected as the optimal or-
ganic phase.
3.3. Effect of Extraction Time
LLLME is dependent on equilibrium rather than exhaus-
tive extraction [17]. The results obviously indicate that
adequate time must be allowed for the system to reach
equilibrium in the partitioning of analytes between the
donor and acceptor phases (see Figure 3). However, when
considering matching the extraction time with the dura-
tion of spectroscopic analysis, an extraction period of 45
min was chosen for subsequent extractions.
3.4. Effect of Acceptor Phase Volume
The acceptor phase was a basic solution (0.1 M NaOH).
Generally, in the three-phase LLLME systems, a smaller
volume of acceptor phase involves a higher analyte con-
centration (or enrichment) in the acceptor phase [18].
However, the important factor for LLLME is not con-
centration, but the total mass of the analytes in the ac-
ceptor phase. Accordingly, the acceptor phase should be
of large volume to promote analyte transport to the ac-
ceptor phase. The acceptor phase volume was examined
over the range of 7 - 17 µL. As trial results indicated,
15.0 µL of acceptor phase provided superior operation
and this volume was used for subsequent extractions.
3.5. Effect of the Donor Phase PH on the
Extraction
To extract the chlorophenol, a weak acid, into the organic
Figure 2 . Effect of organic solvent on the extraction proce-
dure.
Figure 3. Effect of extraction time on the method efficiency.

4 Z. ES’HAGHI
phase from the donor phase, the pH of the donor phase
was acidified to convert the analytes into their molecular
(i.e., uncharged) form [19].
As trial results indicated, the extraction efficiencies of
the chlorophenols varied slightly with HCl(aq) concen-
tration of the donor phase from 0.01 M to 0.5 M. The
pKa values of the chlorophenols studied range from 6.23
to 8.49. Theoretically, a pH value of the donor phase of
4.0 would be sufficiently acidic. In order to deal with
potential matrix interference in real field samples, HCl
solution with pH = 3 was used.
3.6. Effect of Acceptor Phase PH on the
Extraction
Chlorophenols are weakly acidic in character, so the ac-
ceptor solution must be sufficiently alkaline to convert
them to the ionic form in order to extract them from the
organic phase. On examining NaOH(aq) concentrations
from 0.01 M to 1 M, the results indicate that 0.05 M
NaOH(aq) is satisfactory. Nevertheless, it is not of suffi-
ciently high alkalinity for the extraction of real field
samples. Many contaminants could be extracted into the
acceptor phase from a real field sample matrix; these
contaminants could neutralize the basicity of the acceptor
phase. Because of this, 0.1 M NaOH(aq) was selected as
the acceptor phase.
3.7. Effect of Agitation Speed
Agitation of the donor solution reduces the required ex-
traction time and increases extraction efficiency. Stirring
provides fresh donor solution for the organic phase to
extract and reduces the effect of the stationary boundary
layer zone (Nernestian layer) produced close to the or-
ganic phase; these factors promote analyte transport from
the donor phase to the organic phase [20].
However, agitation in excess of the optimal stirring
rate may also cause lower extraction efficiency because
the hollow fiber is vibrated by the surrounding turbulent
flow and air bubbles reduced absorption for each analyte
and a decrease in the precision of the method [21].
To evaluate the effect of stirring, donor solution was
extracted at varying stirring rates (300 - 1000 rpm). It
was found that the extraction efficiencies of these chloro-
phenols increased at higher stirring rates and reached a
maximum at 900 rpm. Consequently, 900 rpm was cho-
sen for subsequent extractions.
3.8. Effect of Salt Addition
Adding salt to the analytes may have contradictory ef-
fects. First, one expects a positive effect from salting out
when the ionic strength of the donor phase is increased.
This is due to the decrease in the solubility of the ana-
lytes in the aqueous phase and enhances their partition-
ing into the organic phase. Secondly, a negative effect is
attributed to changes in the physical properties of the
Nernst diffusion film, which results in reducing the rate
of diffusion of the analytes from the donor phase into the
organic phase. In addition, the salting out phenomenon
would also reduce the solubility of organic solvent in
water, causing the organic phase to exhaustively wall off
the acceptor phase from the donor phase, a quite desir-
able result.
Therefore, 0.0 - 2.0 g of NaCl(s) was added to 14 mL
of the donor phase to determine the effect of salt addition .
As the experimental results indicated, the addition of 1.5
g NaCl(s) (9.7%, w/w) of sodium chloride optimally
enhanced the extraction of the chlorophenols in water.
This quantity of NaCl was therefore used in subsequent
experiments.
3.9. Quantitative Aspects
Absorbance measurements at the band maxima of the
UV spectra obey the linear Beer’s law more accurately
than measurements off the band maxima. The UV spec-
tra of the analytes can be utilized to identify target ana-
lytes in the UV-Vis spectrophotometer. Accordingly,
each extracted chlorophenol was quantified at its own
maximum adsorption wavelength (as shown in Figure
4).
The chlorophenols were determined under selected
experimental conditions to assess repeatability, linearity,
coefficient of determination, and detection limit. The
results are shown in Table 1.
Figure 4. UV spectra of the various analytes, obtained with
Diod Array Detector (DAD) .

Z. ES’HAGHI
5
Table 1. Method perfor mance.
Compound UV Wavelengh
(nm) Linear range (ng/mL) R2 LODa (ng/mL) RSD(%)b RSD(%)c
2-CP 197 0.5-100 0.9978 01 7.3 3.8
2,4-DCP 201 nm 0.5-100 0.9991 0.08 3.5 2.7
2,4,6-TCP 204 nm 0.5-100 0.9986 0.09 3.9 2.7
a) LODs are calculated as three times the standard devia tion o f seven replica ted runs o f a mixtur e co ntainin g 0.5 ng · mL
-1
each of the co mp oun d; b) Data
are based on the absorbance of seven replica ted runs of a mixture contain ing 0.5 ng · mL
-1
eac h of the compounds; c) Data is based on the absorbance of
seven replicated runs of a mixtur e containi ng 5 ng · mL
-1
ea ch of the co mp ound.
Linearity was established o ver the concentratio n range
of 0.5 - 150 ng/mL. The coefficient of determination r2
varied from 0.9978 to 0.9991. The limit of detection
(LOD) values were calculated as three times the standard
deviation of seven replicate runs of water spiked with
analytes at 1.0 ng/mL each. The relative standard devia-
tion (RSD) values were calculated for seven replicate
runs, and these values were less than 7.4%.
3.10. Real Sample Analysis
Two samples, waste-water (collected from an industrial
center near Mashhad, Iran) and tap water (from Mashhad,
Iran), were analyzed to demonstrate the practical appli-
cability of this technique.
These two samples were filtered with a Milli-Q filter-
ing system (Millipore), and the tap water sample was
overdosed with sodium thiosulfate (80 µg/mL) to neu-
tralize residual chlorine or hypochlorite. The analytical
results of these samples indicated that th e tap water sam-
ple is free from chlorophenols, but a target analyte is
present in the waste-water sample. The detected analyte,
whose concentration is 0.8 ng/mL, coincides with 2,4,5-
trichlorophenol.
For recovery, this LLLME method depends on equi-
librium rather than exhaustive extraction.
Instead of absolute recoveries, relative recoveries de-
termined as the ratio of the real field sample outcome to
that of the deionized water sample are summarized in
Table 2. These results indicate that the relative recover-
ies of chloropheno ls from the spiked waste-water and tap
water samples (performed in three replicates on 0.5
ng/mL of each compound spiked) were 94.1% - 101.4%
and 87.8% - 99.2%, respectively.
3.11. Chemometrics Evaluation of the Method
The main problem of spectrophotometric multicompo-
nent analysis is the simultaneous determination of two or
more compounds in the same mixtures without prelimi-
nary separation.
Table 2. Relative recovery (%) of practical water samples
(N = 3).
Compound WasteA water TapB water
2-CP 94.1 99.2
2,4-DCP 99.9 87.8
2,4,6-TCP 101.4 99.0
A) The sample was obtained from an industrial center, Mashhad, Iran; B)
The tap water was from Mashhad, Iran.
Several spectrophotometric determination methods
have been used for resolving mixtures of compounds
with overlapping spectra. When these methods are com-
pared with each other, the range of application of deriva-
tive spectrophotometry is more reliable with respect to
utility and sensitivity than normal spectrophotometry.
Other spectrophotometric methods, such as partial least
squares regression (PLSR), principal component regres-
sion (PCR), multi-wavelength linear regression analysis
(MLRA), H-point standard addition method (HPSAM)
for binary and ternary mixtures have been utilized [22-
25].
Recently, Bahram et al. proposed a new spectropho-
tometric method for the simultaneous determination of
ternary mixtures, without prior separation steps. This
method is called the successive derivative ratio spectra
[26,27]. The method is based on the successive deriva-
tives of ratio spectra in two steps. The method was
evaluated by model data and also by application to the
simultaneous spectrophotometric determination of cobalt,
nicked and zinc based on their complexes with 1-(2-
pyridylazo)2-naphthol in micellar media as experimental
data .
In our research this new and very simple method was
developed for the simultaneous determination of ternary
chlorophenols mixtures, with prior separation steps. This
method is based on the mean centering of ratio spectra.
This method eliminates derivative steps and therefore
signal-to-noise ratio is enhanced.

6 Z. ES’HAGHI
4. Modeling
4.1. Validation of the Method
Validation of the method with model data to demonstrate
the analytical applicability of the proposed method for
the analysis of ternary mixtures, three spectra were cre-
ated. The three curves form model of the overlapping
spectra of three compounds X, Y and Z in the range 190 -
230 nm (Figure 4).
The mathematical explanation of the procedure was
illustrated by the authors. After modeling procedure, the
method has been applied to the simultaneous analysis of
ternary mixtures of chloprophenols. The analytical char-
acteristics of the method such as detection limit, accu-
racy, precision, relative standard deviation (R.S.D.) and
relative standard error (R.S.E.) was calculated. The re-
sults showed that the proposed method is accurate and
precise method for analysis of ternary mixtures (see Ta-
bles 3 and 4).
Limit of detection of the method for determination of
chloprophenols in their ternary mixtures (defined as the
concentration equivalent to three times the standard de-
viation of five replicate measurements of the blank) are
also shown in Table 4.
The effect of divisor concentration on the analytical
parameters such as detection limit, slope, intercept and
correlation coefficient of the calibration equations was
tested. It was observed that changing the concentration of
divisors in their linear calibration range had no signifi-
cant effect on the analytical parameters.
Therefore, a normalized spectrum of each of the ana-
lytes was used as divisor spectrum in the proposed me-
thod.
In order to obtain the accuracy and precision of the
method, several synthetic mixtures with different con-
centration ratios of chlorophenols were analyzed using
the proposed method.
The prediction error of a single component in the mix-
tures was calculated as the relative standard error (R.S.E.)
Table 3 . Results for several experiments for analysis of chlorophenols in ternary mixtures in different concentration ratios by
proposed method, HF-LPME-MCRS.
Conc. (ng/mL) (in mixture) Estimated Conc. (ng/mL) Recovery (%)
2-CP 2,4-DCP 2,4,6-TCP 2-CP 2,4-DCP 2,4,6-TCP 2-CP 2,4-DCP 2,4,6-TCP
0.7 0.5 0.1 0.76 0.58 0.11 108.57 116 110
0.2 0.9 0.1 0.2 0.81 0.15 100 90 100
0.4 0.8 0.2 0.41 0.93 0.28 102.5 98.75 90
0.4 0.8 1 0.33 0.73 1 82.5 98.75 100
0.8 0.2 0.3 1 0.24 0.29 125 110 95
1 0.1 0.6 0.96 0.1 0.63 96 100 95
0.9 0.4 0.2 0.9 0.45 0.2 100 95 100
0.9 0.9 0.8 0.85 0.86 0.88 94.44 95.55 105
0.6 0.6 0.1 0.57 0.61 0.1 95 101.66 100
0.9 0.5 0.4 0.91 0.59 0.33 101.11 98 82.5
Mean recovery % 100.51 100.37 97.92
R.S.E. single % 9.879 6.751 5.862
R.S.E. total % 7.5
Table 4
. Analytical aspects of results in ternary mixture by HF-LPME-MCRS methods.
Analyte Wavelength max Calibration slope Calibration
intercept R2 Linear range
(ng·mL-1) LOD
(ng·mL-1)
2-CP 197 9.99 –0.0066 0.9999 0.01 - 100 0.007
2,4-DCP 201 86.26 –14.446 0.9960 0.01 - 100 0.008
s2,4,6-TCP 204 8.27 +0. 0139 0.9998 0.01 - 100 0.008

Z. ES’HAGHI
7
of the prediction concentration. Table 4 also shows the
reasonable single and total relative errors for such sys-
tem.
5. Conclusions
The results obtained in this work ind icate the application
of hollow fiber three liquid phase microextraction of
chlorophenols from water samples, with the obvious ad-
vantages of higher enrichment factors and lower detec-
tion limits. This technique is simple, economical, safe fo r
the examiner, rapid, sensitive and easy to understand and
apply.
The application of UV-Vis spectrophotometry for the
simultaneous determination of analytes in water samples
is feasible by using homemade software. This technique
consists of a fast, inexpensive proc edure and provides an
easy method with which to assay the target analytes in
environmental samples.
6. References
[1] R. C. C. Wegman and A. W. M. Hofster, “Chlorophenols
in Surface Waters of the Netherlands (1976-1977),” Wa-
ter Research, Vol. 13, 1979, pp. 651-657.
doi:10.1016/0043-1354(79)90015-0
[2] J. Lezamiz and J. A. Jönsson, “Development of a Simple
Hollow Fibre Supported Liquid Membrane Extraction
Method to Extract and Preconcentrate Dinitrophenols in
Environmental Samples at ng•L(-1) Level by Liquid
Chromatography,” Journal of Chromatography A, Vol.
1152, No. 1-2, 2007, pp. 226-233.
doi:10.1016/j.chroma.2006.11.104
[3] T. Mathialagan and T. Viraraghavan, “Biosorption of
Chlorophenols: A Review,” International Journal of En-
vironment and Pollution, Vol. 34, No. 1-4, 2008, pp.
164-194. doi:10.1504/IJEP.2008.020790
[4] A. Ribeiro, M. H. Neves, M. F. Almeida, A. Alves and L.
Santos, “Direct Determination of Chlorophenols in Land-
fill Leachates by Solid-Phase Micro-Extraction-Gas
Chromatography-Mass Spectrometry,” Journal of Chro-
matography A, Vol. 975, No. 2, 2002, pp. 267-274.
doi:10.1016/S0021-9673(02)01280-3
[5] F. Bianchi, M. Careri, C. Mucchino and M. Musci, “Im-
proved Determination of Chlorophenols in Water by
Solid-Phase Microextraction Followed by Benzoylation
and Gas Chromatography with Electron Capture Detec-
tion,” Chromatographia, Vol. 55, 2002, pp. 595-600.
doi:10.1007/BF02492907
[6] M. C. Quintana and L. Ramos, “Sample Preparation for
the Determination of Chlorophenols,” Trends in Analyti-
cal Chemistry, Vol. 27, No. 5, 2008, pp. 418-436.
doi:10.1016/j.trac.2008.03.009
[7] P. Verónica, A. Juan, G. Venerando and A. Ana, “Moni-
toring Chlorophenols in Industrial Effluents by Solid-
Phase Microextraction-Gas Chromatography-Mass Spec-
trometry,” International Journal of Environmental Ana-
lytical Chemistry, Vol. 87, No. 3, 2007, pp. 159-175.
doi:10.1080/03067310600847252
[8] Q. B. Cass, L. G. Freitas, E. Foresti, M. Zamariolli and M.
H. R. Aminaovic, “Developmet of HPLC Method for the
Analysis of Chlorophenosl in Samples from Anaerobic
Reactors for Waste Water Treatment,” Journal of Liquid
Chromatography & Related Technologies, Vol. 23, No. 7,
2000, pp. 1089-1097. doi:10.1081/JLC-100101510
[9] T. Araki, M. Chiba, S. Tsunol and M. Tanaka, “Separa-
tion of Chlorophenols by HPLC and Capillary Electro-
chromatography Using beta-Cyclodextrin-bonded Sta-
tionary Phases,” Analytical Sciences, Vol. 16, No. 4,
2000, pp. 421-424. doi:10.2116/analsci.16.421
[10] L. W. Chung and M. R. Lee, “Evaluation of Liquid-Phase
Microextraction Conditions for Determination of Chloro-
phenols in Environmental Samples Using Gas Chroma-
tography-Mass Spectrometry without Derivatization,”
Talanta, Vol. 76, 2008, pp. 154-160.
doi:10.1016/j.talanta.2008.02.018
[11] P. Tölgyessy, B. Vrana, M. Bartal, Z. Krascsenits and K.
Šilhárova, “Determination of Chlorophenols in Sedi-
ments Using Ultrasonic Solvent Extraction Followed by
Stir Bar Sorptive Extraction Coupled to TD-GC-MS,”
Chromatographia, Vol. 69, 2009, pp. 389-392.
doi:10.1365/s10337-008-0877-y
[12] J. A. Jönsson and L. Mathiasson, “Liquid Membrane
Extraction in Analytical Sample Preparation: I. Princi-
ples,” Trends in Analytical Chemistry, Vol. 18, No. 5,
1999, pp. 318-325. doi:10.1016/S0165-9936(99)00102-8
[13] E. Psillakis and N. Kalogerakis, “Developments in Sin-
gle-Drop Microextraction,” Trends in Analytical Chemis-
try, Vol. 21, No. 1, 2002, pp. 53-63.
doi:10.1016/S0165-9936(01)00126-1
[14] M. A. Jeannot and F. F. Cantwell, “Solvent Microextrac-
tion into a Single Drop,” Analytical Chemistry, Vol. 68,
1996, pp. 2236-2240. doi:10.1021/ac960042z
[15] J. Lee, H. K. Lee, K. E. Rasmussen and S. Peder-
sen-Bjergaard, “Environmental and Bioanalytical Appli-
cations of Hollow Fiber Membrane Liquid-Phase Micro-
extraction: A Review,” Analytica Chimica A cta, Vol. 624,
No. 2, 2008, pp. 253-268. doi:10.1016/j.aca.2008.06.050
[16] K. E. Rasmussen and S. Pedersen-Bjergaard, “Develop-
ments in Hollow Fibre-Based, Liquid-Phase Microextrac-
tion,” Trends in Analytical Chemistry, Vol. 23, No. 1,
2004, pp. 1-10. doi:10.1016/S0165-9936(04)00105-0
[17] C. C. Chen, M. B. Melwanki and S. D. Huang, “Liq-
uid-Liquid-Liquid Microextraction with Automated
Movement of the Acceptor and the Donor Phase for the
Extraction of Phenoxyacetic Acids Prior to Liquid Chro-
matography Detection,” Journal of Chromatography A,
Vol. 1104, No. 1-2, 2006, pp. 33-39.
doi:10.1016/j.chroma.2005.11.122
[18] A. Sarafraz-Yazdi and Z. Es’haghi, “Liquid-Liquid-Liquid
Phase Microextraction of Aromatic Amines in Water Us-
ing Crown Ethers by High-Performance Liquid Chroma-

8 Z. ES’HAGHI
tography with Monolithic Column,” Talanta, Vol. 66, No.
3, 2005, pp. 664-669. doi:10.1016/j.talanta.2004.12.026
[19] A. Gjelstad, T. M. Andersen, K. E. Rasmussen and S.
Pedersen-Bjergaard, “Microextraction across Supported
Liquid Membranes Forced by pH Gradients and Electri-
cal Fields,” Journal of Chromatography A, Vol. 1157, No.
1-2, 2007, pp. 38-45. doi:10.1016/j.chroma.2007.05.007
[20] W. Xie, J. Pawliszyn, W. M. Mullett and B. K. Matus-
zewski, “Comparison of Solid-Phase Microextraction and
Liquid-Liquid Extraction in 96-Well Format for the De-
termination of a Drug Compound in Human Plasma by
Liquid Chromatography with Tandem Mass Spectromet-
ric Detection,” Journal of Pharmaceutical and Biomedi-
cal Analysis, Vol. 45, No. 4, 2007, pp. 599-608.
doi:10.1016/j.jpba.2007.08.029
[21] A. S. Yazdi and Z. Es’haghi, “Two-step Hollow Fiber-
Based, Liquid-Phase Microextraction Combined with
High-Performance Liquid Chromatography: A New Ap-
proach to Determination of Aromatic Amines in Water,”
Journal of Chromatography A, Vol. 1082, No. 2, 2005,
pp. 136-142. doi:10.1016/j.chroma.2005.05.102
[22] M. Palit, D. Pardasani, A. K. Gupta and D. K. Dubey,
“Application of Single Drop Microextraction for Analy-
sis of Chemical Warfare Agents and Related Compounds
in Water by Gas Chromatography/Mass Spectrometry,”
Analytical Chemistry, Vol. 77, 2005, pp. 711-717.
doi:10.1021/ac0486948
[23] E. Dinç and F. Onur, “Application of a New Spectropho-
tometric Method for the Analysis of a Ternary Mixture
Containing Metamizol, Paracetamol and Caffeine in Tab-
lets,” Analytica Chimic a Acta, Vol. 359, No. 1-2, 1998, pp.
93-106. doi:10.1016/S0003-2670(97)00418-2
[24] S. Wold, M. Sjostrom and L. Eriksson, “PLS-Regression:
A Basic Tool of Chemometrics,” Chemometrics and In-
telligent Laboratory Systems, Vol. 58, No. 2, 2001, pp.
109-130. doi:10.1016/S0169-7439(01)00155-1
[25] E. Dinc, C. Yucesoy and F. Onur, “Simultaneous Spec-
trophotometric Determination of Mefenamic Acid and
Paracetamol in a Pharmaceutical Preparation Using Ratio
Spectra Derivative Spectrophotometry and Chemometric
Methods,” Journal of Pharmaceutical and Biomedical
Analysis, Vol. 28, No. 6, 2002, pp. 1091-1100.
doi:10.1016/S0731-7085(02)00031-6
[26] M. Bahram, “Mean Centering of Ratio Spectra as a New
Method for Determination of Rate Constants of Consecu-
tive Reactions,” Analytica Chimica Acta, Vol. 603, No. 1,
2007, pp. 13-19. doi:10.1016/j.aca.2007.09.041
[27] A. Afkhami and M. Bahram, “Mean Centering of Ratio
Spectra as a New Spectrophotometric Method for the
Analysis of Binary and Ternary Mixtures,” Talanta, Vol.
66, 2005, pp. 712-720. doi:10.1016/j.talanta.2004.12.004