Professional Documents
Culture Documents
a r t i c l e i n f o a b s t r a c t
Article history: The ionic liquid-based microwave-assisted dispersive liquid–liquid microextraction (IL-based MADLLME)
Received 4 July 2011 and derivatization was applied for the pretreatment of six sulfonamides (SAs) prior to the determina-
Received in revised form tion by high-performance liquid chromatography (HPLC). By adding methanol (disperser), fluorescamine
13 September 2011
solution (derivatization reagent) and ionic liquid (extraction solvent) into sample, extraction, derivati-
Accepted 14 September 2011
zation, and preconcentration were continuously performed. Several experimental parameters, such as
Available online 19 September 2011
the type and volume of extraction solvent, the type and volume of disperser, amount of derivatization
reagent, microwave power, microwave irradiation time, pH of sample solution, and ionic strength were
Keywords:
Ionic liquid-based microwave-assisted
investigated and optimized. When the microwave power was 240 W, the analytes could be derivatized
dispersive liquid–liquid microextraction and extracted simultaneously within 90 s. The proposed method was applied to the analysis of river
In situ derivatization water, honey, milk, and pig plasma samples, and the recoveries of analytes obtained were in the range of
Sulfonamides 95.0–110.8, 95.4–106.3, 95.0–108.3, and 95.7–107.7, respectively. The relative standard deviations varied
High-performance liquid between 1.5% and 7.3% (n = 5). The results showed that the proposed method was a rapid, convenient and
chromatography-fluorescence detection feasible method for the determination of SAs in liquid samples.
© 2011 Elsevier B.V. All rights reserved.
0003-2670/$ – see front matter © 2011 Elsevier B.V. All rights reserved.
doi:10.1016/j.aca.2011.09.018
X. Xu et al. / Analytica Chimica Acta 707 (2011) 92–99 93
2.3.3. Milk
Whole milk samples were purchased in a local supermarket.
The fat content of the milk was 3.5 g/100 mL. 10 mL of homogenized
milk was accurately transferred to a 15 mL polypropylene tube. The
spiked sample was prepared by spiking the mixed working solu-
tion into the milk sample. Then, 1.0 mL trichloroacetic acid (15% in
water) was added, and the solution was shaken and centrifuged at
15,000 rpm for 10 min [41]. The supernatant was collected, diluted
to 10 mL with sodium acetate buffer (pH 3.5). The resulting solution
was referred to as sample solution, filtered through 0.45 m filters
to remove the denatured proteins and then stored at 4 ◦ C.
Fig. 2. Effect of type of disperser. Spiked concentration, 0.5 g L−1 ; extraction sol- Fig. 5. Effect of microwave power. Spiked concentration, 0.5 g L−1 ; extraction
vent ([C6 MIM][PF6 ]) volume, 100 L; disperser volume, 0.75 mL; microwave power, solvent ([C6 MIM][PF6 ]) volume, 100 L; disperser (methanol) volume, 0.75 mL;
240 W; microwave irradiation time, 90 s; derivatization reagent volume, 200 L; pH, microwave irradiation time, 90 s; derivatization reagent volume, 200 L; pH, 3.5.
3.5.
are largest when the volume of methanol is 0.75 mL. The volume
of disperser can directly affect the solubility of IL in aqueous phase
and the volume of IL phase. When the volume of methanol was
excessively small, cloudy suspension could not be formed well.
However, when the volume of methanol was excessively large, both
the volume of IL phase and extraction efficiency of target analytes
decrease due to the increase of the solubility of IL in the aque-
ous phase. Therefore, 0.75 mL methanol was selected in the further
experiments.
Fig. 4. Effect of the volume of disperser. Spiked concentration, 0.5 g L−1 ; extrac- Fig. 6. Effect of microwave irradiation time. Spiked concentration, 0.5 g L−1 ;
tion solvent ([C6 MIM][PF6 ]) volume, 100 L; microwave power, 240 W; microwave extraction solvent ([C6 MIM][PF6 ]) volume, 100 L; disperser (methanol) volume,
irradiation time, 90 s; derivatization reagent volume, 200 L; pH, 3.5. 0.75 mL; microwave power, 240 W; derivatization reagent volume, 200 L; pH, 3.5.
96 X. Xu et al. / Analytica Chimica Acta 707 (2011) 92–99
decrease when the time exceeds 90 s. The increase of extraction the other hand, when the concentration of NaCl was excessively
time was beneficial to the decrease of the viscosity of ILs, the accel- high, the ion exchange between IL and chloride occurs, which could
eration in the rate of derivatization reaction and the mass transfer make [C6 MIM][Cl] soluble in water, leading to the decrease of the
of analytes. However, the increase of extraction time can result in amount of IL phase and the poor extraction performance [44]. On
the increase of solubility of [C6 MIM][PF6 ] in the sample solution, basis of these results, concentration of NaCl was chosen as 3% in all
which resulted in the decrease of extraction efficiency. Based on subsequent experiments.
the experimental results, the irradiation time selected was 90 s.
3.2. Method evaluation
3.1.6. Effect of derivatization reagent volume
Fluorescamine and its hydrolytic products are nonfluorescent. 3.2.1. Analytical performances
This property eliminates extensive clean-up and chromatographic Under the optimal experimental conditions, a series of experi-
separation of the fluorescent derivatives from the excess reagent. ments were performed for obtaining linear ranges, precision, and
To investigate the effect of derivatization reagent volume on extrac- the limits of detection (LODs) and quantification (LOQs). The work-
tion efficiency for in situ derivatization of SAs, different volumes of ing curves were constructed by plotting the peak areas measured
derivatization reagent ranging from 50 to 300 L were used. As versus the concentrations of analytes in the samples. All the experi-
shown in Fig. 7, the peak areas of SAs derivatives increase with the ments were performed in triplicate. The linear regression equations
increase of the volume of derivatization reagent from 50 to 200 L, and correlation coefficients are listed in Table 2. The correlation
and then slightly decrease, which is probably due to quenching coefficients (r2 ) ranging from 0.9989 to 0.9998 are obtained for
effect produced by fluorescamine hydrolytic products [43]. There- all the analytes. The working curves are applied to evaluating the
fore, 200 L of derivatization reagent was selected in the following method LODs and LOQs. The LODs and LOQs were calculated by the
experiments to ensure quantitative derivatization of SAs. following equations:
s s
LODs = 3 ; LOQs = 10
3.1.7. Effect of pH of sample solution k k
The pH of sample solution plays an important role in the extrac- where s is the standard deviation of blank signal, and was obtained
tion of organic compounds because the pH value of the solution by analyzing the blank sample 11 times. k is the slope of the
determines the present state of analytes. Additionally, the deriva-
tization reaction of SAs with fluorescamine needs an acidic medium
[16–18]. On the other hand, when the pH value was too low, the
emulsive phenomenon did not occur, and the volume of IL phase
was sharply reduced. In this experiment, effect of pH on the extrac-
tion performance was investigated, and the results are shown in
Fig. 8. It can be seen that the peak areas of SA derivatives are highest
when pH is 3.5, which is in agreement with the results previously
reported for SA derivatization [18]. Hence, pH 3.5 was chosen in the
following experiments.
Table 2
Analytical performance.
Sample Analyte Liner rangea Regression equationb A = (a ± SDa ) + (b ± SDb )ca Correlation coefficient (r2 ) EF LODsa LOQsa
Honey SMZ 1.00–50.00 A = (2.962 ± 0.904) × 103 + (1.837 ± 0.007) × 104 c 0.9994 26 0.233 0.778
SMP 1.00–50.00 A = (2.959 ± 0.916) × 103 + (1.615 ± 0.007) × 104 c 0.9993 28 0.269 0.898
SMD 1.00–50.00 A = (3.483 ± 1.147) × 103 + (3.133 ± 0.009) × 104 c 0.9998 32 0.174 0.579
SMX 1.00–50.00 A = (3.481 ± 1.111) × 103 + (3.483 ± 0.009) × 104 c 0.9994 33 0.151 0.505
SDM 1.00–50.00 A = (4.276 ± 0.981) × 103 + (3.312 ± 0.008) × 104 c 0.9993 40 0.141 0.469
SPP 1.00–50.00 A = (5.481 ± 0.885) × 103 + (3.492 ± 0.007) × 104 c 0.9990 37 0.120 0.401
Milk SMZ 0.10–5.00 A = (3.529 ± 1.005) × 103 + (1.795 ± 0.008) × 105 c 0.9994 26 0.027 0.089
SMP 0.10–5.00 A = (3.845 ± 1.023) × 103 + (1.571 ± 0.008) × 105 c 0.9998 28 0.031 0.095
SMD 0.10–5.00 A = (8.155 ± 1.482) × 103 + (3.112 ± 0.012) × 105 c 0.9997 32 0.023 0.075
SMX 0.10–5.00 A = (6.553 ± 1.473) × 103 + (3.557 ± 0.011) × 105 c 0.9991 34 0.020 0.066
SDM 0.10–5.00 A = (6.472 ± 1.227) × 103 + (3.308 ± 0.010) × 105 c 0.9989 40 0.018 0.059
SPP 0.10–5.00 A = (5.164 ± 1.318) × 103 + (3.458 ± 0.010) × 105 c 0.9993 38 0.018 0.060
Pig plasma SMZ 0.10–5.00 A = (5.431 ± 0.958) × 103 + (1.651 ± 0.008) × 105 c 0.9997 24 0.028 0.092
SMP 0.10–5.00 A = (3.928 ± 0.996) × 103 + (1.451 ± 0.008) × 105 c 0.9995 25 0.033 0.104
SMD 0.10–5.00 A = (9.275 ± 1.556) × 103 + (2.878 ± 0.013) × 105 c 0.9991 30 0.026 0.086
SMX 0.10–5.00 A = (1.090 ± 0.151) × 104 + (3.176 ± 0.012) × 105 c 0.9994 31 0.023 0.075
SDM 0.10–5.00 A = (9.924 ± 1.291) × 103 + (3.120 ± 0.010) × 105 c 0.9990 38 0.020 0.065
SPP 0.10–5.00 A = (9.679 ± 1.314) × 103 + (3.221 ± 0.011) × 105 c 0.9992 35 0.018 0.062
a
The units of concentrations are g L−1 for river water, milk, pig plasma and g kg−1 for honey.
b
A, peak area of SA; c, SA concentration in g L−1 for river water, milk, pig plasma and g kg−1 for honey; a, intercept; b, slope; SDa , SDb , standard deviations of intercept
and slope, respectively.
working curve. The method LODs and LOQs for all types of matri- the analytes, five replicates were conducted for the sample solu-
ces are shown in Table 2, and the results indicate that the proposed tions containing 5 g L−1 of the analytes. It was found that under
method should be a feasible method in the determination of trace optimal conditions, EFs in the range of 24–44 were obtained.
SAs in various matrices.
Enrichment factor (EF) was calculated by the following equa-
3.2.2. Analysis of samples
tions:
To evaluate the applicability of the proposed method, some real
ca
EF = samples, including environmental water (river water), food (honey
cs
and milk), and biological fluid (pig plasma) were analyzed. The typ-
where ca and cs are the concentrations of the analyte in analytical ical chromatograms of the blank and spiked samples are shown in
and sample solutions, respectively. In order to calculate the EF for Fig. 10. As can be seen, the SAs in the samples are not detectable,
Fig. 10. Typical chromatograms for spiked (A) and blank (B) samples. (a) River water, (b) honey, (c) milk, and (d) pig plasma. The concentrations of analytes are 0.1 g L−1 in
spiked river water, milk, and pig plasma samples and 1 g kg−1 in spiked honey sample. 1: SMZ; 2: SMP; 3: SMD; 4: SMX; 5: SDM; 6: SPP.
98 X. Xu et al. / Analytica Chimica Acta 707 (2011) 92–99
Table 3
RSD (%)
The intra- and inter-day precision and recoveries of the assay.
3.62.3
5.53.9
3.14.2
5.23.5
6.44.1
Sample Analytes Spiked Intra-day Inter-day
concentrationsa
Recovery (%)
Recovery RSD Recovery RSD
103.6105.7
101.3104.9
99.8101.5
106.0104.2
102.298.6
(%) (%) (%) (%)
SPP
1.00 101.0 4.3 99.8 5.6
SMP 0.10 100.5 1.9 100.1 5.5
RSD (%)
1.00 96.4 2.7 98.0 4.0
1.83.1
3.64.1
3.96.4
5.23.3
3.01.9
SMD 0.10 104.5 3.5 106.3 3.8
1.00 110.8 4.6 108.2 5.2
SMX 0.10 97.0 4.1 95.9 5.1
Recovery (%)
1.00 96.1 5.3 95.0 4.0
104.4102.1
104.1105.9
98.4100.1
99.597.6
99.097.3
SDM 0.10 101.8 1.8 104.4 3.1
1.00 98.1 2.6 99.5 5.3
SDM
SPP 0.10 103.6 3.4 105.7 4.2
1.00 105.7 3.1 104.8 6.5
RSD (%)
1.92.6
3.13.6
2.95.1
4.53.0
3.63.0
10.00 106.0 2.1 104.8 3.9
SMP 1.00 95.9 2.6 97.1 5.4
10.00 96.4 3.4 98.4 4.3
SMD 1.00 104.7 2.6 106.3 3.0
Recovery (%)
101.8103.7
104.3101.2
105.3102.0
106.1102.0
10.00 104.1 3.1 103.8 5.8
96.899.4
SMX 1.00 101.8 1.5 99.8 3.4
10.00 104.0 2.5 106.1 5.6
SMX
SDM 1.00 97.3 3.0 95.4 4.7
10.00 96.7 4.6 98.0 7.3
SPP 1.00 100.5 1.9 103.1 4.5
RSD (%)
3.44.2
4.52.9
6.22.5
4.63.0
2.63.0
10.00 102.9 3.1 104.0 5.1
Recovery (%)
SMP 0.10 101.5 2.5 99.8 6.1
98.6102.8
104.2106.0
102.999.1
97.195.6
103.199.7
1.00 103.0 3.4 105.6 6.8
SMD 0.10 96.7 2.5 95.0 5.2
SMD
1.00 97.3 4.0 99.4 4.6
SMX 0.10 99.6 2.3 102.1 4.2
1.00 104.2 3.6 106.9 6.5
RSD (%)
SDM 0.10 103.9 1.9 101.1 4.8
2.65.8
3.63.1
2.82.1
4.56.4
7.86.4
1.00 107.9 2.7 106.3 6.7
SPP 0.10 105.2 1.5 107.0 4.4
1.00 106.8 3.3 108.3 5.4
Recovery (%)
The units of concentrations are g L−1 for river water, milk, pig plasma and g kg−1 for honey.
102.9103.4
102.6103.9
105.2105.9
104.1106.5
Pig plasma SMZ 0.10 96.1 2.4 98.5 5.8
95.798.3
1.00 101.0 3.8 99.6 7.2
SMP 0.10 104.5 3.6 105.8 6.3
SMP
3.92.9
5.62.3
3.23.5
3.63.4
5.24.0
SMX 0.10 104.2 2.1 102.8 5.8
1.00 101.8 4.1 103.4 4.9
SDM 0.10 95.7 3.5 97.0 5.9
1.00 98.4 2.0 96.4 7.3
Recovery (%)
104.6101.1
103.4105.7
98.599.1
97.399.0
a
The units of concentrations are g L−1 for river water, milk, pig plasma and
g kg−1 for honey.
concentrationsa of
1.0010.00
0.101.00
0.101.00
0.101.00
Spiked
was determined by analyzing the samples five times in one day. The
inter-day precision was achieved by analyzing the samples once a
The stability of SAs in the sample.
1-Month
1-Month
1-Month
Pig plasma
spiked samples stored for one month were analyzed. The spiked
samples were all stored at −4 ◦ C except that the spiked pig plasma
Sample
Honey
Table 4
Milk
sample was stored at −20 ◦ C. The results from Table 4 indicate that
a
Table 5
Comparison of this method with other extraction methods for determination of SAs.
Method Matrix Extraction time (min) Recovery (%) Enrichment factor LODsa LOQsa Ref.
Freeze–thaw stability was especially evaluated for the analytes in [2] Z.X. Cai, Y. Zhang, H.F. Pan, X.W. Tie, Y.P. Ren, J. Chromatogr. A 1200 (2008)
plasma samples. The sample was submitted to three freeze–thaw 144–155.
[3] M.A. Raviolo, M. Rambla-Alegre, J. Clausell-Tormos, M.E. Capella-Peiró, S. Carda-
cycles. The each cycle consisted of removing sample from freezer, Broch, J. Esteve-Romero, Anal. Chim. Acta 593 (2007) 152–156.
thawing at room temperature, keeping at room temperature and [4] A. Gentili, D. Perret, S. Marchese, Trends Anal. Chem. 24 (2005) 704–733.
refreezing at −20 ◦ C. The results shown in Table 4 indicate that SAs [5] M. Gros, M. Petrovic, D. Barceló, Anal. Bioanal. Chem. 386 (2006) 941–952.
[6] N.A. Littlefield, W.G. Sheldon, R. Allen, D.W. Gaylor, Food Chem. Toxicol. 28
in pig plasma have an acceptable stability after three freeze–thaw (1990) 157–167.
cycles. [7] Code of Federal Regulations 21 CFR Parts 510 and 556, US Government Printing
Office, Washington, DC, 1987.
[8] Council Regulation (EEC) No. 2377/90 of 26 June 1990 laying down a Commu-
3.2.3. Comparison of IL-based MADLLME and derivatization with nity procedure for the establishment of maximum residue limits of veterinary
other methods medicinal products in foodstuffs of animal origin, Off. J. Eur. Commun. 224
(1990) 1–8.
Some other methods reported in literature, such as solid-phase [9] U.S. Food and Drug Administration (FDA), General Principles for Evaluating the
extraction liquid chromatography with fluorescence detection Safety of Compounds Use in Food-Producing Animals, FDA, Rockville, MD, 1986.
(SPE-LC-FD) [17], sugaring-out extraction HPLC with fluorescence [10] N. Assassi, A. Tazerouti, J.P. Canselier, J. Chromatogr. A 1071 (2005) 71–80.
[11] A. Cannavan, S.A. Hewitt, W.J. Blanchflower, D.G. Kennedy, Analyst 121 (1996)
detection (SAE-HPLC-FD) [18], solid-phase microextraction HPLC
1457–1461.
with ultraviolet detection (SPME-HPLC-UV) [13], stir bar sorp- [12] T. Li, Z.G. Shi, M.M. Zheng, Y.Q. Feng, J. Chromatogr. A 1205 (2008) 163–170.
tive extraction HPLC with ultraviolet detection (SBSE-HPLC-UV) [13] Y. Wen, M. Zhang, Q. Zhao, Y.Q. Feng, J. Agric. Food Chem. 53 (2005) 8468–8473.
[14], and hollow fiber supported liquid phase microextraction [14] X.Y. Huang, N.N. Qiu, D.X. Yuan, J. Chromatogr. A 1216 (2009) 8240–8245.
[15] Y. Tao, J.F. Liu, X.L. Hu, H.C. Li, T. Wang, G.B. Jiang, J. Chromatogr. A 1216 (2009)
HPLC with diode array detection (HF-LPME-HPLC-DAD) [15], were 6259–6266.
compared with the proposed method, and the results are pre- [16] J. Raich-Montiu, J. Folch, R. Compañó, M. Granados, M.D. Prat, J. Chromatogr. A
sented in Table 5. Compared with the reported methods, when the 1172 (2007) 186–193.
[17] A. Zotou, C. Vasiliadou, Chromatography 70 (2009) 389–397.
proposed method is applied, the LODs are lower and the extrac- [18] W.H. Tsai, H.Y. Chuang, H.H. Chen, Y.W. Wu, S.H. Cheng, T.C. Huang, J. Chro-
tion time is shorter. Those reported methods normally involved matogr. A 1217 (2010) 7812–7815.
multi-step operation, which would make the extraction proce- [19] A. Zotou, C. Vasiliadou, J. Sep. Sci. 33 (2010) 11–22.
[20] M.M. Zheng, M.Y. Zhang, G.Y. Peng, Y.Q. Feng, Anal. Chim. Acta 625 (2008)
dure complex and time-consuming. The proposed method was 160–172.
timesaving and convenient because IL-based microwave irradia- [21] L. Verzegnassi, M.C. Savoy-Perroud, R.H. Stadler, J. Chromatogr. A 977 (2002)
tion could obviously improve extraction efficiency and two-step 77–87.
[22] M.Y. Haller, S.R. Müller, C.S. McArdell, A.C. Alder Marc, J.F. Suter, J. Chromatogr.
operation, including preparation of sample solution, and extrac- A 952 (2002) 111–120.
tion, derivatization and preconcentration of the analytes, was [23] M. Rezaee, Y. Assadi, M.R.M. Hosseini, E. Aghaee, F. Ahmadi, S. Berijani, J. Chro-
involved. matogr. A 1116 (2006) 1–9.
[24] D. Nagaraju, S.D. Huang, J. Chromatogr. A 1161 (2007) 89–97.
[25] L.Y. Fu, X.J. Liu, J. Hu, X.N. Zhao, H.L. Wang, X.D. Wang, Anal. Chim. Acta 63
4. Conclusion (2009) 289–295.
[26] Z.G. Shi, H.K. Lee, Anal. Chem. 82 (2010) 1540–1545.
[27] H.X. Chen, H. Chen, J. Ying, J.L. Huang, L. Liao, Anal. Chim. Acta 632 (2009) 80–85.
In this work, a new sample pretreatment method, IL-based [28] S. Pandey, Anal. Chim. Acta 556 (2006) 38–45.
MADLLME and derivatization combined with HPLC-FD, was suc- [29] C.F. Poole, S.K. Poole, J. Chromatogr. A 1217 (2010) 2268–2286.
[30] M.T. Pena, M.C. Casais, M.C. Mejuto, R. Cela, J. Chromatogr. A 1216 (2009)
cessfully applied to the determination of trace SAs in various
6356–6364.
matrices such as river water, honey, milk, and pig plasma. When [31] L.J. He, X.L. Luo, H.X. Xie, C.J. Wang, X.M. Jiang, K. Lu, Anal. Chim. Acta 655 (2009)
IL was used as the extraction solvent not only the consumption of 52–59.
[32] H. Abdolmohammad-Zadeh, G.H. Sadeghi, Talanta 81 (2010) 778–785.
organic solvent can be reduced but also the extraction time can be
[33] L.M. Ravelo-Pérez, J. Hernández-Borges, M. Asensio-Ramos, M.á. Rodríguez-
shortened. Compared with other pretreatment methods reported, Delgado, J. Chromatogr. A 1216 (2009) 7336–7345.
the proposed method can be performed in two steps and has some [34] K.J. Huang, C.Y. Wei, W.L. Liu, W.Z. Xie, J.F. Zhang, W. Wang, J. Chromatogr. A
advantages in extraction time, extraction efficiency and organic 1216 (2009) 6636–6641.
[35] M. Cruz-Vera, R. Lucena, S. Cárdenas, M. Valcárcel, J. Chromatogr. A 1216 (2009)
solvent consumption. 6459–6465.
[36] P. Berton, R.G. Wuilloud, Anal. Chim. Acta 662 (2010) 155–162.
[37] C.L. Ye, Q.X. Zhou, X.M. Wang, Anal. Chim. Acta 572 (2006) 165–171.
Acknowledgements [38] F.Y. Du, X.H. Xiao, G.K. Li, J. Chromatogr. A 1140 (2007) 56–62.
[39] W.Y. Ma, Y.B. Lu, R.L. Hu, J.H. Chen, Z.Z. Zhang, Y.J. Pan, Talanta 80 (2010)
This work was supported by the National Natural Science Foun- 1292–1297.
[40] S.Q. Gao, J.Y. You, X. Zheng, Y. Wang, R.B. Ren, R. Zhang, Y.P. Bai, H.Q. Zhang,
dation of China (No. 20905030) and the China Postdoctoral Science Talanta 82 (2010) 1371–1377.
Foundation (No. 20090461039). [41] J. Han, Y. Wang, C.L. Yu, C.X. Li, Y.S. Yan, Y. Liu, L. Wang, Anal. Chim. Acta 685
(2011) 138–145.
[42] M. Baghdadi, F. Shemirani, Anal. Chim. Acta 613 (2008) 56–63.
References [43] S. de Bernardo, M. Weigele, V. Toome, K. Manhart, W. Leimgruber, P. Böhlen, S.
Stein, S. Udenfriend, Arch. Biochem. Biophys. 163 (1974) 390–399.
[1] S. Yang, J. Cha, K. Carlson, J. Chromatogr. A 1097 (2005) 40–53. [44] Q.X. Zhou, H.H. Bai, G.H. Xie, J.P. Xiao, J. Chromatogr. A 1177 (2008) 43–49.