Académique Documents
Professionnel Documents
Culture Documents
DOI 10.1007/s10337-016-3022-3
ORIGINAL
Received: 14 August 2015 / Revised: 12 November 2015 / Accepted: 21 December 2015 / Published online: 18 January 2016
© Springer-Verlag Berlin Heidelberg 2016
Abstract A simple microextraction, namely low-density Keywords Low-toxicity organic solvent-based dispersive
solvent with low-toxicity organic solvent-based dispersive liquid–liquid microextraction · QuEChERS · Extraction ·
liquid–liquid microextraction (LDS-DLLME) and modi- HPLC · Neonicotinoid pesticides
fied QuEChERS sample preparation procedures were opti-
mized and validated for preconcentration of neonicotinoid
pesticides prior to the determination using HPLC. The Introduction
experimental parameters affecting the extraction efficiency,
including salt addition, type of disperser solvent and its Neonicotinoids have been the fastest growing class of
volume, effect of extraction solvent and its volume, and insecticides in modern crop protection [1]. There are seven
extraction time were investigated. Under the selected LDS- commercial neonicotinoids: imidacloprid, acetamiprid,
DLLME and HPLC conditions, separation of seven neonic- nitenpyram, thiacloprid, thiamethoxam, clothianidin and
otinoid pesticides (imidacloprid, acetamiprid, clothianidin, dinotefuran. These insecticides are active against numerous
thiacloprid, thiamethoxam, dinotefuran, and nitenpyram) sucking and biting pests and insects, including whiteflies,
was achieved within 26 min. 1-Octanol (extraction solvent) aphids, beetles and some lepidoptera species [2]. They act
and acetonitrile (disperser solvent) were used for extrac- as agonists at the insect nicotinic acetylcholine receptors
tion of the target analytes. Under the optimum condition, (nAChRs), which play an important role in synaptic trans-
linearity was obtained within the range of 0.1–1000 ng g−1 mission in the central nervous system [3]. They can give
with a correlation coefficient more than 0.999. The high rise to serious risks for the health and safety of the consum-
enrichment factor of the target analytes was 50-fold and ers of the agricultural products due to their distribution over
a low limit of quantitation (0.80–2.50 ng g−1) could be large areas of agricultural land [2]. Many countries have
obtained. The fruit samples (at fortified levels of 10, 30, formulated strict limits for neonicotinoids in various matri-
and 50 ng g−1) were successfully analyzed, and relative ces to ensure that the residues are below the safety limit for
recoveries were obtained in the range of 90.07–112.22 %. maximum residue levels (MRLs). The MRLs of neonicoti-
noids range between 0.1 and 1 mg kg−1 [4].
Due to their low volatility and high polarity, neonicoti-
noid insecticides are unsuitable for direct analysis by gas
* Jitlada Vichapong
chromatography [5]. Nowadays, high-performance liquid
jitlada.v@msu.ac.th; jitlada_v@yahoo.com
chromatography (HPLC) coupled with various detection
1
Creative Chemistry and Innovation Research Unit, systems, including electrochemical (ECD) [6], fluorescence
Department of Chemistry, Center of Excellence (FLD) [7], UV/diode array (DAD) [8, 9] and mass spec-
for Innovation in Chemistry, Faculty of Science,
trometry [10], is the favored technique for multi-analysis of
Mahasarakham University, Mahasarakham 44150, Thailand
2
neonicotinoid pesticides. Although a MS detector provides
Materials Chemistry Research Center, Department
more sensitivity and selectivity than UV for monitoring tar-
of Chemistry, Center of Excellence for Innovation
in Chemistry, Faculty of Science, Khon Kaen University, get compounds in complex samples, it suffers from being a
Khon Kaen 40002, Thailand very expensive and complex instrument [11]. Due to their
13
286 J. Vichapong et al.
13
Alternative Liquid–Liquid Microextraction as Cleanup… 287
Results and Discussion 0
0.0 0.5 1.0 1.5 2.0 2.5
The simultaneous separation of the studied neonicotinoids Fig. 2 Effect of concentration of salt on the extraction of neonicoti-
was carried out on a reversed-phase HPLC system with noids
13
288 J. Vichapong et al.
limits when the amount of salt was more than 2.5 g. There- because 1-octanol has a lowrt density than other extraction
fore, 2.5 g Na2SO4 was selected for further studies. solvents (data not shown). Consequently, 1-octanol was
In LDS-DLLME, the dispersive solvent is one of the selected as the extraction solvent for further study.
most important factors that accelerates the emulsification of The effect of extraction solvent volume was studied for
water-immiscible extraction solvent affecting the extraction 1-octanol volume in the range from 50 to 750 µL. It was
efficiency. For the investigation, various types of disperser found that when the extraction solvent volume is 50 µL, the
solvent (50 µL) including acetonitrile, methanol and etha- solution cannot complete phase separation. Moreover, the
nol were studied. As shown in Fig. 3a, the results showed extraction solvent volume of more than 100 µL decreased
that acetonitrile gave a higher extraction efficiency in term the peak area of neonicotinoids. As can be seen from Fig. 4,
of peak area compared to ethanol and methanol; therefore, the highest extraction efficiency was obtained using 100 µL
it was selected as the disperser solvent. of 1-octanol. Therefore, 100 µL of 1-octanol was selected
To investigate the effect of the disperser solvent (ace- for further study.
tonitrile) volume on the extraction efficiency, several vol- Centrifugation time is another important step in LDS-
umes of acetonitrile in the range of 25–250 µL were studied DLLME for achieving phase separation. Therefore, the
(Fig. 3b). It was found that, if the disperser solvent volume effect of centrifugation time was studied in the range of
is 25 µL, the solution cannot complete phase separation. It 1–15 min at 3500 rpm. It was found that the peak areas
was noted that a disperser solvent volume of 50 µL provide of all analytes increased by applying centrifugation up to
Dinotefuran Dinotefuran
Nitenpyram
a 1200000
Nitenpyran
Thiamethoxam
b 900000 Thiamethoxam
Clothianidin Clothianidin
1000000 Imidacloprid 750000 Imidacloprid
Acetamiprid Acetamiprid
Thiacloprid Thiacloprid
800000 600000
Peak area
Peak area
600000 450000
400000 300000
200000 150000
0 0
Ethanol Acetonitrile Methanol 0 50 100 150 200 250
Kind of disperser solvent Volume of acetonitrile (µL)
Fig. 3 Effect of a the kind of disperser solvent and b the volume of extraction solvent on the extraction of neonicotinoids
13
Alternative Liquid–Liquid Microextraction as Cleanup… 289
10 min, and then gradually decreased. Consequently, a cen- 0.999. The enrichment factors, defined as the concentration
trifugation time of 10 min was selected. ratio of the analytes in the settled phase (Cset) and in the
aqueous sample (Co), were 50 for all compounds. The chro-
Analytical Performance of the Method matograms obtained from the separation of standard neo-
nicotinoids by HPLC without and with the LDS-DLLME
Under the above-mentioned conditions, the proposed LDS- procedure are compared in Fig. 5a and b, respectively. The
DLLME combined with the HPLC method for the determi- LOD and LOQ were determined based on the concentra-
nation of neonicotinoid pesticides was validated by deter- tion giving a signal-to-noise ratio of 3 (S/N = 3) and 10
mining its analytical characteristics, i.e. linearity, precision, (S/N = 10), respectively. LOD were found to be in the
limit of detection (LOD), limit of quantitation (LOQ) and range of 0.25–0.80 ng g−1 and LOQ in the range of 0.80–
enrichment factor. The analytical performances of the pro- 2.50 ng g−1. The RSDs of five consecutive extractions of
posed method are summarized in Table 1. The calibration the standard solution containing 50 µg kg−1 nenonicoti-
curves of all the analytes were linear in the range from 0.1 noids were in the ranges of 0.52–1.97 and 2.59–6.38 % for
to 1000 ng g−1 with a correlation coefficient (R2) more than retention time and peak area, respectively.
LOD limit of detection in ng g−1, LOQ limit of quantitation in ng g−1, RSD relative standard deviation (n = 5), EF enrichment factor
a
Values reported in parentheses obtained from the standard neonicotinoids without preconcentration (direct injection)
Fig. 5 Chromatograms of 6
standard neonicotinoids: a 0.040
4
without LDS-DLLME and b
with LDS-DLLME; concen- 0.035
tration of all standards was 5
500 ng g−1. Peak assignments: 7
1 dinotefuran, 2 nitenpyran, 3 0.030
thiamethoxam, 4 clothianidin, 5 3
Voltage (mV)
0.015
0.010
(b)
0.005 1
3
2 4 5 6
7 (a)
0.000
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 21.00 22.00 24.00 26.00 28.00
Time (min)
13
290 J. Vichapong et al.
Table 2 Recovery of the studied neonicotinoids spiked in fruit samples obtained by the proposed method
Analyte Spiked (ng g−1) Watermelon (n = 3) Longan (n = 3) Grape (n = 3)
Recovery (%) RSD (%) Recovery (%) RSD (%) Recovery (%) RSD (%)
Dinotefuran 0 – – – – – –
10 102.78 2.02 98.93 2.54 92.60 3.66
30 98.82 0.78 97.82 2.54 98.93 1.82
50 110.53 0.81 95.28 0.67 90.07 0.10
Nitenpyram 0 – – – – – –
10 94.90 0.75 93.47 2.35 92.97 2.26
30 98.89 2.26 97.72 1.51 100.64 3.21
50 93.79 0.90 99.32 1.79 101.52 0.34
Thiamethoxam 0 – – – – – –
10 97.60 1.42 93.27 2.61 105.60 3.34
30 96.78 0.84 91.78 1.50 104.89 1.73
50 99.25 1.03 100.49 1.12 102.69 1.08
Clothianidin 0 – – – – – –
10 104.75 0.45 100.03 1.30 99.70 1.40
30 97.78 1.49 97.78 1.49 107.78 2.12
50 99.69 2.13 101.02 1.10 105.35 1.20
Imidacloprid 0 – – – – – –
10 104.03 1.25 102.37 3.46 103.37 3.73
30 112.22 2.38 100.22 2.93 102.11 1.50
50 110.69 2.09 99.82 0.33 100.62 1.00
Acetamiprid 0 – – – – – –
10 107.75 2.83 97.70 2.33 97.03 3.03
30 93.22 1.79 99.78 0.33 102.33 2.11
50 91.69 1.95 100.22 0.43 100.09 0.09
Thiacloprid 0 – – – – – –
10 101.7 2.53 105.7 1.82 100.37 4.26
30 93.67 0.51 101.11 0.77 102.33 1.92
50 99.42 1.16 108.69 1.12 100.22 0.63
– not detected
The proposed LDS-DLLME procedure was applied for the A simple miniaturized extraction method, namely low-
determination of neonicotinoid pesticides in fruit samples to density solvent with a low-toxicity organic solvent-based
elucidate the applicability and reliability of this method. In dispersive liquid–liquid microextraction (LDS-DLLME)
this study, no residues of the neonicotinoids were detected and a modified QuEChERS method was developed for the
in the samples (Table 2). In order to validate the accuracy extraction and preconcentration of seven neonicotinoid pes-
of the established method, fruit samples were spiked with ticides in fruit samples before their analysis by HPLC. The
neonicotinoid pesticides at concentration levels of 10, 30, proposed extraction method is simple, fast and effective
and 50 ng g−1. As indicated in Table 2, the recoveries of the and convenient in operation. The developed method shows
studied neonicotinoid pesticides were between 90.07 and good analytical features providing low limits of detection
112.22 % with RSDs of less than 4.26 %. Good recoveries at the level of 0.25–0.80 ng g−1 which is below the accept-
were obtained, indicating that the performance of the method able MRLs for neonicotinoids. High preconcentration fac-
was not affected by the sample co-extractives, at least for the tors, good recoveries and high reproducibility were also
matrices studied in this work. Figure 6 shows a typical chro- obtained. The proposed method has the potential to be used
matogram of watermelon and spiked watermelon samples as an alternative green extraction method for the determina-
using the LDS-DLLME procedure and HPLC analysis. tion of neonicotinoids in fruit samples.
13
Alternative Liquid–Liquid Microextraction as Cleanup… 291
Fig. 6 Chromatograms of a
0.10
watermelon and b watermelon
spiked with 100 ng g−1 of each
neonicotinoid. Peak assign-
ments are described in Fig. 5 0.08
0.06
Voltage (mV)
0.04
1 3 4
0.02 2 6 7
5
(b)
0.00
- 0.02 (a)
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 21.00 22.00 24.00 26.00 28.00
Time (min)
Acknowledgments This article is dedicated to Professor Dr. Kate 10. Jovanov P, Guzsvány V, Franko M, Lazić S, Sakač M,
Grudpan (Chiang Mai University, Thailand) in celebration of his 60th Milovanović I, Nedeljković N (2014) Food Res Int 55:11–19
birthday. The authors gratefully acknowledge the financial supports 11. Vichapong J, Burakham R, Srijaranai S (2013) Talanta
of this research by the Thailand Research Fund (TRF) and Mahasara- 117:221–228
kham University, through the TRF Grant for New Scholars (Grant no. 12. Jansson C, Pihlstrom T, Öterdahl BG, Markides KE (2004) J
TRG5780060). The authors also gratefully acknowledge the partial Chromatogr A 1023:93–104
supports for this research from the Center for Innovation in Chem- 13. Zhou Q, Ding Y, Xiao J (2006) Anal Bioanal Chem
istry (PERCH-CIC), Commission on Higher Education, Ministry of 385:1520–1525
Education, and Materials Chemistry Research Center, Khon Kaen 14. Štajnbaher D, Zupančič-Kralj L (2003) J Chromatogr A
University. 1015:185–198
15. Cazorla-Reyes R, Fernandez-Moreno JL, Romero-Gonzalez R,
Compliance with Ethical Standards Frenich AG, Vidal JL (2011) Talanta 85:183–196
16. Liu S, Zheng Z, Wei F, Ren Y, Gui W, Wu H, Zhu G (2010) J
Conflict of Interest The authors declare that they have no conflict Agric Food Chem 58:3271–3278
of interest. 17. Chitescu CL, Oosterink E, de Jong J, Stolker AAML (2012) Tal-
anta 88:653–662
18. Yáñez KP, Bernal JL, Nozal MJ, Martín MT, Bernal J (2013) J
Chromatogr A 1285:110–117
References 19. Campillo N, Viñas P, Férez-Melgarejo G, Hernández-Córdoba M
(2013) J Agric Food Chem 61:4799–4805
1. Sánchez-Bayo F, Hyne RV (2014) Chemosphere 99:143–151 20. Jovanov P, Guzsvány V, Franko M, Lazić S, Sakač M, Šarić B,
2. Zhang S, Yang X, Yin X, Wang C, Wang Z (2012) Food Chem Banjac V (2013) Talanta 111:125–133
133:544–550 21. Leong M-I, Huang S-D (2008) J Chromatogr A 1211:8–12
3. Di Muccio A, Fidente P, Barbini DA, Dommarco R, Seccia S, 22. Vera-Avila LE, Rojo-Portillo T, Covarrubias-Herrera R, Peña-
Morrica P (2006) J Chromatogr A 1108:1–6 Alvarez A (2013) Anal Chim Acta 805:60–69
4. Pesticide EUMRLs Database (2008) 23. Ahmadi-Jouibari T, Fattahi N, Shamsipur M (2014) J Pharm
5. Wu Q, Li Z, Wang C, Wu C, Wang W, Wang Z (2011) Food Anal Biomed Anal 94:145–151
Methods 4:559–566 24. Sanagi MM, Abbas HH, Ibrahim WAW, Aboul-Enien HY (2012)
6. Rancan M, Sabatini AG, Achilli G, Galletti GC (2006) Anal Food Chem 133:557–562
Chim Acta 555:20–24 25. Xu H, Ding Z, Lv L, Song D, Feng Y-Q (2009) Anal Chim Acta
7. Gil MD, Martínez M, Santiago R, Galanti A, Girotti S (2007) J 636:28–33
Chromatogr A 1147:17–23 26. Nguyen TD, Yu JE, Lee DM, Lee G-H (2008) Food Chem
8. Pareja L, Colazzo M, Pérez-Parada A, Niell S, Carrasco-Letelier 110:207–213
L, Besil N, Cesio MV, Heinzen H (2011) Int J Environ Res Pub- 27. Banerjee K, Oulkar DP, Dasgupta S, Patil SB, Patil SH, Savant
lic Health 8:3844–3858 R, Adsule PG (2007) J Chromatogr A 1173:98–109
9. Tapparo A, Giorio C, Soldà L, Bogialli S, Marton D, Marzaro M, 28. Seebunrueng K, Santaladchaiyakit Y, Srijaranai S (2012) Anal
Girolami V (2013) Anal Bioanal Chem 405:1007–1014 Bioanal Chem 404:1539–1548
13