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 Author's personal copy

Food Chemistry 141 (2013) 2675–2681

Contents lists available at SciVerse ScienceDirect

Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Evaluation of an analytical methodology using QuEChERS and GC-SQ/MS

for the investigation of the level of pesticide residues in Brazilian melons
Jonas da Silva Sousa a, Rubens Carius de Castro b, Gilliane de Albuquerque Andrade b,
Cleidiane Gomes Lima b, Lucélia Kátia Lima b, Maria Aparecida Liberato Milhome b,
Ronaldo Ferreira do Nascimento c,⇑
a
Departamento de Química Orgânica e Inorgânica, Universidade Federal do Ceará, Rua Humberto Monte, S/N Campus do Pici, Bl. 935, CEP 60451-970 Fortaleza, CE, Brazil
b
Fundação Núcleo de Tecnologia Industrial do Ceará (NUTEC), Rua Rômulo Proença S/N, Campus do Pici, CEP 60451-970 Fortaleza, CE, Brazil
c
Departamento de Química Analítica e Físico Química, Universidade Federal do Ceará, Rua Humberto Monte, S/N Campus do Pici, Bl. 940, CEP 60451-970 Fortalezam, CE, Brazil

a r t i c l e i n f o a b s t r a c t

Article history: A multiresidue method based on the sample preparation by modified QuEChERS and detection by gas
Received 21 February 2013 chromatography coupled to single quadruple mass spectrometers (GC-SQ/MS) was used for the analysis
Received in revised form 27 April 2013 of 35 multiclass pesticides in melons (Cucumis melo inodorus) produced in Ceara-Brazil. The rates of
Accepted 9 May 2013
recovery for pesticides studied were satisfactory (except for the etridiazole), ranging from 85% to 117%
Available online 18 May 2013
with a relative standard deviation (RSD) of less than 15%, at concentrations between 0.05 and
0.20 mg kg1. The limit of quantification (LOQ) for most compounds was below the MRLs established
Keywords:
in Brazil. The combined relative uncertainty (Uc) and expanded uncertainty (Ue) was determined using
Multiresidue
Melon
repeatability, recovery and calibration curves data for each pesticide. Analysis of commercial melons
QuEChERS samples revealed the presence of pesticides bifenthrin and imazalil at levels below the MRLs established
GC-SQ/MS by ANVISA, EU and USEPA.
Uncertainty Ó 2013 Elsevier Ltd. All rights reserved.

1. Introduction The quality control of pesticide residues becomes important

Melon is a crop of great economic importance and prominence
in the national and international market. Melons are first in terms
of the volume of fresh fruit exported from Brazil, the main market
being the European Union, followed by North America. The North-
east of Brazil holds about 95% of melon production, mainly in the
states of Rio Grande do Norte, Ceara, Pernambuco and Bahia, which
are major exporters of melons (Neitzke et al., 2009).
A major concern of the government in relation to agricultural
growth is the indiscriminate use of pesticides. In the last 10 years
the market has increased by 190%. In 2008, Brazil surpassed the
United States and assumed the position of the largest global mar-
ket for pesticides (ABRASCO., 2012).
The melon crop demands large pesticide consumption. In Brazil,
about 95 products are registered and authorised for this crop, rep-
resenting 58 active ingredients and 28 different chemical groups.
About 30% of these products are classified as highly toxic, 46% as
moderately toxic and 25% as slightly toxic. Within the class of
insecticides, there are various products of chemical grouping of
organophosphates and pyrethroids, which have high toxicity
(ANVISA – Agência Nacional de Vigilância Sanitária., 2012).
⇑ Corresponding author. Tel.: +55 85 3366 9958; fax: +55 85 3366 9982.
E-mail address: ronaldo@ufc.br (R.F. do Nascimento).
mainly in fruits like melon that are consumed ‘‘in natura’’, in salads
or minimally processed. The ingestion of foods containing high lev-
els of residues can cause serious problems of human poisoning,
which can lead to dermatitis, respiratory problems, eye damage
and mutagenic and teratogenic changes (Nougadère, Reninger, Vol-
atier, & Leblanc, 2011). Several studies have reported effects and
risks of contamination from pesticides residues (Camino-Sánchez
et al., 2011; Jardim & Caldas, 2012; Kruve, Lamos, Kirillova, & Her-
odes, 2007; Milhome, Sousa, Lima, & Nascimento, 2009).
In Brazil, monitoring programs have been implemented since
2001 with the aim of evaluating the quality of food and implement
control actions for pesticide residues (Jardim & Caldas, 2012). At
least one pesticide residue was found in 48.3% of the samples
analysed from 2001 to 2010 in the Brazilian monitoring program.
However, due to the wide variety of fruits and vegetables
consumed by the population and the increasing production of
new active principles, the implementation and adaptation of meth-
odologies for multiresidue analysis becomes relevant (Milhome,
Sousa, Keukeleire, & Nascimento, 2011; Rodrigues et al., 2011).
The analysis of pesticides in complex matrices, such as fruits,
require sample preparation methods that are very efficient, and
are able to extract the contaminants at low levels (Prestes, Friggi,
Adaime, & Zanella, 2009). In recent years, changes in the original
QuEChERS method, have been suggested to improve the extraction

0308-8146/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodchem.2013.05.027
 Author's personal copy

2676 J. da Silva Sousa et al. / Food Chemistry 141 (2013) 2675–2681

of multiresidues. In particular adding salt to the extract in acetoni-
trile aims to promote the effect of ‘‘salting out’’, and improve the
efficiency in separation and recovery of the analytes (González-
Curbelo, Hernández-Borges, Ravelo-Pérez, & Rodríguez-Delgado,
2011; Koesukwiwat, Lehotay, Miao, & Leepipatpiboon, 2010; Lu
et al., 2012; Prestes et al., 2009).
Different types of detectors have been used for pesticide
residues analysis (Camino-Sánchez et al., 2011; Furlani, Marcilio,
Leme, & Tfouni, 2011; Iñigo-Nuñez, Herreros, Encinas, &
Gonzalez-Bulnes, 2010). Rahman et al. (2013) used HPLC/UVD
and LC–MS–MS for the determination of dinotefuran and its
metabolites in melons. Frenich, Vidal, Ĺopez, Cort́es Aguado, and
Salvador (2004) developed a new analytical method using liquid
chromatography with mass spectrometry (LC–MS–MS) for the
analysis of 31 pesticide residues in samples of green beans, cucum-
ber, pepper, tomato, eggplant, watermelon, melon and zucchini.
Gas chromatography coupled to single quadruple mass spec-
trometers (GC-SQ/MS) has been used for multiresidue analysis
due to its good resolution, selectivity and sensibility expected gi-
ven purpose, in addition to its versatility, as it can be used to ana-
lyse different matrices (Menezes Filho, Santos, & Pereira, 2010;
Yang et al., 2011). Camino-Sánchez et al. (2011) used the QuE-
ChERS method and detection by GC–MS–MS for the quantification
of 121 pesticide residues in samples of tomato, pepper, lettuce,
cucumber, eggplant, zucchini, melon, watermelon and apple ac-
quired from Spain.
This paper describes the application of the QuEChERS method
for the evaluation of the level of 35 pesticide residues in samples
of melons marketed in Fortaleza-CE-Brazil, during a period of
3 months. For this study different classes of pesticides (inseticides,
fungicides, herbicides and acaricides) were selected considering
the following aspects: (a) pesticides authorised by ANVISA for
the melon crop (b) application in fruits produced in Ceará state,
based on previous studies Milhome et al. (2009), (b) feasibility
analysis by GC-SQ/MS. Among the pesticides analysed, 15 active
ingredients are approved for use by ANVISA in melon crops.
Parameters validation and the calculation of uncertainty were
determined to ensure the reliability of the method using GC-SQ/
MS system. The assessment of the levels of exposure to pesticide
residues in fruits consumed by the population is of fundamental
importance to food security.
2. Materials and methods
2.1. Chemicals and reagents
Certified standards for 35 pesticides studied were purchased
from Sigma–Aldrich (Brazil) and Dr. Ehrenstorfer (Brazil). All the
standards had purities exceeding 97.0%.
HPLC grade methanol and acetonitrile were acquired from
Merck (Brazil), ethyl acetate and cyclohexane from J.T. Baker
(U.S). The reagents anhydrous magnesium sulphate P.A. (Vetec,
Brazil), sodium chloride P.A. (Vetec, Brasil), trisodium citrate dihy-
drate P.A. (Vetec, Brazil), sodium hydrogen citrate sesquihydrate
P.A (Sigma–Aldrich, Brazil), Bondesil Primary/Secundary Amine
(PSA) 40 lm (Supelco, EUA) were analytical purity. Standard stock
solutions were prepared in methanol or acetone, at 1000 lg mL1.
A standard mixture solution, with all 35 pesticides was prepared in
ethyl acetate:cyclohexane (1:1) at 1 lg mL1 of each pesticide. This
solution was used as spiking solution and also to prepare the
standard solutions to obtain the analytical curves by dilution with
acetate:cyclohexane (1:1) or matrix extract.

Table 1
The retention times of 35 pesticides analysed and their fragments quantitative (ion target) and qualifiers (ions Q1, Q2 and Q3).

Number Pesticide Chemical group Class Retention time (min) Ion T Ion Q1 Ion Q2 Ion Q3
1 Etridiazole Aromatic hydrocarbon F 8.23 211 183 185 213
2 Chloroneb Substituted benzene F 8.77 193 141 191 208
3 Molinate Thiocarbamate H 9.12 126 98 187 –
4 Propachlor Chloroacetamide H 9.97 120 169 176 –
5 Trifluralin Dinitroaniline H 10.69 306 248 264 290
6 Hexachlorobenzene Organochlorine F 11.40 286 142 282 284
7 Atrazine Triazine H 11.70 200 173 202 215
8 Chlorothalonil Chloronitrile F 12.82 266 264 267 268
9 Parathion-methyl Organophosphate A/I 13.91 263 109 125 –
10 Ametryn Triazine H 14.16 227 170 185 212
11 Metalaxyl Phenylamide F 14.34 160 130 132 206
12 Fenitrothion Organophosphate F/I 14.80 277 109 125 260
13 Malathion Organophosphate A/I 15.19 173 125 127 158
14 Chlorpyrifos Organophosphate A/F/I 15.56 314 197 199 316
15 Dimethyl tetrachloroterephthalate (DCPA) Chlorinated benzoic acid H 15.73 301 299 303 332
16 Pendimethalin Dinitroaniline H 16.73 252 162 191 281
17 Cis-chlordane Organochlorine I 17.61 373 371 375 377
18 Trans-chlordane Organochlorine I 18.19 373 371 375 377
19 Imazalil Imidazole F 18.73 215 173 175 217
20 Buprofezin Thiadiazinone A/I 19.38 105 104 106 172
21 Kresoxim-methyl Strobilurin F 19.60 116 131 132 206
22 Cyproconazole Triazole F 19.80 222 125 139 224
23 Chlorobenzilate Organochlorine I/A 20.24 139 141 251 253
24 Triazophos Organophosphate A/I 21.35 161 134 162 172
25 Bifenthrin Pyrethroid A/F/I 24.44 181 165 166 182
26 Pyriproxyfen Pyridyl ether I 25.96 136 96 137 226
27 Lambda-cyhalothrin Pyrethroid I 26.75 181 197 208 –
28 Fenarimol Pyrimidine F 27.81 139 107 219 –
29 Trans-permethrin Pyrethroid F/I 28.36 183 163 164 165
30 Cis-permethrin Pyrethroid F/I 28.66 183 163 164 165
31 Boscalid Anilide F 30.45 140 112 114 142
32 Pyraclostrobin Strobilurin F 32.68 132 111 133 164
33 Deltamethrin Pyrethroid F/I 34.72 181 172 251 –
34 Azoxystrobin Strobilurin F 35.67 344 345 372 388
35 Imibenconazole Triazole F 37.44 125 127 251 376
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2.2. Samples extraction and clean-up procedures 2.3. GC-SQ/MS analysis

The melon samples were purchased at supermarkets in Fort-
aleza, Ceará, Brazil, obtained from three different suppliers (S1,
S2 and S3). Analyses were conducted between October 2011 and
December 2011. Three samples were collected monthly, one in
each supplier, for monitoring 35 pesticides probably used in fruits.
The entire process of receiving, processing and storage of the fruits
was performed according to the SANCO Guidelines (European
Commission., 2009).
Melon samples were analysed based on QuEChERS method
described by Anastassiades, Lehotay, Tajnbaher, and Schenck
(2003). A portion of 10.0 g crushed and homogenised sample was
weighed into a 50.0 mL PTFE centrifuge tube and 10.0 mL of aceto-
nitrile were added, soaking for 1 min by using a vortex mixer. 4.0 g
MgSO4, 1.0 g NaCl, 1.0 g trisodium citrate dihydrate and 0.5 g
disodium hydrogen citrate sesquihydrate were added and then
vortexed immediately for 1 min to prevent the formation of coag-
ulated sodium sulphate.
The extract was centrifuged for 10 min at 3600 rpm, using a
centrifuge Fanen (São Paulo, Brazil). 4.0 mL aliquots of the extracts
were transferred to 15 mL tubes and 600.0 mg of magnesium sul-
fate and 100.0 mg of PSA were added. After centrifugation at
3600 rpm for 10 min, 10 lL of 5% formic acid solution (for the sta-
bilization of the extracts) were added in 3.0 mL aliquots of the ex-
tract and put into a vacuum concentrator to dryness. The pesticide
residues were reconstituted in 3.0 mL of cyclohexane:ethyl acetate
(1:1) and the final extracts were directly analysed by GC-SQ/MS
(Thermo, USA).
A gas chromatograph coupled to a single quadrupole mass spec-
trometer(GC-SQ/MS, DSQII model, Thermo, USA), of the Fundação
Núcleo de TecnologiaIndustrial do Ceará – NUTEC was used.
A Vertibond 50 m  0.25 mm i.d. capillary column with a
0.10 lm film was used in the pesticide separation and helium as
carrier gas at a constant flow of 1 mL min1. The temperature pro-
gram was the following: initial temperature 100 °C held for 1 min,
15 °C min1 rate to 180 °C, then 4 °C min1 rate to 280 °C and held
for 14 min. The injection temperature was 250 °C, and 1 lL volume
was injected in splitless mode (45.3 min). The mass spectrometer
was operated in electron impact (EI) mode, ion source temperature
200 °C, MS Transfer Line 270 °C, electron multiplier voltage 1295 V,
scanning from m/z 50 to 500 at 2.0 s/scan; solvent delay 6.0 min.
Quantitative analysis was performed in the selected ion moni-
toring mode (SIM) based on the use of one ion target (T) and two
or three ions qualifiers (Q1, Q2 and Q3). Table 1 shows the retention
times of pesticides and their fragments quantitatively and qualita-
tively used in SIM method.
2.4. Method validation
The range of linearity and the calibration curves were estab-
lished by injecting a standard solution containing the 35 pesticides
in concentrations ranging from 0.02 to 0.75 mg kg1. The limit of
detection (LOD) and the limit of quantification (LOQ) were esti-
mated in the SIM mode analysis as the lowest concentration in-
jected that yielded S/N ratio of three and ten, respectively. The

Table 2
Linearity data (equation, and correlation coefficient, R2), LOD, LOQ and MRLs (mg kg1).

Pesticide Equation R2 LOD (mg kg1) LOQ (mg kg1) MRL (mg kg1)
ANVISA EU USEPA
Etridiazole Y = 4.66  104 + 3.18  106x 0.9910 0.010 0.030 N.A. 0.05 N.E.
Chloroneb Y = 1.25  105 + 5.80  106x 0.9946 0.010 0.030 N.E. N.E. N.A.
Molinate Y = 1.48  105 + 1.50  107x 0.9908 0.005 0.015 N.A. 0.05 N.A.
Propachlor Y = 2.49  105 + 1.27  107x 0.9924 0.010 0.030 N.E. 0.10 N.A.
Trifluralin Y = 4.22  103 + 4.19  106x 0.9946 0.010 0.030 N.A. 0.50 0.05
Hexachlorobenzene Y = 2.31  104 + 4.52  106x 0.9952 0.005 0.015 N.E 0.01 N.E.
Atrazine Y = 5.24  104 + 7.97  106x 0.9930 0.010 0.030 N.A. 0.05 N.A.
Chlorothalonil Y = 5.49  104 + 5.13  106x 0.9909 0.050 0.150 0.50 2.00 5.00
Parathion-methyl Y = 4.42  104 + 5.21  106x 0.9915 0.005 0.015 N.A. 0.02 N.A.
Ametryn Y = 2.06  105 + 1.07  107x 0.9931 0.010 0.030 N.A. N.E. N.A.
Metalaxyl Y = 1.34  105 + 2.48  106x 0.9904 0.050 0.150 0.10 0.20 1.00
Fenitrothion Y = 2.07  103 + 3.34  106x 0.9931 0.005 0.015 N.A. 0.01 N.A.
Malathion Y = 2.34  104 + 6.99  106x 0.9922 0.005 0.015 N.A. 0.02 8.00
Chlorpyrifos Y = 1.32  104 + 1.59  106x 0.9919 0.010 0.030 N.A. 0.05 N.A.
DCPA Y = 2.99  104 + 4.54  106x 0.9910 0.010 0.030 N.A. 1.00 1.00
Pendimethalin Y = 3.55  104 + 1.24  107x 0.9910 0.010 0.030 N.A. 0.05 0.10
Cis_Chlordane Y = 4.32  104 + 2.51  106x 0.9982 0.005 0.015 2.00 0.01 N.E.
Trans-Chlordane Y = 2.60  104 + 2.72  106x 0.9904 0.005 0.015 2.00 0.01 N.E.
Imazalil Y = 4.02  104 + 3.01  106x 0.9940 0.010 0.030 0.10 2.00 N.A
Buprofezin Y = 2.91  105 + 7.38  106x 0.9910 0.005 0.015 N.E 1.00 0.50
Kresoxim-methyl Y = 3.53  103 + 1.09  107x 0.9905 0.005 0.015 N.E. 0.20 0.40
Cyproconazole Y = 1.12  104 + 7.38  106x 0.9916 0.005 0.015 1.00 0.05 N.A
Chlorobenzilate Y = 1.74  105 + 1.01  107x 0.9918 0.005 0.015 0.30 0.02 N.E.
Triazophos Y = 4.59  104 + 4.87  106x 0.9927 0.050 0.150 0.10 0.01 N.E.
Bifenthrin Y = 6.12  104 + 2.02  107x 0.9921 0.005 0.015 0.01 0.05 0.40
Pyriproxyfen Y = 9.19  104 + 3.14  106x 0.9935 0.010 0.030 N.E. 0.05 0.10
Lambda-cyhalothrin Y = 4.97  103 + 3.29  106x 0.9932 0.010 0.030 N.A. 0.05 0.05
Fenarimol Y = 3.18  104 + 6.42  106x 0.9934 0.010 0.030 0.05 0.05 N.A.
Trans-permethrin Y = 3.17  104 + 7.31  106x 0.9916 0.010 0.030 0.05 0.05 1.50
Cis-permethrin Y = 2.62  105 + 1.78  107x 0.9906 0.010 0.030 0.10 0.05 1.50
Boscalid Y = 2.33  105 + 1.28  107x 0.9942 0.010 0.030 0.05 3.00 1.60
Pyraclostrobin Y = 2.77  104 + 4.17  106x 0.9948 0.010 0.030 N.A. 0.50 0.50
Deltamethrin Y = 3.86  104 + 1.23  106x 0.9904 0.050 0.150 N.A. 0.20 0.20
Azoxystrobin Y = 5.52  104 + 3.01  106x 0.9907 0.050 0.150 0.05 1.00 0.30
Imibenconazole Y = 1.81  104 + 2.28  106x 0.9937 0.050 0.150 0.10 N.E. N.E.

MRL – maximum residue limit; N.E. – not established; N.A. – not authorised for melon; ANVISA – Agência Nacional de Vigilância Sanitária (2012); EU – European Union (EU).
(2012); USEPA – United States Environmental Protection Agency (2013).
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2678 J. da Silva Sousa et al. / Food Chemistry 141 (2013) 2675–2681

accuracy and precision of the method were tested with recovery
experiments, performed with five replicates of blank samples
spiked with pesticides at 0.05, 0.10 and 0.20 mg kg1 and determi-
nation of relative standard deviation (RSD, %). The analytical qual-
ity control was maintained through the analysis of spiked sample
(0.1 mg kg1) simultaneously with the analysis of samples of mel-
on. Results were considered unsatisfactory if the spiked sample
presented a low recovery value.
The uncertainty was determined for all of the pesticides,
according to the procedures recommended by EURACHEM/CITAC
and Quantifying Uncertainty in Analytical Measurement (2000).
The uncertainty of measurement was obtained at the level of
0.05 mg kg1 using validation data. Three sources of uncertainty
were taken into account: uncertainty associated with precision
(u1), uncertainty associated with bias (u2) and uncertainty associ-
ated with the calibration curve (u3). The combined relative uncer-
tainty (Uc) was calculated using the expression:
1=2
Uc ¼ ðu21 þ u22 þ u23 Þ
The expanded uncertainty (Ue) was obtained by multiplying Uc by a
coverage factor k = 2 (confidence of 95%).
3. Results and discussion
3.1. Method validation
The QuEChERS method and te detection by ⁄GC-SQ/MS were
validated for the analysis of 35 pesticides in melon. Validation
parameters (selectivity, linearity, LOD, LOQ, accuracy and preci-
sion) were determined according the SANCO Guidelines (European
Commission, 2009).
3.1.1. Selectivity, linearity, LOD and LOQ
A standard matrix has been widely applied to avoid the
problems associated with raising or suppression of ionisation of
pesticides in chromatographic systems coupled to mass spectrom-
eters. Thus, the standard matrix was used for determining the lin-
earity range of pesticides studied. The linearity data (equation,
correlation coefficient), are shown in Table 2. The linearity was ob-
served in the range 0.02–0.75 mg kg1 and all correlation coeffi-
cients were >0.99.
Table 2 shows the LODs and LOQs obtained for each pesticide.
The LOD is the lower level of pesticide detected in instrument
(GC-SQ/MS) and the LOQ is considered the lowest level at which
the criteria for accuracy and precision has been reached. The LOD
and LOQ values obtained ranged from 0.005 to 0.050 mg kg1.
Most components presented LOQs values below LRM established
by USEPA, EU and ANVISA. However, some pesticides, such hexa-
chlorobenzene, Metalaxyl, Fenitrothion, cis-Chlordane, trans-
chlordane, Triazophos, Bifenthrin, imibenconazole presented val-
ues above those established by ANVISA and the EU, but all were
in accordance with the USEPA.
Lesueur, Knittl, Gartner, Mentler, and Fuerhacker (2008) ana-
lysed 105 pesticides with GC-SQ/MS after extraction with the QuE-
ChERS method in four matrices (grape, lemon, onion and
tomatoes); the LOD and LOQ values obtained ranged from 0.4 to
48.2 lg kg1 and from 1.2 to 161 lg kg1, respectively. Nguyen,
Yu, Lee, and Lee (2008) used QuEChERS and GC–MS for the deter-
mination of 107 pesticides in cabbage and radish, and obtained
LOQ values ranging from 0.002 to 0.1 mg kg1.
3.1.2. Accuracy, precision and uncertainty
The study of method recovery was carried out for three levels of
concentration (0.05, 0.1 and 0.2 mg kg1) by comparing the

Fig. 1. Total Ion Chromatogram (TIC) of (a) standard solution of 35 pesticides (0.5 mg kg1); (b) blank melon matrix and (c) sample of melon containing imazalil residue.
Identified compounds: (1) etridiazole, (2) chloroneb, (3) molinate, (4) propachlor, (5) trifluralin, (6) hexachlorobenzene, (7) atrazine, (8) chlorothalonil, (9) Parathion-methyl,
(10) ametryn, (11) metalaxyl, (12) fenitrothion, (13) malathion, (14) chlorpyrifos, (15) DCPA, (16) pendimethalin, (17) cis-chlordane, (18) trans-chlordane, (19) imazalil, (20)
buprofezin, (21) kresoxim-methyl, (22) cyproconazole, (23) chlorobenzilate, (24) triazophos, (25) bifenthrin, (26) pyriproxyfen, (27) lambda-cyhalothrin, (28) fenarimol, (29)
trans-permethrin, (30) cis-permethrin, (31) boscalid, (32) pyraclostrobin, (33) deltamethrin, (34) azoxystrobin, (35) imibenconazole.
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J. da Silva Sousa et al. / Food Chemistry 141 (2013) 2675–2681 2679

Table 3
Average recovery (n = 5), RSD (%), combined standard uncertainty (Uc) and expanded uncertainty (Ue) values.

Pesticide 0.05 mg kg1 0.10 mg kg1 0.2 mg kg1 Uc Ue

Recovery (%) RSD (%) Recovery (%) RSD (%) Recovery (%) RSD (%)
Etridiazole 68.8 8.2 65.8 6.4 63.6 6.2 7.4 14.7
Chloroneb 115.4 5.2 114.12 6.3 110.0 4.1 17.8 35.6
Molinate 112.6 10.1 98.1 8.1 105.4 9.4 6.8 13.6
Propachlor 114.4 6.8 103.1 4.2 108.5 5.5 5.9 11.7
Trifluralin 102.4 5.3 108.3 4.1 112.4 5.6 7.2 14.5
Hexachlorobenzene 98.9 4.1 85.7 5.8 92.8 5.3 8.9 17.8
Atrazine 103.7 5.4 89.3 4.6 108.2 4.1 9.3 18.5
Chlorothalonil 99.5 4.5 86.7 4.1 93.1 3.8 12.3 24.6
Parathion-methyl 109.8 4.6 101.8 4.4 108.4 4.8 10.1 20.2
Ametryn 115.3 8.1 102.9 6.7 111.3 6.4 8.9 17.9
Metalaxyl 94.8 6.5 106.7 5.1 106.8 5.5 18.1 36.1
Fenitrothion 110.2 5.0 102.3 3.7 107.5 4.6 6.5 13.1
Malathion 108.2 6.2 105.8 5.5 108.4 6.7 8.3 16.6
Chlorpyrifos 103.6 7.4 113.7 6.4 109.9 5.3 4.9 9.8
DCPA 104.6 7.3 96.1 4.1 114.8 3.9 6.3 12.5
Pendimethalin 109.5 6.1 103.8 4.9 116.9 6.3 7.6 15.2
Cis-Chlordane 107.6 8.2 95.7 7.8 105.4 8.1 8.2 16.4
Trans-Chlordane 113.2 4.1 102.8 4.8 117.9 3.8 7.2 14.5
Imazalil 106.4 10.7 110.3 11.2 115.5 12.5 9.8 19.6
Buprofezin 104.8 5.9 103.8 6.8 116.4 5.1 6.8 13.7
Kresoxim-methyl 115.1 5.3 109.6 6.2 116.5 5.7 5.8 11.6
Cyproconazole 108.5 6.9 94.7 4.8 98.9 7.1 11.2 22.4
Chlorobenzilate 116.5 6.2 109.3 5.3 112.7 5.8 7.5 15.0
Triazophos 108.6 5.4 102.8 5.2 115.1 6.7 8.8 17.6
Bifenthrin 107.8 4.6 102.3 5.8 110.4 4.6 7.1 14.3
Pyriproxyfen 108.7 6.7 88.7 6.3 97.9 5.1 7.9 15.8
Lambda-cyhalothrin 114.9 7.4 99.2 6.5 108.5 5.2 9.7 19.4
Fenarimol 109.6 6.2 112.8 4.4 101.6 6.7 8.6 17.3
Trans-permethrin 110.3 4.7 108.9 6.8 103.4 5.7 8.1 16.2
Cis-permethrin 114.1 4.8 107.6 5.5 102.2 6.2 8.0 16.0
Boscalid 111.8 10.4 102.8 8.7 114.9 9.1 8.2 16.4
Pyraclostrobin 114.3 12.2 102.5 10.8 103.8 10.6 4.6 9.1
Deltamethrin 109.7 10.3 112.5 9.5 105.1 7.2 4.3 8.6
Azoxystrobin 103.3 12.0 99.4 8.7 102.3 10.3 4.0 7.9
Imibenconazole 113.7 10.5 110.1 12.2 115.3 14.4 9.9 19.9

Uncertainty values estimated at the first level of concentration (0.05 mg kg1) and expressed as %.

concentration of each pesticide obtained after extraction and con-
centration in fortified matrix.
The method precision is expressed as the repeatability (RSD, %)
of the recovery determinations at the three different spiking levels
(0.05, 0.1 and 0.2 mg kg1). The recovery averages and the values
of relative standard deviations (RSD, %) of the individual pesticides
are shown in Table 3.
All pesticides studied (except etridiazole) showed recovery
rates ranging from 85% to 117% with a relative standard deviation
less than 15%. The SANCO Guidelines recommend recovery per-
centages of 70–120% and RSD 6 20% (European Commission,
2009). Only etridiazole not reached the established standards,
requiring modifications to the method.
The QuEChERS method was previously used by Barakat, Bada-
wy, Salama, Attallah, and Maatook (2007) for the extraction of 36
pesticides from honey samples, using the GC–ECD and GC–NPD
systems. A recovery percentage between 70% and 120% was ob-
tained, with an RSD 6 22%. Nguyen et al. (2008) achieved satisfac-
tory recoveries (80–115%) for 107 pesticides in cabbage and radish.
Hernández-Borges, Cabrera, Rodríguez-Delgado, Hernández-Suar-
ez, and Saúco (2009) using the method QuEChERS and GC–NPD
for the determination of 11 pesticides in banana obtained recovery
percentages between 67% and 118% with RSD 6 16%.
The uncertainty of measurement represents a quantitative indi-
cator of the reliability of the analytical results, expressed as a range
which is estimated to be the real value, usually associated with a
confidence level. In an analytical procedure, the uncertainty about
the result may arise from many possible sources, including sam-
pling, matrix effects, environmental conditions, glassware, method
of measurement and random variation. In this work, the combined
relative uncertainty (Uc) and expanded uncertainty (Ue) was
determined using repetitivity, recovery and calibration curves data
for each pesticide (Table 3). The Ue values (Table 3) ranged from
7.9% to 36.1%. Chloroneb and Metalaxyl showed higher levels of
uncertainty (>30%). The repeatability was the largest contribution
to the measurement uncertainty. Other sources, such as purity of
the reference standard feature small contribution in the
uncertainty values, not exerting considerable influence on the final
value.
3.2. Analyses of commercial melon samples
The developed method was used for monitoring and analysing
pesticides residues in melon samples, which were acquired at
three commercial establishments in Fortaleza, state of Ceara (Bra-
zil) during the period of 3 months and transported to the labora-
tory where they were analysed.
The results revealed the presence of bifenthrin and imazalil res-
idues in the melon samples. About 67% of the samples investigated
presented these residues. Fig. 1 shows the chromatogram of a sam-
ple of melon containing imazalil residue and the chromatogram of
the blank and the pesticides standard solution.
Imazalil and bifenthrin residues were detected in the 3 months
analysed, at the concentrations ranged from 0.030 to 0.100 and
0.005 to 0.010 mg kg1, respectively. Only in the melons provided
by S1, the pesticides were not detected. The pesticide imazalil was
detected during the 3 months monitored in the samples provided
by S2. All samples showed levels of imazalil below the MRL
 Author's personal copy

2680 J. da Silva Sousa et al. / Food Chemistry 141 (2013) 2675–2681

results showed the presence of the pesticide imazalil at levels be-

Fig. 2. Pesticide levels detected (mg kg1) in samples from Brazilian melons
analysed during the period of Oct/2011–Dec/2011.
Table 4
Imazalil and Bifenthrin levels detected in foods by other authors.
low the MRL established for orange (5 mg kg1).
Imazalil is a systemic fungicide used in Brazil on crops of bana-
na, citrus, apple, papaya, mango and melon. It is classified by ANVI-
SA as moderately toxic (Class III). The Acceptable Daily Intake (ADI)
for imazalil is 0.025 mg kg1 (PPDB – Pesticide Properties Data-
Base., 2011). According to the results of an individual who con-
sumes 400 g of fruit/day (recommended by WHO), with a level of
0.1 mg kg1 imazalil, a daily intake of 0.0007 mg kg1 is estimated
(assuming 60 kg b.w.), which is below the value of ADI.
Bifenthrin is a member of the pyrethroid chemical class. It is an
insecticide and acaricide, usually applied to control ants. ANVISA
has classified bifenthrin as highly toxic (Class II). It is highly toxic
to fish, aquatic organisms and mammals (PPDB, 2011). Large doses
may cause incoordination, tremor, salivation, vomiting, diarrhoea,
and irritability to sound and touch. The Acceptable Daily Intake
(ADI) for bifenthrin is 0.015 mg kg1 (PPDB, 2011).
The increased frequency of detection of pesticide residues in
food samples shows the need for pollution control by government
agencies to ensure food security for the population.

Method Pesticide Food Detected Country Refs.

Level
4. Conclusion
(mg kg1)
The QuEChERS method for the extraction of pesticides in sam-
GC– Bifenthrin Melon, 0.11 Spain Camino-
MS/ watermelon, Sánchez et al. ples of melon and subsequent quantification by GC-SQ/MS showed
MS cucumber, (2011) good selectivity, linearity and sensitivity for the 35 pesticides
tomato analysed, with precision and accuracy within the ranges recom-
GC–MS Imazalil Orange, 0.08– Croatia Knežević and
mended for most pesticides. Analysis of melon samples commer-
apple, 27.9 Serdar (2009)
peache, pear
cialised in Fortaleza-Ceara State showed the presence of
and pesticides, such as bifenthrin and imazalil at levels below the MRLs
grapefruit established by ANVISA, EU and USEPA. The validated method
HPLC/ Imazalil Apples 0.05–0.6 Spain Iñigo-Nuñez proved to be fast and effective, and provide reliable monitoring
UV et al. (2010)
of pesticide residues in melon. The method can be efficiently ap-
GC-TOF Bifenthrin Apples 0.03 Spain Cervera,
Portoles, plied to increase the scope of monitoring programs for various
Pitarch, types of foods, and thus improve food security.
Beltran, and
Hernandez Acknowledgements
(2012)
GC–MS Bifenthrin Mango 0.02–0.06 Brazil Menezes Filho
et al. (2010)
The authors gratefully the FUNCAP (Proc.: BFP-785.935.063-72)
LC– Imazalil Orange 0.04–0.64 Estonia Kruve et al. e CNPq (Proc.: 302300/2011-2) for financial support of this search
MS– (2007) and the NUTEC for providing the infrastructure of laboratories for
MS chromatographic analysis.
LC– Bifenthrin Fruits and 0.01–0.08 France Nougadère
MS– vegetables et al. (2011)
MS
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