Food Chemistry 245 (2018) 1018–1024

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

Analytical Methods

Silica/graphene oxide nanocomposites: Potential adsorbents for solid phase T

extraction of trace aflatoxins in cereal crops coupled with high performance
liquid chromatography
⁎ ⁎ ⁎
Li Yua,b,c,d,1, , Fei Maa,b,c,e,1, , Xiaoxia Dinga,d,e, Hengling Wanga,e, Peiwu Lia,b,c,d,e,
a
Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan 430062, PR China
b
Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China
c
Key Laboratory of Detection for Mycotoxins, Ministry of Agriculture, Wuhan 430062, PR China
d
Laboratory of Risk Assessment for Oilseeds Products, Ministry of Agriculture, Wuhan 430062, PR China
e
Quality Inspection and Test Center for Oil seeds and Products, Ministry of Agriculture, Wuhan 430062, PR China

A R T I C L E I N F O A B S T R A C T

Keywords: Graphene oxide wrapped silica nanocomposites were synthesized and selected as solid phase extraction ad-
Aflatoxins sorbents for high performance liquid chromatography analysis of aflatoxins. The major parameters affecting
Silica/graphene oxide nanocomposites extraction efficiency were optimized, including the amount of adsorbents, extraction time and desorption
Solid phase extraction conditions. The limit of detections and the limit of quantifications were from 0.1 to 0.3 µg/kg and from 0.3 to
High performance liquid chromatography
1.0 µg/kg, respectively. The recoveries of aflatoxins in the spiked maize and rice samples were in the range of
Cereal crops
76.8–104.7% and 81.1–106.9%, respectively, and with the RSDs less than 12.4%. The proposed method was
proven to be simple, rapid and reliable for routine analysis of aflatoxins in crops.

1. Introduction chromatography coupled with various detectors (La Barbera et al.,

Aflatoxins, which have been proven to be carcinogenic, teratogenic,
and mutagenic to humans, are a group of toxic metabolites produced by
Aspergillus flavus and Aspergillus parasiticus. Among these secondary
metabolites, aflatoxin B1 (AFB1) is the most common and toxic, and it
was classified as a group I carcinogen by the International Agency for
Research on Cancer (IARC) in 1993. These toxic metabolites could
contaminate agriculture products during harvest, storage, processing
and transportation, such as wheat, peanut, maize, rice and edible nuts
(Pereira, Fernandes, & Cunha, 2014). Serious health problem and eco-
nomic losses are caused by these toxic metabolites in Asia, Africa and
South America (Temba, Njobeh, & Kayitesi, 2017). Many countries have
established strict maximum residue limits (MRLs) of aflatoxins to con-
trol contamination in the food chain; in the European Union, the MRLs
are 2 µg/kg for B1 and 4 µg/kg for the total amount of all aflatoxins in
foodstuffs (EC, 2006, 2010). Therefore, it is important to develop rapid
and simple quantification method for monitoring and understanding
the occurrence of aflatoxins in cereal crops.
The determination of aflatoxins are achieved by various instru-
ments, including thin layer chromatography, enzyme-linked im-
munosorbent assays, antibody-based test strips, and liquid
2017; Li, Zhang, & Zhang, 2009; Li et al., 2013). Based on the gold
standard and versatility of aflatoxins analysis, high performance liquid
chromatography coupled with fluorescence detector (HPLC-FLD) is the
most widespread and routine method (Iqbal, Asi, Hanif, Zuber, & Jinap,
2016; Zhou, Xu, Huang, Cai, & Ren, 2017) . To achieve low limit de-
tection and good selectivity of aflatoxins in complex matrix, the pre-
concentration procedure is usually required prior to qualitative and
quantitative analysis.
Different extraction techniques have been used to enrich and purify
aflatoxins from crop samples, including liquid–liquid extraction, im-
munoaffinity chromatography and solid phase extraction (SPE)
(Lattanzio, Ciasca, Powers, & Visconti, 2014; Ok et al., 2016; Skendi,
Irakli, & Papageorgiou, 2016). Owing to the rapid, efficient, low cost
and less cross-contamination risks, the SPE method has been widely
used in control laboratories and proven to be one of the preferred
techniques for sample preparation in agri-products. Currently, Mycosep
columns, Supel Tox SPE cartridges and Isolute multi-mode SPE clean-up
column, are used for removing interferences associated with aflatoxin
analysis (Khayoon, Saad, Lee, & Salleh, 2012). However, analytes
cannot be concentrated simultaneously while the clean-up procedures
have been done. Faced with this challenge, C18, hydrophilic-lipophilic-

⁎
Corresponding authors at: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan 430062, PR China.
E-mail addresses: yuli01@caas.cn (L. Yu), mafeicpu@163.com (F. Ma), peiwuli@oilcrops.cn (P. Li).
1
These authors contributed equally to this study.

https://doi.org/10.1016/j.foodchem.2017.11.070
Received 29 May 2017; Received in revised form 16 November 2017; Accepted 17 November 2017
Available online 21 November 2017
0308-8146/ © 2017 Elsevier Ltd. All rights reserved.
L. Yu et al.
balanced (HLB) and Sep-Pak cartridges were investigated as antibody-
free adsorbents for aflatoxins (Chen & Zhang, 2013; Li et al., 2014).
Different types of polymeric materials have also been synthesized to
purify aflatoxins, including 3-(trimethoxysilyl)-1-propanthiol (TMSPT)
modified with 2-amino-5-mercapto-1,3,4-thiadiazole, ethylene glycol
bis-mercaptoacetate modified TMSPT grafted Fe3O4 nanoparticles and
poly dopamine-coated magnetic nanoparticles (Hashemi, Taherimaslak,
& Rashidi, 2014; Liu et al., 2013; McCullum, Tchounwou, Ding, Liao, &
Liu, 2014).
Traditional carbon materials, graphitized carbon black, and acti-
vated carbon were mainly used to clean up. Recently, carbon nanotube
and graphene oxide (GO), with high specific surface area and good
compatibility for organic and inorganic materials and solvents, play an
important role in sample extraction, especially as the adsorbents ap-
plied in the trace analysis of small molecular analytes (Es’haghi,
Sorayaei, Samadi, Masrournia, & Bakherad, 2011; Yu et al., 2013).
According to our previously reported work (Yu et al., 2013), GO could
be an effective adsorbents for aflatoxins. However, there were several
problems associated with using GO as adsorbents directly for enrich-
ment of aflatoxins. Although high-speed centrifugation was performed,
the existence of tiny sheets of GO could not be removed completely. To
avoid this problem, GO was bonded on silica nanoparticles as SPE ad-
sorbents for chlorophenols, polycyclic aromatic hydrocarbons and
heavy metals (Luo, Zhu, Li, Yuan, & Feng, 2013; Su, Chen, He, & Hu,
2014; Xu, Feng, Li, Liu, & Jiang, 2012). To the best of our knowledge,
GO-wrapped silica nanocomposites coupled with HPLC-FLD have not
been applied to analyze total aflatoxins in cereal crops.
The aim of this work was to synthesize GO-wrapped silica nano-
composites as solid phase adsorbents for the determination of aflatoxins
in cereal. The extraction parameters were optimized and successfully
applied to quantify analytes by HPLC-FLD.
2. Experimental
2.1. Chemicals and standards
N-hydroxysuccinimide (NHS), 1-ethyl-3-(3-dimethylaminopropyl)
carbodiimide hydrochloride (EDC), AFB1, AFB2, AFG1, and AFG2 stan-
dards were purchased from Sigma-Aldrich (St. Louis, MO, USA).
Methanol, acetonitrile and acetone of HPLC grade were obtained from
Anhui Fulltime Specialized Solvents & Reagents Co., Ltd. (Anqing,
China). Hydrochloric acid (HCl), n-hexane, and trifluoroacetic acid
were purchased from Tianjin Kermel Chemical Reagent Co., Ltd.
(Tianjin, China). Unless otherwise stated, all other inorganic chemicals
and organic solvents were of analytical reagent grade or better.
Ultrapure water (18 mΩ) was obtained from Milli-Q purification system
(Millipore Co., Ltd., Milford, MA, USA). Aminopropyl silica was sup-
plied by Sphere Scientific Co., Ltd. (Wuhan, China).
The stock solutions of AFB1, AFB2, AFG1 and AFG2 were prepared by
accurately weighing out 10 ± 1 mg of each standard and dissolving it
separately in 50 mL of methanol. A series of standard solutions were
prepared by diluting the stock solutions with methanol. All standard
solutions were sealed with parafilm, covered with aluminum foil and
stored in the dark at 4 °C until use. Certified reference material
(T04291QC Maize, FAPAS, UK) was used in the optimized procedures.
2.2. Instruments
The vacuum freeze drier was from Thermo Electron Corporation
(Rockford, Illinois, USA). The CF16RX high-speed freezing centrifuge
was from Hitachi (Tokyo, Japan). HPLC-FLD were performed on an
Agilent 1100 LC system (Agilent Tech., Palo Alto, USA). The ultrasonic-
assisted GO preparation was carried out by a KQ-800KDE ultrasonic
device (Kunshan Ultrasound Instrument Company, Kunshan, China).
1019
Food Chemistry 245 (2018) 1018–1024
2.3. Methods
2.3.1. Preparation of graphene oxide
According to previous GO synthesis procedure (Marcano et al.,
2010), 500 mg of GO was ultrasonicated in 1 L of water for 4 h at room
temperature. Then, the GO solution was centrifuged at 10,000 rpm for
10 min. The precipitate was discarded, and the unexfoliated GO sheets
could be removed from the GO solution. The final supernatant was
retained as GO precursor for the composite material.
2.3.2. Pretreatment of silica particles
To activate the amino groups and remove the free amino groups, the
pretreatment of aminopropyl silica particles was done before the
synthesis of silica/GO nanocomposites. Aminopropyl silica particles
(10.0 g) were swollen thoroughly in 100 mL of hydrochloric acid
(0.05 mol/L), successively washed with 100 mL of pure water 100 mL of
methanol and 100 mL of pure water 3 times, and the final products
(9.0 g) were collected by vacuum freeze drying for assembly and named
CK.
2.3.3. Synthesis and characterization of silica/GO nanocomposites
GO contains hydroxyl and carboxyl groups at the edge of the basal
plane (Dreyer, Park, Bielawski, & Ruoff, 2010). The hydroxyl group was
negatively charged and could be attracted to the positively charged
aminopropyl silica. Aminopropyl silica particles could be easily
wrapped in GO sheets by electrostatic interactions (Liu, Zhang, Chen, &
Wang, 2011a). 3.5 g of the previously prepared aminopropyl silica
particles were added into 120 mL of GO solution (0.5 mg/mL) and
gently shaken for 24 h at various temperatures (25 °C, 40 °C and 80 °C).
The silica particles were collected by centrifugation at 4500 rpm for
3 min and washed with deionized water several times until the super-
natant was clean. After vacuum freeze drying, the brown powders were
silica/GO nanocomposites RT, 40 and 80.
Owing to the presence of the carboxyl hydroxyl groups, GO could
also covalently bond with the aminopropyl silica particles by using EDC
and NHS as coupling agents (Liu et al., 2011b). 400 μL of 0.2 g/mL EDC
were added slowly into 160 mL of GO solution (0.5 mg/mL) with stir-
ring for 20 min. Then, 400 μL of NHS solution (0.1 g/mL) was added
into the mixture with continuous stirring for 1 h. Finally, 2.0 g of
aminopropyl silica particles were added into the same system and the
mixture was gently shaken for 24 h at room temperature. The brown
product was collected by centrifugation at 4500 rpm for 3 min and
washed with deionized water several times until the supernatant was
clean. After vacuum freeze drying, brown powders were obtained, and
were named High for short. Under the otherwise identical experimental
conditions, another two kinds of silica/GO nanocomposites were ob-
tained by reducing the added volumes of EDC and NHS solution to
200 μL and 40 μL, denoted as Mid and Low for short respectively. The
preparation scheme of silica/GO nanocomposites is shown in Fig. 1. The
microstructures of silica/GO nanocomposites were observed using
scanning electron microscopy (JSM-5610LV, 20 kV). The measurements
Fig. 1. Scheme for synthesis of silica/GO nanocomposites.
L. Yu et al.
were taken at room temperature.
2.3.4. Sample preparation
Rice and maize samples were purchased from local markets in
Wuhan, China. The rice and maize samples were ground until they
could pass through 0.850 mm. Two grams of the sample was weighted
into a centrifuge tube (polypropylene, 50 mL), homogenized in 10 mL
of methanol/water (70:30, v/v) for 3 min, and filtered; 5 mL of extract
was diluted to 28 mL with deionized water. Finally, the resulting aqu-
eous sample extract was filtered through 0.45 μm PTFE membrane and
stored in glass bottles in the dark at 4 °C.
2.3.5. Extraction procedure
To optimize the adsorbents, the adsorption efficiencies of as-syn-
thesized silica/GO nanocomposites (RT, 40, 80, High, Mid and Low)
and the aminopropyl silica particles (the original material without
loading GO, control blank, CK) for aflatoxins were studied. In the ad-
sorbent screening experiment, 2 g of spiked maize (spiked with 25 µg/
kg each of the four aflatoxins) was prepared and weighed. Then the
extract was prepared according to the method described in Section
2.3.4. A set of centrifuge tubes (polypropylene, 10 mL) contained 7 mL
sample of the prepared extract. To each tube, the same amount
(100 mg) of adsorbent material (RT, 40, 80, High, Mid, Low and CK)
was added. Each tube was vortexed for 30 s and shaken on a slow-
moving platform shaker for 30 min. Then, they were centrifuged at
4500 rpm for 2 min. Finally, the supernatant was collected from each
tube and the concentrations of aflatoxins in the supernatant were de-
termined by HPLC following the ISO16050:2003(E) method. The afla-
toxin adsorption efficiency (E) was calculated by using the equation
that follows, in which C0 and C1 represent the original and residual
concentrations of aflatoxins in the solution:
C0−C1
E= × 100%
C0 (1)
Extraction parameters were also optimized including the amount of
adsorbents (50 mg, 100 mg, 150 mg and 200 mg), extraction time
(5 min, 10 min, 20 min and 30 min), the desorption solvents (acetone,
acetonitrile and methanol) and the desorption volume (1 mL, 2 mL,
3 mL and 4 mL). Briefly, 2 g of maize reference material was prepared
and weighed. Then the extract was prepared according to the method
described in Section 2.3.4. A set of centrifuge tubes (polypropylene,
10 mL) contained 7 mL sample of the prepared extract. To each tube,
different amounts of adsorbent were respectively added. The mixture
was vortexed for 30 s and extracted by a slow-moving platform shaker
for different time. Then the tubes were centrifuged at 4500 rpm for
2 min. The supernatant was discarded. Subsequently, different type and
volume of desorption solvents were added into the tube and the mixture
was vortexed for 30 s. After centrifugation at 4500 rpm for 2 min, the
desorption solvent was collected and was dried with nitrogen at 60 °C,
after which it was derivatized with 100 µL of trifluoroacetic acid in
200 µL of n-hexane at 40 °C for 20 min. The reactant was dried with
nitrogen at 50 °C and dissolved in 0.1 mL acetonitrile/water mixture
(15:85, v/v) for HPLC-FLD analysis.
2.3.6. HPLC analysis
Chromatographic separation was performed using a C18 column
(4.6 mm × 150 mm, 5 μm) with mobile phase at a flow rate of 1.0 mL/
min and the column temperature was set at 40 °C. The mobile phase
consisted of A, acetonitrile, and B, water. The gradient elution program
was described in previous literature (Yu et al., 2013). The fluorescence
detection wavelength was set at λex/λem = 360/440 nm, and the in-
jection volume was 10 µL.
1020
Food Chemistry 245 (2018) 1018–1024
2.4. SPE HPLC-FLD method validation
2.4.1. Limit of detections and limit of quantifications
The limit of detections (LODs) and the limit of quantification (LOQs)
were measured as the minimum concentration of aflatoxins in the
spiked samples of rice and maize. The LODs and LOQs were defined as
the lowest detectable concentration with a signal-to-noise ratio (S/N) of
3 and 10, respectively.
2.4.2. Calibration curves and linear range
The calibration study was performed in triplicate using blank
maize/rice samples spiked at six different concentrations of AFG1 (1.0,
2.5, 5.0, 10, 15 and 20 µg/kg), AFB1 (0.5, 1.0, 2.5, 5.0. 10 and 20 µg/
kg), AFG2 (0.8, 1.25, 2.5, 5.0, 10 and 20 µg/kg) and AFB2 (0.3, 1.0, 2.5,
5.0, 10 and 20 µg/kg), respectively. All calibration curves were estab-
lished by plotting the average peak areas versus the corresponding
analytes concentration in the spiked samples.
2.4.3. Accuracy and precision
The accuracy of the SPE HPLC-FLD method was studied as trueness
(systematic error) and expressed in terms of the mean recovery. A batch
of 3 blank maize/rice samples were fortified with different amounts of
aflatoxins (1 µg/kg, 2 µg/kg and 5 µg/kg, respectively). After keeping in
the dark overnight at 25 °C, the spiked samples with complete eva-
poration of organic solvent and sufficient equilibration with cereal
matrix were prepared. Then, the spiked samples were applied to study
the recoveries of aflatoxins and the mean values of samples were cal-
culated.
The reproducibility of the SPE HPLC-FLD method was recorded as
the intra-day and inter-day precision detection of blank rice samples
spiked with aflatoxins at 10 µg/kg. The intra-day precisions were de-
termined by five parallel spiked rice samples within one day, and the
inter-day precisions were determined by four parallel spiked rice sam-
ples for four consecutive days.
2.4.4. Application to cereal samples
A total of 22 rice and 20 maize samples were collected from the
local markets (Wuhan, China). Under the optimized conditions, the
proposed method was applied to determine aflatoxins in the cereal
crops.
2.5. Statistical analysis
All the experiments were conducted in triplicate, and the average
values ± relative standard deviations (RSDs) were reported. The sta-
tistical analyzes were performed by using the @Risk 5.5.1 software
package (Palisade, Australia). The statistical difference was tested using
the Student t-test at a significance level of 0.05.
3. Results and discussion
3.1. Optimization of solid phase extraction adsorbents
To optimize the adsorbents, the adsorption efficiencies of as-syn-
thesized silica/GO nanocomposites (RT, 40, 80, High, Mid and Low)
and aminopropyl silica particles (the original material without loading
GO, CK) for aflatoxins were studied. As shown in Fig. 1s, the adsorption
of aflatoxins increased remarkably when the adsorbents varied from CK
to the silica/GO nanocomposites, which could be attributed to the in-
troduction of GO. Previous studies showed that GO was high efficiency
adsorbents for aflatoxins (Yu et al., 2013). Also, the adsorption effi-
ciencies of all silica/GO nanocomposites present high values (all higher
than 80%), indicating that varied amount of GO was coated on the
aminopropyl silica particles. When silica/GO nanocomposites (40) were
used as the adsorbents, aflatoxins could not be detected in the super-
natant and the adsorption efficiency reached almost 100%. Thus, silica/
L. Yu et al.
Fig. 2. Optimized extraction conditions: (a) effect of the adsorbent amount on the recoveries of aflatoxins, (b) effect of extraction time on the recoveries of aflatoxins, (c) effect of
desorption solvent on the recoveries of aflatoxins, and (d) effect of desorption volume on the recoveries of aflatoxins.
GO nanocomposites (40) were selected as the extraction adsorbent for
further studies.
The SEM images of the prepared silica/GO nanocomposites and
aminopropyl silica particles were shown in Fig. 2s. The size of the
prepared silica/GO nanocomposites were not changed basically after
loading with GO. The presence of wrinkles on the matrix surface could
be caused by the self-assembly of few layers of GO during the drying
process. In Fig. 2s (c and d), it showed that the surfaces of silica/GO
nanocomposites (40) were rougher than those of aminopropyl silica
particles, and for the nanocomposites, GO nano sheet-like structures
(wrinkles and thin grooves) could be observed, indicating that GO
sheets were attached to the surface of silica particles and enhances the
adsorption efficiency of GO.
3.2. Extraction optimization
The amount of adsorbents ranged from 50 mg to 200 mg in the
process of extracting aflatoxins. The results were presented in Fig. 2a,
illustrated that 100 mg of silica/GO adsorbents possessed the satisfac-
tory recoveries, and no increases were found from 100 to 200 mg. The
large surface afforded by silica/GO adsorbents facilitates strong ad-
sorption for π – π electrostatic interactions with the furan and coumarin
rings of the analytes. Thus, 100 mg was used in the SPE procedure.
The influence of extraction time was investigated, and the shaking
time was from 5 min to 30 min. Fig. 2b indicated that as the extraction
time increased from 5 to 10 min, the extraction recoveries of aflatoxin
G1 increased, while the other aflatoxins recoveries remained almost
constant (P < .05). Prolonging the extraction time did not increase the
extraction efficiency, thus the extraction time was fixed at 10 min.
1021
Food Chemistry 245 (2018) 1018–1024
The effect of desorption solvent was evaluated using different types
of solvents, and high desorption recoveries were achieved when acet-
onitrile was selected (Fig. 2c). This phenomenon could be attributed to
the π – π electrostatic interactions between lower unoccupied mole-
cular orbital and the higher occupied molecular orbital of the furan and
coumarin rings of aflatoxins (Hii, Basheer, & Lee, 2009). Therefore,
acetonitrile was adopted as the optimal desorption solvent. The deso-
rption volume was also investigated from 1 to 4 mL. As presented in
Fig. 2d, the results showed that the recoveries increased from 1 to 3 mL,
and no significant changes were observed when the volume was in-
creased. Consequently, 3 mL of acetonitrile was chosen in the following
experiments.
3.3. Analytical evaluation
The analytical parameters of the silica/GO SPE HPLC-FLD method
were studied under the optimized conditions. In Table 1, the correlation
coefficients (R2) of aflatoxins were ranging from 0.99651 to 0.99983.
The LODs and LOQs of aflatoxins were in the ranges 0.1–0.3 µg/kg and
0.3–1.0 µg/kg, respectively. The intra-day and inter-day of the relative
standard deviations (RSDs) were less than 3.9% and 6.4% respectively,
which indicated that this method could be used for routine analysis of
aflatoxins in cereals.
To evaluate the practicability and reliability of the method, the
proposed SPE HPLC-FLD method was used to analyze the aflatoxins in
maize and rice. The blank maize and rice samples were spiked at three
levels. The recoveries were 76.8–104.7% for aflatoxins in maize and
81.1–106.9% in rice sample, and with the RSDs less than 12.4%, which
were indicating good accuracy of the method.
L. Yu et al. Food Chemistry 245 (2018) 1018–1024

Table 2

Inter-day
Precision (RSD, %)b

(n = 4)
Results for the determination of aflatoxins in cereal samples.

6.4
3.2
2.5
3.9
Intra-day Sample Number of samples Analytes (µg/kg)a Detection rate %
(n = 5)
AFG1 AFB1 AFG2 AFB2

3.9
2.0
1.7
0.5
Rice 22 NDb ND ND ND –
(2.7) Maize 1 20 ND 4.1 ND ND 25%
(4.3)
(6.4)
(7.6)
Maize 4 ND 2.5 ND ND
5 µg/kg
Spiked

Maize 5 ND 3.8 ND ND
88.3
93.6
94.8
81.1 Maize 12 ND 1.8 ND 0.8
Maize 16 ND 1.2 ND ND
(6.8)
(4.8)
(5.1)
(6.9)
2 µg/kg
Spiked

a
Mean value of aflatoxins were analyzed in cereal samples.
94.6
95.1
93.4
97.5

b
Aflatoxins were no detect in samples.
106.9 (8.5)

3.4. Analysis of real samples

91.7 (5.2)
89.8 (8.2)
90.8 (5.7)
1 µg/kg
Spiked
Rice

The SPE HPLC-FLD method was applied to analyze aflatoxins in

cereal crops (22 rice and 20 maize). The results of determination of
(4.7)
(5.2)
(3.3)
(4.1)

aflatoxins in these samples were listed in Table 2. Only 25% of the

5 µg/kg
Spiked

maize samples contained AFB1 and AFB2, and the amounts of aflatoxins
Recovery (%) ± RSD (%, n = 3)a

96.4
95.7
95.3
76.8

were all under the MRLs set by EU and China. AFB1 was detected in five
maize samples, ranging from 1.2 µg/kg to 4.1 µg/kg, and AFB2 was
(12.4)
(7.5)
(9.4)
(6.7)

detected in only maize 12 (0.8 µg/kg). Typical HPLC chromatograms of

2 µg/kg
Spiked

aflatoxins analysis in a blank rice sample and a spiked maize sample

88.5
90.8
84.5
94.8

were provided in Fig. 3.

A comparative study of this developed method for aflatoxins ana-
100.9 (8.2)
104.7 (5.3)
95.3 (3.6)
96.8 (7.5)

lysis in cereal matrices to previously reported methods was performed,

1 µg/kg
Spiked
Maize

and the results were illustrated in Table 3 (Andrade, da Silva, & Caldas,
2013; Hashemi et al., 2014; Khayoon et al., 2012; Liu et al., 2013; Ma
et al., 2013). The developed method showed good sensitivity and ex-
(µg/kg)
LOQ

cellent recoveries, which could avoid the complicated and laborious

1.0
0.5
0.7
0.3

synthetic steps for magnetic solid phase adsorbents and polymers. The
LOQs in the developed SPE HPLC-FLD method were comparable with
(µg/kg)
LOD

HPLC-MS/MS for aflatoxins (Khayoon, Saad, Salleh, Manaf, & Latiff,

0.3
0.2
0.2
0.1

2014). Moreover, this is the first time that GO-wrapped silica nano-
composites were applied to extract aflatoxin in cereal crops, which
Y = 21.10335X + 3.66836 (0.99746)

Y = 18.36108X + 8.26801 (0.99949)

Rice matrix-matched calibration (R2)

Y = 3.52807X + 0.54905 (0.99976)

Y = 9.37904X + 0.76506 (0.99889)

shows great potential for sample preparation in complex matrix.

The precisions of aflatoxins were evaluated at 10 µg/kg in blank rice samples by calculating the RSDs.

4. Conclusions

Silica/GO nanocomposites were successfully applied for extraction

and quantification of aflatoxins in cereal. The strong adsorption prop-
erty made silica/GO nanocomposites an excellent candidate for SPE
Analytical performance data for the aflatoxins by the silica/GO SPE-HPLC method.

adsorbents. The silica/GO nanocomposites were synthesized at 40 °C by

The recovery of aflatoxins were evaluated in blank maize and rice samples.
Maize matrix-matched calibration (R2)

Y = 18.79886X + 10.04061 (0.99673)

Y = 21.66605X + 7.11175 (0.99651)
Y = 3.93708X + 0.68932 (0.99983)

Y = 9.66109X + 1.33041 (0.99956)

Linear range

1.0–20.0
0.5–20.0
0.8–20.0
0.3–20.0
(µg/kg)
Compound

Fig. 3. Typical chromatograms of aflatoxins analysis in maize (a) and rice (b) samples
Table 1

AFG1

AFG2
AFB1

AFB2

(dash line-unspiked samples; solid line-samples spiked with aflatoxins at the concentra-
b
a

tion of 2 µg/kg (rice) and 4 µg/kg (maize), respectively).

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L. Yu et al. Food Chemistry 245 (2018) 1018–1024

Table 3
Comparison of sample preparation procedures and recoveries among different methods.

Matrix Absorbents Sample pretreatment Derivatisation Determination Recovery (%) LOQs (µg/kg) Ref.
condition technique

Breast milk – Liquid–liquid extraction with Post-column HPLC-FLD 73.0–99.5 0.005–0.03 Andrade et al.
low temperature purification derivatisation (2013)
Agri-product mAbs-amino silica gel Immunoaffinity Pre-column HPLC-FLD 90.1–104.4 0.10–0.30 Ma et al.
microparticles chromatography cleanup derivatisation (2013)
Chilli, peanut ISOLUTE-multimode Solid phase extraction Pre-column HPLC-FLD 86.3–104.5 Peanut 0.23–0.58 Khayoon et al.
and rice absorbents derivatisation Rice 0.27–0.62 (2012)
Chilli 0.64–5.59
Coffee and malt C8 sorbents Porous membrane-protected – HPLC-MS/MS 86.0–109.5 Malt 0.40–1.11 Khayoon et al.
beverage micro-solid phase extraction Coffee 0.76–2.52 (2014)
Grains and grain Oasis HLB and Bond Elut Solid phase extraction Pre-column HPLC-FLD 77.9–111.6 Corn 0.13–0.18 Chen and
products cartridges derivatisation Wheat 0.11–0.15 Zhang (2013)
Cereal Hyperbranched Solid phase extraction – HPLC-FLD 82.7–103.0 0.04–0.40 Liu et al.
polymers (2013)
Cereal product AMT-TMSPT MNPs Magnetic solid phase – HPLC-FLD 90.0–97.0 Rice 0.215 (B1) & Hashemi et al.
nanoparticles extraction 0.046 (B2) (2014)
Corn 0.219 (B1) &
0.048 (B2)
Peanut Graphene oxide Solid phase extraction Pre-column HPLC-FLD 85.1–100.8 0.08–0.65 Yu et al.
adsorbents derivatisation (2013)
Cereal crops Silica/graphene oxide Solid phase extraction Pre-column HPLC-FLD 76.5–119.0 Rice 0.30–1.0 This work
nanocomposites derivatisation

electrostatic interactions. This is the first report about the character-
istics of silica/GO nanocomposites in extracting the aflatoxins in cereal
samples. Under the optimized conditions, the method for quantification
of aflatoxins in rice and maize samples was established. The validation
results indicated that the developed method was suitable for the de-
termination of aflatoxins in cereals. Owing to its simplicity, rapidity,
and economical nature, the proposed method represents a valuable
method for the routine analysis and survey of aflatoxins and its deri-
vatives in protein-based food.
Acknowledgments
This work was supported by the National Natural Science
Foundation of China (31601576, 31540048 & 31201447), the Natural
Science Foundation of Hubei Province – China (2016CFB612), the
Fundamental Research Funds for Central Non-profit Scientific
Institution, the National Key Research and Development Program of
China (2016YFF0201901) and the International Science & Technology
Cooperation Program of China (2016YFE0112900).
Conflict of interest
The authors declare no conflict of interest.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in the
online version, at http://dx.doi.org/10.1016/j.foodchem.2017.11.070.
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