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Degradation kinetics of aflatoxin B 1 and B 2 in solid medium by using pulsed

light irradiation

Article  in  Journal of the Science of Food and Agriculture · April 2018

DOI: 10.1002/jsfa.9058

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Research Article
Received: 6 October 2017 Revised: 31 January 2018 Accepted article published: 10 April 2018 Published online in Wiley Online Library:

(wileyonlinelibrary.com) DOI 10.1002/jsfa.9058

Degradation kinetics of aflatoxin B1 and B2 in

solid medium by using pulsed light irradiation
Bei Wang,a,b Noreen E Mahoney,c Ragab Khir,a,d Bengang Wu,b Cunshan
Zhou,b Zhongli Pana,c* and Haile Mab

Abstract
BACKGROUND: Pulsed light (PL) is a new potential technology to degrade aflatoxin. The objective of this study was to investigate
the degradation characters of aflatoxin B1 (AFB1 ) and B2 (AFB2 ) treated under PL irradiation. A kinetic degradation study of
AFB1 and AFB2 in solid medium was performed under PL irradiation at different initial concentrations of AFB1 (229.9, 30.7
and 17.8 𝝁g kg−1 ) and AFB2 (248.2, 32.2 and 19.5 𝝁g kg−1 ) and irradiation intensities (2.86, 1.60 and 0.93 W cm−2 ) of PL. A
second-order reaction model was applied to describe degradation of AFB1 and AFB2 .

RESULTS: The results showed that the degradation of AFB1 and AFB2 followed the second-order reaction kinetic model well
(R2 > 0.97). The degradation rate was proportional to the intensities of PL irradiation and the initial concentrations of aflatoxins.

CONCLUSION: It is concluded that the degradation of AFB1 and AFB2 with the use of PL could be accurately described using the
second-order reaction kinetic model.
© 2018 Society of Chemical Industry

Keywords: pulsed light; aflatoxin; photodegradation; kinetics; second-order reaction

INTRODUCTION
According to an FAO report, each year 25% crops are contami-
nated by mytoxins,1 among which the most harmful is aflatoxin.
Aflatoxin B1 and B2 (AFB1 and AFB2 ) are a group of highly toxic,
mutagenic and carcinogenic compounds produced by Aspergillus
flavus and Aspergillus parasiticus. The International Agency for
Research on Cancer (IARC) has classified AFB1 in Group 1 as a
human carcinogen.2 Consumption of aflatoxin-contaminated food
causes serious diseases in humans. Both acute and chronic toxicity
can result from aflatoxin accumulation in the body, causing acute
liver damage, liver cirrhosis, tumors and teratogenic effects.3 To
solve this problem, numerous methods, including chemical, phys-
ical and biological, have been studied to eliminate aflatoxins.4–6
However, most of them have limitations, such as low degradation
efficiency, chemical (alkaline) residue or the creation of undesir-
able residual by-products. Therefore, there is a pressing need to
investigate alternative methods that can effectively degrade afla-
toxins.
Pulsed light (PL) is an FDA-approved technology for food
decontamination.7 PL is a technique to decontaminate the sur-
face of food by killing microorganisms or degrading chemicals
using short-duration pulses of an intense broad-spectrum light.
Researchers reported that PL could produce high performance
and low secondary pollution to degrade mycotoxins such as AFB,
zearalenone, deoxynivalenol and ochratoxin.8,9 Moreau et al.8
reported that eight flashes of PL (1 J cm−2 per pulse) degraded
AFB1 solution (5 𝜇g mL−1 ) by 93%. Our previous study revealed
that PL treatment achieved an effective degradation of AFB1 and
AFB2 in rice bran and rough rice.9 The results showed that after
15 s of PL treatment at an intensity of 0.52 J cm−2 per pulse, 90.3%
and 86.7% of AFB1 and AFB2 were reduced in rice bran, respec-
tively. Under the same conditions, 80 s of treatment achieved
75.0% and 39.2% of AFB1 and AFB2 reduction in rough rice. The
high-intensity flashlight created by PL could destruct the structure
of mycotoxins. Research proved that PL irradiation could destroy
the structures of AFB and degrade it into harmless substances.
The treated samples did not show cytotoxicity or mutagenicity.8,9
However, the characteristics of aflatoxins degraded by PL irra-
diation have rarely been reported. To the best of the authors’
knowledge, no kinetic study has been performed on solid sam-
ples, although the aflatoxin exists widely in solid media. In fact,
most of the photodegradation kinetic studies are on liquid media.
The objectives of this research were therefore to (i) evaluate the
effectiveness of PL treatment on the degradation of AFB1 and AFB2
under different concentrations and intensities, and (ii) study the
characteristics of the degradation kinetics of AFB1 and AFB2 in solid
medium.
∗ Correspondence to: Z Pan, Healthy Processed Foods Research Unit,
USDA-ARS-WRRC, Albany, CA 94710, USA. E-mail: zhongli.pan@ars.usda.gov;
or E-mail: zlpan@ucdavis.edu
a Department of Biological and Agricultural Engineering, University of California,
Davis, CA, USA
b School of Food and Biological Engineering, Jiangsu University, Zhenjiang,
China
c Healthy Processed Foods Research Unit, USDA-ARS-WRRC, Albany, CA, USA
d Department of Agricultural Engineering, Faculty of Agriculture, Suez Canal
University, Ismailia, Egypt

J Sci Food Agric (2018) www.soci.org © 2018 Society of Chemical Industry

 www.soci.org
Figure 1. Molar absorption spectra of AFB1 and AFB2 and emission spectra
of PL lamp.
MATERIALS AND METHODS
Chemicals
AFB1 (2,3,6𝛼,9𝛼-tetrahydro-4-methoxycyclopenta[c]furo[3′ ,2′ :4,5]
furo[2,3-h]chromene-1,11-dione; purity > 98%) and AFB2
(2,3,6𝛼,8,9,9𝛼-hexahydro-4-methoxy-cyclopenta[c]furo[3′ ,2′ :4,5]
furo[2,3-h] [1]benxopyran-1,11-dione; purity > 98%), analytical-
grade acetonitrile, benzene and methanol were purchased from
Sigma-Aldrich (St Louis, MO, USA).
Standard stock solutions (1 𝜇g mL−1 ) of AFB1 and AFB2 were
separately prepared in benzene–acetonitrile 98:2 (v/v) according
to the protocol of AOAC10 and stored in the dark at 4 ∘ C. The
stability of the stock solution was checked every 3 months. Three
concentrations of AFB1 and AFB2 were prepared according to
the description of Murata et al.11 Immediately before use, 0.50,
0.25 and 0.10 mL of AFB1 and AFB2 stock solutions (1 𝜇g mL−1 )
were absorbed on 1 × 3 cm filter paper strips (Grade 1, Whatman
International Inc., Maidstone, UK) and placed in glass Petri dishes,
respectively. The Petri dishes with lids partly covered, were placed
in a fume hood for 5 min to evaporate the solvent.
Pulsed light treatment
The SteriPule-XL® 3000 sterilization system (Xenon Corp., Wilm-
ington, MA, USA) was used as the PL generator for conducting
this study. A detail schematic diagram of the system was rep-
resented in our published paper.12 The system generated three
pulses per second of polychromatic light in the wavelength range
of 100–1100 nm at an input voltage of 3800 V. The PL fluence has
overlapping AFB1 and AFB2 absorbance spectra with the afore-
mentioned wavelengths (Fig. 1). The emission spectrum of PL was
provided by the manufacturer. The irradiated energy to the sam-
ples could be adjusted by changing the distance between the
samples to the lamp. As per the manufacturer’s specifications, the
energy produced at distances of 10.8, 14.8 and 22.8 cm from lamp
to sample were 2.86, 1.60 and 0.93 W cm−2 .
Before treatment, the dried strips absorbing different concen-
trations of AFB1 and AFB2 were placed in glass Petri dishes with
lids off. The Petri dishes were then exposed under PL at distances
of 10.8, 14.8 and 22.8 cm to achieve intensities of 2.86, 1.60 and
0.93 W cm−2 for different durations of 1, 2, 4, 6, 8, 10 and 12 s,
wileyonlinelibrary.com/jsfa © 2018 Society of Chemical Industry
B Wang et al.
respectively. The temperature was controlled at ambient tempera-
ture (25 ± 2 ∘ C). After treatment, each strip was vortexed with 2 mL
methanol to collect the residual aflatoxins in preparation for anal-
ysis by high-performance liquid chromatography (HPLC).
HPLC aflatoxin assay
The residual aflatoxin after treatment were analyzed by
reversed-phase HPLC (Agilent 1100, Santa Clara, CA). Condi-
tions for HPLC analyses (Inertsil ODS-3, 4.6 × 250 mm) were:
mobile phase, water–acetonitrile–methanol (45:25:30, v/v/v);
flow 1.0 mL min−1 ; temperature 25 ∘ C; fluorescence detector
at excitation wavelength of 365 nm and emission wavelength
455 nm; derivatization using photochemical reactor (PHRED, Aura
Industries), 25 m × 0.25 mm i.d. coil; injection volume 20 𝜇L; and
retention times for AFB2 of 9.4 min and AFB1 of 10.6 min. All
aflatoxin experiments were performed in triplicate.
Kinetic modeling
The degradation ratio of aflatoxin was calculated using following
equation: [( ) ]
E = C0 − Ct ∕C0 × 100% (1)
where E is the degradation ratio of aflatoxin (%); C 0 is the initial afla-
toxin concentration (𝜇g kg−1 ) and C t is the aflatoxin concentration
(𝜇g kg−1 ) at time t (s).
The reaction rate equation of the photodegradation of aflatoxin
can be expressed as follows:13
r = −dC∕dt = kapp C n (2)
where r is the reaction rate (𝜇g kg−1 s), C is the concentration of
aflatoxin (𝜇g kg−1 ), t is time (s), kapp is the apparent reaction rate
constant and n is the reaction order.
Using integral method for linear fitting, for the second-order
kinetics, n = 2, Eqn (2) can be rearranged as follows:
1∕Ct − 1∕C0 = k2 t (3)
where C t is the concentration (𝜇g kg−1 ) at time t, C 0 is the initial
concentration (𝜇g kg−1 ) and k2 is the second-order reaction rate
constants (kg 𝜇g −1 s−1 ).
Statistical analysis
Linear regression analysis was carried out using GraphPad Prism
5.0 (GraphPad Software, Inc., USA). Statistical analyses were con-
ducted using the SPSS 19.0 software (SPSS, Chicago, IL, USA). A
one-way analysis of variance (ANOVA) was conducted with signifi-
cance level of 0.05 to compare the results of different groups.
RESULTS AND DISCUSSION
Effect of PL on degradation of AFB1 and AFB2
The effect of PL irradiation time on the degradation of AFB1
and AFB2 is presented in Fig. 2. In general, at the beginning of
the treatment, the degradation rate was high. After only 1 s of
treatment at an intensity of 2.86 W cm−2 , 61.2 ± 7.5% of AFB1 and
57.3 ± 8.3% of AFB2 were degraded, respectively. At the beginning
of the treatment, high reactant concentration led to high efficiency
of the reaction. When the treatment time was prolonged, the
degradation rate decreased. This was due to the reduction of
the reactant concentration and the production of intermediate
J Sci Food Agric (2018)
Degradation kinetics of aflatoxins in solid medium using pulsed light irradiation
Figure 2. Degradation curves of AFB1 and AFB2 under different PL intensities and initial concentrations.
products that absorbed the light energy. After 12 s of PL irradiation,
AFB1 and AFB2 were reduced by 96.6 ± 4.8% and 91.7 ± 2.3%,
respectively. No further significant reduction was obtained due to
the low degree of penetration of the PL or a shadow effect of the
filter papers.14–16
The results illustrated that PL treatment was a promising poten-
tial technology in rapidly degrading aflatoxin. The commonly used
method to degrade pollution is short-wave ultraviolet (UV-C), cur-
rently. However, UV-C has much lower degradation efficiency.
Murata et al.11 treated 30 mg kg−1 zearalenone (ZEN) and deoxyni-
valenol (DON) in filter paper with 0.1 and 24 mW cm−2 of mild
and strong UV-C, respectively. The level of ZEN and DON became
undetectable at mild intensity after 60 min of treatment and at
strong intensity after 15 min and 20 min of treatment, respectively.
This illustrated that intensity plays an important role in myco-
toxin degradation. PL produces high-intensity light of very short
duration. The energy the photons carry would greatly accelerate
the process of the reaction.17 Therefore, the reaction rate of pho-
todegradation produced by PL was much higher than that pro-
duced by the UV-C.
Second-order reaction kinetic models
Photodegradation by PL of aflatoxin is a complex dynamic process.
In this study, a second-order reaction kinetic model was applied
to regress aflatoxin degradation under PL irradiation. Plots of 1/C t
− 1/C 0 versus irradiation time for AFB1 and AFB2 at different PL
intensities and initial concentrations are shown in Fig. 3 and the
estimated parameters are presented in Table 1. It can be seen that
the second-order reaction kinetic model fitted well. In Fig. 3, the
degradation curves show clear trends with initial concentrations
and intensities. Table 1 shows a high average R2 value of 0.97 for
the second-order reaction kinetic model. By comparing the R2 ,
root mean square error (RMSE) and P-values, it could be concluded
J Sci Food Agric (2018) © 2018 Society of Chemical Industry
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that the second-order reaction kinetic model was a good fit for
describing the degradation of AFB1 and AFB2 .
Effect of PL intensity on degradation of AFB
Degradations of AFB1 and AFB2 under different intensities are
presented in Fig. 2. AFB1 and AFB2 began with the highest ini-
tial concentrations of 229.9 and 248.2 𝜇g kg−1 , respectively. As
Table 1 reveals, higher PL power intensities showed higher degra-
dation rate of AFB1 and AFB2 . Figure 3 shows that there is a
strong correlation between regressed curves and PL intensity.
These findings revealed that, with decreased intensity, the pho-
todegradation rate decreased, which is in agreement with Liu
et al.18,19 Their results showed the degradation rate had a pos-
itive relationship with the intensity of UV-C irradiation. In gen-
eral, increasing intensity of PL increases the energy of the emit-
ted light and more photons could be produced. Thus the reac-
tion rate could be accelerated by high-energy photons. In the
present study, all half-life values were shorter than 5 s. This is
much shorter than the half-life times obtained by many reports
using continuous UV or visible light.18–21 The results illustrated
that the degradation rate had a positive relationship with the
intensity of PL.
Effect of initial concentration on degradation of AFB1 and AFB2
The effect of initial concentration on the degradation of AFB1
and AFB2 by PL irradiation was investigated at an intensity of
2.86 W cm−2 at a constant temperature of 25 ± 2 ∘ C. The results
are shown in Fig. 2. It can be seen that the high initial concen-
tration led to high degradation rates for both AFB1 and AFB2 .
Table 1 shows that a higher initial concentration corresponds to a
shorter half-life t1/2 . These findings revealed that with decreased
initial aflatoxin concentration the degradation rate decreased.
In contrast to UV treatment, Jing et al.22 reported that when
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Figure 3. Second-order reaction kinetic model regression of AFB1 and AFB2 under different PL intensities and initial concentrations.

Table 1. Second-order kinetic of AFB1 and AFB2 degradation under different conditions

Sample Concentration and intensity Equation R2 RMSE P-value k2 (kg 𝜇g−1 s−1 ) t1/2 (s)

AFB1 0.92 J cm−2 per pulse 1/C t − 1/C 0 = 0.0067 t 0.9885 0.0033 <0.0001 0.0067 ± 0.0002 0.64
0.52 J cm−2 per pulse 1/C t − 1/C 0 = 0.0053 t 0.9885 0.0033 <0.0001 0.0053 ± 0.0002 0.81
0.30 J cm−2 per pulse 1/C t − 1/C 0 = 0.0030 t 0.9840 0.0021 <0.0001 0.0030 ± 0.0001 1.46
229.9 𝜇g kg−1 1/C t − 1/C 0 = 0.0067 t 0.9885 0.0033 <0.0001 0.0067 ± 0.0002 0.64
30.7 𝜇g kg−1 1/C t − 1/C 0 = 0.0183 t 0.9704 0.0150 <0.0001 0.0183 ± 0.0008 1.74
17.8 𝜇g kg−1 1/C t − 1/C 0 = 0.0196 t 0.9252 0.0248 <0.0001 0.0174 ± 0.0013 2.85

AFB2 0.92 J cm−2 per pulse 1/C t − 1/C 0 = 0.0048 t 0.9691 0.0037 <0.0001 0.0048 ± 0.0023 0.86
0.52 J cm−2 per pulse 1/C t − 1/C 0 = 0.0030 t 0.9556 0.0040 <0.0001 0.0030 ± 0.0002 1.38
0.30 J cm−2 per pulse 1/C t − 1/C 0 = 0.0021 t 0.9888 0.0014 <0.0001 0.0021 ± 0.0001 1.91
248.2 𝜇g kg−1 1/C t − 1/C 0 = 0.0048 t 0.9691 0.0037 <0.0001 0.0048 ± 0.0023 0.86
32.2 𝜇g kg−1 1/C t − 1/C 0 = 0.0432 t 0.9716 0.0434 <0.0001 0.0432 ± 0.0023 0.72
17.8 𝜇g kg−1 1/C t − 1/C 0 = 0.0479 t 0.9492 0.0673 <0.0001 0.0479 ± 0.0035 1.17

naphthalene is treated in seawater by UV irradiation, the average
reaction rate constant at high concentration was slightly lower
than that at low concentration. Conversely, Liu et al.18 studied the
effect of initial concentration on degradation of AFB1 in peanut
oil using continuous UV irradiation and found that initial con-
centration was not related to k, which is a feature of first-order
reaction kinetics.
Generally, under constant temperature, the photoreaction rate
mainly depended on the concentration of reactants and increased
with increased concentration of the reactants.3 According to the
collision theory, the rate of a photoreaction is directly proportional
to the number of effective collisions per second between the reac-
tant molecules. However, the medium of the reaction influences
the reaction rate. Most of the previous photodegradation studies
were carried out in a liquid environment. Cooper et al.23 reported
that in aqueous medium UV could produce active particles such as
hydrated electrons, single-state oxygen, peroxy radical, hydroxyl
radical and hydrogen peroxide that promoted the degradation of
organisms. Jing et al.22 explained that when the initial concentra-
tion of sample was low the number of reactive radicals generated
from UV irradiation in water was sufficient to trigger the photore-
actions. In contrast, if the initial concentration was much higher,
using the same number of radicals may not be sufficient to sustain
the reaction at the desired rate. Therefore, in aqueous medium, the
photodegradation rate showed a reverse relationship with the ini-
tial concentration. Conversely, in Liu’s research,18 the photodegra-
dation occurred in peanut oil, where fewer active particles were
produced by UV than that in water. In the primary photochemi-
cal reaction, a photon was an active reactant molecule. Therefore,
the reaction rate depended on the absorption rate of photons that
were produced by the UV light, regardless of the initial concentra-
tion of reactants.

wileyonlinelibrary.com/jsfa © 2018 Society of Chemical Industry J Sci Food Agric (2018)

Degradation kinetics of aflatoxins in solid medium using pulsed light irradiation
In the current study, photodegradation was performed in solid
stable medium. The reaction came to completion in a couple of
seconds. The short time frame might suggest that not enough
active particles were produced to influence the degradation.
Meanwhile, overproduction of photons may result during PL
irradiation. The reaction was dependent on the concentration
of aflatoxins. Therefore, the obtained results indicated that the
degradation rate under PL irradiation had a positive relation-
ship with initial concentrations of AFB1 and AFB2 . The reac-
tion rate was directly related to the reaction medium and the
number of photons.
CONCLUSIONS
The photodegradation behavior of AFB1 and AFB2 in solid medium
has been investigated in this study. PL irradiation as a new tech-
nology could effectively degrade AFB1 and AFB2 in seconds. The
degradation of AFB1 and AFB2 in filter paper has been proved to
follow second-order reaction kinetics well. The power intensity
and initial concentration had a positive relationship with the reac-
tion rate. This study would help predict the final content of AFB1
and AFB2 after PL treatment and provide important information
for reducing the risk of aflatoxin contamination in foods and agri-
cultural products.
ACKNOWLEDGEMENTS
The research was conducted at UC Davis and WRRC, USDA-ARS.
The authors thank the National Natural Science Foundation of
China for Youth Scholars (No. 31601516), Jiangsu Province Nat-
ural Science Foundation for Youth Scholars (No. BK20150499),
National Key Research and Development Program of China
(2016YFD0400705-04) and Scientific Research Foundation
Research for Advanced Talents of Jiangsu University (No.
16JDG048) for grant support.
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