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Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in greenhouse chrysanthemum
Journal Pre-proof Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in greenhouse chrysanthemum Wenwen Gong, Mengyun Jiang, Tingting Zhang, Wei Zhang, Gang Liang, Bingru Li, Bin Hu, Ping Han PII:
S0269-7491(19)33362-7
DOI:
https://doi.org/10.1016/j.envpol.2019.113499
Reference:
ENPO 113499
To appear in:
Environmental Pollution
Received Date: 12 July 2019 Revised Date:
25 September 2019
Accepted Date: 25 October 2019
Please cite this article as: Gong, W., Jiang, M., Zhang, T., Zhang, W., Liang, G., Li, B., Hu, B., Han, P., Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in greenhouse chrysanthemum, Environmental Pollution (2019), doi: https://doi.org/10.1016/j.envpol.2019.113499. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.
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Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in
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greenhouse chrysanthemum
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Wenwen Gonga,*, Mengyun Jianga,b, Tingting Zhangb, Wei Zhangc, Gang Lianga, Bingru Lia, Bin
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Hud, Ping Hana
5
a
Beijing Research Center for Agriculture Standards and Testing, Beijing 100097, China
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b
College of Chemical Engineering, Beijing University of Chemical Technology, Beijing 100029,
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China
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c
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48824, USA
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d
Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI
Beijing Plant Protection Station, Beijing 100029, China
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*Corresponding author.
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Dr. Wenwen Gong, E-mail: [email protected]
1
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ABSTRACT
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Production of chrysanthemum (Dendranthema grandiflora) in greenhouses often requires
16
intensive pesticide use, which raises serious concerns over food safety and human health. This study
17
investigated uptake, translocation and residue dissipation of typical fungicides (metalaxyl-M and
18
fludioxonil) and insecticides (cyantraniliprole and thiamethoxam) in greenhouse chrysanthemum
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when applied in soils. Chrysanthemum plants could absorb these pesticides from soils via roots to
20
various degrees, and bioconcentration factors (BCFLS) were positively correlated with lipophilicity
21
(log Kow) of pesticides. Highly lipophilic fludioxonil (log Kow = 4.12) had the greatest BCFLS (2.96 ±
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0.41 g g-1), whereas hydrophilic thiamethoxam (log Kow = -0.13) had the lowest (0.09 ± 0.03 g g-1).
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Translocation factors (TF) from roots to shoots followed the order of TFleaf > TFstem > TFflower.
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Metalaxyl-M and cyantraniliprole with medium lipophilicity (log Kow of 1.71 and 2.02, respectively)
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and hydrophilic thiamethoxam showed relatively strong translocation potentials with TF values in
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the range of 0.29–0.81, 0.36–2.74 and 0.30–1.03, respectively. Dissipation kinetics in
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chrysanthemum flowers followed the first-order with a half-life of 21.7, 5.5, 10.0 or 8.2 days for
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metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively. Final residues of these
29
four pesticides, including clothianidin (a primary toxic metabolite of thiamethoxam), in all
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chrysanthemum flower samples were below the maximum residue limit (MRL) values 21 days after
31
two soil applications each at the recommended dose (i.e., 3.2, 2.1, 4.3 and 4.3 kg ha-1, respectively).
32
However, when doubling the recommended dose, the metabolite clothianidin remained at
33
concentrations greater than the MRL, despite that thiamethoxam concentration was lower than the
34
MRL value. This study provided valuable insights on the uptake and residues of metalaxyl-M,
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fludioxonil, cyantraniliprole and thiamethoxam (including its metabolite clothianidin) in greenhouse
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chrysanthemum production, and could help better assess food safety risks of chrysanthemum
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contamination by parent pesticides and their metabolites.
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Keywords：Chrysanthemum, Pesticide, Uptake, Translocation, Greenhouse, Residue dissipation 2
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1. Introduction
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Chrysanthemum (Dendranthema grandiflora) is widely grown in China. More than 26 billion
42
chrysanthemum flower heads were sold in China in 2016 with approximately $2.2 billion in sales [1].
43
It is commercially valuable as both cut flowers and potted plants. Chrysanthemum is also used in
44
food supplement, herbal tea, and medicine [2]. Its flower head is a popular traditional medicine due
45
to its unique flavor, color and health benefits [3] and is listed in Chinese Pharmacopoeia as
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Chrysanthemi Flos [4]. Previous studies have indicated that chrysanthemum tea has calming,
47
heat-clearing and detoxifying effects and can help reduce physical and mental stress [5, 6].
48
Chrysanthemum flowers are most often consumed via tea infusion, which is prepared from extraction
49
of chrysanthemum flowers with hot water by consumers at home or sold on the market as
50
ready-to-drink tea [7, 8]. Thus, certain pesticide residues in contaminated tea or herbs (including
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chrysanthemum flowers) may be transferred to tea infusion, which may then pose health risks to tea
52
drinkers [8-10].
53
Chrysanthemum is often cultivated in greenhouses for protection against inclement weather.
54
However, high humidity and warm temperature in greenhouses may cause severe infestations by
55
insects (e.g., aphids and larvae of various Lepidoptera species), fungi and oomycetes [11]. Pest
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management with pesticides is often used to reduce crop loss from insects and plant diseases, but
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also exposes chrysanthemum crops to potential pesticide residues [12]. When pesticides are used in
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greenhouses, their residual concentrations in soils and plants may be greater than those in open fields,
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largely due to unique greenhouse conditions such as inadequate ventilation and high humidity that
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could affect pesticide dissipation (e.g., vaporization of volatile/semi-volatile active ingredient)
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[12-14]. However, knowledge on dissipation behaviors of pesticides during chrysanthemum
62
production in greenhouses is very limited.
63
Dissipation behaviors of pesticides is dependent on methods of pesticide application. Foliar
64
application of pesticides is the most widely used treatment in field and greenhouse conditions, but 3
65
often results in several negative consequences, such as off-target drift, killing pests’ natural enemies,
66
secondary pest outbreaks, as well as potential exposure by farmer workers [15]. In contrast,
67
application of pesticides to soils via irrigation water (i.e., chemigation) is an alternative that could
68
avoid or reduce adverse impact mentioned above [15, 16]. Soil application can be effective not only
69
for soilborne pests and pathogens, but also for protecting the upper part of plants through uptake and
70
translocation of active pesticide ingredients to plant shoots via transpiration stream [17]. Thus, soil
71
application is becoming increasingly popular. In addition, previous studies indicated that
72
combination formulation of several pesticides can be more effective than single ingredient
73
formulation. Therefore, combination pesticides are often applied to crops [18]. However, the uptake
74
and residue dissipation of mixed pesticides in chrysanthemum plants and soils after soil application
75
have rarely been studied.
76
Therefore, this study aimed to investigate the uptake, translocation and dissipation of two typical
77
fungicides (metalaxyl-M and fludioxonil) and two insecticides (cyantraniliprole and thiamethoxam)
78
in chrysanthemum grown in greenhouses after pesticide application to soils at the
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manufacturer-recommended dose and double of the recommended dose. A modified quick, easy,
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cheap, effective, rugged and safe (QuEChERS) extraction method was established to extract
81
pesticides from chrysanthemum root, stem, leaf and flower samples and soil samples, and the
82
extracted pesticides were then analyzed with UPLC-MS/MS. In addition, a main metabolite of
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thiamethoxam (clothianidin) in chrysanthemum was identified and measured throughout the
84
experiment as its analytical-grade standard was commercially available.
85
2. Materials and methods
86
2.1. Reagents
87
Standards of metalaxyl-M (purity of 99.5%), fludioxonil (purity of 99.0%), cyantraniliprole
88
(purity of 99.2%), thiamethoxam (purity of 98.2%) and its metabolite clothianidin (purity of 99.0%)
89
were purchased from China Standard Material Center (Beijing, China). Properties of tested pesticides 4
90
are shown in Supplemental Table S1. A commercial formulation of 62.5 g L-1 metalaxyl-M
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fludioxonil SC (37.5 g L-1 for metalaxyl-M and 25 g L-1 for fludioxonil), and a commercial
92
formulation of 40% cyantraniliprole thiamethoxam SC (20% for each) were obtained from
93
Syngenta (Beijing, China). HPLC-grade acetonitrile and formic acid were purchased from
94
Sigma-Aldrich (St. Louis, MO, USA) and used as received. Analytical reagent-grade sodium
95
chloride (NaCl) and magnesium sulfate (MgSO4) were purchased from Sinopharm Chemical
96
Reagent (Beijing, China) and heated for 12 h at 120 °C to remove residual water before use. Primary
97
secondary amine (PAS, 40 µm), graphitized carbon black (GCB) and C18 were purchased from
98
Agela Technologies Ltd. (Tianjin, China) and used without pretreatment.
99
2.2. Greenhouse experiments
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Greenhouse experiments were performed in a greenhouse at Beijing Plant Protection Station,
101
Beijing, China from September to November 2018. The greenhouse was under natural lighting and
102
maintained at average temperature of 21 ± 2 °C and relative humidity of 75 ± 2%. Chrysanthemum
103
plants were cultivated in plastic pots (500 × 340 × 263 mm) each filled with dry soil mass of 6 ± 0.5
104
kg by experienced gardeners. The soil had 2.6% of sand, 86.4% of silt and 11.0% of clay, and was
105
classified as silt as per the USDA classification. The soil had an organic matter of 262.3 g kg-1, a
106
cation exchange capacity of 38.1 cmol kg-1 and a pH of 7.1. A randomized block design
107
(Supplemental Table S2) was used where two adjacent block were separated by a 0.5-m alley.
108
Experiment A was designed to investigate the uptake, translocation and dissipation kinetics of
109
metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam, and clothianidin in chrysanthemum plants.
110
Chrysanthemum plants in 120 pots (four plants per pot) at the flower bud initiation stage received a
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mixed pesticide solution (200 mL) once at the manufacturer-recommended dose (3.2, and 2.1, 4.3
112
and 4.3 kg ha-1 for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively; low
113
dose). Chrysanthemum samples were collected from three random pots taken at 0 (about 2 h), 1, 2, 3,
114
5, 7, 14 and 21 days after pesticide application. Meanwhile, 50 pesticide-free Chrysanthemum pots 5
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(Experiment C) received pure water of the same volume. Samples were collected before the pesticide
116
application and then at harvest.
117
Experiment B was conducted to study final pesticide residues in chrysanthemum flowers.
118
Chrysanthemum plants in 50 pots (four plants per pot) received a mixed solution of pesticides at the
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low dose or the high dose (i.e., doubling the recommended dose) two or three times with a 7-day
120
interval. Chrysanthemum flowers from three randomly selected pots in each treatment were collected
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at pre-harvest interval (PHI) of 7, 14 and 21 days after the last soil application. Extra pots were
122
included and not all planted pots were used in the experiments.
123
2.3. Sample collection, extraction and analysis
124
The chrysanthemum samples were divided into four parts, including flowers, leaves, stems and
125
roots. Chrysanthemum roots were rinsed in acetonitrile under shaking for 20 s followed by washing
126
with distilled water to remove pesticides residue on root surface [15]. Then, each portion of
127
chrysanthemum was separately cut into small pieces and homogenized using a blender
128
(BUCHI/B-400, China) for 1–2 min until a completely homogenized sample was obtained. All
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samples were transported to the laboratory and stored in a freezer at −20 °C until extraction. The
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QuEChERS method modified from Golge and Kabak [19] was used for extracting the
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chrysanthemum samples. The choices of extraction solvents (acetonitrile, acetone or ethyl acetate)
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and clean-up reagents were optimized by measuring recoveries of spiked pesticides.. Acetonitrile was
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shown as the most suitable solvent to extract the tested pesticides from chrysanthemum matrix. A
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combination of 20, 30, or 50 mg of PSA and C18 (or GCB) as well as 100 mg of anhydrous MgSO4
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were tested as clean-up reagents. Satisfactory recoveries were obtained by the combination of 20 mg
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PSA, 20 mg GCB and 100 mg anhydrous MgSO4 for leaf and flower samples and 50 mg PSA, 20 mg
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GCB and 150 mg anhydrous MgSO4 for root and stem samples, respectively. The QuEChERS
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method is detailed as follows: 5 g of each flower or leaf sample were added to a 50-mL centrifuge
139
tube with 10 mL acetonitrile and shaken for 20 min, followed by adding 2 g of anhydrous MgSO4 6
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and 1 g of NaCl. The mixture was vortexed for 1 min, and then centrifuged at 8000 rpm for 5 min.
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Afterwards, 1 mL of the supernatant was transferred to a 2-mL clean-up tube containing 20 mg PSA,
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20 mg GCB and 150 mg anhydrous MgSO4. After mixing, the tube was centrifuged at 10000 rpm for
143
10 min. The supernatant was collected again and filtered through 0.22-µm Whatman GF/C glass
144
fibre filters for further analysis. In addition, 2 g of stem and root samples were extracted using the
145
abovementioned procedure except that each clean-up tube had 50 mg PSA and 150 mg anhydrous
146
MgSO4.
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Soil samples were thoroughly mixed and passed through a 2-mm sieve. Subsequently, 5 g soil
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samples were weighed into the 50-mL centrifuge tube. The extraction and clean-up procedures were
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the same with the abovementioned protocol, except that each clean-up tube contained 50 mg PSA, 50
150
mg C18 and 150 mg anhydrous MgSO4.
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The analyses of metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam and clothianidin
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concentrations in extracted samples were carried out with a Waters ACQUITY UPLC-MS/MS
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system equipped with an ACQUITY BEH C18 column (2.1 mm × 100 mm × 5 µm) (Milford, MA,
154
USA). The main metabolite of thiamethoxam (clothianidin) was analyzed as we had access to its
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analytical-grade standard. Other metabolites were not included because they were either not
156
identified or no analytical-grade standards were available. Full metabolite identification and
157
quantification were out of scope for this study and could be explored in future studies. Detailed
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information regarding the optimized UPLC-MS/MS condition is provided in Supporting information
159
(SI). Analysis was performed in multiple reaction monitoring (MRM) mode, with operational
160
parameters shown in Supplemental Table S3.
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2.4. Data analysis
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To estimate accumulation potential of organic compounds from soils into plant tissues,
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bioconcentration factors (BCFs) are calculated as the ratios of concentrations in plant tissues to that
164
in soils and have been widely used as indexes in studying plant uptake of organic contaminants [20, 7
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21]. However, some studies have suggested that BCFs based on concentrations in bulk soils (BCFsoil)
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may not accurately represent the uptake potential because chemical concentrations in bulk soils
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could not directly reflect the bioavailability of chemicals for plant uptake [22, 23]. In addition, the
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calculation of BCFsoil is generally based on an assumption that chemical concentrations in soils
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remain relatively constant [24, 25]. However, a significant decrease of pesticide concentrations in
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soils (approximately by 28% to 43%) was observed during chrysanthemum cultivation in this study.
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To overcome the large variation (uncertainty) of pesticide concentrations in bulk soils, BCFs based
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on the loss of pesticides in soils (BCFLS) were previously proposed by Sun et al. [14]. We assume
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that pesticides are first released from soils to soil pore water, their losses due to leaching,
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volatilization, biodegradation, or plant root uptake. Therefore, the total decrease of pesticide
175
concentrations in soils represent the total amount of pesticides available for plant uptake. Thus, root
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BCFLS (g g-1) was calculated using Eq. (1). = ⁄
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(1)
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where Croot (mg kg-1) is the pesticide concentration in roots, and Closs in soil (mg kg-1) is the decrease of
179
pesticide concentration in soils, i.e., the maximum concentrations of pesticides in soils subtracted by
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the concentrations of pesticides in soils measured at time t (14 and 21 days in this study).
181 182
To better evaluate translocation potential of measured pesticides from chrysanthemum roots onto shoot (stem, leaf, and flower), translocation factors (TF) were calculated as follows.
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TF = /
(2)
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TF = /
(3)
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TF = /
(4)
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where Cstem, Cleaf, and Cflower (mg kg-1) are the pesticide concentration in stem, leaf and flower,
187
respectively.
188
As the pesticide residues in chrysanthemum flowers are key to health risk assessment, the
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dissipation kinetics of measured pesticides in chrysanthemum flowers were fitted to the first-order 8
190
kinetic equation (Eq. 5). The dissipation half-life (t1/2) was calculated via Eq. (6).
191
=
(5)
192
/ = 2/"
(6)
193
where k (day-1) is the dissipation rate constant, Ct (mg kg-1) and C0 (mg kg-1) are the pesticide
194
concentration at a given time t and at the beginning. In this study, the pesticide concentration in
195
chrysanthemum flowers initially increased for a few days (1 to 3 days) after the final pesticide
196
application (Supplemental Table S2). Therefore, the peak concentration was used as the initial point
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for fitting the dissipation kinetics [12].
198
The uptake, translocation and dissipation kinetics of target pesticides in chrysanthemum plants
199
and linear regression analyses between log BCFLS with log Kow values of pesticides were analyzed
200
with the Origin software 9.0 (OriginLab Inc., Northampton, MA, USA). One-way analysis of
201
variance (ANOVA) was performed to test significant difference in means between sample groups
202
using SPSS v20.0 (IBM, New York, NY, USA). According to Tukey’s multiple comparison tests, p <
203
0.05 and p < 0.01 were considered as significant and highly significant, respectively. Correlation
204
analyses were performed using Pearson’s test with SPSS v20.0, and correlation was considered
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statistically significant when p value was less than 0.01.
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2.5. Quality assurance and quality control (QA/QC)
207
The QA/QC of pesticide analysis included determination of linearity, recovery, precision and
208
accuracy. Linearity was assessed with matrix-matched standards in the range of 0.5 µg L-1–0.2 mg
209
L-1 for chrysanthemum (including flower, leaf, stem and root) and soil samples. The correlation
210
coefficients (R2) of matrix-matched calibration curves were all greater than 0.991, demonstrating a
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good linearity (Supplemental Table S4). Recovery and precision tests were conducted using blank
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chrysanthemum samples (flower, leaf, stem and root) fortified with target compounds at 0.005, 0.05
213
and 0.2 mg kg-1 levels or blank soil samples fortified at 0.002, 0.05 and 0.2 mg kg-1 levels with five
214
replicates. The average recoveries of the fortified samples ranged from 78% to 114%, with relative 9
215
standard deviation (SD) varying between 0.05% to 13.06% (Supplemental Table S5). In all cases, the
216
recovery results were acceptable and thus confirmed that the analytical method was reliable for
217
analyzing metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam and its metabolite clothianidin.
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The limit of quantification (LOQ) was determined as the lowest concentration of a given analyst at
219
which an acceptable recovery (70–120%) and a relative SD of ≤ 20% can be achieved [17]. The
220
LOQs of five pesticides were 0.005 mg kg-1 in chrysanthemum tissues and 0.002 mg kg-1 in the soils,
221
based on the analysis of five replicates.
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3. Results
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3.1 Uptake and bioaccumulation from soils into chrysanthemum roots
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The four pesticides were detected in chrysanthemum roots at 0.63–2.28 mg kg−1 on day 14 (the
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pre-anthesis stage) and at 0.46–1.33 mg kg−1 on day 21 (the anthesis stage,) after the last soil
226
application (Experiment A, Supplemental Table S6), whereas none was found in the roots for the
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blank control (Experiment C). Thus, chrysanthemum roots could absorb and accumulate these
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pesticides from soils. Fludioxonil exhibited the highest concentrations of 2.28 and 1.33 mg kg-1 on
229
day 14 and 21, respectively, which was up to 3.6 times of the concentrations of other three pesticides
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(Supplemental Table S6). The calculated BCFLS values for each pesticide are shown in Fig. 1A. The
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BCFLS values ranged between 0.15–2.96 g g-1 at day 14, and between 0.09–2.59 g g-1 at day 21.
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Among all the pesticides, fludioxonil demonstrated the highest BCFLS values, whereas thiamethoxam
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had the lowest BCFLS values (p < 0.01). There was a strong positive correlation between log BCFLS
234
and log Kow (i.e., octanol-water partition coefficient, Table S1), as shown in Fig. 1B (R2 = 0.92, p <
235
0.01).
236
3.2 Root-to-shoot translocation
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In this study, the residue concentrations of metalaxyl-M, fludioxonil, cyantraniliprole, and
238
thiamethoxam in different parts of chrysanthemum (flower, stem and leaf) over the testing period 10
239
were measured and shown in Fig. 2. The concentrations of all the measured pesticides were much
240
higher in chrysanthemum leaves than in stem, and were the lowest in flowers. The pesticides also
241
remained longer in leaves than in stems and flowers (Fig. 2). Fludioxonil increased in
242
chrysanthemum leaves during the entire cultivation period but were much less than the other three
243
pesticides. The residue concentrations of metalaxyl-M, cyantraniliprole, and thiamethoxam increased
244
and reached their maximum concentrations in leaves 7 days after the last application, and then
245
gradually decreased.
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The average TF values of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam are shown in
247
Fig. 3. The TFstem, TFleaf and TFflower values of four pesticides during the cultivation period were in
248
the range of 0.01–2.22, 0.07–3.88, 0.002–0.77, respectively (Supplemental Table S7). Interestingly,
249
the TF values initially increased after pesticide applications (1–5 days) and then decreased over time.
250
Generally, TF of each tissue for all pesticides followed the order of TFleaf > TFstem > TFflower (p <
251
0.05), suggesting that chrysanthemum leaves are the main reservoir for absorbed pesticides. 3.3
252
Dissipation kinetics in chrysanthemum flowers
253
Normally, the residues of four pesticides in chrysanthemum flowers increased from 1 to 3 days
254
after the last application, and then decreased gradually thereafter (Fig. 2). Their dissipation
255
percentages in chrysanthemum flowers 21 days after the last application were 49.0%, 90.9%, 72.4%
256
and 81.6%, respectively. The dissipation of each pesticide in chrysanthemum flowers was
257
satisfactorily fitted with a first-order kinetic equation, with R2 ranging from 0.744 to 0.943 (Table 1).
258
The calculated half-life values t1/2 in chrysanthemum flowers were 21.7, 5.5, 10.0 and 8.2 days for
259
metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively.
260
In this study, clothianidin was detected in chrysanthemum flowers with the application of
261
thiamethoxam. Clearly, thiamethoxam was metabolized to clothianidin, resulting in a considerable
262
concentration of clothianidin in chrysanthemum plants. The residues of clothianidin increased with
263
the degradation of thiamethoxam (Fig. 2). Clothianidin concentration reached a maximum level in 11
264
chrysanthemum flowers (0.06 mg kg-1) 5 days after the last application and then declined eventually
265
to 0.02 mg kg-1. The dissipation of clothianidin residues was fitted to a first-order kinetic equation
266
Ct=0.075 e-0.061t (R2 = 0.975). The calculated half-life t1/2 value of clothianidin was 11.4 days,
267
suggesting that the dissipation rate of thiamethoxam (t1/2 = 8.2 days) in chrysanthemum flowers was
268
greater than that of its metabolite clothianidin.
269
3.4 Final residues
270
The final residues of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in
271
chrysanthemum flowers on day 7, 14 and 21 after the last application are presented in Table 2. Their
272
residues in chrysanthemum flowers were 0.05–0.38, <0.005–0.11, 0.04–0.27 and 0.09–0.35 mg kg-1,
273
respectively, after 2–3 times of application at the recommended dose. When applied at a higher dose,
274
their final residues were increased to varying degrees. Greater concentrations were observed in the
275
treatments with more applications and shorter pre-harvest intervals.
276
4. Discussion
277
Our studies showed that chrysanthemum root could uptake metalaxyl-M, fludioxonil,
278
cyantraniliprole, and thiamethoxam from soils to different degrees (Fig. 1A and Supplemental Table
279
S6 ), and the calculated BCFLS values were related positively to log Kow of pesticides (Fig. 1B). This
280
observation was in a good agreement with other studies [26, 27]. Thus, partitioning was the dominant
281
mechanism controlling the root uptake of these nonionic organic compounds [28]. Recently, several
282
studies have found ion trap effect for ionizable organic compounds [23, 29, 30], which would not be
283
applicable to non-ionic compounds that do not change their charge speciation as a function of
284
solution pH. In addition, molecular weight is regarded as an important factor for plant uptake,
285
probably due to its effect on the passage through cell membrane or Casparian strip [31-34]. For
286
example, Li et al. [23] investigated the uptake of 15 chemicals with a wide range of molecular
287
weight by radish roots, and found that most of the chemicals with molecular weight <500 g mol-1 12
288
showed a greater bioconcentration than those with molecular weight > 500 g mol-1. For the pesticides
289
with low molecular weight (Table S1), no correlation was observed between BCFLS and molecular
290
weight, suggesting that there may be threshold molecular weight below which plant uptake was not
291
influenced by molecular weight. In addition to physicochemical properties of chemicals, other
292
factors such as crop species, cultivation methods and environmental conditions may collectively
293
affect the uptake and bioconcentration of chemicals [35]. For example, studies have showed that
294
crops could absorb and concentrate relatively more organic pollutants (e.g., clarithromycin and
295
sulfadiazine) when grown in hydroponic systems than in soils [36-38].
296
For most plants, pesticides applied through chemigation to soils could be absorbed by plant roots
297
and then translocated to plant shoots through transpiration stream [15, 17, 39-41]. Overall, the
298
concentrations of pesticides in chrysanthemum leaves took some time to reach the maximum, were
299
dissipated slowly and could remain in the plant for a long time (Fig. 2). In stems, the pesticide
300
residues researched their maximum earlier than in leaves, but remained at a largely stable and low
301
level throughout the experimental period. These results were consistent with previous studies,
302
reporting that the stems more likely serve as a transport channel with little retention of pesticides
303
[17]. Several studies suggested that deposition of gaseous pesticides volatilized from soils or
304
deposition of resuspended soil particles on aerial parts of plants also contributed to the accumulation
305
of pesticides in plant shoots [14, 42]. However, none of the pesticides was detected in the shoots of
306
chrysanthemum planted in pesticide-free soils in the same greenhouse, indicating that these two
307
pathways were negligible for the accumulation of pesticides with low vapor pressure (Supplemental
308
Table S1) in chrysanthemum shoots.
309
TF values could reveal the tendency of pesticides to be translocated into a component of the
310
shoot (stem, leaf, and flower) from the root. Our results showed that the TF values initially increased
311
after pesticide applications and then decreased over time (Table S7). Similar results were also
312
observed for the translocation and accumulation of imidacloprid in six leafy vegetables [43] and five 13
313
neonicotinoids in komatsuna [44]. In addition, the TF values of each tissue (0.07–0.17) of highly
314
lipophilic fludioxonil (log Kow = 4.12) were significantly lower than the other three pesticides,
315
indicating that it was mainly accumulated in chrysanthemum roots. In contrast, metalaxyl-M and
316
cyantraniliprole with medium lipophilicity and hydrophilicity (log Kow of 1.71 and 2.02, respectively)
317
and thiamethoxam with low lipophilicity (log Kow = −0.13) showed relatively stronger translocation
318
potentials with the TF values in the range of 0.29–0.81, 0.36–2.74 and 0.30–1.03, respectively. It was
319
previously suggested that there is an optimum lipophilicity of organic compounds for their
320
translocation. Highly hydrophilic compounds may have difficulty to cross the hydrophobic root
321
membranes, whereas highly lipophilic compounds on the other hand are concentrated in roots by
322
binding with root tissues (e.g., lipids) and are less likely to be translocated upward to shoots [27, 45].
323
Therefore, it is possible that a highly lipophilic pesticide (e.g., fludioxonil) can have the highest root
324
bioconcentration, but the lowest translocation to the shoots (Fig. 1 and Fig. 3), suggesting minor
325
health risks from consuming chrysanthemum flowers.
326
Previous studies suggested that the dissipation pattern of pesticides varies with plant species,
327
dose and interval of pesticide application, and climatic conditions [46, 47]. The half-life values of
328
pesticides such as pyrethrins and dechlorane plus in plants or soils under open field conditions were
329
generally less than that under greenhouse conditions, due to factors such as rainfall, solar radiation
330
and crop growth rate, etc. [14, 48]. In addition to physical and chemical factors, such as light,
331
temperature, pH, and moisture, pesticide application methods (e.g., foliar spray, soil application of
332
chemigation, or seed treatment) might also play an important role in the dissipation of pesticides in
333
plants. For example, the observed half-life of thiamethoxam in chrysanthemum flowers in this study
334
was much greater than that reported in teas (1.6 days) [49] and tobacco leaves (3.9–4.4 days) [50] by
335
foliar spraying in the field.
336
The final residues of metalaxyl-M, fludioxonil, and thiamethoxam in chrysanthemum flowers
337
were below the MRL values (Table 2) according to the MRLs for metalaxyl-M (3 mg kg-1), 14
338
fludioxonil (20 mg kg-1) and thiamethoxam (1.5 mg kg-1) in herbs or edible flowers established by
339
the UK/EC MRL Database [51]. However, the final residues of metalaxyl-M, fludioxonil, and
340
thiamethoxam in chrysanthemum flowers were below the MRL values. However, the final residues
341
of cyantraniliprole were greater than the MRL value (0.05 mg kg-1) except for the residue
342
concentrations at the 14 and 21 days after two applications at the low dose. Normally, the MRL value
343
of a hazardous chemical is established based on its acceptable daily intake (ADI). On the basis of the
344
toxicological and ecotoxicological data, the World Health Organization (WHO) and the Food and
345
Agriculture Organization of the United Nations (FAO) established the ADI of cyantraniliprole at 0.03
346
mg kg-1 bw, which is much lower than that of metalaxyl-M, fludioxonil and thiamethoxam (0.08, 0.4,
347
and 0.08 mg kg-1 bw, respectively) [52]. Therefore, cyantraniliprole has much lower MRL, and is
348
thus more likely to exceed the MRL than other pesticides.
349
It has been suggested that organic compounds accumulated in plants may be metabolized, and
350
thus, further influence their dissipation pattern in plants [23, 53]. It should be noted that in some
351
cases metabolites can even be more toxic or at higher residue levels than parent pesticides [54, 55].
352
Therefore, there is increasing concern over the residues of some toxicologically significant
353
metabolites that remain in harvested crops and can then be ingested by humans and animals via food
354
or feed. Uptake of pesticide metabolites by chrysanthemum roots was not quantified in this study,
355
because we did not identify all the metabolites and it is also difficult to distinguish whether the
356
metabolites originate from in-plant metabolism or the uptake from soils [56]. However, we identified
357
and measured clothianidin, a main metabolite of thiamethoxam, in chrysanthemum as its
358
analytical-grade standard was commercially available. The final residue levels of clothianidin in
359
chrysanthemum flowers ranged from 0.045 to 0.36 mg kg-1 (Table 2) and were greater than the MRL
360
for clothianidin in flowers (0.05 mg kg-1) listed in the UK/EC MRL database except for its
361
concentration at 14 and 21 days after 2 applications and at 21 days after 3 applications at the low
362
dose. In contrast, the residues of parent pesticide thiamethoxam in all samples were lower than the 15
363
MRL values, indicating that risks may be underestimated if only thiamethoxam was measured.
364
Therefore, when assessing the food safety and human health risks of thiamethoxam application, it
365
would not be adequate to measure only the residue concentrations of thiamethoxam, and the
366
metabolite (clothianidin) should be included as well. Finally, when applied at a higher dose or at a
367
low dose with more frequent applications during chrysanthemum clutivation, the final residues of
368
tested pesticides were likely to exceed the MRL (Table 2), which warrants close attention to resultant
369
risks to food safety.
370
5. Conclusion
371
The findings of this study have several important implications to the food safety and quality of
372
chrysanthemum production in greenhouses. Metalaxyl-M, fludioxonil, cyantraniliprole and
373
thiamethoxam had varying potential to be absorbed by chrysanthemum roots, and then translocated
374
to shoots (stems, leaves and flowers), following their soil application. Pesticide residues in
375
chrysanthemum flowers increased at the first few days after application, and then decreased
376
gradually thereafter, followed the first-order kinetics. Moreover, clothianidin, a primary metabolite of
377
thiamethoxam, was detected in both chrysanthemum roots and shoots throughout the experiment.
378
The final residues of these four pesticides and clothianidin in chrysanthemum flowers were
379
significantly affected by the application times, doses and preharvest intervals. The final residues of
380
metalaxyl-M, fludioxonil, and thiamethoxam after 2–3 applications at either low dose or high dose
381
were blow the MRL in chrysanthemum flowers, suggesting the lower food safety risks of these
382
parent pesticide compounds. However, cyantraniliprole exceeded the MRL more often than other
383
pesticides, suggesting that close attention should be paid to its residues in chrysanthemum flowers.
384
Furthermore, the main metabolite of thiamethoxam (clothianidin) had the final residues greater than
385
the MRL in most treatments, highlighting the need to monitoring the concentrations of toxic
386
metabolites in addition to parent compounds. This study was limited in the sense that only one
387
metabolite was included because other metabolites could not be identified or their analytical-grade 16
388
standards were not available to us. Thus, future study should be directed to characterizing and
389
quantifying toxic metabolites of parent pesticide compounds. This study provided useful data for
390
assessing the safe use of typical fungicides and insecticides by soil application in greenhouse
391
chrysanthemum cultivation. Future work should also investigate the transfer of pesticide residues
392
from chrysanthemums flowers into tea infusion from a food safety and human health perspective.
393
Acknowledgments
394
This work was financially supported by National Natural Science Foundation of China
395
(21806014), Special Projects of Construction of Science and Technology Innovation Ability of
396
Beijing Academy of Agriculture and Forestry Sciences (KJCX20190405; KJCX20170419), and
397
Project of Beijing Excellent Talents (2017000020060G127).
398
17
399
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Table 1. Dissipation kinetic equation, correlation coefficient and half-life of target pesticides in chrysanthemum flowers.a
a
Pesticides
Regression equation
Correlation coefficient (R2)
Half-life (t1/2)
Metalaxyl-M
Ct=0.148 e-0.032t
0.835
21.7
Fludioxonil
Ct=0.046 e-0.127t
0.762
5.5
Cyantraniliprole
Ct=0.096 e-0.069t
0.744
10.0
Thiamethoxam
Ct=0.256 e-0.085t
0.943
8.2
Clothianidin
Ct=0.075 e-0.061t
0.975
11.4
To investigate the dissipation kinetics, a mixed pesticide solution at the manufacturer-recommended dose (3.2, and 2.1, 4.3 and 4.3 kg ha-1 for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively) was applied to chrysanthemum plants one time.
1
Table 2. Final residue of target pesticides in chrysanthemum flowers (mg kg-1, mean ± standard deviation, n = 3) Number of applications 2
3
a b
Metalaxyl-M PHI (d)
Fludioxonil
Cyantraniliprole
Thiamethoxam
Clothianidin
Low dosea
High doseb
Low dose
High dose
Low dose
High dose
Low dose
High dose
Low dose
High dose
7
0.13 ± 0.04
0.69 ± 0.14
0.09 ± 0.05
0.12 ± 0.08
0.10 ± 0.08
0.24 ± 0.04
0.27 ± 0.05
0.36 ± 0.06
0.053 ± 0.02
0.12 ± 0.03
14
0.11 ± 0.04
0.52 ± 0.07
0.01 ± 0.01
0.08 ± 0.12
0.04 ± 0.01
0.22 ± 0.09
0.10 ± 0.001
0.31 ± 0.07
0.047 ± 0.01
0.074 ± 0.01
21
0.05 ± 0.03
0.29 ± 0.03
< 0.005
0.02 ± 0.03
0.04 ± 0.003
0.14 ± 0.10
0.09 ± 0.003
0.29 ± 0.03
0.045 ± 0.01
0.057 ± 0.01
7
0.38 ± 0.09
0.90 ± 0.12
0.11 ± 0.05
0.10 ± 0.03
0.27 ± 0.08
0.40 ± 0.15
0.35 ± 0.01
0.78 ± 0.09
0.14 ± 0.02
0.36 ± 0.22
14
0.28 ± 0.10
0.86 ± 0.03
0.01 ± 0.001
0.09 ± 0.01
0.13 ± 0.01
0.23 ± 0.06
0.21 ± 0.03
0.75 ± 0.15
0.11 ± 0.03
0.15 ± 0.03
21
0.20 ± 0.03
0.50 ± 0.04
< 0.005
0.03 ± 0.01
0.13 ± 0.02
0.15 ± 0.14
0.18 ± 0.01
0.69 ± 0.07
0.048 ± 0.01
0.10 ± 0.03
-1
Low dose is the recommended dose (3.2, and 2.1, 4.3 and 4.3 kg ha for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively); High dose is two times of the recommended dose.
1
2 3
Fig. 1. Root bioconcentration factors based on the loss of pesticides in soils (BCFLS) (A) and relationship between log BCFLS and log Kow (B).
4
5 6
Fig.2. Residue concentrations of metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam, and clothianidin in chrysanthemum shoots (flower,
7
leaf and stem).
Fig.3. Average translocation factors (TFstem, TFleaf and TFflower) of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam. (Difference between means is significant at the 0.05 level*, and at the 0.01 level**)
Highlights 1. Uptake and dissipation of four pesticides in greenhouse chrysanthemum were studied. 2. Root bioconcentration factors were positively related to pesticide lipophilicity. 3. Root-to-shoot translocation factors followed the order of leaf > stem > flower. 4. A main toxic metabolite of thiamethoxam (clothianidin) showed greater health risks.
S0269-7491(19)33362-7
DOI:
https://doi.org/10.1016/j.envpol.2019.113499
Reference:
ENPO 113499
To appear in:
Environmental Pollution
Received Date: 12 July 2019 Revised Date:
25 September 2019
Accepted Date: 25 October 2019
Please cite this article as: Gong, W., Jiang, M., Zhang, T., Zhang, W., Liang, G., Li, B., Hu, B., Han, P., Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in greenhouse chrysanthemum, Environmental Pollution (2019), doi: https://doi.org/10.1016/j.envpol.2019.113499. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.
1
Uptake and dissipation of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in
2
greenhouse chrysanthemum
3
Wenwen Gonga,*, Mengyun Jianga,b, Tingting Zhangb, Wei Zhangc, Gang Lianga, Bingru Lia, Bin
4
Hud, Ping Hana
5
a
Beijing Research Center for Agriculture Standards and Testing, Beijing 100097, China
6
b
College of Chemical Engineering, Beijing University of Chemical Technology, Beijing 100029,
7
China
8
c
9
48824, USA
10
d
Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI
Beijing Plant Protection Station, Beijing 100029, China
11 12
*Corresponding author.
13
Dr. Wenwen Gong, E-mail: [email protected]
1
14
ABSTRACT
15
Production of chrysanthemum (Dendranthema grandiflora) in greenhouses often requires
16
intensive pesticide use, which raises serious concerns over food safety and human health. This study
17
investigated uptake, translocation and residue dissipation of typical fungicides (metalaxyl-M and
18
fludioxonil) and insecticides (cyantraniliprole and thiamethoxam) in greenhouse chrysanthemum
19
when applied in soils. Chrysanthemum plants could absorb these pesticides from soils via roots to
20
various degrees, and bioconcentration factors (BCFLS) were positively correlated with lipophilicity
21
(log Kow) of pesticides. Highly lipophilic fludioxonil (log Kow = 4.12) had the greatest BCFLS (2.96 ±
22
0.41 g g-1), whereas hydrophilic thiamethoxam (log Kow = -0.13) had the lowest (0.09 ± 0.03 g g-1).
23
Translocation factors (TF) from roots to shoots followed the order of TFleaf > TFstem > TFflower.
24
Metalaxyl-M and cyantraniliprole with medium lipophilicity (log Kow of 1.71 and 2.02, respectively)
25
and hydrophilic thiamethoxam showed relatively strong translocation potentials with TF values in
26
the range of 0.29–0.81, 0.36–2.74 and 0.30–1.03, respectively. Dissipation kinetics in
27
chrysanthemum flowers followed the first-order with a half-life of 21.7, 5.5, 10.0 or 8.2 days for
28
metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively. Final residues of these
29
four pesticides, including clothianidin (a primary toxic metabolite of thiamethoxam), in all
30
chrysanthemum flower samples were below the maximum residue limit (MRL) values 21 days after
31
two soil applications each at the recommended dose (i.e., 3.2, 2.1, 4.3 and 4.3 kg ha-1, respectively).
32
However, when doubling the recommended dose, the metabolite clothianidin remained at
33
concentrations greater than the MRL, despite that thiamethoxam concentration was lower than the
34
MRL value. This study provided valuable insights on the uptake and residues of metalaxyl-M,
35
fludioxonil, cyantraniliprole and thiamethoxam (including its metabolite clothianidin) in greenhouse
36
chrysanthemum production, and could help better assess food safety risks of chrysanthemum
37
contamination by parent pesticides and their metabolites.
38
Keywords：Chrysanthemum, Pesticide, Uptake, Translocation, Greenhouse, Residue dissipation 2
39
40
1. Introduction
41
Chrysanthemum (Dendranthema grandiflora) is widely grown in China. More than 26 billion
42
chrysanthemum flower heads were sold in China in 2016 with approximately $2.2 billion in sales [1].
43
It is commercially valuable as both cut flowers and potted plants. Chrysanthemum is also used in
44
food supplement, herbal tea, and medicine [2]. Its flower head is a popular traditional medicine due
45
to its unique flavor, color and health benefits [3] and is listed in Chinese Pharmacopoeia as
46
Chrysanthemi Flos [4]. Previous studies have indicated that chrysanthemum tea has calming,
47
heat-clearing and detoxifying effects and can help reduce physical and mental stress [5, 6].
48
Chrysanthemum flowers are most often consumed via tea infusion, which is prepared from extraction
49
of chrysanthemum flowers with hot water by consumers at home or sold on the market as
50
ready-to-drink tea [7, 8]. Thus, certain pesticide residues in contaminated tea or herbs (including
51
chrysanthemum flowers) may be transferred to tea infusion, which may then pose health risks to tea
52
drinkers [8-10].
53
Chrysanthemum is often cultivated in greenhouses for protection against inclement weather.
54
However, high humidity and warm temperature in greenhouses may cause severe infestations by
55
insects (e.g., aphids and larvae of various Lepidoptera species), fungi and oomycetes [11]. Pest
56
management with pesticides is often used to reduce crop loss from insects and plant diseases, but
57
also exposes chrysanthemum crops to potential pesticide residues [12]. When pesticides are used in
58
greenhouses, their residual concentrations in soils and plants may be greater than those in open fields,
59
largely due to unique greenhouse conditions such as inadequate ventilation and high humidity that
60
could affect pesticide dissipation (e.g., vaporization of volatile/semi-volatile active ingredient)
61
[12-14]. However, knowledge on dissipation behaviors of pesticides during chrysanthemum
62
production in greenhouses is very limited.
63
Dissipation behaviors of pesticides is dependent on methods of pesticide application. Foliar
64
application of pesticides is the most widely used treatment in field and greenhouse conditions, but 3
65
often results in several negative consequences, such as off-target drift, killing pests’ natural enemies,
66
secondary pest outbreaks, as well as potential exposure by farmer workers [15]. In contrast,
67
application of pesticides to soils via irrigation water (i.e., chemigation) is an alternative that could
68
avoid or reduce adverse impact mentioned above [15, 16]. Soil application can be effective not only
69
for soilborne pests and pathogens, but also for protecting the upper part of plants through uptake and
70
translocation of active pesticide ingredients to plant shoots via transpiration stream [17]. Thus, soil
71
application is becoming increasingly popular. In addition, previous studies indicated that
72
combination formulation of several pesticides can be more effective than single ingredient
73
formulation. Therefore, combination pesticides are often applied to crops [18]. However, the uptake
74
and residue dissipation of mixed pesticides in chrysanthemum plants and soils after soil application
75
have rarely been studied.
76
Therefore, this study aimed to investigate the uptake, translocation and dissipation of two typical
77
fungicides (metalaxyl-M and fludioxonil) and two insecticides (cyantraniliprole and thiamethoxam)
78
in chrysanthemum grown in greenhouses after pesticide application to soils at the
79
manufacturer-recommended dose and double of the recommended dose. A modified quick, easy,
80
cheap, effective, rugged and safe (QuEChERS) extraction method was established to extract
81
pesticides from chrysanthemum root, stem, leaf and flower samples and soil samples, and the
82
extracted pesticides were then analyzed with UPLC-MS/MS. In addition, a main metabolite of
83
thiamethoxam (clothianidin) in chrysanthemum was identified and measured throughout the
84
experiment as its analytical-grade standard was commercially available.
85
2. Materials and methods
86
2.1. Reagents
87
Standards of metalaxyl-M (purity of 99.5%), fludioxonil (purity of 99.0%), cyantraniliprole
88
(purity of 99.2%), thiamethoxam (purity of 98.2%) and its metabolite clothianidin (purity of 99.0%)
89
were purchased from China Standard Material Center (Beijing, China). Properties of tested pesticides 4
90
are shown in Supplemental Table S1. A commercial formulation of 62.5 g L-1 metalaxyl-M
91
fludioxonil SC (37.5 g L-1 for metalaxyl-M and 25 g L-1 for fludioxonil), and a commercial
92
formulation of 40% cyantraniliprole thiamethoxam SC (20% for each) were obtained from
93
Syngenta (Beijing, China). HPLC-grade acetonitrile and formic acid were purchased from
94
Sigma-Aldrich (St. Louis, MO, USA) and used as received. Analytical reagent-grade sodium
95
chloride (NaCl) and magnesium sulfate (MgSO4) were purchased from Sinopharm Chemical
96
Reagent (Beijing, China) and heated for 12 h at 120 °C to remove residual water before use. Primary
97
secondary amine (PAS, 40 µm), graphitized carbon black (GCB) and C18 were purchased from
98
Agela Technologies Ltd. (Tianjin, China) and used without pretreatment.
99
2.2. Greenhouse experiments
100
Greenhouse experiments were performed in a greenhouse at Beijing Plant Protection Station,
101
Beijing, China from September to November 2018. The greenhouse was under natural lighting and
102
maintained at average temperature of 21 ± 2 °C and relative humidity of 75 ± 2%. Chrysanthemum
103
plants were cultivated in plastic pots (500 × 340 × 263 mm) each filled with dry soil mass of 6 ± 0.5
104
kg by experienced gardeners. The soil had 2.6% of sand, 86.4% of silt and 11.0% of clay, and was
105
classified as silt as per the USDA classification. The soil had an organic matter of 262.3 g kg-1, a
106
cation exchange capacity of 38.1 cmol kg-1 and a pH of 7.1. A randomized block design
107
(Supplemental Table S2) was used where two adjacent block were separated by a 0.5-m alley.
108
Experiment A was designed to investigate the uptake, translocation and dissipation kinetics of
109
metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam, and clothianidin in chrysanthemum plants.
110
Chrysanthemum plants in 120 pots (four plants per pot) at the flower bud initiation stage received a
111
mixed pesticide solution (200 mL) once at the manufacturer-recommended dose (3.2, and 2.1, 4.3
112
and 4.3 kg ha-1 for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively; low
113
dose). Chrysanthemum samples were collected from three random pots taken at 0 (about 2 h), 1, 2, 3,
114
5, 7, 14 and 21 days after pesticide application. Meanwhile, 50 pesticide-free Chrysanthemum pots 5
115
(Experiment C) received pure water of the same volume. Samples were collected before the pesticide
116
application and then at harvest.
117
Experiment B was conducted to study final pesticide residues in chrysanthemum flowers.
118
Chrysanthemum plants in 50 pots (four plants per pot) received a mixed solution of pesticides at the
119
low dose or the high dose (i.e., doubling the recommended dose) two or three times with a 7-day
120
interval. Chrysanthemum flowers from three randomly selected pots in each treatment were collected
121
at pre-harvest interval (PHI) of 7, 14 and 21 days after the last soil application. Extra pots were
122
included and not all planted pots were used in the experiments.
123
2.3. Sample collection, extraction and analysis
124
The chrysanthemum samples were divided into four parts, including flowers, leaves, stems and
125
roots. Chrysanthemum roots were rinsed in acetonitrile under shaking for 20 s followed by washing
126
with distilled water to remove pesticides residue on root surface [15]. Then, each portion of
127
chrysanthemum was separately cut into small pieces and homogenized using a blender
128
(BUCHI/B-400, China) for 1–2 min until a completely homogenized sample was obtained. All
129
samples were transported to the laboratory and stored in a freezer at −20 °C until extraction. The
130
QuEChERS method modified from Golge and Kabak [19] was used for extracting the
131
chrysanthemum samples. The choices of extraction solvents (acetonitrile, acetone or ethyl acetate)
132
and clean-up reagents were optimized by measuring recoveries of spiked pesticides.. Acetonitrile was
133
shown as the most suitable solvent to extract the tested pesticides from chrysanthemum matrix. A
134
combination of 20, 30, or 50 mg of PSA and C18 (or GCB) as well as 100 mg of anhydrous MgSO4
135
were tested as clean-up reagents. Satisfactory recoveries were obtained by the combination of 20 mg
136
PSA, 20 mg GCB and 100 mg anhydrous MgSO4 for leaf and flower samples and 50 mg PSA, 20 mg
137
GCB and 150 mg anhydrous MgSO4 for root and stem samples, respectively. The QuEChERS
138
method is detailed as follows: 5 g of each flower or leaf sample were added to a 50-mL centrifuge
139
tube with 10 mL acetonitrile and shaken for 20 min, followed by adding 2 g of anhydrous MgSO4 6
140
and 1 g of NaCl. The mixture was vortexed for 1 min, and then centrifuged at 8000 rpm for 5 min.
141
Afterwards, 1 mL of the supernatant was transferred to a 2-mL clean-up tube containing 20 mg PSA,
142
20 mg GCB and 150 mg anhydrous MgSO4. After mixing, the tube was centrifuged at 10000 rpm for
143
10 min. The supernatant was collected again and filtered through 0.22-µm Whatman GF/C glass
144
fibre filters for further analysis. In addition, 2 g of stem and root samples were extracted using the
145
abovementioned procedure except that each clean-up tube had 50 mg PSA and 150 mg anhydrous
146
MgSO4.
147
Soil samples were thoroughly mixed and passed through a 2-mm sieve. Subsequently, 5 g soil
148
samples were weighed into the 50-mL centrifuge tube. The extraction and clean-up procedures were
149
the same with the abovementioned protocol, except that each clean-up tube contained 50 mg PSA, 50
150
mg C18 and 150 mg anhydrous MgSO4.
151
The analyses of metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam and clothianidin
152
concentrations in extracted samples were carried out with a Waters ACQUITY UPLC-MS/MS
153
system equipped with an ACQUITY BEH C18 column (2.1 mm × 100 mm × 5 µm) (Milford, MA,
154
USA). The main metabolite of thiamethoxam (clothianidin) was analyzed as we had access to its
155
analytical-grade standard. Other metabolites were not included because they were either not
156
identified or no analytical-grade standards were available. Full metabolite identification and
157
quantification were out of scope for this study and could be explored in future studies. Detailed
158
information regarding the optimized UPLC-MS/MS condition is provided in Supporting information
159
(SI). Analysis was performed in multiple reaction monitoring (MRM) mode, with operational
160
parameters shown in Supplemental Table S3.
161
2.4. Data analysis
162
To estimate accumulation potential of organic compounds from soils into plant tissues,
163
bioconcentration factors (BCFs) are calculated as the ratios of concentrations in plant tissues to that
164
in soils and have been widely used as indexes in studying plant uptake of organic contaminants [20, 7
165
21]. However, some studies have suggested that BCFs based on concentrations in bulk soils (BCFsoil)
166
may not accurately represent the uptake potential because chemical concentrations in bulk soils
167
could not directly reflect the bioavailability of chemicals for plant uptake [22, 23]. In addition, the
168
calculation of BCFsoil is generally based on an assumption that chemical concentrations in soils
169
remain relatively constant [24, 25]. However, a significant decrease of pesticide concentrations in
170
soils (approximately by 28% to 43%) was observed during chrysanthemum cultivation in this study.
171
To overcome the large variation (uncertainty) of pesticide concentrations in bulk soils, BCFs based
172
on the loss of pesticides in soils (BCFLS) were previously proposed by Sun et al. [14]. We assume
173
that pesticides are first released from soils to soil pore water, their losses due to leaching,
174
volatilization, biodegradation, or plant root uptake. Therefore, the total decrease of pesticide
175
concentrations in soils represent the total amount of pesticides available for plant uptake. Thus, root
176
BCFLS (g g-1) was calculated using Eq. (1). = ⁄
177
(1)
178
where Croot (mg kg-1) is the pesticide concentration in roots, and Closs in soil (mg kg-1) is the decrease of
179
pesticide concentration in soils, i.e., the maximum concentrations of pesticides in soils subtracted by
180
the concentrations of pesticides in soils measured at time t (14 and 21 days in this study).
181 182
To better evaluate translocation potential of measured pesticides from chrysanthemum roots onto shoot (stem, leaf, and flower), translocation factors (TF) were calculated as follows.
183
TF = /
(2)
184
TF = /
(3)
185
TF = /
(4)
186
where Cstem, Cleaf, and Cflower (mg kg-1) are the pesticide concentration in stem, leaf and flower,
187
respectively.
188
As the pesticide residues in chrysanthemum flowers are key to health risk assessment, the
189
dissipation kinetics of measured pesticides in chrysanthemum flowers were fitted to the first-order 8
190
kinetic equation (Eq. 5). The dissipation half-life (t1/2) was calculated via Eq. (6).
191
=
(5)
192
/ = 2/"
(6)
193
where k (day-1) is the dissipation rate constant, Ct (mg kg-1) and C0 (mg kg-1) are the pesticide
194
concentration at a given time t and at the beginning. In this study, the pesticide concentration in
195
chrysanthemum flowers initially increased for a few days (1 to 3 days) after the final pesticide
196
application (Supplemental Table S2). Therefore, the peak concentration was used as the initial point
197
for fitting the dissipation kinetics [12].
198
The uptake, translocation and dissipation kinetics of target pesticides in chrysanthemum plants
199
and linear regression analyses between log BCFLS with log Kow values of pesticides were analyzed
200
with the Origin software 9.0 (OriginLab Inc., Northampton, MA, USA). One-way analysis of
201
variance (ANOVA) was performed to test significant difference in means between sample groups
202
using SPSS v20.0 (IBM, New York, NY, USA). According to Tukey’s multiple comparison tests, p <
203
0.05 and p < 0.01 were considered as significant and highly significant, respectively. Correlation
204
analyses were performed using Pearson’s test with SPSS v20.0, and correlation was considered
205
statistically significant when p value was less than 0.01.
206
2.5. Quality assurance and quality control (QA/QC)
207
The QA/QC of pesticide analysis included determination of linearity, recovery, precision and
208
accuracy. Linearity was assessed with matrix-matched standards in the range of 0.5 µg L-1–0.2 mg
209
L-1 for chrysanthemum (including flower, leaf, stem and root) and soil samples. The correlation
210
coefficients (R2) of matrix-matched calibration curves were all greater than 0.991, demonstrating a
211
good linearity (Supplemental Table S4). Recovery and precision tests were conducted using blank
212
chrysanthemum samples (flower, leaf, stem and root) fortified with target compounds at 0.005, 0.05
213
and 0.2 mg kg-1 levels or blank soil samples fortified at 0.002, 0.05 and 0.2 mg kg-1 levels with five
214
replicates. The average recoveries of the fortified samples ranged from 78% to 114%, with relative 9
215
standard deviation (SD) varying between 0.05% to 13.06% (Supplemental Table S5). In all cases, the
216
recovery results were acceptable and thus confirmed that the analytical method was reliable for
217
analyzing metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam and its metabolite clothianidin.
218
The limit of quantification (LOQ) was determined as the lowest concentration of a given analyst at
219
which an acceptable recovery (70–120%) and a relative SD of ≤ 20% can be achieved [17]. The
220
LOQs of five pesticides were 0.005 mg kg-1 in chrysanthemum tissues and 0.002 mg kg-1 in the soils,
221
based on the analysis of five replicates.
222
3. Results
223
3.1 Uptake and bioaccumulation from soils into chrysanthemum roots
224
The four pesticides were detected in chrysanthemum roots at 0.63–2.28 mg kg−1 on day 14 (the
225
pre-anthesis stage) and at 0.46–1.33 mg kg−1 on day 21 (the anthesis stage,) after the last soil
226
application (Experiment A, Supplemental Table S6), whereas none was found in the roots for the
227
blank control (Experiment C). Thus, chrysanthemum roots could absorb and accumulate these
228
pesticides from soils. Fludioxonil exhibited the highest concentrations of 2.28 and 1.33 mg kg-1 on
229
day 14 and 21, respectively, which was up to 3.6 times of the concentrations of other three pesticides
230
(Supplemental Table S6). The calculated BCFLS values for each pesticide are shown in Fig. 1A. The
231
BCFLS values ranged between 0.15–2.96 g g-1 at day 14, and between 0.09–2.59 g g-1 at day 21.
232
Among all the pesticides, fludioxonil demonstrated the highest BCFLS values, whereas thiamethoxam
233
had the lowest BCFLS values (p < 0.01). There was a strong positive correlation between log BCFLS
234
and log Kow (i.e., octanol-water partition coefficient, Table S1), as shown in Fig. 1B (R2 = 0.92, p <
235
0.01).
236
3.2 Root-to-shoot translocation
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In this study, the residue concentrations of metalaxyl-M, fludioxonil, cyantraniliprole, and
238
thiamethoxam in different parts of chrysanthemum (flower, stem and leaf) over the testing period 10
239
were measured and shown in Fig. 2. The concentrations of all the measured pesticides were much
240
higher in chrysanthemum leaves than in stem, and were the lowest in flowers. The pesticides also
241
remained longer in leaves than in stems and flowers (Fig. 2). Fludioxonil increased in
242
chrysanthemum leaves during the entire cultivation period but were much less than the other three
243
pesticides. The residue concentrations of metalaxyl-M, cyantraniliprole, and thiamethoxam increased
244
and reached their maximum concentrations in leaves 7 days after the last application, and then
245
gradually decreased.
246
The average TF values of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam are shown in
247
Fig. 3. The TFstem, TFleaf and TFflower values of four pesticides during the cultivation period were in
248
the range of 0.01–2.22, 0.07–3.88, 0.002–0.77, respectively (Supplemental Table S7). Interestingly,
249
the TF values initially increased after pesticide applications (1–5 days) and then decreased over time.
250
Generally, TF of each tissue for all pesticides followed the order of TFleaf > TFstem > TFflower (p <
251
0.05), suggesting that chrysanthemum leaves are the main reservoir for absorbed pesticides. 3.3
252
Dissipation kinetics in chrysanthemum flowers
253
Normally, the residues of four pesticides in chrysanthemum flowers increased from 1 to 3 days
254
after the last application, and then decreased gradually thereafter (Fig. 2). Their dissipation
255
percentages in chrysanthemum flowers 21 days after the last application were 49.0%, 90.9%, 72.4%
256
and 81.6%, respectively. The dissipation of each pesticide in chrysanthemum flowers was
257
satisfactorily fitted with a first-order kinetic equation, with R2 ranging from 0.744 to 0.943 (Table 1).
258
The calculated half-life values t1/2 in chrysanthemum flowers were 21.7, 5.5, 10.0 and 8.2 days for
259
metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively.
260
In this study, clothianidin was detected in chrysanthemum flowers with the application of
261
thiamethoxam. Clearly, thiamethoxam was metabolized to clothianidin, resulting in a considerable
262
concentration of clothianidin in chrysanthemum plants. The residues of clothianidin increased with
263
the degradation of thiamethoxam (Fig. 2). Clothianidin concentration reached a maximum level in 11
264
chrysanthemum flowers (0.06 mg kg-1) 5 days after the last application and then declined eventually
265
to 0.02 mg kg-1. The dissipation of clothianidin residues was fitted to a first-order kinetic equation
266
Ct=0.075 e-0.061t (R2 = 0.975). The calculated half-life t1/2 value of clothianidin was 11.4 days,
267
suggesting that the dissipation rate of thiamethoxam (t1/2 = 8.2 days) in chrysanthemum flowers was
268
greater than that of its metabolite clothianidin.
269
3.4 Final residues
270
The final residues of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam in
271
chrysanthemum flowers on day 7, 14 and 21 after the last application are presented in Table 2. Their
272
residues in chrysanthemum flowers were 0.05–0.38, <0.005–0.11, 0.04–0.27 and 0.09–0.35 mg kg-1,
273
respectively, after 2–3 times of application at the recommended dose. When applied at a higher dose,
274
their final residues were increased to varying degrees. Greater concentrations were observed in the
275
treatments with more applications and shorter pre-harvest intervals.
276
4. Discussion
277
Our studies showed that chrysanthemum root could uptake metalaxyl-M, fludioxonil,
278
cyantraniliprole, and thiamethoxam from soils to different degrees (Fig. 1A and Supplemental Table
279
S6 ), and the calculated BCFLS values were related positively to log Kow of pesticides (Fig. 1B). This
280
observation was in a good agreement with other studies [26, 27]. Thus, partitioning was the dominant
281
mechanism controlling the root uptake of these nonionic organic compounds [28]. Recently, several
282
studies have found ion trap effect for ionizable organic compounds [23, 29, 30], which would not be
283
applicable to non-ionic compounds that do not change their charge speciation as a function of
284
solution pH. In addition, molecular weight is regarded as an important factor for plant uptake,
285
probably due to its effect on the passage through cell membrane or Casparian strip [31-34]. For
286
example, Li et al. [23] investigated the uptake of 15 chemicals with a wide range of molecular
287
weight by radish roots, and found that most of the chemicals with molecular weight <500 g mol-1 12
288
showed a greater bioconcentration than those with molecular weight > 500 g mol-1. For the pesticides
289
with low molecular weight (Table S1), no correlation was observed between BCFLS and molecular
290
weight, suggesting that there may be threshold molecular weight below which plant uptake was not
291
influenced by molecular weight. In addition to physicochemical properties of chemicals, other
292
factors such as crop species, cultivation methods and environmental conditions may collectively
293
affect the uptake and bioconcentration of chemicals [35]. For example, studies have showed that
294
crops could absorb and concentrate relatively more organic pollutants (e.g., clarithromycin and
295
sulfadiazine) when grown in hydroponic systems than in soils [36-38].
296
For most plants, pesticides applied through chemigation to soils could be absorbed by plant roots
297
and then translocated to plant shoots through transpiration stream [15, 17, 39-41]. Overall, the
298
concentrations of pesticides in chrysanthemum leaves took some time to reach the maximum, were
299
dissipated slowly and could remain in the plant for a long time (Fig. 2). In stems, the pesticide
300
residues researched their maximum earlier than in leaves, but remained at a largely stable and low
301
level throughout the experimental period. These results were consistent with previous studies,
302
reporting that the stems more likely serve as a transport channel with little retention of pesticides
303
[17]. Several studies suggested that deposition of gaseous pesticides volatilized from soils or
304
deposition of resuspended soil particles on aerial parts of plants also contributed to the accumulation
305
of pesticides in plant shoots [14, 42]. However, none of the pesticides was detected in the shoots of
306
chrysanthemum planted in pesticide-free soils in the same greenhouse, indicating that these two
307
pathways were negligible for the accumulation of pesticides with low vapor pressure (Supplemental
308
Table S1) in chrysanthemum shoots.
309
TF values could reveal the tendency of pesticides to be translocated into a component of the
310
shoot (stem, leaf, and flower) from the root. Our results showed that the TF values initially increased
311
after pesticide applications and then decreased over time (Table S7). Similar results were also
312
observed for the translocation and accumulation of imidacloprid in six leafy vegetables [43] and five 13
313
neonicotinoids in komatsuna [44]. In addition, the TF values of each tissue (0.07–0.17) of highly
314
lipophilic fludioxonil (log Kow = 4.12) were significantly lower than the other three pesticides,
315
indicating that it was mainly accumulated in chrysanthemum roots. In contrast, metalaxyl-M and
316
cyantraniliprole with medium lipophilicity and hydrophilicity (log Kow of 1.71 and 2.02, respectively)
317
and thiamethoxam with low lipophilicity (log Kow = −0.13) showed relatively stronger translocation
318
potentials with the TF values in the range of 0.29–0.81, 0.36–2.74 and 0.30–1.03, respectively. It was
319
previously suggested that there is an optimum lipophilicity of organic compounds for their
320
translocation. Highly hydrophilic compounds may have difficulty to cross the hydrophobic root
321
membranes, whereas highly lipophilic compounds on the other hand are concentrated in roots by
322
binding with root tissues (e.g., lipids) and are less likely to be translocated upward to shoots [27, 45].
323
Therefore, it is possible that a highly lipophilic pesticide (e.g., fludioxonil) can have the highest root
324
bioconcentration, but the lowest translocation to the shoots (Fig. 1 and Fig. 3), suggesting minor
325
health risks from consuming chrysanthemum flowers.
326
Previous studies suggested that the dissipation pattern of pesticides varies with plant species,
327
dose and interval of pesticide application, and climatic conditions [46, 47]. The half-life values of
328
pesticides such as pyrethrins and dechlorane plus in plants or soils under open field conditions were
329
generally less than that under greenhouse conditions, due to factors such as rainfall, solar radiation
330
and crop growth rate, etc. [14, 48]. In addition to physical and chemical factors, such as light,
331
temperature, pH, and moisture, pesticide application methods (e.g., foliar spray, soil application of
332
chemigation, or seed treatment) might also play an important role in the dissipation of pesticides in
333
plants. For example, the observed half-life of thiamethoxam in chrysanthemum flowers in this study
334
was much greater than that reported in teas (1.6 days) [49] and tobacco leaves (3.9–4.4 days) [50] by
335
foliar spraying in the field.
336
The final residues of metalaxyl-M, fludioxonil, and thiamethoxam in chrysanthemum flowers
337
were below the MRL values (Table 2) according to the MRLs for metalaxyl-M (3 mg kg-1), 14
338
fludioxonil (20 mg kg-1) and thiamethoxam (1.5 mg kg-1) in herbs or edible flowers established by
339
the UK/EC MRL Database [51]. However, the final residues of metalaxyl-M, fludioxonil, and
340
thiamethoxam in chrysanthemum flowers were below the MRL values. However, the final residues
341
of cyantraniliprole were greater than the MRL value (0.05 mg kg-1) except for the residue
342
concentrations at the 14 and 21 days after two applications at the low dose. Normally, the MRL value
343
of a hazardous chemical is established based on its acceptable daily intake (ADI). On the basis of the
344
toxicological and ecotoxicological data, the World Health Organization (WHO) and the Food and
345
Agriculture Organization of the United Nations (FAO) established the ADI of cyantraniliprole at 0.03
346
mg kg-1 bw, which is much lower than that of metalaxyl-M, fludioxonil and thiamethoxam (0.08, 0.4,
347
and 0.08 mg kg-1 bw, respectively) [52]. Therefore, cyantraniliprole has much lower MRL, and is
348
thus more likely to exceed the MRL than other pesticides.
349
It has been suggested that organic compounds accumulated in plants may be metabolized, and
350
thus, further influence their dissipation pattern in plants [23, 53]. It should be noted that in some
351
cases metabolites can even be more toxic or at higher residue levels than parent pesticides [54, 55].
352
Therefore, there is increasing concern over the residues of some toxicologically significant
353
metabolites that remain in harvested crops and can then be ingested by humans and animals via food
354
or feed. Uptake of pesticide metabolites by chrysanthemum roots was not quantified in this study,
355
because we did not identify all the metabolites and it is also difficult to distinguish whether the
356
metabolites originate from in-plant metabolism or the uptake from soils [56]. However, we identified
357
and measured clothianidin, a main metabolite of thiamethoxam, in chrysanthemum as its
358
analytical-grade standard was commercially available. The final residue levels of clothianidin in
359
chrysanthemum flowers ranged from 0.045 to 0.36 mg kg-1 (Table 2) and were greater than the MRL
360
for clothianidin in flowers (0.05 mg kg-1) listed in the UK/EC MRL database except for its
361
concentration at 14 and 21 days after 2 applications and at 21 days after 3 applications at the low
362
dose. In contrast, the residues of parent pesticide thiamethoxam in all samples were lower than the 15
363
MRL values, indicating that risks may be underestimated if only thiamethoxam was measured.
364
Therefore, when assessing the food safety and human health risks of thiamethoxam application, it
365
would not be adequate to measure only the residue concentrations of thiamethoxam, and the
366
metabolite (clothianidin) should be included as well. Finally, when applied at a higher dose or at a
367
low dose with more frequent applications during chrysanthemum clutivation, the final residues of
368
tested pesticides were likely to exceed the MRL (Table 2), which warrants close attention to resultant
369
risks to food safety.
370
5. Conclusion
371
The findings of this study have several important implications to the food safety and quality of
372
chrysanthemum production in greenhouses. Metalaxyl-M, fludioxonil, cyantraniliprole and
373
thiamethoxam had varying potential to be absorbed by chrysanthemum roots, and then translocated
374
to shoots (stems, leaves and flowers), following their soil application. Pesticide residues in
375
chrysanthemum flowers increased at the first few days after application, and then decreased
376
gradually thereafter, followed the first-order kinetics. Moreover, clothianidin, a primary metabolite of
377
thiamethoxam, was detected in both chrysanthemum roots and shoots throughout the experiment.
378
The final residues of these four pesticides and clothianidin in chrysanthemum flowers were
379
significantly affected by the application times, doses and preharvest intervals. The final residues of
380
metalaxyl-M, fludioxonil, and thiamethoxam after 2–3 applications at either low dose or high dose
381
were blow the MRL in chrysanthemum flowers, suggesting the lower food safety risks of these
382
parent pesticide compounds. However, cyantraniliprole exceeded the MRL more often than other
383
pesticides, suggesting that close attention should be paid to its residues in chrysanthemum flowers.
384
Furthermore, the main metabolite of thiamethoxam (clothianidin) had the final residues greater than
385
the MRL in most treatments, highlighting the need to monitoring the concentrations of toxic
386
metabolites in addition to parent compounds. This study was limited in the sense that only one
387
metabolite was included because other metabolites could not be identified or their analytical-grade 16
388
standards were not available to us. Thus, future study should be directed to characterizing and
389
quantifying toxic metabolites of parent pesticide compounds. This study provided useful data for
390
assessing the safe use of typical fungicides and insecticides by soil application in greenhouse
391
chrysanthemum cultivation. Future work should also investigate the transfer of pesticide residues
392
from chrysanthemums flowers into tea infusion from a food safety and human health perspective.
393
Acknowledgments
394
This work was financially supported by National Natural Science Foundation of China
395
(21806014), Special Projects of Construction of Science and Technology Innovation Ability of
396
Beijing Academy of Agriculture and Forestry Sciences (KJCX20190405; KJCX20170419), and
397
Project of Beijing Excellent Talents (2017000020060G127).
398
17
399
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Table 1. Dissipation kinetic equation, correlation coefficient and half-life of target pesticides in chrysanthemum flowers.a
a
Pesticides
Regression equation
Correlation coefficient (R2)
Half-life (t1/2)
Metalaxyl-M
Ct=0.148 e-0.032t
0.835
21.7
Fludioxonil
Ct=0.046 e-0.127t
0.762
5.5
Cyantraniliprole
Ct=0.096 e-0.069t
0.744
10.0
Thiamethoxam
Ct=0.256 e-0.085t
0.943
8.2
Clothianidin
Ct=0.075 e-0.061t
0.975
11.4
To investigate the dissipation kinetics, a mixed pesticide solution at the manufacturer-recommended dose (3.2, and 2.1, 4.3 and 4.3 kg ha-1 for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively) was applied to chrysanthemum plants one time.
1
Table 2. Final residue of target pesticides in chrysanthemum flowers (mg kg-1, mean ± standard deviation, n = 3) Number of applications 2
3
a b
Metalaxyl-M PHI (d)
Fludioxonil
Cyantraniliprole
Thiamethoxam
Clothianidin
Low dosea
High doseb
Low dose
High dose
Low dose
High dose
Low dose
High dose
Low dose
High dose
7
0.13 ± 0.04
0.69 ± 0.14
0.09 ± 0.05
0.12 ± 0.08
0.10 ± 0.08
0.24 ± 0.04
0.27 ± 0.05
0.36 ± 0.06
0.053 ± 0.02
0.12 ± 0.03
14
0.11 ± 0.04
0.52 ± 0.07
0.01 ± 0.01
0.08 ± 0.12
0.04 ± 0.01
0.22 ± 0.09
0.10 ± 0.001
0.31 ± 0.07
0.047 ± 0.01
0.074 ± 0.01
21
0.05 ± 0.03
0.29 ± 0.03
< 0.005
0.02 ± 0.03
0.04 ± 0.003
0.14 ± 0.10
0.09 ± 0.003
0.29 ± 0.03
0.045 ± 0.01
0.057 ± 0.01
7
0.38 ± 0.09
0.90 ± 0.12
0.11 ± 0.05
0.10 ± 0.03
0.27 ± 0.08
0.40 ± 0.15
0.35 ± 0.01
0.78 ± 0.09
0.14 ± 0.02
0.36 ± 0.22
14
0.28 ± 0.10
0.86 ± 0.03
0.01 ± 0.001
0.09 ± 0.01
0.13 ± 0.01
0.23 ± 0.06
0.21 ± 0.03
0.75 ± 0.15
0.11 ± 0.03
0.15 ± 0.03
21
0.20 ± 0.03
0.50 ± 0.04
< 0.005
0.03 ± 0.01
0.13 ± 0.02
0.15 ± 0.14
0.18 ± 0.01
0.69 ± 0.07
0.048 ± 0.01
0.10 ± 0.03
-1
Low dose is the recommended dose (3.2, and 2.1, 4.3 and 4.3 kg ha for metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam, respectively); High dose is two times of the recommended dose.
1
2 3
Fig. 1. Root bioconcentration factors based on the loss of pesticides in soils (BCFLS) (A) and relationship between log BCFLS and log Kow (B).
4
5 6
Fig.2. Residue concentrations of metalaxyl-M, fludioxonil, cyantraniliprole, thiamethoxam, and clothianidin in chrysanthemum shoots (flower,
7
leaf and stem).
Fig.3. Average translocation factors (TFstem, TFleaf and TFflower) of metalaxyl-M, fludioxonil, cyantraniliprole and thiamethoxam. (Difference between means is significant at the 0.05 level*, and at the 0.01 level**)
Highlights 1. Uptake and dissipation of four pesticides in greenhouse chrysanthemum were studied. 2. Root bioconcentration factors were positively related to pesticide lipophilicity. 3. Root-to-shoot translocation factors followed the order of leaf > stem > flower. 4. A main toxic metabolite of thiamethoxam (clothianidin) showed greater health risks.