2014 Volume 39 Issue 2 Pages 69-75
Greenhouse cultivation supplies fresh vegetables even in the winter season by protecting a crop from the cold environment. Greenhouse cultivation also influences the decline of pesticide residues in crops due to runoff by rainfall1) and photodegradation by sunlight.2) There are many reports on the effects of greenhouses on pesticides in crops; however, those reports provide only fragmentary knowledge of individual test conditions.1–6) There are few comparative studies on multi-class pesticides. Therefore, these reports are insufficient for clarifying the influence of complex factors of pesticide residues involved in common agricultural practices, such as the intervals between pesticide application and harvest and the effects of the physicochemical properties of pesticides.
In our previous two studies, we observed different residue tendencies assumed to be influenced by growing conditions. The first study estimated variations in pesticide residue levels in crops7); the individual residue levels in grapes grown in a greenhouse had a good agreement between acetamiprid and cypermethrin. In contrast, the distribution of cypermethrin residue levels in cabbage grown in an open field was slightly skewed at higher residue levels as compared to that of acetamiprid. The second study estimated the effect of field-to-field variation in pesticide residue levels in different years;8) the correlation of the pesticide residue levels in spinach grown in greenhouses between the data obtained in different years from the same six greenhouses was in good agreement. In contrast, there was no clear correlation between the pesticide residue levels in Chinese cabbage samples grown in open fields. Unfortunately, these results are insufficient for clarifying the influence of growing conditions on pesticide residue levels in crops. Our previous studies were not designed to focus on that matter.
The Japanese test guideline for pesticide residue trials recommends greenhouse cultivation as a basic field condition.9) Recently, the Japanese test guideline for pesticide residue trials for major and semi-major crops was revised to increase the number of test fields from the previous regulation of more than two test fields.9) For example, the pesticide residue trial for lettuce heads, as a major crop, should be conducted at more than six fields. The expected effect of increasing the number of test fields is a more accurate understanding of the behavior of pesticide residue in crops. In practice, an unexpected suspicious behavior is sometimes observed in the residue data of large wrapped leafy vegetables such as cabbage, Chinese cabbage, and lettuce head samples.
In consideration of the above mentioned, we conducted decline trials in six open fields and six greenhouses to evaluate the influence of growing conditions on the pesticide residue levels in lettuce heads. Head lettuce is one of the most popular vegetables cultivated in greenhouses and plastic tunnels in Japan. According to the status survey report for glass or plastic-covered greenhouses in Japanese agriculture, approximately one quarter of lettuce fields are covered by greenhouses, including plastic tunnels.10)
This study was also undertaken to estimate the effect of the sample size reduction process on measured pesticide residue levels in lettuce heads, which are representative of large wrapped leafy vegetables. There was no significant effect of the sample size reduction process for fruits and fruiting vegetables.11–13) In contrast, this process had a significant effect on large wrapped leafy vegetables, such as Chinese cabbage, in our previous studies.8)
Trials involved four pesticides that varied greatly in their respective physicochemical properties. Dinotefuran is a neonicotinoid insecticide that has a relatively low log POW of −0.549 and a relatively high water solubility of 39.8 g/L.14) Permethrin is a pyrethroid insecticide that has a relatively high log POW of 6.1 and a relatively low water solubility of 6 µg/L.14) The other two pesticides, acetamiprid and azoxystrobin, are a neonicotinoid insecticide and a fungicide, respectively, that have intermediate physicochemical properties of log POW and water solubility (log POW of 0.80 and 2.5, and water solubility of 4250 and 6 mg/L, respectively).14) Planting hole application for neonicotinoid insecticides is widely used under modern Japanese agricultural practices due to easy handling and safety for farmers. Therefore, of the two kinds of pesticides, dinotefuran, with the higher water solubility and the weak soil adsorption, was applied to the planting hole, whereas the other three pesticides were applied to foliage, allowing us to compare the efficacy of the types of application.
These investigations provided valuable information for estimating the effect of different plant cultivation techniques on the levels of pesticide residues in lettuce heads in modern agricultural practice in Japan.
The field experiments on head lettuce were conducted in six greenhouses and six open fields near greenhouses across Japan, in accordance with Japanese common agricultural practices.9) Portions (3 g) of Starcle® granules (dinotefuran 1% a.i.) were incorporated into planting holes for each nursery head lettuce. Three formulations of acetamiprid, azoxystrobin, and permethrin were applied in the field experiment for head lettuce to evaluate pesticide decline over time through three sampling intervals, including the shortest pre-harvest interval of 7 days. After diluting with water by factors of 1 : 1000 or 1 : 2000, the pesticides were sprayed three times at a rate of 200 L/10 a per treatment at approximately 7-day intervals between each treatment. The sprays were administered with a tank-mix combination of Mospiran® water-soluble granules as acetamiprid 20% a.i. (Nippon Soda Co., Ltd.), Amistar 20® flowable as azoxystrobin 20% a.i. (Syngenta Japan K.K.), and Adion® emulsifiable concentrate as permethrin 20% a.i. (Sumitomo Chemical Co., Ltd.), by using back-carried sprayers connected to cone nozzles. Lettuce heads were collected 1, 3, and 7 days after the final application. The intervals between Starcle® treatment and harvest ranged from 48, 50, and 54 days in the Kochi field to 70, 72, and 76 days in the Mie field. Each sample consisted of 5–10 lettuce heads taken at random from the test field. The samples, under chilled conditions at 3°C, were carried to our laboratory by a commercial delivery service. Information on the head lettuce fields is summarized in Table 1.
| Field location | Variety | Planting datea) | Sample weight | Weather datab) |
|---|---|---|---|---|
| Open field | ||||
| Ibaraki | Mizusawa | Sep. 11−Nov. 10, 2009 | 517 g | 16.9°C, 293 mm |
| Gunma | Sysco | Sep. 1−Nov. 10, 2009 | 421 g | 18.5°C, 165 mm |
| Nagano | Shinano-summer | Sep.1−Oct. 26, 2009 | 736 g | 17.3°C, 147 mm |
| Mie | Sysco | Sep. 15−Nov. 30, 2009 | 563 g | 16.4°C, 616 mm |
| Kochi | Sysco | Oct. 9−Dec. 8, 2009 | 320 g | 15.6°C, 371 mm |
| Miyazaki | Sysco | Sep. 25−Nov. 23, 2009 | 1051 g | 18.3°C, 613 mm |
| Greenhouse | ||||
| Ibaraki | Falcon | Oct. 13−Dec. 27, 2010 | 652 g | 12.4°C |
| Gunma | Sysco | Sep. 20−Nov. 29, 2010 | 818 g | 23.2°C |
| Nagano | Shinano-summer | Aug. 30−Oct. 25, 2010 | 989 g | 19.7°C |
| Mie | Sysco | Oct. 1−Dec. 10, 2010 | 962 g | 16.6°C |
| Kochi | Sysco | Oct. 20−Dec. 13, 2010 | 374 g | 16.7°C |
| Miyazaki | Sysco | Oct. 26−Dec. 20, 2010 | 590 g | 16.1°C |
a) The range represents the period from the planting date of nursery head lettuce in the fields (the plant hole application date) to the final sampling date.b) The numbers indicate the mean temperature and the total precipitation amount from the first application date to the final sampling date.
After measuring the weight of each sample, we removed the cores and any obviously decomposed or withered leaves from the lettuce heads. Each unit was divided into four or eight pieces vertically by cutting, and a pair of two opposite pieces from each sample was selected as a size-reduced sample (sample-A). For the greenhouse samples, we also examined the other two pieces at 90° angles from sample-A as an additional size-reduced sample (sample-B) in order to investigate the influence of the sample size reduction process. The number of cut pieces was optimized to make each sample a suitable total weight (ca. 800 g). Those size-reduced samples were stored in a freezing room set at −20°C until analyzed. The frozen size-reduced samples were homogenized using the BLIXER 5 Plus (Robot Coupe, MS, USA) before analysis.
3. Residue analysisThe residue analytical methods were optimized to allow rapid analysis of each pesticide in the lettuce head sample, as described below.
3.1 Chemicals and reagentsStandards for acetamiprid, azoxystrobin, dinotefuran, and permethrin were purchased from Hayashi Pure Chemical (Japan) and Wako Pure Chemical Industries, Ltd. (Japan). Pesticide analysis-grade acetone, acetonitrile, and methanol; HPLC-grade tetrahydrofuran; LC-MS-grade methanol; and analytical-grade ammonium formate were purchased from Wako Pure Chemical Industries, Ltd.. Water used for the experiments was purified by a Milli-Q® system (Millipore, MA, USA).
Standard stock solutions (200 mg/L) of each pesticide were separately prepared with acetone or acetonitrile. Portions of each stock solution were diluted with an acetonitrile/water (8 : 2, v/v) solution to make standard solutions in the range of 0.05–2 µg/L to prepare a calibration curve.
3.2 ExtractionA portion (20 g) of the homogenized sample was weighed in an Erlenmeyer flask and extracted with 100 mL of acetonitrile by shaking for 30 min using a reciprocal shaker. The mixture was then filtered by vacuum suction, and the residual cake was washed with 50 mL of acetonitrile and then filtered again. The filtrates were combined and made up to a 200-mL volume with acetonitrile.
3.3 Clean-up procedure 3.3.1 Acetamiprid, azoxystrobin, and permethrinTwo milliliters of the acetonitrile extract (eq. sample of 0.2 g) was cleaned up using solid-phase extraction with an octadecyl silica cartridge (conditioned with acetonitrile and water). Eight milliliters of water was added to the 2 mL of acetonitrile extract, and the mixture was loaded onto the cartridge. A 10-mL mixture of acetonitrile : water (6 : 4, v/v) was passed through the cartridge, and the eluate was collected in a test tube (acetamiprid and azoxystrobin fraction). The cartridge was aspirated for 1 min and dried. Then 10 mL of tetrahydrofuran was passed through the cartridge, and the eluate was collected in a round-bottom flask (permethrin fraction). The acetamiprid and azoxystrobin fraction was diluted to a suitable volume (20–1000 mL) with a mixture of acetonitrile : water (6 : 4, v/v). The permethrin fraction was evaporated until it was dry under a nitrogen stream, and then the residue was dissolved and diluted with a suitable volume (20–1000 mL) of acetonitrile. An aliquot (5 µL) of each diluted test solution was injected into the LC-MS/MS system.
3.3.2 DinotefuranTwo milliliters of the acetonitrile extract (eq. sample of 0.2 g) was cleaned up using solid-phase extraction with a styrene-divinylbenzene copolymer cartridge (1000 mg/6 mL, InertSep™ PLS-2; GL Sciences, conditioned with methanol and water). One milliliter of water was added to the 2 mL of acetonitrile extract, and the mixture was concentrated. Then, 5 mL of water was added to the concentrated solution, and the mixture was loaded onto the cartridge. A 10-mL mixture of methanol : water (6 : 4, v/v) was passed through the cartridge, and the eluate was collected. The eluate was diluted to a suitable volume (20–1000 mL) with water, and then an aliquot (5 µL) of the diluted test solution was injected into the LC-MS/MS system.
3.4 LC-MS/MS analysisEach amount of pesticide in the injected solution was determined using linear regression analysis of each standard calibration curve by comparing the peak area to each concentration of the pesticide in the sample. An LC-MS/MS (Model 1200 Pumping System; Agilent, CA, USA; MS/MS, Model 6410 Triple Quadrupole Tandem Mass Spectrometer; Agilent) equipped with an electrospray interface operating in a positive ion mode was used. Data were processed using the Agilent MassHunter (version B03.01).
3.4.1 Acetamiprid, azoxystrobin, and permethrinLiquid chromatography separation was performed on an Atlantis dC18 column (150 mm×2.1 mm, 5 µm; Waters, USA) at 40°C. The pump was set in an isocratic mode at the rate of 0.2 mL/min with a mobile phase of methanol and 5 mmol/L ammonium formate aqueous solution (6 : 4 for acetamiprid and azoxystrobin, 9 : 1 for permethrin, v/v). The retention times of acetamiprid and azoxystrobin were 3.1 and 11.1 min, respectively. The retention time of permethrin was 6.2 and 7.0 min (isomers separated into two peaks). The mass spectrometry parameters were as follows: capillary voltage, 4000 V; nebulizer gas, 35 psi; drying gas, 11 L/min (350°C); and fragmentor voltage, 100 V. Nitrogen was used as the collision gas for acetamiprid, azoxystrobin, and permethrin at 20, 10, and 15 V, respectively. Precursor ions of acetamiprid, azoxystrobin, and permethrin were selected for m/z 223 ([M+H]+), 404 ([M+H]+), and 408 ([M+NH4]+), respectively. Product ions of acetamiprid, azoxystrobin, and permethrin for m/z 126, 372, and 183, respectively, were detected in the multiple-reaction monitoring mode.
3.4.2 DinotefuranThe analytical column and the column temperature were the same as above. The mobile phase was a linear gradient starting with 10% methanol(A) in 5 mmol/L ammonium formate aqueous solution(B) to reach 95% A in B in 6 min at the flow rate of 0.2 mL/min. The retention time was 5.0 min. The mass spectrometry parameters were as follows: capillary voltage, 4000 V; nebulizer gas, 45 psi; drying gas, 5 L/min (350°C); and fragmentor voltage 50 V. Nitrogen was used as the collision gas at 5 V. A precursor ion was selected for m/z 203 ([M+H]+) and a product ion for m/z 129 was detected in the multiple-reaction monitoring mode.
4. Estimate of the effect of sample size reductionThe means of duplicate measured values were applied to the pesticide residue data for lettuce heads from greenhouses. The overall means of duplicate measured values were applied to the pesticide residue data of lettuce heads from greenhouses because residue levels from the two size-reduced samples were analyzed to provide an additional estimation regarding the sample size reduction.
Results from the recovery and stability tests for lettuce heads, which were used to verify the residue analytical method applied in this study, are summarized in Supplemental Table 1. The accuracy and precision of the analytical method were confirmed by the recovery tests of pesticides fortified at 0.01, 0.25, and 5 mg/kg, which exceeded the highest residue levels. The mean recoveries of 36 spiked samples ranged from 90 to 101%, and their relative standard deviations were equal to or less than 10.8%. The specificity of the analytical method was confirmed by analyzing duplicate blank samples, which were obtained from each field sample. No interference peak was observed around the retention time of each pesticide on chromatograms from the blank samples.
Accurate and consistent instrument performance was ensured using additional recovery samples (QC: quality control samples spiked at 0.1 mg/kg of pesticides) and blank samples and by running a control after every twenty samples. All of the recoveries, from a total of seven additional recovery samples, ranged from 84 to 104%. No interference peak was observed around the retention time of each pesticide on chromatograms from the seven additional blank samples.
Portions (20 g) of the homogenized samples of lettuce heads were weighed in Erlenmeyer flasks. The stability test samples were fortified at 0.25 mg/kg of each pesticide and stored for 34–97 days at −20°C. All of the recoveries from the stability test samples taken from six open fields ranged from 86 to 104%.
These results show that the residue analytical methods applied in this study provided adequate datasets to evaluate the variation in residual pesticide levels in the lettuce heads.
2. Overview of pesticide residue variationStatistical analytical results of pesticide residue datasets in lettuce heads from the six open fields and six greenhouses are summarized in Table 2. The overall mean residue levels in lettuce heads ranged from 0.05 mg/kg for dinotefuran (all samples from the open fields) to 2.04 mg/kg for acetamiprid (1 day after final application in the greenhouses). The overall mean residue levels of each pesticide in lettuce heads were similar to the overall median residue levels.
The relative standard deviation of mean residue levels ranged from 44% for acetamiprid (3 days after final application in the greenhouses) to 98% for azoxystrobin (1 day after final application in the open fields). The highest residue level of azoxystrobin in the samples from the open fields was 119 times higher than the lowest one (0.02–2.38 mg/kg; Mie/Kochi, both samples from 1 day after final application). This was the greatest variation in pesticide residue levels under the same test conditions and was a result of field-to-field variability.
Four varieties of head lettuce were cultivated in six test fields. The mean weights of lettuce heads ranged from 320 to 1051 g. The sample growing tendencies according to the sampling intervals were not observed in all lettuce heads. The mean temperature from the first application date to the final harvest ranged from 12.4 to 23.2°C. The total precipitation amount from the first application date to the harvest date ranged from 147 to 616 mm. The variability of these field conditions represented the wide range of common agricultural practices in Japan.
The residue levels of four pesticides in lettuce heads grown in open fields and greenhouses are shown in Supplemental Fig. 1. The total precipitation amounts at the Mie and Miyazaki fields were larger than those at the other test fields, and the residue levels of four pesticides in the open fields of Mie and Miyazaki are lower than those of the greenhouse in most of the samples (excluding permethrin in a sample from the Mie field from 3 days after final application). However, we could not confirm any clear correlations between the field experimental data, such as sample weights and weather conditions, including a rainfall effect, and the residue levels of pesticides.
At first, we predicted that the pesticide residue level in lettuce heads grown in a greenhouse would be higher than the level in crops grown in an open field, because runoff by rainfall1) and/or photodegradation by sunlight2) might be obstructed in a greenhouse. For example, Stensvand and Christiansen4) investigated the residue levels in strawberries grown in a commercial greenhouse in Norway, who applied eight different fungicides to their samples at the rate recommended for field-grown strawberries. Then the residue levels of some fungicides were above the maximum residue limits (MRLs) of the harmonized EU standards. In our study, all pesticide residue levels in the lettuce heads were lower than the MRLs, as specified by the Japanese Food Sanitation Law15) (MRLs of acetamiprid, azoxystrobin, dinotefuran, and permethrin in the lettuce heads were 5, 30, 25, and 3 mg/kg, respectively).
We observed no clear differences in the residue levels of acetamiprid, azoxystrobin, and permethrin in lettuce heads between the two growing conditions. As compared to the other three pesticides applied to the foliage, a clear difference in the two growing conditions was observed when dinotefuran, the pesticide with the highest water solubility and the longest interval between application and harvest, was applied to the planting hole (excluding a single set of samples from the Koch field). This result suggests that the influence of the growing condition on pesticide residue levels may be smaller than the effect of variations in residues caused by different application methods.
4. Tendency of pesticide residues to declineOne of the important objectives of crop field trials is to determine the rate of decline of the pesticide residue to conduct a dietary risk assessment.16) However, it was difficult to evaluate a tendency to decline (or increase) based on the residue data from each field. The statistical analytical results of the Mann–Whitney U-test indicate that there is a significant difference between the residue data in lettuce heads grown in greenhouses and open fields only for dinotefuran, while no significant difference was confirmed between the residue data in the different growing conditions for the other three pesticides applied to foliage (Supplemental Table 3).
The residue levels of acetamiprid, azoxystrobin, and permethrin in the lettuce heads from the greenhouses were almost as high as those from the open fields in the same location for 7 days after the final application (excluding acetamiprid in a sample from the Gunma field and permethrin in a sample from the Nagano field). Therefore, regarding the tendency of pesticide residues to decline (or increase) over time, the residue levels of four pesticides in the lettuce heads are plotted against the days after final application in Fig. 1. The results, based on the overall mean residue levels of pesticides, indicate that the residue levels of pesticides correlated negatively with the intervals between pesticide application and harvest. There were approximate correlations between the overall mean residue levels of all investigated pesticides in the lettuce heads and the days after final application (R2>0.6956).
Regression line slopes for each residue dataset revealed decline tendencies of pesticides under the different growing conditions. Dinotefuran showed the largest difference between the tendency to decline in the open fields and in greenhouses. This difference reflects the difference in the method of application and the number of days after final application, as described in the previous section. In contrast, the residue dataset of permethrin showed that the tendency of the pesticide to decline was similar in open fields and greenhouses. This result is due to the physicochemical properties of permethrin, which has relatively low water solubility (6 µg/L) and vapor pressure (approx. 2×10−4 mPa) and is relatively stable to light in field situations.14) Therefore, the influence of the greenhouse was not observed on the residue levels of permethrin. The other two pesticides, acetamiprid and azoxystrobin, with intermediate physicochemical properties, showed an intermediately different tendency to decline in the open fields and in greenhouses.
Fantke and Juraske6) reported that the variability of the decline tendency due to growing conditions is mainly a function of substance solubility where, for soluble pesticides, higher temperatures in greenhouses reduce their dissipation half-lives, while for less soluble pesticides, wash-off and leaching into the soil are reduced, thereby leading to increased half-lives. In this study, the decline tendency of soluble pesticides, such as dinotefuran, in greenhouses was attenuated, while there is no clear difference in residue levels between the two growing conditions for less soluble pesticides, such as permethrin. These incompatible results indicate that complex factors affect the pesticide residue levels in different growing conditions.
5. Effect of sample size reduction on residue dataThe percentages of the mean residues of Sample A/Sample B (A/B) and the coefficient of variations (CV) are summarized in Supplemental Table 2. The largest coefficient of variation value in size-reduced samples of 78% was observed for the residue levels of azoxystrobin in lettuce heads from the Gunma field 7 days after final application; the A/B value was 44%. The maximum difference in residue levels of azoxystrobin between two size-reduced samples from the same lettuce head was obtained from a head in which one sample was 2.3 times greater than the other one (0.64 mg/kg / 0.28 mg/kg).
| Growing conditionsa) | Residue (mg/kg)b) | RSD (%) | ||||
|---|---|---|---|---|---|---|
| LR | HR | HR/LR | Median | Mean | ||
| Acetamiprid | ||||||
| O/1-d | 0.08 | 3.95 | 49.4 | 1.62 | 1.58 | 91 |
| G/1-d | 0.90 | 3.64 | 4.0 | 1.94 | 2.04 | 46 |
| O/3-d | 0.24 | 2.34 | 9.8 | 0.92 | 1.12 | 77 |
| G/3-d | 0.29 | 2.01 | 7.1 | 1.61 | 1.44 | 44 |
| O/7-d | 0.09 | 1.84 | 20.4 | 0.53 | 0.64 | 97 |
| G/7-d | 0.39 | 2.45 | 6.3 | 1.00 | 1.14 | 63 |
| Azoxystrobin | ||||||
| O/1-d | 0.02 | 2.38 | 119.0 | 0.85 | 0.92 | 98 |
| G/1-d | 0.53 | 2.40 | 4.5 | 1.12 | 1.22 | 56 |
| O/3-d | 0.08 | 1.50 | 18.8 | 0.82 | 0.77 | 79 |
| G/3-d | 0.13 | 1.36 | 10.9 | 0.84 | 0.80 | 53 |
| O/7-d | 0.03 | 0.94 | 31.3 | 0.36 | 0.37 | 92 |
| G/7-d | 0.39 | 1.89 | 4.8 | 0.55 | 0.73 | 77 |
| Dinotefuran | ||||||
| O/49–70-d | 0.02 | 0.11 | 5.5 | 0.04 | 0.05 | 69 |
| G/48–69-d | 0.12 | 0.74 | 6.2 | 0.40 | 0.44 | 58 |
| O/51–72-d | 0.02 | 0.10 | 5.0 | 0.04 | 0.05 | 59 |
| G/50–71-d | 0.09 | 0.64 | 7.5 | 0.54 | 0.42 | 62 |
| O/55–76-d | 0.01 | 0.10 | 10.0 | 0.04 | 0.05 | 75 |
| G/54–75-d | 0.08 | 0.64 | 7.9 | 0.41 | 0.39 | 59 |
| Permethrin | ||||||
| O/1-d | 0.03 | 1.81 | 60.3 | 0.87 | 0.82 | 83 |
| G/1-d | 0.45 | 2.27 | 5.0 | 0.93 | 1.08 | 61 |
| O/3-d | 0.24 | 1.48 | 6.2 | 0.89 | 0.90 | 57 |
| G/3-d | 0.12 | 1.17 | 9.7 | 0.76 | 0.68 | 51 |
| O/7-d | 0.18 | 1.12 | 6.2 | 0.33 | 0.47 | 76 |
| G/7-d | 0.34 | 1.49 | 4.4 | 0.44 | 0.59 | 75 |
a) Growing conditions of lettuce heads: open fields (O) or greenhouses (G) with the days after each of final application.b) Lowest residue (LR), highest residue (HR), median residue, mean residue, and relative standard deviation (RSD).
Box plots of the percentages of difference in the residue levels of pesticides between size-reduced sample-A and sample-B (A/B) in greenhouses are shown in Fig. 2. The box plots clearly show that the range of A/B values of dinotefuran (72–120%) was narrower than the values of the other three pesticides (44–178%). Acetamiprid has a relatively low log POW of 0.80; however, the range of A/B values of acetamiprid was at the same level as the results of the other two pesticides.
The narrow range of dinotefuran residue among the size-reduced samples may reflect the application method of pricking the pesticide into each planting hole. In contrast, the other pesticide formulations were directly sprayed on the lettuce heads. Therefore, the days after applications of dinotefuran were 48–75 days, significantly longer than those of the other pesticides (1–7 days). The distributions of pesticide residues in individual crop units usually depend on the intervals between application and harvest, route of absorption, and penetration and translocation abilities of the pesticides; therefore, we expect the distribution of dinotefuran residues in lettuce heads following soil treatment to be more uniform than that of other pesticide residues, which have shorter intervals between application and harvest following foliage application. In contrast, the majority of sprayed pesticides were present on the surface of outer leaves, especially in wrapped leafy vegetables such as lettuce heads and cabbage.7,17) The assumption just described is supported by our previous study,8) in which the Starcle® water-dispersible granules of dinotefuran were applied to Chinese cabbage using sprayers, and the variations in residue levels of dinotefuran in size-reduced Chinese cabbage were at the same level as the other pesticides.
As explained above, the size reduction process for sample preparation affected the variations in this residue data. However, the differences in the pesticide residue levels among the size-reduced samples were significantly smaller than were the differences due to field-to-field variability described in the above section.
We conclude that the present study demonstrates that the pesticide residue levels in lettuce heads under different growing conditions were influenced by complex factors, such as differences in pesticide application methods, intervals between pesticide application and harvest, and physicochemical properties of the pesticides. For example, the residue levels of dinotefuran with application to the planting hole were higher in lettuce heads grown in greenhouses than in those grown in the open fields. Although the investigation was conducted on limited pesticides and growing conditions, the results indicate that weather conditions may affect the pesticides through runoff by rainfall. In addition, the ranges between two size-reduced samples of dinotefuran applied to the planting hole were narrower than were the ranges of values of the other three pesticides that were applied to the foliage.
We thank the members of the committee for this research (Dr. M. Ueji, Dr. Y. Ishii, and Dr. K. Nakamura) for their valuable suggestions and comments. We also thank the staff at the Japan Plant Protection Association for their cooperation in the field experiments. This study was supported by a grant from the Japanese Ministry of Agriculture, Forestry and Fisheries in 2009 and 2010.
