[Simultaneous Determination of Pesticide Residues in Brown Rice Using GC-MS/MS and LC-MS/MS].

This study developed a method that simultaneously detected 283 pesticide residues in brown rice using GC-MS/MS and LC-MS/MS. In this method, we examined the desirable amount of sodium chloride required for salting out and the SPE cartridge required for clean-up. Pesticide residues from the sample were extracted with acetonitrile using a homogenizer and mixed with salts including anhydrous magnesium, two types of citrate, and sodium chloride. The sample solution of the acetonitrile layer was cleaned up using the GCB/NH2 (200 mg/200 mg, 6 mL) SPE cartridge. The determination method was validated using two concentrations (0.01 and 0.1 µg/g) of 283 pesticides based on the validation guideline of the Ministry of Health, Labor and Welfare in Japan. Of the 283 pesticides, 250 were detected satisfactorily. In addition, 59 brown rice samples sold in Tokyo were surveyed using the same method. Out of 44 samples, 12 pesticide residues below MRLs were detected. Therefore, this developed method is useful for the simultaneous determination of pesticide residues in brown rice.

[Evaluation on a Simultaneous Analytical Method of Pesticide Residues in Limes].

A LC-MS/MS simultaneous analytical method for screening 191 pesticide residues in limes had been developed and validated. Pesticides were extracted with acetonitrile from samples. Then mixed salts, which were anhydrous magnesium sulfate for dehydration, sodium carbonate for adjusting pH, and sodium chloride for salting out, were added to the sample. After centrifugation, supernatant was transferred to a tube. The sample solution was cleaned up using solid phase extraction (SPE) with C18/GC/PSA for the determination by LC-MS/MS. The developed method was improved the recovery rate of thiabendazole, which had a low recovery rate by the conventional method. Validation study, which was following the guidelines of the Ministry of Health, Labor and Welfare, were carried out at 0.01 and 0.1 µg g-1 to evaluate the method. The results of 175 pesticides out of 191 were with satisfactory. A total of 19 imported lime samples sold in Tokyo was analyzed to evaluate the method, then 18 samples contained pesticide residues below MRLs. The developed method is applicable for detection of pesticide residues in lime.

[Comparison of Validation Results Obtained by GC-MS/MS and LC-MS/MS for the Analysis of Residual Pesticides in Agricultural Products].

In the field of food analysis and regulation, different instruments are used to determine the accuracy of quantification values. This is essential, as inconsistencies in values are commonly encountered. To visualize the degree of these discrepancies in each food matrix, we compiled a validation study based on a routine method developed in our laboratory, for 121 pesticides in six agricultural products, namely the grapefruit, potato, paprika, cabbage, spinach, and brown rice. These were analyzed by GC-MS/MS and LC-MS/MS, and the results were compared mainly on the basis of trueness. According to the results of the validation study when using GC-MS/MS, of the 121 pesticides tested in each product class, the number of analytes that satisfied the criteria of the Japanese validation guidelines was 97 in grapefruit, 111 in potato, 110 in paprika, 118 in cabbage, 111 in spinach, and 63 in brown rice. In contrast, in the analysis of the same samples by using LC-MS/MS, the number of analytes that satisfied the criteria of the validation guidelines was 50 in grapefruit, 114 in potato, 103 in paprika, 112 in cabbage, 100 in spinach, and 103 in brown rice. Inconsistences in the differences of trueness were mainly attributed to matrix effects of each instrument, as well as to food matrices, of which the most diverged matrix was that of brown rice (over 20%).

[Validation of a Method for Determining Multiple Residual Pesticides in Dried Red Pepper Using LC-MS/MS].

An analytical method has been developed and validated for determining 107 pesticide residues in dried red pepper using LC-MS/MS. LC method, the clean-up and sample dilution processes were examined to determine their impact on reducing the matrix effects. Clean up was performed using an ENVI-CarbIITM/PSA (300/600 mg, 6 mL) SPE cartridge. In the sample dilution process, eight-fold dilution was used. In the validation of the developed method at two concentrations (0.01 and 0.1 µg/g) for 107 pesticides, 96 pesticides showed recovery rates in the range of 70.1 to 112.6%, RSDs of repeatability of ≤11.5 and 3.4%, and RSDs of within-laboratory reproducibility of ≤24.3 and 19.9%. These values fulfill the criteria of the validation guidelines for pesticide residues in Japan. It is concluded that matrix effects and low recovery rates in the process of extraction are the main factors for values that do not conform to the criteria.

Surveillance of Chlorantraniliprole Residues in Vegetables and Fruits Using LC-MS/MS.

We measured the residual amounts of chlorantraniliprole in various vegetables and fruits. Sample solutions were prepared according to our routine procedure based on the QuEChERS method and analyzed by LC-MS/MS. Performance characteristics were evaluated for 8 kinds of food samples by means of recovery tests of 5 replicates at the concentration of 10 ng/g. Recoveries and RSDs (％) ranged from 50.2 to 93.4％ and from 2.1 to 9.7％, respectively. Application of this method to survey 207 vegetables and 163 fruits gave detection rates of 8.2 and 1.2％, respectively. In vegetables, detection rates were high in okra (4 out of 10 samples), paprika (4 out of 23 samples) and tomato (2 out of 6 samples), and leaf vegetables such as lettuce, mizuna, spinach and wrinkled greens also contained high concentrations of chlorantraniliprole. The highest residual concentration was 571 ng/g in mizuna. The samples containing chlorantraniliprole seemed to be mainly from Asian countries, including samples of domestic Japanese origin. However, none of them contained more than the MRL, which suggests that the use of chlorantraniliprole has been properly controlled.

[Determination of hymexazol in agricultural products by GC-NPD].

A method for the determination of hymexazol in agricultural produdcts by gas chromatography with a highly sensitive nitrogen-phosphorus detector (GC-NPD) was investigated. Hymexazol was extracted with acetonitrile, and the acetonitrile layer was separated by salting-out. The water layer was loaded onto a Chem-Elut column. Hymexazol in the water layer was adsorbed on the column, and eluted with ethyl acetate. The acetonitrile layer and the eluate were mixed and evaporated. The residue was dissolved in ethyl acetate, and the sample solution was cleaned up on a C18 column. Hymexazol in the eluate was analyzed by GC-NPD with a high-polarity capillary column (DB-FFAP) and highly deactivated inlet liner. Recoveries of hymexazol spiked in agricultural products (tomato, lemon, soybean and other samples) at the level of 0.1 mug/g ranged from 65.0 to 84.7%. The limit of detection was 0.02microg/g.

[Survey of pesticide residues in imported cereal products (1994.4 approximately 2006.3)].

A survey of pesticide residues in 490 imported cereal products on the Tokyo market from April 1994 to March 2006 was carried out. Eight kinds of organophosphorus pesticides (chlorpyrifos, chlorpyrifos-methyl, DDVP, diazinon, etrimfos, malathion, MEP and pirimiphos-methyl) were detected at levels between Tr (below 0.01 ppm) and 0.82 ppm from 91 samples. In our investigations, chlorpyrifos-methyl and malathion tended to be detected in samples from America, pirimiphos-methyl in those from Europe, and MEP in those from Oceania. Thus, pesticide residues seemed to be different in produce from different areas. Residue levels of these pesticides were calculated as between 0.08 and 13.2% of their ADI values according to the daily intake of cereal products. Therefore, these cereal products should be safe for normal usage.

[Survey of pesticide residues in rice (April, 1995-March, 2005)].

Pesictide residues in 343 samples of domestic rice and 32 samples of imported rice purchased on the Tokyo market from April 1995 to March 2005 were investigated. Residues of eleven kinds of pesticides (including organophosphorus, organochlorine, carbamate, pyrethroid and organonitrogen pesticides and bromide) were detected at levels between trace and 1 ppm in 47 domestic rice samples. DDVP and bromide residues were detected at levels between 0.01 ppm and 5 ppm in 18 imported rice samples. For rice samples that were found to containing pesticides, the amounts of pesticide intake were calculated according to the daily intake of rice, and they were compared with the ADI values. Since residual pesticide levels were 17/10,000-2/5 of ADI, all the rice samples were considered to be safe for human consumption.

Clean-up method of forchlorfenuron in agricultural products for HPLC analysis.

A simplified method for the determination of forchlorfenuron in agricultural products by HPLC with UV detection was investigated. A chopped sample homogenate from agricultural products was extracted with acetone. The extract was filtrated and concentrated. The residues was loaded onto a Chem Elut column and extracted with ethyl acetate. The crude extract was purified on Oasis HLB and Bond Elut PSA mini-columns using a mixture of methanol and ethyl acetate. Forchlorfenuron was analyzed by HPLC with UV detection (263 nm). HPLC separation was performed on an ODS column with methanol-water as the mobile phase. Recoveries of forchlorfenuron from several agricultural products fortified at the level of 0.1 microg/g were in the range of 87.6-99.5%. The limit of detection (S/N=3) was 0.005 microg/g in the sample.

[Survey of pesticide residues in baby foods (1999.4-2005.6)].

A survey of pesticide residues in 316 baby foods on the Tokyo market from April 1999 to June 2005 was carried out. Fifteen kinds of pesticides, including organophosphorus, organochlorine, carbamate, pyrethroid and others, were detected between at levels Tr (below 0.01 ppm) and 0.55 ppm from 28 samples. The baby foods in which residual pesticides were detected were produced from flour, leaf vegetable and fruits. The pesticides were detected in baby foods produced with minimal processing, such as straining and squeezing. For the baby foods containing pesticides, the amounts of intake of the pesticides were calculated from the recommended servings, and compared with the ADI values. Residual pesticide levels in baby food were between 0.06 and 16.6% of the ADI. Therefore, these foods should be safe in normal usage.

Rapid Multi-Residue Analysis of Pesticides in Pulses by LC-MS/MS.

Rapid multi-residue analysis of pesticides in pulses was developed using LC-MS/MS. Pesticide residues in 5 g of homogenized pulses were extracted with 30 mL of acetonitrile and salted out with 4 g of anhydrous magnesium sulfate and 2 g of sodium chloride in the presence of citrate buffer in a disposable tube. The resulting residues were extracted with 30 mL of acetonitrile, and co-extractives were removed on a handmade four-layer column, consisting of a layer of Z-Sep/C18 (20 mg/50 mg) dry particles on top of a three-layer, custom-made (pre-packed) column (lower bed: 60 mg of PSA, middle bed: 30 mg of GC, and top bed: 60 mg of C18) packed in a 10 mm internal diameter polypropylene column (3 mL). The developed method showed good recoveries of pesticides in soybean, lentil, white kidney bean and garbanzo. According to the method validation guideline of the Ministry of Health, Labour and Welfare of Japan, recovery tests were conducted in soybeans fortified with 107 kinds of pesticides at the levels of 0.01 and 0.1 µg/g, respectively. At each concentration 2 samples were extracted on 5 separate days. Pesticides in the test solution were determined by LC-MS/MS using scheduled MRM. As regards the trueness of this method for 107 pesticides in soybeans, 97 pesticides were in the range of 70-120% with satisfactory repeatability and within-run reproducibility. This new method is expected to be applicable for routine examination of pesticide residues in soybeans.

[Simple analysis of maleic hydrazide in agricultural products by HPLC].

A simplified HPLC determination method for maleic hydrazide in agricultural products was developed, and commercial agricultural crops were investigated. The homogenate of agricultural products was extracted with water. The crude extract was purified on an ACCUCAT Bond Elut extraction cartridge using water. Maleic hydrazide was analyzed by HPLC with UV detection (303 nm). The HPLC separation was performed on a ZORBAX SB-Aq column with acetonitrile-water-phosphoric acid(5:95:0.01) as the mobile phase. Recoveries of maleic hydrazide from 15 agricultural products fortified at 1.0 and 10 micrograms/g were in the ranges of 92.6-104.9% and 94.2-101.3%, respectively. The limit of detection was 0.5 microgram/g in samples. The proposed method was applied to the determination of 242 commercial vegetables and fruits. Maleic hydrazide was detected in 2 samples of imported onion at the levels of 4.9 and 7.2 micrograms/g.

[Identification of chemical detected in kiwi fruit during analysis for residual pesticide].

An unknown peak was detected in a GC chromatogram of many kiwi fruit extracts during analysis for pesticide residues. It was identified by GC/MS as diphenyl 2-ethylhexyl phosphate (DPEHP), used as a plasticizer and flame retardant. The concentration of DPEHP was investigated in 15 samples of kiwi fruit, and it was detected at between 0.02 and 0.14 microgram/g in 10 of the samples. It might be due to migration of DPEHP into the fruit from the printed portion of the polyethylene terephthalate (PET) package.

[Determination of captafol, cyhexatin, 1-naphthylacetic acid and quintozene in apple, Japanese pear and melon by simultaneous extraction].

A simple and rapid method is described for the determination of the non-registered pesticides, captafol, quintozene (PCNB), cyhexatin and 1-naphthylacetic acid (NAA), in fruits. These pesticides were extracted with acidified acetone, then captafol and PCNB were purified with a Florisil mini column and analyzed by GC-ECD. Cyhexatin was ethylated with ethylmagnesium bromide, and the ethyl derivative was analyzed by GC-FPD (Sn filter). NAA was purified with liquid-liquid extraction and determined by HPLC equipped with a fluorescence detector. These analytes were identified with GC/MS or LC/MS. The minimum identified concentration of the pesticides was below 0.2 ng per injection, which corresponds to a detection limit of below 0.02 microgram/g in the original samples. Recoveries of the pesticides spiked at 0.1 microgram/g into apple, Japanese pear and melon were greater than 61%.

[Analysis of pesticides including chlorine in welsh onions and mushrooms using gas chromatograph with an atomic emission detector (GC-AED)].

An analytical method for the determination of 32 kinds of pesticide residues in onions, Welsh onions and mushrooms using gas chromatograph with an atomic emission detector (GC-AED) was developed. The pesticides were extracted with acetone-n-hexane (2:3) mixture. The crude extract was partitioned between 5% sodium chloride and ethyl acetate-n-hexane (1:4) mixture. The extract was passed through a Florisil mini-column for cleanup with 10 mL of acetone-n-hexane (1:9) mixture. Although the sensitivity of GC-AED was inferior to that of GC-ECD, GC-AED has a superior element-selectivity. Therefore pesticide residues in foods could be analyzed more exactly by using GC-AED. Thirty-two pesticides including chlorine in onion, Welsh onion and shiitake mushroom were detected without interference. Recoveries of these pesticides from samples determined by GC-AED were 64-114%, except for a few pesticides.

[Identification of unknown peaks in foods during residual pesticide analysis].

Four unknown peaks were detected in GC chromatograms during analysis of organophosphorus pesticide residues in foods. They were identified by using GC/MS as triethyl phosphate (TEP), tributyl phosphate (TBP), diphenyl 2-ethylhexyl phosphate (DPEHP) and N-ethyltoluenesulfoneamide (NETSA), which are used as plasticizers or flame retardants in food packaging. These chemical substances were detected in the range of tr. (below 0.01 microg/g) to 11 micro/g from 29 samples, and they were also detected in the packaging. It was supposed that they were transferred to the foods from the packaging. Furthermore, they were detected in some cereals and cereal products which contain fat and had been preserved for a long time.

[Studies on rapid multi-residue analysis of pesticides in agricultural products by LC-MS/MS].

Rapid multi-residue analysis of pesticides in agricultural products was studied by using LC-MS/MS. Pesticide residues in 10 g of homogenized agricultural products were extracted with 30 mL of acetonitrile and salted out with 4 g of anhydrous magnesium sulfate and 1 g of sodium chloride in the presence of citrate salts for buffering in a disposable tube. Co-extractives were removed by use of our original triple layered column (C18/GC/PSA; 60/30/60 mg). According to the method validation guideline of the Ministry of Health, Labour and Welfare of Japan, we conducted recovery tests in 5 kinds of agricultural products (brown rice, kiwi, cabbage, sweet potato and spinach) spiked with 60 pesticides at the level of 0.01 or 0.1 µg/g. Each concentration of pesticide spiked was extracted from 2 samples per day on 5 days. Pesticides in the test solution were determined by two types of LC-MS/MS using scheduled MRM. Using this method, 58 out of 60 pesticides satisfied the guideline criteria in brown rice, 59 in kiwi, 55 in cabbage, 55 in sweet potato and 56 in spinach. This method is applicable for routine examination of pesticide residues in agricultural products.

[Survey of pesticide residues in imported tea (1992.4-2010.3)].

A survey of pesticide residues in 116 imported tea samples on the Tokyo market from April 1992 to March 2010 was carried out. Twenty-two kinds of pesticides, including organophosphorus, organochlorine, pyrethroid and others, were detected at levels between trace (below 0.01 ppm) and 4.0 ppm in 76 samples. The rate of detection was 90% in non-fermented tea, 89% in semi-fermented tea, and 49% in fermented tea. Organophosphorus pesticide was not detected in puer tea, which is fermented with bacteria. The pesticide residues tended to be decreased by a fermentation process. However, organochlorine and pyrethroid pesticides appeared to be chemically stable, so they were still detected in fermented teas. Residue levels of these pesticides were calculated as less than 1% of ADI, except for ethion (45% of ADI) based on the daily intake of tea. Therefore, these teas should be safe when drunk in customary amounts.

[Survey of pesticide residues in imported spices and herbs (1997.4-2011.3)].

A survey of pesticide residues in 313 samples of imported spices and herbs on the Tokyo market from April 1997 to March 2011 was carried out. Thirty-seven kinds of pesticides, including organophosphorus, organochlorine, pyrethroid, carbamate and others, were detected between levels of trace (below 0.01 ppm) and 3.3 ppm from 64 samples. The rate of detection was highest in peel (100%) followed by stem (66.7%), fruit (34.5%), bark (33.3%), flower (31.3%) and leaf (14.7%). No residues were detected in root, seed or whole grass. Organochlorine pesticides were detected in all plant parts. The insecticides were detected in products from all production areas, suggesting that their use is common. Residue levels of these pesticides were calculated as less than 1% of their ADI values, based on the daily intake of spices and herbs. Therefore, these spices and herbs should be safe when consumed in customary amounts.

[Survey of pesticide residues in imported frozen vegetables and fruits (1989.4~2008.3)].

A survey of pesticide residues in 595 imported frozen products on the Tokyo market from April 1989 to March 2008 was carried out. Forty three kinds of pesticides, including organophosphorus, organochlorine, carbamate, pyrethroid and others, were detected between levels of trace (below 0.01 ppm) and 4.6 ppm from 162 samples. Chlorpyrifos, cypermethrin and omethoate were frequently detected in green vegetables (komatsuna leaf and spinach), cypermethrin and methamidophos were detected in pods and seeds (green soybean and string pea), chlorpropham (CIPC) was detected in potato, and captan and carbaryl were detected in berries (blueberry, raspberry and strawberry). The hydrophilic pesticide methamidophos was detected in flesh of lychee. Residue levels of these pesticides were calculated as between less than 0.5% and 30% of their ADI values according to the daily intake of frozen products. Therefore, these frozen products should be safe when they were eaten in customary amounts.