aoac study director: guo-fang pang academy of chinese inspection and quarantine

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High-Throughput Analytical Method for Determination of multi-Classes and multi-Kinds of 668 Residue Pesticides and Chemical Pollutants in Tea by GC-MS, GC-MS/MS and LC-MS/MS — AOAC Collaborative Study Protocol (Draft). AOAC Study Director: Guo-Fang Pang - PowerPoint PPT Presentation

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  • *High-Throughput Analytical Method for Determination of multi-Classes and multi-Kinds of 668 Residue Pesticides and Chemical Pollutants in Tea by GC-MS, GC-MS/MS and LC-MS/MS AOAC Collaborative Study Protocol (Draft)AOAC Study Director: Guo-Fang PangAcademy of Chinese Inspection and QuarantineEmail:qhdciqgfpang@yahoo.com.cnCall for Voluntary Collaborators Priority Project of AOAC INTERNATIONAL (2010) Nov. 27, 2010)

  • *1. Purpose of AOAC collaborative study There are more and more and stricter and stricter limit standards for pesticide residues in edible agricultural products in countries all over the world. As everyone knows, the integration of global economy is stepping up, and the strategic positions of the world food safety and pesticide residue limit threshold for international trade are getting higher and higher, which calls for high throughput residue analytical techniques.Comparison of limit standards for pesticides residues in different countries ( Data from 2010)Source of literature: http://www.tbtsps.com/foodsafe/xlbz/Pages/pesticide.aspx The threshold of limit for pesticide residues in tea is high for international trade. At present, there are 17 countries and international organizations that have stipulated MRL levels for over 800 pesticide residues in tea.EU, Germany, Holland, Switzerland, Hungry, Israel, CAC, China, Japan, Chinese Taiwan, Korea, USA, Australia, India, Kenya and South Africa (till 2006).Comparison of pesticide quantities with MRL for different countries USA2001 EU2010 Japan2010 Germany2001 CAC2009 China2005Source of literature: http://ec.europa.eu/sanco_pesticides/public/index.cfmhttp://www.m5.ws001.squarestart.ne.jp/foundation/search.htmlhttp://www.tbtsps.com/foodsafe/xlbz/Pages/pesticide.aspxGB 2763-2005Doctoral Dissertation of Anhui Agricultural University Wu Xueyuan MRL of Pesticides in Tea and Risk Evaluation

  • *2. Application scope of the methodDistribution of pesticide varieties determined by GC-MS & LC-MS/MSLC-MS-MS &GC-MS270LC-MS-MS178GC-MS220LC-MS-MS 448GC-MS 490 The method is applicable to the determination of multi-classes and multi-kinds of 668 residue pesticides and chemical pollutants in Green tea, Red tea, Puer tea and Woolong tea by GC-MS, GC-MS/MS & LC-MS/MS. The limits of detection (LOD) for 490 pesticides determined by GC-MS were between 1.0-500 g/kg, and for 448 pesticides determined by LC-MS/MS were between 0.03-4820 g/kg. There are 482 pesticides with LOD100g/kg for GC-MS method, accounting for 98% of the pesticides tested and there are 417 pesticides for LC-MS/MS, accounting for 93% of the pesticides analyzed; there are 264 pesticides with LOD10g/kg for GC-MS method, accounting for 54% of the pesticides tested, 325 pesticides for LC-MS/MS method, making up 73% of the pesticides analyzed. There are 270 pesticides that can be analyzed by both GC-MS and LC-MS/MS. There are 264 pesticides with LOD100g/kg for GC-MS method, accounting for 98% of the pesticides tested, 247 for LC-MS/MS, making up 91% of the pesticides analyzed; there are, however, 133 pesticides with LOD10g/kg for GC-MS method, accounting for 49% of the pesticides tested, 200 pesticides for LC-MS/MS method, making up 74% of the pesticides analyzed.

  • *3. Analytical proceduresLC-MS-MSGC-MS-MSGC-MS-SIMCentrifugalizeConcentrationHomogenizeSampler vialClean-upConcentration ConcentrationEvaporate with nitrogen DetermineHomogenize at 13500r/min for 1min;Centrifuge at 4200r/min for 5min;Concentrate to ca 1mL at 40;Elute by gravity for clean-up;Concentrate to ca 0.5mL at 40 ; Evaporate to dryness with nitrogen gas at 35 Filter through 0.2m membrane

    The key control points

  • *5g tea sampleAdd 15 mL acetonitrile Add IS Heptachlor epoxide for GC/MS-SIM, GC/MS/MS or Chlorpyrifos-methyl for LC-MS/MS. Evaporate to dryness with nitrogen at 35Cleanert TPT Cartridge(2g,12mL) or Envi-Carb/PSA(0.5g/0.5g,6mL) for clean up. Elute with 25 mL acetonitrile+toluene(3+1), concentrated to about 0.5 mLMake up to 1.5 mL with hexaneMake up to 1.5 mL with acetonitrile+water(3+2)GC/MSDetermination(490 Varieties)GB/T 23204-2008LC-MS/MSDetermination(448 Varieties)GB/T 23205-20083. GC-MS(GC-MS/MS)&LC-MS/MS Analysis programStudy Director would like to remind all the participants, as a binding rule for voluntary participation: this inter-collaborative study is a unified and standardized action for all the collaborators from different countries all over the world, and each step involved in the experiment including reagents and testing materials shall not be altered or replaced without the prior consent of Study Director.Homogenize at 13500r/min for 1min, and centrifuge at 4200r/min for 5 min before the supernatants are taken, while the dregs are re-extracted and the extracts are combined and then concentrated to about 1 mL

  • *4. Shrunken AOAC collaborative study protocol

    It will be extremely difficult in terms of resources, time and personnel for each participant to organize a collaborative study on hundreds of pesticides, so AOAC International has proposed an alternative shrunken protocol, which is to select two teas and two methods of GC-MSGC-MS/MS and LC-MS/MS for a respective determination of 20 pesticides totaling 40 pesticides to be involved in the collaborative study. * Indicates the pesticides to be determined by both GC-MS (GC-MS/MS) and LC-MS/MS4.1 20 pesticides for GC-MS(GC-MS/MS) and LC-MS/MS determinations20 pesticide classes for GC-MS(GC-MS/MS) detemination 20 pesticide classes for LC-MS/MS detemination

    NoGC-MSLC-MS/MSPesticidesMRL/ (g/kg)PesticidesMRL/ (g/kg)12,4'-DDE200phenothrin502benalaxyl100imidacloprid503bromophos-ethyl100tebufenpyrad1004bromopropylate100propoxur1005chlorfenapyr50000bensulide306diflufenican50picoxystrobin1007dimethenamid20chlorpyrifos1008fenchlorphos100diazinon209picoxystrobin100flutolanil5010pirimicarb50pirimiphos methyl5011pirimiphos-methyl50fenazaquin1012bifenthrin5000ethoprophos2013propyzamide50triadimefon20014pyrimethanil100pyriproxyfen5015Quinoxyfen50trifloxystrobin5016tebufenpyrad100kresoxim-methyl10017tefluthrin50acetochlor1018tolclofos-methyl100clomazone2019trifluralin100indoxacarb5020vinclozolin100butralin20

    classified per chemical compositions classified per functionsCarbamate1Acaricide2Organophosphorus4Herbicide4Organohalogen2Insecticide8Pyrethroid2fungicide6Organonitrogen9Other2

    classified per functions classified per chemical compositions Carbamate2Insect growth regulator1Organophosphorus5Plant growth regulator1Organonitrogen11Acaricide2Other2Herbicide3Insecticide8fungicide5

  • *4. Shrunken AOAC collaborative study protocol Each sample is only sufficient for one test, in other words, each participant will have only one testing opportunity for each sample, so he/she will have to make every effort to ensure an utmost success with one shot. Therefore, Study Director will emphasize again that this collaborative study must be dealt with full focus and attention. The compositions of these 16 collaborative study samples are as follows:The concentration range of 20 target pesticides is 10ug/kg-2.0mg/kg equivalent to 1MRL-5MRL.Fortified samples and pesticide aged samples are designed in Youden Pair, with two samples for one pair and the difference of the concentration of the target pesticide is about 5%.*The serial number here represents the design concept of the collaborative study protocol only, the actual serial number of each sample shipped will be determined later. 4.2 Each participating laboratory makes a determination of a total of 16 collaborative study samples.

  • *5. The flow chart of the AOAC collaborative study Excel Worksheet

  • *Step-I Testing materials shipped by Study Director2 ampoules (1.0mL x 2, 1 spare aged) of 20 pesticide mixed standard stock solutions for establishing at least 5 point matrix-matched internal standard calibration curve.2 ampoule (1.0mL x 2, 1 spare aged) of internal standard stock solutions.10 ampoules (0.5mL x 10) of fortified solutions for preparation of fortified samples.10 bags of tea blank samples(5.0 g x 10): No. 01-No.05 is green tea; No.06-No.10 is woolong tea for preparation of fortified samples.6 bags of aged tea samples (5.0 g x 6): No.11-No.13 is green tea; No.14-N0.16 is woolong tea.1 bottle of green tea and woolong tea blank samples each (200g x 2) for preparation of matrix-matched internal standard calibration mixed solutions.(1)For Pre-collaborative study20 pesticide mixed standard stock solutions: 2 ampoules (1.0mL X 2, 1 spare aged).Internal standard stock solutions: 2 ampoules (1.0 mL X 2, 1 spare aged).Green tea and woolong tea blank samples: 1 bottle each (200g x 2).Examination sample materials: 3 ampoules of fortified solutions (0.5mL x 3) for preparation of examination samples. 3 bags of green tea blank samples (5.0g x 3) for preparation of examination samples. 3 bags of woolong tea aged samples (5.0g x3).(2)For Collaborative study(3)For Quality Control Quality control standard stock solutions: 2 ampoules (1.0mL X 2, 1 spare aged), used for checking the instrumental sensitivity and stability.

  • *Step-II The significance Because of the complexity of tea matrices, the magnitude of content of the research and the nature of technology involved, this collaborative study is by no means easy to be undertaken, and the objective of this Pre-collaborative study is well explained in the remarks of AOAC Sr. Director, Stakeholder Communications Krystyna McIve pre-collaborative study by the collaborators to ensure the laboratories are qualified to run the method. Step-II Pre-collaborative studyStudy Director reminds everyone here once again: For each of these 16 collaborative study samples, each participant will have only one testing opportunity, which means success or f