Analysis of Pesticide Residues in Apple Using Agilent Bond Elut QuEChERS AOAC Kits by GC/MS

Similar documents
Anaylsis of Pesticide Residues in Rice Using Agilent Bond Elut QuEChERS AOAC Kit by LC-MS/MS Detection

Analysis of Pesticide Residues in Apple using Agilent Bond Elut QuEChERS European Standard EN Kits by LC/MS/MS Detection

LC/MS/MS Analysis of Pesticide Residues in Apples Using Agilent Chem Elut Cartridges

Agilent Bond Elut QuEChERS Food Safety Applications Notebook: Volume 2 INNOVATIVE APPROACHES FOR TODAY S FOOD ANALYSIS CHALLENGES

Pesticides Analysis Using the Agilent 5977A Series GC/MSD

Macrolides in Honey Using Agilent Bond Elut Plexa SPE, Poroshell 120, and LC/MS/MS

Application Note. Abstract. Authors. Introduction. Food Safety

Determination of Pesticide Residues in Oats by Automated. QuEChERS and LC/QQQ. Application Note. Abstract. Introduction

Rapid Screening and Confirmation of Melamine Residues in Milk and Its Products by Liquid Chromatography Tandem Mass Spectrometry

Fully automated QuEChERS clean-up and LC/MS-QQQ analysis of pesticides in fruits and vegetables.

Validation Report 18

Highly Sensitive and Rugged GC/MS/MS Tool

Aminoglycosides in Milk Using Agilent Bond Elut Plexa SPE, Agilent Poroshell 120, and LC/Tandem MS

Comprehensive Pesticide Analysis in Juice Using a Combination of GC/MS and LC/MS Methods

Application Note. Gas Chromatography/Mass Spectrometry/Food Safety. Abstract. Authors

Screening of Pesticide Residues in Water by Sequential Stir Bar Sorptive Extraction-Thermal Desorption with GC/MS

Determination of Chlorinated Acid Herbicides in Soil by LC/MS/MS Application Note

Quantification of Pesticides in Food without Calibration using GC/FID with the Polyarc Reactor

Determination of Volatile Aromatic Compounds in Soil by Manual SPME and Agilent 5975T LTM GC/MSD

Analysis of Trace (mg/kg) Thiophene in Benzene Using Two-Dimensional Gas Chromatography and Flame Ionization Detection Application

Thermo Scientific HyperSep Dispersive SPE Products. Efficient sample preparation and clean-up using the QuEChERS Method

Application Note. Abstract. Introduction. Determination of Polycyclic Aromatic Hydrocarbons in Seafood by an Automated QuEChERS Solution

The Quantitation and Identification of Coccidiostats in Food by LC-MS/MS using the AB SCIEX 4000 Q TRAP System

Improved GC/MS Analysis of Tomato Pesticides with Agilent Deactivated Silica Tubing

Organophosphorus Pesticides in Apple Matrix by GC/MS/FPD using an Agilent J&W DB-35ms Ultra Inert GC Column

Author. Abstract. Introduction

High Sensitivity HPLC Analysis of Perchlorate in Tap Water Using an Agilent 6460 Triple Quadrupole LC/MS System

Determination of Hormones in Drinking Water by LC/MS/MS Using an Agilent InfinityLab Poroshell HPH Column (EPA 539)

Analysis of Ethanol and Isotopomers by 240 Quadrupole Ion Trap GC/MS

The Importance of Area and Retention Time Precision in Gas Chromatography Technical Note

Validation Report 8 Determination of pesticide residues in rice and wheat by GC-MS/MS and LC-MS/MS (QuEChERS method) Appendix 2

Agilent 6460 Triple Quadrupole LC/MS System with an Agilent 1290 Infinity LC For Multi-Plant Growth Regulator Analysis in Grapes

LC/MS/MS of Fungicides and Metabolites in Apple Juice with Agilent Bond Elut Plexa and Poroshell 120

Analysis of Phthalates in Body Wash using Solid-Supported Liquid-Liquid Extraction

Challenging Pesticide Analysis Using an Agilent J&W DB-35ms Ultra Inert GC Column

Analysis of Metals, Halides, and Inorganic Ions Using Hydrophilic Interaction Chromatography

Analysis of Illegal Dyes in Food Matrices using Automated Online Sample Preparation with LC/MS

Sensitive and Repeatable Analysis of Pesticides in QuEChERS Extracts with APGC-MS/MS

Tyler Trent, Applications Sales Specialist, Teledyne Tekmar P a g e 1

LC/MS/MS of Fungicides and Metabolites in Orange Juice with Agilent Bond Elut Plexa and Poroshell 120

Determination of Total Volatile Organic Compounds in Indoor Air Using Agilent 7667A mini TD and 7820A GC

Separation of Explosives in EPA 8330: Column Choices Optimize Speed, Resolution, and Solvent Use. Application. Authors. Abstract.

Reduce Cost of Pesticide Residue Analysis

The Analysis of Organophosphate Pesticides by LC/MS Application

Amphetamines, Phentermine, and Designer Stimulant Quantitation Using an Agilent 6430 LC/MS/MS

Identification and Quantitation of PCB Aroclor Mixtures in a Single Run Using the Agilent 7000B Triple Quadrupole GC/MS

STANDARD OPERATING PROCEDURES SOP: 1828 PAGE: 1 of 14 REV: 0.0 DATE: 05/12/95 ANALYSIS OF METHYL PARATHION IN CARPET SAMPLES BY GC/MS

Appendix II- Bioanalytical Method Development and Validation

Analysis of Biomarkers in Crude Oil Using the Agilent 7200 GC/Q-TOF

Screening for Water Pollutants With the Agilent SureTarget GC/MSD Water Pollutants Screener, SureTarget Workflow, and Customized Reporting

DIQUAT DIBROMIDE. The Determination of Ethylene Dibromide in Diquat Dibromide and Diquat Dibromide / Paraquat Dichloride SL Formulations

Determination of 17 Organotin Compounds in Beverages Using Triple Quadrupole GC-MS/MS System

Identification and Quantitation of Pesticides in Chamomile and Ginger Extracts Using an Agilent 6460 Triple Quadrupole LC/MS system with Triggered MRM

Agilent s New Weak Anion Exchange (WAX) Solid Phase Extraction Cartridges: SampliQ WAX

Improved Volatiles Analysis Using Static Headspace, the Agilent 5977B GC/MSD, and a High-efficiency Source

Sensitive Detection of 2-MIB and Geosmin in Drinking Water


Sulfotepp impurities in Chlorpyrifos EC formulations

Analysis of Pesticides in Baby Foods Using a Triple-Quadrupole GC/MS/MS

ENVIRONMENTAL analysis

Automated Sample Preparation of Headspace Standards Using the Agilent 7696 WorkBench

Determination of pesticides in baby food by UHPLC/MS/MS using the Agilent 1290 Infinity LC system and the Agilent 6460 triple quadrupole LC/MS

Analysis of USP Method <467> Residual Solvents on the Agilent 8890 GC System

A Direct 5 ms Column Performance Comparison for Active Semi-Volatile Analytes

LC/MS/MS qua ntitation of β-estradiol 17-acetate using an Agilent 6460 Triple Quadrupole LC/MS working in ESI negative ion mode

Application. Gas Chromatography March 1998

Analysis of Pesticides and Contaminants using HS-SPME-GC/MS and DRS Detection. Blair Surridge, Tyler Dyer, Rene Bilideau, Paloma Bertolin Presoti

Agilent 1200 Infinity Series HDR DAD Impurity Analyzer System for the Quantification of Trace Level of Genotoxic Impurity

Analytical determination of testosterone in human serum using an Agilent Ultivo Triple Quadrupole LC/MS

Technical Procedure for Solid Phase Extraction of THC and THC-COOH for GC-MS Analysis

DETERMINATION OF OCs, PCBs AND SYNTHETIC PYRETHROIDS IN ANIMAL FAT

Analysis of Stachydrine in Leonurus japonicus Using an Agilent ZORBAX RRHD HILIC Plus Column with LC/ELSD and LC/MS/MS

UCT. Selectively removing Chlorophyll without Compromising your Planar Pesticides INNOVATION THROUGH CHEMISTRY E N V I R O

Identifying Pesticides with Full Scan, SIM, µecd, and FPD from a Single Injection Application

Analysis of Low-Calorie Sweeteners by Liquid Chromatography-Tandem Mass Spectrometry

Application Note. Authors. Abstract. Xin Sun Shandong Agriculture University, Taian, , China

Advantages of Reversed Sandwich Injection for Pesticide Residue Analysis

Sensitive Femtogram Determination of Aflatoxins B 1 , B 2 , G 1. and G 2. in Food Matrices using Triple Quadrupole LC/MS. Authors.

Analyze Hydrocarbon Impurities in 1,3-Butadiene with an Agilent J&W GS-Alumina PT Column

Quantification of growth promoters olaquindox and carbadox in animal feedstuff with the Agilent 1260 Infinity Binary LC system with UV detection

Using UHPLC-Triple Quadrupole MS/MS to Detect the Presence of Bark Extract and Yohimbine Adulteration in Dietary Supplements and Botanicals

VALIDATION OF A UPLC METHOD FOR A BENZOCAINE, BUTAMBEN, AND TETRACAINE HYDROCHLORIDE TOPICAL SOLUTION

Application. Gas Chromatography February Introduction

Rapid Screening and Analysis of Components in Nonalcoholic Drinks Application

Fast USEPA 8270 Semivolatiles Analysis Using the 6890/5973 inert GC/MSD with Performance Electronics Application

Static Headspace Blood Alcohol Analysis with the G1888 Network Headspace Sampler Application

Validation Report Determination of isoprothiolane residues in rice by GC-MS/MS (QuEChERS method)

METHOD 3600C CLEANUP

1,2-Dibromoethane (EDB) and 1,2-dibromo-3-chloropropane (DBCP), gas chromatography, microextraction

Application Note No Introduction. DMI Instrumentation. DMI Method. DMI for Lettuce and Pea Extracts. Lettuce results

The Analysis of Fluoroquinolones in Beef Kidney Using HPLC Electrospray Mass Spectrometry Application

IMPROVING PESTICIDE RESIDUE DETECTION PROTOCOL FOR FRUIT

Value of Eclipse Plus C18 and ph Selectivity to Analyze Amine-Containing Drugs

Uncontrolled Copy. SOP 109 Ethylene Glycol Screen by Gas Chromatography/Mass Spectrometry. Table of Contents. 1. Principle of Assay...

An Optimal Method for the Analysis of Pesticides in a Variety of Matrices

METHOD 8033 ACETONITRILE BY GAS CHROMATOGRAPHY WITH NITROGEN-PHOSPHORUS DETECTION

Analysis of Polycyclic Aromatic Hydrocarbons in Soil with Agilent Bond Elut HPLC-FLD

Automated Sample Preparation in Quality Control of Eye-Drop Formulation

Determination of Pb, Cd, Cr and Ni in Grains Based on Four Chinese National Methods via Zeeman GFAAS

Routine Multiresidue Pesticide Analysis using the Agilent 6470 Triple Quadrupole Mass Spectrometer

Transcription:

Analysis of Pesticide Residues in Apple Using Agilent Bond Elut QuEChER AAC Kits by GC/M Application Note Food afety Authors Limian Zhao, David chultz, and Joan tevens Agilent Technologies, Inc. 2850 Centerville Road Wilmington, DE 19809-1610 UA Abstract This application note describes the use of a quick, easy, cheap, effective, rugged, and safe (QuEChER) AAC sample preparation approach for extraction and cleanup of 17 GC-amenable pesticide residues from multiple classes, in apple. The method employed involves initial extraction in a buffered aqueous/acetonitrile system, an extraction/ partitioning step after the addition of salt, and a cleanup step utilizing dispersive solid phase extraction (dispersive PE). The two different dispersive PE clean-up approaches used either a 1 ml or 8 ml sample volume and were evaluated in parallel after sample extraction. The target pesticides in the apple extracts were then analyzed by gas chromatography/mass spectrometry (GC/M) operating in selective ion monitoring (IM) mode. The method was validated in terms of recovery and reproducibility. The limit of quantitation (LQ) for most pesticides is 10 ng/g; however, the pesticide Folpet has an LQ of 50 ng/g in apple. This application employing Bond Elut QuEChER kits produced results well below the maximum residue limits (MRLs) for all the pesticides screened. The spiked levels for the recovery experiments were 10, 50, and 200 ng/g. Recoveries ranged between 70 and 136% (92.5% on average), with RD below 15% (5.0% on average).

Introduction The QuEChER method for pesticide analysis was first introduced by UDA scientists in 2003. [1] The method was modified to address problematic pesticides by including a buffered extraction system [2]. After a full validation for more than 200 pesticides, this improved method was formalized and adopted as AAC fficial Method 2007.01. [3] In summary, the method uses a single-step buffered acetonitrile (1% HAc) extraction while simultaneously salting out water from the sample using anhydrous magnesium sulfate (Mg 4 ) to induce liquid-liquid partitioning. For cleanup, a dispersive solid phase extraction (dispersive PE) step is employed using a combination of primary secondary amine (PA) to remove fatty acids as well as other components, and anhydrous Mg 4 to reduce the remaining water in the extract. After mixing and centrifugation, the upper layer is ready for analysis. Gas Chromatography/Mass pectrometry (GC/M) has been widely used in pesticide analysis for many years, because many pesticides are volatile or semi-volatile they are GCamenable. Previously, we evaluated the performance of a Bond Elut AAC buffered extraction kit and Bond Elut AAC dispersive PE kits for the analysis of polar pesticides in apple using LC/M/M for detection and quantification. [4] In this study, the performance of the Bond Elut AAC Buffered Extraction kit (PN 5982-5755) and Bond Elut AAC dispersive-pe kits for General Fruits and Vegetables (p/n 5982-5022 and 5982-5058) were evaluated for the extraction of volatile and semi-volatile pesticides. Analysis was performed by GC/M. eventeen GC-amenable pesticides were selected which represent multiple classes, including nonpolar organochlorine pesticides (Cs), certain organophosphorus pesticides (Ps) and organonitrogen pesticides (Ns). The MRLs of these pesticides are a function of both the pesticide class and food matrix and have been set at 10 ng/g or higher. Table 1 shows the chemical and regulatory information for these pesticides in apple. Experimental Reagents and Chemicals All reagents and solvents were HPLC or analytical grade. Acetonitrile (ACN), and methanol (MeH) were from Honeywell (Muskegon, MI, UA), and acetic acid (HAc) was from igma-aldrich (t Louis, M, UA). Formic acid (FA) was from Fluka (leinheim, Germany). The pesticide standards and internal standard (triphenyl phosphate, TPP) were purchased from igma-aldrich (t Louis, M, UA), Chem ervice (West Chester, PA, UA), or Ultra cientific (North Kingstown, RI, UA). olutions and tandards A 1% acetic acid solution in ACN was prepared by adding 10 ml of acetic acid to 1 L of ACN. tandard and internal standard (I) stock solutions (2 mg/ml of 11 pesticides) were made in MeH, respectively, and stored at 20 ºC. A commercially available mix of 6 pesticides, at 20 µg/ml in hexane was used directly. Three QC spiking solutions of 11 pesticides at 1.5, 7.5 and 30 µg/ml were made fresh daily in 1:1 ACN/H 2 containing 0.1% FA, while the 20 µg/ml of 6 pesticides mix was directly used for QC spike. A 2.5 µg/ml standard solution of 17 pesticides in ACN containing 0.1% FA was used to prepare the calibration curves in the matrix blank extract by appropriate dilution. A 15 µg/ml of TPP spiking solution in 1:1 ACN/H 2 containing 0.1% FA was used as the internal spiking standard (I). Equipment and Material Agilent Gas Chromatograph (Agilent Technologies Inc., anta ara, CA, UA). Agilent 5975C eries GC/MD (Agilent Technologies Inc., anta ara, CA, UA). Agilent Bond Elut QuEChER AAC Extraction kits, p/n 5982-5755 (Agilent Technologies Inc., Wilmington, DE, UA). Agilent Bond Elut QuEChER AAC dispersive PE kits for General Fruits and Vegetables, p/n 5982-5022 and 5982-5058 (Agilent Technologies Inc., Wilmington, DE, UA). CentraCL3R Centrifuge (Thermo IEC, MA, UA) Bottle top dispenser (VWR, o Painfield, NJ, UA) Eppendorf microcentrifuge (Brinkmann Instruments, Westbury, NY, UA) 2

Table 1. Pesticides Chemical and Regulatory Information [5 8] MRLs in apple Name Category Log P pka tructure (ng/g)* s-phenylphenol Phenol 3.18 9.4 20 H Carbaryl Carbamate 2.36 10.4 50 NH Dichlofluanid ulphamide 3.7 NA 5000 Dichlorvos rganophosphate 1.9 NA 10 Diazinon rganophosphate 3.69 2.6 100 N N F P N N P Chlorothalonil Chloronitrile 2.94 NA 10 N N Dichlorobenzo phenone rganochlorine 4.44 NA 500 C (Continued) 3

Table 1. Pesticides Chemical and Regulatory Information [5 8] MRLs in apple Name ass Log P pka tructure (ng/g)* Folpet Phthalimide 3.02 NA 3000 N Chlordane Cyclodiene organochlorine 2.78 NA 20 Endosulfan rganochlorine 3.13 NA 50 Dieldrin Chlorinated hydrocarbon 3.7 NA 10 DDE rganochlorine 6.55 NA 50 Ethion rganophosphate 5.07 NA 300 P P (Continued) 4

Table 1. Pesticides Chemical and Regulatory Information [5 8] MRLs in apple Name ass Log P pka tructure (ng/g)* Endosulfan sulfate rganochlorine 3.13 NA 50 Endrin ketone rganchlorine 4.99 NA 10 Permethrins Pyrethroid 6.1 NA 50 Coumaphos rganothio phosphate 3.86 NA 100 P *The MRLs numbers list in the table are for apple or lowest level in other fruit and vegetables. They could be higher in different commodities. 5

Instrument Condition An Agilent GC/M method for pesticides analysis was used for this study. [9] GC conditions Auto-sampler: Agilent 7683 automatic liquid sampler Inlet: plitless Column: Agilent 30 m 0.25 mm 0.25 µm HP-5M Ultra Inert (p/n 19091-433UI) Carrier gas: Helium in the constant pressure mode Retention time locking: Chlorpyrifos-methyl locked to 16.596 min (nominal column head pressure = 22.0 psi) ven temperature 70 ºC (2 min), 25 ºC/min to 150 ºC (0 min), program: 3 ºC /min to 200 ºC (0 min), 8 ºC/min to 280 ºC (11.5 min) Injection volume: 1.0 µl M conditions Tune file: Atune.u Mode: IM (refer to Table 2 for settings in detail) ource, quad, transfer 230 ºC, 150 ºC and 280 ºC respectively, line temperature: olvent delay: 3.00 min Multiplier voltage: Autotune voltage ample preparation ample comminution rganically grown, pesticide-free apples were purchased from a local grocery store. Approximately three pounds of apples were chopped into small, bean sized cubes. kin was included, but the seeds were discarded. The chopped apple cubes were then placed into a clean plastic bag and frozen at 20 ºC overnight. The bag was massaged occasionally to make sure the cubes remained separate. The following day, only the required amount of frozen apple cubes was removed and thoroughly blended. Dry ice was added while comminuting, when possible. amples were comminuted thoroughly to get the best sample homogeneity, ensuring there were no pieces of apple visible in the final sample. Extraction/Partitioning A 15 g (± 0.1g) amount of previously homogenized sample was placed into a 50 ml centrifuge tube (from the Bond Elut QuEChER extraction kit). QC samples were fortified with 100 µl of appropriate QC spiking solution (11 pesticides) and 7.5, 37.5, and 150 µl of 20 µg/ml stock solution (6 pesticides mixture), respectively, yielding QC samples with concentra- Table 2. Instrument Acquisition Data Used for the Analysis of 17 Pesticides by GC/M. Analyte IM Collection window (min) RT (min) (1) Dichlorvos 184.9 3.0 6.5 5.8 (2) σ-phenylphenol 170.1, 169.1 6.5 9.5 8.8 (3) Diazinon 137.1, 179.1 13.5 14.65 14.5 (4) Chlorothalonil 265.9, 263.9 14.65 16.0 14.8 (5) Carbaryl 144 16.0 17.5 16.8 (6) Dichlofluanid 123, 167.1 17.5 18.8 18.4 (7) Dichlorobenzophenone 139, 249.9 18.8 20.5 19.2 (8) Folpet 259.9, 261.9 21.35 21.8 21.6 (9) γ-chlordane 372.9, 374.9 21.8 22.3 22.0 (10) Endosulfan 240.8, 238.8 22.3 23.2 22.6 (11) Dieldrin 262.8 23.2 25.0 23.9 (12) DDE 245.9, 317.9 23.2 25.0 24.0 (13) Ethion 230.9 25.0 26.4 26.0 (14) Endosulfan sulfate 273.8 26.4 27.2 26.8 TPP (I) 325.1, 326.1 27.2 28.0 27.7 (15) Endrin ketone 316.9 28.0 28.5 28.2 (16) Permethrin 183.1 30.0 32.5 31.4, 31.6 (17) Coumaphos 362.0 30.0 32.5 31.7 6

tions of 10, 50 and 200 ng/g. A 100 µl amount of internal standard spiking solution (15 µg/ml of TPP) was added to all samples except the control blank to yield a 100 ng/g concentration in each sample. Tubes were capped and vortexed for 1 min. A 15 ml amount of 1% HAc in ACN was added to each tube using the dispenser. An Agilent Bond Elut QuEChER extraction salt packet from the kit (PN 5982-5755) containing 6 g of anhydrous Mg 4, and 1.5 g of anhydrous NaAc was added directly to the tubes. The salt bag was massaged carefully to break up any salt clumps before pouring. The tubes were examined to ensure that no powder was left in the threads or rims of the tubes. ample tubes were sealed tightly and shaken vigorously for 1 min by hand to ensure that the solvent interacted with the entire sample and crystalline agglomerates were dispersed. ample tubes were centrifuged at 4000 rpm for 5 min. Dispersive PE eanup A 1 ml aliquot of the upper ACN layer was transferred to an Agilent Bond Elut QuEChER dispersive PE 2 ml tube (p/n 5982-5022). An 8 ml aliquot was transferred to an Agilent Bond Elut QuEChER dispersive PE 15 ml tube (p/n 5982-5058). The 2 ml tube contained 50 mg of PA and 150 mg of anhydrous Mg 4 ; while the 15 ml tube contained 400 mg of PA and 1200 mg of anhydrous Mg 4. The tubes were tightly capped and vortexed for 1 min. The 2 ml tubes were centrifuged with a micro-centrifuge at 13,000 rpm for 2 min, and 15 ml tubes in a standard centrifuge at 4000 rpm for 5 min. An aliquot from the extract, 500 µl was transferred into an autosampler vial, and analyzed by GC/M. Weigh 15 g comminuted sample (±0.01 g) in 50 ml centrifuge tube Add 100 µl of I (TPP) solution, and QC spike solution if necessary, vortex 1 min Add 15 ml of ACN containing 1% HAc Add Bond Elut AAC QuEChER Extraction salt packet Cap and shake vigorously for 1 min Centrifuge at 4000 rpm 5 min Transfer 1 ml of upper ACN layer to Bond Elut AAC dispersive PE 2 ml tube, or 8 ml to Bond Elut AAC dispersive PE 15 ml tube Vortex 1 min, centrifuge at 13000 rpm for 2 min for 2 ml tubes or at 4000 rpm for 5 min for 15 ml tubes Transfer 500 µl extract to autosampler vial Analyze by GC/M Figure 1 shows the flow chart for the QuEChER AAC sample extraction procedure. Figure 1. Flow chart of the Agilent Bond Elut QuEChER AAC extraction procedure. Results and Discussion Using the Bond Elut QuEChER kits, the entire procedure is faster, easier, offers time and labor savings, while ensuring consistency. An analyst can process 40 50 samples in just a few hours. The addition of a food sample with a high content of water directly to the salts creates an exothermic reaction, which can affect analyte recoveries, especially for volatile pesticides. Agilent's Bond Elut extraction salts are uniquely prepared in an anhydrous package. The unique Bond Elut anhydrous salts packet allows addition after adding organic solvent to the sample, as specified in the original QuEChERs method. In our previous study, the new design of Bond Elut QuEChER AAC kits demonstrated excellent recovery and precision for a broad variety of semi-polar to polar pesticides using LC/M/M. [4] There are many semi-volatile and volatile pesticides, so the use of GC/M is applicable for the performance evaluation of the AAC kits for the analysis of these groups of pesticides. The selectivity of GC/M (IM mode) is not as effective as that of LC/M/M (MRM mode). Furthermore, the final QuEChER samples still contained food matrix impurities, which can be observed in the GC/M chromatogram of blank apple extract. Therefore, it is important to carefully choose the monitored ions of each compound when setting up the IM method. In general, the most abundant ions were selected in order to achieve the best sensitivity; however in a few instances the sensitivity was compromised to obtain better selectivity by using more unique but less abundant ions. As shown in Figure 2a, there are interference peaks apparent in the blank chromatogram; fortunately most pesticides are free of co-eluting interferences. There was an interference eluting at a retention time very close to that of s-phenylphenol, and can not be differentiated for quantitation. 7

The response of this interferent within the blank was integrated to be less than 20% response of s-phenylphenol peak at the LQ (10 ng/g) sample. Therefore, the selectivity was considered acceptable for this compound. Figure 2 (a, b) shows the chromatograms of a blank apple extract and 50 ng/g fortified apple extract. Linearity and Limit of Quantification (LQ) The linear calibration range for all of the pesticides was 0 400 ng/g; excluding Folpet at 50 400 ng/g due to poor sensitivity. Two different dispersive PE volumes (1 ml and 8 ml) were used for evaluation and comparison; therefore, two calibration curves were generated from matrix blanks prepared from each size. Each calibration curve was made at levels of 10, 20, 50, 100, 250, and 400 ng/g. The TPP was the internal standard (I) at 100 ng/g in all cases. The calibration curves were generated by plotting the relative responses of analytes (peak area of analyte/peak area of I) to the relative concentration of analytes (concentration of analyte/concentration of I). Table 1 shows that the 10 ng/g quantification limits LQ (10 ppb) and 50 ng/g LQ for Folpet (50 ppb) established for pesticides are substantially lower than many MRLs for the pesticides in fruit and vegetables. The regression fit used for the calibration curves was the average response factor. Table 3 shows the linear term and RF relative standard deviation (%) for both 1 ml and 8 ml dispersive PE. 2000 1800 1600 1400 GC/M Chromatogram of Apple Extracts, Blank Relative to Fortified ample, 50 ng/g after Agilent s Bond Elut QuEChER extraction and dispersive PE, for General Fruits and Vegetables A Abundance 1200 1000 800 600 400 200 I 0 2000 1800 1600 1400 5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 Time (min) B Abundance 1200 1000 800 2 I 600 400 200 0 12 1 3 4 5 6 7 16 8 9 10 11 13 14 15 17 5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 Time (min) Figure 2. GC/M chromatogram of apple extract. (A) apple extract blank; (B) 50-ng/g fortified apple extract. Peak Identification: 1. Dichlorvos, 2. s-phenylphenol, 3. Diazinon, 4. Chlorothalonil, 5. Carbaryl, 6. Dichlofluanid, 7. Dichlorobenzophenone, 8. Folpet, 9. γ-chlordane, 10. Endosulfan, 11. Dieldrin, 12. DDE, 13. Ethion, 14. Endosulfan sulfate, 15. Endrin ketone, 16, Permethrin, 17. Coumaphos. I. Triphenyl phosphate (TPP). 8

Table 3. Linearity of 17 Pesticides in Apple Extract 1 ml dispersive PE 8 ml dispersive PE Analytes Linear Term RF Rel td Dev (%) Linear Term RF Rel td Dev (%) Dichlorvos 3.47e-001 11.4 3.87e-001 4.6 σ-phenylphenol 1.37e-000 10.7 1.50e-000 11.4 Diazinon 7.04e-001 10.9 7.39e-001 6.5 Chlorothalonil 6.84e-001 13.7 8.02e-001 8.9 Carbaryl 8.07e-001 14.1 1.01e-000 10.8 Dichlofluanid 1.04e-000 12.8 1.08e-000 8.6 Dichlorobenzophenone 4.55e-001 11.4 4.60e-001 8.2 Folpet 3.88e-002 19.5 4.52e-002 20.1 γ-chlordane 3.23e-001 10.4 3.31e-001 9.2 Endosulfan 8.56e-002 15.2 8.26e-002 8.8 Dieldrin 2.71e-001 6.2 2.59e-001 5.9 DDE 1.43e-000 8.4 1.39e-000 7.5 Ethion 5.87e-001 19.7 5.63e-001 17.0 Endosulfan sulfate 2.72e-001 9.6 2.74e-001 9.5 Endrin ketone 2.75e-001 10.1 2.75e-001 7.8 Permethrin 9.71e-001 9.4 9.29e-001 8.0 Coumaphos 2.70e-001 15.6 2.72e-001 15.7 Recovery and Reproducibility The recovery and reproducibility were evaluated by spiking pesticides standards in comminuted apple sample at levels of 10, 50 and 200 ng/g. These QC samples were quantitated against the matrix-spiked calibration curve. The analysis was performed in replicates of six (n=6) at each level. The recovery and reproducibility (shown as % RD) data for 1 ml and 8 ml volume dispersive PE are shown in Table 4 and Table 5, respectively. It can be seen from the results that all of the pesticides give excellent recoveries (average of 90.8% for 1 ml and 94.2% for 8 ml) and precision (average of 5.7% RD for 1 ml and 4.3% RD for 8 ml). As mentioned above, an interferent was eluted very closely with s-phenylphenol. The selectivity was still acceptable because the interferent contributed less than 20% of LQ; however, the contribution of the interference peak resulted in the higher recovery of this compound at low levels. Folpet is a notoriously unstable pesticide, and the main problems dealing with degradation and instability come from the N-trihalomethylthio functional group. [3, 10] Folpet was quantified, but the LQ was found to be 50 ng/g due to poor sensitivity, however, recovery and reproducibility at 50 ng/g and above were acceptable (average recovery 85.5%, average reproducibility 10%). 9

Table 4. Recovery and Repeatability of Pesticides in Fortified Apple With Agilent Bond Elut 2 ml Dispersive PE Tube (p/n 5982-5022); Recovery 90.8%, RD 5.7% (avg) 10 ng/g fortified QC 50 ng/g fortified QC 200 ng/g fortified QC Analytes Recovery RD (n=6) Recovery RD (n=6) Recovery RD (n=6) Dichlorvos 86.8 7.0 83.9 11.6 81.5 5.5 s-phenylphenol 113.4 6.3 96.3 6.5 100.5 3.6 Diazinon 98.6 2.3 87.3 2.8 90.4 4.9 Chlorothalonil 86.1 10.0 84.4 5.3 93.2 7.6 Carbaryl 96.1 9.0 93.8 8.3 99.1 8.2 Dichlofluanid 90.0 7.0 84.6 2.9 94.6 5.0 Dichlorobenzo phenone 97.8 7.6 95.0 6.2 102.2 4.3 Folpet 74.4 9.1 95.7 11.0 g-chlordane 79.6 4.4 88.9 4.3 95.3 4.4 Endosulfan 69.8 9.2 91.2 5.3 96.2 5.2 Dieldrin 90.6 10.9 86.6 3.2 92.8 4.8 DDE 84.0 4.8 89.4 3.8 95.4 4.5 Ethion 90.9 1.8 103.5 1.4 116.5 5.0 Endosulfan sulfate 79.8 1.9 80.4 4.6 86.8 5.6 Endrin ketone 85.2 12.0 80.7 3.6 91.8 4.5 Permethrin 87.9 2.8 93.8 2.0 94.0 4.4 Coumaphos 87.8 5.1 89.7 3.0 90.0 6.4 10

Table 5. Recovery and Repeatability of Pesticides in Fortified Apple With Agilent Bond Elut 15 ml Dispersive PE Tube (p/n 5982-5058); Recovery 94.2%, RD 4.3% (avg) 10 ng/g fortified QC 50 ng/g fortified QC 200 ng/g fortified QC Analytes Recovery RD (n=6) Recovery RD (n=6) Recovery RD (n=6) Dichlorvos 103.4 4.2 85.6 8.1 97.2 7.2 σ-phenylphenol 125.8 8.7 99.2 4.4 105.4 5.0 Diazinon 96.0 4.5 82.3 2.1 88.4 6.3 Chlorothalonil 96.5 3.0 82.8 5.2 97.7 4.5 Carbaryl 97.7 3.9 91.4 4.4 101.9 5.0 Dichlofluanid 91.7 5.1 83.7 1.0 93.7 5.1 Dichlorobenzo phenone 98.8 9.3 96.2 4.7 105.3 4.3 Folpet 88.4 4.0 72.5 6.0 γ-chlordane 80.9 3.5 87.5 3.3 94.8 5.0 Endosulfan 80.3 7.3 84.1 3.6 98.6 3.0 Dieldrin 81.2 3.4 93.1 2.0 98.7 3.9 DDE 86.1 1.8 92.4 3.4 98.9 3.9 Ethion 106.5 3.6 122.2 2.0 136.3 4.2 Endosulfan sulfate 91.6 4.6 87.7 4.0 93.0 4.1 Endrin ketone 76.2 3.3 82.4 3.9 91.8 4.1 Permethrin 97.9 1.6 104.7 1.1 106.6 4.2 Coumaphos 82.3 6.7 86.5 2.5 89.3 5.1 Figure 3 shows the recovery and precision results for 1 ml dispersive PE and 8 ml dispersive PE. The two different dispersive PE clean-ups were performed by transferring 1 ml or 8 ml of ACN extract from the same sample following the extraction step. In order to simplify the comparison, the average recovery and precision of three fortification concentrations were used for all pesticides. The results of each dispersive PE clean-up appeared to be independent of volume used. Both approaches provided efficient and similar sample clean-up, and thus generated relatively equivalent results. 11

Exceptional Recoveries and Precision for 1 and 8 ml Volumes for Agilent Bond Elut Dispersive PE, 2 and 15 ml kits 120 1 ml 8 ml 100 80 60 40 20 0 Dichlorvos σ-phenylphenol Diazinon Chlorothalonil Recovery (%) Carbaryl Dichlorfluanid Dichlorobenzophenone Forpet γ-chlordane Endosulfan Dieldrin DDE Ethion Endosulfan sulfate Endrin ketone Permetrin Coumaphos Figure 3. Recoveries and precision for 1 and 8 ml sample volumes employing Agilent Bond Elut Dispersive PE, 2 and 15 ml kits, respectively. Conclusions Agilent Bond Elut QuEChER AAC method for General Fruits and Vegetables: Extraction and Dispersive PE kits provided a simple, fast and effective method for the purification and enrichment of representative volatile to semi-volatile pesticides in apple. The recovery and reproducibility, based on matrix spiked standards, were acceptable for multiclass, multi-residue pesticide determination in apple. The impurities and matrix effects from apple did not interfere with the quantitation of target compounds. The LQs of the pesticides were lower than regulated MRLs in apple. As the selected pesticides represented a broad variety of different classes and properties, the Agilent Bond Elut QuEChER AAC Extraction and Dispersive PE kits for General Fruits and Vegetables is an excellent choice for other pesticides in similar food matricies. Acknowledgements The authors wish to thank Dr. Meng, Chin K. for his help with the GC/M instrument, and Dr. Ball, Carol H. for many valuable suggestions. 12

References 1. M. Anastassiades,. J. Lehotay, Fast and Easy Multiresidue Method Employment Acetonitrile Extraction/Partitioning and "Dispersive olid-phase Extraction" for the Determination of Pesticide Residues in Produce, J. AAC Int., 2003, 86, 412-431. 2.. J. Lehotay, et al, Use of Buffering and ther Means to Improve Results of Problematic Pesticides in a Fast and Easy Method for Residue Analysis of Fruits and Vegetables, J. AAC Int., 2005, 88, 615-629. 3.. J. Lehotay, et. al, Determination of Pesticide Residues in Foods by Acetonitrile Extraction and Partitioning with Magnesium ulfate, Collaborative tudy, J. AAC Int., 2007, 90, 485-520. 4. L. Zhao, D. chultz, Evaluation of the QuEChER AAC ample Preparation Kit for the Analysis of Pesticide Residues in Apples with LC/M/M Detection, Agilent Technologies publication 5990-3937EN. 5. http://sitem.herts.ac.uk/aeru/footprint/en/index.htm 6. http://www.m5.ws001.squarestart.ne.jp/foundation/ search.html 7. http://www.mrldatabase.com/?selectvetdrug=0 8. http://www.ams.usda.gov/amv1.0/getfile?ddoc Name=PDP1995ummary 9. P. L. Wylie, creening for 926 Pesticides and Endocrine Disruptors by GC/M with Deconvolution Reporting oftware and a New Pesticide Library, Agilent Technologies publication 5989-5076EN. 10. B. Mercedes et.al. Residue determination of captan and folpet in vegetable samples by gas chromatography/ negative chemical ionization-mass spectrometry. J. AAC Int., 2006, 89, 1080-1087. For More Information For more information on our products and services, visit our Web site at www.agilent.com/chem. 13

www.agilent.com/chem Agilent shall not be liable for errors contained herein or for incidental or consequential damages in connection with the furnishing, performance, or use of this material. Information, descriptions, and specifications in this publication are subject to change without notice. Agilent Technologies, Inc., 2012 Printed in the UA January 6, 2012 5990-4068EN