Transfer of pesticides from commodities to natural flavors and aroma chemicals Ian Ronningen Chem 4101 December 9, 2011

Flavors and Fragrances Aroma chemicals are extracted from commodities through a number methods Mechanical pressing Solvent extraction Distillation Expression These extracts are commonly concentrated and then sold as fragrances or flavors AtrazineWarfarin Diazinon

Hypothesis Extraction processes commonly used to separate flavors and aroma chemicals from commodities also extract pesticides Extracted pesticides could make their way to consumers either through foods (flavors) or fragrances (aroma chemicals) Pesticides present have the potential to pose either a chronic or acute health risk 1.Muzaurieta, Annie Bell. "Vitamin C Foods." The Daily Green. Web. 4 Dec "OMG! Cookies." Poached Prose. Web. 04 Dec Orange, Source of Expressed orange oil(1) Potential Final Application (2)

Relevance To gain a competitive advantage flavor houses are increasingly sourcing rare commodities which come from countries that generally have less regulation surrounding pesticides Commodity testing is limited, and screens limited number of pesticides. Screening focus is towards pesticides used in the United States. If any pesticide ended up harming a consumer the producer of the food or fragrance would see massive economic backlash 1. Borneo Rainforest - Borneo Malaysia." Online Travel Guides of Travel Destinations -. Destination 360. Borneo Rainforest(1) Alachor

Analytical Techniques Separation TechniquePositiveNegative HPLCHigh sample throughput, numerous separation mechanisms, large variety of samples analyzed, AOAC sample preparation (QueChers) High solvent costs, co-elution an issue, GCModerate sample throughput, high resolution, numerous separation mechanisms Derivatization maybe required, sample preparation may be laborious, thermal degradation is possible CEGood separation, resolution can be made high, small sample size Poor selectivity for similar analytes, may require numerous separations of one sample Detection Methods MethodPositiveNegative Mass Spectrometry (triple-quad) Established libraries, highly selective, sensitivity adequate for expected concentration Expense, sensitivity may be low without concentration step FlourescenceHigh sensitivityLow selectivity, some compounds may need to be derivitized, no library available

HPLC-MS/MS Analysis Sample Preparation Quechers SPE Cartridge Separation Waters Acquity UPLC System Waters BEH C18 (2.1 x 100 mm) Analysis Waters XEVO TQD Mass Spectrometer High sample throughput, automated, rapid analysis, ability to deal with complex matrices Mass Spectrometer has high analytical selectivity Established Mass Spectra Libraries simplifies data analysis Many stationary phases available for wide application C18 is a good starting point, Phenyl Hexyl also an option Online SPE Also an option for higher sample throughput

Experimentation Incoming Commodities Washing Extraction Final Product Sample and analysis Addition of internal standards of phenolic butyl-parabens Control Experiments: Sample organic or hydroponic commodities (for incoming and extraction, and final product) Sample unused wash water "15 Eccentric Endangered Trees, Plants, and Flowers | WebEcoist." WebEcoist. Web. 07 Dec "Citrus Processing." Triowin. Web. 07 Dec. 2011

HPLC-MS/MS Analysis 1.Morphed, James, and Peter Hancock. “A RAPID METHOD FOR THE SCREENING AND CONFIRMATION OF OVER 400 PESTICIDE RESIDUES IN FOOD“ Waters Application Notes. Tech. Waters, Web. 1 Dec AOAC Official Method , Pesticide Residues in Foods by Acetonitrile Extraction and Partitioning with Magnesium Sulfate 3.QuEChERS-A Mini-Multiresidue Method for the Analysis of Pesticide Residues in Low-Fat Products. ResTek Application Notes. Sample Sample preparation and clean up Separation and characterization Reference to established libraries Unidentified peaks will be further investigated Restek QuEChERS sSPE tubes are an AOAC Method for Prep. Available (1,2). Include: C18, MGSO4, PSA, GCB

HPLC-MS/MS Concept HPLC Separation of compounds from matrix based off interaction of compounds with stationary and mobile phases Mass Spectrometer Ionization of analytes happens at the source, MS can either be used to scan or it can be operated in MS/MS utilizing collision cell and second quadrapole to analyze fragmentation patterns 1.“HPLC Separation Modes” Waters Application Notes, Waters Corporation “Mass Spectrometry Introduction." Department of Chemistry, University of Pittsburgh. Diagram of Separation (1) Diagram of Triple-quad (2)

Figures of Merit (1)Xu, Y. "Simultaneous Determination of Flonicamid and Its Metabolites in Vegetables Using QuEChERS and Reverse-Phase Liquid Chromatography Tandem Mass Spectrometry." Journal of chromatography. (2011). FigureLevel Expected analyte concentration after SPE ppm Desired Limit of detection*10 ppb Xevo TQD limit of detection (1)0.2 ppb Required Accuracy*1-2 ppb *Figure and requirements depends on the analyte, since instrument response and level in extract is dependent on pesticide

Summary Sampling along the line of extraction of aroma chemical from commodities will allow investigation of safety of these products HPLC-MS/MS Analysis allows for effective separation of pesticides from complex matrix and effective analysis of pesticides in natural flavors and fragrances extracted from commodities Although added cost of a MS/MS instrument is high added selectivity and sensitivity justify cost

Additional References AOAC Official Method , Pesticide Residues in Foods by Acetonitrile Extraction and Partitioning with Magnesium Sulfate. Culliney, T W. "Pesticides and natural toxicants in foods." Agriculture, ecosystems & environment (1992): Davies J, Barquet A, Vaclavek C, et al. Human Pesticide Poisonings by a Fat-Soluble Organophosphate Insecticide. Archives of Environmental Health. December 1975;30(12):608. Picó, Y. "Environmental and food applications of LC–tandem mass spectrometry in pesticide ‐ residue analysis: An overview." Mass spectrometry reviews 23.1 (2003):45. Reineccius, Gary. Flavor Chemistry and Technology. Boca Raton : Taylor & Francis, c2006. Schreiber, A. "Application of spectral libraries for high-performance liquid chromatography- atmospheric pressure ionization mass spectrometry to the analysis of pesticide and explosive residues in environmental samples." Journal of chromatography (2000):411 Seiber, J N. "Contributions of Pesticide Residue Chemistry to Improving Food and Environmental Safety: Past and Present Accomplishments and Future Challenges." Journal of agricultural and food chemistry (2011): Thurman, E M. "Choosing between atmospheric pressure chemical ionization and electrospray ionization interfaces for the HPLC/MS analysis of pesticides." Analytical chemistry (2001):5441.