1. INTRODUCTION TO PESTICIDE RESIDUES ANALYSIS SUNDAY J KERKULA

2. PRESENTATION OUTLINE Introduction The analyst The laboratory EQUIPMENT AND SUPPLIES Avoidance of contamination Process steps Some guidance documents

3. INTRODUCTION Pesticide residues are one of the major chemical contaminants in food They are usually of very minute quantities – ppb, ppm Thus, the need for reliable analytical methods Food safety Facilitate trade

4. THE ANALYST Residue analysis consists of a chain of procedures mostly readily understood by a trained chemist. Due to the low levels of analyte concentrations in the range of μ g/kg to mg/kg and because the analyses can be challenging, attention to detail is essential. The analyst in charge should have an appropriate professional qualification and be experienced and competent in residue analysis. Staff must be fully trained and experienced in the correct use of apparatus and have appropriate laboratory skills

5. THE LABORATORY The laboratory and its facilities must be designed to allow tasks to be allocated to well-defined areas where maximum safety and minimum chance of contamination of samples. Laboratories should be constructed of, and utilise, materials resistant to chemicals likely to be used within them. Ideally, separate rooms should be designated for sample receipt and storage, sample preparation, standards preparation, extraction and clean-up, and instrumentation Sample receipt, storage, and preparation should be handled in areas devoted to working at residue levels Maintenance of sample integrity is very essential Extraction and clean-up area/room must be suitable for solvents to be used High quality fume extraction facilities are needed

6. THE LABORATORY 2 No smoking, eating, drinking, or application of cosmetics should be permitted in the working area. Only small volumes of solvents should be held in the working area. Bulk of the solvents stored separately, away from the main working area Avoid or minimize the use of highly toxic solvents Safety screens must be provided for use of glassware under pressure Provision of safety glasses, gloves, and other protective clothing, emergency washing facilities Fire fighting equipment must be available Staff must be aware that many pesticides have acutely or chronically toxic properties and therefore, great care is necessary when handling reference standard compounds

7. EQUIPMENT AND SUPPLIES Adequate and reliable supplies of electricity and water Adequate glassware, reagents, and solvents of suitable quality Equipment such as homogenizers, analytical balances, shakers, centrifuges, nitrogen or rotary evaporators, refrigerators and freezers Analytical equipment such as GC, GC-MS, HPLC, LC-MS Pesticide reference standards of known and acceptably high purity Parent compounds as well as those metabolites that are included in MRLs All analytical standards, stock solutions, working solutions and reagents should be properly labelled including preparation date, analyst’s identification, solvent used, expiry date and stored under appropriate conditions: low temperature, exclusion of moisture and light

8. AVOIDANCE OF CONTAMINATION Avoidance of contamination and interference is very critical. Trace amounts of contamination in the final samples used for the determination stage of the method can give rise to errors such as false positive or false negative results or to a loss of sensitivity that may prevent the residue from being detected. Contamination may arise from almost anything that is used for, or is associated with, sampling, sample transport and storage, and the analyses. All glassware, reagents, organic solvents and water should be checked for possible interfering contaminants before use, by analysis of a reagent blank. Polyvinylchloride (PVC) apparatus should not be used Ban the use of polishes, barrier creams, soaps containing germicides, insect sprays, perfumes, and cosmetics as they can give rise to interference problems and especially with the use of ECD. All glassware should be cleaned with detergent solution, rinsed thoroughly with distilled (or other clean) water and then rinsed with the solvent to be used Separate glassware used from reference standards from those used for samples Do not store standards solutions with extracts

9. PROCESS STEPSSampling Drawing of representative sample Sample preparation Removal of parts not to be analyzed Sample size Reduction Grinding, homogenization etc. Extraction and clean-up Maceration Shaking/vertexing Partitioning SPE dSPE Determination (instrumentati on ) Chromatography GC GC-MS HPLC LC-MS Data processing and reporting

10. PROCESS STEPS 2 Sampling – taking a representative is essential for reliable results. Use appropriate sampling protocol Subsampling should be appropriate by following relevant procedures Samples received must be accompanied by the relevant information All samples must be assigned a unique identification code immediately. This should accompany it through all stages of the analysis to the reporting of the results Samples to be analysed within a few days should be stored at (1 - 5 °C), away from direct sunlight Samples received deep-frozen must be kept at ≤ -16 o c until analysis. Temperature for longer storage period before analysis storage should be approximately - 20 °C, at which temperature enzymic degradation of pesticide residues is usually extremely slow. If prolonged storage is unavoidable, the effects of storage should be checked by analysing fortified samples stored under the same conditions for a similar period. Sample preparation – removal of portions not to be analysed, size reduction, grinding Extraction and clean-up – separation of the residue from matrix and removal of interfering components Instrumentation – determination of analytes (residues) and their quantitation

11. SOME GUIDANCE DOCUMENTS Guidelines on good laboratory practice in pesticide residue analysis, CXG 40-1993 Guidelines on performance criteria for methods of analysis for the determination of pesticide residues in food and feed, CXG 90-2017 Analytical quality control and method validation procedures for pesticide residues analysis in food and feed, SANTE 11312/2021 Recommended methods of sampling for the determination of pesticide residues for compliance with MRLs, CXG 33-1999 Portion of commodities to which maximum residues limits apply and which is analyzed, CXG 41-1993 Principles and guidance on the selection of representative commodities for the extrapolation of maximum residue limits for pesticides to commodity groups, CXG 84-2012 Guidelines on performance criteria for methods of analysis for the determination of pesticide residues in food and feed, CXG 90-2017

12. Thank you