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Analytical Toxicology

Analytical Toxicology

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Analytical Toxicology

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  1. Analytical Toxicology Instrumentation and Methodologies

  2. Sampling from liquid phases • Liquid extractions: • Batch liquid-liquid extraction • Solid phase extractions: • SPE – Solid Phase Extraction (transfer to another solvent) • SPME – Solid Phase Micro-Extraction (solventless) • Headspace sampling: • Static headspace (high concentrations) • Dynamic headspace (low-moderate concentrations) • Purge-and Trap (low concentrations)

  3. 2 Shake or mix thoroughly 3 Separate the 2 phases Combine the extracts and evaporate to reduce the volume Solvent Extraction 1 Add immisible extracting solvent Repeat these steps 1 to 3 times

  4. Solvent Extraction Distribution coefficient: Fraction remaining after n extractions:

  5. Miscibility of solvents Miscible if the two solvents can be mixed in all proportions without forming two phases

  6. Solid Phase Extraction - SPE Conditioning The sorbent is wetted and rinsed by the eluting solvent Acetonitrile Water Water samples Methanol/ water Acetonitrile The sorbent is conditioned by a pure solvent like the matrix Retention A liquid sample passed through a short column of solid sorbent, where the desired compounds are sorbed Rinse Unwanted compounds are rinsed by elution with a suitable solvent Elution The analytes are eluted by a suitable eluent

  7. Solid Phase Extraction - SPE

  8. Solid phases for SPE Most materials are bonded phases attached covalently to big porous silica particles (o.d.  50 m surface 50 m2g-1) • Non-polar sorbents (reversed phase): • C18 – C8 – C6 – C4 – C2 • Cyclohexyl • Phenyl • Cyanoprolyl • Polar sorbents (normal phase): • Cyanopropyl • Bare Silica • Diol • Aminoalkyl • Ion exchange sorbents: • Strong Anion eXchanger • Strong Cation eXchanger • Weak anion- and cation exchangers • Endcapped sorbents: • accessible –SiOH are reacted with trimethyl silane

  9. SPE – Modes and eluents • Change of sample polarity: • dilute with appropriate solvent • exchange solvent by SPE

  10. Solid Phase Micro-Extraction - SPME Metal rod • Sorption of analytes: • In situ extraction from headspace or liquid samples • 1-60 min • NO SOLVENTS USED ! Protecting metal tube • Thermal desorption: • Splitless injection in GC • (Interface for HPLC are known) Silica fiber Solid sorbent coating 1 cm

  11. Solid Phase Micro-Extraction - SPME

  12. P Po UV-Vis Absorption Spectroscopy • A = ebc

  13. A l Measure at lmax

  14. Tungsten Halogen Lamp D2 Lamp Grating Cell Detector Single beam spectrophotometer

  15. Reference Cell D2 Lamp Detector Chopper Beam Splitter Sample Cell Double beam Spectrophotometer

  16. Energy s p n p* s* s-s* s-s* p-p* p-p* n-s* n-s* n-p* n-p* Which drugs can be determined using UV-Vis? Energy s pnp*s*

  17. VR IC T1 S2 ISC S1 FL IC and EC Absorption Ph S0 Luminescence Spectroscopy

  18. Absorption Fluorescence I Phosphorescence Wavelength F = KP0ebc

  19. Sample cell Source Fex Fem Black Surface Detector Fluorometers

  20. Source Sample cell Black Surface lex lem Detector Spectrofluorometers

  21. 000000000 Luminometers

  22. 6s 5s 3d 4s 3p E 3s Atomic Absorption Spectrometers

  23. Solution of Analyte Nebulization Spray Desolvation Solid/Gas Aerosol Volatilization Excited Molecules Gaseous molecules hn molecular Atoms Excited Atoms hn Atomic Excited ions hn Ionic Atomic Ions

  24. High Pressure Gas Flow Solution sample Solution sample High Pressure Gas Flow Sample introduction

  25. Detector Pr P Half-Silvered Mirror Flame or Graphite Furnace Atomizer AAS Spectrometers

  26. Atomic Emission Spectroscopy

  27. DCP

  28. Detector ICP Spectrometer

  29. CCD or CID Detector Grating Mutichannel ICP Spectrometer

  30. ICP-MS

  31. Syringe Septum Carrier Gas Vaporization Chamber To Column Injector

  32. Thermal Conductivity detector

  33. Flame Ionization Detector

  34. Packed Columns • These columns are fabricated from glass, stainless steel, copper, or other suitable tubes. Stainless steel is the most common tubing used with internal diameters from 1-4 mm. The column is packed with finely divided particles (<100-300 mm diameter) which is coated with stationary phase. However, glass tubes are also used for large scale separations. Several types of tubing were used ranging from copper, stainless steel, aluminum and glass. Stainless steel is the most widely used because it is most inert and easy to work with. The column diameters currently in use are ordinarily 1/16" to 1/4" 0.D.

  35. Capillary/Open Tubular Open tubular or capillary columns are finding broad applications. These are used for fast and efficient separations but are good only for small samples. The most frequently used capillary column, nowadays, is the fused silica open tubular column (FSOT) which is a WCOT column. The external surface of the fused silica columns is coated with a polyimide film to increase their strength. The most frequently used internal diameters occur in the range from 260-320 micrometer.

  36. Liquid Stationary Phases • In general, the polarity of the stationary phase should match that of the sample constituents ("like" dissolves "like"). Most stationary phases are based on polydimethylsiloxane or polyethylene glycol (PEG) backbones:

  37. Stationary phases

  38. Typical Separation

  39. TPGC versus Isothermal

  40. Head Space GC