2. ENVIRONMENTAL ANALYSIS I. Solid Phase Extraction fundamentals of SPE

3. SPE versus HPLC Elution Chromatography: V r = V m + KV s 0.2 << K << 20 Extraction Chromatography: K = > 1000 Retention K = < 0.001 Elution [stat] [matrix] [stat] [eluent]

4. SPE versus HPLC v HPLC ä continuous elution v SPE ä on-the-phase retention ä off-the-phase digital elution v Phase reproducibility ä top complaint in both techniques

5. Comparison of SPE and HPLC

6. Phase Usage in SPE and HPLC

7. Definition of SPE Separation or removal of an analyte or analytes from a mixture of compounds by selective partitioning of the compounds between a solid phase (sorbent) and a liquid phase (solvent).

8. SPE Analyte Partitioning Immiscible Phases Immiscible Phases

9. Why Silica? v Controlled particle/pore size v Controlled surface area v Surface can be modified v Mechanical strength/rigidity v Chemical stability v Wide solvent compatibility

10. Silica Surface Variations SiOH SiOHO H H Si OH OH Free silanolAdsorbed water Geminal silanol SiloxaneBound silanols

11. The Silica Surface

12. Monochlorosilane Chemistry R 3 SiCl + O Si O Si O OHOH O Si O Si O OHO Si RR R

13. Trichlorosilane Chemistry RSiCl 3 H 2 O RSi(OH) 3 OSiOSiO R OH R OH + OH Si O Si OO Si O OHOH OHSi R O Si OO Si O Si O O SiHO R OSiO HO OH R

14. Silica Particles (1)

15. Silica Particles (2)

16. ISOLUTE Bonded Silicas  50  m, 60 Å ä Narrow particle size distribution ä Easy, Rapid Flow v 550 m 2 /g Surface Area ä High Capacity v Rigid Silica Particles ä Dimensionally Stable ä Rapid Equilibration ä Small bed (void) volume (1.2mL/g) v Sorbent ä Silica pretreatment ä Proprietary Bonded Phase Washing

17. Particle Size Distribution 6.95 8.65 10.67 13.41 16.69 20.78 25.87 32.21 49.1 49.92 62.14 72.36 96.31 I S T ISOLUTE C18 (EC), BATCH NO. 221-3-04 Particle Size,  m Number of Particles Narrow range of particle sizes Negligible fines 1000 900 800 700 600 500 400 300 200 100

18. Particle Size Distribution of another Manufacturer 6.95 8.65 10.67 13.41 16.69 20.78 25.87 32.21 49.1 49.92 62.14 Particle Size,  m Number of Particles 1000 900 800 700 600 500 400 300 200 100 Broad range of particle sizes Significant fines

19. Impact of Fines When fines are present, smaller particles tend to agglomerate together To the right is a sorbent bed having a narrow particle size distribution

20. Impact of Fines The presence of fines results in poor flow characteristics through the column... versus even flow characteristics for a column without fines present

21. Poor Flow Characteristics Conditioning and equilibration solvents don’t saturate areas having a higher pressure drop... versus a well conditioned and equilibrated bed without fines present

22. Sample Loading Analytes are likely to channel through a bed with poor flow characteristics... versus being retained in a tight band

23. Drying Time The presence of fines results in longer drying times Versus a column where there is even gas flow

24. Drying Time, C18(UC)s 0 0.2 0.4 0.6 0.8 1 051015202530 ISTBrand ABrand B Drying Time, min Water, grams

25. Analyte Elution Greater solvent volumes are required to elute analytes... versus when solvent can flow through the bed evenly Larger elution volumes result in a less concentrated analyte and poorer detection limits

26. O OH Silica Surface

27. Porous Silica

28. O Si O O O O O O O O O O O O O O HO O O Si O HO Si O O O OH Ideal C18 Surface Ideal C18 Surface

29. Bonded Silica Surface Residual silanols Si O Si O Si O Si O S O Si Si O Si O Si OH S OH OH OH i i O O pH<3

30. Polar Secondary Interactions Si O Si O Si O Si O Si O Si Si O Si O Si OH Si O H O O H H O H NH 3 + O O pH<3

31. Ionic Secondary Interactions O Si O Si O Si O Si O Si O Si Si O Si O Si O Si O O-O- O pH>7 O H3NH3N +

32. Multiple Interactions Si O Si O Si O Si O Si O Si Si O Si O Si OH Si O H O O H H SO 3 - NH 3 + 3 + O O

33. SPE Mechanism Selection (1) NH 2 NH 3 + NH 2 Functionality AnalyteMechanism Hydrophobic H-Bonding Ionic Non-Polar Polar Ion-Exchange

34. Analyte Characteristics v Functional groups v Solubilities v pKa values... pKa Database v www.zirchrom.com/organic.htm v Chromatographic behavior Hydrophobic H-Bonding CationicAnionic OH HN NH 2 SH NH 3 + NH 2 + NH + COO - S O O O - P O O - O -

35. Molecular Interactions

36. SPE Mechanism Selection (2) OH HN NH 2 SH N H 3 + NH 2 + NH + S O O O - P O O - O - NR 3 + Aqueous Non-Polar Solvent Aqueous (Low Ionic Strength) Aqueous (Low Ionic Strength) Hydrophobic H-Bonding Cations Anions Non-Polar Polar Cation Exchange Anion Exchange C18C8 C2CH PH CN SI 2OH NH2 CBA PRS SCX NH2SAX AnalyteMatrixSorbent