1. ENVIRONMENTAL ANALYSIS I. Solid Phase Extraction
step-by-step
method development

2. Goal of Six-Step Mode
Isolation
Purification
Trace enrichment

3. Six-Step Method Development
Compile known parameters
Tech Support Form
Follow step-by-step guide
Fill in method development form

4. Technical Support Form

5. Pertinent Data The analyte(s) The matrix The analytical method
Existing method(s)

6. The Analyte Structure pKa(s) Concentration Sample volume Solubility
Stability

7. The Matrix
Type of matrix
pH and ionic strength
Interferences

8. The Analytical Method Final technique Compatible solvents
HPLC conditions

9. Existing Method Liquid-liquid method Current SPE method details
Data from current method

10. Method Development Example
Pollution Unlimited
1224 Sunnyside Drive
Elution, Alaska
(907)
(907)
Dr. Anna Light

11. The Analyte

12. The Analyte (cont.)

13. The Matrix

14. The Analytical Method

15. Other Information

16. Following the Guide

17. The “Form” Sample name _________________
Analyte of Interest ________________
Sample size ____________________
Sorbent Bed Mass _______ Cartridge volume_____
Sorbent phases to try 1) _____ 2) _____ 3) _____ 4) _____
1. Sample pretreatment ___________________________
2. Condition column with ____ mL of _________ at ___ mL/min
3. Equilibrate column with ____ mL of _________ at ___ mL/min.
4. Load sample at ______ mL per minute.
5. Remove interference with ____ mL of _________ at ___ L/min.
Drying step _____ minutes
6. Elute analyte with ____ mL of _________ at ___ mL/min.
Remember to include soak step if possible!

18. Filling in the Form

19. Following the Guide

20. Sample Size Determination

21. Filling in the Form

22. 0.3 to 0.6 milli-equivalents per gram
Sorbent Capacity
Hydrophobic phases:
1- 5% of Sorbent Mass
Ion Exchangers
0.3 to 0.6 milli-equivalents per gram

23. Following the Guide

24. Select a Mechanism

25. Select a Mechanism

26. Select a Mechanism

27. Select a Mechanism

28. Following the Guide

29. Selecting a Sorbent

30. Filling in the Form

31. Following the Guide

32. A Reminder...

33. Sample Pretreatment

34. Filling in the Form

35. Following the Guide

36. Conditioning and Equilibration

37. Conditioning and Equilibration

38. Filling in the Form

39. Following the Guide

40. Sample Loading Rate

41. Filling in the Form

42. Following the Guide

43. Removing Interferences

44. Removing Interferences

45. Filling in the Form

46. Following the Guide

47. Drying Time

48. Filling in the Form

49. Following the Guide

50. Selecting an Elution Solvent

51. Elution Solvent Volume

52. The Completed Form

53. What if Low Recovery? Losses during rinse step Analyte breakthrough
Poor elution

54. Losses During Rinse Step
Check rinse for presence of analyte

55. Analyte Breakthrough
Stack Columns
Elute Separately

56. Analyte Breakthrough
Check pH and ionic strength at each step of extraction procedure
Select phase offering greater interaction

57. Impact of pH Analyte loaded at pH=2 Analyte loaded at pH=7
The analyte loaded at pH=2 is well retained on the column. The analyte loaded at pH=7 bleeds through the column.
At a pH=2, the surface of the silica is neutral, since the silanols are protonated (SiOH). At pH=7, the surface is negatively charged, since the silanols are ionized (SiO-). The very water soluble, negatively charged analyte is much more likely to be retained on a surface that is neutral than one that is negatively charged.

58. Selection of Bonded Phase
Analyte poorly retained on C2
Analyte well retained on C18(EC)
Analyte well retained on C8

59. Poor Elution Select stronger elution solvent
pH, ionic strength, % organics
Select less retentive sorbent
Include a soak step

60. Solvent Strength Elution with 5% methanol in water

61. Less Retentive Sorbent
Analyte completely eluted from C8
Traces of analyte remain on C18(EC)

62. Soak Step
The analyte is completely eluted from the column which was soaked for one minute with the elution solvent.
Some analyte remains on the column that was not soaked in the elution solvent.