Solid phase extraction (SPE) is a sample preparation technique that selectively enriches analytes from a sample matrix. It involves passing a sample through a cartridge containing a solid stationary phase. Analytes selectively bind to the phase based on properties like polarity while interfering matrix components are removed. Bound analytes can then be eluted and analyzed. SPE offers advantages over liquid-liquid extraction like higher selectivity and cleaner extracts. It has various applications including impurity profiling, environmental analysis, food chemistry, and biological analysis. Solid phase microextraction (SPME) is a related technique that extracts analytes directly from vapors using a coated fiber for analysis by gas chromatography.

1. seminar onSOLID PHASE EXTRACTION AND APPLICATIONMODH SUDIP C.PA/2010/11NIPER HYDERABAD

2. Introduction

3. Principle

4. Types Solid phases

5. Experimental steps

6. Trace enrichment

7. Solid phase micro extraction (SPME)

8. Comparison with HPLC

9. Application Contents2

10. What is the solid phase extraction?3

11. DEFINATION OF SOLID PHASE EXTRACTION (SPE):“A solid phase extraction consists of bringing a liquid or gaseous test sample in contact with a solid phase, whereby the analyte is selectively adsorbed on the surface of the solid phase” Other solvents (liquids or gases)added to remove possible adsorbed matrix componentsEluting solvent added to desorb analyte selectively4

12. SYNONYMSLiquid-solid extraction Column extraction Digital chromatography Bonded phase extraction Selective adsorption techniques5

13. Strategies for solid phase extraction Active substance can be:Unretained- while matrix interference are adsorbed

14. Retained-while matrix interference are washed through6

15. Retained-while matrix interference are washed through7

16. Principle of Solid Phase Extraction:Partitioning of compounds between two phases of solid and liquid Must having greater affinity for the solid phase than for the sample matrixCompounds retained on the solid phase can be removed by eluting solvent with a greater affinity for the analytes pH changes can be useful8

17. In modern SPE the adsorbent is packed between two flitted disks in polypropylene cartridgeand liquid phases are passed through the cartridge either by suction or by positive pressure9

18. Cartridge10

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20. OVERVIEW OF SPE12

21. Solid phases Activated charcoalAluminaSilica gelMagnesium silicate (Florisil)Chemically bonded silica phases and polymersE.g. styrene divinylbenzene13

22. According to chemical nature of The functional group bonded to the silica or the copolymer The resulting phases are classified as Non-polar

23. Polar

24. Ion exchangers It gives different mode of chromatographyOther solid supports Polymeric resins, cellulose and zirconia14

25. Zirconia coated silica as a stationary phase15

26. Examples of selective stationary phasesReaction of phenobarbitone with pentafluorobenzyl bromide onto the adsorbent Amphetamine by Chiral derivatization of solid PhasesDoxorubicin by the metal-loaded phases in which metal cation is loaded onto a reagent-labelled phaseMolecularly imprinted polymers synthetic polymeric materials with specific cavitiesdesigned for a template molecule16

27. Technical Data - Solid Phase Extraction (SPE) Media ProductSorbent Abbreviations Description ODS Octadecyl silica 5% carbon loadODS-4 Octadecyl silica 14% carbon load, end capped*ODS-5 Octadecyl silica 18% carbon load, end cappedC-8 Octyl silica 8.5% carbon load, end cappedFLO Florisil™ Magnesium silicate NH2 Weak anion exchanger Primary amineSAX Strong anion exchanger Quaternary amine (-NR3+)SCX Strong cation exchanger Aromatic benzene sulfonic acidSIL Normal phase silica * End capping masks residual silanol groups, reducing ionic affinity for amines.17

28. Experimental procedure of five steps:Activation of sorbent by appropriate solvent that conditions the surface of the solid2. Removal of solvent by liquid similar to the sample matrix3. Application of sample, the analytes retained by the sorbent 4. Removal of interfering compounds retained in step 3 with a solvent, but shouldn’t remove the analytes (washing step)5.Elution of the analytes with an appropriate solvent (desorption or elution step) and collecting for analysis 18

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34. Other technique can be usedSupercritical fluid Thermal desorption for analytes of high volatility and thermal stabilityThermal desorption with GC for occupational hygiene analysis24

35. Analyte eluted with an organic, relatively volatile solvent is evaporated to drynessThen residue dissolved in appropriate solventDue to evaporation step, speed with SPE is lost25

36. IMMUNOAFFINITY PHASES:Highly selective packings of Immunoaffinity phases of specific antibody immobilised on solid support such as agarose or silicaUseful for selective extraction of biological importanceSubstanceDiagnosis of cancer ELISA TEST 26

37. IMMUNOAFFINITY PHASES27

38. TRACE ENRICHMENT WITH SPESensitivity depends onPhysicochemical properties of the AnalyteDetection systemSelective clean-up Isolation and concentration step28

39. Solid phase microextraction: SPME is the technique in which by using special instrument, sampling is possible in a vapour state29

40. Design of SPMESyringe like instrumentFused silica fiber of a small size and cylindrical shapeconnected to stainless-steel tube for additional mechanical strength and repeated sampling30

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42. Fused silica fibre coated with thin film of several polymeric stationary phasesReusable and replaceableSmall size and cylindrical geometry of fiberPlacement into sample or headspace is easyLoading in desorption chamber of GC orInterphase of the HPLC without any modification of Plunger 32

43. Working with SPMEFiber is first drawn into the syringe needleLowered into the vial by pressing the plungerFiber cleaned before analysis to remove contaminantsCleaning can be performed in the desorption chamber of HPLC by running solventCleaned fiber coating is exposed to a sample matrix for a predetermined, fixed period33

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45. Limits of detection at the µg/L level with using a flame Ionization detector Limits of detection as low as ng/ L can be reached with an ion-trap mass spectrometer35

46. Extraction can be performed in two ways1) Headspace SPME or HS-SPMEFiber is exposed in the vapour phase above a gaseous, liquid, or solid sample2) Direct immersion or DI-SPMEFiber is directly immersed in liquid samples36

47. Available SPME Fibres, by Film Type•Absorption Fibres-Polydimethylsiloxane (PDMS) 7, 30, and 100μm Unpolar-Polyacrylate (PA) Polar-Polyethylene glycol (PEG) Polar •Adsorption fibres (with particles) -Carboxen-polydimethylsiloxane (CAR-PDMS) Adsorption-Polydimethylsiloxane- divinylbenzene (PDMS-DVB) Adsorption-Divinylbenzene/ Carboxen-Polydimethylsiloxane (DVB-CAR-PDMS) Adsorption 37

48. SPME applied to liquid, gaseous or heavily contaminated samples chemicals likeSubstituted benzene compounds Polyaromatic hydrocarbonsNitro- and chlorophenols Naphthols volatile organochlorine compounds polychlorinated biphenyl congeners caffeineMetallic ions 38

49. The SPE process can be performed in a two ways: On-line Off-lineIn offline SPE eluate from the cartridge is introduced into the chromatograph by means of an injection valveIn on-line SPE the extraction cartridge is inserted as part of chromatographic equipment, as loops or high pressure stream of the mobile phase39

50. Types of online SPE:SPE-GC(SPME-GC)SPE-HPLC40

51. ONLINE SPE-GC41

52. ONLINE SPE-HPLC42

53. 10ppb Nitrosamines in Water: SPME-GC/MSChromatogram courtesy of J. Clark, Liggett Group, Inc.43

54. ONLINE SPE-HPLCTHT= Tetra hydro thiophene44

55. Comparison of SPE-HPLC and SPME-GC SPE-HPLC SPME-GCUniversality Compounds +++ +Detection Sensitivity ++ +++ Selectivity +++ ++ Identification + +++ Detection limit (µ/ L) 0.05-0.8 0.2-5 Reproducibility (%) 1-15 4-14Analysis time 90 20Sample volume (mL) 200 2Automation +++ +++Simplicity + +++ 45

56. Comparison of SPE and HPLC Theoretical basis as HPLCRetention and selectivity remain unaffected by particle sizeEfficiency dependent on:Particle sizeColumn geometryTypical number of platesHPLC ~ 10,000SPE < 50Minimum Selectivity(alpha)for Rs=1.2HPLC 1.06SPE 3.9546