1. ic_sampl_prep_26062003_e.ppt 1 IC Metrohm inline sample preparation Techniques for analytical success Metrohm Ion Chromatography Bart Cleeren, Metrohm België

2. ic_sampl_prep_26062003_e.ppt 2 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

3. ic_sampl_prep_26062003_e.ppt 3 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

4. ic_sampl_prep_26062003_e.ppt 4 IC Metrohm Ion Chromatography Ultrafiltration Inline sample preparation Ultrafiltration cell EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector SSample P2Peristaltic pump A position fill C P1 ED 2

5. ic_sampl_prep_26062003_e.ppt 5 IC Metrohm Ion Chromatography Ultrafiltration Cell Inline sample preparation No particles on the acceptor side Big particles are flushed away and do not clog the membrane High flow on the sample side Low flow on the acceptor side

6. ic_sampl_prep_26062003_e.ppt 6 IC Metrohm Ion Chromatography Ultrafiltration Inline sample preparation # 1 Sample filtration EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector SSample P2Peristaltic pump A position fill C P1 ED 2

7. ic_sampl_prep_26062003_e.ppt 7 IC Metrohm Ion Chromatography Ultrafiltration Inline sample preparation # 2 Sample injection EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector SSample P2Peristaltic pump A position inject C P1 ED 2

8. ic_sampl_prep_26062003_e.ppt 8 IC Metrohm Ion Chromatography Ultrafiltration: vegetable juice Inline sample preparation A Supp 4 – 250 1Organicsn.q. 2Organicsn.q. 3Cl – 4141 4NO 3 – 102 5HPO 4 2– 577 6SO 4 2– 842 7Oxalaten.q. ppt NaHCO 3 /Na 2 CO 3 ; 1.7/1.8 mmol/L; dilution 1:100; 0.15 µm rec. cellul.

9. ic_sampl_prep_26062003_e.ppt 9 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

10. ic_sampl_prep_26062003_e.ppt 10 IC Metrohm Ion Chromatography Inline sample preparation Dialysis cell, acceptor solution EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump P3Peristaltic pump (2 Ch.) Dialysis cell AAcceptor solution Dialysis C P1 ED 2 A E P2 P3 A D C X X

11. ic_sampl_prep_26062003_e.ppt 11 IC Metrohm Ion Chromatography Inline sample preparation # 2 Stopped flow dialysis EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump Injection Valve «B» Dialysis cell AAcceptor solution Dialysis C P1 ED 2 A P3 P2 A P1 C DE X

12. ic_sampl_prep_26062003_e.ppt 12 IC Metrohm Ion Chromatography Dialysis cell Inline sample preparation Continuous flow dialysis Exclusion diameter 0.2 µm Ions can pass Oil droplets, fat micelles, particles are excluded Strong concentration gradient No equilibrium concentration Cellulose acetate membrane

13. ic_sampl_prep_26062003_e.ppt 13 IC Metrohm Ion Chromatography Dialysis cell Inline sample preparation Stopped flow dialysis Dialysis time 2... 8 min Sample volume 10 mL No loss of concentration on the sample side Equilibrium concentration Dialysis rate > 99 % No filtration, no pre concentration Unique technique, that had significantly improves the field of application for IC

14. ic_sampl_prep_26062003_e.ppt 14 IC Metrohm Ion Chromatography Dialysis: milk Inline sample preparation Metrosep Dual 1 – 70 1Chloride938 2Nitraten.q. 3Phosphaten.q 4Sufate103 ppm NaHCO 3 /Na 2 CO 3 ; 2.4/2.0 mmol/L; dilution 1:5; 0.2 µm cellul. acetat

15. ic_sampl_prep_26062003_e.ppt 15 IC Metrohm Ion Chromatography Dialysis: cutting oil emulsion Inline sample preparation Metrosep Dual 2 1Chloride74.7 2Nitrite20.5. 3Nitrate23.9 4n. det.- 5Phosphate25.2 6Sulfate192.7 7n. det.- NaHCO 3 /Na 2 CO 3 ; 2.0/1.3 mmol/L; dilution 1:5; 0.2 µm cellul. acetat

16. ic_sampl_prep_26062003_e.ppt 16 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

17. ic_sampl_prep_26062003_e.ppt 17 IC Metrohm Ion Chromatography Inline sample preparation # 1 Sample preconcentration EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump C P1 D P32 S Inline pre concentration E C P1 P 2 D

18. ic_sampl_prep_26062003_e.ppt 18 IC Metrohm Ion Chromatography Inline sample preparation # 2 Sample analysis EMobile phase: eluent P 1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P 2Peristaltic pump C P1 ED P32 S Inline preconcentration E C D P1P 2

19. ic_sampl_prep_26062003_e.ppt 19 IC Metrohm Ion Chromatography Inline preconcentration: ultra pure water Inline sample preparation Metrosep A Supp 5 – 150 1Acetate2.3 2Formate22.0 3Chloride13.0 4Carbonaten.q. 5Nitrate4.5 6Phosphate16.0 7Sulfate8.5 8Oxalate12.0 ppt NaHCO 3 /Na 2 CO 3 ; 1.0/3.2 mmol/L; 20 mL preconc,

20. ic_sampl_prep_26062003_e.ppt 20 IC Metrohm Ion Chromatography Inline preconcentration: ultra pure water Inline sample preparation Metrosep Dual 2 1Fluoride45 – 150 2Chloride100 – 210 3Bromide0 – 100 4Nitrate10 – 112 5Sulfate40 – 145 ppt NaHCO 3 /Na 2 CO 3 ; 2.0/3.2 mmol/L; 10 mL preconc, spiked 100 ppt

21. ic_sampl_prep_26062003_e.ppt 21 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

22. ic_sampl_prep_26062003_e.ppt 22 IC P3 Metrohm Ion Chromatography Inline sample preparation Transfer solution «T» EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump Injection Valve «B» P3Peristaltic pump TTransfer solution A position fill, B position fill C P1 ED 2 T Matrix elimination

23. ic_sampl_prep_26062003_e.ppt 23 IC P3 Metrohm Ion Chromatography Inline sample preparation # 1 Preconcentrator column inline EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump Injection Valve «B» P3Peristaltic pump TTransfer solution A position fill, B position fill C P1 ED 2 T Matrix elimination

24. ic_sampl_prep_26062003_e.ppt 24 IC P3 Metrohm Ion Chromatography Inline sample preparation # 2 Sample transfer, matrix elim. EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump Injection Valve «B» P3Peristaltic pump TTransfer solution A position fill, B position inject C P1 ED 2 T Matrix elimination

25. ic_sampl_prep_26062003_e.ppt 25 IC P3 Metrohm Ion Chromatography Inline sample preparation # 3 Sample inject EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2Peristaltic pump Injection Valve «B» P3Peristaltic pump TTransfer solution A position inject, B position inject C P1 ED 2 T Matrix elimination

26. ic_sampl_prep_26062003_e.ppt 26 IC Metrohm Ion Chromatography Matrix elimination: organic solvents Inline sample preparation 50 µL iPrOH Bwith matrix elimination Awithout matrix elimination ppt F–F– Ac – 443 Cl – 30 NO 3 – 28 SO 4 2– 425 Cl – 24 NO 3 – 17 SO 4 2–

27. ic_sampl_prep_26062003_e.ppt 27 IC Metrohm Ion Chromatography Matrix elimination: 35 % H 2 O 2 Inline sample preparation left: without m.e. Loop 20 µL right: with m.e. Loop 20 µL transfer vol. 2 mL

28. ic_sampl_prep_26062003_e.ppt 28 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

29. ic_sampl_prep_26062003_e.ppt 29 IC P3 Metrohm Ion Chromatography Inline sample preparation 833.0030 IC Sample Prep. Unit EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2, 3Peristaltic pumps Injection Valve «B» TTransfer solution 833 IC Sample Prep A position inject, B position fill C P1 ED 2 T Matrix neutralization

30. ic_sampl_prep_26062003_e.ppt 30 IC P3 Metrohm Ion Chromatography Inline sample preparation # 1 Preconcentrator column inline EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2, 3Peristaltic pumps Injection Valve «B» TTransfer solution 833 IC Sample Prep A position fill, B position fill C P1 ED 2 T Matrix neutralization

31. ic_sampl_prep_26062003_e.ppt 31 IC P3 Metrohm Ion Chromatography Inline sample preparation # 2 Sample transfer/neutralization EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2, 3Peristaltic pumps Injection Valve «B» TTransfer solution 833 IC Sample Prep A position fill, B position inject C P1 ED 2 T Matrix neutralization

32. ic_sampl_prep_26062003_e.ppt 32 IC P3 Metrohm Ion Chromatography Inline sample preparation # 4 Sample inject EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2, 3Peristaltic pumps Injection Valve «B» TTransfer solution 833 IC Sample Prep A position inject, B position inject C P1 ED 2 T Matrix neutralization

33. ic_sampl_prep_26062003_e.ppt 33 IC 0.74 F – 3.6 Cl – 0.97 NO 3 – 0.98 HPO 4 2– 1.1 SO 4 2– Metrohm Ion Chromatography Matrix neutralization: caustic soda Inline sample preparation 30 % NaOH, 20 µL loop Bwith matrix elimination Awithout matrix elimination ppm

34. ic_sampl_prep_26062003_e.ppt 34 IC Metrohm Ion Chromatography Matrix neutralization: caustic soda Inline sample preparation Metrosep Dual 2 1Chloride25.08 2Carbonaten.q. 3Chlorate4.76 4Sulfate10.43 ppm NaHCO 3 /Na 2 CO 3 ; 1.7/1.8 mmol/L; 50 % NaOH, 20 µL loop

35. ic_sampl_prep_26062003_e.ppt 35 IC 1 Metrohm Ion Chromatography Matrix neutralization: ammonia Inline sample preparation Metrosep A Supp 4 – 250 1Formaten.q. 2Chloride0.03 3Nitriten.q. 4Bromide0.95 5Nitrat0.04 6Carbonaten.q. 7Sulfate0.13 ppm NaHCO 3 /Na 2 CO 3 ; 1.7/1.8 mmol/L; 26 % NH 3, 20 µL loop 2 3 4 5 6 7

36. ic_sampl_prep_26062003_e.ppt 36 IC P3 Metrohm Ion Chromatography Inline sample preparation OH cartridge EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P2, 3Peristaltic pumps Injection Valve «B» TTransfer solution Cartridge A position fill, B position fill C P1 ED 2 T Matrix neutralization Ctr

37. ic_sampl_prep_26062003_e.ppt 37 IC Metrohm Ion Chromatography Matrix neutralization: HCl Inline sample preparation 10 µL HCl direct injection ppm Metrosep C2 – 250

38. ic_sampl_prep_26062003_e.ppt 38 IC Metrohm Ion Chromatography Matrix neutralization: HCl Inline sample preparation Metrosep C2 – 250 1Nickeln.q. 2Sodium5.4 3Ammonium1.3 4Potassium0.8 5Magnesium1.5 6Calcium0.8 ppm Tartaric-/dipicolinic acid; 4.0/0.75 mmol/L; 7 mol HCl, 10 µL loop 1 2 3 456

39. ic_sampl_prep_26062003_e.ppt 39 IC Metrohm Ion Chromatography Inline sample preparation Ultra filtration Stop flow dialysis Pre concentration technique Matrix elimination Matrix neutralization In- line calibration

40. ic_sampl_prep_26062003_e.ppt 40 IC Metrohm Ion Chromatography Inline sample preparation Standard S, flow path EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P1,2,3Peristaltic pumps Injection Valve «B» SStandard A position inject, B position fill Inline calibration C P1 ED P32 S

41. ic_sampl_prep_26062003_e.ppt 41 IC Metrohm Ion Chromatography Inline sample preparation # 1 Measurement of standard EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P1,2,3Peristaltic pumps Injection Valve «B» SStandard A position inject, B position fill C P1 ED P32 S Inline calibration

42. ic_sampl_prep_26062003_e.ppt 42 IC Metrohm Ion Chromatography Inline sample preparation # 2 Preconcentrator inline EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P1,2,3Peristaltic pumps Injection Valve «B» SStandard A position fill, B position fill C P1 ED P32 S Inline calibration

43. ic_sampl_prep_26062003_e.ppt 43 IC Metrohm Ion Chromatography Inline sample preparation # 3 Transfer of standard EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P1,2,3Peristaltic pumps Injection Valve «B» SStandard A position fill, B position inject C P1 ED P32 S Inline calibration

44. ic_sampl_prep_26062003_e.ppt 44 IC Metrohm Ion Chromatography Inline sample preparation # 4 Inject of standard EMobile phase: eluent P1High pressure pump Injection valve «A» CAnalytical column »MSM» Suppressor DConductivity detector P1,2,3Peristaltic pumps Injection Valve «B» SStandard A position inject, B position inject C P1 ED P32 S Inline calibration

45. ic_sampl_prep_26062003_e.ppt 45 IC Metrohm Ion Chromatography Inline sample preparation # 1: Transfer of 2 mL ultrapure water to the preconcentrator Blank value # 2: Transfer of 2 mL ultrapure water to the preconcentrator. Valve B is thrown from fill to inject and back once Calibration level 1 # 3: Transfer of 2 mL ultrapure water to the preconcentrator. Valve B is thrown from fill to inject and back twice Calibration level 2 # 4: Transfer of 2 mL ultrapure water to the preconcentrator. Valve B is thrown from fill to inject and back three times Calibration level 3, etc Inline calibration

46. ic_sampl_prep_26062003_e.ppt 46 IC Metrohm Ion Chromatography Inline sample preparation Inline calibration 100 ppt 100... 400 ppt

47. ic_sampl_prep_26062003_e.ppt 47 IC inline sample preparation ? Metrohm can do that Metrohm Ion Chromatography