1. SoSe 2006 Alexander Semmler: HPLC 1 Alexander Semmler HPLC High pressure liquid chromatography High performance -‘‘-

2. SoSe 2006Alexander Semmler: HPLC2 Basic Principle Sample Injection Separation Detection Utilisation by integration and calculation Injector ColumnColumn Detector EDU

3. SoSe 2006Alexander Semmler: HPLC3 Device

4. SoSe 2006 Alexander Semmler: HPLC 4 Step 1 Sample Injection

5. SoSe 2006Alexander Semmler: HPLC5 Sample injection Sample is inserted into the device Syringe with or without needle Sample is mixed with eluent solution Mixture is inserted into column

6. SoSe 2006Alexander Semmler: HPLC6 Injector

7. SoSe 2006Alexander Semmler: HPLC7 Injection (II) I N J E C T O RSeparation Elu- ent Sample PUMP syringe bottle with eluent solution

8. SoSe 2006 Alexander Semmler: HPLC 8 Step 2 Separation

9. SoSe 2006Alexander Semmler: HPLC9 Separation mobile phase: eluent-solution, liquid stationary phase: column, solid separation by different polarities packed columns

10. SoSe 2006Alexander Semmler: HPLC10 Separation (II) Pre- column C O L U M N Det Waste to save the column

11. SoSe 2006 Alexander Semmler: HPLC 11 Step 3 Detection

12. SoSe 2006Alexander Semmler: HPLC12 Detection one substance group after the other flows through the detector detection by intensity of different detectors sending data to data utilisation

13. SoSe 2006Alexander Semmler: HPLC13 Possible detectors refractive index – RI UV/VIS IR conductivity mass spectroscopy – MS (coupled)

14. SoSe 2006Alexander Semmler: HPLC14 Detection (II) Detector Solution separated components DU Waste

15. SoSe 2006 Alexander Semmler: HPLC 15 Step 4 Data Utilisation

16. SoSe 2006Alexander Semmler: HPLC16 Data Utilisation PC/integrator – printer/plotter intensity against time graph calculation of retention time & integration of the peaks qualitative information: retention time (Rf-value) quantitative information: peak size (area)

17. SoSe 2006Alexander Semmler: HPLC17 Data Utilisation (II) Detector Computer Result- database Printer Calibration- database

18. SoSe 2006Alexander Semmler: HPLC18 Chromatogramm baseline peaks

19. SoSe 2006 Alexander Semmler: HPLC 19 Answering Questions For further information please contact your analytical chemistry teacher

20. SoSe 2006 Alexander Semmler: HPLC 20 Thank you for listening Visit semmler.samcal.org