PowerPoint PresentationSlide 2Slide 3Slide 4Slide 5Slide 6Slide 7Slide 8Slide 9Slide 10Slide 11Slide 12Slide 13Slide 14Slide 15Slide 16Slide 17Slide 18Slide 19Slide 20Slide 21Slide 22Slide 23Slide 24Liquid Chromatography1. Introduction and Column Packing Material2. Retention Mechanisms in Liquid Chromatography4. Column Preparation6. Detectors(Chapter 4 and 5 in The essence of chromatography)3. Method Development5. General Instrumental aspectsColumn Preparation1. Column is the central part of liquid chromatography.2. Column Packing MethodsA. Dry-Packing Procedures: for rigid particles with diameter greater than 20 m. B. Down-Fill Slurry Packing: for rigid particles with diameter smaller than 20 m.C. Up-Fill Slurry Packing: for rigid particles with diameter smaller than 20 m.Down-Fill Slurry PackingUp-Fill Slurry PackingThis method is used for conventional Diameter columns. It is unsuitable for packing small diameter columns.3. Evaluation of column qualityMajor parameters: Capacity factor, plate numbers, separation factor, and asymmetry factor.Routine Column Quality EvaluationAsymmetry Factor Asymmetry factor = ABSpecific column quality test for chemically bonded phasesGeneral Instrumental aspects 1. Pumps2. Injectors3. DetectorsDirect pressure pumpGas pressure is applied from anexternal gas tank using a high Pressure regulator.No pressure pulses are producedThe solvent reservoir is limitedA major problem is introductionOf gas into the solvent.Reciprocating Pumpa. Motorb. Gearc. Seald. Pistone. Solvent in f. Check valvesg. Solvent outReciprocating PumpOne of the most common type of systemUnlimited reservoir systemA major problem is that it produces variable pressurePressure Variances Since the pump must spend at least a portion of its time filling, there is a pressure drop during the phase. This effect must be minimized because it would greatly affect your sensitivity and detection limit.Reciprocating PumpOne approach is to have a more rapid fill cycleThis does not eliminate the problem by reduce itReciprocating PumpOne could also use two or more pumps working in tandemPulse DampersThe Popular Approach to minimize flow pulse associated withReciprocating pumpsMechanism: Absorb the peak and valley of the pressure pulseGradient ControllerDual pumping systemsA valve system can be sued on each pump that can Provide a different solvents.Valve InjectorsAutomatic InjectorsAutomatic syringeThis method allow for adjustment of sample size.The motor driven syringe can provide sufficient pressureTo inject sample past the check valve.Trace Enrichment SamplingMulti-Dimensional LC Dimension -- column2-D Liquid ChromatographyGeneral Instrumental aspects 1. Pumps2. Injectors3. Detectors1. Solute 1 and 2 are eluted from a reversed-phase column with retention time of 18.1 min and 23.5 min at a flow-rate of 1 mL/min, using a 10% 2-propanol: 90%water mixture as the mobile phase. The void time of the column at this flow-rate is 0.9 min. The strength of 2-propanol and water are 3.9 and 0, respectively.a. What % 2-propanol must be used in order to elute 1 with a retention time of 10.0 min? What will be the retention time of solute 2 under this condition.b. What mixture of water and THF (S’ = 4.4) will allow elution of 1 with a retention time of 10.0