Speakers. Moderator. John V Hinshaw GC Dept. Dean CHROMacademy. Tony Taylor Technical Director CHROMacademy. Dave Walsh Editor In Chief LCGC Magazine
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3 Speakers John V Hinshaw GC Dept. Dean CHROMacademy Tony Taylor Technical Director CHROMacademy Moderator Dave Walsh Editor In Chief LCGC Magazine
4
5 An Introduction to Column Selection for Capillary GC
6 Aims & Objectives 1. Important analyte / stationary phase interactions 2. GC stationary phase polymer types 3. Selecting an appropriate stationary phase 4. Stationary Phase Selectivity Effects tuning the chemistry 5. Choosing column dimensions 6. Effects of column dimensions on selectivity, retention, efficiency and resolution 7. Bringing it all together from a practical standpoint
7 Capillary GC column selection - what's important? Stationary Phase Length (m) x Internal Diameter (mm) x Film Thickness (mm) L r c d f 5% Diphenyl dimethylpolysiloxane 30m x 0.25mm x 0.25mm Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / phase
8 Stationary phase selection: Major analyte / stationary phase interactions Major Analyte / Stationary Phase Interactions Electronegativity / Dipole Moments / Polarity Non-polar Polar - Dipole
9 Stationary phase selection: Dispersive interactions 1. All substances contain small dipoles (small electronegativity differences) 2. Instantaneous dipoles fluctuate throughout the molecule (electron / nuclei vibration) 3. As two molecules approach transient dipoles can induce the opposite dipole in the other molecule and a small attractive effect is seen 4. Often called a dispersive interaction and occurs between compounds which are predominantly non-polar 5. Dispersive interactions occur with all substances, regardless if there is another overriding interaction Van der Waals Forces
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11 Stationary phase selection: Dipole interactions 1. Dipole interactions come in two sorts: Dipole-Dipole / Dipole-Induced Dipole 1. Dipole interactions occur between substances whose permanent dipoles come into close contact with each other 2. Dipole-Induced dipole interactions occur when a polar substance meets a polarisable compound (typically containing pi-electrons) 3. The stronger dipole induces a more permanent dipole in the other substance and an intermolecular attraction occurs Dipole Dipole Interaction
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14 Stationary phase selection: Hydrogen Bonding Interactions 1. A special case of a dipole-dipole interaction 2. Dipoles associated with the functional groups of two molecules come into close proximity 3. Hydrogen bonding interactions are very strong compared to dispersive interactions 4. In the extreme (e.g. the association of water with methanol) the dipole-dipole interaction energy can approach that of a chemical bond 5. Still an underlying weak dispersive interaction occurring simultaneously Hydrogen Bonding Interactions
15 Modern Stationary Phase Chemistry 1. Immobilized Polymeric Liquids bonded to the inner surface of a silica capillary via silyl-ether linkages 2. Deactivation treatments applied before and/or after bonding Polysiloxane Phases Typical Ratio of monomers (X:Y): 5:95 35:65 50:50 Higher percentage of functional monomer indicate higher degree of that interaction 50:50 phase shows stronger Induced Dipole Interactions with Aromatics
16 Modern Stationary Phase Chemistry (III) Polysiloxane Phases Typical Ratio of monomers (X:Y): 6:94 14:86 50:50 Typical Ratio of monomers (X:Y): 35:65 50:50
17 Modern Stationary Phase Chemistry (III) Glycol / Wax Phases Stationary Phase Interaction Summary Lower polarity phases bleed less!
18 Stationary Phase Selection 1. Critical Phase and Temperature directly effect selectivity 2. Principle of like dissolves like holds well 3. Separate polar analytes using a more polar phase and vice versa 4. The skill is knowing the degree of polarity required to avoid overly long retention times whilst still obtaining a satisfactory separation 5. Separating compounds of intermediate polarity or mixed polarity & functionality requires knowledge of the retentivity and selectivity of each phase 6. May require fine tuning of the phase chemistry using the monomeric ratios Like Dissolves Like
19 Stationary Phase Selection: Test Probe Chemistry
20 Stationary Phase Selection: Dispersive Interactions 100% Methyl Polysiloxane Boiling Point Column? Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12) 110 o C 156 o C 182 o C 174 o C 218 o C 216 o C Strong Dispersion No Dipole No H Bonding
21 Stationary Phase Selection: Dipole (Induced Dipole) Phases 5% Phenyl Phase 5% Phenyl 100% Methyl ,6? Strong Dispersion Weak (Induced) Dipole No H Bonding Strong Dispersion No Dipole No H Bonding 1. Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12)
22 Stationary Phase Selection: Dipole (Induced Dipole) Phases 50% Phenyl Phase 50% Phenyl 100% Methyl ? Strong Dispersion Weak (Induced) Dipole No H Bonding Strong Dispersion No Dipole No H Bonding 1. Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12)
23 Stationary Phase Selection: Dipole & Hydrogen Bonding Phases 14% Cyanopropylphenyl Phase 14% Cyano-propylphenyl Strong Dispersion None/Strong Dipole (Ph/CNPr) Weak/Moderate H Bonding (Ph/CNPr) 100% Methyl? Strong Dispersion No Dipole No H Bonding 1. Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12)
24 Stationary Phase Selection: Dipole & Hydrogen Bonding Phases 50% Cyanopropylphenyl Phase 50% Cyano-propylphenyl Strong Dispersion Strong Dipole Moderate H Bonding 100% Methyl? Strong Dispersion No Dipole No H Bonding 1. Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12)
25 Stationary Phase Selection: Dipole & Hydrogen Bonding Phases 35% Trifluoropropyl Phase 35% Trifluoropropyl 1,2 100% PDMS 1 DB-1 DB Strong Dispersion Moderate Dipole Weak H Bonding 1. p-xylene, 2. m-xylene, 3. o-xylene Column: DB1 / DB200 (30m x 0.25mm, 0.25µm) Carrier: 32 cm/sec. Oven: o 5 o C/min.
26 Stationary Phase Selection: Dipole (& H Bonding) Phases 100% Polyethylene Glycol 100% PEG 100% Methyl ? Strong Dispersion No Dipole No H Bonding Strong Dispersion Strong Dipole Moderate H Bonding 1. Toluene 2. Hexanol 3. Phenol 4. Decane (C10) 5. Naphthalene 6. Dodecane (C12)
27 Stationary Phase Selection: PLOT columns 1. Porous Layer Open Tubular (PLOT) for Gas Solid Chromatographic (GSC) applications 2. PLOT columns are coated with small, solid porous particles using a binder. Particles are Alumina or Molecular sieve 3. Solutes are separated on differences in their adsorption properties, size and shape 4. PLOT columns used to separate highly volatile liquids and permanent gases without the need for cryogenic or subambient cooling of the GC oven
28 Stationary Phase Selection: PLOT columns 5. Alumina columns are well suited to the analysis of C 1 C 10 hydrocarbons and small aromatics 6. KCl derivatised columns produce altered selectivity 7. The Q designated columns show better selectivity for C 1 -C 3 hydrocarbons (not good for >C 6 ) 8. Q columns are also able to separate sulphur gases and most light hydrocarbons. 9. Molecular sieve columns used for noble and permanent gas samples - also good for the separation of solvents. 50m, 0.53mm ID Rt-Alumina PLOT
29 Practically Speaking! 1. If no information or ideas about which stationary phase to use is available, start with a DB-1 or DB-5 2. Low bleed ("ms") columns are usually more inert and have higher temperature limits 3. Use the least polar stationary phase that provides satisfactory resolution and analysis times. Non-polar stationary phases have superior lifetimes to polar phases 4. Use a stationary phase with a polarity similar to that of the solutes. This approach works more times than not; however, the best stationary phase is not always found using this technique
30 Practically Speaking! 5. If poorly separated solutes possess different dipoles or hydrogen bonding strengths, change to a stationary phase with a different amount (not necessarily more) of the dipole or hydrogen bonding interaction 6. If possible, avoid using a stationary phase that contains a functionality that generates a large response with a selective detector % Methyl or 5% Phenyl, 50% Phenyl, 14% Cyanopropylphenyl and WAX (PEG) cover the widest range of selectivities with the smallest number of columns 8. Use PLOT columns for the analysis of gaseous samples at above ambient column temperatures
31 Column Dimensions - Column Length Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 1. Doubling column length doubles efficiency 2. Doubles analysis time (or by x for temperature gradient) 3. Increases column costs 4. Improves resolution by a factor of 1.4 x
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33 Column Internal Diameter (or r c ) Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 1. Affects efficiency, retention, carrier flow rate, capacity and pressure drop across the column 2. Inversely proportional to column efficiency halve diameter, double efficiency, increase resolution by factor of 1.4
34 Column Internal Diameter (or r c ) (II) Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 3. Inversely proportional to analyte retention for isothermal but NOT GRADIENT 4. Consider gradient temperature programming in conjunction with pressure programming for constant flow
35 Column Internal Diameter (or r c ) (III) Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 5. Column head pressure an inverse square function of column radius 6. Column capacity increases with column internal diameter 7. Capacity also depends on the stationary phase type, film thickness and the nature of the analytes
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37 Phase Film Thickness (d f ) Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 1. Affects retention, inertness, capacity, resolution and bleed 2. Film thickness is directly proportional to retention time (1.5:1 for gradient) 3. Thick stationary phase films give retention for highly volatile analytes 4. Increasing film thickness allows retention of volatile analytes at temperatures at or above ambient 5. Doubling d f gives an increase of around 20 o C in elution temperature
38 Phase Film Thickness (d f ) (II) Efficiency (N) carrier gas / L / r c Retention (k) o C / r c / d f Selectivity (a) o C / Phase 6. Retention of late eluting (high boiling point) analytes is reduced using thinner film columns 7. Early eluting analytes (k<2) are better resolved using thicker film columns 8. Resolution may DECREASE for analytes with k values between 5 with INCREASING film thickness 9. Thicker films bleed more - upper temperature limits of thick film columns will be lower 10. Thicker film columns are more inert as the film shields the analyte from active sites on the silica tubing 11. Thicker film columns have higher analyte capacity, and so may reduce peak fronting
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40 Phase Ratio (b) 1. Stationary phase to mobile phase ratio 2. Increasing the phase ratio will result in decreased analyte retention (increasing the column radius or decreasing the film thickness) (reduction in capacity?)
41 Phase Ratio (b) 3. Use to keep retention time approximately constant whilst increasing efficiency (reducing the column internal diameter) and reducing film thickness to keep b constant 4. Net result is a more efficient separation within the same timescale as the original separation!
42 Phase Ratio (b) (II)
43 Practically Speaking! 1. Capillary GC columns are typically 10, 15, 20, 25, 30, 50, 60,120 (m) 2. Extending the column length is the least favoured option for increasing resolution and should be avoided if possible 3. Cost and analysis time are proportional to column length 4. Use the shortest column that will give you the required resolution (begin with m columns if the number/ nature of samples is unknown). 5. To increase resolution try changing the stationary phase or column internal diameter first 6. Narrow internal diameter columns are capable of separating multiple analytes in a single analysis
44 Practically Speaking! 7. Increase film thickness when volatile analytes are involved or reduce film thickness to decrease retention of highly adsorbed analytes 8. Use phase ratio to increase separation efficiency in the same timeframe as the original separation 9. Column head pressure and bleed increase with column length m columns are well suited to samples containing well separated analytes or where the number of analytes is low m columns should be used only where very large numbers of components need to be separated and as a last resort when reducing the column internal diameter and changing the stationary phase and temperature program have failed!
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