Introduction and Principles of Gas Chromatography

Transcription

1 Introduction and Principles of Gas Chromatography Jaap de Zeeuw Restek, Middelburg, The Netherlands

2 Definition and Uses of Gas Chromatography GC Components and Types of Columns Factors Affecting Chromatographic Separation Basic Terminology and Theory

3 Which Industries Use Chromatography? Chemical/Petrochemical Clinical/Forensic Consumer Products Environmental Food Pharmaceutical

4 Why Gas Chromatography? Simple Cheap (can be automated) Short analysis times High Accuracy Qualitative and Quantitative analysis Applicable in % to ppb level

5 General Nitrogen Hydrocarbons PCBs* Tars, Oils, waxes Radon Olefins Sulfur components Ethylene Glycol Aldehydes Phenols & cresols Amines Organic sulfur Organo metallic compounds Oxygenates Which components do we see in Natural Gas? * Ref: California proposed Public Gas Warning List

6 Important Separations in Natural Gas analysis Hydrocarbons Rtx-1 Thin-film Nitrogen - methane Rt Q-BOND C5 C6 50 m x 0.32 mm CP-Sil 5 CB, 1.2 um Helium, 160 kpa 40 C, --> 200 C, 5 C/min C8 C9 C10 C11 40 min Sulfur Rtx-1 thick-film Oxygenates Polar phase (wax) CO2 H2S 50 m x 0.32 mm CP-Sil 5 CB, 5 µm 40,2 C H2, 65 kpa; TCD Natural gas peak 10 m x 0.53 mm CP-Lowox 150 C(2 min) -->200 C, 10 C/min Direct Injection, 50 µl CH4 + N2 C2 C3 Methanol 20 ppm i-c4 C4 10 min 9 min Methanol well separated from matrix Symmetrical peak for methanol

7 IUPAC Definition of Chromatography A physical method of separating sample components from a mixture by selective adsorption or partitioning of the analyte between two phases: a mobile phase and a stationary phase

8 Chromatography Phases Mobile Phases Liquids (methanol, water ) Changing dielectric strength, temperature, ph Gases (nitrogen, helium, hydrogen, argon) Stationary Phases Solids (alumina, silica, polymers, carbon ) Adsorption chromatography Liquids (siloxanes, polyethylene glycols ) Partition (distribution) chromatography

9 stationary phases Commercial phases are very dirty.. Specialized polymer chemists Lowest specifications possible Innovation, unique phase technologies High purity /viscosity polymers for lowbleed and stable stationary phases

10 GC Components and Types of Columns Components of a Gas Chromatograph Types of GC Columns Types of GC Capillary Columns

11 Components of a Gas Chromatograph

12 Gas Purification Equipment Triple Filter 12

13 New Filter Triple filter O2 indicator Used Filter New O2 Indicator Saturated O2 Indicator Color change

14 New Filter Triple filter H2O indicator Used Filter New H2O Indicator Saturated H2O Indicator Color change

15 Components of a Gas Chromatograph

16 Types of GC columns

17 Types of GC Columns Packed Capillary Length, [meters] ID, [millimeters] Theoretical plates 5,000 (2m) 120,000 (30m) Capacity [ng] 10, (0.25mm ID) Amount of Liquid phase 1-30 % μm Price (30m, 0.25 mm ID)

18 Capillary Column Materials Fused Silica Synthetic, amorphous glass with very low (<1ppm) metallic oxide impurities Protective outer coating of polyimide resin imparts flexibility but coil diameter is limited Excellent inertness and useable up to approximately 380 o C(400 o C) Metal Tubing-MXT MXT, stainless steel, surface coated, Siltek deactivated Can be coiled to small diameter Almost as inert as fused silica but useable up to approx. 450ºC

19 Types of GC Capillary Columns WCOT (Wall Coated Open Tubular) Partition chromatography Typical phases: Siloxanes and Polyethylene glycols 0.10 to 0.53mm internal diameters PLOT (Porous Layer Open Tubular) Adsorption chromatography Gases, light hydrocarbons/solvents analysis Adsorbents: molecular sieve, porous polymers, alumina (<1 um particle diameter) 0.25 to 0.53mm internal diameter

20 Compounds Amenable to Gas Chromatography Should be thermally stable Should be un-reactive and non-absorptive to chromatographic system Should be volatile at temperatures below C Presence of polar groups reduces volatility

21 Sample Transfer Injection: how the sample is transferred to the column As a liquid via syringe Non-liquid techniques Purge & trap Headspace Gas sample loop NOTE: It is critical to get the sample into the column in a focused band

22 Peak width of eluting component σ injection + σ column + σ detection = Σ peak + + = + + =

23 Sample introduction Need to realize smallest possible sample band on the capillary column.. Injection Techniques commonly used: Split Splitless Thermal desorption (PTV) Headspace On-Column % - 5 ppm levels Small amount injected Narrow injection band ppm-ppb levels Large amount injected Must use a focusing mechanism..

24 Sample Transfer How to Get a Focused Initial Band Analyte Focusing Higher boiling components Increase retention (stationary phases type, film and temperature) Solvent Focusing All components, but must elute later then solvent itself

25 Analyte focusing If analytes are high boiling, they will be focused by the retention of the stationary phase If analytes have lower boiling points this will show itself as smeared peaks

26 Analyte focusing: example focused peaks Smeared peaks Not enough retention for focusing

27 Solvent-Focusing in splitless injection In splitless injection some solvent condensation is required to create the solvent effect. This solvent will trap (= focus) compounds and makes sure that a narrow band is formed. Realization: Oven temperature during injection must be 20 C below the Bp of the solvent.

28 Splitless Injection at oven temperature 20 C BELOW BP of solvent The solvent effect makes sure all peaks are focused

29 Focusing in Splitless injection No Focusing: Long Initial Sample Band, broad peaks Focusing: Correct solvent peak and narrow peaks

30 Split/Splitless injection system Silicone septum

31 Split injection Considerations 1-3 ml/min into the column ml/min in the inlet (mostly split vent flow) Split Ratio: Column flow rate : 1 ml/min Split vent flow rate : 100 ml/min Split ratio = Column Flow rate = 1 Split-vent + Column flow rate Split ratio ~ 1:100

32 Impact of Split Ratio Increasing the split ratio decreases the peak area, if all other variables are equal. 25:1 Split 50:1 Split SPLIT Column Head Pressure Total Flow Septum Purge Split Vent Purge Vent

33 Factors impacting the separation

34 Non-Column Factors Affecting Separation Carrier gas: type & linear velocity Temperature Injection bandwidth

35 Gas Carrier and Linear Velocity Hydrogen : cm/sec Helium : cm/sec Nitrogen : cm/sec

36 Carrier Gas and Linear Velocity Isothermal Analysis Hydrogen is 2x faster than helium and 4x faster than nitrogen Temperature Programmed Analysis Need to optimize temperature program to get SAME elution temperatures; Also here Hydrogen is 2x faster then helium

37 Hydrogen is of interest Fastest analysis Availability ( can be generated) Need less sample for same signal (maintenance) Deal with safety issues Setting of constant flow (impossible to built up high H2 levels Use H2-detection (will cost $, but makes safety officer happy) Use metal (MXT) columns (also VERY inert)

38 Column length 10/15 m < 10 components and Fast analysis 25/30 m components 50/60 m complex mixtures: > 20 components Most widely used is 30m Do we need extreme LONG columns?

39 Summary: Effect of Length Retention time proportional to length in isothermal analysis but not proportional in temperature program analysis Gain in resolution is not double, but are The BETTER the Chromatographer..... the SHORTER the column..

40 Examples where we need LONG columns Detailed Hydrocarbon Analysis (>300 components) Bio-ethanol (to get isobutane-methanol separation) Separation of PCB isomers (209 congeners) Separation of Cis and Trans FAME isomers

41 Column Internal Diameter 0.53 mm High flow and loadability Direct injection via insert or valve (analyzer) TCD detection Retention gap for On-column 0.32 mm On-column injection; Thick films are possible Electronic Gas / Pressure Control 0.25 mm Ideal for split and splitless injection Relatively high plate number 0.18/0.15/0.10 mm Short analysis times Low bleed / GC x GC

42 Internal diameter 0.10mm 0.15/0.18mm 0.25mm 0.32mm 0.53 mm Practically the following dimensions are used : 0.10 <1 % 0.15/0.18 mm 3 % 0.25 mm 45 % 0.32 mm 35 % 0.53 mm 10 %

43 Basic Terminology and Theory Resolution (R) Theoretical Plates (N eff ) Height Equivalent to a Theoretical Plate (HETP) Phase Ratio (ß) Retention (Capacity) Factor (k) Retention Time (t) Column Selectivity Factor (α)

44 The Resolution R s Quality of Separation between 2 Peaks Dimensionless Parameters: - Selectivity - Retention - Efficiency

45 Resolution Resolution depends on: α : Selectivity k : Retention Factor N th : Plate Number

46 Impact of Nth on resolution Increase N: Longer column Smaller Internal diameter Impact of Higher N using 2x smaller diameter, same length

47 Impact of k on resolution Increase k: use thicker film (same column dimensions) decrease oven temperature (ever 15C, k changes a factor 2) CH4 Impact of using 2x thicker film, Same column dimensions and temp.

48 Impact of α on resolution Increase alpha: use different stationary phase (same column dimensions) CH4 Impact of using different phase with higher selectivity

49 Effective Theoretical Plates (N eff ) N eff = 16 t' ( ) W R b 2 Dead Time W b Adjusted Retention Time = Retention Time Dead Time W b = Width between tangents of a peak at baseline intercept t' R

50 Theoretical plates 1 touch = 1 plate This nr is the number of touches of a component with the stationary phase, while it moves through the column.

51 One theoretical plate Two Theoretical plates Three Theoretical plates Carrier gas

52 Theoretical plates 1 touch = 1 plate This nr is the number of touches of a component with the stationary phase, while it moves through the column. The more touches, the more plates, the better the separation Impacted by: Column diameter Column length

53 Height Equivalent to a Theoretical Plate (h) h = L N H depends on Flow and Column ID Lower h = Improved Separation Basic equation used in GC: Van Deemter equation

54 Van Deemter equation H = A + B/u + Cu H = height of a theoretical plate U = average linear gas velocity A, B, C = different contribution factors to peak broadening

55 The A - term Contribution to peak broadening due to different path length (eddy diffusion) For capillary columns A = 0

56 The B - term Contribution to peak broadening due to multidirectional diffusion in the Gas phase Indirect proportional to flow rate

57 The B - term Contribution to peak broadening due to multidirectional diffusion in the Gas phase Indirect proportional to flow rate

58 The B - term Contribution to peak broadening due to multidirectional diffusion in the Gas phase Indirect proportional to flow rate

59 The C - term Resistance to mass transfer in the liquid phase Direct proportional to flow rate

60 The C - term Resistance to mass transfer in the liquid phase Direct proportional to flow rate

61 The C - term Resistance to mass transfer in the liquid phase Direct proportional to flow rate

62 The C - term Resistance to mass transfer in the liquid phase Direct proportional to flow rate

63 Van Deemter: there is an optimal flow

64 Van deemter: Gas Carrier and Linear Velocities Van Deemter Plot 1.0 N 2 HETP (mm) 0.6 He H Average Linear Velocity (cm/sec)

65 Stationary phase Film Thickness Kapacity factor K and Retention

66 Retention (Capacity) Factor : k Practical the most effective separation occurs when the k value for an analyte is minimal 5.

67 Retention (k) can be influenced By film thickness Retention is LINEAR with film thickness By temperature Every 15 C change in oven temp. the k will change about a factor 2

68 Film Thickness and Beta (phase ratio) 0.25 μm 1.0 μm 3.0 μm 0.5 min 2 min k is linear with Film thickness 6 min

69 Phase Ratio (ß) β = mobile phase volume stationary phase volume = column radius 2x film thickness Phase ratio is important if you want to change column internal diameter; For the most easy method conversions, one should try to keep the phase ratio the same

70 Film Thickness Effects : 0.25µm Rtx-1 30m, 0.32mm ID, 0.25µm Rtx-1 70ºC isothermal 1. 1-butanol benzene 3. 2-pentanone 7 4. C 7 8 K C10 = nitropropane 6. pyridine 9 7. C 8 8. C 9 9. C min [min]

71 Film Thickness Effects: 1.0µm Rtx-1 30m, 0.32mm ID, 1.00µm Rtx-1 70ºC isothermal K C10 = butanol 2. benzene 3. 2-pentanone 4. C nitropropane 6. pyridine 7. C 8 8. C 9 9. C min [min]

72 Film Thickness Effects : 3.0µm Rtx-1 30m, 0.32mm ID, 3.0µm Rtx-1 70ºC isothermal 1,2, K C10 = butanol 2. benzene 3. 2-pentanone 4. C nitropropane 6. pyridine 7. C 8 8. C 9 9. C min [min] Peak 9 elutes at 55 min..

73 Stationary Phase: Column Selectivity (α) Interactions with the stationary phase pi pi Van der Waals (london) Hydrogen bonding Depends upon the Chemical composition of the phase

74 Column Selectivity Chemical Composition of Phases Rtx -1 Stationary Phase 100% dimethylpolysiloxane

75 Column Selectivity Chemical Composition of Phases Rtx -5 Stationary Phase 5% diphenyl 95% dimethylpolysiloxane

76 Column Efficiency, Selectivity, and Peak Symmetry Not Efficient, not Selective Not Efficient, but Selective Efficient, but not Selective Efficient and Selective

77 Examples of selectivity..

78 Selectivity via geometry Separation of Para - en Meta Xylenes P M O P + M O P + M O P M O P M O Squalane 100 % methyl Rtx-1 50 % phenyl 50 % methyl Rtx-17 PEG Rtx-Wax Rt-TCEP Non-Polar Polar

79 Shape selectivity Using selected cyclodextrins as modifiers p-xylene m-xylene o-xylene

80 Basic rule in GC stationary phase.. Solubility of sample component in the stationary phase based upon likes dissolve likes. choose a stationary phase that looks like the components you want to separate.. Hydrocarbons 100% PDMS Rtx-1 Aromatic subst. Phenyl subst. PDMS Rtx-5, 17, 35 RT-Dioxins Halogenates Arom. Fluorimated-phenyl Rtx-440, Cl-Pesticides, Rtx-200 Solvents Cyano /phenyl Rtx-1301/624 Alcohols PEG Stablewax Double bonds Cyano propyl Rt 2330, 2460

81 (GC-MS) Presence of Diethylene Glycol and Ethylene Glycol in Toothpaste Column: Stablewax Dissolves : likes like

82 Hydrocarbons on 100% PDMS n-c4 Column n-c5 C3 n-c6 n-c7 : 100 x 0.25 mm Rtx-1 PONA CB, tuned 5%phenyl PDMS Oven : 5 C, 10 min -> 50 C, 5 C/min, 54 min, --> 200 C, 1.3 C/min Carrier gas : He, 24 cm/s, 39.3 Psi; Injection Split, 1 : 150; Detection : FID; n-c9 n-c10 n-c

83 PAHs using an Rxi-17Sil MS (30m x 0.25mm x 0.25μm) ,15,16 benzo(b)fluoranthene benzo(k)fluoranthene benzo(j)fluoranthene / /

84 Detection in GC