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MSC: Analytical Methods > Introduction to Gas Chromatography Mass Spectrometry > Flashcards

Introduction to Gas Chromatography Mass Spectrometry Flashcards

1
Q

What are Gas Chromatography and Mass Spectrometer?

A

Gas Chromatograpy

  • Separation technique for volatile organic compounds
  • Can be used as a qualitative or quantitative tool
  • Pharmaceuticals, environmental pollutants, drugs, metabolic compounds

Mass Spectrometer

  • Used as the detector
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2
Q

How is the sample used for Gas Chromatography Mass Spectrometer?

A
  • Usually a mixture of several components
  • Sample usually introduced as a liquid
  • Components of interest (analytes) usually in low concentrations (<1% to ppb levels)
  • Samples dissolved in volatile solvent
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3
Q

What are key components of GC-MS?

A

Hardware to introduce the sample

  • Injector

Technique to separate the sample into components

  • Column
  • Oven
  • Carrier Gas: Nitrogen or Helium (1-2 mL/min)

Hardware to detect the individual components

  • Detector

Data processing

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4
Q

What is the seperation process?

A
  • Sample is introduced into system via hot, vaporising injector. Typically 1uL injected
  • Flow of “Carrier Gas” moves vaporised sample (i.e. gas) onto column
  • Column is coated with wax type material with varying affinity for components of interest
  • Components are separated in the column based on this affinity.
  • Individual analytes are detected as they emerge from the end of the column through the detector
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5
Q

How does the Chromatogram appear?

A
  • Different analytes have different affinity for the column and so elute at different times – rt
  • Response relative to amounts – qual and quant dep on application
  • Detectors number of detectors – but for this MS, also FID (alcohols), TCD (CO
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6
Q

What is a GC step by step?

A
  • Carrier Gas
  • Injector
  • Column (Capillary, Stationary Phase)
  • Oven
  • Detectors - Mass Spectrometer
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7
Q

What is the carrier gas?

A

Inert gases such as Helium or nitrogen

  • Choice dictated by detector, cost, availability
  • Pressure regulated for constant inlet pressure
  • Flow controlled for constant flow rate
  • Chromatographic grade gases (high purity)
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8
Q

What is the process of Injection of the sample?

A
  • A GC syringe penetrates a septum to inject sample into the vaporization chamber
  • Instant vaporization of the sample, 280 °C
  • Carrier gas transports the sample into the head of the column
  • Purge valve controls the fraction of sample that enters the column
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9
Q

What is the purpose of Injection?

A
  • Deposit the sample into the column in the narrowest band possible
  • The shorter the band at the beginning of the chromatographic process (tall narrow peaks) gives maximum resolution and sensitivity
  • Therefore type of injection method and operating conditions is critical in obtaining precise and accurate results
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10
Q

What is a split injection?

A

Mechanism by which a portion of the injected solution is discarded. Only a small portion (1/1000 - 1/20) of sample goes through the column

  • Used for concentrated samples (>0.1%)
  • Can be performed isothermally
  • Fast injection speed
  • Injector and septa contamination not usually noticed
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11
Q

What is a splitless injection?

A

Most of the sample goes through to the column (85-100%)

Used for dilute samples (<0.1%)

  • Injection speed slow
  • Should not be performed isothermally
  • Solvent focusing is important
  • Controlled by solenoid valve. Requires careful optimisation
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12
Q

What is an on column injection?

A

All of the sample is transferred to the column. Needle is inserted directly into column or into insert directly above column

  • Trace analysis
  • Thermally labile compounds e.g Pesticides, Drugs
  • Wide boiling point range
  • High molecular weight
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13
Q

What is a Large Volume Injection nd the steps involved?

A

To enhance sensitivity in Environmental applications.

  • Uses 100µL syringe: Inject up to 70 µL
  • Very slow injection with injector temperature a few degrees below solvent boiling point, split open, flow at about 150 mL/ min
  • Solvent vents out of split vent, thus concentrating the analytes
  • Close split
  • Fast temperature ramp to top column temperature +20°C
  • Column programming as per sample requirements
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14
Q

What are types of Columns?

A

Packed (Preparative)

  • Glass or stainless steel
  • 1-5m length and 5µm ID
  • Larger capacity, low resolution

Capillary (analytical)

  • Thin-fused silica
  • 1—100m length, 250µm ID
  • Smaller capacity, high resolution
  • Higher separation efficiency
  • Easily overloaded by sample
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15
Q

What are characteristics of a capillary column to consider?

A
  • Length (10m - 100m)
  • Internal Diameter (0.1mm - 0.53mm)
  • Liquid Stationary Phase
  • Film Thickness (0.1um - 5um)
  • Polarity (non-polar - polar)
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16
Q

What are advatages of shorter and longer column lengths?

A

Shorter (<15m)

  • Screening samples with few analytes
  • Faster runtimes, higher efficiency separations, good for high MW compounds
  • Resolution decreases as length decreases

Longer (>50m)

  • Separation of complex mixtures with closely eluting peaks
  • Higher resolution as length increases, suitable for low boilers, less active samples or complex temperature ramps
  • Slow run times
17
Q

What are advatages of shorter and longer internal diameter?

A

Smaller ID

  • Good resolution of early eluting compounds
  • Lower sample capacity
  • Limited dynamic range
  • Easily overloaded

Larger ID

  • Have less resolution of early eluting compounds
  • Increased sample capacity
  • Sufficient resolution for complex mixtures
  • Greater dynamic range
18
Q

What are characterisitics of Stationary Phases?

A
  • Choice of phase determines selectivity
  • Hundreds of phases available. Many phases give same separation
  • Same phase may have multiple brand names. Stationary phase selection for capillary columns much simpler
  • Like dissolves like: Use polar phases for polar components and Use non-polar phases for non-polar components
19
Q

What is the Film thickness?

A
  • Amount of stationary phase coating
  • Affects retention and capacity
  • lStandard capillary columns typically 0.25µm
20
Q

What are the differences found between using thicker films and thinner films?

A
  • Thicker films increase retention and capacity but typically have slower runtimes, higher bleed and lower temperature limits. Used for low boilers, gases, solvents and volatiles.
  • Thin films are useful for high boilers, have high efficiency, lower bleed, faster run times and higher temperature limits, however they have limited retention.
21
Q

What is column capacity?

A

The maximum amount that can be injected without significant peak distortion

22
Q

How does column capacity increase?

A

Column capacity increases with :

  • Film thickness
  • Temperature
  • Internal diameter
  • Stationary phase selectivity
23
Q

What happens is the column capacity is exceeded?

A

If exceeded, results in:-

  • Peak broadening
  • Asymmetry
  • Tailing
24
Q

What is a common issue is encoutered in GC-MS?

A

Column Bleed

  • One of the most common issues encountered in GC-MS
  • Stationary phase degrades over time; components pass through the column contributing to the signal, usually minimal and of no consequence
  • Excessive bleed is often caused by something damaging the stationary phase of the column, oxygen from a leak or some component in the sample
  • Can result in a higher baseline signal, peak tailing, poor reproducibility, presence of unusual fragments at high levels and ghost peaks
  • Check for leaks, use inline oxygen filters on carrier gas, review sample preparation and loading requirements
25
Q

What are advantages to temperature programming of the column oven?

A

Start at low temperature and gradually ramp to higher temperature

  • More constant peak width
  • Better sensitivity for components retained longer
  • Improved chromatographic resolution
  • Peak refocusing at head of column
26
Q

How does high and low oven temperature result in?

A
  • High - co-elution, poor resolution but faster separation
  • Low - longer elution times with improved resolution

Compromise temperature to optimise program

27
Q

What are the types of detector and their use?

A
  • Thermal conductivity: Universal, Carbon monoxide
  • Flame ionisation: Hydrocarbons, Ethanol, methanol, acetone, isopropanol
  • Electron capture: Chlorine containing compounds
  • Thermionic: Nitrogen- and phosphorus-containing compounds
  • Mass spectrometer: Drugs, steroid hormones, vitamins, metabolic products
28
Q

What is Mass Spectrometry?

A
  • Mass spectrometry is a micro-analytical technique used to obtain information regarding structure and molecular weight of an analyte
  • In all cases some form of energy is transferred to analyte to cause ionisation
  • In principle each mass spectrum is unique and can be used as a “fingerprint” to characterise the sample
  • Destructive method - sample consumed during analysis
29
Q

What is GC-MS?

A

GC-MS is a hyphenated technique that combines the separation ability of the GC with the detection qualities of mass spectrometry

30
Q

What is the process of GC-MS ?

A
  • Sample injected onto column via injector
  • GC then separates sample molecules
  • Effluent from GC passes through transfer line into the Ion Trap/Ion source
  • Molecules then undergo electron /chemical ionisation
  • Ions are then analysed according to their mass to charge ratio
  • Ions are detected by an electron multiplier which produces a signal proportional to ions detected
31
Q

What are components of a Mass Spectrometer?

A
  • Sample introduction
  • Source (ion formation)
  • Mass analyzer (ion separation) - high vacuum
  • Detector (electron multiplier tube)
32
Q

What are the fdifferent sample introduction/sources?

A

Volatiles

  • Electron impact (EI) – hard ionisation
  • Chemical ionization (CI) – soft ionisation

Non-volatiles

  • Direct infusion/electrospray (ESI)
  • Matrix Assisted Laser Adsorption (MALDI)

Elemental mass spectrometry

  • Inductively coupled plasma (ICP)
  • Secondary Ion Mass Spectrometry (SIMS)
33
Q

How does Electron Impact take place?

A
  • Gas-phase molecules enter source through heated probe or GC column
  • 70 eV electrons bombard molecules forming M+* ions that fragment in unique reproducible way to form a collection of fragment ions
  • EI spectra can be matched to library standards
34
Q

How does chemical ionisation take place?

A

CI (soft ionization)

  • Higher pressure of methane leaked into the source (mtorr)
  • Reagent ions transfer proton to analyte
35
Q

What does an electron multipier do?

A
  • Passes the ion current signal to system electronics
  • Signal is amplified
  • Result is digitised
  • Results can be further processed and displayed
36
Q

What is the advanatge of GC-MS?

A
  • Best technique for volatiles, non-polar analytes
  • Highly robust reproducible mass spectra (EI) - Universal fragmentation conditions
  • Libraries. Transferable among instruments (universal) and commercially available mass spectral libraries
  • Automated identification by mass spectra through deconvolution
37
Q

What is deconvolution?

A

Deconvolution is the process of computationally separating co-eluting components and creating a pure spectrum for each component.

38
Q

What are limitations of GC-MS?

A
  • Analyte must be volatile or made volatile by chemical derivatisation
  • Analyte or its derivative should be thermally stable
  • Sample preparation is generally longer
  • Larger sample volume
  • Lack of direct sample analysis
  • Molecular ion is often lost when electron impact (EI) ionization is used

MSC: Analytical Methods (13 decks)

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