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QUESTION 1
Gas chromatography (GC), liquid chromatography (LC), and supercritical fluid chromatography
(SFC) are three types of separation methods with different mobile phases.
Compared with liquids and gases, the diffusion rates of solutes in supercritical
fluids is about ten times greater than that in liquids ( and about three times less
than in gases). Diffusion coefficient of solutes is highest in gas, followed by
supercritical fluid and liquid. Therefore, also able to dissolve large, non volatile
molecules. Gas with the lowest density can only elute volatile solutes with
small molecular weight.
ii) the types of sample that are suitable for each of the above separation method.
(6 marks)
Liquid has the highest density, therefore able to dissolve and elute large and
non volatile solutes as well small and volatile solutes. Compared with gas,
supercritical fluid has higher density (liquid-like), therefore also able to dissolve
large, non volatile molecules. Gas with the lowest density can only elute volatile
solutes with small molecular weight.
b) For each of the above separation methods, discuss one main parameter (exclude flow
rate) that can be manipulated to improve the separation of analytes.
(6 marks)
In GC, temperature: low T for volatile, high T for less volatile analytes.
In LC, the composition of mobile phase affects the elution of analytes. The higher
the eluent strength, the faster the elution.
QUESTION 2
For each of the following, give a reason to the mentioned problem and suggest an approach to
overcome the problem without changing the extraction method used.
Alkanes can be extracted from urban air particulates using supercritical fluid extraction (SFE)
with CO2 at 75 bar (45C), whereas the polyaromatic hydrocarbons (PAHs) remain unextracted.
(3 marks)
CO2 at 75 bar (45C) has low density therefore could not dissolve higher molecular
weight molecules. Therefore, increase the pressure. As the pressure in the system is
increased, density of with CO2 increases and able to dissolve and extract the PAHs.
Or
CO2 is non polar therefore not able to extract PAHs which are slightly more polar than
alkanes. Add a small amount of polar modifier into with CO2 to increase its polarity and
enhance the solvating power. Thus PAHs can be extracted.
QUESTION 3
b) Explain the differences in properties of the mobile phase in GC, HPLC and supercritical
fluid chromatography (SFC).
(6 marks)
Compared with gas, supercritical fluid has high density (liquid-like)(higher than
gas, lower than liquid), therefore able to dissolve large, non volatile molecules and
increase column loading.
Compared with liquid, supercritical fluid has low viscosities, therefore able to use
higher flow rate for faster analysis.
The diffusion rates of solutes in supercritical fluids is about 10x greater than that
in liquids ( and about 3x less than in gases). This results in a decrease in
resistance to mass transfer in the column and allows for fast separations (faster
than HPLC).
d) Discuss the advantages of supercritical fluid extraction (SFE) over solvent extraction.
(4 marks)
Faster extraction: due to greater diffusion rate and lower viscosity.
Solvent strength: can be varied with pressure and temperature.
Recovery of analytes: By releasing pressure of CO2 (environmental friendly).
Less organic solvent (organic modifier).
(Q4 JAN 2012)
QUESTION 4
a) Compare the diffusion coefficient for gas, liquid and supercritical fluid and relate this
property to band broadening in gas chromatography, liquid chromatography and
supercritical fluid chromatography (SFC).
(3 marks)
Diffusion rate: liquid <SF< gas (1)
Therefore band broadening is greater in SFC than LC but less than in GC (2)
A modifier is required when CO2 is used to separate polar compounds (1). Since
CO2 is non polar (1), a polar modifier is added to enhance the solvating power
(making it polar) and enhance the separation (1).
c) Supercritical fluid extraction is faster with easy analyte recovery method compared to
liquid-liquid extraction. Explain.
(4 marks)
Faster due to the higher diffusion coefficient and lower viscocity (1) for fluid
compared to liquid causing greater mass transfer (1).
Many supercritical fluids are gasses at ambient condition (1). Thus, recovery of
analyte is simple compared to organic liquids, as no vaporization step is required
(1).
d) You are given a water sample contaminated with several relatively non polar pesticides.
Outline the steps required for an efficient a solid phase extraction technique. Please
suggest specific sorbent and solvent in each step.
(6 marks)
1. Column (non polar such C18) () must be conditioned (1) using methanol or
and water ();
2. Sample loading (1)
3. Inteference elution (1) using polar solvent such as water ()
4. Elution of analytes (1) using non polar solvent such as hexane ().
QUESTION 5
a) Give two characteristics of a supercritical fluid which can enhanced the separation of
analytes compared to gas.
(2 marks)
Higher viscosity
Higher density
Diffusion rate for supercritical fluid is 3x less than gas (1), thus less band
broadening due to B term compared to GC (1).
Viscosity of supercritical fluid is less than liquid (1), therefore faster analysis
compared to HPLC (1).
CO2 is non polar (1), therefore have ability to extract non-polar compounds only
(1). Need to add a strongly polar modifier to enhance the polarity of CO 2 and
improve the extraction (1).
Use pressure programming (1). Start with low pressure to separate volatile
compounds (1), then increase pressure to increase solvating power (1), therefore,
non volatile compounds can be eluted faster (1).
a) The analysis of eight pesticides mixture (of various polarities and volatilities) was
conducted using supercritical fluid chromatography (SFC). The separation was done
using carbon dioxide as the mobile phase, isobaric condition of 300 bar and isothermal
condition of 100 C.
i) Only 6 peaks were observed. The polar pesticides were not detected. Explain
and suggest an approach to rectify the problem.
(4 marks)
Non polar carbon dioxide (1) is not suitable for polar analytes (1).
Add polar modifier (methanol) to make the mobile phase more polar for the
polar analytes (2).
ii) In the above analysis, it was found that the non volatile pesticides were retained
too long resulting in a very broad peaks. Explain the use of pressure
programming approach in solving this problem.
(4 marks)
Use lower pressure for volatile analytes (1). Increase pressure will increase
the density of mobile phase and correspondingly its solvating power
increases (1), thus able to solvate the non volatile analytes (1). Faster
elution (1).
b) Supercritical fluid extraction (SPE) is considered as a fast and clean extraction technique
compared to liquid-liquid extraction. Explain.
(4 marks)
Use (1) (no/less organic solvent), hence produced clean extracts (1). Fast
extraction due to greater diffusion rate (1) and lower viscosity (1) of supercritical
fluid.
QUESTION 7
a) Compare the density and viscosity of supercritical fluid with liquid and gas. Discuss how
these properties are considered an advantage for supercritical fluid chromatography
(SFC) compared to HPLC and GC.
(6 marks)
Viscosity of SF is lower than liquid but higher than gas (1). Density of SF is lower
than liquid but higher than gas (1).
Compared with gas, supercritical fluid has high density (liquid-like)(higher than
gas, lower than liquid), therefore able to dissolve large non volatile molecules (2).
b) T
he separation of a mixture of polychlorinated biphenyl (PCBs) using SFC with carbon
dioxide as the mobile phase resulted in long analysis time. Discuss an approach to
reduce the analysis time of the separation.
(4 marks)
SFC with CO2 limted to separation of non polar compounds due to non polar
character of CO2. (1)
To reduce the analysis time, the mobile phase must be made more polar (1) by
adding polar modifier (1), e.g. methanol (1).
QUESTION 8
Describe the fundamental difference between ion exchange chromatography (IEC) and size
exclusion chromatography (SEC) in term of its principle of separation.
(4 marks)
Or
IEC relies on charge-charge interactions between the charges in your sample and the
charges immobilized on the resin of your choice (1). Separations based upon exchange
of ions (cations or anions) on an ion exchange stationary phase (resin) column (1).