Chap. 1 (Introduction), Chap. 2 (Components and Circuits)
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Transcript Chap. 1 (Introduction), Chap. 2 (Components and Circuits)
Supercritical Fluid Chromatography
• Theory
• Instrumentation
• Properties of supercritical fluid
Critical temperature
Above temperature liquid cannot exist
Vapor pressure at critical temperature is
critical pressure
T and P above critical T and P
Critical point
Supercritical fluid
19-1
Supercritical fluid
• Above the critical temperature
no phase transition regardless of
the applied pressure
• supercritical fluid is has physical and
thermal properties that are between
those of the pure liquid and gas
fluid density is a strong function
of the temperature and pressure
diffusivity much higher a liquid
readily penetrates porous
and fibrous solids
Low viscosity
Recovery of analytes
Return T and P
19-2
Typical Supercritical Solvents
Compound
Tcº C
Pc atm
d*
CO2
C2H4
N2O
NH3
31.3
9.9
36.5
132.5
72.9
50.5
72.5
112.5
0.96
--0.94
0.40
n-C5
n-C4
CCl2F2
196.6
152.0
111.8
33.3
37.5
40.7
0.51
0.50
1.12
CHF3
H2O
25.9
374.1
46.9
218.3
------19-3
Supercritical fluid chromatography
• Combination of gas and
liquid
• Permits separation of
compounds that are not
applicable to other
methods
Nonvolatile
Lack functional
groups for detection
in liquid
chromatography
19-4
Supercritical Fluid Extraction
•
•
•
•
near the critical point properties
change rapidly with only slight
variations of pressure.
inexpensive,
extract the analytes faster
environmentally friendly
sample is placed in thimble
supercritical fluid is pumped through
the thimble
extraction of the soluble
compounds is allowed to take
place as the supercritical fluid
passes into a collection trap
through a restricting nozzle
fluid is vented in the
collection trap
solvent to escapes or is
recompressed
material left behind in the collection
trap is the product of the extraction
batch process
19-5
Capillary Electrophoresis
• Separations based on different rate of ion
migration
Capillary electrochromatography separates
both ions and neutral species
Electroosmotic flow of buffer acts as pump
• Principles
• Applications
19-6
Planar electrophoresis
• porous layer
• 2-10 cm long
paper
cellulose acetate
polymer gel
soaked in
electrolyte
buffer
• slow
• difficult to automate
19-7
Capillary Electrophoresis
• narrow (25-75 mm diameter)
silica capillary tube
40-100 cm long
• filled with electrolyte buffer
• fast
• complex but easy to automate
• quantitative
• small quantities
nL
19-8
Separation
• Movement of ions function of different parameters
molecular weight
charge
small/highly-charged species migrate rapidly
pH
Deprotonation HAH+ + A
ionic strength
low m
few counter-ions
low charge shielding
high m,
many counter-ions
high charge shielding
19-9
Migration rate
• v= migration velocity
me=electrophoretic mobility (cm2/Vs)
• E=field strength (V/cm)
• For capillary
V=voltage
L=length
• Electrophoretic mobility depends on net charge and
frictional forces
Size/molecular weight of analyte
Only ions separated
• Plate height (H) and count (N)
19-10
Function of diffusion and V
Plates
• Planar electrophoresis
large cross-sectional area
short length
low electrical resistance, high currents
Sample heating Vmax=500 V
N=100-1000 low resolution
• Capillary electrophoresis
small cross-sectional area
long length
• high resistance
• low currents
Vmax=20-100 kV
• N=100,000-10,000,000 high resolution
As comparison, HPLC N=1,000-20,000
19-11
Zone Broadening
• Single phase (mobile phase) - no partitioning
• three zone broadening phenomena
longitudinal diffusion
transport to/from stationary phase
multipath
• planar
no stationary phase
• capillary
no stationary phase or multipath
19-12
Transport
•
•
•
•
ions migrating in electric field
cations to cathode (-ve)
anions to anode (+ve)
Electroosmosis movement in one
direction
anode (+ve) to cathode (-ve)
Components
Analyte dissolved in
background electrolyte and
pH buffer
Silica capillary wall coated
with silanol (Si-OH) and SiO
Wall attracts cations double-layer forms
Cations move towards
cathode and sweep fluid in
one direction
Electroosmotic flow proportional
to V
usually greater than
electrophoretic flow
19-13
Bulk flow properties
hydrodynamic
ion buffer
19-14
Techniques
• Electropherogram
migration time
analogous to
retention time in
chromatography
• Isoelectric focusing
Gradient
No net migration
pH gradient with
weak acid
19-15
Techniques
19-16