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Supercritical Fluid Chromatography (SFC)
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Principle-Supercritical Fluid Chromatography (SFC) operates on the principle
of using a supercritical fluid (SCF) as the mobile phase to separate
compounds in a mixture
...
This unique state allows supercritical
fluids to diffuse like gases but dissolve materials like liquids
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The separation process in SFC occurs
because different compounds in the sample interact with the supercritical fluid
and stationary phase to different extents, depending on their chemical
properties such as polarity, size, and volatility
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

Components
1
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CO₂ is chosen due to its ability to be easily pressurized and its
low toxicity
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The density and viscosity of
CO₂ can be altered by changing the pressure and temperature, making it
a highly versatile solvent for a wide range of compounds
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Syringe Pump-The syringe pump is responsible for introducing the
supercritical fluid into the chromatographic system
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Accurate control over the flow rate is
essential for reproducible results and efficient separation of components
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3
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The column plays a critical role
in determining the separation efficiency by providing a surface for
interactions between the analytes and the stationary phase
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4
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Common detectors used in
SFC include:

 UV-Visible (UV/VIS) Detector: This is often used when the
compounds of interest absorb UV or visible light
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 Fluorescence Detector: This is employed when compounds
naturally fluoresce or can be derivatized to fluoresce
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 Refractive Index (RI) Detector: This is useful for compounds that
do not absorb UV light or fluorescence, such as sugars or
certain lipids
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Pressure Regulator-The pressure regulator ensures that the supercritical
fluid remains at a constant, appropriate pressure throughout the
chromatographic process
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6
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Temperature influences the
solubility of analytes in the supercritical fluid and affects the retention time
of the analytes
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Procedure –
 A small volume of the sample is injected into the system using an
injection port
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 The supercritical fluid is pumped through the chromatographic column
containing the stationary phase
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Compounds that
interact more strongly with the stationary phase will be retained longer,
while those that interact weakly will elute faster
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The resulting signals are recorded as peaks in a
chromatogram, with the area under each peak correlating to the
quantity of each component
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The retention time (the time it takes
for a compound to elute) and the peak area provide critical information
about the identity and concentration of each compound
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Pharmaceutical Analysis-SFC is used for the purification and analysis of
active pharmaceutical ingredients (APIs), drug formulations, and impurities
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2
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It allows for high-throughput analysis and purification of these substances,
which is critical in drug development and biomarker discovery
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Food and Beverage Industry-SFC is used to extract and analyze compounds
such as essential oils, flavors, and fragrances
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4
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SFC is used to separate
complex environmental contaminants, including pesticides, herbicides, and
industrial chemicals, with high sensitivity
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Forensic Science-In forensic toxicology, SFC is used to identify and quantify
illicit drugs, poisons, and metabolites from biological samples such as blood,
urine, and tissues
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6
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It is particularly beneficial for separating non-polar compounds like
essential oils, which are often challenging to isolate using traditional solventbased extraction methods

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