Ch' 23 Analytical Separations

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Ch. 23 Analytical Separations
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Separation of Silicones
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Why a Separation?

• Most real-world samples are complex mixtures
• To identify and quantify components of a mixture,
  they compounds must be separated
• There are a number of different separation
  methods
• Extraction
• Chromatography
• Electrophoresis

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Extraction

• Extraction is the transfer of a solute from one
  phase to a second
• Extractions are performed to concentrate analytes
  or isolate them for interferences

We can define an equilibrium constant, K, for
distribution of the chemical species between
phases If we extract from one solution to the
other the fraction remaining in solution 1,
q A q of 1/2, means 1/2 remains in solution 1
after extraction
of extractions
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Extraction Efficiency Example

• A solute has a partition coefficient of 7 between
  toluene and water. If you have 100 mL of a 0.010
  M solution of the solute in water.
• What fraction of the solute remains in H2O after
  a 500 mL extraction with toluene?
• What fraction of the solute remains in H2O after
  a 5-100 mL extractions with toluene?

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pH Effects on Extraction

• If you have an acid or base analyte, the charge
  changes dependent on pH
• Generally, a neutral species is more soluble in
  organic solvents, while a charged species is more
  soluble in water
• To extract an acid or base you must do so at an
  appropriate pH
• For bases, pH low enough to convert B to BH
• For acids, pH high enough to convert HA to A-

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Crown Ethers for Extraction of Metals
dibenzo-30-crown-10
Because charged species are difficult to dissolve
in organic solvents, charged complexes (like
M-EDTA) are difficult to use with organic
solvents. Crown ethers envelop a metal ion and
can bring them even into non-polar solvents.
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Chromatography

• Chromatography operates on the same principle as
  extraction
• One phase is held in place while one moves past
  it
• It can be thought of as repeated extractions or
  as a continuous extractions

Separation occurs based on the various
partitioning coefficients of different solutes
between the two phases. The mobile phase is
the solvent that moves through the column, while
the stationary phase remains in place.
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Chromatographic Interactions

• Chromatography techniques are divided into
  categories based on how the solute interacts with
  the stationary phase

Adsorption chromatography relies on a solute
adsorbing onto the surface of the stationary
phase. The stronger a solute adsorbs, the longer
it takes to travel through the chromatography
column
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Chromatographic Interactions (2)
Ion-exchange chromatography uses ionic
interactions to separate ions. A stationary
phase of anions will separate cations and vice
versa.
Size exclusion chromatography uses size to
separate molecules. Big molecules pass quickly
because they do not get caught up in pores, while
small ones do.
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Chromatographic Interactions (3)
Partition chromatography relies on partitioning
of solutes between a mobile phase and bonded
liquid stationary phase. This is virtually
identical to the interaction in a typical
extraction.
Affinity chromatography uses interactions
specific to a compound or class of
compounds. For instance, it could use
antibodies to select out one protein from a
mixture of hundreds
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The Chromatogram
Adjusted Retention time of solute
Retention time of mobile phase (CH4 for GC)
Retention time of solute
A chromatogram is a plot of detector response
with time. The retention time for a component is
the time it takes from injection for the compound
to reach the detector.
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Retention Paramaters

• Adjusted retention time-additional time for
  solute to travel through column
• Relative retention-ratio of adjusted retention
  times for any two components
• Capacity factor-partition ratio

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Retention Example

• What is the adjusted retention time and capacity
  factor for benzene and toluene in a GC? Methane
  was retained at 42 s benzene at 251 s and
  toluene at 333 s.

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Partition Coefficient and Retention

• The capacity factor is equivalent to the time the
  solute spends in the stationary phase over the
  mobile phase and can be related to the partition
  coefficient
• Retention time, capacity factor, and partition
  coefficient are all proportional to each other,
  so this means that relative retention can be
  related to any of the three terms

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Determining Volume of Column

• Any calculation of retention parameters requires
  knowing the volume of the stationary and mobile
  phases
• More frequently we calculate the relative volume
  of the two

If the outer diameter (OD) of the column is 250
µm, and the stationary phase thickness is 1 µm
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Industrial Size Chromatographs
Most analytical chromatographs deal with µL
volumes. For synthetic work, purifying new
compounds, industrial chromatographs can separate
kg of material.
300 L column