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Analytical Chemistry: Part I Chemical Engineering Section 2 (ex-ET)
Analytical Chemistry: Part I Chemical Engineering Section 2 (ex-ET)
ANALYTICAL CHEMISTRY
8 lectures, Michaelmas Term 2013
Course outline
1. What is Analytical Chemistry?
2. General features of molecular spectroscopy
3. Ultraviolet/visible spectroscopy
4. Infrared spectroscopy
5. Microwave spectroscopy
6. Nuclear magnetic resonance spectroscopy
7. Methods of elemental analysis
8. Mass spectrometry
9. Chromatography
Text books
These lecture notes contain all you need to know about analytical chemistry for examination
purposes. You can find out more (if you want to) from almost any textbook with “Physical
Chemistry” or “Analytical Chemistry” in the title.
Examples paper: One examples paper will be issued to test understanding and aid exam
preparation.
Analytical Chemistry is thus vital in the process industries and in research laboratories.
• Qualitative analysis is the identification of elements, functional groups, or
particular compounds in a sample.
• Quantitative analysis is the determination of the amount of a particular
element, species or compound in a sample.
ΔE
hν
hν hν hν
absorption spontaneous stimulated
emission emission
sample monochromator
cell e.g. prism
lens
detector
• Transition Probability
o This depends on the precise quantum mechanical wavefunctions of the initial and
final states (beyond the level of this course).
❧ Some transitions may have zero probability – in that case, they are said to
violate “selection rules”.
• The Population of States
o The initial population of an energy level obviously affects spectral intensities.
o At thermal equilibrium, the relative populations of two energy levels may be
obtained from the Boltzmann factor:
upper
N upper ⎛ − ΔE ⎞
Δ = exp⎜ ⎟
N lower ⎝ kT ⎠
E
L
Io I
Incident Transmitted
light light
• A variety of different units are commonly used in spectroscopy to represent the energy
difference between the levels. You need to be familiar with:
o J (Joules), the SI unit of energy
o ν (frequency, often expressed in MHz); related by ΔE = h ν
o λ (wavelength, often expressed in nm), related by ΔE = hc / λ.
o 1/λ or ν~ (reciprocal wavelength, or wavenumbers, often expressed in cm–
1
); related by ΔE = h c (1/λ).
o eV (electron volts), related by 1 eV = 1.602 x 10–19 J (i.e. the charge of an
electron)
-2
10
Microwaves Molecular rotation
-3
10
-4
10
-6
10
Visible
-7 Electronic excitation
10
Ultraviolet
-8
Increasing 10
energy
-9
10
-11
10
-12
10 γ-ray nuclear excitation
(Mössbauer spectroscopy)
-13
10
cosmic rays
LUMO
ΔE for transition of interest
HOMO
• Aside: due to simultaneous vibrational and rotational transitions (see Sections 3 and 4),
UV spectra normally consist of fairly broad peaks.
• The absorbing groups in a molecule are called chromophores. Two isolated
chromophores in a molecule give roughly independent absorptions for each one:
o e.g. CH3CH2–CNS: λmax= 245 nm and ε = 800
SNC–CH2CH2CH2–CNS: λmax= 247 nm and ε = 2000
• In organic chemistry, π-conjugated systems (when multiple bonds are separated
by a single bond) tend to give particularly informative spectra.
o Overlap of adjacent π orbitals results in a decrease in the energy gap between the
occupied π orbital and the unoccupied π* antibonding orbital.
o This results in an increase in absorption wavelength (even into the visible region
for greatly conjugated systems e.g. organic dyes), and normally an
increase in the intensity as well.
❧ General rule: increased conjugation increases λmax and ε.
o Aromatic systems exhibit conjugation, but tend to give complex spectra,
frequently with more than one absorption band.
• Conjugation with lone pairs (n-π conjugation) can also result in spectral transitions,
though these are much weaker than those originating from overlap of π orbitals.
227 22,000
275 30,000
310 77,000
O
225 13,000
184 60,0000
203 7,000
254 200
O
OH
203 shif ted 12,000
to 230
O
OH
230 shif ted 21,000
to 273
• Absorption in the infrared region is associated with transitions between vibrational energy
levels.
• Let us consider a diatomic molecule first (such as H–Cl) as it contains just a single bond.
• Imagine the bond behaves like a perfect spring with a force constant k
m1 m2
spring constant = k
v=3 E = 7/2 h ν0
v=2 E = 5/2 h ν0
v=1 E = 3/2 h ν0
ΔE
v=0 =0 E = 1/2 h ν0
• At room temperature, ΔE >> kT, implying that only the v = 0 quantum level is
significantly populated.
• Real bonds do not behave as ideal springs; they behave as anharmonic oscillators.
• Potential energy diagram:
0
0 1 2 3 4 5 6
Potential Energy
-100
• The simple harmonic oscillator SHO model discussed earlier provides a good
approximation to the real case at the lowest energy level when r is always close to
requilibrium.
v=4
v=3 E = ~ 7/2 h ν0
v=2 E = ~ 5/2 h ν0
v=1 E = ~ 3/2 h ν0
ΔE
v=0 =0 E = 1/2 h ν0
• Note that the ground state of the molecule (i.e. the state of lowest energy) doesn’t have
E = 0. Two consequences of this are:
o Even at a temperature of absolute zero, bonds will still have a non-zero vibrational
energy that depends on k and μ:
❧ The molecule is said to possess zero-point energy.
❧ Atoms still move by vibrations at absolute zero; whilst seen here from the
maths, it’s a consequence of the Heisenberg uncertainty principle.
o Consider bonds involving different isotopes, e.g. compare
O–H and O–D bonds:
❧ They involve the same number of electrons, and so have identical force
constants and bond lengths.
❧ However, they have different zero-point energies because of different μ.
❧ They will have different bond dissociation energies, as this is
the energy required to take the bond from its zero-point energy up to an
energy corresponding to the atoms being widely separated.
H H H
O O O
H H H
–1
cm 3500 2500 2000 1500 400
(from http://www.cem.msu.edu/~reusch/VirtualText/Spectrpy/InfraRed/infrared.htm#ir1)
•
• The sample for IR spectroscopy may be solid/liquid/gas.
• For lab measurements, the sample is held in a special cell:
o the windows shouldn’t absorb above ~500 cm–1 (e.g. can’t use glass; KBr is quite
common)
o the path length is usually short (because absorbances tend to be strong).
• Beer-Lambert law can be used to estimate concentration; this is difficult in practice to do
accurately due to scattered radiation affecting the baseline.
• Limitation: need to identify a band from the component of interest that isn’t overlapping
with bands from any other components that may be present; usually okay for simple
mixtures.
• Rough cost estimate of basic spectrometer: £12k
• Timescale: Typically 10 s for a modern instrument, but it depends on sensitivity and
resolution required.
• On-line measurement of process fluids is not straightforward:
o Most of the materials used for cell windows aren’t resistant to chemicals (e.g. KBr
dissolves in water).
o Normal fibre optic cables absorb in the infrared region so we can’t use them.
o Special materials for fibre optic cables that don’t absorb above, say, 800 cm–1 are
being developed; thus far they tend to be expensive and react with acid/alkali.
Example: NIR spectra of C-H stretching overtone region for water-ethanol mixtures.
(www.axsun.com)
• New techniques are being devised for on-line process analysis – one called Encoded
Photometric Infrared (EPIR) spectroscopy has great potential, but it’s too early to say
how useful it will be.
• Another advantage: light at the laser frequency can be focussed on to a small area of
sample easily – Raman microscopy can record a vibrational spectrum on just a
small part of the sample (down to ~1 μm).
• Main limitations:
o if the laser also promotes electrons into a higher energy level, then light will be
emitted as the electron relaxes back down – hence there may be
interference from sample fluorescence
o interference from background radiation
o quantifying signal – the Beer-Lambert law doesn’t apply
o it’s not good for complex mixtures
• Rough cost estimate of basic spectrometer: £15k ?
• Raman spectroscopy is beginning to be used for on-line process analysis:
o the laser can be in the near-infrared region meaning that it can travel through glass
windows, or be transported by fibre optic cables; the latter makes possible remote
on-line sensing of the process fluid.
• This technique is not really useful in analytical chemistry, but is covered briefly here for
completeness and because it can impact vibrational spectra.
• Absorption in the microwave region is associated with transitions between rotational
energy levels.
o This is how microwave ovens work.
• We shall only consider linear molecules in this section (e.g. diatomic molecules, or CO2).
• In the same way that we write down and solve the Schrödinger equation for vibrations we
can also do so for pure rotational motion.
• For rigid linear molecules this gives energy levels characterised by the quantum numbers
J and MJ
o E = B J (J+1) J = 0, 1, 2, 3, …
o MJ = J, J–1, J–2, …, –J [i.e. there are 2J+1 values of MJ]
h2
o B= where I is the moment of inertia
8π 2 I
❧ Note that I = μ r2 for a diatomic molecule, where μ is the reduced mass
J=2 E = 6B 5 levels
J=1 E = 2B 3 levels
J=0 E=0 1 level
• Selection rules:
o ΔJ = ±1 [to conserve angular momentum]
o Molecule needs a permanent dipole moment (to interact with EMR)
• Population of levels depends on the degeneracy (number of levels having the same
energy) and the Boltzmann factor:
• Vibrations and rotations can be treated as being independent of each other as they occur
on different timescales.
• For a diatomic molecule, the selection rule is:
Δv = 0, ±1 (for SHO) and ΔJ = ±1 and molecule needs a permanent dipole moment
• Thus pure rotational spectra may be observed, but pure vibrational spectra
are forbidden despite our discussion in Section 4!
o Each transition from vibrational level v = 0 to v = 1 has to be accompanied by
a rotational change ΔJ = ±1 in order to conserve angular momentum.
J=5
v=1 J=4
vibrational
level J=3
Energy
J=2
J=1
J=0
J=5
v=0 J=4
vibrational
level J=3
J=2
J=1
J=0
• For liquid samples, intermolecular collisions usually mean that the linewidths are
too broad to see rotational fine structure in infrared spectra.
r
Energy
• This is usually the “best” method for the structural identification of organic
molecules.
• The sample normally needs to be a liquid.
o Where possible, dissolve solids to make a solution before doing NMR
spectroscopy.
o Far more limited information can be obtained on solids (or gases).
• In the same way that an electron has angular momentum (described by “spin”), nuclei
may also possess angular momentum and so be described as having spin.
o The nuclear spin quantum number, I, gives the total angular momentum.
• The value of I for a particular nucleus depends on the detailed arrangement of
protons and neutrons within the nucleus.
o 1H has I = 1/2 2
H (0.015% natural abundance) has I = 1
12 13
o C has I = 0 C (1.1% natural abundance) has I = 1/2
o 16O has I = 0 17
O (0.04% natural abundance) has I = 5/2
• The z-component of nuclear angular momentum is given by the quantum number mI,
which can take values I, I–1, ..., –I.
• Ordinarily the mI quantum levels all have the same energy (i.e. are degenerate).
• However they will split into 2I+1 different energies if a large magnetic field B0 is applied.
• Transitions between these energy levels is termed nuclear magnetic resonance (NMR)
spectroscopy. The selection rule is ΔmI=±1.
ΔE = hν
4 degenerate levels
when B0 = 0 mI = 1/2
ΔE = hν
mI = 3/2
Increasing B0
• We shall concentrate only on the case of nuclei having I = ½, the most important of which
are 1H and 13C.
o Nuclei with I = 0 will not show NMR spectra.
o Nuclei with I > ½ tend to give broad uninformative spectral lines.
Example:
an I = 1/2 nucleus
Energy
mI = – 1/2
ΔE = (h/2π) γ B0 (1–σ)
No magnetic
field mI = 1/2
Sample in strong
magnetic field B0
• The separation of the two energy levels for a spin I = ½ nucleus is given by:
ΔE = (h/2π) γ B0 (1–σ)
o h = Planck’s constant
o γ = magnetogyric ratio of the nucleus (ratio of magnetic moment to angular
momentum):
❧ For 1H: γ = 26.752 x 107 rad T–1 s–1
❧ For 13C: γ = 6.7283 x 107 rad T–1 s–1
e–
B0
δB
• There are also some other effects that cause chemical environment to give a σ
shielding term that we don’t have time to discuss (e.g. hydrogens attached to
aromatic rings have less shielding than might be expected).
• Hence nuclei in different chemical environments in the molecule will have peaks at
different frequencies in the NMR spectrum.
o Those with fewer electrons around them will have less σ shielding.
• The peaks in NMR spectra are usually quoted using the “chemical shift”
scale in parts per million (ppm).
o This is effectively a dimensionless frequency scale relative to a standard reference
substance:
νsample − νreference
δ / ppm = × 106
νreference
o The reference sample for both 1H and 13C NMR spectra is a compound known as
TMS, tetramethylsilane, Si(CH3)4,
o The chemical shift scale is used because it is independent of B0 value –
this is helpful because B0 is rarely known exactly as it changes slightly day by day
for a superconducting magnet.
chemical shift, δ
frequency, ν
shielding, σ
6 5 4 3 2 1 0
1H chem ical shift (ppm )
HA HB
o Consider nucleus HA:
❧ Spin HB may be in the same or the opposite direction to it.
❧ Hence HA sites in a sample have two slightly different energy states.
o NMR signal from HA will therefore be a 1:1 doublet due to coupling to HB.
o Similarly, the NMR signal from HB will be a 1:1 doublet due to coupling to HA.
J /Hz J /Hz
δ /ppm HA HB
HA HB
o Nucleus HA can now couple to two chemically equivalent HB nuclei:
❧ Both HB spins may be in same direction as HA (probability ¼).
❧ Both HB spins may be in opposite direction as HA (probability ¼).
❧ One HB may be in same direction as HA, and one opposite (probability ½).
o NMR signal from HA will therefore be a 1:2:1 triplet due to coupling to HB.
o Note that one HB nucleus does not couple to the other HB nucleus because they are
in chemically equivalent environments.
o The NMR signal from HB will therefore be a 1:1 doublet due to coupling to HA
J /Hz
J J
δ /ppm HA HB
• Example 3:
X HB
X HB
HA HB
o Nucleus HA now can couple to three chemically equivalent HB nuclei.
o The resulting pattern for HA will be a 1:3:3:1 quartet (reflecting the probabilities
of the different possible spin states), while the signal from HB will be a 1:1 doublet
due to coupling to HA.
J /Hz
J J J
δ /ppm HA HB
7 Hz
6 5 4 3 2 1 0
1H chem ical shift
100 Hz
7 Hz
6 5 4 3 2 1 0
1H chem ical shift 500 Hz
• Timescale for 1H NMR experiment is ~1 minute, but setting up may take more time
depending on the experiment being performed.
• Each chemically distinct environment gives rise to a chemical shift (fixed in ppm), which
may then split up due to J-coupling (fixed in Hz).
• Peak areas give relative amounts of each H environment.
• The first use of 13C NMR spectroscopy is to give the number of chemically inequivalent
carbon atoms in the sample.
• Note that symmetry considerations may mean that this is less than the
number of carbon atoms in the molecule.
CH3 CH3 CH3
CH3
CH3
CH3
CH CH2 CH 3
Example 13C NMR spectrum (proton decoupled): riboflavin (vitamin B2, C17H20N4O6)
(from http://riodb01.ibase.aist.go.jp/sdbs/)
• The NMR techniques described above are those used in research and specialist analytical
laboratories.
• They can’t be implemented on a chemical plant: they’re too expensive and the hardware
is not robust enough.
• There’s also a problem measuring NMR spectra of flowing samples – the nuclei under
investigation need to be in the magnet for a certain length of time before they can be
measured by NMR.
o Slow flow only, or need to “stop” the flow for the measurement.
• On a chemical plant, we can use permanent magnets of far lower magnetic field strength
(corresponding to a 1H frequency of, say, 20 MHz)
o Can’t do high-resolution spectroscopy but can sometimes separate low-resolution
signals: for instance, we can get a measurement of the ratio of aromatic to
aliphatic 1H environments in gasoline.
o 1H NMR signal intensity can be used to estimate the water content in solids.
o 1H NMR relaxation times (how long spins take to move from the upper energy
level to the lower energy level) of liquids in porous solids gives some information
on the pore sizes present; this is used in oil-well logging.
• Magnetic resonance imaging (MRI) is also being developed for use in a process plant
setting.
Atomic orbitals
Light at ν
(chosen for specific
F Measure absorption
element of interest)
L
A hν
M
E
Solution of sample
• Advantages of AAS:
o Sensitivity to a wide range of elements (typically down to 1 ppm)
o High accuracy if care is taken over sample preparation and calibration.
• Disadvantages of AAS:
o Some solid samples are difficult to get into solution form.
o Need a hollow cathode lamp for sharp monochromatic lines for each element.
o Different atoms require different flame temperatures to achieve reliable results
(e.g. air/acetylene 2250°C; NO/acetylene 2955°C).
• This is a method of elemental analysis for solid samples. It’s useful for those that don’t
easily dissolve, e.g. some minerals and ores.
• X-rays are fired at the sample, and these knock out a core electron from an inner quantum
level (e.g. from the 1s, 2s or 2p atomic orbital).
• An electron then falls from an upper energy level to fill the core-level vacancy; this is
accompanied by the emission of a photon of frequency hν.
• The wavelengths of emitted photons enable the elements present in the sample to be
determined, while the intensities give the amount of each element present (after careful
calibration).
• This method is only appropriate for elements with atomic numbers greater
than ~20. Gram quantities of a solid sample are required.
hν
4s4p
3d
3p
3s
2p
2s
1s
Hole created
by X-rays
• Electron Ionisation (EI) is the most common method of ionisation for small organic
molecules:
o The sample must first be vaporized (by heat or a spark) if it isn’t already in gas
phase.
❧ Some sample decomposition may occur for thermally unstable samples
during this step.
o The sample is then bombarded with electrons that knock an electron out:
M + e– M+ + 2e–
o The parent ion is always produced in a vibrationally excited state and so might
fragment (often in a fairly predictable manner) to smaller ions before it reaches
the detector.
o EI is a harsh ionisation method: fragmentation may be so extensive that the parent
ion is absent from the spectrum.
o The fragmentation pattern provides a fingerprint method of sample
identification by comparison of results with MS databases.
o Example: EI MS of vinyl chloride
• For very large macromolecules such as proteins, the above methods don’t work well:
o multiple fragmentation makes interpretation difficult/impossible.
o it’s difficult to separate species with high m/z ratios (e.g. above 5000 Da).
• Once the ions have been formed, we need to separate them according to m/z ratio.
• This part of the spectrometer will need to be at a good vacuum because we don’t want the
ions to hit other molecules before reaching the detector.
• The analysis method used will depend on mass resolution required, mass range, scan rate,
and detection limit required.
[from http://www.cem.msu.edu/~reusch/VirtualText/Spectrpy/MassSpec/masspec1.htm#ms1]
8.3 Uses of MS
• All the techniques discussed so far are limited in one important respect:
o They can only easily analyse pure compounds, or simple mixtures at best.
• What do we do if the sample is a complicated mixture?
• Chromatography is the key separation technique used by analytical chemists when
the sample is a mixture.
• After calibration, chromatography can be used for structural identification and
quantitative measurement as well as simply being a separation technique.
• Chromatography is also a chemical engineering unit operation for purification of high-
value chemicals, particularly in the pharmaceutical and biotechnology industries.
• Chromatography is based on the physical separation of individual chemical components
in a sample:
o The sample is present in a mobile or carrier phase: may be gas, liquid, or even
supercritical fluid.
o The sample is separated into components due to differences in affinity for a
stationary phase.
• For GC, the carrier phase is an inert gas (e.g. He, Ar, N2).
• The sample needs to be vaporised if it’s not already a gas, and injected as a pulse into the
carrier stream.
• The stationary phase is usually a column containing the stationary phase on a fused silica
support:
o The column is usually very narrow (say 2 mm diameter) and may be 1-10 m long;
physically it will look like a coiled loop.
o There are many types of silica and modified silica so different separations can be
achieved.
• Separation is based on the components having different retention times on the column:
o affected by boiling points of the substances to be separated.
o affected by selective adsorption of a component onto the stationary phase.
• The column is located in an oven, the temperature of which can be controlled.
• For good separations in a reasonable length of time, it’s common for the temperature of
the oven to be increased over the course of the experiment.
[from http://teaching.shu.ac.uk/hwb/chemistry/tutorials/chrom/gaschrm.htm]
• There are a range of detectors available. We’ll mention the two most common.
• Flame ionisation detectors (FID):
o These are widely used for analysis of organic compounds.
o Gas at the column exit is mixed with hydrogen and air and burnt. Any organic
compounds present produce ions and electrons in the flame making it capable of
conducting electricity. The FID measures the current response to an electric
potential at the burner tip.
o The FID response has high sensitivity, a large linear response range, and low
noise. It is also robust and easy to use, but it destroys the sample.
o For hydrocarbons, the FID peak areas are proportional to the number of carbon
atoms present in that component of the sample.
• Thermal conductivity detectors (TCD):
o These compare the thermal conductivity of the gas at the column exit with a
reference flow of carrier gas (usually He). Any change is due to the presence of
sample compounds.
o Disadvantage: TCDs are slightly less sensitive than FIDs and have slightly lower
resolution (as they have a larger dead volume).
o Advantage: TCDs can be used to detect any compound (i.e. not just
hydrocarbons), and the sample isn’t destroyed.
o Because the thermal conductivity of organic compounds tend to be similar to each
other, TCD peak areas for hydrocarbons are roughly proportional to the
concentration of that component.
45°C
time,min
145°C
time,min
• Note modern instruments often use the acronym HPLC (for “high-performance” or
“high-pressure” LC).
• In this case the sample is in the liquid phase (by dissolving into solution if it’s not already
a liquid).
• The stationary phase is a solid packed into a column; it can be a liquid-coated solid.
• Different detector systems are used (e.g. UV, fluorescence, refractometry).
• A variety of different separation mechanisms are used.
• Liquid-Solid separations are based on the intermolecular interactions between sample
molecules and the solid phase.
o For instance, these may be polar interactions or hydrogen bonding
interactions.
o The “normal” case is that the solid has hydroxyl groups at the surface and so has
an affinity for polar groups:
❧ Less polar molecules will pass through the column faster than polar
molecules if the surface of the solid likes polar species.
o In “reverse phase” chromatography, the stationary phase is made hydrophobic
(e.g. silica with n-alkyl chains covalently bound to its surface):
❧ In this case, hydrophobic compounds will have longer retention times than
hydrophilic ones.
o The retention times are critically affected by the polarity of the solvent (carrier
phase).
Collect
product 1
• Liquid-Liquid separations are based on the partition of the sample between two liquid
phases.
• Size-Exclusion chromatography is based on the molecular size of the compounds
present.
o The stationary phase consists of solid beads containing small pores.
o Large compounds can’t enter inside the beads and thus will elute first.
o Smaller compounds enter the beads and will have longer retention times.
• Ion-exchange chromatography operates on the basis of selective exchange of ions
in the sample with those in the stationary phase.
• These are simply a combination of the techniques that we’ve discussed so far.
• Examples include:
o GC – MS
o LC – MS
o LC – NMR
• With these techniques, separation and sample identification of complex mixtures can be
performed in a single piece of equipment.
• This represents the “state of the art” in modern analytical chemistry.