Vibrational Spectroscopy

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Vibrational Spectroscopy

• Physical Biochemistry, November 2006
• Dr Ardan Patwardhan, a.patwardhan_at_imperial.ac.uk,
  Faculty of Natural Sciences, Imperial College
  London

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Molecular motion
In general, molecules can undergo three types of
motion
Translation, i.e., displacement of the whole
molecule
Rotation
Vibration
All three types of motion have associated
energies, but only rotational and vibrational
motion have quantized energies
Under certain circumstances, the rotational and
vibrational motion of a molecule can exchange
(absorb or emit) energy with EMR this is the
basis for rotational and vibrational
spectroscopies
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Born-Oppenheimer approximation

• The time-scale for rotations(10-11s) is several
  orders of magnitude longer than the time-scale
  for vibrations (10-13s), which in turn is several
  orders of magnitude longer than electronic
  transitions(10-15s)
• The influence of each of these phenomena can
  therefore be viewed separately, and the energy
  levels are simply the sum of the energy
  contributions from the three phenomena

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Combining the electronic, vibrational and
rotational energy levels

• The energy differences between rotational energy
  levels is smaller than the differences between
  the vibrational levels, which in turn are smaller
  than the differences in energy between electronic
  levels
• To describe a molecules energy state, simply add
  the three contributions together
• Combine the vibrational energy levels with each
  of the electronic energy levels, and then combine
  the rotational energy levels with every
  combination of these

Electronic
Vibrational
Rotational
Total
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Molecular rotation

• Can be probed with microwave radiation in the
  case of molecules with electric dipole moments,
  e.g. heteronuclear diatomic molecules
• Can be used to determine bond lengths in small
  molecules

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Vibrating diatomic molecule Simple harmonic
oscillator
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Simple harmonic oscillator

• Note that E0 ? 0 ? molecule is always vibrating
  and never completely still ! Manifestation of
  Heisenberg uncertainty principle

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The anharmonic oscillator

• In reality, stretching and compressing a bond are
  not energetically equivalent
• The spacing between energy levels decreases with
  increasing u

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Normal modes
Symmetric stretch3652 cm-1
Antisymmetric stretch3756 cm-1
Symmetric bend1595 cm-1

• Normal modes have energy levels that are
  independent of each other and can interact with
  EMR independently
• Any molecular vibration can be described as a
  linear combination of the normal modes

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Polyatomic molecules

• A varying electric dipole is necessary for a
  normal mode of vibration to produce a spectra
• The symmetric stretch in the above example will
  not produce a vibrational spectra

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Regions of the spectrum

• Fingerprint region600-1500cm-1- Complex, can be
  used to identify compound
• Functional groups have typical absorptions in
  1000-4000cm-1 region

IR spectrum of 1-hexene
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Functional groups

• Groups with light atoms have a higher frequency
  than groups with heavier atoms
• Stretching modes usually have higher frequencies
  that bending modes
• The stronger the bond the higher the frequency
  (triple bond gt double bond gt single bond)

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Important characteristic vibrations

• N-H One or two sharper peaks between
  3500-3300cm-1
• O-H Broad peak between 3650-3200cm-1
• CO Strong and sharp peak between 1820-1660cm-1

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Basic IR spectrometer

• Measurement is alternated between sample and
  no-sample to minimize influence of background
  radiation and environmental effects
• If the sample is in a solvent, then a reference
  cell containing only solvent can be used to
  cancel the solvent absorption spectrum

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Sample Handling

• Solvents water and alcohol are seldom used as
  they absorb strongly and attack cell window
  materials. No solvent is transparent through-out
  the entire mid-IR region. CCl4 and chloroform are
  common solvents
• Liquid samples are often analyzed neat for this
  reason and cell thicknesses can be very small
  ranging from 0.1 to 1.0 mm
• Cells NaCl and KBr often used as a transparent
  material sample holder. Both glass and quartz
  are opaque in the IR region

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Attenuated Total Reflection (ATR) Infrared
Spectroscopy

• If the refractive index of the block is higher
  than the sample, given certain constraint on the
  angle of the incident beam, it will be reflected
  at the block sample interface
• For each reflection, the beam only penetrates
  1-10um into the sample, and will be attenuated
  depending on what is in this thin layer of the
  sample
• Multiple reflections helps amplify the signal
• Excellent technique for IR spectroscopy of thin
  layered samples such as membranes

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Fourier Transform Infrared (FTIR) spectroscopy
More specifically, the interference imposes a
cosine modulation on the spectrum, which has a
periodicity that is depended on the path
difference
This encoding is known as the Fourier transform
(FT)
To reconstruct the absorption spectrum we need to
decode the signal recorded as a function of
path difference
This decoding is performed computationally by
imposing cosine modulations onto the detected
signal and is known as the inverse Fourier
transform (IFT)
The sample is irradiated by the full spectrum of
the source

• Due to interference between the two split beams,
  for a given path difference, some wavelengths
  will interfere constructively, others
  destructively, and the rest somewhere in the
  middle

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Effect of interference on the spectrum

• The frequency of the modulation imposed on the
  spectrum increases with the path difference dx

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FTIR Advantages

• Spectrum over entire frequency range obtained at
  once
• For a given recording time, FTIR is less noisy
  a 10 min recording with conventional techniques
  takes only 10s
• Uniform resolution over entire spectrum

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Case Study

• Isotope editing modifying the isotope of an atom
  in a functional group is a method for
  specifically shifting the absorption frequency of
  a targeted group
• By examining the difference spectra between
  edited and unedited specimens, very precise
  information about specific groups can be obtained
• In the example, the amide I band of a-crystallin
  has been labelled (12C to 13C) leading to a shift
  in absorption from 1632cm-1 to 1591cm-1

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Main Points

• Molecular vibrations can be probed using IR
  radiation
• Only normal modes that represent varying electric
  dipoles will interact with EMR
• The FTIR technique provides speed and resolution
  in acquiring spectra
• Isotope editing allows one to study specific
  functional groups in a complicated spectrum