Spectroscopy Flashcards
What do the following types of energy involve?
- Nuclear
- electronic
- vibrational
- rotational
- translational
- Nuclear: protons and neurtrons within the nucleus
- Electronic: Electrons between a few atoms
- Vibrational: Few atoms
- Rotational: Whole molecule
- Translational: Whole molecule moving large distance
Whats the formula for the Boltzmann distribution of molecules over energy levels?
What makes molecules move up into a higher energy level?
As temperature increases, the moelcules inhibit higher energy levels
What is electromagnetic radiation?
What is the energy for a given frequency described as?
Electromagnetic radiation has an electric and magnetic component. where the electric component is oscillating and the magnetic component is rotating from north and south
Energy (E) for a given frequency (u) is
E= hu
where h is the Planks constant (6.626x10-34 J s)
Whats the order of the electromagnetic spectrum from the smallest frequency to the largest?
What are the conditions in order for the electromagnetic radiation to be absorbed?
- must be two or more energy levels
- Charge displacement must occur:
a. Linear displacement: electric transition dipole
b. Rotation of charge: magnetic transition dipole
* ∆E= hu (must be between the ground and excited state)
Once the incident radiation, I0, is absorbed what are the types of radiation you can get from this?
- Transmitted radiation, It
- Emitted radiation, Ie
What are the characteristics of a spectrum?
What does the intensity depend on?
The number of moelcules that can be excited from ground state
What determines if population different is small and therefore sensitivity to be low?
∆E is roughly kT
Name 3 types of spectroscopy and which one sees more absorption of the electromagnetic radiation?
- NMR
- rotational
- electronic (sees more absorption)
What is absorption dependent on?
What equation describes this?
What does each part of the equation represent? Units?
The concentration of absorbing species
The Beer Lambert law:
Log10(I0/ It) = OD = ecl
e= the extinction coefficient (absorbing power of a species at a particular wavelength)
C= the concentration of absorbing species (mol dm-3)
l= length of solution of light passes through (cm)
In the Beer Lambert law, when OD >1 what does this mean?
It is no longer linear–> not accurate
What is the Beer Lambert law?
Its combined of 2 laws and each are correlates which state that, the absorbance of light is proportional to the thickness of the sample or absorbance is proportional to the concentration of the sample
When is maximum absorbance seen in intensity?
When the transition dipole is aligned with the polarised light
The orientations of light leads to differential…
absorption
What is natural linewidth determine by?
Lifetime of excited state (t)
Whats the Heissenberg uncertainty principle?
it states that its impossible to know simultaneously the exact position and momentum of a particle
That is, the more exactly the position is determines, the less known the momentum and vice versa
How does decay from an excited state occur?
- stimulated emission: through application of external field
- conversion to other excited state
- non-radiative loss to environment
- resonance energy transferred to nearby molecule
- chemical reactions
Why is spectroscopy important?
- Study conformation of molecules
- Structure of molecules
- Reports on their environment
- Quantitation
- Protein/ DNA concentration determination
- Biochemical assays
- Detection
- DNA sequencing
- Microscopy
- Cytometry
Learn absorbance/ emittence
Coupling to vibrational states
What are the types of electronic transitions?
1. Transition involving p, s and n electrons
Organic compounds:
- Saturated (σ –> σ (< 150 nm)*, n –> σ*(<250 nm))
leads to displacement of charge when σ –> σ*
- unsaturated (π- π*, n–> π* (200-700nm))
2. Transitions involving charge transfer
- electrons move from ligand donor to metal acceptor
3. Transitions involving d and f electrons
- transitions between d orbitals split by the presence of a ligand
Whats determines
- Dependent on molecular structure
- No easy rule for assignment, however:
- Presence of resonance >300 nm suggests presence of conjugated polyene chain/ polycyclic aromatic
- Conjugated systems typically have larger extinction coefficients
- Refer to databases (as above doesn’t give much detail)
What are the changes that can occur in a UV/ visible spectra?
Name the chromophores found in biology?
- Proteins
- Nucleic acids
- Metal ions
Whats the most common protein chromophore?
Peptide bonds
How many transition states are associated with this molecule?
2
In proteins sidechains are there transition dipoles and conjugates bonds present?
What determines a higher extinction coeffient in a sidechain?
yes
The number of delocalised electrons
What can influence energy levels? and how?
Solvent polarity by stabilising or destabilising the excited state
How does changing the exposure to light influence the wavelength in prosthetic groups?
When in dakr conditions and then exposed to light, the wavelength can decrease
Do nucleic acids have more transition dipoles then protens?
Yes
How does nucleic acid transition coupling effect the excited state?
What does coupling depend on?
When two transition dipoles are coupled, they interact, splitting the energy levels (excitation coupling)
The coupling depends on relative orientation
- When the transition dipole is parallel to the induced dipole what occurs?
- When the transition dipole os collinearl to the induced dipole what occurs?
- Hypochromism
- Hyperchromism
- What effect does structured DNA cause
- What effect does unstructured DNA cause
- Structured DNA –> parallel dipoles Hypochomism
- Unstructured DNA –> Dipole unaligned hyperchromic shift
