Types Of Spectroscopy:

Question 1

Fluorescence spectroscopy:

How does it work:
What it uses:
Sensitivity:

Answer 1

Takes certain wavelengths of light and transmit it through sample.
Light detected by detector.
Causes ER to match to sample.
Energy absorbed and given off at different wavelengths.
Fluorophores e.g. Rhodamine (aromatic).
Can be sensitive.
Can detect single molecule and analyse by fluorescently tagging molecule.
Can see where fluorescently tagged molecule goes to.

Question 2

NMR spectroscopy:

Sensitivity:
How it works:
Frequency range:
What it identifies:

Answer 2

Insensitive
Nuclear spin occurs due to magnetic field. H exists with mass number of 1 so is magnetically active.
Frequency is in radio frequency to microwave region.
Different H show different frequency.
Can identify chemical compounds. Can look at individual atoms in molecule.
Determine 3D structure of proteins and nucleic acids.

Question 3

Mass spectroscopy:

Answer 3

Measures mass of molecule.
Can identify and characterise sample.
Insensitive.

Question 4

X-Ray crystallography:

Answer 4

Uses diffraction pattern of crystalline solids exposed to X-Ray. Determined 3D structure.

Question 5

Absorbance and transmission spectroscopy:

Answer 5

Light is absorbed at certain series of frequency.
Absorption is positive signal.
Transmission is negative signal.
E.g. Chlorophyll absorbs blue and transmits green.
Heme absorbs blue and transmits red.
Doesn’t interact with other wavelength of light.

Question 6

UV-Vis spectroscopy:

Answer 6

Detects aromatic AA so gives particular absorption features.

Beer-Lambert law - amount of absorption you have on a given wavelength is a product of molar absorptivity, pathlength and conc.

A = εlc

Molar absorptivity is ε
Pathlength is l
Concentration is M

Pathlength depends on content of material. Less light absorbed if thick.

Usually part of another process (e.g. Chromatography)
Can find separate compounds and know how much you have of that compound.

Question 7

Infrared spectroscopy:

Transmission or absorption:
Comparison to UV-Vis spec:
Resonance frequencies:
What it identifies and doesn’t identify:

Answer 7

Shows transmission rather than absorption.
Uses longer wavelengths and lower frequency than UV-Vis spec.
Resonance frequencies are associated with bond vibrations: stretching, bending and twisting.
Shows characteristic patterns of signal.
Can identify chemical compounds.
But can’t use it to study macromolecules e.g protein. Due to unique structures of monomers so produce different frequencies/vibrations.