Fluorescence

Question 1

What is a fluorophore, what are its characteristics?

Answer 1

Fluorescent molecules
Typically contain aromatic groups or have several Pi bonds

Question 2

What is the process of fluorescence?

Answer 2

1. Photon of excitation light is absorbed (raises everyday level of an electron)
2. Some energy is lost
   (Molecular collisions transfer to proximal molecules)
3. Remaining energy is emitted as a proton as molecule returns to the ground state

Question 3

How do emission and absorption relate with respect to wavelength and energy? Why?

Answer 3

Absorption Absorbance is at a higher energy
Therefore the higher the frequency, the lower the wavelength
Emission is a higher wavelength because energy lowers before emitting a photon

Question 4

Label an emission and excitation spectra

Answer 4

Intensity vertical
Left - excitation (lambda max incident light) light on molecule
Right - emission (lambda max emitted light) light off of molecule

Question 5

What is the stokes shift? What does the size relate to?

Answer 5

The stokes shift is the difference in the excitation and emission lambda maxima
Emissions maxima do not change w/ varying excitation wavelengths
Emission intensity changes with varying excitation wavelengths

Question 6

What is quenching? What causes it?

Answer 6

Reactions increasing or changing fluorescence
Causes:
pH (aqueous)
Adjacent molecules or fluorophores (formation of complex in the ground state)
Solvent polarity (aggregation)

Question 7

What do we use for quantitation?

Answer 7

Fluorescence
(Lower concentration - emission intensity is proportional to analyte concentration.