Absorption, luminescence, lasers

Question 1

What is monochromatic light?

Answer 1

Light is monochromatic if its spectrum consists of a single wavelength only

Question 2

What kind of special characteristics does laser light have?

Answer 2

• monochromatic
• coherence in time and distance - small divergence
• high light density.

Question 3

What kind of special characteristics does laser light have?

Answer 3

• monochromatic
• coherence in time and distance - small divergence
• high light density.

Question 4

List the types of interactions laser light can have with tissues.

Answer 4

• photothermal (laserthermy, coagulation, vaporization, carbonization)
• fluorescence, photochemical reactions
• photodissociation
• multiphoton ionization

Question 5

When is electromagnetic radiation coherent?

Answer 5

If it consists of photons capable of forming observable interference fringes.

Question 6

What basic phenomena is the generation of laser emission based on?

Answer 6

• population inversion is needed for light amplification to occur, and it is only possible in systems with 3 or more energy levels
• stimulated emission is needed to give rise to coherent monochromatic light.

Question 7

What is the approximate coherence length of a laser and that of a classical light source?

Answer 7

10 ele. 10 cm and a couple of cm, respectively

Question 8

Align in ascending order the following transitions according to their energy difference: vibrational, rotational and electronic!

Answer 8

rotational < vibrational < electronic

Question 9

Write the Lambert-Beer law and interpret the variables in the formula!

Answer 9

lg J0/J = EcL = A

J - intensity of light after passing through a material with thickness L
Jo- incident intensity of light when it enters the sample A – absorbance (optical density or extinction)
E - molar extinction coefficient
c - concentration in mol/liter
L - optical path length.

Question 10

What does the molar extinction coefficient depend on?

Answer 10

It depends on the type of the absorbing material, the wavelength of the light, temperature, the type of the solvent and the environment.

Question 11

How many fold does the intensity of light decreases if the absorbance (optical density, extinction) of a solution is 1?

Answer 11

It decreases 10-fold.

Question 12

What is the definition of the molar extinction coefficient?

Answer 12

It is the absorbance (optical density) of a solution with a concentration of 1M and an optical path length of 1 cm.

Question 13

At what wavelength are the characteristic absorption maxima of proteins and nucleic acids?

Answer 13

proteins 280 nm, nucleic acids 260 nm

Question 14

Which amino acids have reasonably high absorption?

Answer 14

Tyr, Trp, Phe

Question 15

What is the definition of a singlet and a triplet state?

Answer 15

In a singlet and a triplet state the number of unpaired electrons is zero and two, respectively. In a singlet and a triplet state, the value of the resultant spin multiplicity is 1 and 3, respectively.

Question 16

What are the possible ways of relaxation of an excited electron in a molecule? (List at least 5 of them!)

Answer 16

• vibrational relaxation
• internal conversion
• intersystem crossing
• fluorescence
• phosphorescence
• delayed fluorescence
• energy transfer to another molecule.

Question 16

What are the possible ways of relaxation of an excited electron in a molecule? (List at least 5 of them!)

Answer 16

• vibrational relaxation
• internal conversion
• intersystem crossing
• fluorescence
• phosphorescence
• delayed fluorescence
• energy transfer to another molecule.

Question 17

What is the definition of fluorescence lifetime?

Answer 17

The time during which the number of excited molecules decreases to 1/e-times (37 %) of its initial value.

Question 18

What is a., scintillation, b., chemiluminescence, c., photoluminescence?

Answer 18

Processes where photon emission is elicited by
a., ionizing radiation
b., chemical reaction
c., excitation by photon

Question 19

How can fluorescence quantum efficiency (yield) be defined?
(One definition is sufficient.)

Answer 19

The fraction of excited molecules emitting a fluorescent photon, OR the number of fluorescence photons divided by the number of absorbed photons, OR the rate constant of fluorescence divided by the rate constants of all possible de-excitation processes.

Question 20

Why is the fluorescence quantum yield always smaller than one?

Answer 20

Because relaxation from the excited state can be accomplished not only by fluorescence emission.

Question 21

What is the lifetime range of fluorescence?

Answer 21

t = 10 -9 – 10 -7s

Question 22

What is the lifetime range of phosphorescence?

Answer 22

t=10 -6 –10s

Question 23

Why is phosphorescence lifetime longer than fluorescence lifetime?

Answer 23

Because phosphorescence is the result of spin- forbidden transitions.

Question 24

Why is Förster type resonance energy transfer a sensitive method for distance measurements?

Answer 24

Because its probability is proportional to the inverse sixth power of the separation between the donor and the acceptor.

Question 24

Why is Förster type resonance energy transfer a sensitive method for distance measurements?

Answer 24

Because its probability is proportional to the inverse sixth power of the separation between the donor and the acceptor.

Question 25

What can Förster-type resonance energy transfer be used for in biology?

Answer 25

For measuring inter- and intramolecular distances.

Question 26

List at least five parameters which can be determined using fluorescent measurements!

Answer 26

• DNA, RNA, protein and lipid content of a cell, or the
  quantity of any kind of material that we tagged with a
  fluorescent label.
• permeability of the cell membrane
• intracellular enzyme activities
• membrane potential
• intracellular calcium level
• intracellular pH
• presence and density of cell surface antigens and
  receptors
• mitochondrial potential and the number of
  mitochondria per cell.