Intro To Spec Flashcards
Electromagnetic radiation consists of what fields and in which direction
Oscillating electric and magnetic fields that are perpendicular
What is the wavelength of visible light
700nm to 400nm
Why is the Change in wavelength an order of 3 for each type of wave in the electromagnetic spectrum (radio waves, or waves, uv , etc.
Because of the wavelengths relationship to the speed of light (3x10^8m/s)
Humans can only see a small part of the
Electromagnetic spectrum
Equations for energy and energy per molecule
On cheat sheet
Does vocable light have enough energy to break typical chemical bonds
No, should be nondestructive
Slide 5
What is colour perceived as
What gives colour
The average of the wavelengths of light that is perceived by the eye
The light transmitted/reflected from a substance it what travels to our eyes and let us see a colour
Why is chlorophyll green
This is because green light is not strongly absorbed by chlorophyll
This means it is instead transmitted and reflected
What implies that energy levels are quantized
The fact that only some frequencies of light are absorbed
What has to happen for absorption to MAYBE occur
The change in energy between energy levels has the be equal to the energy of the photon
But other criteria has to be satisfied
Does absorption of a photon happen quick or slow
Quick
In uv/vis spec what nature of levels does the molecule change
What about IR
Electronic levels
Vibrational or rotational levels
What is the difference in vibration and rotational levels vs electronic levels
vibration and rotational levels are closer together than electronically levels
Which transitions use mainly kinetic energy
Translational (moving in x y or z direction)
Rotational
Vibrational
Which transitions use mainly potential energy
Electronic
Nuclear
Is the change in energy of an electronic transition greater or lower than that of a vibrational transition
Greater
What is wave number proportional to
1/ wavelength
What is the order from highest change in energy to lower for each transition
N E V R T
What is an example of a electronic transition
Vibrational transition
Chlorophyll absorbing at 660nm and having a energy of 181kJ/mol
The vibrating aldehyde c=o in chlorophyll B at 1655cm-1
What is an example of a electronic transition
Vibrational transition
Chlorophyll absorbing at 660nm and having a energy of 181kJ/mol
The vibrating aldehyde c=o in chlorophyll B at 1655cm-1
What happens after a molecule gets excited
Can emit a photon (as either fluorescence or phosphorescence)
Can lose that energy
What ways can molecules lose energy after being excited
Internal conversion (so losing heat)
Redox reaction (losing an electron)
Energy transfer (to another acceptor molecule)
The lifetime of the excited state (time before energy is lost) is longer for fluorescence or phosphorescence
Phosphorescence
The emmited photon has _____ energy than the excitation photon
Less
Single beam absorption spec details
The monochromator scans across a wavelength region then only allows a specific wavelength of the incident light through
I0 is intensity from incedent light
If is intensity at detector (final instensity)
The I0 is measured with a blank to get absorbance when there is no sample
Only then can you measure sample
Why are aqueous buffers used in measuring absorbace of samples
To keep the protien in a pH range that is works at
Double beam absorption spec details
The I0 and If are measured at the same time through use of a beam splitter
The I0 is adjusted to equal If with solvent in both cuvettes (so that the Intensity of both are the same and can be easily compared)
The wavelength and path length for both cuvettes is the same
What is special for the monochromater is double beam spec
Can automatically scan the wavelgngh region of interest
What is the final output of a double beam spec
Log (I0/If) versus the wavelength
For accurate absorbances measurements what should the absorbance be between
0.1 and 1.0
When is absorbance less than zero
When the blank absorbs more than the sample
What causes small changes in Absorbances
Large changes
Temp
Solvent used
Ph
Path length
Concentration of molecule
Molecule used
Protiens are measured as
Percentage (w/v)
Or mg/ml
Slide15
When there are two non interacting absorbing species what is their total absorbances equal to
The sun of the absorbance for both species
Find absorbances and concentrations using the equations
Slides 16 and 17
