Measuring Absorbance Flashcards
The absorbance at any wavelength is equal to what
The sum of all absorbances or all species in the solution
Equation on slide 3
What is the isobestic point
If you have a reaction that is linked stoichiometrically, the isobestic point is the wavelength where where the sum or the
A absorbance= B absorbance
If you see this it’s a good indicator that the reaction is linked
What is fluorescence quenching
Adding something to the solution that takes energy from the excited state of the analyte and releases it nonradiatively
Ex. O2 takes energy from other atoms
Fluorescent quenching equation
Slide 5 OKAY
What is photo bleaching
When something loses fluorescence when exposed to light
It’s permanent and the molecule does it to itself (unlike quenching)
It breaks down the fluorophore
What is fret
Instead of quenching, the energy from one species is transferred to another fluorescent species
Ex. One thing emits blue after absorbing purple.
A k+ binging protien is attached to it and another species that emits yellow.
When k is in the solution the binding protien brings the two species together and the energy from the first species excites the second species and the second species absorbed blue and emits yellow
What are the parts of a spectrophotometer
- A light source that is ploychromatic
- A monochrometer that take one specific wavelength from the light source
- Sample
- Detector (senses light intensity and gives p and p0)
What are the 4 types of light sources
Arc sources
Continuum sources
Line sources
IR sources
What are arc sources
Emits like a continuum source
They have a lamp that holds high pressure gas, ex. Xe, D2, Hg
This high pressure gas create “band broadening”
The lamp is named by the type of gas that’s in it and the wavelength is determined by the gas as well
What are continuum sources
Basically light bulbs
Ex. Tungsten lamp (quartz, halogens)
The max wavelength empires depend on the T (transmittance)
What are line sources
Specific narrow wavelength ranges
Laser: give stimulated emmision, had narrow wavelength and the light is coherent
LED: had a broader wavelength and the light is not coherent (still narrower than continuum)
low pressure gas (HCL): atomic line emmision, it’s the narrowest with 0.1nm or less
What are IR sources
They’re mostly continuum sources and weaker than UV-vis light sources
Ex. Glober (ceramic)
Nernst glower (ceramic)
Nichrome wire (wire)
What type of excitation are laser and LED light sources
Molecular excitation
