spectroscopy part 2

what must the chromophore contain

double bonds (pi system)

what will increasing conjugation reduce

the energy gap

what can modify the chromophore absorption

functional groups

what is absorbance the logarithm of 

the ration of light before and after passing through the sample 

what is absorbance proportional to

the chromophore concentration

what is beer-lamberts law

A= e x l x c
where:

• A = absorbance

• e = molar absorptivity or extinction coefficient 

• l = path length of cuvette

• c = conc. of the solution 

what will fluorescent molecule emit light of when excited with light 

will emit light of lower energy (longer gamma) when excited with light 

what is fluorescence quenching 

the light emission is stopped 

what does adding NaCl to tonic water do

quenches quinine fluorescence

what is the excitation and emission of quinine in UV-Vis fluorescence 

• excitiation- 349 nm

• emission- 481 nm 

when is ethidium bromide fluorescent

when dissolved in organic solvent but not in water (quenching)

in the presence of DNA, what does ethidium bromide do 

• ethidium bromide slides between the base pairs acting as an intercalator

• when bound, the fluorescence is recovered, and the dye is used for the detection of DNA 

what will the frequency of the vibration in vibrational transitions depend on

the bond nature and strength

how are IR spectra generally plotted as

transmittance vs wavenumber

what does conjugation do to the bond and what does this mean for IR spec 

• ‘weakens’ the bond

• reduces the IR absorption wave number

what is mass spec used to measure

measures the mass of molecules by ionising them and measuring their mass-to-charge ratio (m/z)

is mass spec a spectroscopy

• no

• there is no electromagnetic radiation involved 

• but it generates spectra with the distribution of mass of the ions generated

what is mass spec used as

• both a stand-alone analytical instrument and as a detection method in chromatography

• its importance is due to its high sensitivity 

what are the applications of mass spec determined by

the combination of different ways of generating and analysing of ions

what is electron impact

when a sample is vaporised and bombarded with electrons, it charges the molecules in the sample and normalising it makes them break down into small fragments (fragmentation)

what is chemical ionisation

• when a sample is bombarded with cations (H+, K+, Na+)

• these ions bind to the sample molecules, forming combined ions

what is electrospray ionisation

• when a sample is mixed with some salt or acid and is vaporised, and the molecules get the charge from the ions

• it can form cations or anions 

what is a matrix-assisted laser desorption/ionisation

• sample is dissolved in a non-volatile matrix and put in a vacuum chamber

• then it is hit with a laser that desorbs the sample molecules

• it is mainly used for macromolecules and peptides

iii elements have multiple isotopes, how do you detect these ions

ions will correspond to each isotope corresponding to their abundance

what are the 2 non-radioactive isotopes of carbon

• C12 and C13

• abundance of C13 is only 1.11%

what are the 2 isotopes of bromine

• Br79 and Br81

• in an approximate abundance of 1:1

how do you calculate the amount of carbon atoms in a molecule 

number of C = abundance [M + 1]+/abundacne M+ x 100

what are the detectors for HPLC

• UV-Vis

• mass spec

• light scattering

• refraction index

• fluorescence

• conductivity