Chapter 29 - Analytical techniques Flashcards
what is the purpose of chromatography and what do all types have
“Chromatography is used to separate individual components from a mixture of substances”
All forms have:
- a stationary phase- usually a solid or liquid supported by a solid, the stationary phase does not move
- a mobile phase - usually a gas or liquid, the mobile phase does move
state the advantages and disadvantages to TLC
advantages:
- quick, inexpensive
- indicates number of components
disadvantages
- can be difficult to identify substances
- sometimes not full seperation
explain how TLC works
it uses a TLC plate
- made of glass or plastic and coated in a silica (or adsorbant surface)
- the silica is the stationary phase
different components have different attractions for the adsorbant
so they bind differently to it and seperate out
explain the method for TLC
1) take TLC plate, draw a line 1cm from the bottom in pencil
2) use a capiliary tube, spot the sample onto the line
3) prepare chromatography tank (beaker with watch glass on)
4) fill to 0.5cm with solvent (usually ethanol)
5) put in TLC plate
6) allow solvent to rise to about 1cm from top then remove and mark the solvent front immediately
7) mark any visible spots and let dry
what do we use to analyse TLC and how can we use these values
Rf values can be measured and compared to standard values
Rf = distance moved by component/distance moved by solvent front
they should never be >1
when do we tend to use gas chromatography and what are the stationary and mobile phases
- we tend to use it for complex mixtures
- mobile phase = inert gas
- stationary phase = adsorbed liquid (high bpt)
how does gas chromatography work
- a small amount of a sample is injected, it is vaporised and carried through the long column using a carrier gas (inert)
- the column contains the stationary phase, a high bpt liquid
- the sample will repeatedly dissolve into the stationary phase and evaporate out into the mobile phase
- depending on their solubility, their retention time values are different
- the higher the retention time, the greater the solubility
what are the advantages and disadvantages of gas chromatography
advantages:
- it has better seperation than other techniques
- only very small samples are required
disadvantages:
- can only be used where the substance is volatile
- doesn’t work well with biological substances
- if there are small amounts of a substance with the same retention time as another, they can be hidden
how can we analyse the results of gas chromatography
- if a component is more soluble, it will have a greater retention time
- these can be compared to known retention times to identify substances
- the area under the curve produced by the detector is directly proportional to the concentration of the substance (peak integration)
how do calibration curves work and how can we make them
1) prepare standard solutions of different concentrations
2) get gas chromatograms of each
3) plot a calibration curve of relative peak area (on Y) against concentration (on X)
4) this can then be used in reverse, peak integration area –> true concentration
what is the test for an alkene
- add bromine water
- if the bromine water is decolourised, there is a C=C bond present
- orange to colourless
what is the test for a haloalkane
- add ethanol and silver nitrate and heat to 50 degrees
- if a white, cream, or yellow precipitate is formed then a haloalkane is present
what is the test carbonyl
add 2,4 dinitrophenylhydrazine
if an orange precipitate forms, a carbonyl group is present
what is the test for an aldehyde
add Tollens’ reagent and warm
if a silver mirror is produced, an aldehyde group is present
what is the test for primary and secondary alcohols, and aldehydes
add acidified potassium dichromate and heat
they will turn orange to green if the groups are present
what is the test for carboxylic acids
- add aqueous Na2CO3
- if CO2 is formed then it was a carboxylic acid
what is the test for phenol
- add bromine
- it will decolourise and form a white precipitate
what is NMR and what does it use
- NMR is a form of spectroscopy
- it uses a strong magnetic field and radio waves
what is nuclear spin and when does it occur
- like electrons, nuclei can have opposite spin states
- this is only relevant when there are an odd number of nucleons, C13, H1 are the main examples
what is resonance and how is it important
- if the nucleus’ spin state is opposed to the external magnetic field then it has high energy
- if the nucleus’ spin state is aligned with the external magnetic field then it has low energy
- at any given point there are slightly more at the low energy state
- at a certain frequency, the nucleus rapidly flips between spin states, this is resonance
- because of the greater number of low energy spin states there is a slight absorption of energy at this point
how is the magnetic field produced and what is it proportional to
strength of external magnetic field is directly proportional to the frequency required for resonance
to get the strong external magnetic field, superconducting electromagnets cooled to 4K are used
what is the reference compound that we measure chemical shifts against and what is the importance of its structure
TMS
tetramethylsilane
- the protons in the centre are the most shielded they can be
what factors affect the chemical shift of a substance and why
- its chemical environment
- electrons have a spin and set up a small magnetic field themselves
- this means for a more shielded nucleus, the magnetic field experienced by the nucleus is lower and therefore the frequency at which it resonates is lower
- so the chemical shift is lower
how is an NMR spectrum run
- the sample is dissolved in a substance and placed in an NMR tube
- it is spun to even out impurities and zeroed to TMS then a range of radio freqs are fired at it with a uniform magnetic field
what are the two important pieces of information given by a C13 NMR spec
- the number of carbon environments
- the types of carbon environments
what is the carbon environment and what important thing is there
- carbon atoms bonded to different groups or near a major functional group have different shifts
- if two carbons are positioned symmetrically in a molecule then they will have the same chemical shift
which 4 pieces of information does a proton NMR give us
- the type of proton (hydrogen) environments
- the number of proton environments
- the relative numbers of each type of proton environment
- the number of non-equivalent protons on adjacent carbons due to spin-spin coupling
when do protons have different chemical shifts
- protons in the same environment or symmetrical will have the same chemical shift
- protons in different environments will have different chemical shifts
e.g.
butanoic acid = 4 chemical environments
butandioic acid = 2 chemical environments
what does the area of the peak tell us in proton NMR
- in C13 it tells us nothing
- in H1 it tells us the relative numbers of each proton environment
what is spin-spin coupling
- spin-spin coupling is when a proton’s spin interacts with the spin states of protons on adjacent carbons
what does spin-spin coupling tell us and what is the n+1 rule
- it tells us the number of protons on adjacent carbons
- for a proton where there are n protons on the adjacent carbon, it will have n+1 peaks
what is the significance of OH and NH in H1 NMR
apart from when they’re as part of a COOH group, they can appear anywhere on the spectrum
they don’t have spin-spin coupling
how can we tell if a proton peak is OH/NH or just another proton, what happens with COOH
- the protons on these groups are labile so they will dissapear if the spectrum is run with D2O (heavy water) as 2D (2H) doesn’t appear on the spectrum as it has an even number of neucleons
1) run spectrum as normal
2) re-run spectrum with D2O
3) any peaks that disappear are OH or NH
COOH acts the same as OH
what is the generalised method for identifying a substance form its H1 NMR spectrum
1) analyse the types of proton environment and numbers present
2) analyse splitting patterns
3) analyse chemical shifts and combine all this info
what are the general steps for predicting a C13 spectrum
1) draw structure
2) identify number of carbon environments
3) predict shifts (approx.)
what are the general steps for predicting a proton NMR spectrum
1) draw out structure, identify number of proton environments
2) predict their shifts
3) predict peak heights
4) predict splitting patterns and combine all info
what term is given to the process by which separation is achieved in gas chromatography
partition
define retention time
Retention time is the time between injection and emergence (or detection) of a component
What is a way to distinguish between carbonyls e.g. ketones, aldehydes, hydroxyketones without using spectroscopy or chromatography
use the 2,4 DNP method, where you filter and recrystallise the precipitate and compare to known values
which peaks only ever form singlets
COOH/OH and NH
