29 - chromatography and spectroscopy Flashcards by Priya Brana

overall function of chromatography

Used to separate individual components from a mixture of substances

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practical uses of chromatography

analysis of drugs, plastics, flavourings, air samples and forensic science

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stationary phase in TLC

solid silica (SiO2) or alumina (Al2O3) held on an inert support material (glass/plastic)

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mobile phase of TLC

Liquid solvent

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advantages of TLC

Quick

inexpensive

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how does separation occur in TLC

via relative adsorption

Components in the mixture adsorb differently to the surface of the stationary phase which result in different rf values

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outline the method for TLC

Draw a base line approx. 1 cm from the bottom in pencil

Using a capillary tube, spot small sample of solution to be tested onto the baseline

Pour some solvent into the chromatography tank.

Ensure the depth is lower than the pencil line.

Place the TLC plate into the solvent, ensuring the solvent does not cover the spot, and that the edges of the TLC plate do not touch the sides of the tank.

Place a lid on the solvent tank.

Allow the solvent to rise until it is about 1 cm below the top of the plate

Remove the plate, marking the solvent level immediately with pencil

Allow the plate to dry

Circle any visible spots.

may need to hold a UV lamp over the TLC plate or spray the plate with a locating agent to show the position of the spots

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compare TLC and paper chromatography

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rf formula

distance moved by component / distance moved by solvent front

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limitations of TLC (4)

Difficult to measure the exact centre of the component ‘spot’

Similar compounds have similar Rf values – difficult to tell them apart

Difficult to find a solvent which dissolves all components in a sample

may not be a reference chromatogram that exists in the database

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overall function of gas chromatography

Separate volatile components in a mixture

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what types of chemicals can gas chromatography analyse

components with low boiling points

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stationary phase of gas chromatography

high boiling liquid adsorbed onto an inert solid support

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mobile phase of gas chromatography

inert carrier gas

e.g. helium / neon

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how does separation occur in gas chromatography

Separation via relative solubility

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what is retention time

time taken for each component to travel through the column

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what are the conditions for adsorption and solubility

Adsorption = solid stationary phase = TLC

Solubility = liquid stationary phase = gas chromatography

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explain separation by adsorption

component molecules bind to surface of stationary phase

stronger the adsorption, more the component molecules are slowed down

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explain separation by relative solubility

components dissolve in liquid stationary phase

the greater the solubility in the stationary phase, the more the component molecules are slowed down

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outline the method for gas chromatography

A small amount of volatile mixture is injected into the gas chromatograph

inert carrier gas carries the components in the sample through the capillary column -contains the liquid stationary phase adsorbed onto the solid support.

components slow down as they interact with the liquid stationary phase

4 reach the detector at different times depending on their interactions with the stationary phase in the column

Component that is in the column for the shortest time has the lowest retention time + detected first

what information does a gas chromatogram give us

each component = detected as a peak

Retention time – used to identify the components present in the sample by comparing these to known components.

Peak integration – can be used to determine the concentrations of components in the sample

(area under the peak)

formula for percentage of component in a mixture

how to plot a calibration curve for concentration of substance against peak area

Prepare standard solutions with known concentrations of the compound being investigated.

Run gas chromatograms for each standard solution.

Plot a calibration curve of peak area against concentration

Run a gas chromatogram of sample.

Use the calibration curve to measure the concentration of the compound.

limitations of gas chromatography

Compounds may have the same retention times

Small amount of one component may hide behind a component with a higher concentration

Unknown components will not have any retention time data to compare it to

test + positive observation for alkenes

add bromine water drop-wise should decolourise from orange to yellow

test + positive observation for haloalkane

add silver nitrate + ethanol and warm to 50°C in water bath chloroalkane - white precipitate bromoalkane - cream precipitate iodoalkane - yellow precipitate

test + positive observation for carbonyls

add 2,4 DNP orange precipitate

test + positive observation for aldehyde

add tollens reagent + warm silver mirror

test + positive observation for primary / secondary alcohol and aldehyde

add acidified potassium dichromate + warm in a water bath colour change from orange to green

test + positive observation for carboxylic acids

add aqueous sodium carbonate effervescence

property of a nucleus

has nuclear spin which is ONLY significant if there is odd number of nucleons (particles in the nucleus) E.g. H1 and C13

how does NMR work

Uses strong magnetic field strength + radio frequency radiation Causes nuclei of some atoms to absorb radiation + flip between spin states = resonance Energy absorbed measured + recorded

describe the spectrometer used

strong superconducting electromagnet Cooled by liquid helium

what is TMS used for

standard reference chemical which all chemical shifts are measured against

what ppm does TMS have

why do we use TMS

Chemically unreactive 1 carbon environment – one peak Nuclei heavily shielded – rarely peaks below it Volatile + easily removed from sample after

preparation of sample for NMR

dissolve in a solvent

what is the issue with solvents for NMR + how do we overcome this

Most solvents have H1 and C13 would produce a peak Must use deuterated solvent – all 1H replaced with 2H Produces no NMR signal in frequency range

what is the solvent used for NMR

Deuterated solvents CDCl3 D is an isotope of H with 2 neutrons

describe the method of NMR

Sample dissolved in deuterated solvents + put into NMR sample tube Tube inside spectrometer + spun to even out imperfections in magnetic field Spectrometer zeroed against TMS standard Given pulse of radiation whilst maintaining constant magnetic field Absorptions of energy from resonance are detected

what information does carbon NMR provide

Number of different carbon environments = number of peaks Types of carbon environment present = chemical shift

how to identify carbon environments

If two atoms symmetrical – equivalent + have same chemical environment Carbon environment with higher chemical shift takes priority

what information does proton NMR decide

Number of different proton environments Types of proton environments Relative number of each type of proton Number of non-equivalent protons adjacent to a given proton

what happens if 2+ protons are equivalent

absorb at same chemical shift + increase size of peak

what are splitting patterns caused by

Caused by protons spin interacting with the spin of nearby protons that are in a different environment

what is the rule of splitting patterns

Number of peaks is 1 more than number of adjacent protons causing the splitting e.g. 4 peaks (quartet) means 3 protons in adjacent environment

how to identify O-H and N-H specifically

proton exchange

describe the process of proton exchange and the end result

Add small volume of deuterium oxide - D2O Shake mixture Run NMR as normal Replaces the OH and NH protons with deuterium atoms The peak disappears