NMR + chromatography Flashcards
2 types of NMR
- 13C NMR
- 1H (proton) NMR
number of signals = methylpropene
2
number of signals in methylamine
2
number of signals in propene
CH2=CH-CH3
3
calibration
-small quantity of TMS (tetramethylsilane) is added to samples as it produces a single providing internal standard to which other peaks are compared
-only givies one single, non-toxic, inert, low boiling point
NMR
-number of peaks in an NMR spectrum refers to Hydrogen atoms
-area under the peak = proportional to the number of hydrogen atoms
Chemical shift = closer the hydrogen is to an electronegative atom the greater the shift
why compounds absorb radiowaves
-the nucleus of some atoms have a nuclear spins e.g 1H, 13C, 19F etc)
-a nucleus with spin generates a small magnetic field
-there is a small difference in energy ebtween these 2 alignments that corresponds to the energy of radiowaves
how NMR is done
-the sample is usually dissolved in a solvent in a long thin tube and placed inside the machine where it is bombarded with radiowaves in a strong magnetic field
-the stronger the magnetic field the better the resolution of the signals
solvents
-if the solvent contains any 1H atoms then these will appear in the spectrum
-common solvents used = CCl4
chemical shifts
-the chemical shift depends on what other atoms/groups are near the H. The closer the H to electronegative atoms the greater the shift.
scales
-horizontal scale = chemical shift
-vertical axis = intensity of the absorption
signals
-same distance of CH3 = 1 signal. Difference = multiple signals
-in 1 H NMR the area of the signal is proportional to the number of hydrogen atoms is represents
-the height of the signal doesn’t matter but rather the area which the signal covers.
At A-level to indicate the relative intensity of the signals from which the simplest whole number ratio can be calculated:
e.g 1.2 : 1.2 : 1.8 = 2:2: 3
calculating signals
1) Divide all values with the smallest number
2) Find whole number
splitting patterns
-the number of lines in a signal is split into and gives information about neighbouring H atoms
-usually only H atoms on the neighbouring C atom cause splitting
-number of lines = 1 + number of H atoms on adjacent carbon atoms (n+1)
-H atom on OH group in alcohols rarely causes splitting or is split itself.
Doublet = 1:1
Triplet = 1:2:1
Quartllet = 1: 3: 3: 1
outline the steps needed to locate positions of the amino acid on TLC plate and determine their RF values
-spray with developing agent
-measure distances from initial pencil line to spots of pigment (x)
-measure distance from initial pencil line to solvent front (Y)
-Rf value = X / Y
why do amino acids have different Rf values
amino acids have different polarities
-different solubilities in the developing solvent (mobile phase) and retention values in the stationary phase
solid vs liquid stationary phase
A solid stationary phase separates by adsorption,
A liquid stationary phase separates by relative solubility
polar vs non-polar stationary phase
If the stationary phase was polar and the moving
phase was non- polar e.g. hexane. Then non-polar
compounds would pass through the column more
quickly than polar compounds as they would have
a greater solubility in the non-polar moving phase.
Precuations for chromotaography
Wear plastic gloves to prevent contamination
from the hands to the plate
pencil line –will not dissolve in the solvent
tiny drop – too big a drop will cause different
spots to merge
Depth of solvent– if the solvent is too deep it
will dissolve the sample spots from the plate
Will get more accurate results if the solvent is
allowed to rise to near the top of the plate but
the Rf value can be calculated if the solvent
front does not reach the top of the plate
lid– to prevent evaporation of toxic solvent
dry in a fume cupboard as the solvent is toxic
UV lamp used if the spots are colourless and
not visible
Practical procedure for TLC
a) Wearing gloves, draw a pencil line 1 cm above the
bottom of a TLC plate and mark spots for each sample,
equally spaced along line. b) Use a capillary tube to add a tiny drop of each solution to a
different spot and allow the plate to air dry.
c) Add solvent to a chamber or large beaker with a lid so that
is no more than 1cm in depth
d) Place the TLC plate into the chamber, making sure that
the level of the solvent is below the pencil line. Replace
the lid to get a tight seal.
e) When the level of the solvent reaches about 1 cm from
the top of the plate, remove the plate and mark the solvent
level with a pencil. Allow the plate to dry in the fume
cupboard.
f) Place the plate under a UV lamp in order to see the spots.
Draw around them lightly in pencil.
g) Calculate the Rf values of the observed spots.
procedure for column chromatography
- A glass tube is filled with the stationary phase usually silica
or alumina in powder form to increase the surface area. * A filter or plug is used to retain the solid in the tube.
Solvent is added to cover all the powder. * The mixture to be analysed is dissolved in a minimum of a
solvent and added to the column. * A solvent or mixture of solvents is then run through the
column. * The time for each component in the mixture to reach the
end of the column is recorded (retention time)
types of chromatography
-Thin layer chromatography = plate is coated with a solid and a solvent moves up the plate
-Column chromatography –> column is placed with a solid and solvent moves down the column
-Gas chromotography –> gas is passed through column under pressure at high temperature
separation dependency
Separation depends on a balance between solubility in moving phase and retention in stationary phase
mobile phase
Mobile phase = carries soluble component (more soluble = moves faster)
stationary phase
Stationary phase = phase which holds back the component (more affinity = moves slower)
retention time
Retention time = how long it takes to travel through the chromatogram. Depends on affinity for mobile and stationary phase
Rf value = between 0 and 1
equation for retention values
Rf = distance moved by component / distance moved by solvent
most polar =
ionic substances = NaCl
what does retention time depend on
Retention time depends on the polarity of a substance
The stationary phase attracts polar substances = longer retention time as the substance moves slower
detection
Amino acids can be stained using ninhydrin
Colourless samples can be viewed using UV light
phases
-Mobile phase = solvent that moves up the plate = solvent soaks up the TLC plate and dissolves the compounds at the baseline.
-Separation occurs based on different attractions to the stationary and mobile phase
-Adsorption = compound temporarily binds to silica gel surface
-more strongly absorbed = less distance it will travel
factors affecting separation
-higher solubility = faster movement up the plate
-stronger adsorption = slower movement e.g compound forming hydrogen bonds with silica gel will move slower than van der waals
column chromatography
-tube is usually filled with silica or aluminia in powder form to increase surface area
-used for separating mixtures on a larger scale
-stationary phase = silica gel
-mobile phase = liquid solvent
-chromatography column is filled with silica gel
-concentrated sample solution is added at the top
-solvent is added continiously to wash compounds through the column
-more polar compounds interact more with the stationary phase = move slower
Gas chromatography
-uses an inert gas e.g He,H2,N2
-long coiled tube placed in powder
-samples analysed using mass spec
-mobile phase = gas solvent
-stationary phase = high boiling point liquid adsorbed on a solid
-used for analysing small volatile compounds
1) Tiny sample is injected with a syringe
2) sample vaporises instantly in a heated injector
3) sample travels through a coiled column coated with stationary phase liquid
4) separation depends on boiling point and solubility in the stationary phase
5) results appear as peaks on a chromatogram
separation is too slow
-if separation is too slow, use more polar solvent not compound
factors affecting retention time
-higher boiling point = longer retention time
-solubility in the stationary phase = more soluble = longer retention times
-higher temperature = shorter retention time
gas chromatogram
-peak position = retention time to identify the compound
-peak area = proportional to the amount of compound present
when calculating Rf values
measure up to the middle (midpoint) of the spot not the top
ethanol forms hydrogen bond with stationary phase =
adsorbs strongly = longer retention time
wider + tall =
high abundance = chromatogram
each peak =
each environment that absorbs specific amount of energy
1 peak = 1 environment
e.g ethene
maximum environments in carbon 13 NMR
depends on the carbon atoms present
butane =
2 peaks in C13 due to equivalent
C13 for chlorobenzene
4 environments
1H for chlorobenzene
3 environments
polar sample =
dissolve in a polar solvent
non-polar solvent
tetrachloromethane
CCl4
polar solvent
CDCl3
tetramethylsilane
Si(CH3)4
-only gives a single peak
-far away from peaks produced by the sample
-inert = wont react with sample
-non-toxic
-highly volatile = remove from sample easily
larger chemical shift =
closer to an electronegative atom
turn decimal ratio into whole number ratio =
multiply all ratios by the smallest value
integer ratio =
adds up to show all hydrogen atoms in the molecule
splitting pattern in high resolution NMR
singlet = single line/spike
doublet = 2 lines/spike split into 2
etc
triplet, quartet, multiplet
n + 1
n = number of non-equivlaent hydrogen atoms
n =1 hydrogen = doublet forms
splitting only occurs
for hydrogen atoms on carbon atoms
e.g hydrogen in OH group = singlet + NH2
example = propanoic acid
3 peaks = 3 environments = types of non-equivalent 1H
integration ratio = 1:2:3
O-H = singlet
CH3 = triplet as split by adjacent CH2
CH2 = quartlet
no splitting =
singlet
triplet next to a quartet =
CH3 - CH2
two triplets =
CH2-CH2
number of environments =
number of peaks
tetramethylsilane formula
Si(CH3)4
CHCl4 not used for 1H
would show hydrogen peak
any ratio with 9 =
3 CH3 groups surrounding a carbon atom
ratio based on number of hydrogens
e.g 3 : 3 : 2
3 hydrogens , 3 hydrogens, 2 hydrogens
largest energy gap =
fewest electrons
carbon 12 nuclei
dont have spin
stronger magnets =
higher frequencies interact with the same sample
higher peak
more molecules in the same environment
never use peak heights in C13 NMR analysis
3 rules for hydrogen NMR
-hydrogens bonded to the same carbon are in the same environment
-hydrogens bonded to carbons in the same environment are in the same environment
-
hydrogens on double bond
not necessarily the same environment due to restricted rotation
need to look at groups on the opposite side of the double bond
