Spectroscopy and Chromatography Boardwork

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Transcript Spectroscopy and Chromatography Boardwork

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Mass spectrometry Mass spectrometry

is an analytical technique that can be used to deduce the

molecular formula

of an unknown compound.

Gaseous molecules of the compound are bombarded with high-speed electrons from an electron gun. These knock out an electron from some of the molecules, creating

molecular ions

(

M +

), which travel to the detector plates:

M (g) + e -



M + (g) + 2e -

The relative abundances of the detected ions form a

mass spectrum

: a kind of molecular fingerprint that can be identified by computer using a spectral database.

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The molecular ion peak

The peak with the highest

mass-to-charge ratio

(

m/z

) is formed by the heaviest ion that passes through the spectrometer. This value of m/z is equal to the

relative molecular mass

of the compound.

100 80 60 40 20

molecular ion peak

High resolution mass spectrometry can be used to determine the molecular formula of a compound from the accurate mass of the molecular ion.

0 0 4 of 45 40 80 120

mass spectrum of paracetamol

160

m/z

What is fragmentation?

A molecular ion is a positively-charged ion, which is also a radical as it contains a single unpaired electron. It is therefore sometimes represented as

M +•

During mass spectroscopy, the molecular ion can

fragment

into a positive ion and a radical:

M +• → X + + Y •

For example, in the case of propane:

CH 3 CH 2 CH 3 +• → CH 3 CH 2 + + CH 3 •

CH 3 CH 2 CH 3 +• → CH 3 CH 2 • + CH 3 +

NB: Only the

ions

are detected by the mass spectrometer. This fragmentation process gives rise to characteristic peaks on a mass spectrum that can give information about the

structure

of the molecule.

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Fragmentation and molecular structure

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Fragmentation of carbonyl compounds

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Interpreting mass spectra

Origins of some common peaks in mass spectra:

methyl positive ion (15) loss of a methyl ion phenyl positive ion (77)

Mass differences

between

peaks indicate the loss of groups of atoms (fragments). For example, loss of a methyl group leads to a mass difference of 15 between peaks.

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Interpreting mass spectra activity

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Uses of mass spectrometry

Some uses of mass spectrometry:  Identifying elements in new or foreign substances, for example analysing samples from the Mars space probe.

 Monitoring levels of environmental pollution, for example amounts of lead or pesticide in a sample.

 In biochemical research, for example determining the composition of a protein by comparing it against a database of known compounds.

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Infrared spectroscopy Infrared spectroscopy

is an analytical technique that provides information about the a compound.

functional groups

present in Certain groups of atoms absorb characteristic frequencies of infrared radiation as the bonds between them undergo transitions between different

vibrational

energy levels. Infrared energy is only transferred to a bond if the bond contains a dipole that changes as it vibrates. Symmetrical molecules such as O 2 or H 2 , are therefore IR inactive.

bonds absorb IR energy

The particular wavelengths absorbed are specific to that particular configuration of bonds and atoms (functional group).

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Infrared spectra

infrared spectrum

is a plot of transmission of infrared radiation against

wavenumber

(1 / wavelength). Any wavelength that is absorbed by the sample will transmit less than the others, forming a dip in the graph.

The pattern in the

fingerprint region

(1500-400 cm -1 ) is unique to each molecule, and so can be used for identification purposes.

wavenumber (cm -1 ) IR absorption spectrum for chloroethane

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IR spectra of different functional groups

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Interpreting IR spectra activity

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Uses of IR spectroscopy

Some uses of infrared spectroscopy:  Use of IR spectra to follow the progress of a reaction involving change of functional groups (e.g. in the chemical industry to determine the extent of the reaction).

 Use of IR spectra to assess the purity of a compound.



Breathalyzers

: modern breathalyzers calculate the percentage of ethanol in the breath by looking at the size of the absorption caused by the C –H bond stretch in the alcohol.

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How does NMR spectroscopy work?

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What does an NMR spectrum tell us?

Different

chemical environments

(bonds and atoms surrounding a nucleus) affect the strength of magnetic field that must be applied to a nucleus in order for it to enter the resonance state.

By measuring the strength of magnetic field that must be applied, NMR spectroscopy gives us information about the

local environment

of specific atoms in a molecule. This can be used to deduce information about molecular structure.

The environments of 13 C and 1 H atoms are most commonly studied in NMR spectroscopy.

Carbon-13 NMR spectroscopy

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Interpreting 13 C NMR spectra

The 13 C NMR spectrum of ethylamine contains two peaks. This is because ethylamine has two unique 13 C environments, each requiring the application of a different magnetic field strength for that carbon nucleus to enter the resonance state. H H H C H C H N H H One peak is due to the carbon atom with three hydrogen atoms attached to it, and the second to the carbon atom with two hydrogen atoms and an amine group attached to it.

Interpreting 13 C NMR spectra activity

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Chemical shift and TMS

The horizontal scale on an NMR spectrum represents

chemical shift

(

). Chemical shift is measured in

parts per million

(

ppm

) of the magnetic field strength needed for resonance in a reference chemical called

TMS

H TMS (tetramethylsilane) is universally used as the reference compound for NMR as its methyl H H C H H groups are particularly well shielded and so it produces a strong, single peak at the far right H C H H Si C C H H H of an NMR spectrum.

H The signal from the carbon atoms in TMS is defined as having a chemical shift of 0. © Boardworks Ltd 2010 23 of 45

13 C NMR chemical shift assignment

The chemical shift values of peaks on the 13 C NMR spectrum can help us identify the types of carbon atom in a compound. The likely source of spectrum peaks can be identified using a data table of typical chemical shift values.

Type of carbon δ/ppm

5 –40 20 –50 190 –220

chemical shift

13 C NMR chemical shift activity

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Proton NMR spectroscopy

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Interpreting 1 H NMR spectra activity

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Integration and the number of hydrogens

The height of the peaks in an NMR spectrum does not give us any useful information.

However, the

area under the peaks

on a 1 H NMR spectrum is proportional to the number of hydrogen atoms causing the signal. The ratio of the areas under the peaks tells you the ratio of 1 H atoms in each environment.

2 1 3

The spectrum can be

integrated

to find this information.

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Spin coupling

Splitting pattern activity

1 H NMR chemical shift assignment

The chemical shift values of peaks on an 1 H NMR spectrum give information about the likely types of proton environment in a compound.

Type of proton δ/ppm

0.7

–1.2

2.1

–2.6

9.0

–10.0

1 H NMR chemical shift assignment activity

Uses of NMR spectroscopy

NMR spectroscopy uses the same technology as

magnetic resonance imaging

(MRI). This is an important non-invasive method of gaining information about internal structures in the body used in diagnostic medicine and scientific research.

NMR spectroscopy is also used in the pharmaceutical industry to check the purity of compounds.

Often, a combination of mass spectrometry, infrared spectroscopy and NMR spectroscopy is used in modern analysis to elucidate the structure of organic molecules.

What is chromatography?

Chromatography

is a series of analytical techniques that can be used to separate mixtures of compounds for further use or for analysis.

In all forms of chromatography, a

mobile phase

moves through or across a

stationary phase



stationary phase

– this phase does not move. Compounds in the mixture are attracted to it (

adsorbed

) and slowed down. Either a solid or a liquid.



mobile phase

– this phase moves. The more soluble compounds in the mixture are carried faster as the mobile phase moves. Either a liquid or a gas.

Thin layer chromatography

Column chromatography

Gas –liquid chromatography

High performance liquid chromatography High performance liquid chromatography

(

HPLC

) is a development of column chromatography in which the eluent is pumped through the column at

high pressure

eluent reservoir data analysis pump injector column

This results in better and faster separation than can be achieved in standard column chromatography.

detector components collected

Gas chromatography –mass spectroscopy

In both GL chromatography and HPLC, the output from the chromatography column can be passed through a mass spectrometer. The spectra obtained can be compared to spectra of known compounds.

Gas chromatography – mass spectroscopy

(

GC –MS

) is used extensively in forensics, environmental monitoring and in airport security systems. It is sensitive enough to detect minute quantities of substances © Boardworks Ltd 2010 40 of 45

Chromatography: true or false?

Glossary

What’s the keyword?

Multiple-choice quiz