Mass spectrometry

Question 1

What are OE ions and EE ions?

Answer 1

OE ions have an odd number of electrons. EE ions have an even number of electrons.

Question 2

How are OE ions formed during ionisation?

Answer 2

M + e^- → M^+● + 2e^-

M +e^- → M^-●

Question 3

How are EE ions formed during ionisation?

Answer 3

M + H⁺ → [M+H]⁺

M + Na⁺ → [M+Na]⁺

M - H⁺ → [M-H]^-

M + Cl^- → [M+Cl]^-

Question 4

How do you identify the molecular ion?

Answer 4

It is the peak with the highest m/z value, ignoring isotopes. It is usually at an even m/z. It is usually an OE ion (radical cation).

Question 5

What happens during electron ionisation?

Answer 5

The molecule is ionised when the energy supplied equals the enthalpy of electron attachment. The ionising energy populates vibrational states within the energy well so ions of various energies are formed. The electron of lowest ionisation energy is removed as it is held less tightly (non-bonding > π > σ).

Question 6

Why are fragment ions formed during ionisation?

Answer 6

At 70eV, there is excess energy so the ion can access higher vibrational levels, which can cause bond cleavage to occur. The weakest bonds in the molecule break to form fragment ions. At 16eV, fragmentation is unlikely.

Question 7

Why are soft ionisation methods used?

Answer 7

They produce ions from non-volatile or unstable molecules. They generate ions with less residual energy, so less fragmentation occurs.

Question 8

What is chemical ionisation?

Answer 8

It uses a reagent gas (CH₄, NH₃, isobutane), which is ionised in the ion source. The ionised reagent gas then transfers a proton to the molecule to be analysed. For example:

CH₄ + e^- → CH₄^+● + 2e^-

CH₄^+● + CH₄ → CH₅⁺ + CH₃^●

CH₅⁺ + M → CH₄ + [M+H]⁺

Proton transfer occurs is the proton affinity (PA) of the analyte is higher than the PA of the reagent gas. The greater the difference between the PA of the reagent gas and the PA of the analyte, the greater the energy transfer on ionisation.

Question 9

What is atmospheric pressure chemical ionisation?

Answer 9

It is ionisation at atmospheric pressure (10⁵Pa). A liquid is introduced into the source. The heater and N₂ gas volatilise and remove the solvent. Ionisation occurs by corona discharge due to the high electric potential on the needle. This generates protonated water clusters, which transfer protons to the molecules to be analysed. A counter current of N₂ gas is also used to prevent any neutral molecules from entering the mass analyser.

Question 10

What are isotopologues?

Answer 10

They are isomers that have different isotope combinations. The relative intensities of their peaks reflect the natural abundances of the isotopes.

Question 11

What are the classifications given to elements to identify them in a mass spectrum?

Answer 11

A is an element with one significant isotope. A+1 is an element with two significant isotopes separated by 1Da. A+2 is an element with at least two significant isotopes separated at least once by 2Da.

Question 12

How do you calculate the abundance of a peak?

Answer 12

For each element in the molecule, multiply the number of times is occurs in the molecule by its relative abundance. Add these values together.

Question 13

What are some common isotope profiles?

Answer 13

2 peaks in a 1:1 ratio, separated by 2Da is characteristic of 1 Br atom. 3 peaks in a 9:6:1 ratio, separated by 2Da is characteristic of 2 Cl atoms. 4 peaks in a 1:4:4:1 ratio is characteristic of 3 Br atoms.4 peaks in a 4:6:4:3 ratio is characteristic of 5 Cl atoms.

Question 14

How do you find the probability of an isotope occurring in a molecule?

Answer 14

Multiply the number of times the element occurs in the molecule by its relative abundance. To find the probability of the isotope occurring twice, square its relative abundance and multiply it by the number of time the element occurs.

Question 15

How do you find the number of carbon atoms in a molecule?

Answer 15

Scale the molecular ion cluster so the A peak becomes 100%. Divide the abundance of the A+1 peak by the relative abundance of ¹³C.

Question 16

How can you estimate for error when calculating the number of carbon atoms?

Answer 16

Calculate the values at +10% and -10% for the abundance of the A+1 peak. Determine if n+1 or n-1 fall within this tolerance. If they do not, then it is a reasonable formula.

Question 17

How do you calculate the degrees of unsaturation?

Answer 17

du (r + db) = x - 0.5y + 0.5z + 1

x = the number of tetravalent atoms

y = the number of monovalent atoms

z = the number of trivalent atoms

An OE ion gives an integer value. An EE ion gives a value ending in 1/2.

Question 18

What is the nitrogen rule?

Answer 18

Nitrogen has an even mass and an odd valence. Compounds with an odd number of N atoms give the molecular ion at an odd m/z. Compounds with an even number of N atoms give the molecular ion at an even m/z.

Question 19

What are the different types of fragmentation?

Answer 19

If one bond is broken, EE ions are formed. For example: M^+● → A⁺ + BC^●

M^+● → A^● + BC⁺ → B + C⁺

If two bonds are broken, rearrangement of the atom’s bonds can occur and OE ions are formed. For example: M^+● → C^+● + D

Question 20

What are the main factors affecting ion abundance?

Answer 20

Cation stability, electron sharing and resonance stabilisation.

Question 21

What is Stevenson’s rule?

Answer 21

The most probable fragment is where the positive charge is on the fragment with the lowest ionisation energy. If ions have similar stability, loss of the largest radical is favoured, which may lead to the loss of the least stable ion.

Question 22

How are alkanes ionised and how do they fragment?

Answer 22

Ionisation of alkanes removes an electron from a σ-bond. Usually the molecular ion is weak, and the charge is not localised at any specific site so it can fragment at many position along the chain. Fragmentation occurs by σ-cleavage (the radical in the σ-bond moves to a carbon atom producing an alkyl cation and an alkyl radical). Secondary fragmentation can occur when the cation produces an alkene and another cation. The ion series is C_nH_2n+1.

Question 23

What is radical site initiation?

Answer 23

The radical site has a strong tendency for electron pairing, which results in a-cleavage. This is homolytic cleavage.

Question 24

How do aliphatc alcohols fragment?

Answer 24

They undergo a-cleavage and the loss of the largest alkyl is favoured die to the radical stability. They can also rearrange to remove water.

Question 25

What are neautral losses?

Answer 25

Neautral species like radicals or molecules can be lost during fragmentation. They are not detected because they are not charged. They are recognised by the difference in m/z between ions. Only a few can be lost by cleavage of one bond.

Question 26

How are alkenes ionised and how do they fragment?

Answer 26

Alkenes are ionised by the loss of a π-electron from a π-bond. They fragment by allylic cleavage, which is similar to a-cleavage, except the radical is on the bond. This forms a positively charged alkene and an alkyl radical. The alkene is stabilised by resonance so it is favourable. Alkenes are in the C_nH_2n-1⁺ ion series, with some C_nH_2n+1.

Question 27

How are aromatic hydrocarbons ionised and how do they fragment?

Answer 27

Aromatic hydrocarbons are ionised by the loss of a π-electron from a π-bond. They fragment by benzylic cleavage, which is made more favourable by the alkyl group. This is because it can form a tropylium cation (a seven membered, fully aromatic structure).

Question 28

What is charge-site initiation?

Answer 28

The positive charge strongly attracts an electron pair, which causes inductive cleavage. This is heterolytic cleavage. It is used for EE ions. For OE ions, inductive cleavgae competes with a-cleavage.

Question 29

How does fragmentation occur in unsaturated heteroatomic molecules?

Answer 29

This includes carbonyls, amides, carboxylic acids and esters. The main driving force is the formation of a stable carbenium (acylium) ion. For aldehydes, hydrogen loss occurs if the acylium ion is stabilised (e.g. by resonance). For ketones, a-cleavage is more important.

Question 30

How does fragmentation occur in cycloalkanes?

Answer 30

If one bond is broken, a stable OE ion is formed. It is a diastonic radical (there is a distance between the charge and the radical). For fragmentation to occur, two bond must be broken.

Question 31

What is the Diels Alder reaction?

Answer 31

It is a cycloaddition reaction between a diene and an alkene to form cyclohexene, using heat. It is the key reaction to forming a six membered ring.

Question 32

What is a retro Diels Alder rearrangement?

Answer 32

It affects structures containing cyclohexene rings.

Question 33

What is McLafferty reaarangment?

Answer 33

It is y-reaarangement and it is important if the transition state is allowed. It is a radical site reaarangement on unsaturated compounds. The y-hydrogen is transferred and B-cleavage occurs. It forms a strong OH bond. The radical in the fragment ion is resonance stabilised.

Question 34

What is double McLafferty rearrangement?

Answer 34

If the unsaturated compound has a long enough chain length on both sides of the unsaturation, then two McLafferty rearrangments can occur in sequence.

Question 35

How does rearrangement occur in cyclohexanone?

Answer 35

McLafferty rearrangement is not possible because the y-hydrigen is held too far away.

Question 36

What are charge-site rearrangements?

Answer 36

Hydrogen rearrangments driven by the charge site are favoured when the hydrogen is transferred to an unstaturated group containing a heteroatom. Stabilisation by resonance can lead to this being favourable. For example, the elimination of a ketene from phenolates is fvourable as it produces a resonance stabilised OE ion.

Question 37

What are common difficulties when interpreting a mass spectrum?

Answer 37

The molecular ion might be missing, or the molecule might be above the instrument mass range. The mass spectrum can be too complex with many similar isomers, or it can be too simple (e.g. with soft ionisation). Even with couple chromatographic separation (e.g. GC or HPLC), the components can co-elute.

Question 38

How can mass spectrometry be used for macromolecules?

Answer 38

Macromolecules can have masses beyond the normal range of an instrument, but they can be anaylsed by ESI as they form multiply charged ionised molecular species. This gives infomration about mass. Selected enzymatic cleavage can be used to get structural information.

Question 39

How do you find the position of unstaurated bonds?

Answer 39

Locating the position of double or triple bonds is hard because of migration. To locate them a label is introduced at the position of the bond. This can be done by reduction with a deuteriated reagent or by reaction with a sulfide. In the isotope method, they are labelled with D₂ or Wilkinson’s catalyst. Double bonds add two D and triple bonds add 4D. In the reaction method, the bond is fixed with dimethyl sulfie/ I₂. This forms 1,2-dithiane and the molecule fragment between the two thioether groups.

Question 40

What is tandem mass spectrometry?

Answer 40

It is performed using specialised mass spectrometers that can perform more than one stage of mass analysis (e.g. triple quaprupole mass spectrometer). In this, the first and last quadrupole perform the mass analysis. The collisions between the ions and the gas energise the ions (collision induced dissociation).

Question 41

What are the scans used in tandem mass spectrometry?

Answer 41

Product ions are those formed by dissociation. The product ion scan fixes Q1 on the precursor m/z, dissociates in Q2 and scans in Q3 to generate a sepctrum of the product ions. This gives a spectrum for species that do not produce fragment ions. Precursor ions are the tandem MS ions. The precursor ion scan scans in Q1, dissociates in Q2 and fixes Q3 on the m/z for a specific product ion. The signal appears only when the correct percursor ion is dissociated. The neutral loss scan scans in Q1 and Q3 with a constant m/z offset, and fragments in Q2. This allows recogonition of neutral losses.

Question 42

What are GDGTs?

Answer 42

Glycerol dialkyl glycerol tetraethers contain 2 hydrocarbons etherfied to 2 glycerol groups. They have between 0 and 8 cyclopentyl rings. They are large, non-volatile structures so are introduced in liquid form (LC-MS with an APCI source).It has an odd mass but does not contain nitrogen.