Mass Spec 5&6 (Flo-3)

Question 1

What are the two reasons for derivatisation?

Answer 1

1 - make molecule more volatile

2- direct fragmentation

Question 2

Why do hydrophobic molecules work better in MS?

Answer 2

They are easier to ionise because they cannot form H-bonds are so are more volatile

Question 3

Give an example of derivatisation to form a hydrophobic molecule.

Answer 3

trimethysiylation, addition of a S(CH3) group to an -OH group (O-S(CH3))

Question 4

What general equation demonstrates radical elimination (radical ion eliminating radical molecules)?

Answer 4

A+. => B+ + C.

Question 5

What general equation demonstrates neutral elimination (radical ion eliminating neutral molecules)?

Answer 5

A+. => B+. + C

Question 6

Are neutral/radical species detected?

Answer 6

No, only charged species are detected

Question 7

What is the even electron rule?

Answer 7

An even electron ion will not normally fragment to two odd electron species (two radicals)

Question 8

Is radical elimination described as

a) one electron movement
b) two electron movement

Answer 8

a) one electron movement

Question 9

Is neutral elimination described as

a) one electron movement
b) two electron movement

Answer 9

b) two electron movement

Question 10

In radical elimination what is alpha cleavage?

Answer 10

Cleavage of carbonyl compound after ionisation of oxygen. The odd electron is donated to form a new bond to the adjacent atom and cleavage of the original bond to give:
R-C(=O+)-R –> R-C≡O+ + R.

Question 11

Give an example of beta cleavage?

Answer 11

CH3-CH2-O+-CH3 –> CH2=O+-CH3 + .CH3

+N-CH2-R –> N+=CH2 + R.
Normally, the charged molecule is O/N

Question 12

When does neutral elimination occur?

Answer 12

Occurs when a stable neutral molecule (eg. water) can be readily formed via 1,2 elimination mechanism
(often removal of water)

Question 13

What is inductive cleavage?

Answer 13

Initiation of cleavage by the +ve charge involves attraction of electron pair.
e.g.
R-C(=O+)-R –> R-C.(=O) + R+

Question 14

What is the name of the rearrangement when fragmentation occurs via bond breaking and forming?

Answer 14

McLafferty Rearrangement

Question 15

What are CAD experiments?

Answer 15

Collisional activation decomposition - can select an ion and give it more energy to fragment it.

Question 16

How does MS/MS create fragmentation?

Answer 16

Ions are separated and then selected by the first MA and selected ions are activated by exposure to collision gas (transfer of kinetic energy) in the collision chamber (between two MAs).

Question 17

What gases could be used as collsion gases in MS/MS?

Answer 17

Helium, nitrogen, argon

Question 18

What is the purpose of the second analyser in MS/MS

Answer 18

The second analyser separates based on their m/z ratios to produce a MS/MS spectrum

Question 19

In the triple quadrapole MS/MS what does each QP do?

Answer 19

Q1 - separates molecular ions based on m/z
Q2- ions pass through collision chamber
Q3- resolves fragments based on m/z

Question 20

Why is the MS/MS combination of QP-TOF used?

Answer 20

It is a very powerful tool in terms of sensitivity and resolution.

Question 21

What are the advantages of using ESI instead of MALDI for MS/MS peptide sequencing?

Answer 21

• Multiply charged ions produced which requires less energy to fragment.
• Also majority of impurity derived signals are singly charged.
• Gives better signal to noise ratio if you can distinguish between singly and multiply charged ions.

Question 22

Why do peptides digested with trypsin have a minimum of 2 charges?

Answer 22

N-terminus and C-terminal K or R

Question 23

How are isotope clusters used to identify the charge on multiply charged ions?

Answer 23

The distance between the peaks with all 12C will be one mass lower than with 1 13C.
This mass difference (according to m/z) is divided by the charge. therefore if the difference is 1 the charge is 1. If the difference is 0.5 the charge is 2. If the difference is 0.33 the charge is 3.