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MSF2 -Mass Spec > Mass Spec 1&2 (Flo-1) > Flashcards

Mass Spec 1&2 (Flo-1) Flashcards

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1
Q

Who showed that electron had a m/z ratio?

Briefly outline his method.

A

Sir J.J Thompson - used cathode ray tubes to show that electrons were deflected by magnetic and electric fields relative to m/z.

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2
Q

What are the two properties that make mass spec so powerful?

A
  • Sensitivity (structural information from femtomoles)

- Can handle complex mixtures of sample (no purification required)

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3
Q

What are the 3 basic arts of a MSR?

A
  • Ion source
  • Mass analyser
  • Detector
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4
Q

What is the best way to visualise data?

A

Spectrum

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5
Q

In a mass spectrum what intensity on the graph is given to the highest peak?

A

100% intensity- this can be altered if you want to zoom in on a certain section

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6
Q

What are the 3 types of ion source?

A
  • Electron Impact (EI)
  • Matrix Assisted Laser Desorption ionisation (MALDI)
  • Electrospray Ionisation (ESI)
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7
Q

What are the soft techniques of Ionisation?

A

ES and MALDI

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8
Q

What is meant by a soft ionisation technique?

A

Ionisation which doesn’t result in fragmentation of the molecule.

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9
Q

What mass range can Electron Impact ionise?

A
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10
Q

Why is fragmentation of molecules caused in Electron ionisation?

A

The system provides excess energy to the molecules causing fragmentation. (Bond Energy of average biomolecule = 5eV, electron beam provides 70eV)

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11
Q

What are the two ways of generating gas phase molecules in EI?

A
  • Sample on metal and heat

- Couple MSR to gas chromatographer

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12
Q

How does EI generate ions from gas phase molecules?

A

Stream of electron beam focused by magnets come into close proximity with sample, repelling electrons from the molecules.

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13
Q

Specifically why does EI cause fragmentation of the gas phase molecules?

A

There is a large excess of energy provided. The electron beam has an energy of 70eV (electron volts) and the average biomolecule has 5eV in its bonds.

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14
Q

Where do the electrons in the EI electron beam from?

A

Tungsten or Rhenium

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15
Q

What type of ions are generated by EI?

A

Radical Cation

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16
Q

Which ionisation techniques are performed in vacuum?

A

ALL 3 OF THEM

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17
Q

What is used to direct the cations from EI into the MSR?

A

Strong positive repeller plate

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18
Q

MALDI uses energy pulses of laser to generate gas phase ions. True or False?

A

True

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19
Q

What does the crystalline sample produced in the first step of MALDI consist of?

A

Solid phase biological sample embedded in the matrix

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20
Q

What factor do you have to consider before choosing your matrix in MALDI?

A

The matrix has to have a maximum of absorption near the wavelength of the laser

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21
Q

What type of ions are generated by MALDI?

A

Positive and negative, but these have to be analysed separately.

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22
Q

What is used to direct the ions from MALDI into the MSR?

A

An appropriate repeller plate is used - charge depending on whether you want to analyse anions or cations

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23
Q

What is the modern laser used for MALDI? Why is it better?

A

Solid State UV laser - higher repetition rate (0.5-1Hz)

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24
Q

What is the wavelength of the Solid State UV laser?

A

355nm

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25
What is the older laser used for MALDI? Why is it not as good?
Nitrogen gas UV laser - lower repetition rate (1-20Hz)
26
What is the wavelength of Nitrogen gas UV laser?
337nm
27
What would you use a Alpha-cyano-4-hydroxycinnamin (CHCA) matrix to embed?
Peptides and proteins
28
What would you use a 2,5 dihydroxybenzoic acid (2,5-DHB) matrix to embed?
Carbohydrates
29
What was the problem with MALDI 30/40 years ago?
Ionising from solid phase (3D lattice). Ions near the surface are ionised first and have more energy. More energy ruired to ionise molecules further down. So ions with same mass will have different speeds.
30
What is delayed extraction in MALDI?
Delaying the potential on the repeller plate to have discrete packages of ions fired with the same and m/z ratio?
31
What is the mass limit of MALDI?
High
32
Why is ESI starting liquid phase an advantageous?
Biological matter often exists in liquid state
33
What is placed on the needle capillary (narrow glass capillary) of ESI?
The sample dissolved in a solvent.
34
Why is the thin metal film on the tip of the needle in ESI required?
Required to produce an electrical conductance. Once the needle is put into electrode an electrical potential is generated between the needle (metal film) and the electrode.
35
How are ions created from the biological sample?
Pressure is applied to the needle producing tiny ionised droplets.
36
What type of ions are generated by ESI?
Both positive and negative ions depending on the pH of the droplet and molecule.
37
What is the effect of blowing inert drying gas (nitrogen) over droplets?
The droplets get smaller and smaller, charged species repel causing droplet to explode. This occurs until gas phase ions are produced.
38
What is the Rayleigh limit? FLO NEEDS TO ADD TO NOTES (REMIND FLO)
The point at which ions get expelled from the droplets and become gas phase
39
Does electrospray make singly charged ions?
NO!! they make multi charged ions and so spectra looks different because you get different peaks for the same species dependant on the charge. Computer algorithms can output this into a single peak
40
What is the flow rate of normal ESI?
2.5 microliters/min
41
What is the flow rate of nano ESI?
10-30 nanolitre/min --> it is more sensitive, smaller initial droplets and improved ionisation efficiency
42
What volume solution is required for nano ESI?
1 microliter - adapted needles. Capillary effect is used so no 'plunger' is required to apply pressure. Also --> smaller droplet = less salt = less contamination Fission takes less time
43
What is nanoLC?
Because ESI uses liquid phase it is possible to connect it directly to liquid chromatography - separation techniques.
44
What is Z-SPRAY-nano-ESI?
The most modern technique. The original system had the end of the needle directly in the MSR. The modern system uses an e field to direct the electrons in a Z shape so only correctly charged ions enter the MSR. This reduces contamination.
45
What mass analyser is connected to Z-SPRAY-nano-ESI?
Q-TOF
46
What are the 3 factors
- The Upper Mass Limit - Ion Transmission - Resolution
47
When was the quadrapole first invented?
mid 1950's
48
How many parallel rods are used in the quadapole mass filter?
4 rods, these allow for different e fields and RF to be applied - setting up n electrical field between them
49
What is E field produced by?
Application of a Direct current (DC) to one pair and RF potential to the other pair of poles
50
Some ions will be in harmony with the E field. What happens to these ions?
These ions carry on in a straight line and hit the detector
51
What happen when the ions are not in harmony with the E field?
These will hit the quadrapoles resulting in annihilation
52
You can vary the strength of the QP to get different what?
Different ions (m/z) detected
53
What is the lowest performing Mass analysers?
Quadrapole mass filter
54
What is the upper mass limit of Quadrapole mass filter?
m/z up to ~4000, quite small
55
What is the resolving power of Quadrapole mass filter?
unit resolution ~3000, lowest of all mass analysers
56
What is the ion transmission like in a quadrapole mass filter?
The scanning of the QP means that you are not detecting all the ions hence sensitivity is lowered.
57
Why is the quadrapole mass filter still the most widely used mass analyser?
Cheap, reliable, robust and ideal for GC-MS
58
Describe the structure of the Ion trap mass analyser.
4 metal rods are squashed together to form a small hole between cap electrodes at each end.
59
What generates the E field in the ion trap mass analyser?
RF voltages
60
How do ions enter the ion Trap?
Ions pass through a central chamber via entrance electrode. E field applied is generated by RF voltages.
61
How does the Ion trap analyse ions?
Ions circulate and are trapped in an E field. Altering the E field effects the diameter the ions circulate with (relative to m/z). Some have a wider diameter and have enough energy to exit the chamber. The E field can be altered to sequentially exit all ions.
62
How does the general performance of Ion trap relate to the performance of the quadrapole mass filter?
Slightly improved on all performances of QP
63
What are the issues with the Ion Trap analyser?
- lots of ions required for high sensitivity - too many ins overloads the chamber and it becomes difficult to control circulating pattern (ions many hit each other and rods)
64
What improvements have been made with ion trap analyser?
A linear ion trap has been developed which has a larger volume and so more ions an be trapped and controlled. This means they have a higher sensitivity.
65
What does TOF stand for?
Time of flight analyser
66
What is the principle for how TOF analyses ion m/z?
Ions are accelerated down a metal tube. The time take to leave the source and reach the detector is proportional to the m/z. Ions with larger m/z are slower. All molecules must have the same energy.
67
What is the upper mass limit of TOF?
There is not upper mass limit in theory!
68
What is the transmission of TOF?
100%!!
69
What device is used to increase resolving power of TOF?
The reflectron - like an ion mirrror Ions will leave source at the exact same time with the same energy so m/z should only be proportional to speed but energy is hard to control. reflection helps account for this.
70
How does the reflectron work?
A series of electrodes at the end of the tube. Ions with the same m/z will fly down. If one has more energy it will get closer to the reflectron before it is repelled. This means the ion with more energy will then have to 'catch up' with the other ion. So ions with same m/z will arrive at the detector at the same time
71
What is the highest resolution and sensitivity mass analyser?
Fourier Transform Ion Cyclotron Resonance (FT-ICR) | For these reasons it is very expensive (and not robust)
72
Which mass analyser also doubles as the detector?
- FT-ICR, this is very powerful - has suer conducting magnets and liquid helium cooling (cools magnet to absolute 0) - Oribtrap
73
How does the FT-ICR analyse ions?
Ions pass through metal box which inside a super conducting magnet. Ions circulate and a burst of Rf is introduced. Causes ions to circulate wider and move closer to the bottom and top plates. Because of the charge applied this completes the circuit between the top and bottom plates which is detected. FT Relates RF to m/z
74
Why does FT-ICR have high sensitivity?
No ions are destroyed so MS can be performed infinite times
75
What is the physical set up of the orbitrap?
There is an outside barrel electrode and a spindle electrode running though the centre.
76
How does the orbitrap analyse ions?
Ions are held up by electrostatic forces and spin around and move up and down the central electrode. The rate of up/down movement is related to m/z ratio. FT relates this movement to m/z ratio.
77
What is the general performance of orbitrap like?
It has very high performance. | 2nd highest resolving power and mass accuracy, good upper mass range and good ions transmission.

MSF2 -Mass Spec (3 decks)

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