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Maas spectroscopy

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MASS SPECTROMETRY
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Page 1: Maas spectroscopy

MASS SPECTROMETRY

Page 2: Maas spectroscopy

Index

Introduction

Principle

General Instrumentation

=> Sample Introduction (Inlet)

{A} Ionization

1. Electron Ionization

2. Electro Spray

3. Matrix – Assisted Laser Desorption/ Ionization (MALDI)

{B} Mass Analyzers

1. Magnetic Sector Analyzer

2. The Quadrupole Analyzer

3. Time of Flight Analyzer

{C} Detectors

1. Faraday cup

2. Electron Multiplier

3. Photomultiplier Conversion Dynode

Applications of Mass Spectrometery

Reference

Page 3: Maas spectroscopy

• MS is powerful analytical

technique.

• Identify unknown compounds.

• Quantify known materials.

• Elucidation of structural and

chemical properties.

•Requires minute quantities

(<pg).

• High sensitivity , selectivity

and specificity – provides

valuable information in various

branches of science.

Introduction

Mass Spectrometer is an instrumental technique in which, sample is

converted to rapidly moving positive ions by electron bombardment and

charged particles are separated according to their masses.

Page 4: Maas spectroscopy

PRINCIPLE

Principal based on:-

1. First event is to ionize the molecule.

2. Ions move into mass analyzer where the ions are differentiated according to their mass to charge ( m/z ) ratio.

3. The differentiated ion beams now fall onto the detector , which measures the ion beam current.

Page 5: Maas spectroscopy

General Insrumentation

A sample introduction system (inlet system)

1. An ion source,

2. A mass analyzer and

3. A detector

Page 6: Maas spectroscopy

Sample Introduction

To introduce a sample two things must be achieved here:-

• One is that the sample must be introduced into vacuum

• Secondly , the sample must be vaporized prior to ionization, so that proper

ionization take place.

To introduce sample one approach is by placing a sample on a probe, which is than

inserted through a vacuum lock into the ionization region of MS.

Another method of introducing the sample is capillary infusion.

This delivers small quantities of sample to the ionization chamber without

disturbing the vacuum.

The capillary can be a column from a gas /liquid chromatography.

Page 7: Maas spectroscopy

Ionization

Ionization means placing a charge on an otherwise neutral molecule.

Several different methods :-

1.Electron Ionization

2.Electro spray Ionization

3.Matrix- Assisted Laser Desorption/ Ionization (MALDI)

Page 8: Maas spectroscopy

Electron Impact (EI)

•The original mass spectrometry (MS) ionization method

•The most widely used

•Used for ionization and fragmentation the sample molecules before mass analysis

•Firstly the sample is vaporized and then enters into the ion source

•A beam of electrons help the sample be impacted with sufficient

energy to ionize the molecule

Page 9: Maas spectroscopy

Electrospray Ionization (ESI)

• Soft ionization technique, desorption ionization method

• Proteins, peptides, and other biological macromolecules

• No fragmentation of the macromolecules into smaller charged particles; turning the

macromolecule being ionized into small droplets

• Large mass molecules are detected in the small mass range of the instrument

•As the m/z value increases, the number of protons attached to the molecular ion decreases

Page 10: Maas spectroscopy

Matrix-Assisted Laser Desorption Ionization (MALDI)

• A laser-based soft ionization method

• Important for protein analysis.

• Ionization occurs with bombarding the sample with laser light

• The sample is embedded in a chemical matrix (UV absorbant)

• The matrix makes the production of intact gas-phase ions from large, nonvolatile, and

thermally decomposed compounds such as proteins, oligonucleotides, synthetic polymers

easy

• The matrix absorbs the laser light energy thus small amount is vaporized

Page 11: Maas spectroscopy

Mass Analyzer

Mass Analyzer is a region of the mass spectrometer is used to separate

ions within a selected range of mass – to – Charge (M/Z) ratio.

To separate ions different method used:-

1.Magnetic Sector Analyzer

2.The Quadrupole Analyzer

3.Time of Flight Analyzer

Page 12: Maas spectroscopy

Magnetic Sector Analyzer

• J.J. Thompson (1897), built the mass spectrometer , used a magnet to measure the

M/Z value .

• Magnetic Sector instrument used today evolved from this concept.

• Separates ions in a magnetic field according to the momentum & charge of the ion.

• An electric field (1-10 kv) accelerates ions

from source region intomagnetic sector.

• When it reaches magnetic field ,the ion

beam bent in an arc by the

magnetic field.

• Greater the momentum of ions, larger

their arc radius.

• Instrument have a set of slites at a fixed radius, transmitt a single M/Z to detector.

Page 13: Maas spectroscopy

Time-of-Flight Analyzer

• Used with the MALDI technique for analysis of large biomolecules

• All ions have same kinetic energy.

• Thus having same kinetic energy leads different velocities depending on their masses

• Masses affect their arrival time at the detector, smaller ions will reach the detector first

due to they have greater velocity and vise versa.

• Since, time of reaching the detector is of essense in this analyzer, hence name given.

• Arrival time of an ion at the detector is based upon mass, charge & kinetic energy of the

ion.

Page 14: Maas spectroscopy

Quadrupole Analyzer

• An electric field accelarates ions

into the quadrupole analyzer.

• Consist of 4 rodes/ electrodes

arranged across from each other.

• Ions made to travels through

the Quadrupole.

• They get filtered according to their M/Z ratio.

• M/Z ratio is based upon the radio frequency & direct current voltages applied to

these electrodes.

• These voltage produce an oscillating electric field ,transmitt ions according to their

M/Z value.

Page 15: Maas spectroscopy

MS Detector

Detector generates a signal current from incident ions by generating secondary

electrones which are further amplified.

Most commenly used detectors are :-

1.Faraday Cup

2.Electron Multiplier

3.Photomultiplier Conversion Dynode

Page 16: Maas spectroscopy

Faraday Cup

• A change in charge on a metal plate results in a flow of electrons. The flow creates a

current. This is the concept on which faraday cup detector operates.

• When a single ion strike the surface of a dynode in the faraday cup, it results in

ejection of several electrons.

• This ejection of electrons induces a current in the cup.

• A small amplification may also results because of secondary electrons.

Page 17: Maas spectroscopy

Electron Multiplier

•Concept same as faraday cup.

•Only difference is that faraday cup uses a

single dynode, an electron multiplier uses a

series of dynodes maintained at successively

higher potential.

•Electrons released by the first dynode when

the ion strike on it are dragged to second

dynode because it has a higher potential.

•These primary electrons strike the second

dynode with a force and the collision releases

even more secondary electrons which too are

dragged and so on.

•Typical amplification of an electron multiplier

is one million.

Page 18: Maas spectroscopy

Photomultiplier

•Similar to electron multiplier in that the ion strike and resulting in the emission of

electrons.

•These electrons now made to strike on a phosphorus screen. This screen releases

photons when electron strike it.

•This photons are now detected by a photomultiplier.

•Photomultiplier tube housed in vacuum , removes the possibility of any

contamination from the internal environment.

•It has a higher life for this reason this detector are becoming more popular

Page 19: Maas spectroscopy

Applications of MS

• Protein characterization.

• Peptide mass finger printing and protein identification.

• Applications in virology.

• Sequencing peptide and oligonucleotides.

• Determination of higher order protein structure.

• Analysis of biological noncovalant complexes.

Page 20: Maas spectroscopy

REFERENCES

Avinash Upadhyay, Himalaya publishing house, Biophysical Chemistry, Fourth edition, chapter 15, Mass spectrometry, pg no. 576-585.

http://www.google.com

http://www.wikipedia.com

Page 21: Maas spectroscopy

.

Thank you


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