Gas chromatography mass spectrometry (GC-MS)

Gas chromatography-mass spectrometry (GC-MS) is the synergistic combination of two analytical method to separate and identify different substances within a test sample.

Gas chromatography separates the components of a mixture in time.

Mass spectrometer provides information that aids in the identification and structural elucidation of each component.

1. GAS CHROMATOGRAPHY-MASS SPECTROMETRY (GC-MS) Presented By- Sudipta Nandi M. Pharm 1 st Sem Department of Pharmaceutical Sciences Dr. Harisingh Gour Vishwavidyalaya Sagar, M.P. ( A Central University )

2. Content INTRODUCTION HISTORICAL BACKGROUND OF GC-MS PRINCIPLE INSTRUMENTATION GAS CHROMATOGRAPHY INTERFACE MASS SPECTROMETER WORKING LIMITATIONS APPLICATION CONCLUSION REFERENCES

3. INTRODUCTION Gas chromatography-mass spectrometry (GC-MS) is the synergistic combination of two analytical method to separate and identify different substances within a test sample. Gas chromatography separates the components of a mixture in time. Mass spectrometer provides information that aids in the identification and structural elucidation of each component. HYPHENATED TECHNIQUES

4. Gas Chromatography-Mass Spectrometry (GC-MS) GAS CHROMATOGRAPHY MASS SPECTROMETRY GAS CHROMATOGRAPHY-MASS SPECTROMETRY Separated mixture of chemicals so each can be identified individually Identifies (detects)chemicals based on their molecular weight or mass A Chemical Analysis Technique combining two instruments to provide for powerful separation and identification capabilities

5. Historical Background of GC-MS

6. The sample solution is injected into the GC inlet where it is vaporized and swept onto a chromatographic column by the carrier gas. The sample flows through column and compounds are separated by virtue of their relative interaction with the coating of the column and the carrier gas. Latter part of column passes through a heated transfer line and ends at entrance to ion source where compounds eluting from the column are converted to ions and detected according to their mass to charge m/z ratio. Principle of GC-MS

7. INSTRUMENTATION LAYOUT Fig : Principle of GC-MS

8. Carrier Gas : Served as mobile phase supplied in the steel tank under high pressure. At pressure of 40-80 psi this passes into flow controllers. Example- nitrogen, helium; hydrogen and argon can also be used. Requirements – Inert Column requirements Detectors Purity- better than 99.995% Cost effective & available GAS CHROMATOGRAPHY

9. Pneumatic Control : Gas supply is regulated to the correct pressure and then fed to the required part of instrument. Older instruments – manual pressure control via regulators. Modern GC instruments – Electronic Pneumatic pressure controller. Oven : Temperature programmable, typically range from 5°C - 400°C but can go as low as -25°C with cryogenic cooling.

10. Sample Injection Port : Sample is made to vaporized rapidly before entering to column. Various kind of injectors : Packed column injectors Split injection Splitless injection Programmed Split/ Splitless injection Programmed On-Column injector Column : Two kinds of column used : Packed column Capillary column Gas chromatograph GC-MS utilizes capillary column. Here, stationary phase has been chemically bonded to the fused silica, e.g., DB-5.

11. Packed Column :- Less commonly used, having diameter of 2-3 mm and length of 0.5-10 m. Manufactured from steel or glass, internal wall is treated to avoid catalytic effect with the sample. They can withstand a carrier gas flow rate within range 10-40ml/min. Capillary Column :- Consist of long capillary tubing 10-100 m in length. Made up from stainless steel & coil. Shorter columns - for fast analysis Larger columns – for high resolution separation

12. Detectors : Simple and reliable Sensitive to electronegative groups (halogens) Largely non-destructive Limited dynamic range (10 ²) Mass sensitive detectors -Thermal Conductivity Detector (TCD) -Flame Ionization Detector (FID) -Electron Capture Detector (ECD)

13. INTERFACE Pressure incompatibility problem between GC and MS was solved by inserting an interface. Mainly three types of interfaces are there commercially available : Jet/Orifice separator Direct capillary infusion interface Watson- Biemann effusion separator Fig : Direct capillary infusion Fig : Jet separator Fig : Watson- Biemann effusion

14. MASS SPECTROMETER “ Mass spectrometry is a technique used for measuring the molecular weight and determining the molecular formula of an organic compound.” In general a mass spectrometer consists of An ion source High-vacuum system A mass-selective analyzer An ion collector Data system

15. Ion source : sample introduction/ ionization method Ionization Method Typical Analytes Sample Introduction Mass Range Method Highlights Chemical Ionization (CI) Relatively small, volatile GC or liquid/solid probe Upto 1000 Daltons Soft method, molecular ion peak [M+H] + Electron Impact Ionization (EI) Relatively small, volatile GC or liquid/solid probe Upto 1000 Daltons Hard method, versatile, provides structure info Electrospray Ionization (ESI) Peptides, proteins, nonvolatile Liquid chromatography Upto 200000 Daltons Soft method, ions often multiply charged Fast Atom Bombardment (FAB) Carbohydrates, organometallics, peptides, nonvolatile Sample mixed in viscous matrix Upto 6000 Daltons Harder than ESI or MALDI Matrix Assisted Laser Desorption Ionization (MALDI) Peptides, proteins, nucleotides Sample mixed in solid matrix Upto 500000 Daltons Soft method, very high mass

16. Mass Selective Analyzer They deflects ions down a curved tubes in a magnetic fields based on their kinetic energy determined by the mass, charge and velocity. The magnetic field is scanned to measure different ions. Mass Analyzers Quadrupole Ion Trap Time of flight

17. Quadrupole Analyzer : Also known as ‘Hewlett-Packard’ or ‘Mass Selective Detector’. In quadrupole mass analyser a set of four rods are arranged parallel to the direction. Only m/z is been determined and stable oscillation takes place. Ions travels in quadrupole axis with cork screw type of trajectory. It functions as a mass filter

18. Ion Trap Mass Analyzer : The ion trap mass analyzer operates by similar principles where it consists of circular ring electrode and two end caps that form a chamber. AC or DC power along RF potential is applied between the cups and the ring electrode. Ions entering into chamber Trapped by electromagnetic fields Ions oscillate in concentric trajectories Resonant Ejection

19. Time-of-flight Analyzer : The time-of-flight (TOF) analyser uses an electric field to accelerate the ions through the same potential. Then it measures the time take to reach the detector. If the particles all have the same charge, the kinetic energies will be identical and their velocities will depend upon only on their masses.

20. Mass spectrometers employ computer control of same functions and also use a computerised display and output. It is used to identify and measure the concentration of one or more analytes in a complex mixture Advantages – Amount of data generated is very large It stores every sec for upto 90 min. Data Handling

21. GC TRACE OF A THREE COMPONENT MIXTURE. THE MASS SPECTROMETER GIVES A SPECTRUM FOR EACH COMPONENT

22. Consider the mass spectrum of CH 4 below:

23. LIMITATIONS Only compounds with vapour pressure exceeding about 10 -10 torr can be analysed by gas chromatography-mass spectrometry (GC-MS). Determining positional substitution on aromatic ring is often difficult. Certain isomeric compounds cannot be distinguished by mass spectrometry. Non-volatile matrices require additional preparation (extraction, outgassing, etc.) Atmospheric gases are challenging ( CO 2 , N 2 , O 2 , Ar , CO, H 2 O ).

24. Applications of GC-MS

25. CONCLUSION As we can see, GC-MS is used both in qualitative and quantitative measurement. Being hyphenated technique, the cost of GC-MS becomes very high. Nowadays, cost is also cheaper than before as quadrupole and ion trap instruments are used, which does not only save space but also save manufacturing cost. GC-MS is an analytical method of increased sensitivity and reliability even in very small quantities ( ng ).

26. Braun R., Introduction To Instrumental Analysis, Second Edition, PharmaMed Press, Hyderabad, Page no. 251-270. Chatwal G.R., Anand S.K., Instrumental method of Chemical Analysis, Himalaya Publishing House, Fifth Edition-2012, New Delhi, Page no. 420-449. Gohlke R.S.(1959), Analytical Chemistry, 36, Page no. 759-764. REFERENCES

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