Gas chromatography-Mass spectrometry (GC-MS)

PRESENTED BY
SAIRA FATIMA
SABAHAT MEHMOOD
SANA USMAN
MSc 4 (2018-2020)
Department of MicroBiology & Molecular Genetics
University of the Punjab
Lahore, Pakistan

1. GAS CHROMATOGRAPHY AND MASS SPECTROMETRY PRESENTED BY SAIRA FATIMA SABAHAT MEHMOOD SANA USMAN Department of MicroBiology & Molecular Genetics University of the Punjab Lahore, Pakistan MSc 2018-2020

2. Introduction: Gas chromatography-mass spectroscopy (GC-MS) is a hyphenated analytical technique. It is sensitive but also specific and reliable. GC can separate volatile and semi-volatile compounds with great resolution, but it cannot identify them. MS provide detailed structural information on most compounds such that they can be exactly identified, but can’t readily separate them. Therefore, both instruments have been proposed shortly after the development of GC in the mid-1950s. We obtain both qualitative and quantitative information of our sample in a single run within the same instrument. Today computerized GC/MS instruments are widely used in environmental monitoring ,in the regulation of agriculture and food safety , and in the discovery and production of medicine.

3. It is used in identification of unknown samples, including that of material samples obtained from planet Mars during probe missions as early as the 1970s. It can also be used in airport security to detect substances in luggage or on human beings. Additionally, it can identify  trace elements  in materials that were previously thought to have disintegrated beyond identification. Like   liquid chromatography–mass spectrometry , it allows analysis and detection even of tiny amounts of a substance.

4. History The use of mass spectrometer as a detector in gas chromatography was developed during the 1950s by Roland. Miniaturized computers has helped in the simplification of instrument In 1968, the Finnigan Instrument Corporation delivered the first quadrupole GC/MS By the 2000s computerized GC/MS instruments using quadrupole technology had become essential

5. Principle of GC-MS The sample solution is injected into the GC inlet where it is vaporized and swept onto a chromatographic column by the carrier gas (usually helium). The sample flows through the column and the compounds comprising the mixture of interest are separated by virtue of their relative interaction with the coating of the column (stationary phase) and the carrier gas (mobile phase). The latter part of the column passes through a heated transfer line and ends at the 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

6. Instrumentation The GC-MS is composed of two major building blocks, the  gas chromatograph  and the  mass spectrometer . The gas chromatograph utilizes a capillary column which depends on the column's dimensions (length, diameter, film thickness) as well as the phase properties (e.g. 5% phenyl polysiloxane ). The difference in the chemical properties between different  molecules  in a mixture and their relative affinity for the stationary phase of the column will promote separation of the molecules as the sample travels the length of the column. The molecules are retained by the column and then elute (come off) from the column at different times (called the retention time ), and this allows the mass spectrometer downstream to capture, ionize, accelerate, deflect, and detect the ionized molecules separately. The mass spectrometer does this by breaking each molecule into  ionized  fragments and detecting these fragments using their mass-to-charge ratio.

7. GC-MS

8. Steps:- Inject sample into Gas Chromatograph (GC) – The sample is injected into a port which is heated to up to 300° C where the material is then volatilized . Separation of gaseous components as they flow through the column – The column is wound within a special oven which controls temperatures from -20° to 320°.  The column surface is coated with a material which will separate the various chemical compounds in the sample based on size and/or polarity. Sample components that are more volatile and smaller in size will travel through the column more quickly than others. Analysis in the Mass Spectrometer (MS) – The separated components flow directly out of the column and into the MS which has three internal steps: Ionization source – components are blasted with electrons, causing them to break up and turn into positively charged ions

9.       Filter – the ions pass through an electromagnetic field and filter through based on mass. Analysts set a predetermined range of masses to be allowed through as they pass from the ionization source.        Detector – counting the number of filtered ions, the information is sent to a computer and a mass spectrum, a distribution of ions of different sizes, is generated.

10. Gas chromatography Technique used for the separation of volatile compounds that were easily vaporized at room temperature. Components Column Mobile phase Stationery phase Detector

11. C olumn :- U sed in gas chromatography very long and arranged in a coil Two types of column Packed column Consist of glass and stainless steel having Length 1 to 3 meters and internal diameter 2 to 4 millimeter Capillary column It made of fused quartz having a length 10 to 100 meters and internal diameter 0.1 to 1 mm Column is placed in a chamber so that uniform temperature can be maintained

12. Stationary phase Stationary phase is packed in the inner wall of the column Stationary phase is made of silicon grease or wax which can stand at high temperature .

13. Mobile Phase Inert gas helium usually is used as a mobile phase Non reactive gas is also used as a mobile phase The mobile gas is packed in a cylinder which connected to the column via molecular sieve which is used to remove unwanted gasses such as hydrocarbon, water vapours and oxygen

14. Detector Detector present at the end of column, used to detect test samples Flamed ionization detector is commonly used in gass chromatography FID have three inlet Attached with column With hydrogen cylinder Attached with oxygen cylinder Hydrogen oxygen ignite produces flame when sample molecule reached ionized and detect on detector in the form of electric current

15. Diagram

16. Working of gas chromatography Separated sample mixed with appropriate volatile such as heptane, acetone and methanol Just before the column septum is present which is used to inject sample Temperature of the injection region is kept 20 to 50 degree centigrade higher than the column Allowed the rapid volatilization of the sample This sample pass through the column where separation occurs During analyses temperature of the column is 150 to 300 degree centigrade .

17. Separation principles Separation occurs on the base of interaction of the molecule with stationary phase and volatile phase More volatile molecule less interact with stationary phase and move fast Less volatile molecule more interact with stationary phase and move slowly When the separation is completed , we can seen results on detector.

18. Diagram

19. Mass spectrometry The mass spectrometer is an instrument which help in separating the individual atoms or molecules because of difference in their masses. A mass spectrometer generates multiple ions from the sample under investigation It then separates them according to their specific mass-to-charge ratio (m/z), and then records the relative abundance of each ion type Produce ions from the sample in the ionization source. Separate these ions according to their mass-to-charge ratio in the mass analyzer.

20. Components:- The instrument consists of three major components: Ion Source: For producing gaseous ions from the substance being studied. Analyzer: For resolving the ions into their characteristics mass components according to their mass-to-charge ratio. Detector System: For detecting the ions and recording the relative abundance of each of the resolved ionic species.

21. Coupling of Gc-Ms Gas chromatograph and Mass spectrometry GC-MS Separates mixture of components into individual Identifies molecules based on their mass A chemical analysis technique combining two instruments to provide for powerful separation & identification Interface play important role in GCMS

22. Types of interface Capillary direct interface Jet separator (packed column) Watson & Biermann effusion separator

23. Capillary direct interface Today most GC-MS systems use capillary columns & fused silica tubing Permits an inert, high efficiency, direct transfer between the 2 systems. Fused silica tubing permits an inert, high efficiency, direct transfer between the 2 systems. Flow rates is 5ml/min

24. Jet separator (packed column) The separator consist of two glass tubes aligned with a Small distance between them. Carrier gas entering from the GC column is pumped away by a separate vacuumed system. The larger sample molecules maintain their momentum and pass preferentially in to the second capillary. Sample enrichment occurs & the initial atmospheric pressure is reduced.

25. Watson & Biermann effusion separator It consists of a sintered glass tube . The carrier usually Helium, passes preferentially through the sintered glass tube & the effluent in concentrated by a factor of up to 100. The gas flow rates in the order of 20-60ml/min.

26. Applications Advantages Disadvantages Summary

27. Applications Criminal forensics Law enforcement Spot anti doping analysis Chemical warfare agent detection Chemical engineering Food , beverage and perfume analysis Astro chemistry Medicine

28. GC-MS can analyze the particles from a human body in order to help link a criminal to a crime there is even an established American Society for Testing and Materials (ASTM) standard for fire debris analysis GCMS/MS is especially useful here as samples often contain very complex matrices and results, used in court, need to be highly accurate . Criminal forensics

29. Law enforcement GC-MS is increasingly used for detection of illegal narcotics, and may eventually supplant drug-sniffing dogs GC-MS is also commonly used in forensic toxicology to find drugs and/or poisons in biological specimens of suspects, victims, or the deceased.

30. Sports anti-doping analysis GC-MS is the main tool used in sports anti-doping laboratories to test athletes' urine samples for prohibited performance-enhancing drugs, for example anabolic steroids security

31. Chemical warfare agent detection:- As part of the post-September 11 drive towards increased capability in homeland security and public health preparedness, traditional GC-MS units with transmission quadrupole mass spectrometers, as well as those with cylindrical ion trap (CIT-MS) and toroidal ion trap (T-ITMS) mass spectrometers have been modified for field portability and near real-time detection of chemical warfare agents (CWA) such as sarin, soman , and VX These complex and large GC-MS systems have been modified and configured with resistively heated low thermal mass (LTM) gas chromatographs that reduce analysis time to less than ten percent of the time required in traditional laboratory systems.

32. Chemical engineering GC-MS is used for the analysis of unknown organic compound mixtures. One critical use of this technology is the use of GC-MS to determine the composition of bio-oils processed from raw biomass.

33. Food, beverage and perfume analysis Foods and beverages contain numerous aromatic compounds, some naturally present in the raw materials and some forming during processing. GC-MS is extensively used for the analysis of these compounds which include esters, fatty acids, alcohols, aldehydes, terpenes etc .

34. Astro chemistry Several GC-MS have left earth. Two were brought to Mars by the Viking program. Venera 11 and 12 and Pioneer Venus analyzed the atmosphere of Venus with GC-MS The Huygens probe of the Cassini–Huygens mission landed one GC-MS on Saturn's largest moon, Titan. The MSL Curiosity rover's Sample Analysis at Mars (SAM) instrument contains both a gas chromatograph and quadrupol mass spectrometer that can be used in tandem as a GC-MS. The material in the comet 67P/ Churyumov – Gerasimenko was analyzed by the Rosetta mission with a chiral GC-MS in 2014

35. ADVANTAGES

36. It is very useful technique and have a very advantage given following, It is important for identification of compound. It can Provides sensitive response to most analytes . It is important to provide information of particular or specific class of compound. It can provide information of structure or different structure of compound. It is having high resolution and separation capacity. It is time saving technique, having a high resolution capacity

37. Continue…….. It is simple, rapid, reproducible technique.  It play important role for determination of fragmentation pattern of compounds such as protein, peptides, amino acids and all biological or natural compounds.

38. Disadvantages Higher capital cost (approx. $ >85 K vs. $15 K for GC). Higher maintenance (time, expertise and money). For optimum results requires analyst knowledgeable in both chromatography and mass spectrometry Very complex process and tricky process

39. Summary GC/MS begins with the gas chromatograph, where the sample is volatized. This effectively vaporizes the sample (the gas phase) and separates its various component using a capillary column packed with stationary (solid) phase. The compounds are propelled by an inert carrier gas such as argon, helium or nitrogen. As the components become separated, they elute from the column at different times, which is generally referred to as their retention time.

40. Continue…. Once the components leave the GC column, they are ionized by mass spectrometer using chemical or electron ionization source. Ionized molecules are then accelerated through the instrument’s mass analyzer, which quite often is ion trap. It is here that ions are separated based on their different mass-to-charge (m/z) ratios. The final step of the process involve ion detection and analysis, with compound peaks appearing as a function of their (m/z) ratios. Peak heights, meanwhile, are proportional to the quantity of the corresponding compound. A complex sample will produce several different peaks, and the final readout will be a mass spectrum. Using computer libraries of mass spectra for different compounds, researchers can identify and quantitate unknown compounds and analytes .

41. Reference: https://en.wikipedia.org/wiki/Gas_chromatography%E2%80%93mass_spectrometry  Subramani Parasuraman1*, Anish R2 , Subramani Balamurugan3 , Selvadurai Muralidharan4 , Kalaimani Jayaraj Kumar5 and Venugopal Vijayan5  Sloan, K. M.; Mustacich , R. V.; Eckenrode , B. A. (2001). "Development and evaluation of a low thermal mass gas chromatograph for rapid forensic GC-MS analyses". Field Analytical Chemistry & Technology. 5 (6): 288–301. doi:10.1002/fact.10011  http://cires1.colorado.edu/jimenez/CHEM-5181/Labs/Gas_Chromatography.pdf

42. Continue……. https://www.slideshare.net/sagarsavale1/gcms-56112017  https://www.googleadservices.com/pagead/aclk?sa=L&ai=DChcSEwj90pvBxvTnAhWGjFEKHYpnB58YABABGgJ3cw&ohost=www.google.com&cid=CAESQOD25uy1FHG-Ed4aFPxROh7lfZI9XueoTcLNJG88jae38uXwuI4HSj8tdIuezuWgLnmPK-Mwom15XqeCAiyt0uc&sig=AOD64_3w1ixh5YDhJcRMAWVHDCfc0rsyLA&q=&ved=2ahUKEwiK4ZLBxvTnAhXp8eAKHfY0Bbo4ChDRDHoECA4QAQ&adurl  https://www.innovatechlabs.com/newsroom/642/stuff-works-gcms-analysis/

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