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= '''Introduction''' = | = '''Introduction''' = | ||
Chromatography is a technique in the field of analytical chemistry that separates a fluid mixture into its parts for analysis. The field of chromatography is very broad and is used in many different ways. Chromatography can be done on a small strip of paper to large industrial columns for pharmaceutical purification [1]. | Chromatography is a technique in the field of analytical chemistry that separates a fluid mixture into its parts for analysis. The field of chromatography is very broad and is used in many different ways. Chromatography can be done on a small strip of paper to large industrial columns for pharmaceutical purification.<sup>[[https://www.khanacademy.org/science/class-11-chemistry-india/xfbb6cb8fc2bd00c8:in-in-organic-chemistry-some-basic-principles-and-techniques/xfbb6cb8fc2bd00c8:in-in-methods-of-purification-of-organic-compounds/a/principles-of-chromatography 1]]</sup> Types of chromatography include gas chromatography (GC), high performance liquid chromatography (HPLC), size exclusion, ion exchange, hydrophobic interaction, and affinity chromatography. The different types of chromatography, use different properties of molecules to separate and identify mixtures.<sup>[[https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 2]]</sup> | ||
The analytical tool known as gas chromatography | [[Image:GC_Image.jpg|300 px|right|thumbnail|'''Figure 1''' A high performance gas chromatography machine (center) that is used in conjunction with a mass spectrometer (left).<sup>[[https://ppmu.utm.my/laboratory-services/advance-mass-spectroscopy-lab/gcms-high-performance-gas-chromatography-mass-spectrometry/ 3]]</sup>]] | ||
The analytical tool known as gas chromatography was founded by Archer Martin and Richard Synge in 1941. The basis of this new technique, separating itself from other forms of chromatography at the time, was the concept that chromatography was not limited to just the liquid phase. It took nearly 10 years after the idea of gas chromatography, in 1951, for the concept to be proven as an effective analytical tool.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> Figure 1 shows a state of the art GC machine that is paired with a mass spectrometer for more streamline analysis. | |||
='''How It Works'''= | ='''How It Works'''= | ||
[[Image:GC_Pic.PNG|300 px| | [[Image:GC_Pic.PNG|300 px|left|thumbnail|'''Figure 2''' Flow sheet of a gas chromatography system with carrier gas, injector, column, detector and data system.<sup>[[https://www.chromedia.org/chromedia?waxtrapp=wlqdcDsHqnOxmOlIEcClBwFjE&subNav=rwhpbjDsHqnOxmOlIEcClBwFjEQ 5]]</sup>]] | ||
Gas chromatography can be broken down into a flow sheet diagram like in Figure 2. Gas chromatography uses a gas mobile phase and a liquid stationary phase for separation. The gas mobile phase is made up of a carrier gas, usually an inert gas like hydrogen, and the sample. The sample is injected into a vessel where it is turned into a gas through heat and is carried through the machine to the detector. Once the sample is ready, it is opened to the carrier gas and sent through the column with the help of pneumatic valves. Once the sample is carried into the column, it is allowed to interact with the stationary phase through different molecular interactions. The sample then is broken down into its components to be analyzed in the column. The analyzer instrument in the system detects the chemical properties of the sample and generates an electronic signal to be read. The electronic signal is then converted into a physical graph called a chromatogram for interpretation.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> | |||
[[Image:GC Tube.PNG|400 px|right|thumbnail|'''Figure 3''' The inside of a GC column detailing the mobile (flow) phase in an analyte and the stationary phase as they move toward the detector. <sup>[[https://openwetware.org/wiki/File:GC_Tube.PNG 6]]</sup>]] | |||
Components in the flow are traditionally eluted in order of descending vapor pressure. Therefore, the molecules that are usually seen in the gaseous phase are seen first on the chromatograms, followed by the heavier components of the analyzed mixture. To actively change the elution of gas chromatography, the stationary phase shown in Figure | Components in the flow are traditionally eluted in order of descending vapor pressure. Therefore, the molecules that are usually seen in the gaseous phase are seen first on the chromatograms, followed by the heavier components of the analyzed mixture. <sup>[[https://jackwestin.com/resources/mcat-content/separations-and-purifications/chromatography-basic-principles-involved-in-separation-process 6]]</sup> To actively change the elution of gas chromatography, the stationary phase shown in Figure 3 can be modified. Increasing the thickness will allow for more gases to be analyzed but with decreased resolution. Decreasing the thickness will increase resolution but at the expense of retention time. The temperature and pressure of the system can be changed to get different results, where increasing the temperature of the column or pressure of the sample can decrease the retention time but greatly increase resolution. By modifying equipment conditions, gas chromatography can be used in a variety of analytic work. | ||
= '''Advantages''' = | = '''Advantages''' = | ||
* Gas chromatography is fast, efficient, and can analyze many components at once. | * Gas chromatography is fast, efficient, and can analyze many components at once. | ||
* | * The resolution of the chromatogram is very high, giving very clear data and results. | ||
* GC analysis is very accurate and reliable for sample analysis. | * GC analysis is very accurate and reliable for sample analysis. | ||
* The volume of sample needed to analyze the solution is small. | * The volume of sample needed to analyze the solution is small. | ||
* Gas chromatography has been around for over 50 years and commercially available to most laboratories [ | * Gas chromatography has been around for over 50 years and is commercially available to most laboratories.<sup>[[https://ppmu.utm.my/laboratory-services/advance-mass-spectroscopy-lab/gcms-high-performance-gas-chromatography-mass-spectrometry/ 4]]</sup> | ||
='''Shortcomings'''= | ='''Shortcomings'''= | ||
* Samples need to be able to be stable in the gas phase. | * Samples need to be able to be stable in the gas phase. | ||
* The system is run at a high temperature so samples may break down during the analysis. | * The system is run at a high temperature so samples may break down during the analysis. | ||
* Cannot determine the structure of the sample [2]. | * Cannot determine the structure of the sample.<sup>[[https://ppmu.utm.my/laboratory-services/advance-mass-spectroscopy-lab/gcms-high-performance-gas-chromatography-mass-spectrometry/ 4]]</sup> | ||
='''Analysis'''= | |||
The analysis of chromatography starts at the molecular scale, looking at the separation of molecules in the mobile and stationary phases. The value that describes this partition between the two phases is called the retention value (Rf).<sup>[[https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 2]]</sup> | |||
<math>Rf=\frac{(distance \, traveled \, by \, sample)}{(distance \, traveled \, to \, solvent \, front)}=1-\alpha</math> | |||
Where the closer Rf gets to one, the faster the separation happens, while the closer it gets to zero, the slower the separation happens. To determine the Rf value, α, the partition coefficient needs to be determined.<sup>[[https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 2]]</sup> | |||
<math>\alpha=\frac{(Concentration \, in \, stationary \, phase)}{(Concentration \, in \, stationary \, and \, mobile \, phase)}</math> | |||
Chromatography runs are analyzed through a plot of detector signal versus time they produce called a chromatogram. The different components can be seen as peaks in the graph. The separation between two peaks, or components of a mixture, can be defined as the resolution.<sup>[[https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 2]]</sup> | |||
[[Image:GC_Chroma.PNG|300 px|right|thumbnail|'''Figure 4''' An example of what a chromatograph would look like plotted vs time.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup>]] | |||
<math> Resolution = \frac{0.589\Delta t_{r}}{1/2w_{av}} </math> | |||
Where Δt<sub>r</sub> is the difference in time between the two peaks and w<sub>av</sub> is the width of the peak in units of time. The area under the peak can generally tell how much of a compound or molecule is in a sample solution. Figure 4 shows what a typical GC chromatograph would look like. To determine which peak corresponds to which compound, a known sample can be run through the system as a reference or a standard from literature to best analyze the results.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> | |||
='''Applications'''= | ='''Applications'''= | ||
'''Medicine''' | '''Medicine''' | ||
Gas chromatography is a valuable resource to the medical field due to its quick and reliable analysis. By developing sample preparation and validation techniques, biological samples can be tested for drugs and proteins for medical testing. It was found in a 2004 study, that gas chromatography, in conjunction with mass spectroscopy, can determine concentrations of lidocaine, prilocaine, and other drugs. This sampling of blood can be done in just one minute, providing quick results to patients and doctors [ | Gas chromatography is a valuable resource to the medical field due to its quick and reliable analysis. By developing sample preparation and validation techniques, biological samples can be tested for drugs and proteins for medical testing. It was found in a 2004 study, that gas chromatography, in conjunction with mass spectroscopy, can determine concentrations of lidocaine, prilocaine, and other drugs. This sampling of blood can be done in just one minute, providing quick results to patients and doctors.<sup>[[https://doi.org/10.1016/j.jchromb.2004.09.020 7]]</sup> | ||
'''Pharmaceuticals''' | '''Pharmaceuticals''' | ||
Uses for gas chromatography in pharmaceuticals | Uses for gas chromatography in pharmaceuticals happen in not only development but also in post-market screening. Due to regulations, drugs need to be extremely pure with consistent properties. Using gas chromatography to analyze drug product solutions when developing an industrial purification process is important due to its quick and accurate analysis. The faster the properties of a drug product can be determined, the quicker a company can bring a drug to market.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> | ||
Pharmaceutical companies also have to do research into how effective their products are | Pharmaceutical companies also have to do research into how effective their products are. Testing for efficacy can include urine drug screening of patients on different drugs to see how it's working. Gas chromatography can be used to find drug fragments or different biological indicators to best determine the drug's true effect on patients.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> | ||
'''Food''' | '''Food''' | ||
Characterization of food molecules is critical for food companies | Characterization of food molecules is critical for food companies due to the many tight regulations. Analyzing food mixtures for quality and consistency is important for the company's image. Unilever research and development, has looked into oil compositions using gas chromatography. Oil and fats are usually not made up of a single molecule but a collection of many different variations of similar molecules. Companies need to know the number of different types of oils and fats, so the contents can be labeled on the products mandated by the regulatory agencies. Gas chromatography is used as a quick and effective analysis of different types of food.<sup>[[https://doi.org/10.1002/ejlt.200900074 8]]</sup> | ||
'''Petroleum''' | '''Petroleum''' | ||
Due to the large variations in chemicals in petroleum in diesel, it is commonly seen for product validation and characterization. Octane numbers for gasoline are determined through the composition of gas. Providing as close to real time data on the composition of affluent gas streams of a refinery | Due to the large variations in chemicals in petroleum in diesel, it is commonly seen for product validation and characterization. Octane numbers for gasoline are determined through the composition of gas. Providing as close to real-time data on the composition of affluent gas streams of a refinery is key to providing reliable products to consumers. Due to the quick characterization of gas chromatography, it is one of the most popular analysis techniques used in the petroleum industry.<sup>[[https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf 4]]</sup> | ||
='''References'''= | ='''References'''= | ||
[1] | [1] Principles of Chromatography. 2016. https://www.khanacademy.org/science/class-11-chemistry-india/xfbb6cb8fc2bd00c8:in-in-organic-chemistry-some-basic-principles-and-techniques/xfbb6cb8fc2bd00c8:in-in-methods-of-purification-of-organic-compounds/a/principles-of-chromatography (accessed 2023-03-26). | ||
[2] Kumar, P. Top 12 Types of Chromatographic Techniques | Biochemistry. biologydiscussion.com. https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 (accessed 2023-03-26). | |||
[3] HIGH PERFORMANCE GAS CHROMATOGRAPHY MASS SPECTROMETRY, 2018. https://ppmu.utm.my/laboratory-services/advance-mass-spectroscopy-lab/gcms-high-performance-gas-chromatography-mass-spectrometry/ (accessed 2023-03-26). | |||
[4] "Theory and Instrumentation of GC." GC's CHROM Academy (n.d.): 1-24. Crawford Scientific. Web. https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf (accessed 2023-03-26). | |||
[ | [5] Snow, N. Introduction to Capillary GC Injection Techniques; Chromedia Analytical Sciences. https://www.chromedia.org/chromedia?waxtrapp=wlqdcDsHqnOxmOlIEcClBwFjE&subNav=rwhpbjDsHqnOxmOlIEcClBwFjEQ (accessed 2023-03-26). | ||
[ | [6] Chromatography Basic Principles Involved In Separation Process - Separations And Purifications - MCAT Content. Jack Westin. https://jackwestin.com/resources/mcat-content/separations-and-purifications/chromatography-basic-principles-involved-in-separation-process (accessed 2024-03-10). | ||
[ | [7] Altun, Z.; Abdel-Rehim, M.; Blomberg, L. G. New Trends in Sample Preparation: On-Line Microextraction in Packed Syringe (MEPS) for LC and GC Applications Part III: Determination and Validation of Local Anaesthetics in Human Plasma Samples Using a Cation-Exchange Sorbent, and MEPS-LC-MS-MS. J Chromatogr B Analyt Technol Biomed Life Sci 2004, 813 (1–2), 129–135. https://doi.org/10.1016/j.jchromb.2004.09.020. | ||
[ | [8] Janssen, H.-G.; Steenbergen, H.; de Koning, S. The Role of Comprehensive Chromatography in the Characterization of Edible Oils and Fats. European Journal of Lipid Science and Technology 2009, 111 (12), 1171–1184. https://doi.org/10.1002/ejlt.200900074. | ||
Latest revision as of 11:31, 10 March 2024
Introduction
Chromatography is a technique in the field of analytical chemistry that separates a fluid mixture into its parts for analysis. The field of chromatography is very broad and is used in many different ways. Chromatography can be done on a small strip of paper to large industrial columns for pharmaceutical purification.[1] Types of chromatography include gas chromatography (GC), high performance liquid chromatography (HPLC), size exclusion, ion exchange, hydrophobic interaction, and affinity chromatography. The different types of chromatography, use different properties of molecules to separate and identify mixtures.[2]
The analytical tool known as gas chromatography was founded by Archer Martin and Richard Synge in 1941. The basis of this new technique, separating itself from other forms of chromatography at the time, was the concept that chromatography was not limited to just the liquid phase. It took nearly 10 years after the idea of gas chromatography, in 1951, for the concept to be proven as an effective analytical tool.[4] Figure 1 shows a state of the art GC machine that is paired with a mass spectrometer for more streamline analysis.
How It Works
Gas chromatography can be broken down into a flow sheet diagram like in Figure 2. Gas chromatography uses a gas mobile phase and a liquid stationary phase for separation. The gas mobile phase is made up of a carrier gas, usually an inert gas like hydrogen, and the sample. The sample is injected into a vessel where it is turned into a gas through heat and is carried through the machine to the detector. Once the sample is ready, it is opened to the carrier gas and sent through the column with the help of pneumatic valves. Once the sample is carried into the column, it is allowed to interact with the stationary phase through different molecular interactions. The sample then is broken down into its components to be analyzed in the column. The analyzer instrument in the system detects the chemical properties of the sample and generates an electronic signal to be read. The electronic signal is then converted into a physical graph called a chromatogram for interpretation.[4]
Components in the flow are traditionally eluted in order of descending vapor pressure. Therefore, the molecules that are usually seen in the gaseous phase are seen first on the chromatograms, followed by the heavier components of the analyzed mixture. [6] To actively change the elution of gas chromatography, the stationary phase shown in Figure 3 can be modified. Increasing the thickness will allow for more gases to be analyzed but with decreased resolution. Decreasing the thickness will increase resolution but at the expense of retention time. The temperature and pressure of the system can be changed to get different results, where increasing the temperature of the column or pressure of the sample can decrease the retention time but greatly increase resolution. By modifying equipment conditions, gas chromatography can be used in a variety of analytic work.
Advantages
- Gas chromatography is fast, efficient, and can analyze many components at once.
- The resolution of the chromatogram is very high, giving very clear data and results.
- GC analysis is very accurate and reliable for sample analysis.
- The volume of sample needed to analyze the solution is small.
- Gas chromatography has been around for over 50 years and is commercially available to most laboratories.[4]
Shortcomings
- Samples need to be able to be stable in the gas phase.
- The system is run at a high temperature so samples may break down during the analysis.
- Cannot determine the structure of the sample.[4]
Analysis
The analysis of chromatography starts at the molecular scale, looking at the separation of molecules in the mobile and stationary phases. The value that describes this partition between the two phases is called the retention value (Rf).[2]
[math]\displaystyle{ Rf=\frac{(distance \, traveled \, by \, sample)}{(distance \, traveled \, to \, solvent \, front)}=1-\alpha }[/math]
Where the closer Rf gets to one, the faster the separation happens, while the closer it gets to zero, the slower the separation happens. To determine the Rf value, α, the partition coefficient needs to be determined.[2]
[math]\displaystyle{ \alpha=\frac{(Concentration \, in \, stationary \, phase)}{(Concentration \, in \, stationary \, and \, mobile \, phase)} }[/math]
Chromatography runs are analyzed through a plot of detector signal versus time they produce called a chromatogram. The different components can be seen as peaks in the graph. The separation between two peaks, or components of a mixture, can be defined as the resolution.[2]
[math]\displaystyle{ Resolution = \frac{0.589\Delta t_{r}}{1/2w_{av}} }[/math]
Where Δtr is the difference in time between the two peaks and wav is the width of the peak in units of time. The area under the peak can generally tell how much of a compound or molecule is in a sample solution. Figure 4 shows what a typical GC chromatograph would look like. To determine which peak corresponds to which compound, a known sample can be run through the system as a reference or a standard from literature to best analyze the results.[4]
Applications
Medicine Gas chromatography is a valuable resource to the medical field due to its quick and reliable analysis. By developing sample preparation and validation techniques, biological samples can be tested for drugs and proteins for medical testing. It was found in a 2004 study, that gas chromatography, in conjunction with mass spectroscopy, can determine concentrations of lidocaine, prilocaine, and other drugs. This sampling of blood can be done in just one minute, providing quick results to patients and doctors.[7]
Pharmaceuticals Uses for gas chromatography in pharmaceuticals happen in not only development but also in post-market screening. Due to regulations, drugs need to be extremely pure with consistent properties. Using gas chromatography to analyze drug product solutions when developing an industrial purification process is important due to its quick and accurate analysis. The faster the properties of a drug product can be determined, the quicker a company can bring a drug to market.[4]
Pharmaceutical companies also have to do research into how effective their products are. Testing for efficacy can include urine drug screening of patients on different drugs to see how it's working. Gas chromatography can be used to find drug fragments or different biological indicators to best determine the drug's true effect on patients.[4]
Food Characterization of food molecules is critical for food companies due to the many tight regulations. Analyzing food mixtures for quality and consistency is important for the company's image. Unilever research and development, has looked into oil compositions using gas chromatography. Oil and fats are usually not made up of a single molecule but a collection of many different variations of similar molecules. Companies need to know the number of different types of oils and fats, so the contents can be labeled on the products mandated by the regulatory agencies. Gas chromatography is used as a quick and effective analysis of different types of food.[8]
Petroleum Due to the large variations in chemicals in petroleum in diesel, it is commonly seen for product validation and characterization. Octane numbers for gasoline are determined through the composition of gas. Providing as close to real-time data on the composition of affluent gas streams of a refinery is key to providing reliable products to consumers. Due to the quick characterization of gas chromatography, it is one of the most popular analysis techniques used in the petroleum industry.[4]
References
[1] Principles of Chromatography. 2016. https://www.khanacademy.org/science/class-11-chemistry-india/xfbb6cb8fc2bd00c8:in-in-organic-chemistry-some-basic-principles-and-techniques/xfbb6cb8fc2bd00c8:in-in-methods-of-purification-of-organic-compounds/a/principles-of-chromatography (accessed 2023-03-26).
[2] Kumar, P. Top 12 Types of Chromatographic Techniques | Biochemistry. biologydiscussion.com. https://www.biologydiscussion.com/biochemistry/chromatography-techniques/top-12-types-of-chromatographic-techniques-biochemistry/12730 (accessed 2023-03-26).
[3] HIGH PERFORMANCE GAS CHROMATOGRAPHY MASS SPECTROMETRY, 2018. https://ppmu.utm.my/laboratory-services/advance-mass-spectroscopy-lab/gcms-high-performance-gas-chromatography-mass-spectrometry/ (accessed 2023-03-26).
[4] "Theory and Instrumentation of GC." GC's CHROM Academy (n.d.): 1-24. Crawford Scientific. Web. https://dl.icdst.org/pdfs/files/d6f7ffecfb49f8c43a933328f2db2a8c.pdf (accessed 2023-03-26).
[5] Snow, N. Introduction to Capillary GC Injection Techniques; Chromedia Analytical Sciences. https://www.chromedia.org/chromedia?waxtrapp=wlqdcDsHqnOxmOlIEcClBwFjE&subNav=rwhpbjDsHqnOxmOlIEcClBwFjEQ (accessed 2023-03-26).
[6] Chromatography Basic Principles Involved In Separation Process - Separations And Purifications - MCAT Content. Jack Westin. https://jackwestin.com/resources/mcat-content/separations-and-purifications/chromatography-basic-principles-involved-in-separation-process (accessed 2024-03-10).
[7] Altun, Z.; Abdel-Rehim, M.; Blomberg, L. G. New Trends in Sample Preparation: On-Line Microextraction in Packed Syringe (MEPS) for LC and GC Applications Part III: Determination and Validation of Local Anaesthetics in Human Plasma Samples Using a Cation-Exchange Sorbent, and MEPS-LC-MS-MS. J Chromatogr B Analyt Technol Biomed Life Sci 2004, 813 (1–2), 129–135. https://doi.org/10.1016/j.jchromb.2004.09.020.
[8] Janssen, H.-G.; Steenbergen, H.; de Koning, S. The Role of Comprehensive Chromatography in the Characterization of Edible Oils and Fats. European Journal of Lipid Science and Technology 2009, 111 (12), 1171–1184. https://doi.org/10.1002/ejlt.200900074.
