Determining concentration of compound using #GCMS

shirwansany

31 Mar 202230:39

EducationalLearning

32 Likes 10 Comments

TLDRThis video script offers an in-depth guide on quantifying compound concentrations using Gas Chromatography-Mass Spectrometry (GC-MS). It focuses on the identification and quantification of Fatty Acid Methyl Esters (FAMEs), detailing the preparation of calibration curves and the process of analyzing samples. The tutorial demonstrates the use of software for data analysis, comparing the Selective Ion Monitoring (SIM) mode with the Scan mode for specificity and cleanliness of results. The script promises a follow-up on manual calculation methods for quantification, highlighting the importance of linearity in calibration curves for accurate measurements.

Takeaways

🧪 The video is about using Gas Chromatography-Mass Spectrometry (GC-MS) to identify and quantify compounds, specifically focusing on the quantification of Fatty Acid Methyl Esters (FAME).
🛒 The presenter purchased standard FAME compounds to create a calibration curve for quantification purposes.
🔍 The GC-MS analysis was conducted using a specific instrument, the GT-12 MF Gas Chromatography Time of Flight Mass Spectrometer, to identify and quantify the compounds.
📈 The video explains how to prepare a calibration curve within the software of the GC-MS, which is essential for quantification.
📚 The presenter has previously published a chapter on the identification of 37 FAME compounds, which is referenced for the mass assignments used in the GC-MS.
📝 The process involves creating a calibration curve by analyzing standards at various concentrations and plotting the results to establish a linear relationship.
🔬 The Selected Ion Monitoring (SIM) mode is preferred for quantification due to its specificity and cleaner chromatogram compared to the Scan mode.
📉 The calibration curve's linearity is assessed using the coefficient of determination (R²), with values close to 1 indicating good linearity.
📑 The video demonstrates how to manually input data points for the calibration curve and how to integrate and quantify samples based on peak area responses.
🍪 The presenter applies the quantification method to real-world samples, such as biscuits with added oil, to determine the concentration of specific FAME compounds.
🔑 The video concludes with a teaser for a future video that will cover manual calculations for quantification using GC-MS results.

Q & A

What is the main topic of the video script?
-The main topic of the video script is about how to quantify or determine the concentration of any compound in a sample using Gas Chromatography Mass Spectrometer (GC-MS), specifically focusing on the quantification of Fatty Acid Methyl Esters (FAME).
What is the purpose of the video script?
-The purpose of the video script is to guide viewers on how to perform quantification analysis using GC-MS after identifying compounds, which is a continuation from a previous video focused on qualitative analysis.
What is the difference between SIM (Selected Ion Monitoring) mode and SCAN mode in GC-MS as mentioned in the script?
-In GC-MS, SIM mode is more specific and cleaner than SCAN mode. SIM mode allows for the detection of only specific masses that are of interest, making it ideal for quantification due to its specificity and cleaner chromatogram. SCAN mode, on the other hand, detects a broader range of masses, including impurities, which can make the chromatogram more complex.
What is the role of an internal standard in the calibration curve?
-An internal standard is used in calibration curves to account for any variations in the sample preparation or the analytical process. It helps to increase the accuracy of the quantification by normalizing the response of the target compounds to that of the internal standard.
What does the speaker mean by 'axion equivalation curve'?
-An 'axion equivalation curve' likely refers to a calibration curve that does not include an internal standard. The term might be a mispronunciation or a typo for 'external equivalation curve', which would be a curve established without an internal standard for comparison.
What is the significance of the coefficient of determination (R²) in the calibration curve?
-The coefficient of determination (R²) indicates the goodness of fit of the calibration curve. An R² value close to 1 suggests a high degree of linearity and a good fit, which is desirable for accurate quantification.
How does the speaker plan to introduce the FAME standards to the GC-MS system?
-The speaker plans to introduce the FAME standards by opening the files with the highest concentration of the standards, identifying the retention time and mass-to-charge ratio (m/z) of each compound, and then manually entering these into the system to create a calibration curve.
What is the process of integrating a sample in GC-MS analysis?
-Integration in GC-MS analysis involves identifying and measuring the area under the peak of a compound in the chromatogram, which corresponds to the amount of the compound present in the sample. This step is crucial for accurate quantification.
Why is it important to have multiple points in a calibration curve?
-Having multiple points in a calibration curve is important because it helps to establish a reliable mathematical model for predicting the concentration of unknown samples. More points provide a better representation of the relationship between the detector response and the concentration of the compound.
How does the speaker plan to demonstrate the quantification of FAME in a sample?
-The speaker plans to demonstrate the quantification by integrating the peaks of the FAME compounds in the sample's chromatogram, using the established calibration curve to calculate the concentration of the compounds, and then presenting the results.

Outlines

00:00

🔬 Introduction to Quantification Using GC-MS

The speaker begins by greeting the audience and introducing the topic of quantification of compounds using Gas Chromatography-Mass Spectrometry (GC-MS). They explain that the video will focus on how to determine the concentration of a compound in a sample, specifically fatty acid methyl esters (FAME), using GC-MS. The speaker mentions that they have a GT-12 MF Gas Chromatography Time of Flight Mass Spectrometer in their lab and will demonstrate the process of preparing a calibration curve using the software, without an internal standard, and how to analyze the results to quantify the compounds.

05:03

📊 Selective Ion Monitoring Mode in GC-MS Analysis

In this paragraph, the speaker discusses the use of Selected Ion Monitoring (SIM) mode in GC-MS for a more specific and cleaner analysis. They explain how to identify the mass of individual fatty acids using the certificate of analysis provided with the standards. The speaker references their published work on the antibacterial activity of fatty acid esters and guides the audience through the process of setting up the SIM mode to detect specific masses of the compounds, emphasizing the advantages of SIM over full scan mode for quantification.

10:05

📚 Preparing a Calibration Curve Using GC-MS Data

The speaker outlines the process of preparing a calibration curve using data from the GC-MS analysis. They describe how to open the highest concentration file, set up the quantitation, and introduce the standards into the system. The explanation includes how to identify the retention time and mass of specific compounds, such as C6 and C8, and how to input this information into the software to create a calibration curve. The speaker also mentions that they will cover the process of preparing a calibration curve with an internal standard in a future video.

15:05

📈 Adding Concentration Levels to the Calibration Curve

This section details the step-by-step process of adding different concentration levels to the calibration curve. The speaker demonstrates how to load various concentration files, update the levels, and integrate the data points into the curve. They explain the importance of starting with the lowest concentration and progressively adding higher concentrations, ensuring that the calibration curve is accurate and reliable for quantifying the compounds in the samples.

20:05

📉 Assessing Linearity and Quantifying Compounds in Samples

The speaker discusses how to assess the linearity of the calibration curve by checking the coefficient of determination (R²) for the compounds C6 and C8. They consider values of 0.98 and 0.99 as satisfactory for the calibration curve's linearity. The speaker then moves on to demonstrate how to quantify the compounds in actual samples by integrating the peaks, calculating the response, and determining the concentration of C6 and C8 in the samples, such as biscuits with added oil.

25:07

📝 Conclusion and Preview of Future Content

In the concluding paragraph, the speaker summarizes the video's content and announces that in the next video, they will explain how to quantify the C6 and C8 fatty acids manually using the results from the GC-MS analysis. They end the video with a farewell greeting and express their hope to see the audience in the next installment of the series.

Mindmap

Keywords

💡Gas Chromatography Mass Spectrometer (GC-MS)

Gas Chromatography Mass Spectrometer is an analytical technique that combines the separation capabilities of gas chromatography with the detection and quantification features of mass spectrometry. It is used to identify and quantify compounds in a sample. In the video, the presenter discusses using GC-MS for the identification and quantification of fatty acid methyl esters (FAMEs), highlighting its importance in the qualitative and quantitative analysis of compounds.

💡Quantification

Quantification refers to the process of determining the quantity or concentration of a specific substance within a sample. In the context of the video, the term is used to describe the measurement of the concentration of individual compounds identified by the GC-MS. The script mentions how the presenter will share methods for quantification of compounds, specifically FAMEs, using the GC-MS technique.

💡Fatty Acid Methyl Esters (FAMEs)

Fatty Acid Methyl Esters are the methyl esters of fatty acids, which are commonly used as a standard for analysis in various fields, including biofuels and food chemistry. In the video, the presenter purchases FAMEs in a standard form to use as a reference in their GC-MS analysis, emphasizing the role of these compounds in the quantification process.

💡Calibration Curve

A calibration curve is a graph that represents the relationship between the concentration of a known standard and the response of the analytical instrument, such as the detector signal in GC-MS. The video script describes the creation of a calibration curve within the GC-MS software, which is essential for accurately quantifying unknown samples by comparing their responses to those of known standards.

💡Selected Ion Monitoring (SIM)

Selected Ion Monitoring is a mode in mass spectrometry where the detector is set to record only specific ions, increasing the selectivity and sensitivity of the analysis. The script mentions the use of SIM mode in GC-MS to focus on the ions specific to the FAMEs being analyzed, allowing for a cleaner and more specific detection of the compounds of interest.

💡Retention Time

Retention Time in chromatography is the time it takes for a compound to pass through the system from injection to detection. It is a critical parameter for identifying compounds as different compounds have different retention times. The video script refers to the retention times of FAMEs to identify and quantify them during the GC-MS analysis.

💡Mass Spectroscopy

Mass Spectroscopy is an analytical technique used to identify and quantify substances by measuring the mass-to-charge ratio of ions. In the video, mass spectroscopy is part of the GC-MS process, where the mass of specific ions is used to identify the FAMEs present in the sample.

💡Integration

Integration in the context of chromatography refers to the process of measuring the area under a peak, which corresponds to the amount of a compound present. The video script describes the integration step as a necessary process before quantification, where the area of the peak for each compound is measured to determine its concentration in the sample.

💡Peak Area

Peak Area is the space under a peak in a chromatogram, which is proportional to the amount of a compound in the sample. The script mentions the peak area as the measurement used in the quantification process, where the area of the peak for FAMEs is used to calculate their concentration in the sample.

💡Concentration

Concentration refers to the amount of a substance present in a given volume or mass of a mixture. In the video, the presenter discusses determining the concentration of FAMEs in samples using the GC-MS, with the final aim of quantifying how much of each FAME is present in the analyzed samples.

Highlights

Introduction to quantification of compounds using Gas Chromatography Mass Spectrometer (GCMS).

Explanation of how to identify individual compounds by GCMS for qualitative analysis.

Discussion on the difference between qualitative and quantitative analysis in GCMS.

The process of purchasing and analyzing Fatty Acid Methyl Esters (FAME) using GCMS.

Description of the GT-12 MF Gas Chromatography Time of Flight Mass Spectrometer.

Demonstration of how to operate the GCMS machine for compound identification.

Use of the Selected Ion Monitoring (SIM) mode for compound quantification in GCMS.

Advantages of SIM mode for cleaner and more specific detection in GCMS analysis.

How to prepare a calibration curve within the GCMS software for quantification.

Explanation of the external standard method for preparing calibration curves without an internal standard.

Introduction of the process to manually create a calibration curve using Excel.

Detailed steps for adding compounds to the GCMS system for calibration curve establishment.

Importance of the retention time and mass in identifying individual fatty acid compounds.

The use of a published paper for reference on the mass of individual fatty acids.

Procedure for integrating and quantifying the concentration of compounds in a sample.

Demonstration of how to calculate the concentration of C6 and C8 in a biscuit sample.

The significance of coefficient of determination in assessing the linearity of calibration curves.

Preview of the next video's content on manual quantification calculations using GCMS results.

Conclusion of the video with a summary of the process and a greeting to the viewers.

Transcripts

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Related Tags

GC-MS Analysis Compound Quantification Fatty Acid Methyl Esters Calibration Curve Selected Ion Monitoring Laboratory Techniques Data Integration Scientific Research Video Tutorial Educational Content