How can quantitative analysis be performed based on LC-MS? Or what does quantification mean?

Quantitative analysis by LC-MS (Liquid Chromatography-Mass Spectrometry) is a method that separates components in a sample using liquid chromatography and then detects and quantifies them using mass spectrometry. The core objective of quantitative analysis is to calculate the concentration of the analyte using known standards or established standard curves.

 

Basic process of LC-MS quantitative analysis:

1. Sample Preparation

Dissolve or dilute the sample to be tested, and process it appropriately (such as centrifugation, filtration, purification, etc.) so that it can be effectively analyzed on the LC-MS instrument.

 

2. Liquid Chromatography Separation

Use liquid chromatography (LC) to separate the chemical components in the sample. The chromatographic column separates them based on different compounds' retention times (RT) on the column.

 

3. Mass Spectrometry Detection

Mass spectrometry (MS) detects the components separated by LC. LC-MS can provide highly sensitive qualitative and quantitative information, identifying and quantifying target compounds by their mass-to-charge ratio (m/z).

 

4. Standard Curve Method

(1) The standard curve method is the most commonly used quantitative method in LC-MS analysis. First, use standard solutions of known concentrations (standards) under the same experimental conditions to obtain response signals (usually peak area or peak height) at different concentrations. Then, use this data to plot a standard curve (concentration vs. peak area or peak height).

(2) After performing LC-MS analysis on the sample, compare the measured response signal (peak area or peak height) of the sample with the response signals on the standard curve to determine the concentration of the target analyte in the sample.

 

5. Internal Standard Method

(1) If it is difficult to create a standard curve (e.g., due to expensive or hard-to-obtain standards), the internal standard method can be used for quantification. This method adds a known concentration of an internal standard (a substance structurally similar to the target compound but not interfering with the analysis) to the sample to compensate for instrument fluctuations or losses during sample processing.

(2) Measure the response ratio of the internal standard to the target compound in the sample (i.e., peak area ratio or peak height ratio) and use the concentration and response of the internal standard to calculate the concentration of the target compound.

 

6. Data Analysis

Use specialized software (such as MassLynx, Xcalibur, MZmine, etc.) to process LC-MS data, extract the mass spectrometry and chromatographic data of the target compound, and perform peak identification and quantitative analysis.

 

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