What principles should be followed in the selection of chrom

The quantitative analysis of chromatography is to determine the relative content of each component in the sample according to the size of Du signal detected by component Bai. It is based on Dao that the amount (weight or volume) of each component is proportional to the detection response value (peak height or peak area) generated by the chromatographic detector. Specific to specific methods, there are mainly the following methods:
1. Normalization method
When all the components in the sample can flow out of the column and generate corresponding signals on the detector (get the chromatographic peak), the normalization method is often used.
Ci = mi/m×100％ = fi•Ai /Σ fi•Ai ×100％
*Advantages: simple and convenient, not affected by the fluctuation of injection volume and operating conditions
*Disadvantages: all components must have peaks, and each component must have a correction factor
2. External standard method (also known as standard curve method)
The standard sample with known concentration is prepared for chromatographic analysis. The peak area or peak height of each component is measured, and the standard curve of peak area (or peak height) and concentration is made. Then, under the same operation conditions as the standard sample analysis, the same amount (usually volume) of unknown sample is entered to measure the peak area (or peak height) of the analyzed component, and the corresponding concentration can be found on the standard curve.
The accuracy of the analysis results mainly depends on the repeatability of the injection volume and the stability of the operating conditions.
Ci = mi/m×100％ = fiAi/m×100％
*Correction: standard curve, two point method and single point method
*Advantages: it is simple and does not require all components to peak (correction factor), and is often used for the analysis of several components
*Disadvantages: the fluctuation of operating conditions has a great influence on the injection volume
3. Internal standard method
Because of adsorption or chemical reaction, the chromatographic column can not make all components flow out, or each component can flow out, but the detector can not give the corresponding chromatographic peaks for all components, or some samples contain too much (can not get complete peaks), or too small, or only requires quantitative analysis of several components in complex samples, internal standard method can be used.
Process: if the internal standard s with mass MS is added into the sample with the total amount of M, the content of a component I can be calculated according to the weight of the detected substance and the internal standard and the corresponding peak area ratio on the chromatogram
mi/ms = fiAi / fsAs
Ci = mi/m ×100％
= fiAi ms / fsAs m×100％
*Advantages: it is not necessary for all components to peak (correction factor), and the operation conditions and injection volume have no effect
*Disadvantages: the internal standard is difficult to find - stable, non reactive, structure and performance, similar, retention time interpolation and complete separation
*Note: the internal standard method is more suitable for the analysis of low content components. Generally, the substances selected as internal standard should not exist in the sample, and the retention value should be in the middle of the retention value of all components. The content of the internal standard should not be significantly different from the content of the component to be tested.
4. Superposition method (internal addition)
The internal addition method is suitable for special cases: there is no suitable position to insert the internal standard peak in the spectrum, or there is no suitable internal standard due to various conditions.
At this time, a graph can be drawn from the sample, and then the sample can be injected after adding a certain amount of components to the sample to see how much the peak area has increased, and then the original content can be calculated by the ratio.
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