How Much Internal Standard Should Be Added in Targeted Metabolomics?
The amount of internal standard added in targeted metabolomics analysis depends on several factors, including sample type, analytical method, instrument sensitivity, and the concentration range of the target metabolites. Below are some general guidelines:
Selection of the Appropriate Internal Standard
1. Isotopically Labeled Internal Standards
These are internal standards labeled with isotopes that have a similar structure to the target metabolites but differ in mass.
2. Structurally Similar Internal Standards
In the absence of isotopically labeled internal standards, compounds with similar structures to the target metabolites can be used as internal standards.
Determination of Internal Standard Amount
1. Standard Curve Method
Prepare standard solutions of the internal standard within a known concentration range and add them to the samples, ensuring that the internal standard concentration falls within the linear range of the analytical method.
2. Empirical Approach
Based on previous experience and literature, choose an appropriate amount of internal standard to add. Typically, the internal standard concentration should be similar to that of the target metabolites to ensure the accuracy of quantitative analysis.
Specific Steps
1. Sample Preparation
Add the internal standard early in the sample preparation process to compensate for any losses or changes during sample handling and analysis.
2. Concentration Range
The internal standard concentration should fall within the expected concentration range of the target metabolites, generally 10 to 100 times that of the target metabolites, to ensure reliable quantitative results.
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