1.0 OBJECTIVE
To lay down the procedure for describing types of Chromatographic Methods
2.0 SCOPE
This SOP is applicable for describing types of Chromatographic Methods in company Name.
3.0 BACKGROUND
NIL
4.0 RESPONSIBILITY
4.1 QC/AD-officer
4.2 QC/AD-Executive
4.3 QC/AD-Head
5.0 PROCEDURE
5.1 Chromatographic analytical procedures are analyzed by two types of method there are
5.1.1 External Standard methods
5.1.2 Internal Standard methods
5.1.2 External Standard methods:
5.1.2.1 An external standard method is used when the standard is analyzed on a separate chromatogram from the sample. Quantitation is based on a comparison of the peak area/height (HPLC or GC) or spot intensity (TLC) of the sample to that of a reference standard of the analyte of interest.
5.1.2.2 The external standard method is more appropriate for samples as follows:
1. Sample with a single target concentration and narrow concentration range, e.g., acceptance and release tests.
2. Simple sample preparation procedure.
3. Increased baseline time for detection of potential extraneous peaks, e.g., impurities test.
5.1.2 Internal Standard Methods:
5.1.2.1 With an internal standard method, compound of known purity that does not cause interference in the analysis is added to the sample mixture.
5.1.2.2 Quantitation is based on the response ratio of compound of interest to the internal standard vs the response ratio of a similar preparation of the reference standard (HPLC or GC). This technique is rarely used for TLC methods.
5.1.2.3 The internal standard method is more appropriate for samples as follows:
1. Complex sample preparation procedures, e.g., multiple extractions.
2. Low concentration sample (sensitivity being an issue), e.g., pharmacokinetics studies.
3. Wide range of concentrations expected in the sample for analysis, e.g., pharmacokinetics studies.
HPLC methods for release and stability and TLC methods use external standards; and methods for biological fluids and GC methods use internal standards.
5.1.1 External Standard methods
5.1.2 Internal Standard methods
5.1.2 External Standard methods:
5.1.2.1 An external standard method is used when the standard is analyzed on a separate chromatogram from the sample. Quantitation is based on a comparison of the peak area/height (HPLC or GC) or spot intensity (TLC) of the sample to that of a reference standard of the analyte of interest.
5.1.2.2 The external standard method is more appropriate for samples as follows:
1. Sample with a single target concentration and narrow concentration range, e.g., acceptance and release tests.
2. Simple sample preparation procedure.
3. Increased baseline time for detection of potential extraneous peaks, e.g., impurities test.
5.1.2 Internal Standard Methods:
5.1.2.1 With an internal standard method, compound of known purity that does not cause interference in the analysis is added to the sample mixture.
5.1.2.2 Quantitation is based on the response ratio of compound of interest to the internal standard vs the response ratio of a similar preparation of the reference standard (HPLC or GC). This technique is rarely used for TLC methods.
5.1.2.3 The internal standard method is more appropriate for samples as follows:
1. Complex sample preparation procedures, e.g., multiple extractions.
2. Low concentration sample (sensitivity being an issue), e.g., pharmacokinetics studies.
3. Wide range of concentrations expected in the sample for analysis, e.g., pharmacokinetics studies.
HPLC methods for release and stability and TLC methods use external standards; and methods for biological fluids and GC methods use internal standards.
6.0 ANNEXURES
Nil
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![Difference between Stability[Shelf life] Specification and Release Specification](https://4.bp.blogspot.com/-O3EpVMWcoKw/WxY6-6I4--I/AAAAAAAAB2s/KzC0FqUQtkMdw7VzT6oOR_8vbZO6EJc-ACK4BGAYYCw/w680/nth.png)
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