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How to decide System Suitability Test (SST) in HPLC: Best Practice
Introduction and Outcome
System suitability test (SST) plays a crucial role in chromatographic analysis such as HPLC and GC. Deciding SST during method development is a major challenging task for any chromatographer. A small mistake can lead to a major failure of pharmaceuticals. You can see many 483 observations and deficiency letters on this. That is why I decided to share my skill-based knowledge on this, In this article, I will discuss system suitability test, approaches to decide system suitability criteria, System suitability solution, system suitability marker, system suitability acceptance criteria, retention time marker, case studies and frequently asked questions. After reading you can independently decide system system suitability test during method development.
Table of content
- Introduction and Outcome
- System suitability test
- System suitability test parameters
- Selection of Analytes for System suitability test standard
- Number of components in the System suitability test
- System suitability test marker
- What is the retention time (RT) marker?
- Establishment of system suitability test criteria
- Acceptance criteria for system suitability
- 7 Best practices of system suitability test
- Regulatory requirements on System suitability test
- Case studies
- Conclusion
- FAQs
System suitability test
The system suitability test measures whether the system is suitable to deliver the desired results. Several critical parameters such as the elution ordered of different components and their peak shape are considered during method development to decide the system suitability test. Generally, the chromatographic parameters that affect the column and detector performance are considered while deciding the system suitability test
System suitability test parameters
The following chromatographic parameters are used to define system suitability test;
- Retention time (RT), Relative retention time (RRT)
- Resolution (R)
- Theoretical plate or column efficiency (N)
- Tailing factor (T)
- Signal-to-noise ratio
- Capacity factor (K)
- Precision
Selection of Analytes
The analyte use for system suitability test must have the following characteristics:
- Easily available
- Stable
- Cost effective
- Good UV absorbance or good response
- Good solubility in the common solvents
- Related to process
- Polarity
Number of components in the System suitability test
The system suitably test must contain at least two chromatographic parameters. Generally, 2 to 5 chromatographic parameters are kept in the system suitability test. System suitability test solution is prepared by using individual analyte standard . SST marker standard is also used to prepare the system suitability solution.
System suitability test marker
System suitability marker contains all the components on which sst is decided.
What is the retention time (RT) marker?
If the process contains impurities of different polarities (polar, intermediate polar and nonpolar ) in that case retention time (RT) marker is used as a system suitability test standard. I personally recommend to use RT marker as an SST standard since this approach is cost effective and it avoids all confusion related to variation in the retention time.
Establishment of system suitability test criteria
- System suitability is decided based on the process impurities and their elution pattern.
- If the process contains a close eluting impurity peak, in that case these close eluting analytes must be included in SST acceptance criteria and Resolution will be one of the system suitability test parameter
- If the process does not contain close eluting impurity peak . In that case system suitability will be decided based on column efficiency (N) and tailing factor (T)
- Other chromatographic parameters like precision and signal to noise ratio can be included in the system suitability test.
- System suitability test may be method specific (e.g. assay method, impurity profile method and content test method)
Acceptance criteria for system suitability
- There must be base-to-base separation between the peaks. If analyte chromatogram contains close eluting peak, in that case resolution (R) will be one of the system suitability acceptance criteria.
- The minimum resolution should be not less than 1.5 between adjacent peaks. If 1.5 resolution is not achieved in that case lower resolution can be considered with suitable justification.
- The general limit of the USP tailing factor or symmetry factor is between 0.8 to 1.8. If USP tailing factor or symmetry factor is not achieved in this range in that case lower or higher value can be considered with suitable justification.
- The acceptance criteria of S/N ratio or signal to noise ratio for sensitivity or QL solutop should be more than or equal to 10
- The limit of theoretical plate, tailing factor and capacity factor should be decided based tend data of several batches
The typical example of reporting pattern
| System suitability test parameter | The obtained value | Acceptance criteria | Remarks |
| Resolution (R) | 3 (≥1.5) | The resolution between main analyte (A) and impurity (Y) should not be less than 1.5 | Complies |
| Column efficiency (N) | 5000 (≥4000) | The Column efficiency for main analyte (A) should be more than or equal to 4000 | Complies |
| Tailing factor (T) or symmetric factor | 1.3 (≤1.8) | The Tailing factor for main analyte (A) should be less than or equal to 1.8 | Complies |
| S/N or signal to noise ratio | 21 (≥10) | The S/N for sensitivity solution should be more than or equal to 10 | Complies |
| RSD | 0.7 %(≤2.0%) | The RSD of the area response of main analyte (A) of six replicates should be less than or equal to 2.0 | Complies |
Topical example to report retention time (RT) and relative retention time (RRT)
| Component name | RTs (minutes) | RRTs | Remarks* |
| Main analyte A | 5.4 (limit: about 5) | 1 | Complies |
| Imp. X | 7.7 (limit: about 8) | 1.54 (limit: about 1.5) | Complies |
| Imp.Y | 10.4 (limit: about 10) | 2.08 (limit: about 2) | Complies |
*Note: Refer to this post to handle variation in RTs and RRTs
7 Best practices of system suitability test
Use the following best practices for system suitability test :
- Keep at least two system suitability test criteria in the SST
- Keep resolution in the system suitability test if two peaks elute close to each other in the chromatogram
- Does not keep resolution in the system suitability test if two peaks elute far away to each other in the chromatogram
- Keep either column efficiency (N) or tailing factor in all cases while deciding the system suitability test criteria
- Keep Quantitation limit or sensitivity in the related substances, impurity profile and content test
- Keep precision test in the assay, related substances by external standard method and content test
- Don’t keep quantitation test (QL) or sensitivity test in the assay test
Regulatory requirements on System suitability test
Regulatory agencies like FDA, KFDA, TGA are more cautious about the system suitability testing. System suitability test must be decided scientifically and based on the trend data from multiple lots . Method must contains at lest two system suitability parameters
Case studies
System suitability test in the impurity profile test: A drug substance A contains 3 impurities. Main impurity elutes at 5 minutes, impurity B elutes at 6.5 minutes, impurity C elutes at 10 minutes, impurity D elutes at 25 minutes. What should be system suitability criteria?
Since main analyte A and impurity B elute adjacent to each other. Hence one of the system suitability parameter will be resolution (R). Column efficiency or tailing factor may be kept second system suitability test parameter. Since this is the impurity profile profile method and hence Quantitation limit will also be part of the system suitability test and acceptance criteria will be decided based on S/N ratio. The acceptance criteria of the system suitability will be decided based on trend data of several lots.
System suitability test in Assay test: In a drug substance X having assay limit 98 to 100%w/w. Drug substance X elutes at 6 minutes and its impurity A elutes at 30 minutes. What should be system suitability criteria?
Since impurity A peak elutes far away from the drug substance peak X and therefore Resolution can not be kept in the system suitability test. The system suitability can be decided using column efficiency and tailing factor. Precision will also be part of the system suitability
Inadequate system suitability test and its impact
If system suitability test is inadequate it may
- impact Quality of pharmaceuticals or drug products
- lead to warning letter from regulatory agencies and
- lead to deficiency letters from regulatory agencies
Conclusion
System suitability test measures the capability of the system or method. Therefore, both knowledge and experience are required to decide this. Now I hope this article, cleared all your doubts and now you can decide the system suitability test acceptance criteria for any method. For any opinion or suggestions related to this article, you can write in the comment section. For any further assistance you can contact me using contact form
FAQs
What is the SST in HPLC?
SST or system suitability test decides whether the HPLC system is suitable or not to give the correct result. SST contains two or more than two analytes. SST acceptance criteria.should be decided using at least two chromatographic parameters
What is the SST check?
SST check decides whether the chromatographic system is suitable or not to give the correct result. SST contains two or more than two analytes.
What are the parameters of SST in GC?
The chromatographic parameters like resolution, column efficiency and tiling factor are used in SST in GC
What is the system suitability solution?
System suitability test solution is prepared by using two or more than two analytes. In some of the method SST marker standard is used to prepare the system suitability solution.
What is the USP limit for the tailing factor?
The USP limit for the tailing factor is between 0.8 to 1.8
What is system suitability test marker?
System suitability marker contains all the analytes which decide system suitability acceptance criteria. It may be prepared in the lab or commercial SST marker can be used (if available)
How many chromatographic parameters are included in the system suitability test?
System suitability test acceptance criteria must contains at least two chromatographic parameters
References
- JP pharmacopeia
- USP pharmacopeia, chapter <621>
- https://www.fda.gov/media/163682/download
- https://www.pharmaguideline.net/fda-warning-letter-shilpa-medicare-limited/
Abbreviations
- RTs: Retention time
- RRTs: Relative retention time
- GC: Gas chromatography
- HPLC: High pressure liquid chromatograph
- R: Resolution
- N: Theoretical plate
- T: Tailing factor
- RSD: Relative standard deviation
- SST: system suitability test