SOP for Good Chromatography Practices : Pharmaceutical Guidelines

SOP for Good Chromatography Practices

Standard operating procedure for working with HPLC and GC analysis of pharmaceutical products in quality control laboratory.

1.0 OBJECTIVE

To lay down a procedure for good chromatography practices.

2.0 SCOPE

This SOP shall be applicable for HPLC & GC analysis performed in the quality control department.

3.0 RESPONSIBILITY

QC officer / QC chemist

4.0 ACCOUNTABILITY

HOD QC

5.0 PROCEDURE

5.1. System Suitability

5.1.2 Consider peak area/ peak height as a measure of response from the detector as specified in the method of analysis. While entering the system suitability data in the notebook, only average area and %RSD obtained from the summary report shall be entered.
5.1.3 When system suitability requirements are specified in the method of analysis, only those parameters will be considered.
5.1.4 When limit for %RSD of replicate injections of a standard solution is specified as 2.0% or less, then five consecutive injections of reference solution will be made, unless otherwise specified.
5.1.5 When limit for %RSD of replicate injections of a standard solution is specified as more than 2.0%, then six consecutive injections of the standard solution will be made, unless otherwise specified.
5.1.6 For calculation of %RSD consecutive 5 or 6 sequential standards shall be considered.
5.1.7 Inject bracketing standard as specified in the method of analysis and calculate %RSD with respect to original system suitability. For bracketing standard, the limit or % RSD will be 1.5 times than the original limit
5.2 system suitability parameters are not specified, then following parameters shall be considered.
5.2.1 For tests of assay, dissolution, content uniformity; five replicate injections of reference/ standard solution shall be made & RSD limit will be not more than 2.0%.
5.2.2 For the tests of related substances (other than British Pharmacopeia) where %RSD limit of replicate injections is not specified, only a single injection of the reference / standard solution will be made.
5.2.3 For the tests of related substances (of the British Pharmacopeia) where %RSD limit of replicate injections is not specified, minimum three replicate injections of the reference/ standard solution will be made & %RSD limit will be not more than 5.0%.
5.2.4 For the tests of the residual solvents %RSD limit of six replicate injections will be 15% whenever not specified in the specifications.
5.3 In event of any interruption maximum up to six hours during HPLC run due to
  • Electric interruption
  • Computer interruption
  • The time gap between system suitability injection and sample injection due to delay in sample receipt.
  • The time gap between the two sets of samples
5.3.1 Bracketing standards shall be injected before proceeding with further next sample injection.
5.3.2 The above shall be applicable only in case of the system is in the continuous state of equilibration.

5.4 The deviation between retention times of standard or sample injections should not be more than 15% from the specified retention time in the method of analysis. For related substances test, the retention time of the sample shall be compared with the specified retention time
5.5 In the test of related substances, run the chromatogram for at least 1.5 times the retention time of principal peak unless otherwise specified.
5.6 Adjustment allowed for Liquid chromatography condition.
5.6.1 Following mentioned adjustment to the chromatographic conditions can be made to meet system suitability criteria, Prior permission of QC Head/Designee should be taken before proceeding with any change in the chromatographic conditions.
5.6.2 Composition of the Mobile Phase: the amount of the minor solvent component may be adjusted by ± 10 percent relative or ± 2 percent absolute, whichever is the larger. No other component is altered by more than 10 per cent absolute
5.6.3 pH of the aqueous component of the mobile phase: ± 0.05 pH unless otherwise stated in the monograph, or ± 1.0 pH when neutral substances are to be examined. The concentration of salts in the buffer component of a mobile phase: ± 10 percent.
5.6.4 Detector Wavelength: no adjustment permitted
5.6.5 Stationary Phase:
Column Length: ± 70 percent
Column Internal Diameter: ± 25 percent
5.6.6 Particle Size: maximal reduction of 50 percent, no increase permitted.
5.6.7 Flow Rate: ± 50 percent. When in a monograph the retention time of the principle peak is indicated, the flow rate has to be adjusted if the column internal diameter has been changed. No decrease of flow rate is permitted if the monograph uses an apparent number of theoretical plates in the qualification section.
5.6.8 Temperature: ± 10 percent, to a maximum of 60°C
5.6.9 Injection Volume: may be decreased, provided detection and repeatability of the peak (s) to be determined are satisfactory.
5.6.10 Gradient Elution: the configuration of the equipment employed may significantly alter the resolution, retention time and relative retentions described in the method. Should this occur, it may be due to excessive dwell volume which is the volume between the point at which the 2 eluants meet and the top of the column.
5.7 Adjustment allowed for Gas chromatography
5.7.1 Stationary phase: NO
5.7.2 Column length: ± 70 percent,
5.7.3 Column internal diameter: ± 50 percent
5.7.4 Particle size: maximal reduction of 50 percent, no increase permitted,
5.7.5 Film thickness: -50 percent to + 100 percent
5.7.6 Flow rate: ± 50 percent.
5.7.7 Temperature: ± 10 percent
5.7.8 Injection volume: may be decreased, providing detection and repeatability are satisfactory.

5.8 Data Generation
5.8.1 Follow the method of analysis & respective standard operating procedures to create appropriate method files, data files, sequence files.
5.8.2 Method Files
5.8.1.1 Once the method file is created check it for compliance as per the requirement of the method of analysis & save it.
5.8.1.2 When exact integration parameters or instrument parameters are not known then a trial run can be taken to identify the integration & or instrument parameters. After a trial run, the integration & or instrument parameters can be fixed by saving the method.
5.8.1.3 For related substances / chromatographic purity test, peak up to void volume is to be inhibited, All data must be integrated using the same integration parameters throughout the chromatographic analysis.
5.8.1.4 The integration parameters must be optimized for the entire analysis prior to processing to obtain between the representation of chromatography, While saving changed method, always enter the reason in the software.
5.8.1.5 After the analysis is over, take prints of method parameters & sign
5.9 Data Files
5.9.1 Prior to any sample injection single or sequence, feed the required data file name with required data path as per software requirement. Select the data file name, which is not previously used.
5.9.2 If data reprocessing is required with or without parameter change, then all the related data files like blank, standard, samples generated during that particular analysis will be reprocessed at the same time.
5.9.3 During or after sequence run, data file print will be taken & signed. If printouts are being taken online & after completion of injection but before completion of the entire sequence, ensure that same processing method is being used for all the injections.
5.9.4 During or after sequence run, data file print will be taken & signed. If printouts are being taken online & after completion of injection but before completion of the entire sequence, ensure that the same processing method is being used for all the injections.
5.9.5 Data description should be present on the data file whether printed or written manually on the printed data.
5.10 Sequence Files
5.10.1 While preparing any sequence always ensure that data file names given are not being repeated. The product information like stage, batch number/ A.R. number, test name product name strength, stability conditions etc can be entered in the report through the sequence.
5.10.2 Once the required sequence file is created & saved, load it to run the test.
5.10.3 Sequence can be amended or altered before running any particular analysis and Printouts of sequence parameters are taken & signed after testing.
5.11 Log-Book Entries
5.11.1 Data files, which are generated during analysis & not considered due to failure injections or some other reason, will be attached with the same set of data.
5.11.2 All such extra data will be signed & justification will be written on the printed data about their non-consideration & same shall be approved by QC.
5.11.3 The analyst shall sign the printed data.
5.11.4 The results like dissolution, dissolution profile, content uniformity and K factor obtained using excel sheet can directly be pasted in the notebook as final results.
& processing methods(s) and same shall be reviewed by QC.
5.12 Column Care
5.12.1 All HPLC & GC columns are to be handled with due care.
5.12.2 Always keep columns in their respective casing or box.
5.12.3 Avoid all types of jerks or falls of columns
5.12.4 Always flush columns with miscible solvents if used one after other.
5.12.5 Always replace end fittings of the respective columns after use.
5.12.6 Check the direction of the column before attaching it
5.12.7 Always check for any leakage after attaching the column, & then only close the lid of column compartment

5.13 Precautions
5.13.1 Always ensure that no air bubbles are entrapped in the solvent or mobile phase path.
5.13.2 Prior to injection, use syringe filters or centrifugation of solutions as applicable.
5.13.3 Always use filtered (through 0.45µ or less porosity filters) & sonicated mobile phase only
5.13.4 Before running any sequence, ensure that all the respective solutions (in vials), are located properly in the auto-sampler tray (s) as per the sequence.
5.13.5 Always check mobile phase or solutions for the absence of turbidity or opalescence, before & during use
5.13.6 During GC analysis always ensure that the flow of carrier gas, as well as other gases, is as per the requirement
5.13.7 Inject blank solution intermittently to nullify the effect of carryover if required

6.0 ABBREVIATIONS

6.1 QA: Quality Assurance
6.2 QC: Quality Control
6.3 RSD: Relative Standard Deviation
6.4 No. : Number
6.5 SOP: Standard Operating Procedure.
6.6 HOD: Head of Department
6.7 HPLC: High Pressure Liquid Chromatography
6.8 GC: Gas Chromatography

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Ankur Choudhary is India's first professional pharmaceutical blogger, author and founder of Pharmaceutical Guidelines, a widely-read pharmaceutical blog since 2008. Sign-up for the free email updates for your daily dose of pharmaceutical tips.
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