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SOP for Calibration of High Performance Thin Layer Chromatography


Standard operating procedure to calibrate the High performance thin layer chromatography.

1.0 OBJECTIVE

To check the performance of High performance thin layer chromatography.

2.0 SCOPE

This procedure is applicable to calibrate high performance thin layer chromatography.

3.0 RESPONSIBILITY

3.1 Doing : Technical Assistant
3.2 Checking : Executive /Manager

4.0 ACCOUNTABILITY

Head of the Department

5.0 PROCEDURE

5.1 Check the performance of instrument in terms of
1. Linearity of spotting (Annexure-I)
2. Reproducibility of spotting (Annexure-II)
5.2 Prepare the mobile phase as per method given in Annexure-I.
5.3 Prepare the solution as mentioned in Annexure-I
5.4 Use 10 X 10 cm. HPTLC plate (Kieselgel 60F254 or Identical)
5.5 For Linearity of spotting
5.5.1 Apply 2 µl, 4µl, 6µl ,8µl and 10 µl of solution on HPTLC plate with spotter.
5.5.2 Allow the plate to run in mobile phase.
5.5.3 Dry the plate with drier.
5.5.4 Scan the plate with scanner.
5.5.5 Check the linearity and corelation coefficient.
5.5.6 Fill in the Data as mentioned in Annexure-I
5.6 Reproducibility of spotting
5.6.1 Apply 10µl solution on HPTLC plate for five times in sequence
5.6.2 Allow the plate to run in mobile phase.
5.6.3 Dry the plate with drier.
5.6.4 Scan the plate with scanner.
5.6.5 Calculate the RSD for five track ( RSD Limit : N.M.T 3.0%)
5.6.6 Fill in the data mentioned in Annexure-II


6.0 ABBREVIATIONS

6.1 %= Percentage
6.2 RSD= Relative standard deviation
6.3 N.M.T = Not more than
6.4 N.L.T = Not less than
6.5 µl = Microlitter
6.6 HPTLC = High performance thin layer chromatography
6.7 API= Active pharmaceuticals ingredient

Related: Principle of HPLC


ANNEXURE-I
Q. C. DEPARTMENT
PERFORMANCE CHECK RECORD
(LINEARITY OF SPOTTING)
NAME OF THE INSTRUMENT : HIGH PERFORMANCE THIN LAYER CHROMATOGRAPHY
MAKE                                            :
MODEL NO                                  :
IDENTIFICATION NO.               :
            Application Parameters :
            Plate Size                     : 10 X 10 cms.             Slit Width              :      
            Starting position           : 15 mm                        Migration distance :
            Band  width                 :  6 mm                         Saturation time      :
            Distance space             :  8 mm
            Sec/ ml                         :  15
            Inj.Volume                   :  2,4,6,8,10 ml
            Mobile Phase               : Methanol  : Ethyl acetate : Toluene : Ammonia
                                                          5     :          8          :      4      :   0.4
Sample preparation : 20 mg. Famotidine -----> 100 ml ---> 3 ml----> 10 ml with glacial acetic acid. (working std.)
                              Injection volume                     Area
                                       2.0
                                       4.0
                                       6.0
                                       8.0
                                     10.0   
     
Linearity : ________  ( It should be linear)  Corelation Coefficient : ___ ( Not less than 0.9900)
            Remarks  :  Satisfactory/Not Satisfactory
            Performance check  done by           :                                                      Date  :    

            Checked by                                       :                                                     Date  :

            Next Performance check  due date :

ANNEXURE-II
Q. C. DEPARTMENT
PERFORMANCE CHECK RECORD
(REPRODUCIBILITY OF SPOTTING)
NAME OF  THE INSTRUMENT : HIGH PERFORMANCE THIN  LAYER CHROMATOGRAPHY
MAKE                                             :
MODEL NO                                   :
IDENTIFICATION NO.                :

            Track no.        Applied volume (µl)         Area
               1                           10
               2                           10 
               3                           10
               4                           10
               5                           10

           RSD  =__________ ( Limit: Not more than 3.0%)
           Remarks                     : Satisfactory/Not Satisfactory
           Performance check  done by           :                                                    Date :    

           Checked by                                      :                                                    Date :

           Next Performance check  due date :
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.
Email: .moc.enilediugamrahp@ofni Need Help: Ask Question


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