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MISOPROSTOL SOP


1.0  OBJECTIVE:
To lay down a procedure of analytical report for the active raw material of the Misoprostol from the Pharmacopoeial specifications.
2.0  SCOPE:
This SOP shall be applicable in Q.C laboratory.
3.0  RESPONSIBILITY:
3.1  Q.C Analyst.
4.0  ACCOUNTABILITY:
4.1  Q.C Manager.
5.0  PROCEDURE:
5.1  Characters:
5.1.1        Appearance:
5.1.1.1  Clear.
5.1.1.2  Colourless or yellowish.
5.1.1.3  Oily liquid.
5.1.1.4  Hygroscopic.
5.1.2        Solubility:
5.1.2.1  Material and equipment:
5.1.2.1.1        Glassware (test tubes, spatula).
5.1.2.1.2        Ethanol (96.0%).
5.1.2.1.3        Acetonitrile.
5.1.2.1.4        Purified water.
5.1.2.2  Sample:
5.1.2.2.1        Small quantity.
5.1.2.3  Method:
5.1.2.3.1        Take 3 test tubes and add small quantity of sample for testing solubility according to B.P specifications.
5.1.2.3.2        Add purified water in test tube 1 and observe.
5.1.2.3.3        Add ethanol (96.0%) in test tube 2 and observe.
5.1.2.3.4        Add acetonitrile in test tube 3 and observe.
5.1.2.4  Observations:
5.1.2.4.1        The sample in test tube 1 containing with water is practically insoluble.
5.1.2.4.2        The sample in test tube 2 containing with ethanol (96.0%) is soluble.
5.1.2.4.3        The sample in test tube 3 containing with acetonitrile is sparingly soluble.
5.2  Assay:
5.2.1        Apparatus:
5.2.1.1  Liquid chromatography apparatus.
5.2.1.2  Glassware (according to the requirement).
5.2.1.3  UV/VIS Spectrophotometer (detector).
5.2.1.4  Sonicator.
5.2.2        Material and reagents:
5.2.2.1  Silica gel for chromatography (5μm).
5.2.2.2  Misoprostol CRS.
5.2.2.3  Acetonitrile.
5.2.2.4  Dioxin.
5.2.2.5  Heptane.
5.2.3        Requirements:
5.2.3.1  Sample:
5.2.3.1.1        25.0mg
5.2.3.2  Test solution:
5.2.3.2.1        Test solution:
5.2.3.2.1.1  Take 50.0ml of beaker and dissolve 25.0mg of the substance to be examined in the mobile phase.
5.2.3.2.1.2  Dilute it to 5.0ml with the mobile phase.
5.2.3.3  Reference solutions:
5.2.3.3.1        Reference solution (a):
5.2.3.3.1.1  Take 100ml beaker and dissolve 25.0mg of Misoprostol CRS in the mobile phase.
5.2.3.3.1.2  Dilute it to 5.0ml with the mobile phase.
5.2.3.4  Column:
5.2.3.4.1        Size:
5.2.3.4.1.1  Length=0.25m,
5.2.3.4.1.2  θ=4.6mm.
5.2.3.4.2        Stationary phase:
5.2.3.4.2.1  Silica gel for chromatography (5μm).
5.2.3.5  Mobile phase:
5.2.3.5.1        Mix 5 volumes of acetonitrile, 215 volumes of dioxin and 780 volumes of heptane and sonicate for 10min.
5.2.3.6  Flow rate:
5.2.3.6.1        2ml/min.
5.2.3.7  Detection:
5.2.3.7.1        Spectrophotometer at 210nm.
5.2.3.8  Injection:
5.2.3.8.1        20μL of the test solution and reference solution (a).
5.2.3.9  Run time:
5.2.3.9.1        1.5 times the retention time of the 1st peak of misoprostol.
5.2.3.10    Retention time:
5.2.3.10.1    1.5 times the retention time of misoprostol.
5.2.3.11    System suitability
5.2.3.11.1    Reference solution (a):
5.2.3.11.1.1    Symmetry factor: maximum 3.7 for the peak due to misoprostol.
5.2.4        Method of analysis:
5.2.4.1  Firstly prepare the test solution, reference solution and mobile phase according to the requirements.
5.2.4.2  The solutions must be free from solid particles.
5.2.4.3  Prepare the apparatus.
5.2.4.4  The mobile phase solvent mixtures must be deaerated prior to use either by boiling or by applying a partial vacuum to the solvent reservoir.
5.2.4.5  Equilibrate the column with the prescribed mobile phase, flow rate and at temperature specified until a suitable baseline is achieved.
5.2.4.6  Test solution of the mixture to be separated is now introduced into the mobile phase with the help of an injector just before entering the separating column.
5.2.4.7  As the eluate leaves the column it enters a detector, where it is continuously monitored at the specified λ.
5.2.4.8  The electrical signal obtained from detector is amplified and routes to recorder which record the developed chromatogram.
5.2.4.9  Calculate the percentage content of Misoprostol (C22H38O5) using the declared content of misoprostol CRS.
5.2.5        Limit:
5.2.5.1  96.5% to 102.0% (anhydrous substance).
6.0  REVISION LOG:
Revision No.
Effective Date
Reason
00

New SOP

7.0  REFERENCES:
7.1  The British Pharmacopoeia. Vol II., Official Monograph /Misoprostol: 2015, pp. 298-299.
8.0  ANNEXURES:
Annexure 1: Observations and calculations of HPLC method.






Annexure: 1
Observations and calculations of HPLC method
Analysis on HPLC
Instrument: ___________________                                           Date: _________________
Model: ___________________
Column size:
Length=
θ=
Stationary phase:

Temperature:

Mobile phase:

Flow rate:

Injection size:

Detector:

Wavelength:
λ=

Sample solution: _______________________
Reference standard solution: ______________
Impurities: ____________________________
(calculate each component calculation separately)
OBSERVATIONS:
Attach chromatogram.








CALCULATIONS:
1.      Retention time:                                                                                n= no. of peak
Retention time of unretained peak (tm)= _____________
No. of peaks
Retention time of peak of interest
(tr)n
Height of peak of interest
(h)n
Width of peak of interest
(w)n
Area of peak of interest
A=1/2(h x w)




















2.      Retention volume:
Flow rate= _______________ml/min.
No. of peaks
Retention time of peak of interest
(tr)n
Retention volume = retention time x flow rate












3.      Retention factor:
Retention time of unretained peak (tm)= _____________
No. of peaks
Retention time of peak of interest
(tr)n
Retention factor of a component
k= (tr-tm)/tm














4.      Separation factor (α):
No. of peaks
Retention factor of a component
(kn)
Relative retention of two adjacent peaks
α = k2/k1












5.      Resolution:
Retention time of unretained peak (tm)= _____________
No. of peaks
Retention time of peak of interest
(tr)n
Width of peak of interest
(w)n
Resolution
Rs = 2 (tr2-tr1)
        (w1-w2)
















6.      Efficiency:
No. of peaks or components
Retention time of peak of interest
(tr)n
Width of peak of interest
(w)n
Efficiency
(No. of theoretical plates)
N= 16 (tr/w)2



















7.      Height equivalent to a theoretical plate (HETP):
Length of column = ________________________
No. of peaks or components
No. of theoretical plates
(N)
Height equivalent to a theoretical plate HETP = L/N












8.      Symmetry factor (tailing factor):
No. of peaks or components
Distance from the peak max. to leading edge of the peak
(f)
Width w
Symmetry factor
At 5%
At 10%
As = w5%
       2f
As = w10%
       2f
























9.      Response factor & Relative response factor:
Conc. (mg/ml)= ___________________
No. of peak
Peak area
Response factor = (peak area/conc.)
Relative response factor = (response factor of impurity/response factor of API)


















10.  Relative standard deviation (%RSD):
Use formula of relative standard deviation where it is required i.e.,
PIC
11.  Percentage of content:
Percentage content = (rU/rS) x (CS/CU) x 100.
rU= peak response of substance from the sample solution.
rS= peak response of substance from the standard solution.
CS= concentration of substance in the standard solution (mg/mL).
CU= concentration of substance in the sample solution (mg/mL).


RESULTS:
________________________________________________________________________________________________________________________________________________











9.0  ABBREVIATIONS:
Abbreviation
Expanded Form
SOP
Standard operating procedure
&
And
No.
Number  
Ltd.
Limited
Sr.#
Serial number
Q.C
Quality control
%
Percentage
B.P
British pharmacopoeia
UV/VIS
Ultraviolet/ visible
μm
Micron
CRS
Chemical reference substance
mg
Milligram
ml
Milliliter
m
Meter
θ
Theta
mm
Millimeter
λ
Lambda
Min
Minutes
ml/min
Milliliter per minutes
μL
Microliter
nm
Nanometer
Vol
Volume
QCA
Quality control active ingredient
F
Format


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