1.0 OBJECTIVE:
To
lay down a procedure of analytical report for the active raw material of the Amlodipine
besilate from the Pharmacopoeial specifications.
2.0 SCOPE:
This
SOP shall be applicable in Q.C laboratory.
3.0 RESPONSIBILITY:
3.1 Q.C Analysts.
4.0 ACCOUNTABILITY:
4.1 Q.C Manager.
5.0 PROCEDURE:
5.1 Characters:
5.1.1
Appearance:
5.1.1.1 White or almost white powder.
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
Purified
water.
5.1.2.1.3
Methanol.
5.1.2.1.4
Anhydrous
ethanol.
5.1.2.1.5
2-propanol.
5.1.2.2 Sample:
5.1.2.2.1
Small
quantity.
5.1.2.3 Method:
5.1.2.3.1
Take
4 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
methanol in test tube 2 and observe.
5.1.2.3.4
Add
anhydrous ethanol in test tube 3 and observe.
5.1.2.3.5
Add
2-propanol in test tube 4 and observe.
5.1.2.4 Observations:
5.1.2.4.1
The
sample in test tube 1 containing with water is slightly soluble.
5.1.2.4.2
The
sample in test tube 2 containing with methanol is freely soluble.
5.1.2.4.3
The
sample in test tube 3 containing with anhydrous ethanol is sparingly soluble.
5.1.2.4.4
The
sample in test tube 4 containing with 2-propanol is slightly 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).
5.2.2
Material and reagents:
5.2.2.1 Octadecylsilyl silica gel for
chromatography (5μm).
5.2.2.2 2.3g/L solution of ammonium acetate.
5.2.2.3 Methanol.
5.2.3
Requirements:
5.2.3.1
Sample:
5.2.3.1.1
50.0mg
(sample to be examined).
5.2.3.1.2
50.0mg
(amlodipine besilate CRS) for reference solution.
5.2.3.2
Test solution:
5.2.3.2.1
Test
solution (a):
5.2.3.2.1.1 Take 100ml of beaker and dissolve 50.0mg
of the substance to be examined in the mobile phase.
5.2.3.2.1.2 And dilute to 50.0ml with the mobile
phase.
5.2.3.2.2
Test
solution (b):
5.2.3.2.2.1 Take a beaker of 100ml and dilute 5.0ml of
test solution (a) to 100.0ml with mobile phase.
5.2.3.3
Reference solutions:
5.2.3.3.1
Reference
solution (e):
5.2.3.3.1.1 Take 100ml beaker and dissolve 50.0mg of
amlodipine besilate CRS in the mobile phase.
5.2.3.3.1.2 Dilute 5.0ml of the solution to 100.0ml
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=4.0mm.
5.2.3.4.2
Stationary phase:
5.2.3.4.2.1 Octadecylsilyl silica gel for
chromatography (5μm).
5.2.3.4.3
Temperature:
5.2.3.4.3.1 30oC.
5.2.3.5
Mobile phase:
5.2.3.5.1
2.3g/L
solution of ammonium acetate, methanol (30:70 v/v).
5.2.3.6
Flow rate:
5.2.3.6.1
1.5ml/min.
5.2.3.7
Detection:
5.2.3.7.1
Spectrophotometer
at 237nm.
5.2.3.8
Injection:
5.2.3.8.1
20μL
of the test solution (b) and reference solution (e).
5.2.3.9
Run time:
5.2.3.9.1
Twice
the retention time of amlodipine.
5.2.3.10 Relative
retention:
5.2.3.10.1 With reference to amlodipine (retention
time=about 20min).
5.2.4
Method of analysis:
5.2.4.1 Carry out the test protected from light.
5.2.4.2 Firstly prepare the test solution,
reference solution and mobile phase according to the requirements.
5.2.4.3 The solutions must be free from solid
particles.
5.2.4.4 Prepare the apparatus.
5.2.4.5 The mobile phase solvent mixtures must be
deaerated prior to use either by boiling or by applying a partial vacuum to the
solvent reservoir.
5.2.4.6 Equilibrate the column with the prescribed
mobile phase, flow rate and at temperature specified until a suitable baseline
is achieved.
5.2.4.7 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.
5.2.4.8 As the eluate leaves the column it enters
a detector, where it is continuously monitored at the specified λ.
5.2.4.9 The electrical signal obtained from
detector is amplified and routes to recorder which record the developed
chromatogram.
5.2.4.10 Calculate the percentage content of
Amlodipine besilate (C26H31ClN2O8S)
from the declared content of amlodipine besilate CRS.
5.2.5
Limit:
5.2.5.1 97% to 102% (anhydrous substance).
6.0 REVISION
LOG:
Revision No.
|
Effective Date
|
Reason
|
00
|
New
SOP
|
7.0 REFERENCES:
7.1 The British Pharmacopoeia. Vol I., Official Monograph /Amlodipine
besilate: 2015, pp. 53-54.
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:
___________________
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)= _____________
2.
Retention volume:
Flow
rate= _______________ml/min.
3.
Retention factor:
Retention
time of unretained peak (tm)= _____________
4.
Separation factor (α):
5.
Resolution:
Retention
time of unretained peak (tm)= _____________
6.
Efficiency:
7.
Height equivalent to a theoretical plate
(HETP):
Length
of column = ________________________
8.
Symmetry factor (tailing factor):
9.
Response factor & Relative response
factor:
Conc.
(mg/ml)= ___________________
10. Relative
standard deviation (%RSD):
Use
formula of relative standard deviation where it is required i.e.,
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
|
Q.C
|
Quality
control
|
B.P
|
British
pharmacopoeia
|
μm
|
Micrometer
|
g/L
|
Gram
per liter
|
mg
|
Milligram
|
CRS
|
Chemical
reference solution
|
ml
|
Milliliter
|
m
|
Meter
|
θ
|
Theta
|
mm
|
Millimeter
|
oC
|
Degree
centigrade
|
v/v
|
Volume
by volume
|
ml/min
|
Milliliter
per minute
|
nm
|
Nanometer
|
μL
|
Microliter
|
Min
|
Minute
|
λ
|
Lambda
|
%
|
Percentage
|
UV/VIS
|
Ultraviolet/
visible
|
Vol
|
Volume
|
QCA
|
Quality
control active ingredient
|
F
|
Format
|