1.0 OBJECTIVE:
To
lay down a procedure for the active raw material of the Cycloserine USP 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 Identification
test:
5.1.1
Material
and equipment:
5.1.1.1 Glassware (2 test tubes, 1 spatula, 1
beaker).
5.1.1.2 Analytical weighing balance.
5.1.1.3 0.1N sodium hydroxide.
5.1.1.4 3.0ml of 1N acetic acid.
5.1.1.5 Sodium nitroprusside solution.
5.1.1.6 4N sodium hydroxide.
5.1.2
Sample:
5.1.2.1 1.0mg.
5.1.3
Method:
5.1.3.1 Take a test tube add in it about 1.0mg in
10.0ml of 0.1N sodium hydroxide. Dissolve it thoroughly.
5.1.3.2 Take 1.0ml of this resulting solution in
another test tube and add 3.0ml of 1N acetic acid and 1.0ml of the mixture,
prepared 1 hour before use, of equal parts of sodium nitroprusside solution (1
in 25) and 4N sodium hydroxide.
5.1.3.3 Observe the changes.
5.1.4
Observations:
5.1.4.1 A blue colour gradually develops.
5.2 Specific
rotation:
5.2.1
Material
and equipment:
5.2.1.1 Polarimeter.
5.2.1.2 Analytical weighing balance.
5.2.1.3 Glassware (1 beaker of 50.0ml, 1 stirrer,
1 spatula).
5.2.1.4 Methanol.
5.2.2
Sample:
5.2.2.1 50mg/ml.
5.2.3
Method:
5.2.3.1 Prepare test solution in a beaker of
100.0ml with 50mg/ml, in 2N sodium hydroxide.
5.2.3.2 Firstly clean the Polarimeter with clean
dry cloth,
5.2.3.3 Operate the Polarimeter.
5.2.3.4 Fill the Polarimeter tube with blank
solution (2N sodium hydroxide) and determine the observed optical rotation.
5.2.3.5 Similarly, fill the Polarimeter tube with
sample test solution and determine the observed optical rotation.
5.2.3.6 Note down the values in annexure-1.
5.2.3.7 Calculate the specific optical rotation by
using formula:
[α]λ T = α/lc
5.2.4
Observations:
5.2.4.1 -108o to -114o.
5.3 pH:
5.3.1
Material
and equipment:
5.3.1.1 Glassware (according to the requirement).
5.3.1.2 pH meter.
5.3.2
Sample:
5.3.2.1 Solution (1 in 10).
5.3.3
Method:
5.3.3.1 Firstly clean the pH meter with clean dry
cloth, according to SOP No.
5.3.3.2 Operate the pH meter according to SOP for
operation of pH meter i.e.
5.3.3.3 Rejuvenate the electrode before use
according to SOP instructions, if there is any need of.
5.3.3.4 Calibrate the electrode of the pH meter
according to SOP No.
5.3.3.5 Perform the test on solution (1 in 10).
5.3.3.6 Take a beaker of 100.0ml and add solution
(1 in 10) in it. Such that it immersed electrodes in it completely.
5.3.3.7 Maintain the temperature of sample at 25oC±2oC.
5.3.3.8 Dip the electrode along with temperature
sensor into the sample.
5.3.3.9 When dipping electrode into sample, it
must be completely immersed in it.
5.3.3.10 Stir the probe gently in the sample to
create a homogeneous sample.
5.3.3.11 Allow the reading to stabilize for a time.
5.3.3.12 Record the observed values of pH &
temperature in the respective Annexure-2.
5.3.3.13 Wash the electrodes again after use and
store the electrode in storage solution as recommended in SOP of cleaning i.e.. Calibrate the instrument
before use.
5.3.4
Observation:
5.3.4.1 Between 5.5 and 6.5.
5.4 Assay:
5.4.1
Apparatus:
5.4.1.1 Glassware (according to requirement).
5.4.1.2 Magnetic stirrer.
5.4.1.3 HPLC apparatus.
5.4.1.4 Analytical weighing balance.
5.4.2
pH 6.8 Phosphate buffer:
5.4.2.1 Prepare pH 6.8 phosphate buffer as given
in SOPs of reagents.
5.4.3
Mobile phase:
5.4.3.1 Take a beaker of 1000.0ml and add 0.5g of
sodium 1-decanesulfonate in 800 ml of purified water. Dissolve it by using magnetic
stirrer operate according to SOP
5.4.3.2 Add 50.0ml of acetonitrile and 5 ml of
glacial acetic acid, and mix it again by using magnetic stirrer operate
according to SOP
5.4.3.3 Adjust with 1N sodium hydroxide to pH of
4.4.
5.4.3.4 Filter, and degas. Make adjustment if
necessary.
5.4.4
Standard preparation:
5.4.4.1 Quantitatively dissolve an accurately
weighed quantity of USP Cycloserine RS in pH 6.8 phosphate buffer to obtain a
solution having a known concentration of about 0.4mg per ml.
5.4.5
Assay preparation:
5.4.5.1 Take a volumetric flask of 50.0ml and
transfer about 20.0mg of Cycloserine, accurately weighed.
5.4.5.2 Dissolve it in sufficient quantity of pH 6.8
phosphate buffer by using magnetic stirrer operate according to SOP
5.4.5.3 Dilute it with the same solution to the
volume and mix it by using magnetic stirrer operate according to SOP
5.4.6
Chromatographic system: The
liquid chromatography is equipped with:
5.4.6.1 Mode:
Liquid chromatography.
5.4.6.2 Detector:
219nm.
5.4.6.3 Column:
4.6-mm × 25-cm column that contains 5-μm packing
L1.
5.4.6.4 Column
temperature: Maintained at about 30o
5.4.6.5 Flow
rate: 1.0mL/min.
5.4.6.6 Chromatograph the standard preparation,
and record the peak responses as directed for procedure
5.4.6.6.1
The
tailing factor is not more than 1.8.
5.4.6.6.2
The
relative standard deviation for replicate injections is not more than 2.0%.
5.4.7
Procedure:
5.4.7.1 Equilibrate the column and detector with
mobile phase at specified flow rate until a constant signal is received.
5.4.7.2 Separately inject equal volumes (about
10μL) of the standard preparation and the Assay preparation into the spectrum.
5.4.7.3 Record the spectrum.
5.4.7.4 Measure the peak responses for Cycloserine.
5.4.7.5 Analyze as directed in the monograph.
5.4.7.6 Calculate the quantity, in μg, of C3H6N2O2
in each mg of Cycloserine taken by the formula:
50,000
(C/W) (rU/rS)
In
which,
C
= the concentration, in mg per ml, of USP Cycloserine RS in the standard
preparation.
W
= the quantity, in mg, of Cycloserine taken to prepare the assay preparation.
rU
and rS = are the peak responses for Cycloserine obtained from the
assay preparation and the standard preparation, respectively.
5.4.8
Limit:
5.4.8.1 Cycloserine has a potency of not less than
900μg of C3H6N2O2.
6.0 REVISION
LOG:
Revision No.
|
Effective Date
|
Reason
|
00
|
New
SOP
|
7.0 REFERENCES:
7.1 USP38NF33
Volume-3 Official Monograph/ Cycloserine: 2015, pp.: 2980.
8.0 ANNEXURES:
Annexure 1: Observations
and calculations of specific optical rotation.
Annexure 2:
pH measurement.
Annexure 3:
Observations and calculations of HPLC method.
Annexure:
1
Observations
and calculations of specific optical rotation
Instrument:
___________________
Date: _______________
Model: _______________________ Length of Polarimeter tube:
________________
Sample:
________________________________g.
Solvent:
________________________________ml.
Concentration of
sample solution: ____________g/ml.
Blank solution:
Sr.#
|
Blank solution
|
Temperature
|
Optical rotation
(α)
|
Average: _______________
Optical rotation
of blank solution: _______________
Sample solution:
Sr.#
|
Sample solution
|
Temperature
|
Optical rotation
(α)
|
Average: _______________
Optical rotation
of sample solution: ______________
Optical
rotation of substance = Blank solution - Sample solution.
Specific optical rotation
of sample solution by using formula:
[α]λ T = α/lc
Result: ________________
Remarks: ___________________________________________________________
Annexure:
2
pH
measurement
Sr.#
|
Temperature
|
pH
|
Result: _________________________________________________________________
Annexure:
3
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
spectrum.
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
|
QCA
|
Quality
control active ingredient
|
F
|
Format
|
Q.C
|
Quality
control
|
Vol
|
Volume
|
g
|
Grams
|
ml
|
Milliliter
|
Min
|
Minutes
|
oC
|
Degree
centigrade
|
mg
|
Milligram
|
M
|
Molar
|
%
|
Percentage
|
g/L
|
Gram
per liter
|
N
|
Normal
|
USP
|
United
states pharmacopoeia
|
NF
|
National
formulary
|
cm
|
Centimeter
|
nm
|
Nanometer
|
mm
|
Millimeter
|
Approx.
|
Approximately
|
%
|
Percentage
|
μL
|
Microliter
|
θ
|
Theta
|
λ
|
Lambda
|
w/v
|
Weight
by volume
|
mg/ml
|
Milligram
per milliliter
|
UV
|
Ultraviolet
|
ml/min
|
Milliliter
per minute
|
o
|
Degree
(angle)
|
l
|
Length
|
g
|
Grams
|
c
|
Concentration
(g/ml)
|
g/ml
|
Gram
per milliliter
|
α
|
Alpha
|
T
|
Temperature
|
No comments:
Post a Comment