Isometamidium chloride hydrochloride (Finished product) Assay (By HPLC)
Chromatographic
condition:
Column - C8 (125 x 4.5 mm) 5µ or
equivalent
Flow Rate - 1.0 ml / minute.
Wavelength - 320 nm
Injection Volume - 20 µl
|
Peak Number |
Retention Time |
Relative Retention Time |
|
I |
3.8 |
0.6 |
|
II |
5.5 |
0.87 |
|
III |
6.3 |
1 |
|
IV |
7.5 |
1.17 |
|
V |
15.9 |
2.52 |
Mobile Phase - Buffer: Acetonitrile (77:23)
Buffer Preparation: Dissolve 2.7 g of Potassium Dihydrogen Phosphate in 950ml distil water. Adjust pH to 2.5 with orthophosphoric acid and dilute to 1000 ml with distilled water.
Standard Preparation
Take 100 mg of Isometamidium chloride hydrochloride WS in 100 ml volumetric flask and dilute to 100 ml with 30 % v/v solution of Acetonitrile in water. Stir for 15 minutes on magnetic stirrer to dissolve it completely.
Sample
Preparation
Take sample equivalent to 100 mg of Isometamidium
chloride hydrochloride in 100 ml volumetric flask and dilute to 100 ml with 30%
v/v solution of Acetonitrile in water.
Stir for 15 minutes on magnetic stirrer to dissolve it completely.
Procedure: Inject one blank,
five replicate injection of standard solution, two replicate injection of
sample solution & two bracketing standard solution.
System Suitability:
1) RSD of five replicate injection of
standard solution should not be more than 2.0%
2) RSD of five replicate injection of standard
solution and two bracketing standard solution should not be more than 2.0%
Calculation
Content of Isometamidium chloride hydrochloride in mg/
Mean
peak area of the Spl (sp1+sp2+sp3+sp4+sp5) x weight
of std. taken x100 x Std. Purity of WS x av. Net content
= -------------------------------------------------------------------------------
Mean peak area of standard (std1+std2+std3+std4+std5)
x
100 x weight of sample taken x100
% of Isometamidium chloride hydrochloride
Isometamidium chloride hydrochloride in mg
=---------------------------------------------------------------x
100
Label claim
Where,
sp1 =
Area of sample peak I
sp2 =
Area of sample peak II
sp3 =
Area of sample peak III
sp4 =
Area of sample peak IV
sp5 =
Area of sample peak V
std1 =
Area of standard peak I
std2 =
Area of standard peak II
std3 =
Area of standard peak III
std4 = Area of standard peak IV
std5 = Area of standard peak V
