Protocol
and Analysis of a SKIN-CAP Spray Sample
to Test for the Presence of Clobetasol PropionateApplicant:
KOMVET spol.s r.o.
Sadová 576
675 21 Ok_í_ky
Sample:
SKIN-CAP Spray 100 ml, batch M-10, expiry SEP-03
The sample was handed over to the laboratory on March 9, 1999.
ANALYTICAL
TECHNIQUE
a) preanalytical condition (preparation of the sample before analysis)
The sample was stored at room temperature in the laboratory. Before
handling, it was shaken well. Afterwards, through the applicator which
is an integral part of the spray, some 10 ml were let out into a small
container. Then there followed the degasification of the sample in
ultrasound and the filtration through microfilter (filter units 25
mm - teflon, 5 mm PTFE - fa Shandon).
b) analytical method HPLC
Equipment used:liquid chromatograph HP 1100 with detector DAD (Hewlett-Packard)
Column:LiChroSpher C 18 (250x4,5 mm, Merck) with precolumn (4x4,5
mm)
Eluant:methanol/water (0 - 15 min. 65 % MeOH, 15 - 35 min. 65 - 100
% MeOH)
Flow rate:1 ml/min.
Detection:242 nm
Column temperature:20 ºC
Flow temperature:20 ºC
Injection volume:20 ml
c) analytical standard
Clobetasol propionate CAS: 25122-46-7 (purity 99 %, Sigma)
Concentration of reserve liquid: 1 mg/ml in methanol
Using decimal dilution, there was prepared a five-point scale of standard
solutions (with concentrations between 0.0001 - 1 mg/ml).GRAPHIC EVIDENCE
OF THE TESTS
In order to analyse the sample, an analytical method was developed
and validated.
The following steps were taken (see graphic evidence enclosed):
1. The standard absorption spectrum has been measured (graph 1). For
the proper measurement, there has been determined the correspondingwavelength
242 nm with the maximum absorption.
2. HLPC separation has been optimized (graph 2).
3. There has been determined the extension of the repercussion of
the detector (0.0001 - 1 mg/ml) (graph 3 - calibration).
4. The value of the detection limit is 0.1 mg/ml (graph 4).
5. examination of the real SKIN-CAP Spray sample (100 ml, batch M-10,
expiry SEP-03) (graph 5).
RESULTS
No indication has been found of the presence of CLOBETASOL PROPIONATE
in the preparation SKIN-CAP Spray analysed under the above-mentioned
conditions.
Analysis
made by:Ing. J. Guziur
Consultant:Dr. Ing. V. Schulzova Prof. Ing. J. Haj_lova, CSc
Head of the LaboratoryPrague, 20.03.1999
