EUROPEAN PHARMACOPOEIA 5.

0 Clazuril for veterinary use

Time Mobile phase A Mobile phase B C. R = NH2 : (2RS)-2-[2-chloro-4-(3,5-dioxo-4,5-dihydro-1,2,4-

(min) (per cent V/V) (per cent V/V) triazin-2(3H)-yl)phenyl]-2-(4-chlorophenyl)acetamide,
0 - 20 100 → 0 0 → 100
20 - 25 0 100
25 - 30 0 → 100 100 → 0
30 - 40 100 0

Flow rate : 1.0 ml/min.

Detection : spectrophotometer at 230 nm.
Injection : 5 µl.
System suitability : reference solution (a) :
— peak-to-valley ratio : minimum 1.5, where Hp = height B. R = NH2 : 2-[3-chloro-4-[(RS)-(4-chlorophenyl)cyanometh-
above the baseline of the peak due to impurity G yl]phenyl]-3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-car-
and Hv = height above the baseline of the lowest point boxamide,
of the curve separating this peak from the peak due to
clazuril, D. R = N(CH3)2 : 2-[3-chloro-4-[(RS)-(4-chlorophenyl)cya-
— the chromatogram obtained is concordant with the nomethyl]phenyl]-N,N-dimethyl-3,5-dioxo-2,3,4,5-tetra-
chromatogram supplied with clazuril for system hydro-1,2,4-triazine-6-carboxamide,
suitability CRS.
Limits : E. R = OCH3 : methyl 2-[3-chloro-4-[(RS)-(4-
chlorophenyl)cyanomethyl]phenyl]-3,5-dioxo-2,3,
— correction factors : for the calculation of contents, 4,5-tetrahydro-1,2,4-triazine-6-carboxylate,
multiply the peak areas of the following impurities by
the corresponding correction factor : impurity G = 1.4 ;
impurity H = 0.8 ; F. R = OC2H5 : ethyl 2-[3-chloro-4-[(RS)-(4-
chlorophenyl)cyanomethyl]phenyl]-3,5-dioxo-2,3,
— any impurity : not more than the area of the principal 4,5-tetrahydro-1,2,4-triazine-6-carboxylate,
peak in the chromatogram obtained with reference
solution (b) (0.2 per cent) ;
— total: not more than 3 times the area of the principal peak
in the chromatogram obtained with reference solution (b)
(0.6 per cent) ;
— disregard limit : 0.25 times the area of the principal peak
in the chromatogram obtained with reference solution (b)
(0.05 per cent) ; disregard the peaks due to the solvents.
Loss on drying (2.2.32) : maximum 0.5 per cent, determined
on 1.000 g by drying in an oven at 100-105 °C for 4 h. G. 2-[3-chloro-4-(4-chlorobenzoyl)phenyl]-1,2,4-triazine-3,
5(2H,4H)-dione,
Sulphated ash (2.4.14) : maximum 0.1 per cent, determined
on 1.0 g.

ASSAY
Dissolve about 0.260 g in 35 ml of tetrahydrofuran R and
add 35 ml of water R. Titrate with 0.1 M sodium hydroxide,
determining the end-point potentiometrically (2.2.20). Carry
out a blank titration.
1 ml of 0.1 M sodium hydroxide is equivalent to 37.32 mg
of C17H10Cl2N4O2.

STORAGE
Protected from light. H. [2-chloro-4-(3,5-dioxo-4,5-dihydro-1,2,4-triazin-2(3H)-
yl)phenyl][4-[[2-chloro-4-(3,5-dioxo-4,5-dihydro-1,2,
IMPURITIES 4-triazin-2(3H)-yl)phenyl]cyanomethyl]phenyl](4-
chlorophenyl)acetonitrile,

A. R = OH : (2RS)-[2-chloro-4-(3,5-dioxo-4,5-dihydro-1,2,4- I. (Z)-2-[[3-chloro-4-[(RS)-(4-chlorophenyl)cyanomethyl]phe-
triazin-2(3H)-yl)phenyl](4-chlorophenyl)acetic acid, nyl]diazanylidene]acetamide.

General Notices (1) apply to all monographs and other texts 1313
Clebopride malate EUROPEAN PHARMACOPOEIA 5.0

01/2005:1303 chromatogram obtained with the reference solution (a).

The identification is not valid unless the chromatogram
CLEBOPRIDE MALATE obtained with reference solution (b) shows two clearly
separated bands.

Clebopridi malas TESTS

Solution S. Dissolve 1.0 g in carbon dioxide-free water R
and dilute to 100.0 ml with the same solvent.
Appearance of solution. Examined immediately after
preparation, solution S is clear (2.2.1) and colourless (2.2.2,
Method I).
pH (2.2.3). The pH of solution S is 3.8 to 4.2.
C24H30ClN3O7 Mr 508.0 Related substances. Examine by liquid chromatography
(2.2.29).
DEFINITION Test solution. Dissolve 0.10 g of the substance to be
Clebopride malate contains not less than 98.5 per cent and examined in the mobile phase and dilute to 100.0 ml with
not more than the equivalent of 101.0 per cent of 4-amino- the mobile phase.
N-(1-benzylpiperidin-4-yl)-5-chloro-2-methoxybenzamide acid Reference solution (a). Dilute 1.0 ml of the test solution
(RS)-2-hydroxybutanedioate, calculated with reference to the to 100.0 ml with the mobile phase. Dilute 1.0 ml of this
dried substance. solution to 10.0 ml with the mobile phase.
CHARACTERS Reference solution (b). Dissolve 10.0 mg of clebopride
malate CRS and 10.0 mg of metoclopramide
A white or almost white, crystalline powder, sparingly hydrochloride CRS in the mobile phase and dilute to
soluble in water and in methanol, slightly soluble in ethanol, 100.0 ml with the mobile phase. Dilute 1.0 ml of this solution
practically insoluble in methylene chloride. to 10.0 ml with the mobile phase.
It melts at about 164 °C, with decomposition. The chromatographic procedure may be carried out using :
IDENTIFICATION — a stainless steel column 0.12 m long and 4.0 mm in
First identification : B, C. internal diameter packed with octadecylsilyl silica gel for
chromatography R (5 µm),
Second identification : A, C, D.
— as mobile phase at a flow rate of 1 ml/min a mixture of
A. Dissolve 20.0 mg in water R and dilute to 100.0 ml 20 volumes of acetonitrile R and 80 volumes of a 1 g/l
with the same solvent. Dilute 10.0 ml of the solution solution of heptane sulphonate sodium R adjusted to
to 100.0 ml with water R. Examined between 230 nm pH 2.5 with phosphoric acid R,
and 350 nm (2.2.25), the solution shows two absorption
maxima, at 270 nm and 307 nm. The specific absorbances — as detector a spectrophotometer set at 215 nm.
at the maxima are 252 to 278 and 204 to 226, respectively. Equilibrate the column with the mobile phase for 30 min.
Inject 20 µl of reference solution (b). Adjust the sensitivity
B. Examine by infrared absorption spectrophotometry
of the system so that the heights of the peaks in the
(2.2.24), comparing with the spectrum obtained with
chromatogram obtained are at least 30 per cent of the
clebopride malate CRS. Examine the substances prepared
full scale of the recorder. The test is not valid unless the
as discs.
retention time of the second peak (clebopride) is about
C. Dissolve 20 mg in 1 ml of sulphuric acid R, add 1 ml of 15 min and the relative retention time of the first peak is
β-naphthol solution R1 and mix. The solution examined about 0.45. Inject 20 µl of the test solution and 20 µl of
in daylight has a yellow colour with blue fluorescence. reference solution (a). Continue the chromatography of the
D. Examine by thin-layer chromatography (2.2.27), using test solution for twice the retention time of the principal
as the coating substance a suitable silica gel with a peak. In the chromatogram obtained with the test solution :
fluorescent indicator having an optimal intensity at the area of any peak, apart from the principal peak and the
254 nm. two peaks eluting within 2 min, is not greater than the area
Test solution. Dissolve 5 mg of the substance to be of the principal peak in the chromatogram obtained with
examined in ethanol R and dilute to 10 ml with the same reference solution (a) (0.1 per cent) ; the sum of the areas of
solvent. all peaks, apart from the principal peak and the two peaks
eluting within 2 min, is not greater than three times the area
Reference solution (a). Dissolve 5 mg of clebopride of the principal peak in the chromatogram obtained with
malate CRS in ethanol R and dilute to 10 ml with the reference solution (a) (0.3 per cent). Disregard any peak with
same solvent. an area less than 0.25 times that of the principal peak in the
Reference solution (b). Dissolve 5 mg of clebopride chromatogram obtained with reference solution (a).
malate CRS and 5 mg of metoclopramide
Chlorides. Prepare the solutions at the same time.
hydrochloride CRS in ethanol R and dilute to 10 ml with
the same solvent. Test solution. Dissolve 0.530 g of the substance to be
examined in 20.0 ml of anhydrous acetic acid R, add 6 ml of
Apply to the plate as bands 10 mm by 3 mm, 5 µl of each dilute nitric acid R and dilute to 50.0 ml with water R.
solution. Develop over a path of 15 cm using a mixture
of 2 volumes of concentrated ammonia R, 14 volumes of Reference solution. To 1.5 ml of 0.001 M hydrochloric acid,
acetone R, 14 volumes of methanol R and 70 volumes add 20.0 ml of anhydrous acetic acid R and 6 ml of dilute
of toluene R. Allow the plate to dry in air and examine nitric acid R and dilute to 50.0 ml with water R.
in ultraviolet light at 254 nm. The principal band in Transfer separately both solutions recently prepared to test
the chromatogram obtained with the test solution is tubes. Add to each tube 1 ml of silver nitrate solution R2.
similar in position and size to the principal band in the Allow to stand for 5 min protected from light. Examine the

1314 See the information section on general monographs (cover pages)