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EUROPEAN PHARMACOPOEIA 8.

Diosmin

01/2015:1611 Related substances. Liquid chromatography (2.2.29).


Test solution. Dissolve 25.0 mg of the substance to be
examined in dimethyl sulfoxide R and dilute to 25.0 mL with
DIOSMIN
the same solvent.
Reference solution (a). Dissolve 25.0 mg of diosmin CRS in
Diosminum
dimethyl sulfoxide R and dilute to 25.0 mL with the same
solvent.
Reference solution (b). Dilute 1.0 mL of the test solution to
100.0 mL with dimethyl sulfoxide R.
Reference solution (c). Dissolve 5.0 mg of diosmin for system
suitability CRS (containing impurities A, B, C, D, E and F)
in dimethyl sulfoxide R and dilute to 5.0 mL with the same
solvent.
C28H32O15
Mr 609 Column :
[520-27-4]
size : l = 0.10 m, = 4.6 mm ;
stationary phase : end-capped octadecylsilyl silica gel for
DEFINITION
chromatography R (3 m) ;
7-[[6-O-(6-Deoxy--L-mannopyranosyl)--D temperature : 40 C.
glucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4methoxyphenyl)-4H-1-benzopyran-4-one.
Mobile phase : acetonitrile R, glacial acetic acid R, methanol R,
water R (2:6:28:66 V/V/V/V).
Substance obtained through iodine-assisted oxidation
of (2S)-7-[[6-O-(6-deoxy--L-mannopyranosyl)-Flow rate : 1.5 mL/min.
D-glucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4Detection : spectrophotometer at 275 nm.
methoxyphenyl)-2,3-dihydro-4H-1-benzopyran-4-one
Injection : 10 L of the test solution and reference solutions (b)
(hesperidin) of natural origin.
and (c).
Content : 90.0 per cent to 102.0 per cent (anhydrous substance).
Run time : 6 times the retention time of diosmin.
CHARACTERS
Identification of impurities : use the chromatogram supplied
with diosmin for system suitability CRS and the chromatogram
Appearance : greyish-yellow or light yellow, hygroscopic
obtained with reference solution (c) to identify the peaks due
powder.
to impurities A, B, C, D, E and F.
Solubility : practically insoluble in water, soluble in dimethyl
Relative retention with reference to diosmin (retention
sulfoxide, practically insoluble in ethanol (96 per cent). It
time = about 4 min) : impurity A = about 0.5 ;
dissolves in dilute solutions of alkali hydroxides.
impurity B = about 0.6 ; impurity C = about 0.8 ;
IDENTIFICATION
impurity D = about 2.2 ; impurity E = about 2.6 ;
impurity F = about 4.5.
A. Infrared absorption spectrophotometry (2.2.24).
System suitability : reference solution (c) :
Comparison : diosmin CRS.
resolution : minimum 2.5 between the peaks due to
B. Examine the chromatograms obtained in the assay.
impurities B and C.
Results : the principal peak in the chromatogram obtained
Calculation of percentage contents :
with the test solution is similar in retention time and size
to the principal peak in the chromatogram obtained with
correction factors : multiply the peak areas of the following
reference solution (a).
impurities by the corresponding correction factor :
impurity A = 0.4 ; impurity F = 0.6 ;
TESTS
for each impurity, use the concentration of diosmin in
Iodine : maximum 0.1 per cent.
reference solution (b).
Determine the total content of iodine by potentiometry, using Limits :
an iodide-selective electrode (2.2.36), after oxygen combustion impurity B : maximum 4.0 per cent ;
(2.5.10).
impurities C, E : for each impurity, maximum 3.0 per cent ;
Test solution. Wrap 0.100 g of the substance to be examined in
a piece of lter paper and place it in a sample carrier. Introduce impurity F : maximum 2.0 per cent ;
into the ask 50 mL of a 0.2 g/L solution of hydrazine R. Flush impurity D : maximum 0.6 per cent ;
impurity A : maximum 0.5 per cent ;
the ask with oxygen for 10 min. Ignite the lter paper. Stir
the contents of the ask immediately after the end of the
unspecified impurities : for each impurity, maximum 0.4 per
combustion to dissolve completely the combustion products.
cent ;
Continue stirring for 1 h.
total : maximum 8.5 per cent ;
Reference solution. Dilute 2.0 mL of a 16.6 g/L solution of
reporting threshold : 0.10 per cent.
potassium iodide R to 100.0 mL with water R. Dilute 10.0 mL
The thresholds indicated under Related substances
of this solution to 100.0 mL with water R.
(Table 2034.-1) in the general monograph Substances for
Introduce into a beaker 30 mL of a 200 g/L solution of
pharmaceutical use (2034) do not apply.
potassium nitrate R in 0.1 M nitric acid. Immerse the
Heavy metals (2.4.8): maximum 20 ppm.
electrodes and stir for 10 min. The potential of the solution
2.0 g complies with test C. Prepare the reference solution using
(nT1) must remain stable. Add 1 mL of the test solution and
4.0 mL of lead standard solution (10 ppm Pb) R.
measure the potential (nT2).
Introduce into a beaker 30 mL of a 200 g/L solution of
Water (2.5.12) : maximum 6.0 per cent, determined on 0.300 g.
potassium nitrate R in 0.1 M nitric acid. Immerse the
Sulfated ash (2.4.14): maximum 0.2 per cent, determined on
electrodes and stir for 10 min. The potential of the solution
1.0 g.
must remain stable (nR1). Add 80 L of the reference solution
and measure the potential (nR2).
ASSAY
The absolute value |nT2 - nT1| is not higher than the absolute Liquid chromatography (2.2.29) as described in the test for
value |nR2 - nR1|.
related substances with the following modication.
General Notices (1) apply to all monographs and other texts

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Diosmin

EUROPEAN PHARMACOPOEIA 8.3

Injection : test solution and reference solution (a).


Calculate the percentage content of C28H32O15 taking into
account the assigned content of diosmin CRS.
STORAGE
In an airtight container.
IMPURITIES
Specified impurities : A, B, C, D, E, F.
Other detectable impurities (the following substances would,
if present at a sufcient level, be detected by one or other of
the tests in the monograph. They are limited by the general
acceptance criterion for other/unspecied impurities. It
is therefore not necessary to identify these impurities for
demonstration of compliance. See also 5.10. Control of
impurities in substances for pharmaceutical use) : G.

A. 1-(3-hydroxy-4-methoxyphenyl)ethan-1-one
(acetoisovanillone),

B. (2S)-7-[[6-O-(6-deoxy--L-mannopyranosyl)--Dglucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4methoxyphenyl)-2,3-dihydro-4H-1-benzopyran-4-one
(hesperidin),

C. 7-[[6-O-(6-deoxy--L-mannopyranosyl)--Dglucopyranosyl]oxy]-5-hydroxy-2-(4-hydroxyphenyl)-4H1-benzopyran-4-one (isorhoifolin),

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D. 7-[[6-O-(6-deoxy--L-mannopyranosyl)--Dglucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4methoxyphenyl)-6-iodo-4H-1-benzopyran-4-one
(6-iododiosmin),

E. 7-[[6-O-(6-deoxy--L-mannopyranosyl)--Dglucopyranosyl]oxy]-5-hydroxy-2-(4-methoxyphenyl)-4H1-benzopyran-4-one (linarin),

F. 5,7-dihydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1benzopyran-4-one (diosmetin),

G. (2S)-5,7-dihydroxy-2-(3-hydroxy-4-methoxyphenyl)-2,3dihydro-4H-1-benzopyran-4-one (hesperetin).

See the information section on general monographs (cover pages)