You are on page 1of 2

Dog rose

and anomocytic stomata (2.8.3) [Ga] ; trichomes are of 2


types : a) uniseriate covering trichomes with blunt apex,
usually consisting of 3-5 cells [H, J], often with 1 or more
collapsed cells [Ja], walls mostly nely warty or faintly
striated ; b) glandular trichomes usually with a unicellular
[C, D], sometimes a multicellular, uniseriate [A, B, E] stalk
and a unicellular head [A, B, C, E] or bicellular head, in
side view [D] and in surface view [F] or exceptionally a
tetracellular head.
C. Thin-layer chromatography (2.2.27).
Test solution. To 1.0 g of the powdered herbal drug
(180) (2.9.12) add a mixture of 20 mL of ethanol (50 per
cent V/V) R and 10 mL of lead acetate solution R. Boil for
2 min, allow to cool and centrifuge. Shake the supernatant
solution with 2 quantities, each of 15 mL, of chloroform R ;
separate the 2 layers by centrifugation if necessary. Dry
the chloroform layers over anhydrous sodium sulfate R and
lter. Evaporate 10 mL of the solution to dryness on a
water-bath and dissolve the residue in 1 mL of a mixture of
equal volumes of chloroform R and methanol R.
Reference solution. Dissolve 5 mg of purpureaglycoside A CRS, 2 mg of purpureaglycoside B CRS, 5 mg
of digitoxin R and 2 mg of gitoxin R in a mixture of
equal volumes of chloroform R and methanol R, then dilute
to 10 mL with the same mixture of solvents.
Plate : TLC silica gel G plate R.
Mobile phase : water R, methanol R, ethyl acetate R
(7.5:10:75 V/V/V).
Application : 20 L as bands of 2 cm by 0.3 cm.
Development : over a path of 10 cm.
Drying : until the solvents have evaporated.
Detection : treat with a mixture of 2 volumes of a 10 g/L
solution of chloramine R and 8 volumes of a 250 g/L
solution of trichloroacetic acid R in ethanol (96 per cent) R,
then heat at 100-105 C for 10 min ; examine in ultraviolet
light at 365 nm.
Results : the chromatogram obtained with the reference
solution shows a zone of light blue uorescence in the lower
part of the chromatogram, due to purpureaglycoside B, and,
just above it, a zone of brownish-yellow uorescence due
to purpureaglycoside A ; a zone of light blue uorescence,
due to gitoxin, appears in the middle of the chromatogram
and above it a zone of brownish-yellow uorescence, due
to digitoxin ; the zones in the chromatogram obtained with
the test solution are similar in position, colour and size to
the zones in the chromatogram obtained with the reference
solution. Other zones of uorescence may also appear in
the chromatogram obtained with the test solution.
D. Evaporate 5 mL of the chloroformic solution obtained in
identication test C to dryness on a water-bath. To the
residue add 2 mL of dinitrobenzoic acid solution R and
1 mL of 1 M sodium hydroxide. A reddish-violet colour
develops within 5 min.
E. Evaporate 5 mL of the chloroformic solution obtained in
identication test C to dryness on a water-bath. To the
residue add 3 mL of xanthydrol solution R and heat on a
water-bath for 3 min. A red colour develops.

EUROPEAN PHARMACOPOEIA 8.0

Ash insoluble in hydrochloric acid (2.8.1) : maximum 5.0 per


cent.
ASSAY
Shake 0.250 g of the powdered herbal drug (180) (2.9.12)
with 50.0 mL of water R for 1 h. Add 5.0 mL of a 150 g/L
solution of lead acetate R, shake, and after a few minutes add
7.5 mL of a 40 g/L solution of disodium hydrogen phosphate R.
Filter through a pleated paper lter. Heat 50.0 mL of the
ltrate with 5 mL of hydrochloric acid (150 g/L HCl) under
a reux condenser on a water-bath for 1 h. Transfer to a
separating funnel, rinse the ask with 2 quantities, each of
5 mL, of water R and shake with 3 quantities, each of 25 mL,
of chloroform R. Dry the combined chloroform layers over
anhydrous sodium sulfate R and dilute to 100.0 mL with
chloroform R. Evaporate 40.0 mL of the chloroformic solution
to dryness, dissolve the residue in 7 mL of ethanol (50 per
cent V/V) R, add 2 mL of dinitrobenzoic acid solution R and
1 mL of 1 M sodium hydroxide. At the same time prepare a
reference solution as follows. Dissolve 50.0 mg of digitoxin CRS
in ethanol (96 per cent) R and dilute to 50.0 mL with the same
solvent. Dilute 5.0 mL of the solution to 50.0 mL with ethanol
(96 per cent) R. To 5.0 mL of the resulting solution add 25 mL
of water R and 3 mL of hydrochloric acid (150 g/L HCl). Heat
the solution under a reux condenser on a water-bath for 1 h
and complete the preparation as described above. Measure the
absorbance (2.2.25) of the 2 solutions at 540 nm several times
during the rst 12 min until the maximum is reached, using as
the compensation liquid a mixture of 7 mL of ethanol (50 per
cent V/V) R, 2 mL of dinitrobenzoic acid solution R and 1 mL
of 1 M sodium hydroxide.
From the absorbances measured and the concentrations of
the solutions, calculate the content of cardenolic glycosides,
expressed as digitoxin.
STORAGE
Protected from moisture.
01/2008:1510
corrected 7.5

DOG ROSE
Rosae pseudo-fructus
DEFINITION
Rose hips made up by the receptacle and the remains of the
dried sepals of Rosa canina L., R. pendulina L. and other Rosa
species, with the achenes removed.
Content : minimum 0.3 per cent of ascorbic acid (C6H8O6 ;
Mr 176.1) (dried drug).

IDENTIFICATION
A. It consists of fragments of the eshy, hollow, urceolate
receptacle, bearing the remains of the reduced sepals, light
pink or orange-pink, the convex outer surface shiny and
strongly wrinkled ; bearing on its lighter inner surface
abundant bristle-like hairs.
B. Reduce to a powder (355) (2.9.12). The powder is
orange-yellow. Examine under a microscope using chloral
hydrate solution R. The powder shows the following
TESTS
diagnostic characters : numerous fragments of receptacle,
the outer epidermis with orange-yellow contents and a
Digitalis lanata Ehrh. The presence of leaves with few or no
thick cuticle, the inner epidermis composed of thin-walled
trichomes and with parallel venation or the presence of cells
cells containing cluster crystals and occasional prisms of
of the abaxial epidermis with beaded anticlinal walls and of
calcium oxalate ; scattered lignied cells, isodiametric,
cells of the adaxial epidermis with numerous stomata indicates
with thickened and pitted walls forming the trichome
adulteration by Digitalis lanata Ehrh.
bases ; abundant unicellar trichomes, up to 2 mm long and
Loss on drying (2.2.32): maximum 6.0 per cent, determined
30-45 m thick, tapering towards each end, walls heavily
on 1.000 g of the powdered herbal drug (355) (2.9.12) by
thickened and with a waxy cuticle which may show ssures
drying in an oven at 105 C.
in a spiral arrangement ; numerous oily orange-yellow
globules.
Total ash (2.4.16) : maximum 12.0 per cent.

1228

See the information section on general monographs (cover pages)

Drynaria rhizome

EUROPEAN PHARMACOPOEIA 8.0

C. Thin-layer chromatography (2.2.27).


Test solution. To 5 g of the powdered herbal drug (355)
(2.9.12) add 25 mL of ethanol (96 per cent) R, shake for
30 min and lter.
Reference solution. Dissolve 10 mg of ascorbic acid R in
5.0 mL of ethanol (60 per cent V/V) R.
Plate : TLC silica gel F254 plate R.
Mobile phase : acetone R, glacial acetic acid R, methanol R,
toluene R (5:5:20:70 V/V/V/V).
Application : 20 L of the test solution and 2 L of the
reference solution.
Development : over a path of 15 cm.
Drying : in air.
Detection A : examine in ultraviolet light at 254 nm.
Results A : the chromatogram obtained with the test
solution shows a quenching zone similar in position to
the principal zone in the chromatogram obtained with the
reference solution.
Detection B : spray with a 0.2 g/L solution of
dichlorophenolindophenol, sodium salt R in ethanol (96 per
cent) R. Examine in daylight.
Results B : the chromatogram obtained with the test solution
shows a white zone on a pink background (ascorbic acid)
similar in position and colour to the principal zone in the
chromatogram obtained with the reference solution. The
chromatogram also shows an intense orange-yellow zone
near the solvent front and a yellow zone in the upper third
(carotenoids).

Solution B. Treat 2.0 mL of the reference solution as described


above for solution A.
Calculate the percentage content of ascorbic acid from the
following expression :

A1

= absorbance of the test solution ;

A2
m1

= absorbance of the reference solution ;

m2

= mass of ascorbic acid used, in grams.

= mass of the substance to be examined, in grams ;

07/2012:2563

DRYNARIA RHIZOME
Drynariae rhizoma
DEFINITION
Dried rhizome of Drynaria fortunei (Kunze) J. Sm. The
ramenta may be removed.
Content : minimum 0.5 per cent of naringin (C27H32O14 ;
Mr 580.5) (dried drug).

IDENTIFICATION
A. Long, attened, slat-shaped rhizome, often curved and
TESTS
branched, 5-15 cm long and 1-1.5 cm thick. The surface
Foreign matter (2.8.2) : maximum 1 per cent.
is either completely covered in scaly, dark brown hairs
(rhizome with ramenta) or glabrous with dark brown dots
Loss on drying (2.2.32) : maximum 10.0 per cent, determined
(rhizome without ramenta). The upper surface and both
on 1.000 g of the powdered herbal drug (355) (2.9.12) by
sides show circular frond scars, rarely the frond bases. The
drying in an oven at 105 C.
lower surface shows scars or the remains of brous roots.
Total ash (2.4.16) : maximum 7.0 per cent.
The texture is light, fragile, easily broken. The section is
reddish-brown ; the steles form a ring of small yellow dots.
ASSAY
B. Microscopic examination (2.8.23). The powder is
Test solution. In a round-bottomed ask, weigh 0.500 g
reddish-brown. Examine under a microscope using chloral
of the freshly powdered herbal drug (710) (2.9.12). Add a
hydrate solution R. The powder shows the following
solution of 1.0 g of oxalic acid R in 50.0 mL of methanol R.
diagnostic characters : numerous parenchyma fragments,
Boil under a reux condenser for 10 min, and cool in
consisting of polyhedral cells with slightly and regularly
iced water until the temperature reaches 15-20 C. Filter.
thickened and pitted walls ; scalariform lignied vessels
Transfer 2.0 mL of the ltrate to a 50 mL conical ask.
of variable diameter up to 60 m ; fragments of scaly
Add successively, with gentle shaking after each addition,
hairs forming a tissue consisting of many reddish-brown
2.0 mL of dichlorophenolindophenol standard solution R
cells forming expansions on the margins (rhizome with
and then, exactly 60 s later, 0.5 mL of a 100 g/L solution
ramenta).
of thiourea R in ethanol (50 per cent V/V) R and 0.7 mL of
dinitrophenylhydrazine-sulfuric acid solution R. Heat under a C. Thin-layer chromatography (2.2.27).
reux condenser at 50 C for 75 min, and place immediately
Test solution. To 0.5 g of the powdered herbal drug (355)
in iced water for 5 min. Add dropwise 5.0 mL of a mixture of
(2.9.12) add 5 mL of methanol R and sonicate for 10 min.
12 mL of water R and 50 mL of sulfuric acid R, taking care to
Cool, centrifuge and use the supernatant.
carry out the addition over a period of minimum 90 s and
Reference solution. Dissolve 1 mg of naringin R and 1 mg
maximum 120 s while maintaining vigorous stirring in iced
of hyperoside R in 2 mL of methanol R.
water. Allow to stand for 30 min at room temperature and
Plate : TLC silica gel plate R (2-10 m).
measure the absorbance (2.2.25) at 520 nm using solution A
as compensation liquid.
Mobile phase : acetic acid R, anhydrous formic acid R,
water R, ethyl acetate R (11:11:26:100 V/V/V/V).
Solution A. Treat 2.0 mL of the ltrate obtained during the
preparation of the test solution as described but adding the
Application : 10 L as bands of 8 mm.
dinitrophenylhydrazine-sulfuric acid solution R just before the
Development : over a path of 6 cm.
absorbance is measured.
Drying : in air.
Reference solution. Dissolve 40.0 mg of ascorbic acid R in a
Detection : treat with aluminium chloride reagent R ;
freshly prepared 20 g/L solution of oxalic acid R in methanol R
examine in ultraviolet light at 365 nm.
and dilute to 100.0 mL with the same solvent. Dilute 5.0 mL
of this solution to 100.0 mL with a freshly prepared 20 g/L
Results : see below the sequence of zones present in the
solution of oxalic acid R in methanol R. Treat 2.0 mL of the
chromatograms obtained with the reference solution and
solution as described above for the ltrate obtained during
the test solution. Furthermore, other faint zones may
the preparation of the test solution. Measure the absorbance
be present in the chromatogram obtained with the test
(2.2.25) at 520 nm using solution B as the compensation liquid.
solution.
General Notices (1) apply to all monographs and other texts

1229