Artichoke leaf
Cynarae folium
DEFINITION
Whole or cut, dried leaf of Cynara cardunculusL. (syn.C.scolymusL.).
Content: minimum▶ 0.7◀per cent of chlorogenic acid (C16H18O9; Mr354.3) (dried drug).
IDENTIFICATION
A. The entire leaf may be up to 70cm long and 30cm wide. The lamina is deeply lobed in the upper part to within 1-2cm of the petiole on either side, in the lower part the leaf becomes pinnate; all the segments have markedly dentate margins and taper at the apex. Spines are absent. The upper surface of the lamina is green with a fine covering of whitish hairs, the lower surface is pale green or white and densely tomentose with long, tangled hairs. The petiole and main veins are flat on the upper surface, prominently raised and longitudinally ridged on the lower surface, with conspicuous hairs on both surfaces.
B. Reduce to a powder (1000) (2.9.12). The powder is greenish-grey. Examine under a microscope using chloral hydrate solutionR. The powder shows the following diagnostic characters (Figure 1866.-1): fragments of the epidermises of the lamina, in surface view; the upper epidermis [F] is composed of cells with straight or slightly sinuous walls [Fa], accompanied by palisade parenchyma[Fb]; the lower epidermis [C] is composed of more sinuous-walled cells; abundant anomocytic stomata(2.8.3) on both surfaces [D] and multicellular, uniseriate covering trichomes in felted masses, the majority fragmented [Ca] with a short stalk composed of several cells and a very long, narrow and frequently curled terminal cell, others consisting of 4-6 cylindrical cells; very occasional glandular trichomes with a short stalk and a uniseriate or biseriate head (surface view [E], transverse section [Ba]); abundant fragments of covering trichomes [G]; fragments of the lamina (transverse section[B]); abundant fragments of vascular tissue from the petiole and veins [A].
C.Thin-layer chromatography (2.2.27).
Test solution. To 2.0g of the powdered herbal drug (1000) (2.9.12) add 20mL of ethanol (60per centV/V)R. Allow to stand for 2h with occasional stirring. Filter.
Reference solution. Dissolve 5mg of luteolin-7-glucosideR and 5mg of chlorogenic acidCRS in methanolR and dilute to 10mL with the same solvent.
Plate: TLC silica gel plateR (5-40µm) [or TLC silica gel plateR (2-10µm)].
Mobile phase: anhydrous formic acidR, glacial acetic acidR, waterR, ethyl acetateR (11:11:27:100V/V/V/V).
Application: 10µL [or 2µL] as bands of 10mm [or 8mm].
Development: over a path of 13cm [or 6cm].
Drying: in air.
Detection: heat at 100°C for 5min;▶ treat◀ the warm plate with a 10g/L solution of diphenylboric acid aminoethyl esterR in methanolR followed by a 50g/L solution of macrogol 400R in methanolR; examine in ultraviolet light at 365nm.
Results: see below the sequence of fluorescent zones present in the chromatograms obtained with the reference solution and the test solution. Furthermore, other fluorescent zones may be present in the chromatogram obtained with the test solution.
Top of the plateA light blue fluorescent zone
______/ ______
Luteolin-7-glucoside: a yellow or orange fluorescent zone / A yellow or orange fluorescent zone (luteolin-7-glucoside)
Chlorogenic acid: a light blue fluorescent zone / A light blue fluorescent zone (chlorogenic acid)
______/ ______
Reference solution / Test solution
Figure 1866.-1.– Illustration for identification testB of powdered herbal drug of artichoke leaf
Birch leaf
Betulae folium
DEFINITION
Whole or fragmented, dried leaves of Betula pendula Roth and/or Betula pubescens Ehrh. as well as
hybrids of both species.
Content: minimum 1.5per cent of flavonoids, expressed as hyperoside (C21H20O12; Mr464.4) (dried drug).
IDENTIFICATION
A.The leaves of both species are dark green on the adaxial surface and lighter greenish-grey on the abaxial surface; they show a characteristic dense reticulate venation. The veins are light brown or almost white.
The leaves of B. pendula are glabrous and show closely spaced glandular pits on both surfaces. The leaves of B.pendula are 3-7cm long and 2-5cm wide; the petiole is long and the doubly dentate lamina is triangular or rhomboid and broadly cuneate or truncate at the base. The angle on each side is unrounded or slightly rounded, and the apex is long and acuminate.
The leaves of B.pubescens show few glandular trichomes and are slightly pubescent on both surfaces. The abaxial surface shows small bundles of yellowish-grey trichomes at the branch points of the veins. The leaves of B.pubescens are slightly smaller, oval or rhomboid and more rounded. They are more roughly and more regularly dentate. The apex is neither long nor acuminate.
B. Microscopic examination (2.8.23). The powder is greenish-grey. Examine under a microscope using chloral hydrate solutionR. The powder shows the following diagnostic characters (Figure 1174.-1): numerous fragments of the lamina, in surface view, with straight-walled, adaxial epidermal cells accompanied by underlying palisade parenchyma[E] and cells of the abaxial epidermis surrounding anomocytic stomata (2.8.3) [G]; large, free, glandular trichomes usually measuring 100-120µm [D]; fragments of the lamina in transverse section[B], showing glandular trichomes on the epidermises [Ba], heterogeneous, asymmetrical mesophyll containing cluster crystals[Bb] and prisms[Bc] of calcium oxalate; fragments of spongy parenchyma[A] accompanied by crystal sheaths [Aa] and cells containing cluster crystals of calcium oxalate[Ab]; fragments of vessels and sclerenchyma fibres[C]. If B.pubescens is present, the powder also contains unicellular covering trichomes with very thick walls, about 80-600µm long, usually 100-200µm, numerous on the margin of the lamina[F] or on the epidermises, in surface view[H].
C.Thin-layer chromatography (2.2.27).
Test solution. To 1g of the powdered herbal drug (355) (2.9.12) add 10mL of methanolR and shake. Heat on a water-bath at 60°C for 5min. Cool and filter the solution.
Reference solution. Dissolve 1mg of chlorogenic acidR, 1mg of caffeic acidR, 2.5mg of hyperosideR and 2.5mg of rutinR in 10mL of methanolR.
Plate: TLC silica gel plateR.
Mobile phase: anhydrous formic acidR, waterR, methyl ethyl ketoneR, ethyl acetateR (10:10:30:50V/V/V/V).
Application: 10µL as bands.
Development: over a path of 10cm.
Drying: in a current of warm air.
Detection: treat with a 10g/L solution of diphenylboric acid aminoethyl esterR in methanolR; subsequently treat with a 50g/L solution of macrogol 400R in methanolR; allow to dry in air for 30min and examine in ultraviolet light at 365nm.
Results: the chromatogram obtained with the reference solution shows 3zones in its lower half: in increasing order of RF, a yellowish-brown fluorescent zone (rutin), a light blue fluorescent zone (chlorogenic acid) and a yellowish-brown fluorescent zone (hyperoside), and in its upper third, a light blue fluorescent zone (caffeic acid). The chromatogram obtained with the test solution shows 3zones similar in position and fluorescence to the zones due to rutin, chlorogenic acid and hyperoside in the chromatogram obtained with the reference solution. The zone due to rutin is very faint and the zone due to hyperoside is intense. It also shows other yellowish-brown faint fluorescent zones between the zones due to caffeic acid and chlorogenic acid in the chromatogram obtained with the reference solution. Near the solvent front, the red fluorescent zone due to chlorophylls is visible. In the chromatogram obtained with the test solution, between this zone and the zone due to caffeic acid in the chromatogram obtained with the reference solution, there is a brownish-yellow zone due to quercetin.
Figure 1174.-1. – Illustration for identification testB of powdered herbal drug of birch leaf
Eucalyptus leafEucalyptus leaf
Eucalypti folium
DEFINITION
Whole or cut, dried leaves of older branches of Eucalyptus globulus Labill.
Essential oil content:
–for the whole drug, minimum 20mL/kg (anhydrous drug);
–for the cut drug, minimum 15mL/kg (anhydrous drug).
CHARACTERS
Aromatic odour of cineole.
IDENTIFICATION
A. The leaves, which are mainly greyish-green and relatively thick, are elongated, elliptical and slightly sickle-shaped and usually up to 25cm in length and up to 5cm in width. The petiole is twisted, strongly wrinkled and is 2-3cm, rarely 5cm, in length. The coriaceous, stiff leaves are entire and glabrous and have a yellowish-green midrib. Lateral veins anastomose near the margin to a continuous line. The margin is even and somewhat thickened. On both surfaces there are minute, irregularly distributed, warty, dark brown spots. Small oil glands may be seen in transmitted light.
▶B.Microscopic examination (2.8.23). The powder is greyish-green. Examine under a microscope using chloral hydrate solutionR. The powder shows the following diagnostic characters (Figure 1320.-1): fragments of glabrous lamina (surface view [A, L], transverse section [F, H]), with small, thick-walled epidermal cells bearing a thick cuticle[Fa,Ha], numerous anomocytic stomata (2.8.3) greater than 80µm in diameter [Aa,La] with occasional groups of brown cork cells, 300µm in diameter and brownish-black in their centre, and underlying palisade parenchyma [Ab, Fb]; fragments of bilateral mesophyll (side view [G]), with 2-3 layers of palisade parenchyma [Ga] on each side and in the centre several layers of spongy mesophyll[Gb] with elongated cells having the same orientation as the palisade cells and containing prisms[B,Gd] and cluster crystals of calcium oxalate[Gc,K]; large schizogenous oil glands, whole [E] or usually broken, accompanied by palisade parenchyma[Ea]; fragments of vessels [J] and thick-walled and slightly channelled fibres[C] accompanied by crystal sheaths[Ca, Ja]; crystal sheaths containing prisms of calcium oxalate[D].
C.Thin-layer chromatography (2.2.27).
Test solution. Shake 0.5g of the freshly powdered herbal drug (355) (2.9.12) with 5mL of tolueneR for 2-3min and filter over about 2g of anhydrous sodium sulfateR.
Reference solution. Dissolve 50µL of cineoleR in tolueneR and dilute to 5mL with the same solvent.
Plate: TLC silica gel plateR.
Mobile phase: ethyl acetateR, tolueneR (10:90V/V).
Application: 10µL as bands.
Development: over a path of 15cm.
Drying: in air.
Detection:▶ treat with anisaldehyde solutionR and heat at 100-105°C for 10-15min; examine in daylight.◀
Results: the chromatogram obtained with the reference solution shows in the middle a zone due to cineole. The chromatogram obtained with the test solution shows a principal zone similar in position and colour to the zone due to cineole in the chromatogram obtained with the reference solution, it also shows an intense violet zone (hydrocarbons) near the solvent front and there may also be other fainter zones.
Figure 1320.-1. – Illustration for identification testB of powdered herbal drug of eucalyptus leaf
Ginkgo leaf
Ginkgonis folium
DEFINITION
Whole or fragmented, dried leaf of Ginkgo bilobaL.
Content: not less than 0.5per cent of flavonoids, expressed as flavone glycosides (Mr757) (dried drug).
IDENTIFICATION
A. The leaf is greyish or yellowish-green or yellowish-brown. The upper surface is slightly darker than the lower surface. The petioles are about 4-9cm long. The lamina is about 4-10cm wide, fan-shaped, usually bilobate or sometimes undivided. Both surfaces are smooth, and the venation dichotomous, the veins appearing to radiate from the base; they are equally prominent on both surfaces. The distal margin is incised, irregularly and to different degrees, and irregularly lobate or emarginate. The lateral margins are entire and taper towards the base.
B. Reduce to a powder (355) (2.9.12). The powder is greyish or yellowish-green or yellowish-brown. Examine under a microscope using chloral hydrate solutionR. The powder shows the following diagnostic characters (Figure1828.-1): irregularly-shaped fragments of the lamina[A, B, D,E], with the upper epidermis, in surface view [D] and transverse section [E], consisting of elongated cells with irregularly sinuous walls [Da], often accompanied by palisade parenchyma [Db], and the lower epidermis, in surface view [A] and transverse section [B], consisting of small cells, with a finely striated cuticle and each cell shortly papillose [Aa], and stomata [Ab] about 60µm, wide, deeply sunken with 6-8 subsidiary cells; fragments of vascular tissue from the petiole and veins [C] with xylem [Ca] and parenchyma, some cells containing abundant cluster crystals of calcium oxalate of various sizes [Cb].
C. C.Thin-layer chromatography (2.2.27).
Test solution. To 2.0g of the powdered herbal drug (710) (2.9.12) add 10mL of methanolR. Heat in a water-bath at 65°C for 10min. Shake frequently. Allow to cool to room temperature and filter.
Reference solution. Dissolve 1.0mg of chlorogenic acidR and 3.0mg of rutinR in 20mL of methanolR.
Plate: TLC silica gel plateR.
Mobile phase: anhydrous formic acidR, glacial acetic acidR, waterR, ethyl acetateR (7.5:7.5:17.5:67.5V/V/V/V).
Application: 20µL as bands.
Development: over a path of 17cm.
Drying: at 100-105°C.
Detection: spray the warm plate with a 10g/L solution of diphenylboric acid aminoethyl esterR in methanolR, then with the same volume of a 50g/L solution of macrogol 400R in methanolR; allow to dry in air for about 30min and examine in ultraviolet light at 365nm.
Results: see below the sequence of zones present in the chromatograms obtained with the reference solution and the test solution. Furthermore, other weak fluorescent zones may be present in the chromatogram obtained with the test solution.
Top of the plateA yellowish-brown fluorescent zone
A green fluorescent zone
2 yellowish-brown fluorescent zones
An intense light blue fluorescent zone sometimes overlapped by a greenish-brown fluorescent zone
Chlorogenic acid: a light blue fluorescent zone
A green fluorescent zone
Rutin: a yellowish-brown fluorescent zone / 2 yellowish-brown fluorescent zones
A green fluorescent zone
A yellowish-brown fluorescent zone
Reference solution / Test solution
Figure 1828.-1. – Illustration for identification testB of powdered herbal drug of ginkgo leaf
Hamamelis leafHamamelis leaf
Hamamelidis folium
DEFINITION
Whole or cut, dried leaf of Hamamelis virginianaL.
Content: minimum 3per cent of tannins, expressed as pyrogallol (C6H6O3; Mr126.1) (dried drug).
IDENTIFICATION
A. The leaf is green or greenish-brown, often broken, crumpled and compressed into more or less compact masses. The lamina is broadly ovate or obovate; the base is oblique and asymmetric and the apex is acute or, rarely, obtuse. The margins of the lamina are roughly crenate or dentate. The venation is pinnate and prominent on the abaxial surface. Usually, 4-6 pairs of secondary veins are attached to the main vein, emerging at an acute angle and curving gently to the marginal points where there are fine veins often at right angles to the secondary veins.
B.Reduce to a powder (355) (2.9.12). The powder is brownish-green. Examine under a microscope using chloral hydrate solutionR. The powder shows the following diagnostic characters (Figure 0909.-1): fragments of adaxial epidermis with wavy anticlinal walls, in surface view [C, J], often accompanied by small, cylindrical cells of the palisade parenchyma, in surface view [Ja], or elongated, in transverse section[F]; fragments of abaxial epidermis with stomata mainly paracytic (2.8.3), in surface view [B], which may be accompanied by irregular-shaped cells of spongy mesophyll [K, L]; star-shaped covering trichomes, either entire or broken [A, D, M], composed of 4-12 unicellular branches that are united by their bases, elongated, conical and curved, usually up to 250µm long, thick-walled and with a clearly visible lumen whose contents are often brown; fibres are lignified and thick-walled, isolated or in groups, and accompanied by a sheath of prismatic calcium oxalate crystals [N, P]; sclereids, frequently enlarged at 1 or both ends, 150-180µm long, whole or fragmented [H]; fragments of annular or spiral vessels [E]; isolated prisms of calcium oxalate [G].