Belladonna Leaf— Usually partly matted together, crumpled or broken leaves, together with some smaller stems and a number of flowers and fruits. The leaves are thin and brittle, mostly light green to moderate olive-green. The lamina is mostly from 5 to 25 cm in length and from 4 to 12 cm in width and possesses an ovate-lanceolate to broadly ovate outline, an acute to acuminate apex, an entire margin, an acute to somewhat decurrent base and slightly hairy surface, the hairs being more abundant along the veins; when broken transversely, it shows numerous light-colored dots (crystal cells) visible with a lens. The petiole is slender and usually up to 4 cm in length. The flowers possess a campanulate corolla with 5 small, reflexed lobes, purplish to yellowish purple, becoming faded to brown or dusky yellow or yellow, a green, 5-lobed calyx, 5 epipetalous stamens, and a superior, bilocular ovary with numerous ovules. The fruit is subglobular, dark yellow to yellowish brown to dusky red or black, up to about 12 mm in width and sometimes subtended by the persistent calyx and containing numerous flattened, somewhat reniform seeds, the latter up to about 2 mm in width. The stems are more or less flattened and hollow and finely hairy when young.

Histology—Leaf: The epidermis of the lamina possesses wavy anticlinal walls and a distinctly striated cuticle. Stomata are more numerous in the lower epidermis and are surrounded by 3 or 4 neighboring cells, one of which is smaller than the others. The nonglandular hairs are uniseriate and up to 6-celled. Short club-shaped glandular hairs with a 1-celled stalk and multicellular head and long glandular hairs with a uniseriate stalk and unicellular head occur on both epidermises. The mesophyll consists of a single layer of palisade parenchyma beneath which occurs spongy parenchyma, the latter with scattered cells filled with microcrystals. The midrib contains an arc of bicollateral bundles, collenchyma beneath upper epidermis, and scattered parenchyma cells with microcrystals. Stem: The stem shows an epidermis with striated cuticle and few hairs, a distinct endodermis, small strands of long, thin-walled, slightly lignified pericyclic fibers, and a circle of bicollateral bundles. The parenchyma of the cortex and pith is interspersed with crystal cells. Flower: The calyx possesses numerous glandular hairs with uniseriate stalks and 1- to 3-celled glandular heads. The corolla shows a papillose inner epidermis and an outer epidermis with glandular hairs similar to those of the calyx. The pollen grains, when mounted in chloral hydrate solution, are subspherical, about 40 µm in diameter, tricolpate, having 3 germinal furrows and rows of pits between the ridges on the exine. Fruit: The epicarp exhibits polygonal epidermal cells with a striated cuticle and stomata. The mesocarp consists of large pulp cells some of which contain rosette aggregate crystals of calcium oxalate. Seed: The seed is characterized by an epidermis of large, wavy-walled cells with prominent ridges over the anticlinal walls.

Powdered Belladonna Leaf— Light olive-brown to moderate olive-green in color. The following are among the elements of identification: the separate microcrystals, the dark gray crystal cells, the cuticular striping of the epidermal cells, the vessels with ellipsoidal bordered pits, the fibers of the stem, and occasional hairs and pollen grains. Rosette aggregates of calcium oxalate and fragments of the seed occur when the drug contains belladonna fruits. Examine Belladonna Leaf for hairs having a papillose cuticle and for raphides of calcium oxalate: their presence indicates adulteration.

(Official January 1, 2007)

pH 9.5 Phosphate buffer, Internal standard solution, Standard preparation, Extraction blank, Standard curve, Chromatographic system, and System suitability— Proceed as directed in the Assay under Belladonna Extract.

Assay preparation— Moisten 10 g, previously reduced to a moderately coarse powder and accurately weighed, with a mixture of 8 mL of ammonium hydroxide, 10 mL of alcohol, and 20 mL of ether, and extract the alkaloids by either of the methods given in the following two paragraphs. If necessary, reduce the volume of the extract to 100 mL by evaporation on a steam bath.

I— Place the moistened drug in a continuous-extraction thimble, and allow maceration to proceed overnight, then extract with ether for 3 hours, or longer if necessary to effect complete extraction.

II— Place the moistened drug in a small percolator, and allow maceration to proceed overnight. Percolate slowly with a mixture of 3 volumes of ether and 1 volume of chloroform. Continue the percolation until the residue from 3 to 4 mL of percolate last passed, when dissolved in dilute sulfuric acid (1 in 70) and treated with mercuric iodide TS, shows not more than a faint turbidity.

Procedure— Record from the Standard curves the quantities, in mg, of atropine and scopolamine in the weight of the specimen taken. Proceed as directed for Procedure in the Assay under Belladonna Extract, through the next-to-the-last sentence. Add the quantity, in mg, of atropine and scopolamine, and multiply by 50 to obtain the weight, in mg, of alkaloids in the portion of Belladonna Leaf taken.