Dalmatian Insect Flowers
Preparation
Pyrethrum Extract
DEFINITION
Pyrethrum Flower is the dried flower heads of Chrysanthemum cinerariaefolium Vis. It contains not less than 1.0% of pyrethrins of which not less than half consists of pyrethrin I.
CHARACTERISTICS
Odour, faint but characteristic.
Macroscopical
Capitula occurring loose or compressed into masses; individual capitula more or less flattened, about 6 to 12 mm in diameter, and commonly with a short piece of stalk attached; receptacle almost flat, about 5 to 10 mm in diameter, without paleae, surrounded by an involucre of 2 or 3 rows of brownish yellow lanceolate bracts; ray florets number about 15 to 23, disc florets about 200 to 300; corollas of ligulate florets pale brownish and shrivelled, oblong, about 16 mm long with three rounded apical teeth, the central tooth frequently smaller than the two lateral ones; in the middle region of the corolla about 17 veins present; corollas of the disc florets tubular, yellow, with five short lobes, and enclosing five epipetalous, syngenesious stamens; each ray and disc floret has an inferior five-ribbed oblong ovary about 5 mm long with a filiform style and bifid stigma and surmounted by a membranous tubular calyx about 1 mm long; ovaries and lower part of the corollas covered with numerous scattered shining oil glands.
Microscopical
Loosely arranged large-celled sclereids of the receptacle with moderately thickened walls and few pits; fragments of the corollas of the florets, those of the ray florets showing puckered papillae on the inner epidermis and sinuous-walled cells with a striated cuticle on the outer epidermis, those of the tubular florets composed of cells with slightly thickened walls with papillae occurring only on the lobes; ovoid to spherical glandular trichomes each composed of a short, biseriate stalk and a biseriate head with two or four cells; covering trichomes twisted, T-shaped with moderately thickened walls; numerous pollen grains, spherical, 34 to 40 μm in diameter with three pores and a warty and spiny exine; groups of small rectangular sclereids, some containing prisms of calcium oxalate, from the involucral bracts, ovaries and basal region of the calyx; parenchymatous cells of the calyx and ovaries containing tabular or diamond-shaped crystals of calcium oxalate; occasional cells containing cluster crystals from the base of the corolla of the disc florets; portions of stigmas with papillose tips.
Acid-insoluble ash
Not more than 1.0%, Appendix XI K.
ASSAY
Transfer 12.5 g in No. 1000 powder to an apparatus for the continuous extraction of drugs, Appendix XI F, and extract with aromatic-free petroleum spirit (boiling range, 40° to 60°) for 7 hours. Evaporate the extract to about 40 mL and allow to stand overnight at 0° to 5°. Add 20 mL of 0.5M ethanolic potassium hydroxide and boil under a reflux condenser for 45 minutes. Transfer the solution to a beaker and wash the flask with sufficient hot water, adding the washings to the beaker, to produce a total volume of 200 mL. Boil until the volume is reduced to 150 mL, cool rapidly and transfer the solution to a stoppered flask, washing the beaker with three 20 mL quantities of water and transferring any gummy residue to the flask. Add 1 g of diatomaceous earth (Filtercel is suitable) and 10 mL of barium chloride solution, swirl gently and add sufficient water to produce 250 mL. Stopper the flask, shake vigorously until the separating liquid is clear and filter the suspension through a filter paper (Whatman No. 1 is suitable).
For pyrethrin I
Transfer 200 mL of the filtrate to a separating funnel, rinsing the measuring vessel with two 5 mL quantities of water, and add 0.05 mL of phenolphthalein solution R1. Neutralise the solution by the drop wise addition of hydrochloric acid and add 1 mL of hydrochloric acid in excess. Add 5 mL of a saturated solution of sodium chloride and 50 mL of aromatic-free petroleum spirit (boiling range, 40° to 60°), shake vigorously for 1 minute, allow to separate, remove and retain the lower layer. Filter the petroleum spirit extract through absorbent cotton into a second separating funnel containing 10 mL of water. Return the aqueous layer to the first separating funnel and repeat the extraction with 50 mL and then with 25 mL of aromatic-free petroleum spirit (boiling range, 40° to 60°), reserving the aqueous layer for the assay of pyrethrin II, and filtering the petroleum spirit extracts through the same absorbent cotton into the second separating funnel. Shake the combined petroleum spirit extracts and water for about 30 seconds and allow to separate; remove the lower layer and add it to the aqueous liquid reserved for the assay of pyrethrin II. Wash the combined petroleum spirit extracts with a further 10 mL of water, adding the washings to the reserved aqueous liquid. To the petroleum spirit extracts add 5 mL of 0.1M sodium hydroxide, shake vigorously for 1 minute, allow to separate and remove the clear lower layer, washing the stem of the separating funnel with 1 mL of water. Repeat the extraction by shaking for about 30 seconds with two quantities of 2.5 mL and 1.5 mL of 0.1M sodium hydroxide and add the extracts to the alkaline extract. Add to the flask 10 mL of mercury(II) sulfate solution, stopper, swirl and allow to stand in the dark at 25°±0.5° for exactly 60 minutes after the addition of the mercury(II) sulfate solution. Add 20 mL of acetone and 3 mL of a saturated solution of sodium chloride, heat to boiling on a water bath, allow the precipitate to settle and decant the supernatant liquid through a filter paper (Whatman No. 1 is suitable), retaining most of the precipitate in the flask. Wash the precipitate with 10 mL of acetone, again boil, allow to settle and decant through the same filter paper. Repeat the washing and decanting with three 10 mL quantities of hot chloroform. Transfer the filter paper to the flask, add 50 mL of a cooled mixture of three volumes of hydrochloric acid and two volumes of water, 1 mL of strong iodine monochloride reagent and 6 mL of chloroform. Titrate with 0.01M potassium iodate VS, running almost all the required volume of titrant into the flask in one portion. Continue the titration, shaking the flask vigorously for 30 seconds after each addition of the titrant, until the chloroform is colourless. Repeat the operation without the extract; the difference between the titrations represents the amount of potassium iodate required. Each mL of 0.01M potassium iodate VS is equivalent to 5.7 mg of pyrethrin I.
For pyrethrin II
Transfer the combined aqueous liquids reserved in the Assay for pyrethrin I to a beaker, cover with a watch glass and evaporate to 50 mL within 35 to 45 minutes. Cool, washing the underside of the watch glass with not more than 5 mL of water and adding the washings to the beaker. Filter through absorbent cotton into a separating funnel, washing with successive quantities of 10, 7.5, 7.5, 5 and 5 mL of water. Saturate the aqueous liquid with sodium chloride, add 10 mL of hydrochloric acid and 50 mL of ether, shake for 1 minute, allow to separate, and remove the lower layer. Repeat the extraction successively with 50, 25 and 25 mL of ether. Wash the combined ether extracts with three 10 mL quantities of a saturated solution of sodium chloride and transfer the ether layer to a flask with the aid of 10 mL of ether. Remove the bulk of the ether by distillation and remove the remainder with a gentle current of air and dry the residue at 100° for 10 minutes, removing any residual acid fumes with a gentle current of air. Add 2 mL of ethanol (96%) previously neutralised to phenolphthalein solution R1 and 0.05 mL of phenolphthalein solution R1, swirl to dissolve the residue, add 20 mL of carbon dioxide-free water and titrate rapidly with 0.02M sodium hydroxide VS until the colour changes to brownish pink and persists for 30 seconds, keeping the flask stoppered between additions of alkali. Repeat the operation using the aqueous liquid reserved for the repeat operation in the Assay for pyrethrin I. The difference between the titrations represents the volume of 0.02M sodium hydroxide VS required. Each mL of 0.02M sodium hydroxide VS is equivalent to 3.74 mg of pyrethrin II.
