Pimpinella anisum (fruit)

Nomenclature

Pimpinella anisum L.   Apiaceae  
Syn. Anisum vulgare Gaertn.  
Standardized common name (English): anise

Botanical Voucher Specimen

Source: MOBOT, Tropicos.org

Organoleptic Characteristics

[Pimpinella Anisum:] Odor and taste agreeable and aromatic. Agreeably aromatic odor; taste aromatic and sweet. Source: United States Dispensatory (1918)

Macroscopic Characteristics

Pimpinella Anisum is an annual plant, about a foot in height, with an erect, smooth and branching stem. The leaves are petiolate, the lower roundish-cordate, lobed, incised-serrate, the middle pinnate-lobed with cuneate or lanceolate lobes, the upper trifid, or undivided, linear. The flowers are white, and in terminal compound umbels, destitute of involucres. Anise seeds (botanically, fruit) are officially described as follows: Cremocarp broadly ovoid or pyriform, laterally compressed, from 3 to 6 mm. in length and from 2 to 3 mm. in breadth; mericarps usually cohering and attached to a slender pedicel from 2 to 12 mm. in length; summit witli a ring-like disk and 2 projecting, diverging styles; externally grayish or greenish-gray, seldom grayish-brown, slightly pubescent; each with five light brown filiform ridges and in cross-section with from 15 to 45 vittee. [...] Ovoid, somewhat compressed laterally; about five millimetres long and two millimetres broad. Mericarps usually united and attached to the pedicel. Rough from the presence of short, bristly hairs. Greenish-grey or greyish-brown; primary ridges pale, slender and entire. In transverse section, numerous vittae in each mericarp; commissural surface of the endosperm not deeply grooved. Source: United States Dispensatory (1918) Source: PlantaPhile Source: PlantaPhile

Microscopic Characteristics

Under the microscope transverse sections of Anise show an epidermal layer with numerous papillae and short, one-celled, non-glandular hairs having very thick papillose walls; primary ribs each with a small fibro-vascular bundle, surrounded by a few sclerenchymatous fibers; vittae or oil-tubes from 15 to 45 in number, extending as a more or less interrupted circle in the tissues of the mesoearp on the dorsal side of each mericarp; 2 large vittse on the commissural surface, each separated from the other tissues of the mericarp by a large cavity due to shrinkage of the seed-coat; inner epidermis of pericarp consisting of a layer of narrow, tangentially elongated cells closely united with the 1-layered seed-coat, the inner walls of which are yellowish-brown and considerably thickened; endosperm of polygonal, thick-walled cells filled with spherical or ellipsoidal aleurone grains, each containing a small rosette aggregate of calcium oxalate; the aleurone grains surrounded with an oily protoplasm, the oil of which is liberated upon mounting sections in hydrated chloral T.S., in the form of small globules; epidermal layer near the middle of the commissural surface composed of 2 or 3 rows of cells with thick porous walls, and beneath which occur small groups of thick-walled cells resembling stone cells. The powder is yellowish-brown; consisting of numerous irregular fragments of pericarp showing portions of the yellowish vittae, fragments with tracheae and sclerenchymatous fibers of carpo-phore; cells of endosperm filled with aleurone grains about 0.006 mm. in diameter, each usually enclosing a rosette aggregate crystal of calcium osalate about 0.002 mm. in diameter; non-glandular hairs 1-celled, from 0.025 to 0.2 mm, in length, either straight or curved and with numerous, slight, centrifugal projections on the outer surface. Heat 1 Gm. of the whole drug or powdered drug with 10 mils of potassium hydroxide T.S.; no mouse-like odor develops (fruits of Conium maculatum Linne). Source: United States Dispensatory (1918)

High Performance Thin Layer Chromatographic Identification

Anise (fruit) HPTLC ID - NP and PEG reagent, UV 366 nm (Image electronically enhanced) Anise (fruit) (Pimpinella anisum) Lane Assignments Lanes, from left to right (Track, Volume, Sample): 1 µL Anise fruit 1 3 µL Anise fruit 1 6 µL Anise fruit 1 3 µL Anise fruit 2 1 µL Caraway fruit 1 3 µL Caraway fruit 1 6 µL Caraway fruit 1 3 µL Caraway fruit 2 3 / 1µL Rutin, Caffeic acid (with incr. Rf) 3 µL Bitter Fennel fruit 1 3 µL Bitter Fennel fruit 2 3 µL Sweet Fennel fruit 1 3 µL Sweet Fennel fruit 2 3 µL Wild Fennel fruit 3 µL Fennel tea  Reference Sample(s) Reference: Dissolve 3 mg of rutin in 1 mL of methanol. Dissolve 1 mg of caffeic acid in 1 mL of methanol.  Stationary Phase Stationary phase, i.e. Silica gel 60, F254  Mobile Phase Ethyl acetate, formic acid, water 15:1:1 (v/v/v)  Sample Preparation Method Sample: Mix 500 mg of powdered sample with 5 mL of methanol and sonicate for 10 minutes, then centrifuge or filter the solutions and use the supernatants / filtrates as test solutions. Derivatization reagent: NP and PEG reagent; Preparation NP reagent: 1 g of natural products reagent in 200 mL ethyl acetate; Preparation PEG reagent: 10 g of polyethyleneglycole 400 in 200 mL dichloromethane; Use: Heat for 3 min to 100°C, dip (time 0, speed 5) in NP reagent while still hot, dry, dip in PEG reagent.  Detection Method Saturated chamber; developing distance 70 mm from lower edge; relative humidity 33%  Other Notes Images presented in this entry are examples and are not intended to be used as basis for setting specifications for quality control purposes. System suitability test: Rutin: orange fluorescent zone at Rf ~ 0.07; Caffeic acid: light bluish fluorescent zone at Rf ~ 0.77. Identification: Compare result with reference images. The fingerprint of the test solution is similar to that of the corresponding botanical reference sample. Additional weak zones may be present. In the upper part of the chromatogram there are three prominent zones: a light blue zone at Rf ~ 0.88, a yellow zone at Rf ~ 0.76 and a light blue zone at Rf ~ 0.61. In the lower part of the chromatogram there is a sequence of three zones (yellow, light blue, yellowish) between Rf ~ 0.15 and 0.26. Right above the application position there is a yellow zone. Test for adulteration: In the middle of the chromatogram there are neither yellow zones at Rf ~ 0.32 and Rf ~ 0.38 (red arrows, Caraway fruit) nor a faint dark blue zone at Rf ~ 0.32 (blue arrow, Bitter Fennel fruit) nor a light blue zone at Rf ~ 0.53 (yellow arrow, Sweet Fennel fruit). Source: HPTLC Association

Anise oil HPTLC ID - MAB reagent, white RT Anise oil (Pimpinella anisum) Lane Assignments Lanes, from left to right (Track, Volume, Sample): 1 µL Anise oil 1 (Ph.Eur.) 2 µL Anise oil 1 (Ph.Eur.) 3 µL Anise oil 1 (Ph.Eur.) 1 µL Anise oil 1 2 µL Anise oil 1 3 µL Anise oil 1 2 µL Anethole 2 µL Linalool 2 µL Anisaldehyde 2 µL Anise oil 2 (Ph.Eur.) 2 µL Anise oil 3 (Ph.Eur.)  Reference Sample(s) Reference: Dissolve 65 µL of anethole in 5 mL of toluene; Dissolve 15 µL of linalool in 5 mL of toluene; Optional: Dissolve 10 µL of anisaldehyde in 5 mL of toluene  Stationary Phase Stationary phase, i.e. Silica gel 60, F254  Mobile Phase Toluene, ethyl acetate 93:7 (v/v)  Sample Preparation Method Sample: Dissolve 50 µL of sample in 1mL of toluene Derivatization reagent: MAB reagent; Preparation: 0.25 g methyl 4-acetylbenzoate in a mix of 65 mL cold methanol and 5 mL sulfuric acid; Use: Spray, heat at 100 °C for 8 min  Detection Method Saturated chamber; developing distance 70 mm from lower edge; relative humidity 33%  Other Notes Images presented in this entry are examples and are not intended to be used as basis for setting specifications for quality control purposes. System suitability test: Anethole: yellow zone at Rf ~ 0.71; Linalool: violet zone at Rf ~0.29 Identification: Compare result with reference images. The fingerprint of the test solution is similar to that of the corresponding botanical reference sample. Additional weak zones may be present. The chromatogram of the test solution shows an intense purple zone corresponding to the zone of reference substance anethole, a yellow zone corresponding to reference anisaldehyde and a grey zone corresponding to reference linalool. Between the zones of anethole and anisaldehyde a grey zone is seen at Rf ~ 0.56 (blue arrow). Two other grey zones are detected below the zone of linalool at Rf ~ 0.10 and 0.18 (black arrows). Source: HPTLC Association

Aniseed HPTLC ID - UV 254nm Aniseed (Pimpinella anisum) Lane Assignments Lanes, from left to right (Track, Volume, Sample): 1 µL Aniseed 1 (Ph.Eur. method) 2 µL Aniseed 1 (Ph.Eur. method) 3 µL Aniseed 1 (Ph.Eur. method) 1 µL Aniseed 1 2 µL Aniseed 1 3 µL Aniseed 1 2 µL Anethole 2 µL Olive oil 2 µL Aniseed 2 2 µL Anise oil 1 2 µL Anise oil 2 2 µL Anise oil 3  Reference Sample(s) Reference: Dissolve 10 µL of olive oil in 1 mL of toluene. Dissolve 3 µL of anethole in 1 mL of toluene.  Stationary Phase Stationary phase, i.e. Silica gel 60, F254  Mobile Phase Toluene  Sample Preparation Method Sample: Mix 1 g of powdered sample with 10 mL of methanol and sonicate for 10 minutes, then centrifuge or filter and use the supernatants / filtrates as test solutions. Derivatization reagent: Phosphomolybdic acid reagent; Preparation: 5 g phosphomolybdic acid in 200 mL ethanol; Use: Dip (time 0, speed 5), heat at 120°C for 5 min.  Detection Method Saturated chamber; developing distance 70 mm from lower edge; relative humidity 33%  Other Notes Images presented in this entry are examples and are not intended to be used as basis for setting specifications for quality control purposes. System suitability test: Anethole: under UV 254 nm; a quenching zone at Rf ~ 0.61; Olive oil: after derivatization; a blue spot at Rf ~ 0.17 Identification: Compare result with reference images. The fingerprint of the test solution is similar to that of the corresponding botanical reference sample. Additional weak zones may be present. Under UV 254 nm the chromatogram of the test solution shows a prominent zone corresponding to the zone anethole. After derivatization a bluish zone corresponding to anethole and a zone corresponding to the zone at Rf ~ 0.17 obtained with the reference solution (olive oil) are seen. NOTE: After derivatization anise oil shows a blue zone at Rf ~ 0.81. No zone is seen at Rf ~ 0.17, but below this position two blue zones are detectable. Aniseed HPTLC ID - UV 254nm Aniseed HPTLC ID - Phosphomolybdic acid reagent, white RT Source: HPTLC Association