Other plant parts used, but not included in this monograph: fl owers, seeds, stem bark, oil (1–3, 8, 10, 12).
General appearance
Compound leaves up to 40 cm long composed of 8–18 short-petiolate narrow-ovate, pointed, curved toothed leafl ets, 3–10 cm long and 1–4 cm wide arranged in alternate pairs. Glabrous dark green upper surface, paler underside (1–3).
Organoleptic properties
Odour: characteristic, alliaceous; taste: bitter (1–3).
Microscopic characteristics
Lower epidermis with anomocytic stomata and occasional unicellular tri- chomes. Two layers of palisade cells are found below the upper epidermis. Spongy parenchyma exhibits intercellular spaces and secretory cells, which are abundant on the borderline with the palisade cells. Anticlinal cell walls are almost straight. Mesophyll contains rosette crystals. Col- lenchyma interrupts mesophyll on both upper and lower surfaces in the midrib region. Vascular bundles strongly curved, lignifi ed, collateral (1–3).
Powdered plant material
Green and characterized by the presence of cortical cells of the rachis, fragments of palisade cells, hairs, fi bres, wood fi bres, spiral lignifi ed vas- cular elements, epidermal tissues of the leaf with characteristic anomo- cytic stomata and large pit cells with intercellular spaces. Epidermal cell walls straight (2, 3).
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General identity tests
Macroscopic and microscopic examinations (1–3), microchemical tests (2) and thin-layer chromatography (2).
Purity tests
Microbiological
Tests for specifi c microorganisms and microbial contamination limits are as described in the WHO guidelines on quality control methods for me- dicinal plants (13).
Foreign organic matter
Not more than 2% (4).
Total ash
Not more than 10% (4).
Acid-insoluble ash
Not more than 1% (4).
Water-soluble extractive
Not less than 19% (4).
Alcohol-soluble extractive
Not less than 13% (4).
Loss on drying
Not more than 3% (2).
Pesticide residues
The recommended maximum limit of aldrin and dieldrin is not more than 0.05 mg/kg (14). For other pesticides, see the European pharmacopoeia (14) and the WHO guidelines on quality control methods for medicinal plants (13) and pesticide residues (15).
Heavy metals
For maximum limits and analysis of heavy metals, consult the WHO guidelines on quality control methods for medicinal plants (13).
Radioactive residues
Where applicable, consult the WHO guidelines on quality control methods for medicinal plants (13) for the analysis of radioactive iso- topes.
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Other purity tests
Chemical and sulfated ash tests to be established in accordance with national requirements.
Chemical assays
High-performance liquid chromatography methods are available for the quantitative determination of oxidized tetranortriterpenes (16, 17).
Major chemical constituents
The major characteristic constituents are oxidized tetranortriterpenes including azadirachtin (azadirachtin A), 3-tigloylazadirachtol (azadi- rachtin B), 1-tigloyl-3-acetyl-11-hydroxy-meliacarpin (azadirachtin D), 11-demethoxycarbonyl azadirachtin (azadirachtin H), 1-tigloyl-3- acetyl-11-hydroxy-11-demethoxycarbonyl meliacarpin (azadirachtin I), azadiriadione, azadirachtanin, epoxyazadiradione, nimbin, deacetyl- nimbin, salannin, azadirachtolide, isoazadirolide, margosinolide, nim- bandiol, nimbinene, nimbolin A, nimbocinone, nimbocinolide, nimbo- lide, nimocin, nimocinol and related derivatives (9, 11, 18–20). The structures of azadirachtin, nimbin and deacetylnimbin are presented below.
Medicinal uses
Uses supported by clinical data
External applications for treatment of ringworm (21). However, data from controlled clinical trials are lacking.
Uses described in pharmacopoeias and well established documents
Treatment of worm and lice infections, jaundice, external ulcers, cardio- vascular disease, diabetes, gingivitis, malaria, rheumatism and skin disorders. External applications for treatment of septic wounds and boils (6, 8). O H H O OH H H O CH3 H3C O O O H3C CH3 H O O O OH CH3 CH3 H O OH O O CH3 O H H H azadirachtin O O H3C H CH3 H O CH3 O O H3C O H3C O CH3 H H O R H H nimbin deacetylnimbin R = CO-CH3 R = H SMPvol3 layout.indd 91 SMPvol3 layout.indd 91 10.8.2007 12:10:0610.8.2007 12:10:06
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Uses described in traditional medicine
Treatment of allergic skin reactions, asthma, bruises, colic, conjunctivitis, dysentery, dysmenorrhoea, delirium in fever, gout, headache, itching due to varicella, jaundice, kidney stones, leprosy, leukorrhoea, psoriasis, sca- bies, smallpox, sprains and muscular pain, syphilis, yellow fever, warts and wounds (10, 22). Also used as an antivenin, contraceptive, emmena- gogue, tonic, stomatic and vermicide (9).
Pharmacology
Experimental pharmacology
Anxiolytic and analgesic activities
Intragastric administration of 10.0–200.0 mg/kg body weight (bw) of an aqueous extract of Folium Azadirachti produced anxiolytic effects similar to those of 1.0 mg/kg bw of diazepam in rats in the elevated-plus-maze and open-fi eld behaviour tests (23).
The analgesic effect of an extract of the leaves was assessed in mice using the acetic acid writhing test and the tail fl ick test. Intragastric administration of 10.0–100.0 mg/kg bw of the extract reduced the inci- dence of writhing and enhanced tail-withdrawal latencies (24).
Antiandrogenic activity
Intragastric administration of 20.0 mg, 40.0 mg or 60.0 mg of powdered leaves per day to rats for 24 days resulted in a decrease in the weight of the seminal vesicles and ventral prostate, and a reduction in epithelial height, nuclear diameter and secretory material in the lumen of these organs. De- creases in total protein and acid phosphatase activities were also observed. These regressive histological and biochemical changes suggest that the leaves have an antiandrogenic property (25). Histological and biochemi- cal changes were also observed in the caput and cauda epididymis of rats treated orally with similar doses of the powdered leaves given daily for 24 days. The height of the epithelium and the diameter of the nucleus in both regions were reduced. Serum testosterone concentrations were also reduced in animals receiving the highest dose (26). Intragastric adminis- tration of an aqueous extract of the leaves (dose not specifi ed) to male mice daily for 10 weeks resulted in a signifi cant (P < 0.01) reduction in total serum testosterone and bilirubin (27).
Antihepatotoxic activity
The effect of an aqueous extract of the leaves was evaluated in paracetamol- induced hepatotoxicity in rats. Intragastric administration of 500.0 mg/kg bw of the extract signifi cantly (P < 0.01) reduced elevated levels of serum
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93 aspartate aminotransferase, alanine aminotransferase and γ-glutamyl transpeptidase (28).
Anti-infl ammatory activity
Intragastric administration of 200.0 mg/kg bw of an aqueous extract of the leaves to rats decreased infl ammation and swelling in the cotton pellet granuloma assay (29). Intraperitoneal injection of 200.0–400.0 mg/kg bw of an aqueous extract of the leaves to rats reduced carrageenan-induced footpad oedema (30).
Antihyperglycaemic activity
A hypoglycaemic effect was observed in normal and alloxan-induced dia- betic rabbits after administration of 50.0 mg/kg bw of an ethanol extract of the leaves. The effect was more pronounced in diabetic animals, and reduced blood glucose levels. The hypoglycaemic effect was comparable to that of glibenclamide. Pretreatment with the extract 2 weeks prior to alloxan treatment partially prevented the rise in blood glucose levels as compared with control diabetic animals (31). Intragastric administration of 50.0–400.0 mg/kg bw of a 70% ethanol extract of the leaves signifi - cantly (P < 0.001) reduced elevated blood glucose levels in normal and streptozocin-induced diabetic rats (32–34). A 70% ethanol extract of the leaves signifi cantly (P < 0.05) blocked the inhibitory effect of serotonin on insulin secretion mediated by glucose in isolated rat pancreas (35).
Antimalarial activity
An aqueous or ethanol extract of the leaves inhibited the growth of Plas-
modium falciparum in vitro, with median inhibitory concentrations of
115.0 μg/ml and 5.0 μg/ml, respectively. Nimbolide, a constituent of the extract, inhibited the growth of P. falciparum in vitro with a median effec- tive concentration of 2.0 μg/ml (36). However, intragastric administration of 746.0 mg/kg bw of the aqueous extract, 62.5 mg/kg bw of the ethanol extract or 12.5 mg/kg bw of nimbolide had no such effect in Plasmodium- infected mice (36). P. berghei-infected mice showed parasite suppression after intragastric administration of 125.0–500.0 mg/kg bw of a dried methanol extract of the leaves per day for 4 days, but all the animals died after 5 days (37). A 95% ethanol extract of the leaves at concentrations of up to 500.0 mg/ml did not inhibit the growth of P. falciparum in vitro (38).
Antimicrobial and antiviral activity
A methanol extract of the leaves, 1.0 mg/ml, inhibited plaque formation in six antigenic types of coxsackievirus B at 96 hours in vitro. The mini- mal inhibitory concentrations were not toxic to Vero African green mon-
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key kidney cells. The subtoxic concentration was 8.0 mg/ml and the cyto- toxic concentration was 10.0 mg/ml (39).
An aqueous extract of the leaves, at various concentrations depending on the organism, inhibited the growth of Bacteroides gingivalis, B. inter-
medius, Streptococcus salivarius and S. viridans in vitro (40). A petroleum
ether extract of the leaves, at various concentrations depending on the organism, inhibited the growth of Epidermophyton fl occosum, Microspo-
rum canis, M. gypseum, Trichophyton concentricum, T. violaceum and T. rubrum (41).
Antioxidant activity
The effect of the leaves on hepatic lipid peroxidation and antioxidant sta- tus during gastric carcinogenesis induced by N-methyl-N'-nitro-N-ni- trosoguanidine was assessed in rats. Intragastric administration of 100.0 mg/kg bw of an aqueous extract of the leaves decreased lipid per- oxidation in the liver of tumour-bearing animals, which was accompanied by a decrease in the activities of glutathione peroxidase, glutathione-S- transferase and γ-glutamyl transpeptidase, and a reduction in glutathione level. Administration of 100.0 mg/kg bw of an extract of the leaves sup- pressed lipid peroxidation and increased hepatic levels of glutathione and glutathione-dependent enzymes (42). Intragastric administration of 100.0 mg/kg bw of an aqueous extract of the leaves three times per week to hamsters with buccal pouch carcinogenesis induced by 7,12-dimethyl- benz[α]anthracene reduced lipid peroxidation and increased the glutathi- one concentration in the oral mucosa of tumour-bearing animals (43).
Antiulcer activity
The antiulcer effects of an aqueous extract of the leaves were investigated in rats exposed to 2-hour cold-restraint stress or given ethanol for 1 hour. The extract, administered orally in doses of 10.0 mg/kg bw, 40.0 mg/kg bw or 160.0 mg/kg bw as single- or fi ve-dose pretreatments produced a dose-dependent reduction in the severity of gastric ulcers induced by stress and a decrease in gastric mucosal damage provoked by ethanol. The extract prevented mast cell degranulation and increased the amount of adherent gastric mucus in stressed animals (44). Intragastric administra- tion of 40.0 mg/kg bw of an aqueous extract of the leaves per day for 5 days to rats inhibited stress-induced depletion of gastric wall adherent cells and mucus production (44).
Cardiovascular effects
Intragastric administration of 200.0 mg/kg bw of an alcohol extract of the leaves to anaesthetized rabbits decreased the heart rate from 280 to
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95 150 beats per minute, and had a weak antiarrhythmic effect against oua- bain-induced dysrhythmia (45). Intravenous administration of 100.0 mg/ kg bw, 300.0 mg/kg bw or 1000.0 mg/kg bw of an ethanol extract of the leaves to rats resulted in initial bradycardia followed by cardiac arrhyth- mias. The treatment produced a dose-related fall in blood pressure that was immediate, sharp and persistent. Pretreatment with atropine or me- pyramine failed to prevent the hypotensive effect of the extract (46).
Immune effects
The effect of an aqueous extract of the leaves on humoral and cell-medi- ated immune responses was assessed in mice treated with ovalbumin. At doses of 10.0 mg/kg bw, 30.0 mg/kg bw or 100.0 mg/kg bw, the extract produced no appreciable effects on organ/body weight indices for liver, spleen and thymus compared with controls. In tests for humoral immune responses, IgM and IgG levels, and antiovalbumin antibody titres were higher in mice receiving the highest dose of extract than in animals in the control group. In tests for cell-mediated immune responses, mice receiv- ing the highest dose of extract showed enhancement of macrophage mi- gration inhibition and footpad thickness (47). Intragastric administration of 100.0 mg/kg bw of an aqueous extract of the leaves to normal and stressed rats lowered blood glucose and triglyceride levels, attenuated stress-induced elevations of cholesterol and urea, and suppressed humor- al responses (48).
The effect of powdered leaves on humoral and cell-mediated immune responses was assessed in chickens infected with infectious bursal disease. A dose of 2.0 g/kg bw per day given in the diet increased antibody titres against Newcastle disease virus antigen and enhanced infl ammatory reac- tions to chloro-2,4-dinitrobenzene in the skin contact test (49).
Toxicology
Chickens fed diets containing the powdered leaves, 2% or 5%, from the 7th to the 35th day of age, and then a control diet for 2 weeks, showed a reduction in body weight gain and effi ciency of feed use compared with controls. The main pathological changes observed included an increase in lactic dehydrogenase, glutamic-oxaloacetic transaminase and alkaline phosphatase activities, an increase in uric acid and bilirubin concentra- tions, and a decrease in total serum protein levels. There were marked reductions in the values of erythrocyte count, haemoglobin concentra- tion, packed cell volume, mean corpuscular volume and mean corpuscular haemoglobin, which were associated with yellow discoloration on the legs and hepatonephropathy (50).
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Intragastric administration of 50.0 mg/kg bw or 200.0 mg/kg bw of aqueous suspensions of the leaves per day to goats and guinea-pigs over a period of up to 8 weeks produced a progressive decrease in body weight, weakness, inappetence, loss of condition and decreases in the pulse and respiratory rates. In goats, the higher dose produced tremors and ataxia during the last few days of treatment. No statistically signifi cant haema- tological changes were observed, although there was a tendency towards lowered erythrocyte counts, packed cell volume and haemoglobin levels. The treatment increased aspartate transferase and sorbitol dehydrogenase activities, and concentrations of cholesterol, urea, creatinine and potassi- um in the plasma. No signifi cant changes in the plasma concentrations of sodium, chloride or bilirubin were detected. Autopsy of treated goats re- vealed areas of haemorrhagic erosion. The hearts appeared fl appy and in some animals there was hydropericardium. Histopathologically, there was evidence of various degrees of haemorrhage, congestion, and degen- eration in the liver, kidney, lung, duodenum, brain and seminiferous tu- bules (51).
The effect of intragastric administration of 40.0 mg/kg bw and 100.0 mg/kg bw of an aqueous extract of the leaves per day for 20 days on thyroid function was assessed in male mice. The higher dose decreased serum tri-iodothyronine and increased serum thyroxine concentrations. There was a concomitant increase in hepatic lipid peroxidation and a de- crease in glucose-6-phosphatase activity. The lower dose produced no signifi cant changes (52).
The median lethal dose of a 50% ethanol extract of the leaves in mice was 681.0 mg/kg bw when administered by intraperitoneal injection (53).
Clinical pharmacology
A 70% ethanol extract of the leaves was used for the treatment of ring- worm in seven patients. External applications of a 40% solution of the extract twice per day to the affected areas for 5–10 days were reported to be effective (no further details available) (21).
Adverse reactions
A case of ventricular fi brillation and cardiac arrest due to neem leaf poisoning has been reported (54–56). Contact dermatitis has also been reported (57).
Contraindications
Owing to potential genotoxic effects (58), the leaves should not be administered during pregnancy or nursing, or to children under the age of 12 years.
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Warnings
No information available.
Precautions
Drug interactions
Administration of Folium Azadirachti may reduce blood glucose levels and should therefore be used with caution in insulin-dependent diabetic patients or patients taking oral antihyperglycaemic drugs.
Carcinogenesis, mutagenesis, impairment of fertility
A petroleum ether extract of the leaves was not mutagenic in the
Salmonella/microsome assay at concentrations of 0.1 ml/plate using S. typhimurium strains TA98, TA100, TA1535 and TA1537 (59).
Intragastric administration of 5.0 mg/10 g bw, 10.0 mg/10 g bw or 20.0 mg/10 g bw of an ethanol extract of the leaves per day for 7 days to mice signifi cantly (P < 0.05) increased the incidence of structural and mi- totic disruptive changes in metaphase chromosomes of bone marrow cells on days 8, 15 and 35 (58). Intragastric administration of 100.0 mg/kg bw of an ethanol extract of the leaves per day for 21 days had no effect on spermatogenesis in male rats, and no effect on implantation in female ani- mals mated with treated males (60).
Pregnancy: teratogenic effects
Intragastric administration of 200.0 mg/kg bw of an acetone or 50% etha- nol extract of the leaves to pregnant rats on days 1–7 of pregnancy did not produce any teratogenic or embryotoxic effects (61).
Nursing mothers
See Contraindications.
Paediatric use
See Contraindications.
Other precautions
No information available on general precautions or on precautions con- cerning drug and laboratory test reactions; or non-teratogenic effects in pregnancy.
Dosage forms
Dried leaves for infusions and decoctions, and extracts and tinctures (8). Store leaves in a cool, dry place (3).
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Posology
(Unless otherwise indicated)
Infusion (1:20): 15–30 ml. Tincture (1:5): 4–8 ml (8). External applications: 70% ethanol extract of the leaves diluted to 40%, apply twice daily (21).
References
1. African pharmacopoeia. Vol. 1. Lagos, Organization of African Unity, Scien- tifi c, Technical and Research Commission, 1985.
2. Central Council for Research in Unani Medicine. Standardization of single
drugs of Unani medicine – part II. New Delhi, Ministry of Health and
Family Welfare, 1992.
3. Ghana herbal pharmacopoeia. Accra, Ghana, The Advent Press, 1992. 4. The Ayurvedic pharmacopoeia of India. Part I. Vol. II. New Delhi, Ministry
of Health and Family Welfare, Department of Indian System of Medicine and Homeopathy, 1999.
5. Zahedi E. Botanical dictionary. Scientifi c names of plants in English, French, German, Arabic and Persian languages. Tehran, Tehran University Publica-
tions, 1959.
6. Indian medicinal plants. Vol. I. New Delhi, Orient Longman, 1971.
7. Issa A. Dictionnaire des noms des plantes en latin, français, anglais et arabe.
[Dictionary of plant names in Latin, French, English and Arabic.] Beirut, Dar al-Raed al-Arabi, 1991.
8. Iwu MM. Handbook of African medicinal plants. Boca Raton, FL, CRC
Press, 1993.
9. Farnsworth NR, ed. NAPRALERT database. Chicago, IL, University of Illinois at Chicago, 9 February 2001 production (an online database available directly through the University of Illinois at Chicago or through the Scien- tifi c and Technical Network (STN) of Chemical Abstracts Services).
10. Vijayalakshmi K, Radha KS, Shiva V. Neem: a user’s manual. Madras, Centre for Indian Knowledge Systems; New Delhi, Research Foundation for Science, Technology and Natural Resource Policy, 1995.
11. Medicinal plants in the South Pacifi c. Manila, World Health Organization Regional Offi ce for the Western Pacifi c, 1998 (WHO Regional Publications, Western Pacifi c Series, No. 19).
12. Cambie RC, Ash J. Fijian medicinal plants. University of Auckland, CSIRO Publishing, 1994.
13. Quality control methods for medicinal plant materials. Geneva, World Health Organization, 1998.
14. European pharmacopoeia, 3rd ed. Strasbourg, Council of Europe, 1996. 15. Guidelines for predicting dietary intake of pesticide residues, 2nd rev. ed.
Geneva, World Health Organization, 1997 (WHO/FSF/FOS/97.7; available from Food Safety, World Health Organization, 1211 Geneva 27, Switzerland). 16. Govindachari TR, Suresh G, Gopalakrishnan G. A direct preparative high
performance liquid chromatography procedure for the isolation of major tri-
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terpenoids and their quantitative determination in neem oil. Journal of
Liquid Chromatography, 1995, 18:3465–3471.
17. Schaaf O et al. Rapid and sensitive analysis of azadirachtin and related triterpenoids from neem (Azadiracta indica) by high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectro- metry. Journal of Chromatography A, 2000, 886: 89–97.
18. Bruneton J. Pharmacognosy, phytochemistry, medicinal plants. Paris, Lavoisier Publishing, 1995.
19. Kraus W. Biologically active ingredients: Azadirachtin and other triterpe- noids. In: Schmutterre H, ed. The neem tree Azadirachta indica A. Juss. and
other Meliaceous plants. Weinheim, VCH, 1995.
20. Akhila A, Rani K. Chemistry of the neem tree (Azadirachta indica A. Juss.). In: Herz W, et al. eds. Fortschritte der Chemie Organischer Naturstoffe, 1999, 78:47–149.
21. Singh N et al. Melia azadirachta in some common skin disorders. Antiseptic, 1979, 76:677–680.
22. Perry LM, Metzger J. Medicinal plants of East and Southeast Asia: attributed
properties and uses. Cambridge, MA, MIT Press, 1980.
23. Jaiswal AK, Bhattacharya SK, Acharya SB. Anxiolytic activity of Azadirach-