Medicinal Plant Phyllanthus amarus
for its Botanical, Phytochemical and
Biological Explorations

P. Koul^*, S. Singh, R. Sharma and V. K. Gupta

Indian Institute of Integrative Medicine (CSIR),

Canal Road, Jammu – 180 001, J&K State, India

ABSTRACT

Since pre-historic days attempts are being made to find out suitable drugs from natural source for the treatment of diseases because synthetic medicines cause various inevitable side effects. Recently, much importance has been imparted to develop the formulations from the plant source which are mostly free from toxic actions. The plant families are the rich source of organic compounds, many of which are well known for their therapeutic properties. Phyllanthus amarus (Euphorbiaceae) is an annual herb, which is reported to contain lignans, alkaloids, flavonoids, galloatnoids, glycosides and alkaloids. It is commonly called Bhumi-amala. The plant species is distributed throughout the hotter parts of India. It grows well under tropical conditions, however rarely survives under dry or very low temperature conditions although water logging does not show any lethal effects. The plant grows abundantly throughout India up to 700msl altitude. The plants are propagated through seeds. Since the active constituents of P. amarus are concentrated more in the leaves, production of higher leaf mass is desired for the extraction. Plant bears highest number of leaves during September and this time suitable for harvesting. The therapeutic effects has been acknowledged as antiviral, antiparasitic, antimalarial, antimicrobial, anticancer, antidiabetic, antihypercholesterolemic, cellular protective and wound-healing properties. It also acts on kidney stones and Uric Acid. It protects liver and detoxifies the toxicity and for treating hepatitis B virus.

Keywords:Phyllanthus, Hepatoprotective, Antiviral, Anticancer, Antidiabetic.

Introduction

Phyllanthus amarus belongs to family Euphorbiaceae. Commanly in Hindi it is called Jaramla, Jangli amli, Bhuinanvalah, Bhonyabali, Sada-hasurmani and Bhumiamala and in Sanskrit it is named as Bahuphala, Mahidhatrika, Bhumyamalaki, Bhupatra. The synonyms of Phyllanthus amarus are Phyllanthus niruri L. Phyllanthus fraternus and Phyllanthus nanus. The plant species is distributed throughout the hotter parts of India, in upper Gangetic plain, Bundelkhand, Assam, Bengal, Bihar, Orissa, Punjab, Deccan, Konkan and South Indian states. It is common in Central and Southern India extending to Sri Lanka. The plant grows abundantly throughout India up to 700msl altitude. Whole plant is used as the crude drug. A number of other species of Phyllanthus are found mixed in the commercial samples or used as substitutes. Among these Phyllanthus urinaria L. is used more commonly. This species has large leaves and reddish fruits (on ripening). The commercial samples originating from south India have been found to be a mixture of P. amarus. hyllanthus debilis willd and P. simplex Retz. are also reported to be used as substitute/adulterant to P. amarus. It grows well under tropical conditions, however rarely survives under dry or very low temperature conditions although water logging does not show any lethal effects.

The plants are propagated through seeds. About 1kg seeds are sufficient for seedlings for transplanting in one hectare of land. For raising the seedlings, the seeds are sown in well prepared nursery beds. Well decomposed farm yard manure should be mixed with top layer of the soil while preparing the beds. Being minute, the seeds are mixed with dry soil or sand to allow uniform distribution of seeds on the nursery bed. Later a thin layer of soil is spread to cover the nursery beds. Appropriate moisture is maintained in the beds till the seeds have germinated. In north Indian plains, the month of April-May was found very well for sowing for higher rate of germination of seeds and good herb yield. Approximately 15-30 days old seedlings, which are about 10 cm tall, are transplanted in the field at horizontal and vertical spacing of 15 cm each. Proper irrigation just after transplanting ensures establishment of seedlings. The crop raised by transplanting of seedlings gives improved yield of herbage. The medicinal plants have to be grown without chemical fertilizers and use of pesticides. Organic manures like Farm Yard Manure (FYM), Vermi-Compost, and Green Manure etc. may be used as per requirement of the species. To prevent diseases, bio-pesticides could be prepared (either single or mixture) from Neem (kernel, seeds and leaves), Chitrakmool, Dhatura, Cow’s urine etc. Plants are harvested when the rainy season is over, when they are still green and herbaceous. Since the active constituents of P. amarus are concentrated more in the leaves, production of higher leaf mass is desired for the extraction. Plant bears highest number of leaves during September and this time is suitable for harvesting.

Chemistry

The main active constituents of P. amarus are lignans (phyllanthin, hypophyllanthin, nirurin etc), flavonoids (quercetin, quercetrin, rutin etc), terpens, alkaloids etc. The leaves are the rich source of phyllanthin a diarylbutane and hypophyllanthin an aryltetrahydronaphthalene type of lignin. Enantiomer of norsecurinine, 4–methoxy securinine, 4–methoxy-norsecurinine, nirphyllin and phyllnirurin, phyllantheol, phyllathenol, nirphyllin and phyllnirurin,3,7,11,15,19,23-hexamethyl–2Z, 6Z, 10Z. 14E, 18E, 22E-tetracosahexen-1 ol, tricontanol, phthalic acid bis-ester (phyllester) with β-sitosterol, dotriacontanoic acid and a rare sterol 24-isopropylcholesterol (aerial parts); lintetralin, niranthin, nirtetralin, phyllanthin, hypophyllanthin, phyltetralin, vitamin C (leaves); phyllochrysine alkaloids (leaves, stem); lupa 20 (29)-ene-3-ol, 3,5,7-trihydroxyflavonal-4’-0-α-L(-)–rhamnopyranoside,5,3’,4’-trihydroxy flavanone-7-0-α-L(-)-rhamnopyranoside, nirurin[5,6,7,4’-tetrahydroxy 8-(3-methylbut-2-aryl)flavones-5-0-rutinoside], phyllanterpenyl ester, pentacosanyl ester, heptacosanoic acid, phyllanthusone, phyllanthosterol, phyllanthosecosteryl ester, phyllanthostigmasterol and fraternusterol (root); estradiol (bark,root); ricinoleic, linoleic and linolenic acid (seed oil); corilagin, ellagic acid, gallic acid, geraniin an angiotensin converting enzyme inhibitor, and the flavonoids-FG₁ and FG₂ (plant).

Root

(-)-Epi-gallocatechin-3-O-gallate, Lupeol, Lupeol acetate, Nor-securinine, Phyllanthine.

Root Culture

(+)-Catechin, (+)-Gallocatechin, (-)-Epi-catechin, (-)–Epi-Catechin-3-gallate, (-) Epi-gallocatechin, Gallic acid.

Leaf

4-Hydroxy-lintetralin, 2,3-dimethoxy-iso-lintetralin, Ascorbic acid 0.41 per cent, Asteragalin,β sitosterol, Hydroxy niranthin, Iso-quercitrin, Linnanthin, Lintetralin, Niranthin, Nirtetralin, Phyllanthine, Phyllochrysine, Phyltetralin, Quercetin, Quercitrin, Rutin.

Leaf Ethanolic Extract

(-)-Limonene 4.5 per cent, Cymene 11 per cent.

Plant

(-)-Nor-serurinine, 4-Hydroxy-sesamin, Corilagin, Ellagic acid, Estradiol, Geranin 0.23 per cent, Hinokinin, Iso-lintetralin, Nirtetralin, Nirurine, Nirurinetin, Phyllanthus, Phyltetralin 14 per cent, Quercetin, Repandusinic acid 0.12 per cent, Rutin, Trans-phytol.

Seed Oil

Linoleic acid 21 per cent, Linolenic acid 51.4 per cent, Ricinoleic acid 1.2 per cent.Aerial: 24-Isopropyl cholesterol, Dotriacontanoic acid, Nirphyllin, Nirurine, Phyllanthenone, phyllantheol, phyllester, Phyllnirurin, phylltetrin, Triacontan-1-al, Triancontan-1-ol.

Chemical Markers for Authentication and Identification

The secondary metabolites present in P. amarus are alkaloids, flavanoids, hydrolysable tannins and major lignans. Phyllanthin (bitter constituent) and hypophyllanthin (non-bitter compound) are isolated from the leaves. From the roots rutin, querecetin were isolated. Lintetralin was also isolated from the plant. Amariin, a novel hydrolysable tannin together with geraniin, furosin, phyllanthusin have been isolated from the polar fraction. Several chemical investigations have been conducted where the structures of most of these phytochemicals were determined by UV, IR, Mass and NMR spectroscopy. Houghton et al. (1996) isolated securinega type alkaloids by Column Chromatography (CC) and preparative Thin Layer Chromatography (TLC). This group did qualitative analysis by using TLC, and spots were detected by UV radiation (254 nm and 365 nm). The unknown compounds were determined by means of UV, IR, mass and NMR spectroscopy (Foo, 1992, 1993, 1995).

The extracts (50 per cent methanol in water, 99 per cent methanol and 50 per cent methanol in chloroform) were analyzed by thin layer chromatography based upon the presence of secondary metabolites. P. amarus material was air dried under shade for two weeks at 40°C. The dried plant material was ground with a Wiley Mill grinder to 2 mm or smaller particle size. Plant material and solvent were agitated with a laboratory rotator at 30 rpm for three days at room temperature, i.e. 18-23°C. The supernatant was filtered with Whatman filter paper No. 4 and concentrated with blown air to 1ml. The concentrated liquid extracts were stored at 4°C (May, 2002) until TLC analysis (Rugutt, 1996).

Extraction Method Development

In order to determine the extraction rate, 10 g of ground plant material of P. amarus was extracted with 100 ml (1:10, w/v) of the following solvents: A, 50 per cent methanol in D.D. water; B, 99 per cent methanol; or C, 50 per cent methanol in chloroform. All solvents used were HPLC grade from Fisher Scientific Chemicals. Plant material and solvent were agitated with an orbital shaker at 115 rpm for three days at room temperature, i.e. 18-23°C. The supernatant was drained and the residue was rinsed with 100 ml extraction solvent A, B, or C for 24 hours. The pooled extracts were filtered through Whatman No. 4 filter paper and concentrated at 60 °C using a rotary evaporator. The weight of the powders of P. amarus was recorded (Vitanyi et al., 1997).

Phytochemical Analysis of Crude Extracts by Using TLC Method

Thin layer chromatography (TLC) was employed in this study to analyze the compounds present in the crude plant extracts of the solvents 50 per cent methanol in D.D. water, 99 per cent methanol and 50 per cent methanol in chloroform. Normal phase silica gel GF precoated TLC (scored 10 20 cm) plates; 250 microns (Analtech, Uniplate No. 02521) were used. The solvent extracts (three per plate) were applied as separate spots to a TLC plate about 1.3 cm from the edge (spotting line), using 20 µl capillary tubes (microcaps disposable pipettes, Drummond Scientific Company). The mobile phase, chloroform/methanol= 9:1; 95:5; 98:2 (v/v), for each crude extract from P. amarus was chosen by trial and error. For the powder extracts of P. amarus, mobile phases with different polarities had to be used, namely, chloroform/methanol= 5:5; 7:3; 9:1 and chloroform/methanol= 9:1; 95:5, respectively. All TLC separations were performed at room temperature, i.e. 18-23°C.After sample application of 3 µl for P. amarus, the plates were placed vertically into a solvent vapor saturated TLC chamber. Three mobile phases were respectively used: chloroform/methanol= 9:1; 95:5; 98:2 (volume ratio). The spotting line was about 0.5 cm from the developing solution. After the mobile phase had moved about 80 per cent from the spotting line, the plate was removed from the developing chamber and dried in a fume hood (Houghton, 1996, Wagner and Bladt, 1996).

Phytochemical Analysis of Powder Extracts by Using TLC Method

Extraction solvent B (absolute methanol) was used to produce powder for bio-assay analysis, because this solvent had a good extraction rate for P. amarus. The TLC fingerprint of the produced P. amarus powders was compared with those of the crude solvent extracts. Twenty-five milligrams of crude powder from methanol extract (B) of P. amarus were dissolved in two solvents of different polarities: 1 ml methanoldeionized and distilled water (1); 1 ml absolute methanol (2). The mixture to be analyzed (3 µl P. amarus) was spotted near the bottom of the plate (1.3 cm). The mobile phases in which the P. amarus plates were placed are: CHCl₃

Stay updated, free articles. Join our Telegram channel

Join