Asian Pacific Journal of Tropical Biomedicine (2012)S40-S45 S40

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Document heading Inter-specific variation studies on the phyto-constituents of Christella and Adiantum using phytochemical methods Muraleedharannair Jalajakumari Mithraja1, Johnson Marimuthu @ Antonisamy2, Mony Mahesh1, Zachariah Miller Paul1, Solomon Jeeva1 1 Centre for Biodiversity and Biotechnology, Department of Botany,Nesamony Memorial Christian College, Marthandam - 629 165, Tamil Nadu, India 2 Department of Biology and Plant Biotechnology,St. Xavier’s College (Autonomous), Palayamkottai, Tamil Nadu, India

ARTICLE INFO ABSTRACT

Article history: Objective: Adiantum caudataum (A. caudataum), To examine the phyto-constituents of Received 21 December 2011 Adiantum latifolium (A. latifolium), Adiantum lunulatum (A. lunulatum), 29 2011 (C. dentate) Christella parasitica (C. parasitica) Received in revised form December and , to provide chemical marker and find A 15 F 2012 Methods: ccepted ebruary inter- specific variation between the medicinally important genuses. The dried and A 28 A 2012 vailable online pril powered leaves materials (50 g) were extracted successively with 250 mL of petroleum ether, ethyl acetate, methanol, chloroform, acetone, benzene and water by using Soxhlet extractor for Keywords: 8 h at a temperature not exceeding the boiling point of the solvent. The aqueous extracts were Inter- specific ( 1) 40 variation filtered using Whattman filter paper No. and then concentrated in vacuum at 曟 using Adiantum Rotary evaporator. The residues obtained were stored in a freezer -70 曟 until further tests. Christella Results:Phytoche mical screening of the extracts was carried out according to the standard methods. 30 Phytochemistry A total of fiveA. p caudataum,lants and A.ex tlatifolium,racts were A.ex alunulatum,mined for tC.he dentataphytoch emicC.al sparasiticacreening. Chemical marker The crude extracts of and showed varied degree of phyto-constituents with reference to solvents of the plant extractA.s. 18 caudataum,A total of t hA.re elatifolium, a nA.d lunulatum extracts were examined for the phytochemical screening . The steroid is totA.al lcaudatumy absent in the tested extracts of A. lunulatum. SimilaA.r t ocaudatum. that caob oAx ytotallic a cid also absent in and Coumarin showed its presence only in of two plants and twelve extracts were examined for the 11 phytochemical properties of ChriC.ste lparasiticala. Of which phenol is present in extracC.ts tparasiticaested. Ste roid is presentConclusions: only in one ex tract of and coumarin is present only in five extracts. The present study result provids a simple and cheap tool (Chemical marker) for the characterization and identification of the medicinally important taxa. With the help the phytochemical screenings we can find inter-Adiantum specific v ariatChristellaion between the medicinally important genuses and easily we can differentiate the and genuses.

1. Introduction use of plant extracts and phytochemicals, both with known antimicrobial properties, can be of great significance For a long period of time, plants have been a valuable in therapeutic treatments. In the last few years, a number source of natural products for maintaining human of studies have been conducted in different countries health, especially in the last decade, with more to prove such efficiency [1-15]. Many plants have been A 80% intensive studies for natural therapies. bout of used because of their antimicrobial properties, which individuals from developed countries use traditional are due to compounds synthesized in the secondary medicine, which has compounds derived from medicinal [16] D plants. Therefore, such plants should be investigated to metabolism of plant . iscovery and development of new better understand their properties, safety and efficiency. The therapeutic agents is a continuing process. In spite of the fact that, at present, we have at our command a formidable array of modern drugs, the need to discover and invent C D P B P B * orres’ponding author: epartment of lant iology and lant iotechnology, St. Xavier s College (Autonomous), Palayamkottai, Tamil Nadu, India. new agents is genuine and urgent. Adiantum, commonly Tel: +91 97 86 92 43 34 called as maiden - hair , has about 23 species in India Fax: + 91 46 22 56 17 65 E-mail: [email protected] widely distributed in the Indian subcontinent. Maximum Muraleedharannair Jalajakumari Mithraja et al ./Asian Pacific Journal of Tropical Biomedicine (2012))S40-S45 S41 number (15 species; 78%) of species occurs on the mountains specific difference in biological activity was observed in C. parasitica of south India [17]. All the species are commonly grown as the case of the fern (L.) H. Lev., which is a ornamental plants and majority of them are also used in highly polymorphic species [14]. Some gatherings of this traditional medicines to cure various diseases like cough, species show more biological activity in contrast to some fever, skin diseases, catarrhal affection, throat infection, other gatherings, which are without the same degree of bronchial disorders, dysentery, ulcer, epilepsy, leprosy, biological activity. In order to understand the reason for biliousness, inflammation, tumors of spleen, liver and such differences in biological activity, morphological, other viscera, cold, headache, piles, hair growth etc. It is phytochemical and bioactivity studies were carried out Adiantum lunulatum also considered as tonic and diuretic. among different gatherings based on the information from A. lunulatum Christella Burm. ( ), Adiantum capillus - veneris L and the literatures. It has been known that the fern C. Presl parasitica parasitica A (L ) H L (L ) T diantum radd“ianum ” “are sold in” the market by the . . ev. [ . ard.] is a trade names Hansraj and Paroshan . All the species polymorphic species with the variation in size of the entire are collected from the wild and none of the species is under plant, pubescens, depth of lobing of pinnae etc. In the mean cultivation. time there are two distinct morphotypes. They are glandular Adiantum caudatum A. caudatum L. ( ), rhizomes (are used and eglandular morphotypes. In the former, the leaves bear A. caudatum for cough and fever. Fronds of are used for orange coloured, elongated glands on the lower side of wound healing. It possesses antihelmintic, antimicrobial the costa, costules and veins, but such glands are absent Adiantum activity, antispasmodic, anti-asthmatic etc, in the eglandular morphotype. It is also important to note latifolium A. latifolium Lam ( ) possess antinociceptive and that the glandular morphotype is very rare in occurrence in anti-inflammatory activities. Ayurvedic Vaidyas describe contrast to the common eglandular morphotype [14]. When Adiantum lunulaturm A. lunulaturm the plant Lam ( ) as the gatherings, with and without biological activities, were pungent, alexiteric and used for indigestion. The decoction subjected to morphological studies, it was confirmed that of leaves is useful in dysentery, diseases of the blood, the glandular morphotype shows remarkable bioactivity [14]. ulcers and erysipeleas. Sporophylls are used in leprosy and The availability of such a wide range of variants in these erysipelas. The local people in the Aravalli Hills use the species presents a problem in the selection of suitable decoction of the leaves in cough, asthma and fever. The variant with good medicinal property. Reliable information paste of leaves is used in leprosy and erysipelas. It is also on the existing genetic variation is required for selection, used to overcome hair falling by putting paste of its leave breeding and conservation programmes of genetic resources. on head for an hour or so before taking bath for a fortnight. Phytochemical profile is the first hand indication of all In Mt. Abu area, Bheel uses this plant for urinary diseases the characters. Some belonging to the genera like and nose bleeding. In former the extract of leaves is taken Adiantum, Blechnum, Christella, Dicranopteris etc., from orally and the paste of leaves is applied on the lower Tirunelveli Hills of the Western Ghats were screened for portion of stomach, for clear and early release of urine. The their biological activities, particularly for antibacterial leaf extract is put in drops into the nose to stop bleeding, activity. But there is no report on the Kanyakumari District, during summer months. A. lunulatum possess antimicrobial Western Ghats, India. With this knowledge the present A. property. Leaf and root decoction is used for the chest study was aimed to examine the phyto-constituents of caudataum, A. latifolium, A. lunulatum, C. dentata C. complaints in Malaya. Fresh leaf decoction is given to cure and parasitica irregular menstrual cycle. Plant paste is given to women to . In addition it will provide chemical marker and help them to conceive. Seven triterpenoid are isolated from inter- specific variation between the medicinally important A. lunulatum A. caudataum, A. latifolium, A. lunulatum, C. whole plants of . Christella dentata (Forssk.) plants viz., C. dentate Christella parasitica dentata C. parasitica Brounsey & Jermy ( ) and (L.) and . C. parasitica H. Lev, ( ) extracts are used to treat gout and rheumatism. The decoction orally administered for 10 days 2. Materials and methods to cure spermatorrhea. Chiristella is possessed antifungal and antibacterial properties [18-27]. A. caudatum A. caudatum A. L. is largely used as substitute for Adiantum Healthy, disease free entire plants of L, latifolium A. lunulatum capilllus- veneris L. and there is also possibility for Lam and Burm were collected from the intentional or unintentional adulteration of common from Kakachi, Tirunelveli Hills (1 000 m) on the Western species in the place of rare species. Such adulterant may Ghats. Voucher specimen has been deposited in Centre not have the same medicinal property as in the original for Biodiversity and Biotechnology, Department of Botany, A. lunulatum Adiantum species. Moreover species like and Nesamony Memorial Christian College, Marthandam - 629 raddianum are polymorphic taxa with many morphotypes, 165, Tamil Nadu, India. The fresh materials were washed in cytotypes and ecotypes [28]. Most of the ferns show positive tap water 5 min and dried using blotting papers. The washed results with the difference in the degree of bioactivity plant materials were air and shade dried for two weeks and between species to species. But interestingly, intra- pulverized to powder using mortar. The dried and powered Muraleedharannair Jalajakumari Mithraja et al ./Asian Pacific Journal of Tropical Biomedicine (2012)S40-S45 S42 leaves materials (50 g) were extracted successively with 250 constituents with reference to solvents of the plant extracts. A. caudataum mL of petroleum ether, ethyl acetate, methanol, chloroform, The acetone extracts of showed maximum acetone, benzene and water by using Soxhlet extractor for presence of phyto-constituents (6/12), next to that chloroform, 8 h at a temperature not exceeding the boiling point of the ethanol and petroleum ether (4/12) followed by aqueous (3/12) A. caudataum solvent. The aqueous extracts were filtered using Whattman (Table 1). The benzene extracts of showed only filter paper (No.1) and then concentrated in vacuum at 40 曟 2 constituents presence. The results of the phytochemical using Rotary evaporator. The residues obtained were stored screening revealed that phenol and carbohydrates are A. caudataum in a freezer -70 曟 until further tests [29]. Phytochemical present in all the tested extracts of except A. screening of the extracts was carried out according to the benezene. Tannin also present in all tested extracts of caudataum standard methods [30-35]. except acetone extracts. Saponin is present in chloroform, petroleum ether and benzene extracts of A. A. caudataum caudataum. The acetone extracts of showed 3. Results the carboxylic acid and xanthoprotein presence. Acetone A. caudataum and ethanol extracts of showed the coumarin A. caudataum In the present study, the phytochemical screening was presence. The acetone extracts of showed the performed with acetone, benzene, chloroform, ethanol, unique presence of steroids (Table 1). A. latifolium petroleum ether and aqueous extracts of the whole plants The crude extracts of showed varied chemical A. caudataum, A. latifolium, A. lunulatum, C. dentata of profiles with reference to solvents of the plant extracts. The C. parasitica A. latifolium and . A total of five plants and 30 extracts aqueous and chloroform extracts of showed were examined for the phytochemical screening. The crude maximum occurrence of phyto-constituents (5/12), next to A. caudataum extracts of showed varied degree of phyto- that acetone and petroleum ether (4/12) followed by benzene Table 1 A. caudatum, A. latifolium A. lunulatum Phytochemical variation studies on and from Western Ghats, Kerala, India. A. caudatum A. latifolium A. lunulatum Compounds A B C W E P A B C W E P A B C W E P Alkaloids ------Phenols +++ - +++ ++ ++ +++ +++ + +++ ++ ++ +++ ++ + + + ++ + Flavonoids ------Saponins - +++ ++ - - ++ - +++ +++ - - +++ - ++ ++ - - ++ Proteins ------Quinones ------Steroids ++ ------+ ------Tannins - + ++ + + ++ + - + ++ - - - + - + - - Xanthoprotiens ++ - - - - - ++ - - ++ - - + - - + - - Carboxylic acids ++ ------++ + - - + - - - ++ - Coumarins ++ - - - + ------Carbohydrates ++ - ++ ++ + + +++ - +++ +++ +++ + - - ++ + ++ - Total 6 2 4 3 4 4 4 2 5 5 2 4 3 3 3 4 3 2 A - Acetone; B - Benzene; C - Chloroform; W - Aqueous; E - Ethanol; P - Petroleum Ether Table 2 C. dentata C. parasitica Phytochemical Variation studies on (Forssk.) Brounsey & Jermy and (L.) H. Lev. from Western Ghats, Kerala, India. C. dentata C. parasitica Compounds A B C W E P A B C W E P Alkaloids ------Phenols + +++ ++ - ++ ++ + + + ++ + + Flavonoids ------Saponins - +++ ++ - - ++ - ++ + - - + Proteins ------Quinones ------Steroids ------+ Tannins - + + ++ - - +++ - - ++ +++ - Xanthoprotiens + - - - - - ++ - - - - - Carboxylic acids - - ++ - - - +++ ++ - - +++ - Coumarins ------++ ++ +++ ++ +++ Carbohydrates ++ + +++ + + - - - +++ ++ + - Total 3 4 5 2 2 2 4 4 4 4 5 4 A - Acetone; B - Benzene; C - Chloroform; W - Aqueous; E - Ethanol; P - Petroleum Ether Muraleedharannair Jalajakumari Mithraja et al ./Asian Pacific Journal of Tropical Biomedicine (2012))S40-S45 S43 C. parasitica and ethanol (2/12) (Table 1). The phenol is present in all the ether extracts of (Table 2). A. latifolium tested extracts of . The steroid is present only in A. latifolium the petroleum ether extracts of . The carboxylic A. 4. Discussion acid is present in chloroform and aqueous extracts of latifolium. The tannin is present in acetone, chloroform A. latifolium and aqueous extracts of . The saponin is A total of three plants and 18 extracts were examined for A. A. caudataum, A. latifolium, present in benzene, petroleum ether and chloroform of the phytochemical screening latifolium. A. lunulatum. Xanthopreotein is existed only in acetone and Of which phenol is present in 17 out of A. latifolium aqueous extracts of . Carbohydrate is showed its 18 extracts tested. Saponin showed its presence in nine presence in aqueous, chloroform, petroleum ether, acetone extracts, tannin in 10 extracts, carbohydrates in 13 extracts, A. latifolium and ethanol extracts of (Table 1). xanthoprotein in five extracts, carboxylic acid in 4 extracts A. lunulatum The crude extracts of showed diverse phyto- and steroid and coumarin is present only in two extracts. The profiles with reference to solvents of the plant extracts. phytochemical studies revealed the variation and confirmed A. lunulatum The aqueous extracts of showed maximum the morphological and cytological differences. The steroid is A. lunulatum occurrence of phyto-constituents (4/12), next to that totally absent in the tested extracts of . Similar A. caudatum acetone, benzene, chloroform and ethanol (3/12) followed by to that caoboxylic acid also absent in and A. caudatum petroleum ether (2/12) (Table 1). The phenol is present in all Coumarin showed its presence only in . A. lunulatum the tested extracts of . The carboxylic acid is A total of two plants and twelve extracts were examined A. lunulatum present only in acetone and ethanol extracts of . for the phytochemical properties of Christella. Of which A. The tannin is present in benzene and aqueous extracts of phenol is present in 11 extracts tested. Saponin showed its lunulatum. The saponin is present in benzene, petroleum presence in 6 extracts, tannin in 6 extracts, carbohydrates A. lunulatum ether and chloroform of . Xanthopreotein in 8 extracts, xanthoprotein in 2 extracts, carboxylic acid A. C. parasitica is existed only in acetone and aqueous extracts of in 4 extracts and steroid in one extract of and lunulatum C. parasitica . Carbohydrate is showed its presence in aqueous, coumarin is present only in five extracts. The A. lunulatum chloroform and ethanol extracts of (Table 1). phytochemical studies revealed the variation and confirmed C. dentata The crude extracts of showed various phyto- the morphological and cytological differences. The steroid is C. dentata constituents with reference to solvents of the plant extracts. totally absent in the tested extracts of . Similar to C. dentata C. dentata The chloroform extracts of showed maximum that coumarin also absent in and caroboxylic acid C. dentata occurrence of phyto-constituents (5/12), next to that benzene showed its present only in chloroform extracts of C. parasitica (4/12) followed by acetone (3/12) (Table - 2). The phenol but in caroboxylic acid is absent in chloroform is present in all the tested extracts of C. dentate except extracts and showed its presence in ethanol, acetone and C. parasitica aqueous extract. The carboxylic acid is present only in benzene extracts of . The present study result C. dentata chloroform extract of . The tannin is present in provided a simple and cheap tool for the characterization C. dentata chloroform, benzene and aqueous extracts of . and identification of the medicinally important taxa. The saponin is present in benzene, petroleum ether and With the help the phytochemical screening we can easily C. dentata chloroform of . Xanthopreotein is existed only differentiate the Adiantum and Christella species. C. dentata. in acetone extracts of Carbohydrate is showed There are some studies on phytochemistry and C. parasitica A. lunulatum its presence in aqueous, chloroform, benzene, acetone and pharmacology on and [12-14, 18-20], C. dentata C. dentata, A. caudataum ethanol extracts of . (Table 2) but there is no report on and C. parasitica A. latifolium The crude extracts of illustrated different . In the case of Christella, the present study phyto-profiles with reference to solvents of the plant confirmed the previous observation and provided simple C. parasitica extracts. The ethanol extracts of showed and cheap chemical marker for the differentiation of the C. parasitica C. dentata maximum occurrence of phyto-constituents (5/12), next medicinally important plants from . to that acetone, benzene, chloroform, petroleum ether The presence of antimicrobial activity in a particular part and ethanol (4/12) (Table 2). The phenol is present in all of a particular species may be due to the presence of one or C. parasitica the tested extracts of . The carboxylic acid is more bioactive compounds such as alkaloids, glycosides, C. present in acetone, benezene and ethanol extracts of flavonoids, steroids, saponins etc. [36]. Recently, a number parasitica. The tannin is present in acetone, ethanol and of plants have been reported for antimicrobial properties C. parasitica aqueous extracts of . The saponin is present in across the world [1-16, 18-20]. In the present investigation, five C. parasitica benzene, petroleum ether and chloroform of . plants from India have been screened for phytochemical Xanthopreotein is existed only in acetone extracts of C. properties. The various phytochemical compounds detected parasitica. Carbohydrate is showed its presence in aqueous, are known to have beneficial importance in medicinal C. parasitica chloroform and ethanol extracts of . Coumarin is sciences. Many tannin-containing drugs are used in present in ethanol, chloroform, aqueous and petroleum ether medicine as astringent. They are used in the treatment of C. parasitica extracts of . Steroid is present only in petroleum burns as they precipitate the proteins of exposed tissues to Muraleedharannair Jalajakumari Mithraja et al ./Asian Pacific Journal of Tropical Biomedicine (2012)S40-S45 S44 form a protective covering. They are also medicinally used as healing agents in inflammation, leucorrhoea, gonorrhoea, Conflict of interest statement burns, piles and as antidote. Tannins have been found to have antiviral, antibacterial, antiparasitic effects, anti- inflammatory, antiulcer and antioxidant property for possible We declare that we have no conflict of interest. therapeutic applications. It was also reported that certain tannins were able to inhibit HIV replication selectively References and was also used as diuretic [37,38]. In the present study we revealed the tannins (16/30 extracts) presence in all the S [1] S ingh M, Singh N. Khare PB, Rawat AKS. Antimicrobial activity tested plants. aponins are considered a key ingredient in Adiantum C of some important species used traditionally in traditional hinese medicine and are responsible for most J Ethnopharmacol 115 of the observed biological effects. 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