PHYTOCHEMICAL STUDIES ON SMILAX MACROPHYLLA LINN.; A SOURCE PLANT OF CHOPACHEENI

Abstract

Chopacheeni is an important herbal drug widely used in Ayurveda. Chopacheeni is one among in the red list and in top 20 highly traded medicinal plants in India. Commonly known as Sarsaparilla, various species are available in the market in the name of Chopacheeni and are rarely Smilax china, the official source. The plant is considered as a remedy for Syphilis, Rheumatism, Skin diseases and Gout. Botanically authenticated drug is Smilax china but due to unavailability many adulteration is coming in the market to avoid this it is attempt to look for substitute of the drug of same genus so Smilax macrophylla Linn. was used for the phytochemical analysis. TLC of alcoholic extract of drug on silica gel "G" plate using Toluene (6.5 ml): Ethyl acetate (3.5 ml): Glacial acetic acid (0.2 ml) showed one spot under 366 nm UV, in 254 nm UV no spots, After spraying with Liebermann Burchard reagent followed by heating and then was visualized in day light which showed 2 prominent spots are seen.TLC using Chloroform (9.5 ml): Methanol (0.5 ml) showed two spots under 366 nm UV ,in 254 nm UV no spots were seen. After spraying, it showed one prominent spot. In HPTLC chromatogram showed 2 prominent spots in short wave UV 254 nm, one prominent spot in long wave UV 356 nm and 3 prominent spots at 400 nm. [PUBLICATION ABSTRACT]

Headnote

ABSTRACT

Chopacheeni is an important herbal drug widely used in Ayurveda. Chopacheeni is one among in the red list and in top 20 highly traded medicinal plants in India. Commonly known as Sarsaparilla, various species are available in the market in the name of Chopacheeni and are rarely Smilax china, the official source. The plant is considered as a remedy for Syphilis, Rheumatism, Skin diseases and Gout. Botanically authenticated drug is Smilax china but due to unavailability many adulteration is coming in the market to avoid this it is attempt to look for substitute of the drug of same genus so Smilax macrophylla Linn. was used for the phytochemical analysis. TLC of alcoholic extract of drug on silica gel "G" plate using Toluene (6.5 ml): Ethyl acetate (3.5 ml): Glacial acetic acid (0.2 ml) showed one spot under 366 nm UV, in 254 nm UV no spots, After spraying with Liebermann Burchard reagent followed by heating and then was visualized in day light which showed 2 prominent spots are seen.TLC using Chloroform (9.5 ml): Methanol (0.5 ml) showed two spots under 366 nm UV ,in 254 nm UV no spots were seen. After spraying, it showed one prominent spot. In HPTLC chromatogram showed 2 prominent spots in short wave UV 254 nm, one prominent spot in long wave UV 356 nm and 3 prominent spots at 400 nm.

KEYWORDS: Smilax macrophylla, Chopacheeni, Madhusnuhi, Phytochemical Analysis

INTRODUCTION

Smilax china Linn. of the Family Smilacaceae) is an important herbal drug widely used in Ayurveda. It is used for the treatment of Phiranga roga, Upadamsha, Vatavyadhi, Vrana (Vaidya Bapalal, 2005). It is said that the drug is similar to Ashwagandha (Sharma PV, 2005) in its properties and action. It is one among the red listed plants and in top 20 highly traded medicinal plants in India (http://www.megforest.gov.in). Now a day, due to commercialization and other economic interests, the pharmaceutical industry uses various source plants for the drug Chopacheeni. The availability of genuine samples is a burning issue since it is enlisted as endangered species (www.iucnredlist.org) and demand surpasses production causing confusion in the end users about the quality and safety also therapeutic ambiguity. Various species of Smilax are used as source plant for Chopacheeni; S. China Linn. , S. macrophylla Linn., S. zeylanica Linn. , S. regelli to name a few. S. china is formerly considered as the authentic identification as per The Wealth of India (Anonymous, 1950). Due to non availability and possible adulteration of cheaper substitutes, the therapeutic efficacy is obscure.

Hence there is an urgent need to explore the raw drugs for their quality through phytochemical investigations to establish the authenticity and logical reasoning behind multiple source plants of Chopacheeni. In the current research, roots and rhizomes of Smilax macrophylla Linn. which is a potential source for Chopacheeni was evaluated phyto-chemically for available active ingredients and their strength. Pharmacognostical investigation with macerate and powder study details and HPTLC finger printing of the rhizome and roots of Smilax macrophylla Linn. which helps in identification of crude drug is not available. Hence the present study has been carried out with following Aims & Objectives; Pharmacognostical and Phytochemical analysis of Smilax macrophylla Linn.

The vernacular names (Prajapati ND et al., 2003) of Chopacheeni (Smilax china) are: Sanskrit- Chopacheeni, Dvipantharavacha, Madhusnuhi (Chopra RN et al., 1956); English- Sarsaparilla, China root; Hindi- Jangli, Austibab, Chopachini; Marathi- Guti; Tamil- Malaith, Tamarai, Parangichekkarda; Telugu- Kondata mora, Malkaltamora; Kannada- Kaduhambu; Gujarati- Chopachini; Assamese- Aslussini; Bengali- Topachini

Smilax macrophylla Linn. is a large more or less prickly climber (Haines HH, 2000) growing in Himalaya eastwards from Kumaon at Assam, Bengal, Burma & South to Central Konkan extending to Java. The Stem is Smooth, striate (lines or several angled), armed with a few small distant prickles or almost unarmed; roots are rope-like originate from a short rhizome (Haines HH, 2000).

MATERIALS & METHODS

Plant Material:

Fresh roots and rhizomes were collected from the forests of Odisha during the month of November 2009. Botanical identification was done by expert taxonomists using local floras (Haines HH, 2000) and found to be Smilax macrophylla Linn. Voucher Ref no. IPGT&RA - 301). The collected samples were washed with potable water and chopped in to small pieces which were dried in shade, powdered and used for scientific evaluation.

* Pharmacognostical Study (Kokate CK, et al., 2008, Anonymous 2000, Trease and Evans, 2009): Morphological, Organoleptic, Microscopic and Histochemical study of the powdered drug was done as per the guidelines of Ayurvedic pharmacopoeia of India.

* Phytochemical study (Kasture AV et al., 2009, Anonymous 2000, Stahl E. 2005): Smilax macrophylla Linn. were analyzed by Physicochemical, Qualitative, Quantitative parameters. Chromatographic fingerprinting and Ultra-Violet Spectroscopy were carried out. TLC is mentioned as a primary tool for identification as part of monographs on all medicinal plants. Methanolic extract was used for the spotting of the TLC plate (Silica gel G Pre-coated plates). The solvent systems used in the study were Chloroform: Methanol and Toluene: Ethyl acetate: acetic acid. The sample extract was made to run on silica plate in various ratios. The ratio of 9.5:0.5 and 6.5:3.5:0.2 respectively has given good separation on trial method. Hence these systems are adopted for Chromatographic evaluation of the sample. Acetic acid was added for the second system for better separation. The resulting TLC pattern was viewed under long wave UV light at 366 nm and Short wave at 254 nm. Then after spraying with the Liebermann Burchard reagent and drying in a hot air oven and the number of spots viewed under daylight (Table no.4).

RESULTS & DISCUSSION:

Pharmacognostical Study: (Picture No.2)

* Morphological Study: Rhizomes were 5-6 × 3-4 cm in size, Root 20-23 × 1.5-2 cm in length Cylindrical & tapering towards apex, externally brownish and internally pinkish in color with rough and woody surface, and fracture coarsely fibrous.

* Organoleptic characters: The powder was reddish brown in color, with characteristic odor, slight bitter in taste and fibrous in texture.

* Powder Microscopy: The dried powder was mounted with distilled water to detect the Starch Grains, Cork, Simple Fiber, Acicular crystals, Scalariform vessels, Scleroids, Reticulate Vessels, Pitted Vessels, Parenchyma, Tracheids and Tannin. When stained with Iodine solution, Dil. FeCl, Conc. HCl and Phloroglucinol with Conc. Hcl, Showed Starch grains (Blue), Tannins (Bluish black), Crystals (Effervescence) and Lignified cells (Pink) respectively.

* Physicochemical Parameters: The sample was evaluated for physicochemical parameters like Loss on drying, Total Ash value, Acid insoluble ash, water, alcohol, chloroform, acetone soluble extractive values and for pH value (Table No. 1). The percentage of moisture content was 9.40%w/w, total ash 2.45%w/w, acid insoluble ash 0.15%w/w; Water soluble extractives 31.25%w/w, Alcohol soluble extractives 19.30%w/w, Chloroform soluble extractives 0.1%w/w and Acetone soluble extractives 8.58%w/w. pH was 5.28. Low total ash and Acid insoluble ash signifies low levels of inorganic matter and silica content. The high solubility of the sample in water denotes that drug is best suited for extraction with water or water based preparations. The negligible presence of Volatile oils is also in favor of thermal extractions with water.

* Qualitative chemical tests: Qualitative chemical tests for different functional units were estimated using water, methanol, chloroform and acetone soluble extractives of Smilax macrophylla Linn. Carbohydrates, Reducing sugars, proteins, Amino acids, saponins, alkaloids, Flavonoids and Tannin were qualitatively investigated. All functional units were present in water soluble extractive except amino acids and Flavonoids as these are the basic functional units necessary for metabolism in herbs. (Table No.2)

* Quantitative estimation: Traces of total Volatile oils, 0.08%w/w of total Alkaloids, 8.28%w/w of total Tannins and 22.85%w/w total Saponins were observed in the sample. Saponins are high molecular weight glycosides, consisting of a sugar moiety linked to a triterpene, steroid or steroid alkaloidal aglycone (Natural Remedies). Aglycone portion of the saponin is called as sapogenin. Triterpene saponins are the most common type in the plant kingdom. They show hemolytic activity and have bitter taste. Majority of the pharmacological and clinical action may be linked to these saponins in the case of Chopacheeni. (Table No.3).

* Thin Layer Chromatography Study: Thin layer chromatography of Methanol Extract of Smilax macrophylla Linn. Powder of the sample weighing 5 g were taken with 100 ml of alcohol and kept for twenty-four hours. Filtrate was prepared and evaporated till it was dried in a flat-bottomed shallow dish and concentrated on water bath to volume of requirement. TLC of alcoholic extract of drug on silica gel "G" plate using Toluene (6.5 ml): Ethyl acetate (3.5 ml): Glacial acetic acid (0.2 ml) showed one spot under 366 nm UV at Rf 0.23. Where as in 254 nm UV no spots were seen. After spraying with Liebermann Burchard reagent followed by heating and then was visualized in day light which showed 2 prominent spots at Rf 0.70 and 0.82. TLC using Chloroform (9.5 ml): Methanol (0.5 ml) showed two spots under 366 nm UV at Rf 0.04 and 0.83. Where as in 254 nm UV no spots were seen. After spraying, it showed one prominent spot at Rf 0.83.

* High-Performance Thin Layer Chromatography Study: Methanol extract of Smilax macrophylla Linn. was spotted on pre-coated silica gel GF 60254 aluminium plate as 5 mm bands, 5 mm apart and 1 cm from the edge of the plates, by means of a Camag Linomate V sample applicator fitted with a 100 µL Hamilton syringe. Chloroform (9.5 ml) and Methanol (0.5 ml) (v/v) (20 ml). The Chamber was saturated for 45 min, Development Time taken was 20 min and the development distance was 4.8cm. After development, Densitometric scanning was performed with a Camag T.L.C. scanner III in reflectance absorbance mode at 254 nm, 366 nm and 400 nm under control of win CATS software (V 1.2.1 Camag) (Picture No.2). The slit dimensions were 6 mm x 0.45 mm and the scanning speed was 20 mm s-1.

* However, chromatogram showed 2 prominent spots at Rf 0.08 and 0.56 in short wave UV 254 nm, one prominent spot at Rf 0.08 in long wave UV 356 nm and 3 prominent spots at 0.10, 0.28, 0.70 at 400 nm. (Table No.5 and Picture No. 3)

* UV Spectrophotometry: The spectrum was measured by placing the sample solution into the Shimadzu UV-160 Double beam spectrophotometer. Based on the UV Spectrophotometric analysis, the peaks, wavelengths and absorbance are shown in Table No.6. (Picture No.4)

CONCLUSION

Presence of more acicular crystals, Scalariform vessels, Scleroids, Reticulate Vessels, Pitted Vessels, is the identified character of Smilax macrophylla. The preliminary phytochemical analysis of the rhizome and root of Smilax macrophylla revealed the presence of Carbohydrates, Reducing sugar, saponin, protein, alkaloids and Tannins. The sample has got highest solubility in water followed by methanol. Hence drug is best suited for extraction with water or water based preparations. The Chromatographic finger printing was developed which could be useful for researchers to carry out further researches. The study is expected to be useful for quality control of sample and also will be a useful guide in deciding the source for Chopacheeni looking in to lack of availability and endangered status of the official source plant Smilax china Linn.

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Source of Support: Nil

Conflict of Interest: None Declared

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Cite this article:

Jyothi T, Acharya Rabinanaryan, Shukla C P, Harisha CR (2013), PHYTOCHEMICAL STUDIES ON SMILAX MACROPHYLLA LINN.; A SOURCE PLANT OF CHOPACHEENI, Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 110-117