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About Authors:
Deepak singh1*, Arpit Dixit2, Amir khan3, Vikas singh4, Abhishek sachan4
1*Department of Clinical Research, Jamia Hamdard, Hamdard nagar, New Delhi-110062
2Business executive at Merck, Ghaziabad, India
3Business executive at Cipla, Lucknow, India
4Shri RLT Institute of Pharmaceutical Science & Technology, Etawah(UP), India

The present investigations, which were primarily conducted with the aim of investigating some neuropharmacological activity of Polygonum glabrum (PG), i.e. PG has got anxiolytic activity when tested against open field exploratory behavior, where as elevated plus maze did not show any positive results. The action produced by PG was more than that of diazepam in open field exploratory behaviour. Observations confirms that PG possesses significant antidepressant activity. The observed antidepressant activity of PG was qualitatively comparable to that induced by Imipramine. Pentobarbitone induced hypnosis in mice was significant potentiated by PG.PG at 100 and 200mg/kg, reduced locomotor activity in rats.The PG seems to be little or no motor incoordination effect in mice when tested against rota-rod test.PG had significant analgesic activity which is both centrally and peripherally mediated, when tested against various analgesic models in rodents.The investigations indicates that PG has significant analgesic, anti-inflammatory, antidepressant and anxiolytic actions, some of these actions, including antidepressant and anxiolytic can be rationalized on the basis of the neurochemical data emanating from this study . The present study indicate that PG can be clinically useful not only in inflammation, pain and fever, and worm infestation but also in depression and anxiety. Clinical studies are required to confirm the above mentioned activities.

Reference Id: PHARMATUTOR-ART-2528

Polygonum species are known to ancient communities as useful medicinal plants. Polygonum glabrum Willd is a perennial plant belonging to Polygonaceae family. It is commonly known as Densflower knotweed. The genus Polygonum includes 150 species (Graham 1958)8. According to geographical origin, there are different synonyms for the plant. Polygonum glabrum Willd. Asia is similar to that tropical Africa and Egypt, but the Asian species has absolutely glabrous leaves with eciliate margins. Polygonum senegalense Meisn (Polygonum glabrum Willd) is found in Senegal; the leaves are glabrous with the limb surface exuding a viscous yellow liquid , occasionally, young leaves having deciduous tomentous pubescence are found (Hutchinson &Dalziel 1963; Berhaut 1967)44. In Kenya, there are two forms of  plant ; Polygonum senegalense formasenegalens, distinguished by dark green leaves with a yellow sticky oily residue covering the surface ,and  Polygonum senegalenes forma albotomentosum , distinguished  by the leaves covered with white hairs (Graham 1958)8. The species ofPolygonum glabrum found in India and Sudan seems morphologically comparable.

Indian Polygonum glabrum is a perennial herb growing up to a height of 150cm, distributed widely in the eastern India at altitude of 1900m, especially near liberal water source.Polygonum glabrum contains numerous compounds with documented biological activity.

Constituents that have stimulated the most interest include the sesquiterpenes, polyphenols like vannilic, syringic, p-hydro benzoic, protocatecheuic, galliccis & p-coumaric acids, kaempferol quercitrin and essential oils.

Quercetin, rhametin, quercitrin, avicularin and rutin were isolated from ethanolic extract of the leaves.(Tiwariet al;1979)9

In India the leaves of this plant are used for treatment of colic pain, fever, and for “stich in the side” (Kirtikaret al; 1975)6 while in Sudan they are used to treat roundworm and tapeworm infestations (O.Abdel-Moneim, Sudan National Council for Research 1979, personal communication). The Sudanese species has received little photochemical examination but molluscicidal properties and principals of the Kenya species have been reported (Dossajial1976: Maradufu&Ouma1978; Dossaji& Kubo 1980)10.

It is also reported to possess antiviral (Bhakuniet al; 1985)12 and antibacterial (Krishnamurthi, 1969)12 activities.

Polygonum glabrumhas been studied for anti-inflammatory (Bhupinderet al; 1987)4, analgesic, hypotensive and spasmogenic (Singh et al; 1985)11. The finding show that Polygonum glabrum extract is clinically effective as an anti-inflammatory drug and works by the mechanism of action similar to NSAIDs. Polygonum glabrum also has been researched for anthelmintic activity (Muddathiret al; 1987)13s,which showed activity especially against Hymenolepis nanavar fraternal.The above information reveals that Polygonum glabrum has got extensive pharmacological actions. The lack of viablepharmacological mechanism or the assurance of which components within the plant are critical for therapeutic effect (necessary in order to standardize formulations) . Over the centuries, societies around the world have developed their own traditions to make sense of medicinal plants and their uses. Some of those traditions and medicinal practices may seem strange and magical, others appear rational and sensible, but all of them are attempts to overcome illness and suffering with an aim to enhance the quality of life. Many of the thousands of plant species growing throughout the world have medicinal uses, containing active constituents that have a direct pharmacological action on the body. There is increasing interest in industry, academia and health sciences in medicinal plants57




Pharmacognostical studies:
• Collection and authentication of plant
• Drying and size reduction

• Isolation & Purification
•         TLC
•         HPLC

Phytochemical studies:
• Preliminary Phytochemical studies

Pharmacological studies
• Antidepressant activity

Pharmacognostical studies:

• Collection:
The leaves of Polygonum glabrum Willd were collecte during month of August from the Ayurvedic Garden of National Botanical Research Institute, Lucknow. Authentication was performed by joshi et al., (1987)23.

• Drying and size reduction:
The leaves of  Polygonum glabrum Willd were dried in the shade for about a week followed by drying at 30ºc-40ºc in oven for a day. The leaves were then grinded to coarse powder in Mortar and Pestle. The powdered materials was passed from sieve no. 20(#). Finally the powder is used for extraction.

Extraction was a process where by the desired constituents of a plant are removed using a solvent. The primary ways for extraction of organic molecules of interest to biologists and medical investigators involve breaking open the cells. the classical chemical procedure for obtaining organic constituents from dried plant tissue (heart wood , dried seeds, root , leaf) was to continuously extract powder material  in a soxhlet apparatus with a range of solvents , starting in tern with ether, petroleum and chloroform (to separates lipids and terpenoids ) and then using alcohol and ethyl acetate (for more polar compounds ). This method was useful when working on the gram scale.

Extraction Methods:
These are the extraction methods which were used in phytochemistry.
Maceration:  This method involves shocking and agitating the solvent and plant materials together. The solvent was then drained off. Remaining miscella was removed from the plant material through pressing or centrifuging. This method does not totally extract the active ingredients from the plant materials.

Percolation: With this method, the plant material is moistened with solvent and allowed to swell before being placed in one of a series of percolation chambers. The material is repeatedly rinsed with solvents until all the active ingredients have been removed. A solvent is reused until it is saturated. New solvent is used on plant material that is almost completely exhausted, and then reused on subsequently less exhausted batches. This method is more is more effective in obtaining active ingredients than the maceration technique.

Soxhlet extraction:
The use of commercially available soxhlet extractor was a convenient way to prepare crude plant extracts. This procedure was used mainly with pure solvents, although some although some authors have reported the use of binary or ternary solvent mixtures. Mixed solvents suffer the inconvenience that individual components may distill at different temperature, so that the resulting mixture in the chamber containing the drug was enriched in the solvent of lower boiling point. Thus actual solvent proportions in the extracting chamber differ from that originally used in the collector, and this fact may introduce errors when typing to reproduce the experiment using other extraction methods. The main advantage of extraction using a soxhlet apparatus was that it is an automatic, continuous method that does not require further manipulation other than concentration of the extractive and saves solvents by recycling it over the sample. Moreover, this method is not time-consuming, since for a standard sized sample (500g), the extraction time is less than 24 h. an obvious disadvantage is that the extractives were heated during the period of extraction at the boiling point of the solvent employed and thermally labile compounds such as carotenoids may hydrolyze or decompose.

Infusion and decoction:
Infusion and decoction are simple methods for extraction with water. In the latter, in the former case, hot or cold water is added to the milled drug; in the letter, the sample is boiled for about 15 min. in water. Extraction with water as the role solvent is seldom used for plant material, although some plant constituents are water-soluble, such as carbohydrates, flavonoid polyglycosides, quaternary alkaloids, saponins and tannins. As an example

•Extraction of Polygonum glabrum leaves
Dried leaves of Polygonum glabrum (about 100gms) was extracted with 300ml methanol (90%) in a Soxhlet apparatus for 72hrs. After extraction, the solvent was filtered under reduced pressure.

•Isolation and purification:
After extraction of natural products they are subjected to isolation and purification to ascertain what the natural product is and to carry out sufficient experimental work necessary to biologically characterize or profile the compound. The techniques available include, but are not limited to, solid-phase extraction, high-performance liquid chromatography (HPLC), gradient high-performance liquid chromatography, thin-layer chromatography (TLC), column chromatography, paper chromatography, etc. Chromatography is the method of choice in handling the problem of isolation of a compound of interest from a complex natural mixture 11. Therefore, the chromatographic methods commonly used during an isolation work are briefly described.

Chromatography is a separative technique that has arisen from the chemists’ interest to be able to separate a mixture of compounds into its constituents. The name chromatography was firstly used in 1906 by a botanist called Tswett; who worked to separate colored plant pigments. Chromatography means color-writing in Greek. Tswett made his experiments by allowing a solution of mixed pigments to pass through a column of crushed chalk and than he observed the separated color zones on the column. The international Union of Pure and Applied Chemistry (IUPAC) defined chromatography as follows:

Methods used primarily for the separation of components of a sample, in which components are distributed between two phases, one of which is stationary while other moves. The stationary phase may be a solid, or a liquid supported on a solid, or a gel. The stationary phase may be packed in a column, spread as a layer, or distributed as a film, etc. In these definitions “chromatographic bed” is used as a general term to denote any of the forms in which the stationary phase may be used. The mobile phase may be a gas or a liquid (Sewell, 1987).

Thin layer chromatography:
Thin layer chromatography is a rapid, simple, versatile, sensitive and inexpensive analytical technique for the separation molecules. The stationary phase in TLC is a thin layer of adsorbent that is spread uniformly over a plate. The plate can be prepared by spreading the adsorbent on glass in the lab or commercial pre-coated plates can be used. . The major advantage of TLC is the disposable nature of the plates. Samples do not have to undergo the extensive clean-up steps required for HPLC. The other major advantage is the ability to detect a wide range of compounds cheaply, using very reactive reagents (iodine vapors, sulfuric acid) or indicators. Nondestructive detection (fluorescent indicators in the plates, examination under a UV lamp) also means that purified samples can be scraped off the plate and be analyzed by other techniques. Thin layer chromatography has traditionally been considered as a pilot technique for column chromatography and high performance liquid chromatography. TLC has also been considered to be a suitable pilot technique for solvent or sorbent optimization because it is more flexible, more rapid and cheaper than column techniques8.

Chromatographic solvent "polarity"-
There is four major intermolecular interactions between sample and solvent molecules in liquid chromatography, dispersion, dipole, hydrogen-bonding, and dielectric. Dispersion interactions are the attraction between each pair of adjacent molecules, and are stronger for sample and solvent molecules with large refractive indices. Strong dipole interactions occur when both sample and solvent have permanent dipole moments that are aligned. Strong hydrogen-bonding interactions occur between proton donors and proton acceptors. Dielectric interactions favor the dissolution of ionic molecules in polar solvents. The total interaction of the solvent and sample is the sum of the four interactions. The total interaction for a sample or solvent molecule in all four ways is known as the "polarity" of the molecule. Polar solvents dissolve polar molecules. For normal phase partition chromatography, solvent strength increases with solvent polarity, whereas solvent strength decreases with increasing polarity in reverse-phase systems.



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