Traditional Rigveda, Bhrigutantra, Asvini kumara, Charaksamhita and Sushrutsamhita. The

Traditional medicine used as treatment for Ocular diseases

Medicinal plants have been used as traditional treatment for numerous human diseases for thousands of years in many parts of the world. In rural areas of developing countries, herbal materials continue to be used as the primary source of medicines. The medicinal plants have been used in the world to prevent or cure diseases. Ophthalmic problems affects one-third of the population. Some of these can be treated with antibiotics and steroids but the prolonged use of these drugs has drawbacks. So herbal eye drops preparations (Ophthacare) with basic principles have been used as Ayurvedic system of medicine. Herbal eye drop is a polyherbal formulation used for anti-inflanmatory, antihistaminic effect and degenerative ophthalmic disorders.                         Biswas et al., 2001 reported that many traditional herbal medicines used in curing ocular diseases are now being gradually increased in modern medicine science. The Unani eye drop formulation is also now being used for ocular infections and it is prepared under aseptic conditions as per the method in unani pharmacopeia with slight variations Anonymous. The herbal eye drop formulation is prepared for beneficial effects in allergic and inflammatory conditions of the eyes. The herbal drugs used in the treatment of eye ailments dates to the days of Rigveda, Bhrigutantra, Asvini kumara, Charaksamhita and Sushrutsamhita. The W.H.O has revealed the importance of herbal cures and it has been active in creating guidelines and standards of botanical medicine. Various in-vitro experiments and animal studies described the therapeutic effect of medicinal plants in ophthalmic disorders. Ocular medications such as eye drops or an ointment are mainly used to treat and prevent eye diseases. Many traditional herbal eye drops are prepared from many medicinal plants combination which cures ophthalmic disorders (Klauss and Adala, 1994).

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1.5 Sea grasses

The researcher proposes a natural resources-lased treatment for ocular infections and the researcher identified sea grass as an alternative treatment. Sea grass is a grass-like plant that grows in and around the aquatic marine ecosystems. Sea grasses are the only angiosperms that grow in tidal and subtidal zones of seas, lagoons, gulfs, backwaters, bays and estuaries. They usually grow in muddy, sandy, clayey and coral rubble substrate and sometimes, it may also grow on rocks and in crevices. They grow, either homogeneously or heterogeneously and form thick and dense meadows that produce considerable biomass, provide excellent habitat and perform multiple ecosystem services for the world’s most bio diverse and productive marine ecosystems (Jagtap et al., 2003). However, this group of plants includes 13 genera and approximately 72 species that belong to the families of Cymodoceae, Potamogetonaceae, Posidoniaceae, Zosteraceae, Hydrocharitaceae and Ruppiaceae (Short et al., 2001).                        Sea grasses are flowering plants that are found in the sea and they belong to Monocotyledons. They play an important role in the ecology of various ecosystems. But only a limited number of studies deal with the chemistry of secondary metabolites of sea grasses (Larkum et al., 2006).

 

1.5.1 Taxonomy of Cymodocea serrulata                            

Kingdom                     :           Plantae

Phylum                        :           Tracheophyta

Class                            :           Liliopsida

Order                           :           Najadales

Family                         :           Cymodoceaceae

Genus                          :           Cymodocea                

Species                        :           serrulata

Other Names               :           Karumbupasi (Plate 1)        Plate 1: Cymodocea serrulata   

References                  :           Tanaka and Nakaoka (2006)

1.5.2 Taxonomy of Halodule pinifolia

Kingdom                     :           Plantae

Phylum                        :           Tracheophyta

Class                            :           Liliopsida

Order                           :           Najadales

Family                         :           Cymodoceaceae

Genus                          :           Halodule                                 Species                        :           pinifolia (Miki) Hartog Plate 2:  Halodule pinifolia

Other Names               :           Arugampul passi (Plate 2)

References                  :           Hartog, C. den (1970)

1.5.3 Taxonomy of Halophila ovalis

Kingdom                     :           Plantae

Phylum                        :           Tracheophyta

Class                            :           Liliopsida

Order                           :           Najadales

Family                         :           Cymodoceaceae

Genus                          :           Halophila                       

Species                        :           ovalis (R.Br.) Hook.f              Plate 3: Halophila ovalis

Other Names               :           Kouthupasi, Saethu passi (Plate 3)

References                  :           Kuo and Kirkman (1992)

Sea grasses comprise of few species, their importance to estuarine and coastal marine environments and to pharmaceuticals is significant. Generally, sea grasses are rich source of secondary metabolites which are believed to be a defense mechanism for to these plants. There are varieties of compounds present in it. Phytochemical screening, in other parts of the world, has clearly revealed that sea grasses have pharmaceutically potent secondary metabolites. These metabolites may be useful in containing infection, act as hypolipemic and hypoglycemic agents, reduce blood pressure and regulate blood cholesterol levels (Krishnamurthy, 2005). In folklore medicine, sea grasses have been used for a variety of remedial purposes, such as fever and skin diseases, muscle pains, wounds and stomach problems, remedy against stings of different kinds of rays, tranquilizer for babies (De la Torre castro et al., 2004). Research also depicts the important functional activities of marine seaweeds, such as antioxidant, anti-mutagen and anticoagulant effect, antitumor activity and an important role in the modification of lipid metabolism in the human body.

1.6 Compound Isolation from the sea grasses

The diversity of marine organisms has become an inspiration for researchers, to identify novel marine natural products, that could eventually be developed into therapeutics or pharmaceutical products. Marine organisms such as fungi, bacteria, sea grasses, seaweeds and sponges are taxonomically diverse and biologically active, offering a wide array, for the discovery of new anticancer drugs, derived from bioactive compounds, with medicinal properties, such as terpenoid derivatives, flavonoids, flavones, alkaloids, glycosides, polyphenolics and steroids (Boopathy and Kathiresan, 2010). Marine organisms represent a valuable source of new compounds. The biodiversity of the marine environment and the associated chemical diversity, constitute a practically unlimited resource of new active substances in the field of the development of bioactive products. More than 1,50,000 macro algae or seaweed species are found in oceans of the globe but only a few of them have been identified. Secondary or primary metabolites from these organisms may be potential bioactive compounds of interest for the pharmacological industry. Special attention has been reported for antiviral, anti bacterial and antifungal activities, related to marine algae, against several pathogens. The antimicrobial compounds, derived from the marine flora, consist of diverse groups of chemical compounds. The cell extracts and active constituents of various algae have been shown to have antibacterial activity against Gram positive and Gram negative bacteria (Ramalingam and Amutha, 2013).

1.7 Identification and characterization

The sea grass extracts usually contain and various types of bioactive compounds or phytochemicals, with different polarities and their separation still remains a big challenge for the identification and characterization of bioactive compounds. The isolation of these bioactive compounds contains different separation techniques such as TLC, column chromatography, flash chromatography, Sephadex chromatography and HPLC, to obtain a pure compound. The pure compounds are then used for the determination of structure, by using Fourier-Transform Infrared Spectroscopy (FT-IR), LC/MS and NMR studies.

1.7.1 Thin-Layer Chromatography (TLC)

TLC is a simple, quick, and inexpensive procedure, that gives the researcher a quick answer, as to how many components are in a mixture. TLC is also used to support the identity of a compound in a mixture when the Rf of a compound is compared with the Rf of a known compound. Additional tests involve the spraying of phytochemical screening reagents, which cause colour changes according to the phytochemicals existing in a sea grass extract, by viewing the plate under the UV light. This is used for confirmation of purity and identity of isolated compounds (Striegel and Hill, 1996).

1.7.2 Column Chromatography

Column Chromatography is a method, used to purify individual compounds, from a mixture of compounds. It is often used for preparative applications, on scales from micrograms up to kilograms. The main advantage of column chromatography is the relatively low cost. It is one of the most useful methods for the separation and purification of both solids and liquids. This is a solid – liquid technique in which the stationary phase is a solid and mobile phase is a liquid. The principle of column chromatography is based on differential adsorption of substance by the adsorbent. The usual adsorbents, employed in column chromatography, are silica, alumina, calcium carbonate, calcium phosphate, magnesia, starch, etc., and the selection of solvent is based on the nature of both the solvent and the adsorbent.

 

1.8 Spectral Analysis

1.8.1 Fourier-Transform Infrared Spectroscopy (FT-IR)

FT-IR analysis is used for the characterization and identification of compounds or functional groups (chemical bonds). Samples for FTIR can be prepared in a number of ways. For liquid samples, place one drop of sample between two plates of sodium chloride. The drop forms a thin film between the plates. Solid samples can be milled with potassium bromide (KBr) and then compressed into a thin pellet which can be analyzed. Otherwise, solid samples can also be dissolved in a solvent such as methylene chloride and the solution then placed onto a single salt plate. The solvent is then evaporated off, leaving a thin film of the original material on the plate (Eberhardt et al., 2007; Hazra et al., 2007).

1.8.2 Liquid chromatography/ Mass spectrometry (LC/MS)

Liquid Chromatography Mass Spectrometry (LC/MS) is an analytical chemistry laboratory technique, for identification, quantification and mass analysis of materials. This technique allows for the structural elucidation of unknown molecules through fragmentation. Liquid Chromatography Mass Spectrometry utilizes a compound’s intrinsic affinity for both a “mobile phase” (typically a buffered solvent) and a “stationary phase” (porous solid support with specialized coating). Essentially, a pump is used to provide a continuous flow of a solvent into which a dissolved sample is introduced. Once the sample is in the solvent flow, it travels through an analytical column. The compounds, present in the sample mixture, are then separated, depending on their affinity to the coated particles in the column. After the components in the sample are separated, they pass through a mass detector. The mass detector response and the “retention time” (time it takes for a compound to pass from the injector to the detector) of the compound(s) of interest, may then be compared to a reference material.

1.8.3 Nuclear magnetic resonance spectroscopy(NMR)

Nuclear Magnetic Resonance Spectroscopy, known as NMR Spectroscopy, which exploits the magnetic properties of certain atomic nuclei. This type of spectroscopy determines the physical and chemical properties of atoms or then molecules in which they are contained. It relies on the phenomenon of nuclear magnetic resonance and provides detailed information about the structure, dynamics, reaction state and chemical environment of molecules. The intermolecular magnetic field, around an atom in a molecule, changes the resonance frequency, thus giving access to details of the electronic structure of a molecule and its individual functional groups. Suitable samples range, from small compounds analyzed with 1 – dimensional proton or carbon – 13 NMR spectroscopy to large proteins or nucleic acids, using 3 or 4- dimensional techniques.

1.9 Antibacterial activity of the bioactive compound

Marine and estuarine submerged aquatic angiosperms or sea grasses produce antimicrobial compounds that may act to reduce or control microbial growth. Majority of works, carried out so far on sea grasses, are all in combination with seaweeds and other plants.

Naqvi et al., 1981, tested the extracts of seaweeds and sea grass from Indian coasts, for antiviral, antibacterial, antifungal, antiprotozoa, antifertility and pharmacological properties. Premanathan et al., (1992) tested the extract of the seaweed, sea grasses and mangroves for their antiviral activity. The methanol and hexane extracts of sea grass, E. acoroides and some seaweed were tested against Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Klebsiella pneumonia (Alam et al., 1994). Devi et al., (1997) tested the extract of 16 marine plants for the anti bactericidal activity, against 15 strains of marine fouling bacteria.                       Bhosale et al., 2002, studied the antifouling potential of some marine organisms against Bacillus and Pseudomonas species and reported that the sea grass Cymodocea rotundata exhibited mild activity against all the bacterial strains. Four new metabolites were isolated from organic extract of Cymodocea nodosa, collected from the coastal area of Porto germeno, in Attica, Greece and evaluated for their antibacterial activity against multi drug resistant pathogens, including methicillin-resistant strains of Staphylococcus aureus (Kontiza et al., 2008). The antibacterial properties of three sea grasses, namely, Cymodacea serrulata, Halophila ovalis and Zostera capensis were tested against human pathogens such as Staphylococcus aureus, Bacillus cereus, Bacillu subtilis, Escherichia coli, Salmonella paratyphi, Salmonella typhimurium and Micrococcus luteus, using six different solvents, namely, petroleum ether, chloroform, ethyl acetate, acetone, methanol and water (Sreenath Kumar et al., 2008).

The antibacterial activity of nine seaweeds, collected from the Kanyakumari Coast, inhibited activity against human upper respiratory tract pathogens, which include both gram-positive and gram-negative bacteria (Dharmesh et al., 2014).

Sivakumar and Vignesh 2014 reported that 1:1 proportion of seaweed extract: antibiotic Tetracycline such as Gracillaria grassa, Lobophora variegate, of red algae and    Stoechospermum marginatum, Padinagym nospora of brown algae, Cauler papeltata of green algae was more active compared to other groups of algae tested.