International Journal of Applied Ayurved Research ISSN: 2347- 6362 ANTIBACTERIAL ACTIVITY DECOCTION OF FENESTRATUM ON STAPHYLOCOCCUS AUREUS:AN EXPERIMENTAL STUDY 1Aberathna K.A.S, 2Sakunthala H.S, 1BAMS ,GampahaWickramarachchi institute, University of Kelaniya,Sri lanka , 2M.D (Ayur),Senior lecturer, Department of DravyaGunaVignana,GampahaWickramarachchi Ayurveda institute, University of Kelaniya, Sri lanka ABSTRACT Coscinium fenestratum is one of the most important used in traditional systems of medicine. Pharmacological properties of the have been mentioned as Vranajit and Vrananut in Ayurveda classics especially including wound cleansing property. Unawareness of this, the plant has escaped utilization of C.fenestratum as anti bacterium. The current study focused on the bacterial wounds, by Staphylococcus aureus and the efficacy was evaluated using decoction of C.fenestratum as a washing agent. For antibacterial susceptibility test, testing drug was prepared in six different concentrations homogenously. Prepared decoctions of stem were tested for its potent antibacterial activity against proto type of S.aureus from laboratory specimens. Followed the Antibacterial Sensitivity Test, in duplicate discs of modified Agar well diffusion method on bacterial strains and assessed the efficacy comparing with positive control as Amoxicillin. The mean inhibitory zones in samples were measured as 15.5mm, 14.75mm, 20.25mm, 16.5mm, 12.5mm & 18.75mm respectively. All of the testing samples indicated significant antibacterial potential on laboratory specimens of S.aureus. The current experimental study has revealed that the stems of C. fenestratum have remarkable antibacterial effective on the S. aureus. Keywords: Antibacterial activity, Coscinium fenestratum, S.aureus, stem,decoction INTRODUCTION:There are total of Morphologically , leaves are heart shape some 250,000 species of higher plants in and there are 6 inches length. Outer the world, much less than the species of surface of the leaves are yellow colour and animals (5-10 million). Coscinium there are yellow colour flowers. Fruits are fenestratum (Gaertn) Colebr is one of the small and brown colour. Stem and wood most important plants used in traditional are used for medicinal preparations. This systems of medicine. C. fenestratum is parts dried and taken as a medicine. belonging to family . The Vanuval and banval are the synonyms for vernacular name of the plant is Venivel in the C. fenestratum among the Sri Lankan Sinhala, False calumba or Tree Turmeric in people.3In SriLanka, people use C. 1 English and Atturam or Kadari in Tamil . fenestratum on behalf of Daru haridra This species observe in medium climatic plant (Berberis aristata ) that Indian conditions and occurs in the hills of people use. C.fenestratum is the substitute Malabar region, particularly Western for Daru haridra plant. The main chemical Ghats and in the jungles of South India, active compound of both plants is Malacca, Singapore Sumatra and Sri berberine4. Berberine is one of the major Lanka. It is common in the moist low- alkaloid constituent in C. fenestratum. 2 country forests in Sri Lanka . Berberine has broad spectrum of Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study pharmacological activities. Their Pancha originated due to any reason. Small holes padartha are similar and we can see when in the Vranas get the relations with the we following literature knowledge that external sensations. So that Jivanu (krimi) both plants have the same activity5. alias fungus, bacteria and virus insert to C. fenestratum is used for cleaning of the body through the Vrana according to wounds, for beauty culture, for eye the Ayurveda concept. Following are the diseases for skin diseases. Also C. Jivanu that can be seen with the relations fenestratum is used for making bath the of the Vrana. Staphylococcus aureus, small children. C.fenestratum is the Streptococcus pyogenes, Enterococci, energetic drink in fever conditions and Pseudomonas aeraginosa11. Among this general weakness of the body. organisms, Staphylococcus aureus was Also It has the Raktha shodaka (blood identified as the main organism that purifying) quality6. Such clinical survive in the open wounds12. indications show us that C. fenestratum is Staphylococcus is one of the five most a valuable drug in the preparation of common causes of infections after injury medicines.When there is a wound or surgery. It affects around 500,000 condition, our people used to the C. patients in American hospitals annually. It fenestratum as a drink. The experiments is abbreviated to “S. aureus” or “Staph have been revealed that this plant has the aureus” in medical literature. quality to reduce the tetanus due to In my research work, I attention for berberine chemical compound. Therefore wounds which affected by bacteria like it has been shown that our ancient people Staphylococcus aureus and aim of the concept was very reasonable. Activities of present study was to evaluate the effect of this plant has been mentioned as Vranajit decoction of C. Fenestratum as a washing and Vrananut in the Danwanthari nigandu agent on wound. It is expected by the and raaja nigandu7,8. Vruddatraya also cleaning a wound with the attacked by mentioned that C. fenestratum has the organisms such as bacterial, fungal and ability of Vrana shodana9. viral to remove or control their population. According to the Ayurveda definitions, According to the Ayurveda text, method of Vrana is known as the “Dosas treatment of wounds are sixty in number, samdgathaya” (aggravated dosas) that such as kashaya (washing with astringent originated in the one place of the human liquids), varti (keeping wicks of drug), body. The term “Vrana” is derived from avachurana (dusting powder of drug on the verbal root “vran” meaning “splitting/ wound), ropana (promoting healing), tearing of the body”.(causing discontinuity krimigna(destroying worms/antibacterial) of the skin and other tissues under it); ect. Out of these; kashaya, varti, kalka, since it causes discontinuity of the body it sarpi, taila, rasakriya, avachurana are is called vrana. Dalhana gives the meaning meant for sodhana (purifying) and ropana of the verb “vran” is causing (healing)13.Ayurvedic text explain mode of discolouration of the body. There are eight action washing agent should be mainly Vruna vasthu alias origin places of the shodana, ropana and krumighna. C. vrana as Tvak. mamsa, shira, snayu , fenestratum is such a valuable plant Asthi, sandhi, koshta, marma10. Epidermis medicine in Ayurveda. When we refer the is firstly invaded even the Vrana is Pancha padartha of the C. fenestratum, We

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Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study can see that there is the quality of Krimi prepared drugs in Ayurveda such as nashaka14.According to the Ayurvedic Kasisadi grithaya and gauradi grithaya definitions, the living matters that etc26. The qualities of this drugs are equal deactivate the human body functions were to Kaha (vacha) according to the described under the Krimi. The basic Drvyaguna geethaya27. principles of treatment methods for Krimi There are many information about C. (microorganisms) are krimi Apakarshana, fenestratum on the Vrana shodana effect. prakurthi Vigathaya and Nidana So that It is very important to find out the parivarjana15. it is mentioned under the information of the C. fenestratum in the prakrthi vigathaya chikistha as Materials research manner. Staphylococcus aureus is which have katu, ushna, tiktha and the abundant bacteria that survive with the kashaya rasas are very useful16. relation of the wounds. So that in this C.fenestratum also have the above experiment, that bacteria is used to qualities which are katu, tiktha, kashaya determine the objectives with the chemical and ushna and therefore it is identified as a stream. According to the Qwatha Krimi nashaka drug17.The following books paribasha of the Sharangadhara such as Thalpathe Peliyama, Deshiya samhitha, yellow colour stem (node parts) chikithsa samgrahaya which are the of the C. fenestratum which is the main traditional books in Ayurveda are part is used in the drug preparation. presented Specific yogas of Kriminashaka Prepared the six samples in different ability of C. fenestratum18,19. decoction and named A,B,C,D,E and F. Even the Uses of these plants have That decoction are put into the bacterial mentioned in vruna chikithsa and krimi medium and observe the growth of the chikithsa and both of chikithsa common bacteria. The well diffusion method is factor is to sterilize the organisms. used under the ABST (Anti-Bacterial According to the Ashtangahrdaya Sensitivity Test). After allowing to grow samhitha, C. fenestratum is used as the bacteria in the appropriate medias and Daruharidra for purifying the Siviya20. C. finally measured the inhibitory diameters. fenestratum also mentioned as Methodology “daruhelidda” under the vrana Collection of Plant Material prakshalana kashaya in the Dvi vruniya The node parts of venivel (C. fenistratum) chikithsa adyaya in charaka samhitha21.In were collected from Gampaha susrutha misrakadyaya mentioned C. Wickramaratchi Sidhayurvedha Ausheda fenestratum was used for Daruharidra Co(PVT) LTD, Yakkala who are supply under the vrana shodana kashaya in sri from Emiththirigala division, lanka22. The Shaka samgraha grantha that Sabaragamuwa province. is used abundantly in Ayurveda field also Identification Of Medicinal Plant mentioned about the C. fenestratum for Material cleaning of wounds23.it is also included in The medicinal plant material was the Sharangadara samhitha uththara kanda identified and authenticated by Prof. Dr. under the Vrana shodana yoga24.Vrana A.H.M. Tisera from the department of shothadikara of the Bavaprakasha also Dravya Guna ,Gampahawickramarachchi mentioned it’s action against the Ayurveda Institute, University of wound25. it also mentioned in the lot of Kelaniya, Yakkala, Sri Lanka.

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Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study

INCLUSION CRITERIA 1920ml boiled under moderate heat to Laboratory specimens of Staphylococcus obtain 240ml of decoction. aureus bacteria. Sample F- Put unknown grams of C. EXCLUSION CRITERIA fenistratum (hand measurement) , Water Other bacteria colonies in wounds. two tea cups boiled under moderate heat Drug Preparation for ½ hrs. According to the “Qwatha paribasha” positive controller - dilute the 10mg of of Sharangadhara samhita1 Amoxicillin,1ml of distil water. One pala (60g) of coarsely powdered Negative controller - distil water drugs is boiled with 16 parts of water in an Antibacterial Assay Of Samples earthen pot, over a mild fire till the liquid Followed the Antibacterial Sensitivity Test is reduced to 1/8 of the original quantity. (ABST) (Kirby-Bauer method) The effect this liquid is known as shrta, qwatha, of various plant decoctions on the bacterial kashaya or niryuha (decoction). strains were assayed by Agar well Instruments diffusion method. Measuring scale - Knife Instruments Measuring cylinder - Clay pot Auto clave - Hot air oven - Incubator Piece of clot - Measuring stick Petri dishes - Glass tube - Glass Flask Burner Glass beaker - Measuring cylinders Main Ingredients: Darvi (Coscinium Glass bottles - Micro pipette - Dry fenistratum) Assessory Material - swabs - Foils - Cotton buds - Ruler Water Gloves- Mask -well cutter Drug Preparation Method Main Ingredients First all the ingredients were identified. Nutrient agar - Distill water -Peptone Measured the required amounts of C. water - Muller hinton agar- fenistratum. Then cleaned well.After that Staphylococcus aureus (ATCC 25923) cut them into small pieces. Prepared six bacteria samples in different decoction. Nutrent Broth Preparation Sample A – C. fenistratum 60g, Water One litter of nutrient broth was prepared 120ml boiled under moderate heat to by dissolving 13 g of commercially obtain 15ml of decoction. available nutrient medium in 1000ml Sample B - C. fenistratum 60g, Water distilled water. The medium was dispensed 240ml boiled under moderate heat to as desired and sterilized by autoclaving at obtain 30ml of decoction. 15 lbs pressure (121ºC) for 15 minutes. Sample C- C. fenistratum 60g, Water After that keep the broth to cool in room 480ml boiled under moderate heat to temperature. Staphylococcus aureus obtain 60ml of decoction. (ATCC 25923) bacteria inoculated into Sample D- C. fenistratum60g, Water the broth to growth and incubated at 37ºC 960ml boiled under moderate heat to for 18hrs. obtain 120ml of decoction. MullerHinton Agar Medium Preparation Sample E- C. fenistratum60g, Water The medium was prepared by dissolving 38.16 g of the commercially available 1 SARNGADHARA-SAMHITHA( TEXT WITH Muller Hinton Agar Medium (HiMedia) in ENGLISH TRANSLATION), MADHYAMA KHANDA,QUATHA KALPANA 1000ml of distilled water and boiled to

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Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study dissolve the medium completely. The dissolved medium was autoclaved at 15 lbs pressure at 121°C for 15 minutes. The Table I -Zone Diameter / mm autoclaved medium was mixed well and controller poured onto 100mmsterile Petri plates (25- A B C D E F

Sample + ve -ve

30ml/plate) while still molten and allowed

to solidify at room temperature

Spread the Inoculum 1 16 15 21 16 12 19 40 00 Prepared the dilution series using the broth which are the maximum growth of 2 15 15 20 17 12 18 40 00

Staphylococcus aureus (ATCC 25923) 3 15 15 20 16 13 19 40 00 bacteria. -1 dilution series - put the 1ml broth in to 4 16 14 20 17 13 19 40 00 the 9ml peptone water

-2 dilution series - put the 1ml -1 dilution OBSERVATION & RESULT series in to the 9ml peptone water Fig I : Zone Diameter of sample -3 dilution series - put the 1ml -2 dilution series in to the 9ml peptone water. According to the turbidity standard method (0.5 McFarland standard) select the -1 dilution series. Get 100µl of -1 dilution series and put into the Muller Hinton agar plates. Using a sterile cotton swab spread cultures of the test organism was made on the Muller Hinton agar plates. Follow the Well Diffusion Method Four wells of 8 mm diameter were punched into the agar on each plate using a sterile well cutter. Into two wells in each plate 40 μl of the various plant decoction was added. other wells in each plate positive and negative controller was added. Amoxicillin was used as a positive control and Distil water used as a negative control .The plates were incubated at 37ºC for 24 hrs.The antibacterial activity was evaluated by measuring the zone of inhibition around the well. Used 4 samples in each and repeat the method

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Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study

In the present study, whole decoctions showed zone of inhibition around the well. Table III – T-test chart Sample A mean inhibitory zone diameter Hypothesis was 15.5mm.Sample B mean inhibitory H0 : µ ≤ 19 Vs H1 : µ > 19 zone diameter was 14.75mm.Sample C By considering the above table, A,B,D,E mean inhibitory zone diameter was and F samples mean less than or equal 19 20.25mm.Sample D mean inhibitory zone under 5% significant level. But H0 is rejected under 5% significant level for diameter was 16.5mm.Sample E mean sample C. So sample C mean greater than inhibitory zone diameter was 19.These results indicate that the sample C 12.5mm.Sample F mean inhibitory zone was in standard sensitivity value of diameter was 18.75mm.Thereof Sample C Amoxicillin (= 20 or>20mm). which are the put 8 times water showed CONCLUSION maximum activity against Staphylococcus All of Coscinium fenistratum decoctions aureus. Secondary Sample F showed best have antibacterial effect against laboratory specimens of Staphylococcus aureus. activity and Sample D, A, B showed their activity respectively. Sample E showed Sample C which are the put 8 times water lowest activity against Staphylococcus showed maximum activity against aureus. Staphylococcus aureus.Sample C was in According to Comparative data analyzing, standard sensitivity value of Amoxicillin (= 20 or>20mm). the significant antibacterial activity of the Amoxicillin is normally used as an oral C. Fenistratum decoctions were compared antibiotic for bacterial infections. with the standard antibiotic, Amoxicillin. Coscinium fenistratum decoctions are Standard Amoxicillin Sensitivity chart 2 external application. But Sample C was in standard sensitivity value of Amoxicillin (= 20 or>20mm). So it is better to use a local antibacterial application for comparing the efficacy of Coscinium fenistratum. Acknowledgement I would like to express my utmost gratitude to my supervisor, Senior lecturer Dr. H. S. Sakunthala for her guidance, Table II - Standard Amoxicillin perpetual energy, enthusiasm in research Sensitivity chart and educating me during my research. I According to T-test could not imagine the success of this research without her supervising. Also Test of μ =< 19 vs > 19 thanks to staff of the Laboratory and Pharmaceutical factory of the Gampaha Wickramarachchi Ayurveda Institute, University of Kelaniya, Sri lanka. My deepest gratitude goes to my mother, brothers and sister and relatives for their love, care and support throughout my life. REFERENSES 1) Research article, Berberine Content 2 Fall 2011 – Jackie Reynolds, Richland in Coscinium fenestratum (Gaertn.) Colebr College, BIOL 2421 Grown in Sri Lanka, Volume 3 Issue 12,

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2012, L.D.A.M. Arawwawala and W.A.N. (Chikithsa sthana 01) Department of state Wickramaarachchi language, Colombo, Sri Lanka. 2) Research article, Berberine Content (Buddhadasa , 2007) in Coscinium fenestratum (Gaertn.) Colebr 14) Arya Vaidya sala(1994), Indian Grown in Sri Lanka, Volume 3 Issue 12, medicinal plant ,Volume II,(191page), 2012, L.D.A.M. Arawwawala and W.A.N. Orient Longman,Chennai,India. Wickramaarachchi (Arya ,1994) 3) Osuthuru vesituru(1994), Volume 15) Kumarasingha A. (1994), Charaka iv (320 page), Department of Ayurveda, samhita, Sinhala translation,Volume III , Tharuna seva Publishers, Maharagama, Sri Chapter VII, Department of state language, Lanka (Osuthuru vesituru,1994) Colombo,Sri Lanka. 4) Arya Vaidya sala(1994), Indian (Kumarasingha , 1994) medicinal plant ,Volume II,(191page), 16) Kumarasingha A. (1994), Charaka Orient Longman,Chennai,India. samhita, Sinhala translation,Volume III , (Arya ,1994) Chapter VII, Department of state language, 5) Ayurveda pharmacopoeia(1994), Colombo,Sri Lanka. 1st step,Volume II (93page),Department of (Kumarasingha , 1994) Ayurveda, Tharuna seva Publishers, 17) Ayurveda pharmacopoeia(1994), Maharagama, Sri Lanka 1 st step,Volume II (93page),Department of (Ayurveda pharmacopoeia,1994) Ayurveda, Tharuna seva Publishers, 6) Osuthuru vesituru(1994), Volume Maharagama, Sri Lanka iv (320 page), Department of Ayurveda, (Ayurveda pharmacopoeia,1994) Tharuna seva Publishers, Maharagama, Sri 18) Kumarasingha A. (1984), Deshiya Lanka chikithsa samgrahaya, Department of (Osuthuru vesituru,1994) Ayurveda, Sathosa publisher, Colombo,Sri 7) Ethupa padhadharibi: B.A(1947), Lanka. (Kumarasingha , 1994 Danvanthari Niganduva, Ananda grama 19) Talpate piliyam (1992), Volume Publishers,India. III, Department of Ayurveda, Tharuna (Ethupa padhadharibi: ,1947) seva Publishers, Maharagama, Sri Lanka 8) Pandith Narahari (1982), Raaja (Talpate piliyam ,1992) Niganduva,Oriental Publishers, Varanasi, 20) Buddhadasa R.(2007), India. (Narahari , 1982) Ashatangaharda samhita, Sinhala 9) Kumarasingha A. (1994), Charaka translation, 2nd edition, Department of samhita, Sinhala translation,Volume III , state language, Colombo, SriLanka. Department of state language, (Buddhadasa , 2007) Colombo,Sri Lanka. 21 ) Kumarasingha A. (1994), Charaka (Kumarasingha , 1994) samhita , Sinhala translation,Volume III , Buddhadasa R.(2007), Susruta samhita, Department of state language, Sinhala translation, 2nd edition, Colombo,Sri Lanka. Department of state language, Colombo, (Kumarasingha , 1994) Sri Lanka. 22 ) Buddhadasa R.(2007), Susruta 10) Buddhadasa R.(2007), Susruta samhita, Sinhala translation, 2nd samhita, Sinhala translation, 2nd edition,(Sutrasthana 28) Department of edition,(Sutra sthana 21,28) Department state language, Colombo, Sri Lanka. of state language, Colombo, Sri Lanka. (Buddhadasa , 2007) (Buddhadasa , 2007) 23) Oushadiya shaka samgrahaya 11) Dermnetnz.org//wound.infection.html (2003), Volume III (145page), Department 12) Wiki.Answers.com of Ayurveda, Depani Publishers, 13) Buddhadasa R.(2007), Susruta Nugegoda , Sri Lanka samhita, Sinhala translation, 2nd edition, (Oushadiya shaka samgrahaya ,2003)

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Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study

24) Vithana nagoda P.(2001), Sri Sarangadara samhitha, (Uttharasthanaya, Chapter11), Samayavardana Publisher, Maradana,Colombo, Sri Lanka. (Vithana, 2001) 25) Buddhadasa R.(1982), Bavah Prakashaya, Sinhala translation, Volume II,(Vruna shothadikaraya) Department of Ayurveda, Depani Publishers, Nugegoda , Sri Lanka. (Buddhadasa , 2007) 26) Vithana nagoda P.(2001), Sri Sarangadara samhitha, (Uttharasthanaya, Chapter11), Samayavardana Publisher, Maradana,Colombo, Sri Lanka. (Vithana, 2001) 27) Dr: Veerasingha Leelarathna(1975), Dravya guna geethaya, Volume I (Dr: Veerasingha , 1975)

Corresponding Author: Dr.Aberathna K.A.S,BAMS ,GampahaWickramarachchi Ayurveda institute, University of Kelaniya,Sri lanka , Email;[email protected]

Source of support: Nil Conflict of interest: None Declared Cite this Article as : [Aberathna K.A.S et al;Antibacterial activity Decoction of Coscinium Fenestratum on Staphylococcus Aureus ;An Experimental Study] www.ijaar.in : IJAAR VOLUME III ISSUE II SEP-OCT 2017 Page No:873-880

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