A Study On Evaluation Of Anti Microbial Activity Of Traditional Medicinal Plants

Anti microbial activity refers to the ability of a substance to kill or inhibit the growth of microorganisms such as bacteria, fungi, viruses and protozoa. This property is used to develop drugs and therapies that can help prevent or treat infections. Types of Antimicrobial activity includes Bacteriostatic that inhibits bacterial growth but doesn’t kill bacteria, Bactericidal that kills bacteria, Fungi static that inhibits fungal growth, fungicidalthatkillsfungi,Virustaticthatinhibitsviralreplication,Virucidal that kills viruses.

Examples of Anti microbial agents

Antibiotics-Target bacteria with examples including penicillin, tetracycline

Antifungal–Target fungi with examples flucanozole, clotrimazole

Antiviral–Target viruses includes drugs like acyclovir, valacyclovir

Antiparasite–Target protozoan’sand parasites such as metronidazole and ivermectin

Natural substances like medicinal plants, honey, essential oils also exhibit antimicrobial activity.

Antimicrobials can work through various mechanisms which include cell wall synthesis inhibition, protein synthesis inhibition, nucleic acid synthesis inhibition, cell membrane disruption, metabolic pathway inhibition. Importance of anti microbial activity is crucial in various fields including: Medicine–To treat infections and prevent the spread of diseases. Food industry–To preserve food and prevent spoilage Personal care – To develop products that prevent the growth of microorganisms on skin and wounds Understanding antimicrobial activity is crucial for developing new drugs, therapies, and products to compact microbial infections and promotes public health.

 Medicinal plants have been used for centuries to prevent and treat various infections. Antimicrobial properties have been a key aspect of their therapeutic value. The antimicrobial activity of medicinal plants refers to their ability to inhibit the growth of microorganisms including bacteria, fungi and viruses. This property has been exploited to develop natural remedies, supplements and even conventional drugs. These plants contain active compounds that exhibit anti microbial, anti-inflammatory and other beneficial properties. Natural and safe: Medicinal plants offer a natural and relatively safe way to prevent and treat infections, reducing the risk of adverse effects associated with synthetic drugs. Diverse chemical composition: Medicinal plants contain wide range of bioactive compounds including alkaloids, phenolic and terpenes which contribute to their antimicrobial activity Traditional knowledge: Many medicinal plants have been used in traditional medicine for centuries providing wealth of knowledge and experience. Many modern pharmaceuticals for example, Aspirin was developed based on salicin from willow bark. Ongoing research continues to discover new users and benefits of medicinal plants, as well as validated traditional uses scientific studies. Medicinal plants remain a critical component of health care, offering natural alternatives and complementary therapies for various health conditions. Their continued study and conservation are essential for future medical advancements.

Ocimumtenuiflorum : Ocimumtenuiflorum commonly known as Tulasi or Holy basil is a revered medicinal herb native to Indian sub continent. The aromatic plant has been used for centuries in ayurvedic medicine, prized for its wide range of health benefits. From its anti-inflammatory properties to its ability to support the immune system, Tulasi gained significant attention for  its potential as a natural remedy.

Anti microbial properties of Tulasi : Anti bacterial activity Tulasi extracts have been found to inhibit the growth of various pathogenic bacteria, including Staphylococcus, Escherichia, and salmonella species.

Azadirachtaindica: Azadirachtaindica also known as Neemormargosaornim tree. Native to the Indian sub continent and part of south East Asia, but is neutralized and grown around the world in tropical and subtropical areas.

Anti microbial properties of Neem : Broad spectrum anti microbial activity

Neem extracts have been shown to inhibit the growth of a wide range of bacteria, fungi and viruses making it a versatile antimicrobial agent.

Tinosporacordifolia : Tinosporacardifolia commonly known as Giloy or Tippateega is a well known medicinal plant in traditional Indian medicine, particularly in Ayurveda. It has been extensively studied for its various therapeutic properties, including its antimicrobial activity.

Anti microbial properties of Giloy: Anti bacterial Studies have shown that extracts from Tinosporacordifolia possess significant antibacterial activity against various Gram-positive bacteria such as Staphylococcus aureus and Bacillus subtilis It has also been effective against Gram-negative bacteria like Escherichia coli and Pseudomonas aeruginosa

MATERIALS AND METHODS : Collection of medicinal plants from different areas. Separated parts are washed, dried and grinded for extract formation, extraction by using suitable solvent system to perform anti microbial activity.

 Plant material collection: The plants were identified and collected from the locality of APSP, Kakinada. The fresh leaves of Tulsi, Neem, and Giloy are collected. Preparation of solvent extracts. The leaves of all plants were separated. They were carefully washed under running tap water followed by sterile distilled water to remove debris. They were dried under the sunlight for one week. Dried leaves are made into fine powder using a electric grinder until homogenous powder was obtained. Three different solvents namely acetone, ethanol, methanol were used for the plant extraction preparation.

Three different solvents namely acetone, ethanol, methanol were used for the plant extraction preparation.

(a). Acetone extract : 10 gm of powdered mixture of ocimumtenuiflorum, azdirachtaindica, and tinosporacardifolia was mixed with 100 ml of acetone. This mixture was kept in rotary shaker at100 rpm overnight and filtered with whatmann no.1 filter paper and concentrated to dryness at 40℃. The resulted crude filtrate was dissolved in 50% dimethyl sulphoxide (DMSO) as to get a concentration of 1µg/ml.

(b). Ethanol extract: 10 gm of powdered mixture  of ocimumtenuiflorum, azdirachtaindica, and tinosporacardifolia was mixed with 100 ml of ethanol. This mixture was kept in rotary shaker at 100 rpm over night and filtered with whatmann no.1 filter paper and concentrated to dryness at 40℃. The resulted crude filtrate was dissolved in 50% dimethyl sulphoxide (DMSO) as to get a concentration of 1µg/ml.

(c). Methanol extract: 10 gm of powdered mixture of ocimumtenuiflorum, azdirachtaindica, and tinosporacardifolia was mixed with 100 ml of methanol. This mixture was kept in  rotary shaker at 100 rpm over night and  filtered with whatmann no.1 filter paper and concentrated to dryness at 40℃. The resulted crude filtrate was dissolved in 50% dimethyl sulphoxide (DMSO) as to get a concentration of 1µg/ml.

Procedure: Prepared nutrient agar medium was autoclaved at 15 psi, at 121℃for 15min.Then aseptically transferred it into laminar air flow. Then the media was poured in petri plates and was get solidified in 20 minutes. Later the 10µl inoculum of Pseudomonas aeuroginosa, Staphylococcus aureus was spreaded on different indicating plates. A standard cork borer of 8 mm diameter was used to cut uniform wells on the surface of the nutrient agar. 10-50µl of each concentration of plant extracts was introduced in the wells by taking 20 µl of antibiotics (Tetracycline, Ciproflaxin, and Azithromycin) as control. The plates were incubated for 24 hours at 37℃ and the Zone of inhibition was measured to the nearest millimeter (mm).

Determining the Minimum inhibitory concentration: Determination of MIC s of the plant extracts was done by well diffusion and the concentrations of the extracts used were 10, 20, 30, 40, 50 µl. The lowest concentrations that did not permit any visible growth when compared with the control was considered as the minimum inhibitory concentration. A lower MIC value indicates that less plant extracts is required for inhibiting growth of the organisms therefore, plant extracts with lower MICs cores are more effective anti bacterial agents. Appropriate effective plant extract concentrations were identified.

RESULTS

Table.1.(a)–Zone of inhibition of Ocimum Tenuiflorum against Staphylococuss aureus

MIC Values of Acetone, Methanol and Ethanol are:

Acetone-2mm in 20µl , Methanol-2mmin 10µl,Ethanol-3mm in 30µl.

Table.1.(b)–zone of inhibition Ocimumtenuiflorum against Pseudomonas Aeuroginosa

MIC Values of Acetone, Methanol and Ethanol are:

Acetone-1mm in 10µl , Methanol-3mmin 10µl,Ethanol-2mm in 10µl.

Table.2,(a)–zone of inhibition of Azadirachtaindica against Staphylococuss aureus

MIC Values of Acetone,  Methanol and Ethanol are:

Acetone-4mm in 10µl , Methanol-2mmin 10µl,Ethanol-1mm in 10µl

Table.2.(b)–zone of inhibition of Azadirachtaindica against Pseudomonas Aeuroginosa

MIC Values of Acetone, Methanol and Ethanol are:

Acetone-2mm in 30µl , Methanol-4mmin 20µl,Ethanol-4mm in 10µl.

Table.3.(a)–zone of inhibition of Tinosporacordifolia against  Staphylococcus aureus

MIC Values of Acetone, Methanol and  Ethanol are:

Acetone-2mm in 10µl , Methanol-2mmin 20µl,Ethanol-1mm in 10µl.

Table,3.(b)–zone of inhibition of Tinosporacordifolia against Pseudomonas Aeuroginosa

MIC Values of Acetone, Methanol and Ethanol are:

Acetone-3mm in 40µl , Methanol-1mmin 10µl,Ethanol-1mm in 20µl.

DISCUSSION

This study showed the anti microbial activity of different plant extracts against microorganisms. The leaves of different medicinal plants are collected from the locality and they are made into an extract by using suitable solvent systems. They are tested for the antimicrobial activity against microorganisms. Anti microbial activity of different plant extracts against Staphylococcus aureus and Pseudomonas aeuroginosa were studied. .According to the results, extracts obtained from Ocimum, Azadirachta, tinospora shown to be with antimicrobial properties against Staphylococcus and Pseudomonas. As the concentration of the extract increases, their activity increases thus zone of inhibition also increases. Lower MIC value indicates less plant extract is required for inhibiting growth of organisms. The lowest concentration that did not permit any visible growth is known as Minimum Inhibitory Concentration.

CONCLUSION

In the present study antimicrobial activity of various  medicinal plants against microorganisms was evaluated, since ancient times, several medicinal plants  and their products have been used as sources for  curatives in  both developed and  developing countries. Because of their  potential to prevent diseases attributed to their antimicrobial as well as pharmacological properties. Based on this properties several plants considered to be the crucial sources of traditional medicines and drug manufacturing today and since the very beginning of civilizations. The medical world is on an immense requirement to discover novel antibiotics due to wide spread emergence of resistance among microbial pathogens against currently available antibiotics. However traditional plants have been proved to be better source for novel anti microbial drugs. In this study, antimicrobial activities of three traditional medicinal plants were assessed. The result showed potential antimicrobial effects against bacterial strains tested. They are rich sources of phyto-chemical constituents. These plants possess potent antimicrobial properties. They are easily available and renewable source of antimicrobial agent instead of synthetic chemicals.

REFERENCES

1. Banerjee, S; Kim,.Shariff, M; Khatoon, H., &Yusoff, S.M. (2012). Anti bacterial activity of Neem (Azadrachtaindica). Leaves of Vibrio spp. Isolated from cultured shrimp. Asian 2.  J. Animal VeterinaryADV, 3923: 1-7.
2. Bhargava, K.P., Gupta, M.B., Gupta, G.P., &Mitra, C.R. (1970): Anti-inflammatory activity of saponins and other natural products. Indian J Med Res. 58(6):724.Biswas, K., Chaltoppadhyay., I., Banerjee, R., & Bandhyo padhyay, U. (2002); Biologicalactivitiesand medicinalpropertiesof Neem (Azadirachtaindica). Current Science; 82(11):1336-45.
3. Blum, F.C., Singh, J., &Merrel, D.S. (2019). In vitro activity of neem (Azadirachtaindica) oil extract against Helicobacter pylori. J Ethnopharmacol:236-43.
4. Bodiba,D.C.,Prasad,P.,Srivastava,A.,Crampton,B.,&Lall,N.S. (2018). Antibacterial activity of Azadirachtaindica, Pongamiapinnata, Psidiumguajava and Magnifera Indica and their mechanism of action against Streptococcus mutans. Pharmacogn Mag. 14(54):76-80
5. Campos, E.V; de Oliveira, J.L; pascoli, M, de lima, R., &Fraceto, L.F. (2016). Neem oil and grop protection from now to the future. Front plant Sci;7: 1494.
6. Chaube, S.K., Shrivastav, T.G. Timari, M., Prasad, S., Tripathi, A., & Pandey, A.K. (2014). Neem (Azadirachtaindica)leafextractdeterioratesoocytequalitybyinucingRos-mediatedapoptosis in mammals. Springerplus. 3:464.
7. Chauhan, S., Jindal, M., Singh, P., &Teuiari, S. (2015). Antibacterial activity of Neem nanoemulsion and its toxicity assessment of human lymphocytesinvitro.IntJNanomedicine.10(1),77-86.
8. Jones,I.W.,Denholm,A.A.,Ley,S.V., Lovel, H., Wood, A., &Sinden, R.E. (1994). SharmaR,AminH,GalibR,PrajapatiPK.TherapeuticvistasofGuduchi (Tinosporacordifolia (willd.) Miers): a medico-historical memoir. J Res Educ Indian Med  2014;20 (2):121-35..
9. Sharma R,  Amin H,  Galib R,  Prajapati PK. (2015)Anti diabetic claims of Tinospora cordifolia (Willd.) Miers:critical appraisal and role in therapy. Asian Pac J Trop Biomed 2015;5(1):68-78
10. Sharma R, Kumar V, Ashok BK, Galib R, Prajapati PK, Ravishankar B. (2013) Evaluation of hypoglycaemic and anti-hyperglycaemic activities of Guduchi Ghana in Swiss albino mice. Int J Green Pharm 2013;7:145-8.
11. Sinha K,Mishra NP, Singh J,Khanuja SP.Tinospora cordifolia (Guduchi), (2004)a reservoir plant for therapeutic applications: A  review. Indian J Tradit Know 2004;3:257‑70.
12. AmaneH,Kaore S,KaoreN. (2014) In vitro study of anti microbial properties of Tinosporacordifolia (Guduchi). IntJ Pharm BioSci 2014;5(1):747-53.
13. Rose M, Noorulla KM, Asma M,Kalaichelvi R,Vadivel K,Thangabalan  B,et al. (2010) In-vitro antibacterial activity of methanolic root extract of Tinosporacordifolia (willd). Int J Pharm Res Dev 2010;2(5):1-5.
14. Islam MK, Ashakin K. (2011)Anti microbial screening and brineshrimplethality bioassay of Tinosporacordifolia (fam: Menispermaceae). Int J Pharm Sci Res 2011;2(11):3091-95.
15. Duraipandiyan V,Ignacimuthu S,Balakrishna K,Al-HarbiNA. (2003): Anti microbial activity of Tinosporacordifolia: an ethno medicinal plant. Asian J Trad Med Jeyachandran R, Xavier TF, Anand SP. Antibacterial activity of stem extracts of Tinosporacordifolia (willd) hook. F&thomson. Ancient Sci Life 2003;XXIII(1):40-43.
16. Verami A, Navneet, Gautam SS. (2016): Screening of antibacterial activity of Tinosporacordifolia Miers extracts against dental pathogens. J Pharmacol Mallikarjun S, Rao A,Rajesh G, Shenoy R, Pai M. Antimicrobial efficacy of Tulsi leaf (Ocimumsanctum) extract on periodontal pathogens: An in vitro study. J Indian Soc Periodontol; 2016; 20:145-50.
17. Eswar P,Devaraj CG, Agarwal P (2016);Anti-microbial Activity of Tulsi {OcimumSanctum(Linn.)} Extract on a Periodontal Pathogen in Human Dental Plaque: An Invitro Study; Journal of Clinical and Diagnostic Research. 2016; 10(3):ZC53-ZC56.
18. AliH,Dixit S (2012); In vitro anti microbial activity of flavanoids of Ocimum sanctum with synergistic effect of their combined form; Asian Pacific Journal of Tropical Disease 2012;S396- S398.
19. Hammer KA, Carson CF, Riley TV, (1999) Anti microbial activity of essential oils and other Plant extracts. J Appl Microbiol.1999; 86:985-990.
20. Singh V, Amdekar S, Verma O, Ocimum Sanctum (tulsi) (2010): Biopharmacological activities. Web Med Cent. Pharmacolo. 2010; 1:1-7.
21. Govind P,Madhuri S, (2006): Autochthonous herbal products in the treatment of cancer. Phytomedica. 2006; 7:99-104.
22. Rahal A, Singh V, Mehra D, Rajesh S, Ahmad AH, (2009): Prophylactic efficacy of Podophyllumhexandrum in alleviation of immobilization stress induced oxidative damage in rats. J Nat Prod 2009; 4:110-115.
23. Jirovetz,L.,Buchbauer,G.,Shafi M.P.and,Kaniampady M.M. (2003): Chemo taxonomical analysis of the essential oil aroma compounds of four different Ocimum species from southern India. European Food Research and Technology, 2003;217(2):120-124.
24. Malima, S. M., Massaga, J.J., Malecela, M. N. and Andrew, Y. (2013):  Repellence effectiveness of essential oils from some Tanzanian Ocimum and Hyptis plant species against afro tropical vectors of malaria and lymphatic filariasis. Journal of Medicinal Plants Research, 2013; 7(11):653-660.