INTRODUCTION:

Plants have been generally perceived as a significant source of novel therapeutic compounds since ancient times for the treatment of different illnesses and were accounted for in conventional medication systems like the Siddha and Ayurveda¹.

Bioactive mixtures are referred to for their serum obstruction and as a dependable source of antimicrobial medicines, and are generally utilized as hostile to infective enhancements and aides in mix with other compounds². Be that as it may, the properties of common based combinations should be deliberately explored to decide their pharmacological consequences for natural frameworks^3-4. Roughly 80% of the drugs of the world rely upon plant-based bioactive parts for relieving different diseases^5-7. The effective result of conventional medication has guided the quest for advanced chemotherapeutic substitutes to eliminate the disease brought about by drug-resistant organisms and to downsize the harm brought about by the antibiotic⁸. Notwithstanding, plants have a practically unending capacity to create intensifies that have particular bioactive rules that we can't integrate. The mending property of numerous herbal drugs has been perceived in numerous ancient cultures⁹.

On an exemplification, enormous number of plant species have not been read for likely therapeutic worth or portrayed for it yet different examinations have been distributed, exploring the anti fungal and anti microbial exercises of plant determined mixtures against a scope of pathogens^10-12.

Inflammation is a significant physiological response, which happens because of a wide assortment of harmful specialists (bacterial contamination or actual injury) eventually intending to play out the double capacity of restricting harm and advancing tissue repair^13-15.It requires the investment of different cell types communicating and responding to assorted go betweens along an exact sequence^16-18. In spite of the fact that inflammation is helpful in the setting of safeguard of the host against irresistible trespassers, if unchecked it can add to the pathogenesis of basic persistent incendiary sicknesses like atherosclerosis, obesity induced insulin obstruction, joint pain, chronic inflammatory bowel diseases and multiple sclerosis¹⁹. Chronic inflammation starts 2-4 days after the beginning of intense reaction and can keep going for quite a long time to months or years because of the persistant of the starting improvement, obstruction of the typical mending measure, rehashed episodes of intense irritation or poor-quality seething because of proceeded with creation of resistant reaction mediators²⁰.

Numerous non-steroidal anti-inflammatory drugs (NSAID's) like aspirin, phenylbutazone, indomethacin and so forth are in clinical use however, all these are not totally devoid of side effects²¹.

Hibiscus genus is the flowering plants in the mallow family, Malvaceae. The variety contains a few hundred species that are local to warm-mild, subtropical, tropical areas all through the world and is very enormous. Hibiscus plant has various clinical uses in Chinese Herbology. Their examination demonstrates some potential in restorative skin health management; for instance, a concentrate from the flowers of Hibiscus plant has been appeared to work as sun screen specialist by engrossing bright radiation¹⁸. In the Indian customary arrangement of medication, Ayurveda, Hibiscus, particularly white Hibiscus and red Hibiscus are considered to have therapeutic properties. The roots are utilized to make different creations accepted to fix infirmities like hack, going bald or hair turning gray. The leaves and flowers are ground into a fine glue with a little water, and the subsequent foamy glue is utilized as a cleanser in addition to conditioner. A past creature study showed the impacts of the Hibiscus plant remove on atherosclerosis in rabbits. Prominently, a decrease in fatty oil, cholesterol, and low thickness lipoprotein was seen in rabbits devouring an elevated cholesterol diet (HCD) notwithstanding Hibiscus plant remove contrasted with hares just took care of HCD, proposing a helpful impact^19-21. Besides, the Hibiscus seed is plentiful in phytosterol and tocopherol, plant types of cholesterol that has cell reinforcement and LDL cholesterol bringing down impacts ^23-25. In view of the writing review, so far no work has been completed in Hibiscus hirtus Linn. (HH). Antimicrobial and calming exercises have been accounted for in certain types of the sort Hibiscus.

The current examination is hence attempted to assess the anti microbial and anti-inflammatory effects of ethanolic concentrate of Hibiscus hirtus L.

METHODS AND METHODOLOGY:

The collected plant specimen was pressed against news paper and left for 24 hours. The paper was changed ordinarily till total drying. The dried specimen was mounted on the standard size of herbarium sheet. The specimen was fixed into herbarium sheet and named. The voucher Specimen was saved in the Pharmacognosy Laboratory of A.K.R.G College of Pharmacy, Nallajerla, West Godavari District, Andhra Pradesh. The fresh plant of Hibiscus hirtus with the family Malavaceae was collected from pothavaram, West Godavari District, Andhra Pradesh, on November 2019. The plant was recognized and confirmed by the department of Botany, AKRG College of Arts and Sciences, Nallajerla. Subsequent to collection of the leaves they were washed and left for conceal drying on the floor over the papers for ten days. After that the leaves were dried in hot air broiler at 40°C for an hour not long prior to beginning the extraction interaction to eliminate the harmony dampness content.

Extraction and maceration of leaves:

Subsequent to drying, plant samples were size reduced by grinder. The dried powdered material was gauged and exposed to extraction by the cycle of maceration utilizing organic solvent ethanol for 3 days with intermittent blending. After the extraction, extricates were separated by Whatman channel paper and filtrate was concentrated to acquire rough concentrates. The dried concentrates were gathered, alluded to as ethanolic concentrate of Hibiscus hirtus leaves, moved into a sterile container and put away at 40°C for the phytochemical screening and for the antibacterial and anti-inflammatory activities.

Preliminary phytochemical screening:

The priliminary phytochemical screening of concentrate was done adhering to standard method^25-37. All the chemicals and reagents utilized were of analytical grade.

Table[1]:Chemical tests for the ethanolic extract of leaves of Hibiscus hirsutus L.

Name of the test

Ethanolic extract of leaves of Hibiscus hirsutus L.

Steroids:

A)Salkowski Test:

B)Liebermann Buchard’s Test:

(+) ve

Triterpenes:

A) Salkowski Test:

B)Liebermann- Buchard’s Test:

(-) ve

Alkaloids:

A)Mayer’s Test:

B)Dragendroff’s Test:

C)Wagner’s Test:

D)Hager’s Test:

(-) ve

Carbohydrates

A)Fehling’s Test:

B)Molisch’s Test:

D)Barfoed’s Test:

C)Benedict’s Test:

(+) ve

+) ve

Flavonoids:

A)Shinoda Test:

B)Ferric chloride Test:

C)Lead Acetate Test:

D)Zinc- Hcl reduction Test:

(+) ve

Tannins:

A)Ferric Chloride Test:

B)Gelatin Test:

(+) ve

Glycosides:

A)Baljet Test:

B)Legal Test:

C)Keller- Killiani Test

D)Kedde's test:

(+) ve

(-)ve

Antibacterial activity of the Hibiscus hirtus leaf extract:

The antimicrobial activity of any sample of natural origin is assayed separately using the agar diffusion method employing 24 hours cultures of test bacteria and fungi strains. To carry out these studies, disc diffusion method and agar cup plate method are to be routinely adopted. However, in the present studies only the later method has been chosen. The effect of the extract and the standard drug were analyzed by measuring the diameter of inhibition zones in millimeter are recorded.

Preparation of test solution:

The stock solution of test compounds was prepared by dissolving the dried extract at a concentration of 100 mg/ml in distilled water. This was named as solution-A. From this solution-A, solution-B was prepared by diluting 50ml with 50ml distilled water. Then the required dilutions like 2.5mg/ml, 5.0mg/ml, 7.5mg/ml, 10.0mg/ml, 12.5mg/ml and 15.0mg/ml were prepared by diluting each concentration in 100ml distilled water.

Preparation of standard solution:

The stock solution of standard compounds was prepared by dissolving 250mg in 250ml distilled water to make a concentration of 1 mg/ml. From this solution the required dilutions like 2.5mg/ml, 5.0mg/ml, 7.5mg/ml, 10.0mg/ml, 12.5mg/ml and 15.0 mg/ml were prepared by diluting each concentration in 100ml distilled water.

Method used for the experiment: Cup-Plate Agar Diffusion Method:

Test organisms:

Gram (+) ve bacteria: Staphylococcus aureus,

Gram (-) ve bacteria: Escherichia coli

Composition of Nutrient Agar Medium:

Component	Quantity
Peptone	5g
Sodium chloride	5g
Meat extract	5g
Agar-agar	20g
Distilled water	Upto 1000ml
P^H	7.22-7.4

Procedure:

28g of powdered nutrient agar medium (Hi-Media) was suspended in 1000ml of hot distilled water. The prepared nutrient agar medium was sterilized in autoclave at 120°c (15 lb/sq.in) for 20 minutes. 25 ml of the nutrient agar medium prepared was inoculated with 24 hours old stock cultures of the above-mentioned test organisms and were transferred into sterile 15cm diameter Petri dishes. The medium in the plates were allowed to set at room temperature for about 30 minutes to solidify in laminar air flow unit. 4 cups of 6mm diameter were made in each plate at equal distance. From the ethanolic extract the dilutions (2.5mg/ml, 5.0mg/ml, 7.5 mg/ml, 10.0mg/ml, 12.5mg/ml, 15.0 mg/ml) are prepared. From the standard solution dilutions 2.5mg/ml, 5.0mg/ml, 7.5mg/ml, 10.0mg/ml, 12.5mg/ml and 15.0mg/ml are prepared. The standard and test samples were then added to each cavities with equal volume. The petridishes were left for 1-4hrs at room temperature and then incubated at 30-35⁰C for 18hrs.

Observation:

Fig. 1: Zone of inhibition of S.aureus

After incubation, all plates were analyzed for the appearance of inhibition zone around the extract loaded well and zone of growth inhibition was measured in mm.

Fig. 2: Zone of inhibition of E. coli

Antibacterial activity of ethanolic extracts of the leaves of Hibiscus hirsutus L.

S. No.	Concentration (mg/ml)	Diameter of zone of inhibition(mm)
		*E.coli*		*S.aureus*
		Standard (amoxycillin)	Test	Standard (amoxycillin)	Test
1	2.5	10	12	19	8
2	5	13	14	23	9
3	7.5	15	16	24	12
4	10	18	19	22	16
5	12.5	20	22	25	13
6	15	23	25	27	17

Anti inflammatory activity:

Chemicals:

1. Ibuprofen: It was obtained from Sun pharmaceuticals, New Delhi.

2. Formalin: This phlogestic agent Formalin (prepared in the laboratory) as 2% solution and was used to induce chronic inflammation in the rat’s right hind paw. Not more than 0.1ml was administered by the sub plantar route.

Digital plethysmometer:

The IITC 520 Plethysmometer was used to measure the paw volume. It consists of a water cell which is fitted on to a plexi glass stand. This water cell has an inlet in which the paw is dipped and an outlet with a stopcock. The water cell is connected to an electronic display which shows the volume of displacement of water in millilitre (ml) from the water cell when paw is dipped into water cell which is taken as paw volume. The paw is inserted into water, contained in a special water cell of which the resistance is changed due to the immersion of the animal’s paw. This resistance change is calibrated in ml and displayed on the read out in ml with a resolution of 0.1ml.

Formalin induced paw edema:

This method is also based on the plethysmometeric measurement of oedema produced by injection of 0.1 ml of 2% formalin into the sub-plantar area of hind paw of rat. This method is one of the most suitable test procedures to screen anti arthritic and anti-inflammatory agents as it closely resembles human arthritis.

Procedure:

The involvement of bradykinin and some other inflammatory mediators in formalin-induced edema was examined. Formalin was injected in rat paws at single dose, 0.1ml, the lower dose induced the development of immediate edema associated with accumulation of 125I-labelled albumin in the paws. These changes were suppressed by pre-treatment with and ethanolic extract of Hibiscus hirtus

The formalin was induced inflammation in female albino rats immediately after a single dose of formalin injected through the sub plantar route where the inflammation takes place along with redness. Anti-inflammatory effect was obtained to the rats which are pre-treated with Hibiscus hirtus. Ethanolic extract of Hibiscus hirtus was freshly suspended in distilled water and administered orally using oral feeding needle. The acclimatized animals were divided into 4 groups each consisting of 5 animals and were and are tabulated as below,

S. No.	Group	Treatment	Purpose
I	Normal	Normal saline (p.o)	Serves as normal
II	Inflammatory control	Formalin (0.1 ml, sub plantar)	Serves as disease control
III	Standard ibuprofen (75 mg/kg, p.o)	Formalin (0.1 ml, subplantar) +Ibuprofen tablets (75 mg/kg, p.o)	Serves as standard for Comparison of Anti-inflammatory effect.
IV	Test Hibiscus hirtus (500mg/kg)	Formalin (0.1 ml, subplantar) + Hibiscus hirtus (500 mg/kg, p.o)	Evaluation of anti- inflammatory effect of Hibiscus hirtus

Parameters like paw volume, swelling seeing visually, Redness were monitored for every 7 days of duration of experiment. Paw volume was measured using Plethysmometer. A mark was made above the tibia tarsal part of hind paw. The paw was dipped up to that mark and the displacement in the water level was noted using a scale. Paw volume of all groups were measured and the readings are noted.

Induction of inflammation:

Female albino rats weighing between 150-200g were used in the present study. On the 1st day they were injected with 0.1 ml of Formalin into the sub-plantar region of the right hind paw.

This is synthesized by the oxidation of methanol it is a general purpose chemical reagent and a solution of 35-40 percent of formaldehyde or methanol is formalin. After the single dose of injection the inflammation appears immediately.

Estimation of pharmacological parameters:

Inflammatory parameters:

I. Measurement of paw volume:

Principle: Formalin induced Inflammation is the most reliable model for Inflammation. In our study Inflammation was induced by administration of formalin 0.1 ml was administered through intra plantar route. Paw volume was measured by using Plethysmometer.

Procedure:

Paw volume of right paw were recorded after administration of Formalin injection. A water displacement method with a Plethysmometer was used for recording the paw volumes. The paws were marked with ink at the level of lateral malleolus and immersed in the water column of a Plethysmometer for measuring the paw volume. The paw volume was measured at every 2 hrs in the 1st day. The increased and decreased in paw volumes were noted. The difference of average values between treated animals and control groups were calculated. The percentage inhibition of the growth of edema was calculated by using the Following formula:

Vc- Vt

% Inhibition = ----------×100

Vc - changes in paw volume of control group and Vt - changes in paw volume of test group.

Individual data of formalin-induced paw volume (ml) in rats treated with vehicle (Control)

	Paw Volume in ml										Change in Paw Volume
Rats	0 hr		0.5 hr		1hr		2 hr		3 hr		0.5 hr	1 hr	2 hr	3 hr
	L	R	L	R	L	R	L	R	L	R
1	0.2	0.2	0.2	0.4	0.2	0.6	0.2	0.7	0.2	0.6	0.3	0.4	0.4	0.5
2	0.1	0.1	0.1	0.6	0.1	0.8	0.1	0.7	0.1	0.7	0.5	0.7	0.6	0.6
3	0.2	0.2	0.2	0.6	0.2	0.7	0.2	0.8	0.2	0.9	0.4	0.5	0.6	0.7
4	0.2	0.2	0.2	0.6	0.2	0.7	0.2	0.8	0.2	0.8	0.4	0.5	0.6	0.6
5	0.2	0.2	0.2	0.7	0.2	0.8	0.2	0.8	0.2	0.8	0.5	0.6	0.6	0.6
Mean±SD											0.42	0.52	0.58	0.62

Individual data of formalin-induced paw volume (ml) in rat treated with ibuprofen 75mg/kg (standard)

	Paw Volume in ml										Change in Paw Volume
Rats/	0 hr		0.5 hr		1hr		2 hr		3 hr		0.5hr	1 hr	2 hr	3 hr
	L	R	L	R	L	R	L	R	L	R
1	0.2	0.2	0.2	0.3	0.2	0.5	0.2	0.5	0.2	0.6	0.3	0.2	0.2	0.1
2	0.2	0.2	0.2	0.3	0.2	0.5	0.2	0.3	0.2	0.4	0.2	0.2	0.1	0.2
3	0.2	0.2	0.2	0.3	0.2	0.3	0.2	0.3	0.2	0.3	0.1	0.1	0.1	01
4	0.3	0.3	0.3	0.4	0.3	0.4	0.3	0.4	0.3	0.4	0.1	0.1	0.1	0.1
5	0.2	0.2	0.2	0.3	0.2	0.6	0.2	0.5	0.2	0.6	0.2	0.3	0.3	0.2
Mean±SD											0.22	0.21	0.19	0.18

Individual data of formalin-induced paw volume (ml) in rats treated with hibiscus hirtus 500 mg/kg (test)

	Paw Volume in ml										Change in Paw Volume
Rats/	0 hr		0.5 hr		1hr		2 hr		3 hr		0.5 hr	1 hr	2 hr	3 hr
	L	R	L	R	L	R	L	R	L	R
1	0.3	0.3		0.5		0.6		0.6		0.5	0.4	0.3	0.3	0.2
2	0.2	0.2		0.4		0.4		0.5		0.4	0.4	0.3	0.3	0.2
3	0.1	0.1		0.4		0.5		0.5		0.4	0.5	0.4	0.3	0.3
4	0.2	0.2		0.5		0.4		0.3		0.3	0.3	0.3	0.2	0.1
5	0.1	0.1		0.4		0.3		0.3		0.2	0.4	0.3	0.3	0.2
Mean±SD											0.41	0.38	0.29	0.27

DISCUSSION:

Inflammation represents a protective response designed to rid the body of the initial cause of cell injury and the consequences of the cell injury may occur due to trauma, genetic defects, physical and chemical agents, tissue necrosis, foreign bodies, immune reactions and infections. Inflammation is a local reactive change that involves the release of anti-bacterial agents from near by cells that defend the hoist against infection. The present study was carried out to evaluate the anti-inflammatory effect of ethanolic extract of Hibiscus hirtus against formalin induced inflammation in wistar rats. Administration of formalin caused a prominent increase in paws swelling which indicates the induction if inflammation in experimental rats. Standard drug ibuprofen (mg/kg) and ethanolic extract of Hibiscus hirtus in doses 500 mg/kg significantly decreased the paw volume the anti- inflammatory action of hibiscus hirtus may also be due to presence of flavanoids.

CONCLUSION:

By the present results, it was ratified that the leaf extract of Hibiscus hirtus L has antimicrobial activity. The different concentration of ethanolic extract by soxhlation process shows antimicrobial activity against the tested microorganisms Staphylococcus aureus and Escherichia coli. We tracked down that 15.0 mg/ml convergence of concentrate has shown most extreme zone of hindrance when compared to and 2.5; 5.0; 7.5; 10.0; 12.5mg/ml and 15mg/ml centralization of concentrate. It demonstrated that 15.0mg/ml grouping of concentrate had shown against bacterial action successfully when that 15.0mg/ml compared to other concentrates. This property is adequately found in loop strain. We tracked down that this concentrate additionally shows anti inflammatory property. The conclusion of the present study in comparison with control and standard clearly indicate that the selected plant species’ leaf extract showed anti-inflammatory activity on paw edema in chronic inflammation induced by formalin.

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

REFERENCES:

1. Ahameethunisa A.R., Hopper W. Antibacterial activity of Artemisia nilagirica leaf extracts against clinical and phytopathogenic bacteria BMC Comple. Alt. Med. 2010; 10:6.

2. Al Farug A., Ibrahim M., Mahmood A., Chowdhury M., Rashid R.B., Kuddus R., Rashid M.A. Pharmacological and phytochemical screenings of ethanol extract of Leeamacrophylla Roxb. Innov. Pharm Pharmacother. 2014;2: 321–327.

3. Anosike C.A., Obidoa O., Ezeanyika L.U. Membrane stabilization as a mechanism of the anti-inflammatory activity of methanol extract of garden egg (Solanum aethiopicum) DARU J. Pharmaceut. Sci. 2012; 20(1): 76.

4. Arokiyaraj S., Sripriya N., Bhagya R., Radhika B., Prameela L. Phytochemical screening, antibacterial and free radical scavenging effects of Artemisa nilagirica. Mimosa pudica and Clerodendrum siphonanthus – An in vitro study. Asian Pacific J. Tropic. Biomed. 2012;29: S601–S604.

5. Avula B., Wang Y.H., Smillie T.J., Mabusela W., Vincent L., Weitz F., Khan I.A. Quantitative determination of flavonoids by column high-performance liquid chromatography with mass spectrometry and ultraviolet absorption detection in Artemisia afar and comparative studies with various species of Artemisia plants. J. AOAC Int. 2009;92: 633–644.

6. Banerji A., Luthriya D.L., Kokate S.D. Toxicity of capillin, the insecticidal principle of Artemisia nilagirica Clarke. Indian J. Exp. Biol. 1990;28: 588–589.

7. Blaszyle M., Holley R.A. Interaction of monolaurin, eugenol and sodium citrate on growth of common meat spoilage and pathogenic organisms. Int. J. Food Microbial. 1998;39: 175–183.

8. Nathan C. Points of control in inflammation. Nature. 2002;420:846-52.

9. Gouwy M, Struyf S, Proost P, Van Damme J. Synergy in cytokine and chemokine network amplifies the inflammatory response. Cytokine Growth Factor Rev. 2005;16: 561-80.

10. Wellen KE, Hotamisligil GS. Inflammation, stress and diabetes. J. Clin. Invest. 2005;115: 1111-9.

11. Hanauer SB. Inflammatory bowel disease: epidemiology, pathogenesis and therapeutic opportunities. Inflamm. Bowel Dis. 2006; 12:3-9.

12. Whicher J, Chambers R. Mechanisms in chronic inflammation. Immunol. Today. 1984;5: 3-4.

13. Vinay K, Abdul KA, Nelson F, Aster JC. Robbins and Cotran’s Pathologic basis of diseases. 8th ed Philadelphia: Elsevier Publishers; 2010: 43-77.

14. Laurence LB, Bruce AC, Björn CK. Goodman and Gilman's The pharmacological basis of therapeutics.12th ed China: McGraw-Hill Publishers; 2011.

15. Eisenberg DM, Kessler RC and Foster C. 1993. Unconventional Medicine in the United States: Prevalence, Costs and Patterns of Use. N Eng J Med. 328:246-52.

16. Rakh MS and Chaudhari SR. Evaluation of CNS depressant activity of Momordicadioica Roxb wild fruit pulp. Int. J. Pharm. Pharm. Sci. 2010;2(4):124-6.

17. Perry JM. Medicinal plants of East and Southeast Asia: attributed properties and Uses. MIT Press, Cambridge, MA. 1980;334-60.

18. Mahadevan N, Shivali and Kamboj P, Hibiscus sabdariffa L.-An Overview. Natural Product Radiance. 2009;8(1):77-83.

19. Alviano DS, Alviano CS. Plant extracts: search for new alternatives to treat microbial diseases. Curr Pharm Biotechnol, Jan, 2009; 10(1): 106–121.

20. Malini M, Abirami G, Hemalatha V, Annadurai G. Antimicrobial activity of ethanolic and aqueous ex-tracts of medicinal plants against waste water pathogens. Int J Res Pure Appl Microbiol. 2013; 3(2): 40–42.

21. Zhang R, Eggleston K, Rotimi V, Zeckhauser RJ. Antibiotic resistance as a global threat: evidence from China, Kuwait and the United States. Global Health, Apr 7, 2006; 2: 6.

22. Dorman HJ, Deans SG. Antimicrobial agents from plants: antibacterial activity of plant volatile oils. J Appl Microbiol. 2000; 88(2): 308–316.

23. Talib WH, Mahasneh AM. Antimicrobial, cytotoxicity and phytochemical screening of Jordanian plants used in traditional medicine. Molecules. 2010; 15(3): 1811–1824.

24. Cruz MC, Santos PO, Barbosa Jr AM, de Mélo DL, Alviano CS, Antoniolli AR, Alviano DS, Trindade RC. Antifungal activity of Brazilian medicinal plants involved in popular treatment of mycoses. J Ethnopharmacol. 2007; 111(2): 409–412.

25. Antimicrobial activity of ethanolic extracts from some medicinal plants. Australian Journal of Basic and Applied Sciences. 2011; 5(11): 678-683.

26. Evaluation of antimicrobial activity of various medicinal plants extracts of Latur zone against pathogens. International Journal of Life Sciences and Resource. 2016; 2(5): 612-618.

27. Antimicrobial activity of ethanolic extracts from some medicinal plants. Australian Journal of Basic and Applied Sciences. 2011; 5(11): 678-683.

28. M. Hemalatha, B. Arirudran, A. Thenmozhi, U.S. Mahadeva Rao. Antimicrobial Effect of Separate Extract of Acetone, Ethyl Acetate, Methanol and Aqueous from Leaf of Milkweed (Calotropis gigantea L.). Asian J. Pharm. Res. 2011; 1(4): 102-107.

29. Preeti Tiwari. Antimicrobial Activity of Amritarishta Prepared by Traditional and Modern Methods. Asian J. Pharm. Res. 2014; 4(2): 114-116.

30. A. Sureka, C. Mary Sharmila, R. Chithra Devi, N. J. Muthu Kumar, V. Banumathi. Evaluation of In Vitro Anti-Inflammatory activity of Kusta Gaja Kesari - A Siddha Herbo Mineral Formulation against Albumin Protein Denaturation. Asian J. Pharm. Res. 2018; 8(3): 145-147.

31. Keerti Shrivastava, Sherendra Sahu, Skand K. Mishra, Kantishree De. In vitro Antimicrobial Activity and Phytochemical Screening of Syzygium aromaticum. Asian J. Res. Pharm. Sci. 2014; 4(1): 12-15.

32. Preeti Tiwari. Antimicrobial Activity of Ashwagandharishta Prepared by Traditional and Modern Methods. Asian J. Res. Pharm. Sci. 201; 4(3): 115-117.

33. Nidhi Rao, Sandhya Mittal, Sudhanshu, Ekta Menghani. Assessment of Phytochemical Screening, Antioxidant and Antibacterial Potential of the Methanolic Extract of Ricinus communis L. Asian J. Pharm. Tech. 2013; 3(1): 20-25.

34. Punasiya Rakesh, Pillai Sujit, Yadav Janeshwer. Isolation and identification of compounds from the leaves extract of Hibiscus syriacus L. Asian J. Pharm. Tech. 2015; 5(1): 8-12.

35. Merlin NJ, Parthasarathy V, Manavalan R, Devi P, Meera R. Phyto-Physico chemical evaluation, Anti-Inflammatory and Anti microbial activities of Aerial parts of Gmelina asiatica. Asian J. Research Chem. 2009; 2(1): 76-82.

36. Swapnali Mohite, Rutuja Shah, Naziya Patel. Antimicrobial Activity of Leaves extracts of Cassia tora. Res. J. Pharma. Dosage Forms and Tech. 2018; 10(1): 10-12.

37. R. Suresh, D. Benito Johnson, Appalaraju Gorle, Ashok Kumar Javvadi, Tamil Selvan A. The Wound healing, anti inflammatory and haemostatic effect of Eupatorium odoratum. Research J. Pharmacognosy and Phytochemistry. 2012; 4(2): 75-79.

Received on 13.03.2021 Modified on 14.11.2021

Asian J. Pharm. Res. 2022; 12(1):5-10.

DOI: 10.52711/2231-5691.2022.00002