Characterization of saponins from AcalyphaindicaL and Carica papayaL and its microbial activities

E.Geetha^* , K. Shailaja

Department of Botany, University College of Science, Osmania University, Hyderabad, Telangana-500007, India

Corresponding Author Email: maheshwaram84@gmail.com

DOI : http://dx.doi.org/ 10.5281/zenodo.8146135

Abstract

Keywords

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1. Introduction

Traditional medicinal plants are a therapeutic resourceused by the population of the African continent specificallyfor health care, which may also serve as startingmaterials for drugs[1],Iwu et al.[2] reported that infectious diseases account for one-half ofall deaths in tropical countries. As a result, people ofall continents have long applied poultices and imbibedinfusions of indigenous plants dating back to prehistoryfor health purposes [3]. It comprises therapeuticpractices in existence for hundreds of years beforethe development of modern scientific medicine and is stillin use today without any documented evidence ofadverse effects.

According to the World Health Organization [4] “a medicinal plant” is any plant that in one ormore of its organs contains substances that can be usedfor therapeutic purposes, or which are precursors forthe synthesis of useful drugs. This definition distinguishesplants whose therapeutic properties and constituents have been established scientifically and plants that are regarded as medicinal but have not yet been thoroughly investigated. The term “herbaldrug” determines the part/parts of a plant used for preparingmedicines (for example leaves, flowers, seeds,roots, barks, stems, etc)[5] (Anon, 2007a). Furthermore [6], defines medicinal plants as herbal preparationsproduced by subjecting plant materials to extraction,fractionation, purification, concentration, or other physicalor biological processes which may be produced forimmediate consumption or as a basis for herbal products.

Medicinal and aromatic plants contain biologically activechemical substances such as saponins, tannins, essentialoils, flavonoids, alkaloids, and other chemical compounds[7,8], which havecurative properties. These complex chemical substancesof different compositions are found as secondary plantmetabolites in one or more of these plants. Tyler [9], has reported that plants also contain certain other compoundsthat moderate the effects of the active ingredients.

Medicinal and aromatic plants have demonstrated their contribution to the treatment of diseases such as HIV/AIDS, malaria, diabetes, sickle-cell anemia, mental disorders[10, 11], and microbial infections[2,12]. According to the World Health Organisation [6], 80% of the population of the world uses medicinal plants for the treatment of diseases and in African countries, thisrate is much higher.

Iwu et al.,[2], reported that the primary benefits ofusing plant derived medicines are that they are relativelysafer than synthetic alternatives, offering profound therapeuticbenefits and more affordable treatment. The use of medicinal plants in developing countries as a normativebasis for the maintenance of good health has beenwidely observed[13]. Furthermore, theincreasing reliance on the use of medicinal plants in industrialized societies has been traced to the extractionand development of several drugs and chemotherapeuticsfrom these plants as well as from traditionallyused rural remedies [14].

Moreover, in these societies, herbal remedies have become more popular in the treatment of minor ailments and also on accountt of the increasing costs of personal health maintenance. The survey conducted by the WHO Rollback in malariaprogram in1998, showed that in Ghana, Mali, Nigeriaand Zambia, more than 60% of the children with high fever were treated at home with herbal medicines[15].Saponins are glycosides of triterpenes and steroids characterized by their bitter or astringent taste,foaming property, hemolytic effect on red blood cellsand cholesterol binding properties[16].Saponins have been shown to possess both beneficial(lowering cholesterol) and deleterious (cytotoxic andpermeabilization of intestinal epithelium) properties andto exhibit structure dependent biological activity. In medicine,it is used to some extent as an expectorant and anemulsifying agent[7].

In recent years, research onmedicinal plants has drawn a lot of attention globally for itsversatile applications. Medicinal plants are the wealthiest bioresourcesof drugs of the traditional system of medicines, modernmedicines, food supplements, nutraceuticals, folk medicines,pharmaceutical intermediates, and chemical activities forsynthetic drugs. Scientific experiments on the antimicrobialproperties of plants and their constituents have beendocumented in the late 19th century[17]. Large body of evidencehas been gathered to demonstrate the promising potential ofmedicinal plants used in various traditional, complementaryand alternate systems of treatment of human diseases.

Traditionally used medicinal plants produce differentcompounds of identified therapeutic properties. Thisrevitalization of interest in plant derived drugs is mainly due tothe current extensive belief that, “green medicine is safe”.Many works have been done which aim at knowing thedifferent antimicrobial and phytochemical constituents ofmedicinal plants and using them for the treatment ofmicrobial infections (both topical and systemicapplications) as possible alternatives to chemicallysynthesized drugs to which many infectious microorganismshave become resistant. During the last ten years the paceof development of new antimicrobial drugs has slowedown while the frequency of resistance (especiallymultiple) has increased astronomically [18].

For a long period of time, plants have been precious sources ofnatural remedy for maintaining human health, especially in thelast decade, with more intensive studies for natural therapies.Plants have provided a source of inspiration for novel drugcompounds, as plant derived medicines have made immensecontributions to human health and wellbeing. Plants containnumerous biologically active compounds, many of which haveantimicrobial properties[3].Numerous studies have beenperformed with the extracts of various plants, screeningantimicrobial activity as well as for the discovery of newantimicrobial compounds [19,20-21]. Acalyphaindica(family: Euphobiaceae) is a small annual herb grows up to 60cm along the roadsideshaving medicinal properties [22]. It is distributed in Asian countries such as Sri Lanka, India,Pakistan,Africa and South America [23,24]. It iscommonly known as Indian Copper leaf (T- Kuppaimeni, S- Kuppameniya)[25].Leaves aree little triangular and ovate. Leaf stalks are longer than the 3-5 cm long blades. Flowers are borne on erectof axillary spikes which are stalkless. Male flowers are minute where the female flowers are scattered alongthe inflorescence axis[26,27&28].

The plant extract is used fortreating pneumonia, jaundice, piles, asthma, rheumatism, bedsores, wounds, skin infections and eczema. Has been stated to have wound healing activity, snake venom neutralizing properties, antibacterial activity,and antiurolithiatic activity [29,30,31,32,33&34].The whole plant is diuretic, expectorant, emetic, anthelmintic[35].

The previous findings of Suresh et al, revealed that A. indicashowed considerable antibacterial activityagainstS. aureus and E. coli. The aqueous extracts of A. indicashowed inhibition against E.coli andalcoholic extract show inhibition towards Staphylococcus aureus and Salmonella typhi[36].

The phytochemical content of dried Anting-anting plants has the benefitof treating therapeutic problems, one of which is anthelmintic[37], and also mentioned in the research of Syahiranetet al.,[38], that the leaves of Anting-antings- Anting-antings can be consumed orally as anthelmintic and are moreoften consumed because the amount is abundant and easily separated from plant parts rather than stems rootsand flowers.

According to Nahrstedtet al.,[39], some chemical constituents have been isolated from Acalyphaindica,including kaempferol glycosides, Mauritian, clitorine, nicotiflorin and biorobin, tannins, pyranoquinolinoneflindersin alkaloids. Its ethanol extract is known to contain polyphenols, flavonoids, monoterpenes,sesquiterpenes, steroids, triterpenoids, and quinones[40], also flavonoids, tannins, saponins and glycosides inethanol extract Anting-antings and alkaloids in petroleum ether extract, acetone and methanol [41].

Carica papaya is commonly known as papaya and belongs to family Caricaceae. Active constituent papain induces teratogenic effects and antiovulatory activity in rats. papaya (Caricapapaya) is a special fruit crop having high nutritionalvalue and potential for both fresh and processed marketuses. Papaya exceeds apple, peach, and grapes invitamins, minerals, amino acid, and food energy values asa fruit source. The green fruit is evident to comprise ofprotein, fat, carbohydrate, fiber, ash, Ca, P, Fe, Na, K,beta carotene equivalent, thiamine, riboflavin, niacin,ascorbic acid, and Vitamin E. It has also been found tocontainsinigrin, the enzyme myrosin, and carpasemine[42](Amosu AM etal., 2014).

Papaya has many biologically active compoundsincludingchymopapain and papain, which helps indigestion [43]. Papaya root, seed and leaf extract possess highly anti-tumor and pesticidal properties [44].The green papaya is used for the cure of ulcer andimpotence [45].The unripe fruits have uses asantiseptic, in cleansing the intestines of bacteria andenabling the intestine to absorb vitamins and minerals, especially vitamin B12 [46]. Secondary metabolitessuch as alkaloids, flavonoids, saponins and tannins ingreen papaya are serving as a potent free radicalscavengers and are antimicrobial in action [47]. The leaves, fruits, and latex obtained from the papaya plant havemedicinal uses. The fruit has been found to containcertain immune-stimulating and antioxidant agents.

Antimicrobials from natural products have inspired antibioticdiscovery and have been used for microbial control. These includeplant extracts, small antimicrobial peptides, essential oils, bacteriocinsand various groups of compounds[48]. There areseveral reports showing the antimicrobial activity of free compoundsisolated from natural sources. Natural products such as tannins aregood substances to control microbial growth by interact with bacterialproteins and precipitating them[49].Chalconederivative from Croton anisodontus Mull. Arg. acts as acompetitive inhibitor MepA efflux pump and potentiatesciprofloxacin’s action against multidrug-resistant Staphylococcusaureus. Betulinic acid was reported to have stringinhibition against Candida albicans[50].NimbolidefromAzadiractaindica A. Jusspossesses significant bactericidal activity against Helicobacter pylori by killing free-living bacteriaand cells within biofilm [51].

The objective of this study is to extract and estimate the saponins from Acalyphaindica and Carica papaya, also to assessthesaponins antimicrobial activity and antifungal activity.

• Materials and Methods

2.1 Collection of plant materials. The plants used in this study are Acalyphaindica, Carica papaya. Thedifferent plants were simultaneously collectedfrom forests, open fields,abandoned and cultivated farms in the Hyderabadarea of TelanganaState,India. The plants were identified in their fresh states and collected in sterile polythene zip lock covers to transport the laboratory for analysis.

Figure 1: Pictures of medicinal plants Acalyptaindica and Carica papaya.

2.2 Preparation of plant extracts

The collected plant parts were separated from undesirablematerials and were washed with distilled water. They weresun-dried for one week and ground into fine powder with thehelp of a grinder. The powder was stored in an airtightcontainer and kept in a cool, dark, and dry place until analysiscommenced. The bioactive components were extractedaccording to the methods of Pandey et al.,[52], withslight modification. The powered materials were dissolved in80% methanol (1:10); 1 g sample should be dissolved in 10 mlof solvent Pandey et al. [52]., Mixtures were kept in sterilized beakerswrapped with aluminium foil to avoid evaporation andexposure to light. The beakers were then kept in dark for 3 daysat room temperature accompanying occasional shaking andstirring. After 3 days, mixtures were filtered through Whatmanno. 1 filter paper. The filtrates obtained were concentratedusing a water bath.

2.3 Test for saponins

The extract (50mg) was diluted with 20 ml ofdistilled water, and it was agitated in a graduatedcylinder for 15 minutes. The formation of 1 cm layerof foam showed the presence of saponins.

2.4 Sample Preparation for HPLC analysis

Two grams of each plant sample powder was extracted in soxhlet apparatus with 150 mL of 70% ethanol for 7 hrs at 45°C. The extraction procedure was executed in triplicate for each plant sample. It was transferred into a flat bottom flask and concentrated with a rotary evaporator. The concentrate was then dissolved in 10mL of HPLC-grade methanol.

2.5 HPLC analysis

30μl of these extracts were passed through. 45μm syringe filter and that filtrate was used for HPLC analysis. The HPLC system (Shimadzu lab chromo 2010 HT HPLC, UV detector) was used. The software package used for analyzingresults was Shimadzu lab chromo HPLC control and auto-sampling. Chromatographic analysis was carried out using a c-18 column at 35 ⁰C temperature. Prior to analysis, the column was equilibrated with the corresponding. Running conditions included: injection volume 15μl; mobile phase: acetonitrile: water (40:60) running time 25 min., flow rate 1 ml/min; and detection at 203nm. The separation of filtered methanolic plant extracts, as well as a mixture of authentic standard samples of saponins, was done. The peak area of standards and samples was calculated to determinate concentration.

2.6 Gas Chromatography-Mass Spectroscopy Analysis

To determine the chemical composition of the samples, a Trace GC Ultra-ISQ mass spectrometer was used. The analysis was performed using a TG-5MS column with dimensions of 30m x 0.25mm x 0.25μm film thickness. The temperature of the oven was programmed to increase from 60°C to 150°C at a rate of 5°C/min, then increased to 280°C at a rate of 10°C/min, holding for 2 minutes at 150°C. Helium gas was used as a carrier at a constant flow rate of 1 ml/min, and the inlet and transfer line temperatures were kept at 250°C. A solvent delay of 3 minutes was used, and 1μl of diluted samples were automatically injected in split mode using Auto sample AS3000. Mass spectra were collected in full scan mode at 70 eV ionization voltages over the range of m/z 40-650[53].

2.7 Anti-bacterial activity

Nutrient agar plates are prepared and pathogenic bacterial lab cultures that are Pseudomonas fluorescens: MTCC 9768, E.coli: MTCC 424, Staphylococcus aureus: MTCC 96, Klebsiella pneumoniae: MTCC 272, and Bacillus subtillis: MTCC 3053 were spread in the agar plates. Then activated samples are placed using the paper dip method and incubated for 24hrs. After 24hrs of incubation clear zone of bacterial inhibition was observed around the sample, which was measured, and the measurement of the zone was recorded. Samples which were showing the antibacterial activity are used for further study [54].

2.8 Anti-fungal activity

The anti-fungal activity was detected by the dual culture method. FussariumoxysporumNCIM1008 and Phytophtherainfestans MTCC 8707 were grown on PDA medium. An agar block (five mm dia) was cut from an actively growing (96 h old) fungal culture and placed on the surface of fresh agar medium at the centre of petri plate. A paper disc dipped in the respective sample was placed onto the plate at different locations of a 90 mm dia Petri plate and plates were incubated at 30 ±2 °C. Inhibition zone between two cultures was measured after 5 days of incubation.

Where,

I = Inhibition % of mycelia growth (growth reduction over control)

C = Radial growth of fungus in the control plate (mm)

T = Radial growth of fungus on the plate inoculated with bacteria (mm)

• Results and Discussion:
  • GCMSanalysis of Acalifaindica

Pea	R.Time	Area	Area%	Height	A/H	Basem/z	Name
1	1.073	169501472	6.07	74790793	2.27	48.70	Propanoicacid, 2-chloro-
2	1.172	103581708	3.71	20397489	5.08	59.70	Pentaborane(11)
3	1.215	1331644146	47.65	0	0.00	84.75	Thiophene,2,3-dihydro-
4	1.360	21079744	0.75	27899534	0.76	57.10	Propane,1-(ethenyloxy)-2-methyl-
5	1.436	97250416	3.48	73374675	1.33	84.95	Chloroform
6	1.470	11189036	0.40	13487273	0.83	56.05	Cyclopentane,methyl-
7	1.590	94439361	3.38	55119576	1.71	73.05	Propane,2,2-dimethoxy-
8	2.172	4755265	0.17	1588915	2.99	44.00	Pentanal
9	2.407	433310132	15.51	112846630	3.84	93.80	Pyridine-4-aldehyd,N-ethoxycarbonylhydrazon
10	3.501	27131282	0.97	3847012	7.05	43.05	2-Pentanone,4-hydroxy-4-methyl-
11	21.116	14421802	0.52	4010556	3.60	41.05	n-Hexadecanoicacid
12	29.340	6868000	0.25	1722118	3.99	207.00	(9-Oxo-9,10-dihydroacridin-4-yl)aceticacid
13	29.775	9590520	0.34	1930680	4.97	207.00	Benzene,2-[(tert-butyldimethylsilyl)oxy]-1-isopropyl-4-methyl-
14	30.035	4651958	0.17	2228614	2.09	207.00	Propiophenone,2′-(trimethylsiloxy)-
15	30.162	4979300	0.18	2357177	2.11	133.05	n-Propylamine,N-acetyl-3-[2-acetyl-3,4,5-trimethoxyphenyl]-
16	30.490	9390819	0.34	2325397	4.04	73.05	Silane,trimethyl[(3,7,11-trimethyl-2,6,10-dodecatrienyl)oxy]-
17	30.645	8429432	0.30	2387583	3.53	73.10	Silane,trimethyl[(3,7,11-trimethyl-2,6,10-dodecatrienyl)oxy]-
18	30.825	8002646	0.29	2042767	3.92	253.05	1-Trimethylsilyl-4-(1-methyl-1-silacyclobutyl)benzene
19	30.945	9555031	0.34	2281982	4.19	281.05	1H-Indole-2,3-dione,1-(tert-butyldimethylsilyl)-5-chloro-,3-(O-ethyloxime)
20	31.185	8659981	0.31	2524044	3.43	96.10	Pseduosarsasapogenin-5,20-dienmethylether
21	31.330	4551350	0.16	1773465	2.57	73.05	Silane,[1,4-dioxane-2,3-diylbis(oxy)]bis[trimethyl-,cis-
22	31.485	10450667	0.37	3832685	2.73	281.05	1,3,5,7-Tetraethyl-1,7-dibutoxytetrasiloxane
23	31.600	5559430	0.20	2535191	2.19	281.05	2-Methyl-7-nonadecene
24	31.655	10687457	0.38	3320104	3.22	281.10	Haloxazolam
25	31.696	4426073	0.16	3637583	1.22	209.00	Silane,(9,19-cyclo-9.beta.-lanost-24-en-3.beta.-yloxy)trimethyl-
26	31.885	23580593	0.84	5850074	4.03	67.05	2-(4-Hydroxybutyl)cyclohexanol
27	32.050	37206432	1.33	11388039	3.27	281.10	E-10,13,13-Trimethyl-11-tetradecen-1-olacetate
28	32.225	19344260	0.69	6792112	2.85	69.10	2,10-Dodecadien-1-ol,3,7,11-trimethyl-,(Z)-
29	32.880	5327769	0.19	2568993	2.07	208.00	Pentasiloxane,1,1,3,3,5,5,7,7,9,9-decamethyl-

Figure 2: Chromatogram of Gcms analysis AcalifaIndica.

Table 1: Phytochemicals identified in methanolic plant extract of Acalifaindica by GC-MS.

Interpretation on mass spectrum GC-MS wasconducted using the database of National Institute

Standard and Technology (NIST) having more than62,000 patterns. The spectrum of the unknowncomponent was compared with the spectrum of theknown components stored in the NIST library. Thename, molecular weight and structure of thecomponents of the test materials were ascertained inTable1 figure 2.MethanolicAcalifaindicaplant extractwas subjectedto GC-MS study for identification of medicinalproperties, according to the results, the total 29 Phytocomponents are screened, and most of themedicinal properties areThiophene,2,3-dihydro (47%), Pyridine-4-aldehyd,N-ethoxycarbonylhydrazon (15.5%), Propanoicacid, 2-chloro (6.07%), Pentaborane(11) (3.71%), Chloroform (3.48%), Propane,2,2-dimethoxy- (3.38%), E-10,13,13-Trimethyl-11-tetradecen-1-olacetate (1.33%) relative abundance respectively.

• GCMS  ANALYSIS OF  CARICA PAPAYA

The analysis of extracts using GC MS technique also proved that there are effective compounds in papaya plants according to the solvent used as shown in Table (2) and Figure (3). Table (2) and figure (3) showed the presence of 12 phytocomponentsscreened in the methanolic papaya leaf extract. Through comparative examination, the main components present in the papaya plant extract of the local variety in terms of their relative abundance were 2-Trifluoroacetoxydodecane (38.24%), 1,3,5-Triazine-2,4-diamine, N,N’-bis(1-methylethyl)-6-(methylsulfonyl)-(12.20%), ,5-Heptadien-3-yne (5.71%), 1-Propanethiol (3.73%), Methylene Chloride (3.53%), Chloroform (2.80%) and Hexane (1.32%)relative abundance respectively. 

Figure3 :Chromatogram ofPhytocomponents in the papaya plant extract.

Table 2:Phytochemicals identified in methanolicplantextract of Carica papaya by GC-MS

Peak	R.Time	Area	Area%	Height	A/H	Base m/z	Name
1	0.021	26644071	0.81	48981498	0.54	44.05	2-Heptanamine, 5-methyl-
2	0.133	1254601570	38.24	125255983	10.02	41.10	2-Trifluoroacetoxydodecane
3	1.084	400227497	12.20	102705694	3.90	62.85	1,3,5-Triazine-2,4-diamine, N,N’-bis(1-methylethyl)-6-(methylsulfonyl)-
4	1.166	122228164	3.73	63195535	1.93	46.10	1-Propanethiol
5	1.210	115777816	3.53	81287753	1.42	83.95	Methylene Chloride
6	1.270	22829949	0.70	15520228	1.47	43.05	Pentane, 2-methyl-
7	1.308	16905316	0.52	12578035	1.34	57.15	Pentane, 3-methyl-
8	1.355	43959187	1.34	34446590	1.28	57.15	Hexane
9	1.433	91906495	2.80	65324924	1.41	82.95	Chloroform
10	1.465	13117068	0.40	13106320	1.00	56.10	Cyclopentane, methyl-
11	1.590	10929492	0.33	3578291	3.05	73.10	Propane, 2,2-dimethoxy-
12	2.394	187335547	5.71	105599693	1.77	93.00	1,5-Heptadien-3-yne

3.3 Standard saponin HPLC

Figure 4: Standard saponin chromatogram by using HPLC.

HPLC was used to estimate saponin in plant extracts. NIST traceable standard saponin was procured from the local market. HPLC was run up to 35 minutes and the saponin was eluted at 21.7 minutes. The standard saponin chromatogram was compared with two plant extracts to know the saponin purity.

3.4 HPLC of Acalifaindica

Figure 5: Acalifaindica plant extract chromatogram by using HPLC.

HPLC analysis of acalifaindica plant extract chromatogram, the run time was matched with standard saponin. The purity calculation was revealed that the aclifaindica plant extract is having 43.12% of saponin concentration.

3.5 HPLC analysis of Carica papaya

Figure 6: Carica papaya plant extract chromatogram by using HPLC.

HPLC analysis of Carica papaya plant extract chromatogram, the run time was matched with standard saponin. The purity calculation revealed that the Carica papaya plant extract is having 23.67% of saponin concentration.

3.6 Antibacterial Activities:

Pseudomonas fluorescens: MTCC 9768

1

2

3

4

5

E.coli: MTCC 424

1

2

3

4

5

Staphylococcus aureus: MTCC 96

1

2

3

4

5

Bacillus subtillis: MTCC 3053053

1

2

3

4

5

Here 1: Standard saponin 2: Plant extract of AI with methanol, 3: Plant extract of CP with methanol, 4: Plant extract of AI with water, 5: Plant extract of CP with water.

Figure 7:Antibacterial activity againstbacterial strains

Table 3: Antibacterial activity of plant extracts

	Standard saponin (Zone in cm)	Plant extract of AI with methanol (Zone in cm)	Plant extract of CP with methanol (Zone in cm)
Pseudomonas fluorescens: MTCC 9768	0.8± (0.01)	1.2 ± (0.02)	1.3 ± (0.02)
E.coli: MTCC 424	0.9 ± (0.01)	1.3 ± (0.02)	1.2 ± (0.02)
Staphylococcus aureus: MTCC 96	0.5 ± (0.01)	1.1 ± (0.02)	1.3 ± (0.02)
Bacillus subtillis: MTCC 3053	0.7 ± (0.02)	1.2 ± (0.02)	1.2 ± (0.02)

Antibacterial potentialsof Acalifaindica (AI) and Carica papaya (CP) are appraised using minimum inhibitory concentration (MIC). in vivo,antibacterial potential of AI was assessedagainst Pseudomonas fluorescens and found the MIC as 1.2 cm whereas CP showed the MIC as 1.3cm, the least activity was observed with the standard as 0.8cm.The E.coli bacteria showed significant activities due to the saponins analysed, the highest activity was observed in AI (1.3cm) followed by CP (1.2cm) the least concentration was observed with standard (0.9cm) (CP) displayedsignificant MIC of 1.3cm against clinical isolates of staphylococcus aureusbacteria, whereas AI displayed as 1.1cm zone and lowest was observed in 0.5cmBacillus subtilliswas also checked for its antibacterial activity and the results revealed that the maximum antibacterial activity was observed with both plant extracts (1.2cm), least was observed in standard as 0.7cm. 

3.7 Antifungal Activities

Fussariumoxysporum NCIM1008

1

2

3

4

5

Phytophtherainfestans MTCC 8707

1

2

3

4

5

Here 1: Standard saponin 2: Plant extract of AI with methanol, 3: Plant extract of CP with methanol, 4: Plant extract of AI with water, 5: Plant extract of CP with water.

Figure 8:Antifungal activity against FussariumoxysporumandPhytophtherainfestans.

Table 4: Antifungal activity of plant extracts

	Standard saponin (Inhibition %)	Plant extract of AI with methanol (Inhibition %)	Plant extract of CP with methanol (Inhibition %)
FussariumoxysporumNCIM1008	40± (1.8)	46 ± (1.2)	55 ± (1.8)
Phytophtherainfestans MTCC 8707	50 ± (1.5)	45 ± (1.6)	50± (1.4)

The plant extractsisolated from Acalifaindica (AI) and Carica papaya (CP) exhibit strong antifungal activities. antifungal activities of Acalifaindica against Fussariumoxysporumwas found as 46%, whereas Carica papaya was shown 55% of inhibition lowest inhibition was observed with standard saponin (40%). The fungal strain Phytophtherainfestanswas inhibited 50% by Carica papaya plant extract and standard saponin activity, and the lowest inhibition was observed with Acalifaindica (45%).

Conclusions

The study’s findings indicated that Acalifaindica and Carica papayahave saponins as bioactive constituents, which contribute to their antibacterial properties and antifungal properties. The saponin quantities were identified and conformed with HPLC and GCMS analysis. The antibacterial activity could inhibit Pseudomonas fluorescens, E.coli, Staphylococcus aureusandBacillus subtillisandantifungal activity could inhibit Fussariumoxysporum and Phytophtherainfestanswell with methanolicAcalifaindica and Carica papaya extract.

References

• Sofowora AE (1993). Medicinal Plants and Traditional Medicines in Africa.. 2nd edition. Spectrum Books, Ibadan, Nigeria. p. 289.
• Iwu MM, Duncan AR, Okunji CO (1999). New Antimicrobials of plant origin. In Janick, J. (ed) Perspectives in New crops and New uses. ASHS Press, Alexandria, V.A. pp 457-462.
• Cowan, M. M. (1999). Plant Products as Antimicrobial Agents. Clin. Miicrobiol. Rev. 12(4) : 564-582.
• WHO (1977). Resolution –Promotion and Development of Training and Research in Traditional Medicine. WHO document No: 30-49.
• Anonymous (2007a). Medicinal, Culinary and Aromatic Plants in the Near East-Medic… http://www.faO.Org/dorep/x5402e16.htm.Antibacterial activity of Acalyphaindica L. Eur Rev Med PharmacolSci, 12: 299-302.
• WHO (2001). Legal Status of Traditional Medicine and Complementary/ Alternative medicine: A world wide review. WHO Publishing 1.
• Harborne JB (1973). Phytochemical Methods: A Guide to Modern Techniques of plant Analysis. Chapman and Hall Ltd, London. p. 279.
• Sofowora A. Medicinal Plants and Traditional Medicinal in Africa. 2nd Ed. Sunshine House, Ibadan, Nigeria: Spectrum Books Ltd; 1993. Screening Plants for Bioactive Agents; pp. 134–156.
• Tyler VE (1999). Pyhtomedicines: back to the future. J. Nat. Prod., 62: 1589-1592.
• Elujoba AA, Odeleye OM, Ogunyemi CM (2005). Traditional Medical Development for Medical and Dental Primary Health Care Delivery System in Africa. Afr. J.Trad. Compl. Alter. Med. 2(1):46-61.
• Okigbo RN, Mmeka EC (2006). An Appraisal of phytomedicine in Africa. KMITL Sci. Technol. J. 6 (2): 83-93.
• Okigbo RN, Mbajiuka, Njoku CO (2005). Antimicrobial potentitals of (Uda) Xylopiaaethiopica and Ocimumgratissimum on some pathogens of man. Int. J.Mol. Med. Adv. Sci. 1(4):392-397.
• UNESCO (1996). Culture and Health, Orientation Texts World Decade for cultural Development 1988-1997. Document CLT/ DEC /PRO 1996, Paris, France. p.129.
• UNESCO (1998). FIT/ 504-RAF-48. Terminal Report: Promotion of Ethnobotany and the sustainable use of plant Resources in Africa.. Paris, 1998. p. 60.
• WHO (2004). Guideline on Developing Consumer Information on Proper Use of Traditional, Complementary and Alternative Medicine. Geneva.
• OkwuDE(2005). Phytochemicals, Vitamins and Mineral contents of two Nigeria Medicinal plants. Int. J.Mol. Med. Adv. Sci. 1(4): 375-381.
• Chopra RN, Nayer SL, Chopra IC. (1992). Glossary of Indian Medicinal Plants. Council of Scientific and Industrial Research, New Delhi, 1992, 7-246.
• Hugo WB, Russel AD (1984). Pharmaceutical Microbiology. Edn 3rd, Blackwell Scientific Publications, 1984, 179-200.
• Chah KF, Eze CA, Emuelosi CE, Esimone CO. Antibacterial wound healing properties of methanolic extracts of some Nigerian medicinal plants. Journal of Ethnopharmacology 2006; 104:164-167.
• Nair R, Chanda S. In vitro antimicrobial activity of Psidiumguajava L. leaf extracts against clinically important pathogenic microbial strains. Brazilian Journal of Microbiology 2007; 38(3):452-458.
• Parekh, J. and Chanda, S., (2007). In vitro antimicrobial activity and phytochemical analysis of some Indian medicinal plants. Turkish journal of biology, 31(1), pp.53-58.
• Burkill, H.M. (1985). The useful plants of West Tropical Africa. Royal Botanic Gardens, Kew, UK 2:246.
• Ramachandran J. (2008). Herbs of Siddha Medicine. The First 3D Book OnHerbsWound healing effects of Heliotropiumindicum, Plumbagozeylanicum and Acalyphaindica in rats. J Ethnopharmacol. 79: 249–251.
• Parveen, B.  Upadhyay, S.R. Ashwani, K. (2007).  Traditional uses of  medicinal plants  among the rural communities of Churu district in the Thar Desert, India. Journal of Ethnopharmacol 2 (113):387-9.
• Kirtikar, K.R. Basu, B.D. (1975). Indian Medical Plants. Volume II, 2nd Edition. New Delhi, Jayyed Press 1975; 30-45.
• Kirtikar, K.R. Basu, B.D. (1994). Indian Medicinal Plants, Volume I, 2nd edition. Dehradun pp 314-317.
• Prajapati, N.D. Purohit, S.S. Sharma, A. K. Kumar, T.A. (2003). Handbook of Medicinal Plants,
• Nadkarni, A.K. (1995). Indian Materia Medica, Popular Prakashan (Pvt) Ltd, Bombay, 8-9.
• Reddy, J. S., Rao, P. R., & Reddy, M. S. (2002). Wound healing effects of Heliotropiumindicum, Plumbagozeylanicum and Acalyphaindica in rats. Journal of ethnopharmacology, 79(2), 249-251.
• Govindarajan, M. Jebanesan, A. Reetha, D. Amsath, R. Pushpanathan, T. Samidurai, K. (2008).Antibacterial activity of Acalyphaindica L.European Review for Medical and Pharmacological Sciences.2008; 12: 299-302.
• Sathya, M. Kokilavani, R. Teepa, K.S. Balakrishnan, A. (2011). Bio potency of Acalyphaindica Linn on membrane bound ATPases and marker enzymes urolithic rats. AncSci Life 2011; 31(1): 3-9.
• Jain, S.K. (1987). A Manual of Ethnobotany Scientific Publishers, Jodhpur
• Jain, S.K. (1996). Medicinal Plants Pub. National Book Trust India, New Delhi.
• Sumathi, G. and Puspha, B. (2005). Evaluation of antibacterial activity of some India Medicinal plant. Asian Journal of Microbiology, Biotechnology and Environmental Sci. 9:201-205.
• . Indian medicinal plants: a compendium of 500 species Orient Longman.Publication, Madras, India. 134.
• Divya, N. Thenmozhi, S. Sureshkumar, B.T. and Selvan, M. (2014). Antibacterial Activity Of Medicinal Plant Against Wound Infected Pathogens. International Journal of Pharmaceutical Sciences and Research. 5(11): 4942-4947.
• Mohan C, Dinakar S, Anand T, Elayaraja R, SathiyaPriya B. Phytochemical(2012). GC-MS Analysis and Antibacterial Activity of A Medicinal Plant Acalyphaindica. International Journal Pharmaceutical Technology Research. 2012;1050-1054.
• Syahiran N, Zulkifli R, Saidin S, Muhamad II (2016). A Review of Acalyphaindica L (Euphorbbiaceae) as Tradition Medical Plant and Its Therapeutic Potential. Journal of Ethnopharmacology.
• Nahrestedt A, Hungeling M, Petereit F. (2006). Flavonoids from Acalyphaindica. Institute of Pharmaceutical Biology and Phytochemistry;484-486.
• Febriyanti M, Supriyatna, Abdullah R. Chemical content and cytotoxic activity of extracts and fraction of herbal anting-anting (Acalyphaindica Linn) against breast cancer cells MCF-7 (Kandungan Kimia danAktivotasSitotoksikEkstrakdanFraksiHerba Anting-Anting (Acalyphaindica Linn) terhadapSelKankerPayudara MCF-7). JurnalDarmasi Indonesia. 2014;19-26.
• Vijayarekha P, Sangottaiyan N, Noorjahan A, Ambiga S. Antibacterial Activity of Acalyphaindica Linn. International Journal of Current Microbiology and Applied Science. 2015;1133-1138.
• , Degun AM, Esan EB. 2014. Microbial contaminants in the herbal preparation from green pawpaw fruits. Biomed. Res. 25 (4): 169-178.
• , Lekshmi VD, Prakash SGV. 2011. Bioprocessing of banana peel for amylase production by Sp. Penicillium. Asian J. Biol. Sci. 2:257-264.
• , Ratanyake S, Rupprecht JK, Potter WM. 1992. Evaluation of various parts of paw-paw tree, Asimmatritoba as commercial source of the pestisideannonsceaeacetogenins. J. Econ. Entomol. 85: 2353-2356.
• . 1994. Immense help from nature’s workshop, 1st ed. Elikaf Health Services LTD. Ikeja, Lagos. AGROBIOS, (India) Jodhpur. 3.
• EhiowemwenguanG,Emoghene AO, Inetianbor JE. 2014. Antibacterial and phytochemical analysis of Banana fruit peel. IOSR J Pharm. 4 (8): 18-25.
• Ifesan, O. T.  Fashakin, B., F.  Ebosele, J., Oyerinde, A.S (2013). Antioxidant and Antimicrobial Properties of Selected Plant Leaves. European Journal of Medicinal Plants, 3(3), 465–473. https://doi.org/10.9734/EJMP/2013/3383.
• Stan, D., Enciu, A. M., Mateescu, A. L., Ion, A. C., Brezeanu, A. C., Stan, D., et al. (2021). Natural Compounds with Antimicrobial and Antiviral Effect and Nanocarriers Used for Their Transportation. Front. Pharmacol. 12, 723233. doi:10.3389/fphar.2021.723233.
• Puljula, E., Walton, G., Woodward, M. J., and Karonen, M. (2020). Antimicrobial Activities of Ellagitannins against Clostridiales Perfringens, Escherichia coli, Lactobacillus Plantarum and Staphylococcus aureus. Molecules 25, 3714. doi:10. 3390/molecules25163714.
• Lashin, I. I., Aref, M. S., Abd El-Kader, E. M., Heussin, E. A., and Ewais, E. E.-D. A. (2021). Antimicrobial and In-Vitro Cytotoxic Efficacy of Betulinic Acid from the Callus, Regeneration, and Mother Plant of DilleniaIndica L. Egypt. J. Bot. 62, 213–226. doi:10.21608/ejbo.2021.81121.170610.21608/ejbo.2021.81121.1706
• Wylie, M. R., Windham, I. H., Blum, F. C., Wu, H., and Merrell, D. S. (2022). In Vitro antibacterial Activity of Nimbolide against Helicobacter pylori. J. Ethnopharmacol. 285, 114828. doi:10.1016/j.jep.2021.114828.
• Pandey A, Shweta. Antifungal properties of Psidiumguajava leaves and fruits against various pathogens. Journal of Pharmaceutical and Biomedical Sciences 2011; 13(16):1-6.
• Ayaz, M., Junaid, M., Ullah, F., Subhan, F., Sadiq, A., Ali, G., …& Ahmad, S. (2017). Anti-Alzheimer’s studies on β-sitosterol isolated from Polygonumhydropiper L. Frontiers in pharmacology, 8, 697.
• Mohamed DS, Abd El-Baky RM, Sandle T, Mandour SA, Ahmed EF. Antimicrobial Activity of Silver-Treated Bacteria against other Multi-Drug Resistant Pathogens in Their Environment. Antibiotics. 2020; 9(4):181.