Keywords

Antibacterial; peels; antioxidant

Introduction

Over fifty percent of the western drugs are understood to have derived from plant materials [1]. moreover these antibiotics made from these sources have not been found to cause side effects and are also found to be less expensive [2,3]. Most of the time these wastes are utilized as biofertilizers but using this agrowaste therapeutically is an innovative concept, gradually gaining acceptance moreover, these are novel, natural, eco-friendly and biodegradable. Fruit and beverage industries and distilleries dump most of the fruit peels as agrowastes, which become a source of pollution, calling for bioremediation, which is also expensive. Hence alternative methods are sought after. The current work focuses on the extraction and assay of antimicrobial components from peels of fruits and also evaluate their antioxidant potential, which is probably a new alternative way of waste management. Thus new aspects concerning the use of the wastes therapeutically are precisely interesting and attractive, as mentioned earlier, the present investigation focuses on the possibility of using the peels waste as a source of low-cost natural antimicrobials. Hence, antimicrobial activities of the samples are thought to be reported in a new perspective, thus opening up new frontiers for drug exploration. Further studies are required to determine the nature of compounds responsible for the antimicrobial effects from these extracts. Plant based extracts can be extracted from any part of plant like barks, leaves, fruits, seeds and fruit rinds etc [4]. The natural products have relevant advantages over synthetic compounds; they are easily available, comparatively cheaper, can be consumed without any side effects and are also nutritious.

Materials & Methods

Collection of plant materials

Vitis vinifera and Punica granatum fruit peels were collected from local supermarkets at Tiruchirappalli District, Tamilnadu, India during the month of December, 2013 and authenticated by the botany department of our college and the voucher specimen was deposited. The peels chosen for the study had been washed, macerated and lyophilised. About 500g of the peels yielded 33g and 56g powder respectively. The procedure was repeated to collect the needed quantity.

Preparation of plant extracts

100g of the powdered peels extracted in Soxhlet apparatus separately using 1 L of ethanol for 18h and then filtered. The filtrates were evaporated to dryness under reduced pressure and at a lower temperature in a rotary evaporator7. The dried residues were stored in airtight containers for further use. The procedure was repeated with acetone too.

Culture

Bacterial cultures as E.coli, Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae were obtained from the microbiology department of a local hospital, Tiruchirappalli, India. These clinical isolates were collected and stored in semi-solid agar and were plated out on nutrient agar plates and on MacConkey agar plates to check the viability of the bacteria.

Antimicrobial Susceptibility Testing

Disc Diffusion test [5]

All isolates were tested for susceptibility to the extracts and antimicrobial agents on Mueller Hinton agar (HiMedia India) by the standard disc diffusion method recommended by the National Committee for Clinical Laboratory Standards. The diameter of the zone of inhibition of growth was recorded, and interpreted by the criteria of CLSI [6].

Determination of Minimal Bactericidal concentration (MBC)

The minimal bactericidal concentration (MBC) can be determined by adopting the procedure outlined by CLSI using the microtitre plates and were examined for bacterial growth. The Minimal Bactericidal Concentration (MBC) assay is performed as an adjunct to the MIC and is used to determine the concentration of the extract that is lethal to the target bacteria in vitro.

Table 1

Antibacterial activity of ethanol extracts of the fruit feels by disc diffusion assay

S.NO Name of the organisms Zones of Inhibition in mm
1 E.coli 8 6
2 Klebsiella Pneumonia 10 6
3 Pseudomonas aeroginosa 6 11
4 Proteus sp. 7 9
5 Enterobacter aerogenes 9 6

Table 2

Antibacterial activity of water extracts of the fruit feels by disc diffusion assay

S.NO Name of the organisms Zones of Inhibition in mm
1 E.coli 7 7
2 Klebsiella pneumonia
3 P seudomonas aeroginosa
4 Proteus sp. 8
5 Enterobacter aerogenes

Table 3

Minimum Inhibitory Concentration of ethanol extracts of the fruit feels by broth dilution method

S.NO Bacteria MIC (mg.)
1 E.coli 20 20
2 Klebsiella Pneumonia 2.5 20
3 Pseudomonas aeroginosa 5.0 20
4 Proteus sp. 20 5.0
5 Enterobacter aerogenes 20 20

Results & Discussion

The inhibition was dependent on the type of dissolution solvent used. Inhibition was most favoured when plant sample was dissolved in ethanol. The ethanol extract exhibited good antibacterial activity against the tested gram negative intestinal bacteria, where the zones ranged between 5mm and 14mm (Table 1). The aqueous extract was not effective against several strains as shown in Table 2, indicating the effectiveness of ethanol to dissolve the compounds responsible for antibacterial activity. The minimal inhibitory concentration fell in the range between 2.5mg to 20mg, which was significant. Vitis vinifera demonstrated a good activity against Klebsiella sp., where the MIC value was only 2.5mg (Table 3). Previous studies have also reported bioactive compounds with antimicrobial activity in grape seeds [7] and in their marcs[8], pomegranate peels [9], lemon peels[10], among others. Pomegranate (Punica granatum L.; Punicaceae) has gained popularity in recent years due to its multifunctionality and nutritional benefit in the human diet. The fruit is rich in tannins and other biochemicals, particularly phenolics, which have been reported to reduce disease risk[11,12]. Besides high antioxidant capacity, pomegranate peel extracts have been reported to possess a wide range of biological actions including anti-cancer activity [13-15], antimicrobial activity [16,17], antidiarrheal activity [18], apoptotic and antigenotoxic properties [19,20], anti-tyrosinase activity [21], anti-inflammatory and anti-diabetic activities [22,23]. Polyphenol compounds such as ellagic tannins, flavonols, anthocyanins, catechin, procyanidins, ellagic acid and gallic acid have been implicated in various pharmacological activities in the fruit peel [24]. Grape (Vitis vinifera) are considered rich sources of poly-phenolic compounds, mainly monomeric catechin and epicatechin, gallic acid, and polymeric and oligomeric procyanidins [25,26] skins and seeds . The mode of action may be due to surface interaction of sterol molecules present in the extracts with the bacterial cell wall and membrane leading to alteration in the primary structure of cell wall and membrane, ultimately leading to pore formation and degradation of the bacterial components [27]. In grape skin there is another type of polyphenol, called anthocyanins, which usually have a purple color and amount to ~30% of total polyphenols in grapes. Resveratrol is mainly contained in the skins of grapes. These grape seed and skin extract compounds act as antimutagenic and antiviral agents.