ANTIBACTERIAL AND ANTIOXIDANT PROPERTIES OF SOME PLANT EXTRACTS WITH PROPOLIS

For many years, plants have been utilized in food, healing materials, and curing for many illnesses. Lately, improvements in biological searches have displayed the notable potential of natural compounds. Objective: In this study, biological activities of extracts of herbal mixtures with propolis were investigated. Materials and Methodology: Ethanol and hexane extracts


INTRODUCTION
Plant are valuable sources that can be utilized in many industries.Products based on natural substances attract attention in many industries, such as cosmetics and pharmacy in recent years Stuper-Szablewska et al. (2022).
Antimicrobial agents have important roles against pathogens.On the other hand, using excessive antimicrobial in the world cause drug resistance and undesirable effects.Many bacteria gained resistant to current antibiotics.The increase in untreatable infections require the discovery and development of brand antibacterials.
Plants have been utilized to cure bacterial infections thousands of years ago.Many people in developing countries utilize herbal drugs for antibacterial illnesses Liang et al. (2022).
Oxidative stress may lead Alzheimer's, cancer, and cardiovascular illnesses.Free radicals can be harmful proteins, lipids, nucleic acids and carbohydrates.The utilization of antioxidants can delay or retard the oxidation of biomolecules.Medicinal plants have been utilized as natural antioxidant agents for centuries by people Guchu et al. (2020).
Synthetic antioxidants create negative health effects.Hence, studies about natural antioxidants is valuable.Polyphenolic compounds can behave as antioxidants and free radical scavengers.Studies about polyphenols from plants has now achieve considerable attention Shahinuzzaman et al. (2020).
Papaver somniferum is an annual plant which belons to the family Papaveraceae.P. somniferum use as a foodstuff in manufacturing of bakery products.Moreover, it's oil has various medicinally important metabolites Chmelová et al. (2018).
Syzygium aromaticum possess various medical features like antimutagenic, antibacterial, anti-inflammatory and radical scavenging activity Faujdar et al. (2020).Moreover, It has antioxidant action so it utilizes in preventing some degenerative diseases.Syzygium aromaticum bud oil heals wounds and burns Alanazi et al. (2022).
Foeniculum sp.belongs to family Apiaceae and cultivated in India, China and Egypt.The volatile oil of fennel possess anti-allergic, diuretic, anti-spasmodic, antimicrobial and antioxidant potencies Eliuz et al. (2016).
Bees gather resins from plants, stir them with their own salivary enzymes and beeswax which composes propolis.Propolis possess many important effects such as antibacterial, radical scavenging, antiparasitic, antifungal and antiproliferative Przybylek & Karpinski (2019).
This investigation aims the compare biological features of extracts of propolis-Syzygium aromaticum mixture, propolis-Papaver somniferum mixture, propolis-Foeniculum sp.mixture.

SUPPLYING OF PLANTS USED IN THE STUDY
Propolis, Syzygium aromaticum, Papaver somniferum and Foeniculum sp. were brought from a herbal shop in Giresun, Turkey.

PREPARATION OF THE EXTRACTS
for 24 h utilizing 300 mL ethanol and hexane, separetely.The extracts were filtered through Whatman filter paper No. 1 and residues were evaporated (40 °C) with rotary evaporator Murugan & Parimelazhagan (2014).

ANTIBACTERIAL ACTIVITY
The discs (6 mm diameter) on the petri were impregnated with 20 μL of extracts, separetely.Gentamycine was used as standard antimicrobial agent.DMSO was used as negative control.Plates were incubated for 24 h at 37°C.Diameter of zones were measured with a ruler Murray et al. (1995), Šariš et al. (2009).The tests were carried out twice.

THE DETERMINATION OF MINIMUM INHIBITION CONCENTRATION (MIC)
Minimum inhibition concentration of extracts which created ≥10 mm inhibition zones were determined.Method of Yiğit et al. (2009) were used to reveal MIC values of the tested extracts Yiğit et al. (2009).

ANTIOXIDANT ACTIVITY
All antioxidant tests were carried out three times.Results are expressed as the mean ± standard deviation (S.D.) of each triplicate test.

CUPRAC ACTIVITY
CUPRAC activity of the extracts were studied according to the method of Özyürek et al. (2009).Butylated hydroxytoluene (BHT) was used as a standard antioxidant agent.

DPPH RADICAL SCAVENGING ACTIVITY
Extracts were prepared at 250-1000 µL/mL concentrations by the method of Blois (1958).BHT and Rutin were used as standards.
The DPPH radical scavenging activity was calculated using the following equation:

RESULTS AND DISCUSSION
In the current study it was tested antibacterial efficiencies of ethanol and hexane extracts of propolis-Syzygium aromaticum, propolis-Papaver somniferum and propolis-Foeniculum sp.against test bacteria.Table 1 demonstrates inhibition zones.
Ethanol extracts exhibited higher activity than hexane extracts.Antibacterial activities of mixtures were increased in the following order: propolis-Syzygium aromaticum> propolis-Papaver somniferum> propolis-Foeniculum sp.While inhibition zones was created by ethanol extracts ranges from 7±1.41 mm to 19.5±2.12 mm, inhibition zones was created by hexane extracts ranges from 7±0.00 mm to 14±1.41 mm.Gentamycine showed higher activity when compared with tested extracts.DMSO showed no activity against test bacteria.MIC values of the extracts were given in Table 2. MIC describes as the minimum antimicrobial agent that inhibits the growth of microorganisms.Low MIC values means higher antibacterial effect.While the lowest MIC value was found in PSE as 0.0003 mg/mL against S. aureus, the highest MIC value was found in PSH as 1.5 mg/mL against B. cereus and in PPH as 1.5 mg/mL against Y. pseudotuberculosis.• Antioxidant activity Table 3 summarizes total phenolic content of the extracts.When total phenolic content was compared between extracts, it was found that ethanol extracts have higher phenolic content than hexane extracts.The highest and the lowest phenolic contents were found as 389.81±0.001µg GAE/mL and 100.57±0.012µg GAE/mL in PSE and PFH, respectively.Table 6 shows DPPH radical scavenging activity of the extracts.It was found no activity in PFH and PPH.PSE exhibits higher activity than Rutin and BHT which were used as standard antioxidant agents.CUPRAC activity of the extracts were presented in Table 7.The highest activity was determined in PSE.All extracts were showed higher activity than BHT except for and PFH.There is no study about antibacterial activity of propolis-Syzygium aromaticum, propolis-Papaver somniferum and propolis-Foeniculum sp.On the other hand, there are many studies about propolis, Syzygium aromaticum, Papaver somniferum and Foeniculum sp.
One of the most common and known most studied properties of propolis is its antimicrobial activity.Numerous studies have been conducted on its effects on fungi and viruses Albayrak & Albayrak (2008).Veiga et al. (2017) reported that propolis inhibited gram posivite bacteria and gram negative bacteria such as methicilline resistant Staphylococcus aureus.Yildirim et al. (2016) investigated effect of propolis against tuberculosis and they found that propolis is efficient against many mycobacteria species.
In a study which was carried out by Nzeako et al. (2006)  Like antibacterial studies, there is no study about antioxidant activity of propolis-Syzygium aromaticum, propolis-Papaver somniferum and propolis-Foeniculum sp.On the other hand, there are many studies about propolis, Syzygium aromaticum, Papaver somniferum and Foeniculum sp.Kocot et al. (2018) found that extracts of propolis had better DPPH and ABTS radicals scavenging activities than standard antioxidant agents such as BHT and ascorbic acid.Can et al. (2016) investigated total phenolic content and antioxidant activity of propolis samples from Azerbaijan.It was stated that total phenolic contents of samples ranges from 10.94 to 79.23 mg GAE/g.Propolis samples which was obtained from Ismayilli.Zerdap and Qax had higher antioxidant activity when compared with other districts.
Muhson & Al-Mashkar (2015) investigated total phenolic content, total flavonoid content and DPPH radical scavenging activity and acetone extracts of stem and fruit parts of Syzygium aromaticum.DPPH radical scavenging activity was found %87.50 and %79.41 in fruit and stem, respectively.Elaleem et al. (2017).studied DPPH radical scavenging activity of methanol, petroleum ether and chloroform extracts of seeds of Papaver somniferum.It was found that only methanol extract had activity.

CONCLUSION
The results revealed that these plant-propolis mixtures could play as a promising antibacterial and antioxidant agents, because of their high activities.Moreover, the current study suggests that these mixtures might be developed as pharmaceutical products.Hence, studies on the isolation and identification of substances responsible for biological activities in these mixtures should be expanded.

CONFLICT OF INTERESTS
None.
of control A1: Absorbance of sample or standard

Table 3 Table 3 Total Phenolic Contents of the Extracts (µg GAE/mL)
Total flavonoid content of the extracts were presented in Table4.Ethanol extracts possess higher flavonoid contents than hexane extracts.Total flavonoid contents of the extracts as follows: PSE>PPE> PSH>PFE>PFH>PPH.

Table 4 Table 4 Total Flavonoid Content of the Extracts (µg QE/mL)
Total antioxidant capacity of the extracts were given in Table5.The highest and the lowest values were found as 190.54±0.031µg AAE/mL in PSH and 46.03±0.018µg AAE/mL in PFH.

Table 7 Table 7 CUPRAC Activity of the Extracts and Standard Extract Concentration (µg/mL) CUPRAC Activity (nm)
water extract and essential oil of Syzygium aromaticum showed antibacterial activity against Streptococcus pyogenes, Corynebacterium spp.. Salmonella spp.and Bacteroides fragilis.Gupta & Prakash (2021) used extracts and essential oil of Syzygium aromaticum against Halobacteria sp., Lactobacillus sp., Pseudomonas sp., Micrococcus sp. and Streptococcus mutans which cause dental plaques and cavities.It was concluded that essential oil of Syzygium aromaticum had better activity than extracts of Syzygium aromaticum.Masood et al. (2008) found aqueous infusions of Papaver somniferum seeds had no activity and aqueous decoction had very weak activity against 188 bacterial isolate.Mishra and Pathak (2021) reported methanol and water extracts of Papaver somniferum seeds had activity against Salmonella and Escherichia coli but they were no activity against Pseudomonas.Eliuz et al. (2016) revealed essential oil of Foeniculum sp.had antibacterial effect against E. coli, Klebsiella pneumoniae, Salmonella typhimurium, Bacillus subtilis, Staphylococcus aureus and Enterococcus faecalis.Yıldırım et al. (2010) carried out a study about antibacterial activity of Foeniculum sp.It was reported that ethanol and aqueous extracts of Foeniculum sp.didn't inhibit E. coli, S. aureus, B. cereus, K. pneumoniae and Enterobacter sp. but methanol extract of Foeniculum sp.inhibited S. aureus and Enterobacter sp.