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Antioxidant and Antimicrobial Properties of some Plant Extracts

LILIANA CRISTINA SOARE1,2 *, MARIANA FERDE2, STEFAN STEFANOV3, ZAPRIANA DENKOVA4, RADOSVETA NICOLOVA4,

PETKO DENEV5, CAMELIA UNGUREANU2

1University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca, 3 Mntur, Cluj-Napoca, Romania,2Politehnica University of Bucharest, 313 Splaiul Independenei, 060042, Bucharest, Romania3University of Food Technology, Technical Faculty, 26 Maritza Boulevard, 4002 Plovdiv, Bulgaria,4

University of Food Technology, Technology Faculty, 26 Maritza Boulevard, 4002 Plovdiv, Bulgaria,5Institute of Organic Chemistry with Centre of Phytochemistry BAS, Laboratory of Biologicaly Active Substances, 95 VasilAprilov Boulevard, 4002 Plovdiv, Bulgaria.

The aim of this paper was to test the antioxidant and antimicrobial properties of extracts obtained fromFragaria x ananasa (strawberry, fruits), Paeonia officinalis (peony, petals),Hyacintus orientalis (hyacinth,tepals) and Scilla bifolia (hyacinth, tepals)to find new resources useful in therapy, cosmetics, food technology,etc. The antioxidant activity varied between 11.96mol TE/gFW at Fragaria x ananasa and 555.20 mol TE/gFW at Paeonia officinals (ethanolic extract). The obtained results showed that the antioxidant activity wassimilar to that obtained for different medicinal plants. Almost all tested extracts have antimicrobial activityagainst Escherichia coli ATCC 25922, Pseudomonas aeruginosa NBJMCC 1390, Salmonella abony,Staphylococcus aureus ATCC 25093, Enterococcus faecalis,Brevibacterium flavum ATCC 1406, Sarcina sp.,Bacillus cereus ATCC 1390, Saccharomyces cerevisiae and Aspergillus niger. The best result are obtained forPaeonia officinalis ethanolic extract.

Key words: antioxidant, antimicrobial,Fragaria x ananasa, Paeonia officinalis,Hyacintus orientalis,Scilla bifolia

* email: soleil_cri@yahoo.com; Tel.: +40742097352

Plants produce a wide range of metabolites withbioactive properties, so that they are used inpharmaceuticals, cosmetics, food industries etc. Theanthocyanins constitute a major flavonoid group, which isresponsible for colours ranging from salmon-pink to redand violet to dark blue in plant tissues [1]. They are thelargest group of water-soluble pigments widespread in the

plant kingdom. Chemically, these flavonoids naturally occuras glycosides of flavylium or 2-phenylbenzopyrylium saltsand are most commonly based on six anthocyanidins:pelargonidin, cyanidin, peonidin, delphinidin, petunidin andmalvidin [2]. Anthocyanins are of interest for two reasonsbecause they can be used not only in the technologicalfield as natural colorants but also have importantimplications in the field of human health [3]. Anthocyaninsplay an important role in the promotion of health anddisease prevention. They have been shown to improve riskfactors for cardiovascular health, to aid in preventing obesity,to have antioxidant, anti-inflammatory and anti-cancerproperties, and to improve vision, skin and possibly other

bioactivities [4]. Most phenolic compounds possessantibacterial or antiviral activities [5]. According to thenumbering system used by the Codex AlimentariusCommission, anthocyanins (any anthocyanin-derivedcolorant) are listed as a natural colorant by the EuropeanUnion (EU) legislation as product E163 [2]. Commercialapplications of anthocyanins as food colorants include softdrinks, fruit preserves (jams, canned fruit), sugarconfectionary (jellies), dairy products (essentially yogurts),dry mixes (acid dessert mixes and drink powders) andmore rarely frozen products (ice cream) and few alcoholicdrinks [2]. The aim of this paper was to test the antioxidantand antimicrobial properties of extracts obtained fromFragaria x ananasa,Paeonia officinalis,Hyacintus orientalis

and Scilla bifolia to find new resources useful in therapy,cosmetics, food technology, etc.

Experimental partMaterials and methodsExtracts

The extracts were obtained from cultivated or wildplants: strawberry -Fragaria x ananasa (Weston) Decne.et Naudin (Fa) (red fruits, cultivated plants, ethanolicextract), peony - Paeonia officinalis L. (Po) (red petals,

cultivated plants, ethanolic and methanolic extract),hyacinth - Hyacintus orienta lis L. (Ho) (blue tepals,cultivated plants, ethanolic extract) and violet -Scilla bifoliaL.(Sb) (blue tepals, wild plants, ethanolic extract).

Oxygen Radical Absorbance Capacity (ORAC) assay wasmeasured according to the method of [6] with somemodifications [7]. The method measures the antioxidantscavenging activity against peroxyl radical induced by 2,2-azobis (2-amidinopropane) dihydrochloride (AAPH) at 37C. Fluorescein (FL) was used as the fluorescent probe.The loss of fluorescence of FL was an indication of theextent of damage from its reaction with the peroxyl radical.The protective effect of an antioxidant was measured by

assessing the area under the fluorescence decay curve(AUC) as compared to that of blank in which no antioxidantis present. Solutions of AAPH, fluorescein and Trolox wereprepared in a phosphate buffer (75 mmol/L, pH 7.4).Samples were diluted in phosphate buffer as well. Reactionmixture (total volume 200L) contained FL - (170 L, finalconcentration 5.36x10-8 mol/L), AAPH - (20 L, finalconcentration 51.51 mmol/L), and sample -10 L. FLsolution and sample were incubated at 37 C for 20 min.,and AAPH (dissolved in 37C buffer) was added. Themixture was incubated for 30s before the initialfluorescence was measured. After that, the fluorescencereadings were taken at the end of every cycle after shaking.For the blank, 10L of phosphate buffer was used instead

of a sample. Antioxidant activity was expressed in Troloxequivalents. Trolox solutions (6.25; 12.5; 25; 50 and 100mol/L) were used for defining the standard curve. One

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ORAC unit is assigned to the net protection area, providedby a Trolox solution with concentration of 1mol/L. Thefinal ORAC values were calculated using a regressionequation between the Trolox concentration and the netarea under the curve. The antioxidant activity was

expressed in micromole Trolox equivalents per gram offresh weight (FW) standard deviation (SD).

Antimicrobial assayThe antimicrobial activity of crude extracts were tested

on 10 microorganism: five pathogen bacteria -Escherichiacoli ATCC 25922 (Eco), Pse udomon as aeru ginosaNBJMCC1390 (Pae), Sa lm on el la abon y (Sab),Staphylococcus aureusATCC 25093 (Sau) andEnterococcusfaecalis (Efa), three saprophytic bacteria -BrevibacteriumflavumATCC 14067 (Bfl), Sarcina sp. (Ssp),Bacillus cereusATCC 1390 (Bce), one yeast - Saccharomyces cerevisiae(Sce) and one mould - Aspergillus niger(Ani). The bacterial

strains were cultivated on LBG Agar medium and the yeastand mould on Malt Agar medium. Plant extracts weretested in three concentrations: undiluted extract, 10-1 and10-2. The disc diffusion assay was used to determine thegrowth inhibition of microorganisms [8-9]. Each paper discwas impregned with 6L extract. The antimicrobial activitywas expressed as the zone of inhibition (IZ, mm) producedby the plant extract. The results are presented as mean standard deviation (SD). For each extract was establishedthe minimum inhibitory concentration (MIC, ppm) as thelowest concentration of extract inhibiting the visiblegrowth of microorganism on the agar plate.

Results and discussion

Oxygen Radical Absorbance Capacity (ORAC)Many beneficial activities have been attributed to

anthocyanins and the majority of work investigating theseactivities focuses on the antioxidant characteristics ofanthocyanin rich diets and the enhancement ofcardiovascular health [3]. The oxygen radical absorbancecapacity (ORAC) method has been found to be the mostrelevant one for biologic samples [10-12]. This method ispreferred for the measurement of the antioxidant activityof foods and biological samples. Table 1 shows the ORACantioxidant activity of the plant extracts. The results arepresented as mean standard deviation (SD).

The antioxidant activity varied between 11.96 mol TE/

gFW at Fragaria x ananasa and 555.20 mol TE/gFW atPaeonia officinals (ethanolic extract). The antioxidantactivity of plants varies with species and at the samespecies differences were found depending on the solventextraction, the physical condition of plant material (fresh

or dried) and environmental factors [13]. Also, theantioxidant activity ofPaeonia extract is smaller in themethanolic extract. The obtained results showed that theantioxidant activity was similar to that obtained for differentmedicinal plants. The value of antioxidant activity ofPaeonia officinalis extract is closed to Mentha spicata(leaves, water extract) - 598 mol TE/gDW, Sa lviaofficinalis (leaves, water extract) - 609 mol TE/gDW,Rubusidaeus (leaves, water extract) - 608 mol TE/gDW,Matricaria chamomilla (flowers, water extract) - 469molTE/gDW [13] etc. The value of antioxidant activity ofHyacintus orientalis extract is closed to Betula pendula(leaves, water extract) - 142 mol TE/gDW, Cichoriuminthybus (aerial parts, water extract) - 132 mol TE/g DW),Urtica dioica (leaves, water extract) - 141mol TE/gDW[13]. Moldovan et al. [10] obtained for Urtica dioica extractORAC value: 505.21 mol TE/gDW and for Heleboruspurpurascens extract the ORAC value: 401,97mol TE/gDW.

Antimicrobial assayAll tested extract have antibacterial activity against

Escherichiacoli ATCC 25922. IZ was between 8.50 mm forHyacintus orientalis extract and 16.50 mm for Paeoniaofficinalis ethanolic extract (table 2).

IZ-inhibition zone (mm)The growth ofPseudomonas aeruginosa NBJMCC1390 is

inhibited by all extract, the best result was obtained forPaeoniaofficinalis ethanolic extract (IZ=11.50). The plant extractinhibited the growth ofSalmonella abony andEnterococcusfaecalis. Only the extract obtained fromHyacintus orientalisdid not inhibit the growth ofStaphylococcus aureus ATCC25093.

Table 1

ANTIOXIDANT ACTIVITYTable 2

ANTIBACTERIAL ACTIVITY OF PLANT EXTRACTS

Table 3

MINIMUM INHIBITORY CONCENTRATION (MIC IN PPM) FORPATHOGENIC MICROORGANISM

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The best results are obtained for Paeonia officinalis

ethanolic extract. For this extract the inhibition of bacterialgrowth was obtained at 6 ppm in the case ofSalmonellaabony, at 60 ppm in the case ofEscherichiacoli ATCC 25922,Pseudomonas aeruginosa NBJMCC1390 andEnterococcusfaecalis. 600 ppm is the MIC for Staphylococcus aureusATCC 25093 growth inhibition (table 3). The essential oilsextracted from flowers ofPaeonia daurica have betterantimicrobian activity against Staphyl lococcus aureus(IZ=25 mm), but the IZ is smaller in the case ofEscherichiacoli (IZ=12 mm),Pseudomonas aeruginosa (IZ=10 mm),Enterococcus faecalis (IZ=8 mm)[15].

The growth of all saprophytic microorganisms wasinhibited by the tested extracts (table 4) with one

exception - Bacillus cereus ATCC 1390 and Scilla bifoliaextracts.The best results were obtained for Paeonia officinalis

and Scilla bifolia extracts. The larger IZ (18 mm) wasobtained for Paeon ia of fi ci na li s in the case ofSaccharomyces cerevisiae. ForPaeonia officinalis extractthe inhibition of microorganisms growth was obtained at6 ppm in the case ofBrevibacterium flavumATCC 14067,60 ppm in the case ofSaccharomyces cerevisiae and 600ppm for Sarcina sp., Bacillus cereus ATCC 1390 andAspergillus niger(table 5). The best result for antioxidantactivity and antimicrobials activity are obtained forPaeoniaofficinalis ethanolic extract.

ConclusionsThe extracts obtained from Fragaria x ananasa fruits,

Paeonia officinalis flowers,Hyacintus orientalis flowersandScilla bifolia flowers have antioxidant activity similar tothose of different medicinal plants. Almost all tested

Table 4

ANTIMICROBIAL ACTIVITY OF PLANTEXTRACTS

Table 5

MINIMUM INHIBITORY CONCENTRATION (MIC IN PPM) FORSAPROPHYTIC MICROORGANISM

extracts have antimicrobial activity againstEscherichiacoliATCC 25922, Pseudomonas aeruginosa NBJMCC 1390,Salmonella abony, Staphylococcus aureus ATCC 25093Enterococcus faecalis (Efa),Brevibacterium flavumATCC1406, Sarcina sp., Baci llus ce reus ATCC 1390 ,Saccharomyces cerevisiae andAspergillus niger. The bestresults for antioxidant activity and antimicrobials activityare obtained forPaeonia officinalis ethanolic extract.

Acknowledgments: The work was financially supported by the projectPOSDRU/89/1.5/S/52432 from 1.04.2010 - Inst itutional organization ofa postdoctoral school of national interest Applied biotechnologywith impact in the Romanian economy; the project was co-fundedby the EU Social Fund in the framework of the Sectorial OperationalProgramme 2007-2013 for Human Resources Development.

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Manuscript receuved: 25.11.2011