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Banats Journal of Biotechnology

2012, III(6),

111

COMPARATIVE STUDY OF PHENOLIC CONTENT OF SOME PLANT

EXTRACTS

DOI: 10.7904/ 2068 – 4738 – III(6) – 111

Marian BUTU1, Nicoleta ANDREI1, Alina BUTU1, Steliana RODINO1,2,

1National Institute of Research and Development for Biological Sciences, 060031, Splaiul

Independentei 296, Bucharest, Romania, steliana.rodino@yahoo.com 2University of Agronomic Sciences and Veterinary Medicine, 011464, Mărăşti Blvd. 59, Bucharest,

Romania

Abstract. The aim of the present study was to draw a comparative analysis of the content in flavones and polyphenols of the hydro alcoholic extracts obtained from two medicinal plants, namely artichoke (leaves) and respectively, licorice (root) according to the origin of the vegetal material and the method used in the preparation of the extracts. The plants used in the experiments are two indigenous medicinal plants belonging to wild and cultivated flora in Romania. Artichocke (Cynara scolymus) and licorice (Glycyrrhiza glabra) were purchased from three local companies trading medicinal plants: Plafar, Vitaplant and Franco Impex. It was evaluated the composition of the extracts in terms of the content of polyphenolic compounds expressed in caffeic acid and flavone compounds content expressed in rutin. Plant extracts were obtained by two different methods of preparation: ultrasonation and maceration with intermittent shaking.

Keywords: flavones, polyphenols, vegetal hidroalcoolic extracts

INTRODUCTION Currently, the study of the plant

extracts for isolation of compounds that could be used in various fields is a large concern.

Thus, a large variety of vegetal extracts or bioactive constituents of plants are investigated for their valorification in the development of natural products necessary to treat infectious states–in generally and for antifungal treatment–in particular DULGER and GONUZ, 2004.

Given the national and international context regarding organic farming and environment protection, we strongly believe that there are welcome the investigations in the extraction of active ingredients with antifungal and antimicrobial activity.

In order to find the natural and ecolological methods for the control of diseases caused by fungi and bacteria, the study of antifungal potential of some plant extracts is developing and is part of current research trends on international and national level BUTNARIU, et al., 2011.

Many researches demonstrated that there is a strong relationship between the type and concentration of phenolic compounds from vegetal materials and the antifungal activity of these plants LIANG

et al., 2004; BUTNARIU, et al., 2012. Flavone compounds have been

extensively studied since the middle of last century, and research over time revealed their antifungal, antioxidant, anti–inflammatory, antiallergic and antineoplastic activity GROTEWOLD, 2006.

Plant extracts contain strongly bioactive ingredients and they are used in small concentrations.

They have a much lower impact on plants and environment than synthetic products NASCIMENTO et al., 2000.

Considering their intense antioxidant potential, polyphenolic compounds have a special importance.

In the polyphenolic compounds group the flavonoids family has a special role through its direct impact on fungi HARBRONE and WILLIAMS, 2000.

From the point of view of phytoprotection, the role of active principles can be played by a single

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112

substance or a complex of substancesBUTNARIU, et al., 2012.

The active principle or principles can be extracted and isolated from the plant product and used as pure chemicals.

The two plants taken intro study, artichoke and licorice are native plants spread all over the country.

The choice was made based on their content of secondary metabolites: phenolic compounds, tannins, terpenoids and alkaloids whose effect is known to be antifungal.

Phenolic compounds are represented by polyphenols, phenolic acids, flavonoids LI et al., 2008, BUTNARIU, et al.,

2012. Due to their natural heterogeneity,

the quality the two selected plants has large fluctuations and requires standardization of extracts, authenticity, purity and action testing YADAV and DIXIT, 2008;

HUSSAIN et al., 2009. MATERIALS AND METHODS In this study was realised the

analysis of the composition of the extracts of vegetal origin (100% purity), available in two hydroalcoholic forms (maceration and ultrasonation) derived from the following plant species: artichoke (Cynara scolymus) and licorice (Glycyrrhiza glabra).

The plant material was purchased from local trade companies from different parts of the country namely:

Plafar; Vitaplant; and Franco Impex.

Selection of sample extracts was suggested following a complex bibliographic study RAUHA et al., 2000.

Extraction procedure The plant extracts were obtained by

two different methods: shaking maceration and ultrasonation.

For each method were performed two repetitions.

The solvent used was 70% concentration ethanol, and the ratio of plant material: solvent was 1:10 SUKHDEV et

al., 2008.

Maceration procedure The grounded plant material was

mixed by shaking with the chosen solvent for 5 days and the final extract was filtered.

For ultrasonation was used a frequency of 450 kHz for 6 minutes, repeated in two series.

Spectrofotometric Analysis Absorption spectra were recorded

using the spectrototometer PG INSTRUMENTS T60 and the data obtained were processed using programs UVWin5 Software v 5.2.0 and Origin GONG,

et al., 2005. Were identified specific areas of

absorption at maximum wavelength and absorption intensities were measured FAN

et al., 2006; GONG et al., 2009. Domain 212–214 nm is

attributed to lipid compounds (phytosterols, polar lipids), some vitamins (vitamin C, E) and terpenoids;

Domain 275–290 nm is attributed to phenolic compounds (free phenolic acids and derivatives phenolic acids);

Domain 320–330 nm is attributed to flavonoid compounds in free form or glycosylated;

Domain 390–420 nm is attributed to flavonoid compounds and quinones derived from the oxidation of polyphenols–flavones;

Domain 600–660 nm is attributed to polyphenolic carboxylic acids compounds BUTNARIU, 2012.

Determination of total polyphenolic compounds

The method is based on oxidation reaction with reactive Folin–Ciocalteu (mixture of sodium tungstate and sodium molybdate) phenol groups in a highly alkaline environment.

The following reagents were used: Folin–Ciocalteu reagent (FCR) and anhydrous solution of NaCO3 20%.

Concentration of polyphenolic compounds from the obtained extracts was calculated using a standard curve for caffeic acid.

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Banats Journal of Biotechnology

2012, III(6),

113

caffeic acid

Absorbance was measured at λ = 660 nm.

Formula for determination of total polyphenolic compounds expressed in caffeic acid was determined based on the standard curve (Figure 1).

Figure 1. Standard curve for caffeic acid (mg/ml)

Quantitative spectrophotometric

determination of fatty polyphenol acids is based on oxidation reaction of phenol groups with Folin–Ciocalteu reagent, in a strongly alkaline medium. The product obtained is colored in blue and the absorbance is measured at λ = 660 nm.

1.09705

0.45512Amg/mlAcid Caffeic

Flavone content in vegetal extracts was determined using the standard calibration curve performed for rutin.

rutin

After incubation in the dark at room

temperature for 45 minutes, the sample

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114

absorbance was measured spectrophotometrically at λ = 420 nm.

The formula for determination of total flavone compounds expressed in rutin was established based on standard curve (Figure 2).

56505.0

32407.1A)ml/mg(Rutin

Figure 2. Standard curve for rutin (mg/ml)

RESULTS AND DISCUTIONS The following tables are presenting

the data obtained after the quantitatively determination of the total polyphenols and total flavonoids content for the studied plant extracts (Table1, 2).

MacerationUltrasonication

PlafarVitaplant

Franco Impex0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

Tota

l con

tent

of p

olyp

heno

lic c

ompo

unds

ex

pres

sed

in c

affe

ic a

cid

(mg/

ml)

Figure 3. Total content of polyphenolic

compounds expressed in caffeic acid for C. scolymus

In generally, the results showed that

the total content of polyphenolic compounds expressed in caffeic acid (Figure 3, 4).

MacerationUltrasonication

PlafarVitaplant

Franco Impex0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

Tota

l con

tent

of p

olyp

heno

lic c

ompo

unds

ex

pres

sed

in c

affe

ic a

cid

(mg/

ml)

Figure 4. Total content of polyphenolic

compounds expressed in caffeic acid for G. glabr

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Banats Journal of Biotechnology

2012, III(6),

115

Table 1. Total content of polyphenolic compounds according to the extraction method used and the

place of harvest for each selected plant Total content of polyphenolic compounds (mg/ml) Plafar Vitaplant Franco Impex 0.85698 0.89214 0.92531 maceration 0.84623 0.87896 0.91873 0.61957 0.66982 0.72198 C. scolymus

ultrasonation 0.63187 0.68132 0.73723 0.83751 0.80948 0.87965

maceration 0.84763 0.84521 0.86421 0.48621 0.40463 0.47321

G. glabra ultrasonation 0.42812 0.42451 0.41356

Table 2.

Total content of flavone compounds according to the extraction method used and the place of harvest for each selected plant

Total content of flavone compounds expressed in rutin (mg/ml) Plafar Vitaplant Franco Impex 0.89158 0.93962 0.98476 maceration 0.89332 0.97423 0.97996 0.49028 0.54643 0.68732

C. scolymus ultrasonation 0.51002 0.53982 0.69237

0.93754 0.87266 0.98645 maceration 0.94721 0.88357 0.98421 0.68652 0.60732 0.73654

G. glabra ultrasonation 0.74219 0.67386 0.76321

MacerationUltrasonication

Plafar

VitaplantFranco Impex

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

Tota

l con

tent

of f

lavo

nes c

ompo

unds

exp

ress

ed in

ru

tin (m

g/m

l

Figure 5. Total content of flavone

compounds expressed in rutin for C. scolymus

Figure 6. Total content of flavone

compounds expressed in rutin G. glabra

The total content of flavone

compounds expressed in rutin (Figure 5, 6) were higher in the case of the hydroalcoholic extracts obtained by maceration than by ultrasonation.

CONCLUSIONS The studies presented in this

paper aimed to obtain and to realize a

chemical characterization of medicinal plant extracts rich in phenolic compounds with antifungal potential.

It was aimed the determination of total polyphenolic compounds expressed in caffeic acid and total content of flavone compounds expressed in rutin.

UV–Vis spectroscopic analyses were useful in highlighting the specific

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phytochemical compounds from the investigated plants.

It was observed that the highest total content of polyphenolic compounds expressed in caffeic acid is found in the extract of artichoke, C. scolymus from Franco Impex with a value of 0.92531 mg / ml.

In terms of total content of flavone compounds expressed in rutin was determined that hydroalcoholic extracts obtained by maceration of plants are almost equal: artichokes –0.98476 mg/ml, licorice–0.98645 mg/ml.

The analyses revealed that on the one hand, the method of obtaining hydroalcoholic extracts is important, because macerated plants yielded extracts with a higher content in phenolic compounds, and on the other hand the place of origin of the plants is equally important because the plant material from Franco Impex–Braila resulted in extracts with a higher content in active principles.

The results obtained were compared to other research in the field, both nationally, and internationally, and are comparable with those of other researchers who used similar technology and plants.

Study of antibiotic potential of natural products represents a modern direction of biological research.

The literature describes several methods and techniques of evaluation and certification of this potential, but unfortunately their sensitivity is not constant and comparable.

There is no standardized protocol for screening the antimicrobial activity, but many variants, given techniques modified in working conditions.

For this reason, the results obtained when working with plant extracts are definitely influenced by the method chosen.

Generally speaking, plant extracts are a promising alternative for use in organic farming.

Acknowledgements This work has been funded by the

research contract PN–II–PT–PCCA 106/2012.

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Received: October 3, 2012

Accepted: October 30, 2012