**3.8 Antibacterial activity of** *Cytisus capitatus*

*Cytisus capitatus* extracts showed equable antibacterial activity. MICs were in the interval from 5 mg/ml tо >20 mg/ml for ethanol extract, from 1.25 mg/ml tо >20 mg/ml for ethyl acetate extract and from 1.25 mg/ml to >20 mg/ml for acetone extract. Based on statistic analysis no difference was noted in acting between extracts (*p*<0.05). The obtained results are presented on the Figure 9.

The most sensitive bacteria to tested extracts were *Bacillus subtilis* and *Staphylococcus aureus* ATCC 25923. MIC for *Bacillus subtilis* showed at 5 mg/ml for ethanol extract, 2.5 mg/ml for ethyl acetate extract and 1.25 mg/ml for acetone extract. For *Staphylococcus aureus* ATCC 25923, MIC of ethanol extract was 10 mg/ml, ethyl acetate 1.25 mg/ml and acetone extract 2.5 mg/ml. *Escherichia coli* showed resistance to all three extracts. The results for

Antibacterial Activity of Naturally Occurring Compounds from Selected Plants 17

*Staphylococcus aureus* and *Enterococcus faecalis* are unexpected since Gram-positive bacteria were more sensitive than Gram-negative bacteria, and in this case they showed resistance or lowered sensitivity. Growth of other bacteria was inhibited at approximately the same concentration. Antibacterial activity of *Cytisus capitatus* was tested for the first time in this study. Based on literary data and other species from *Cytisus* genus are less explored as potential antimicrobial agents. Benaiche, 2007 tested antibacterial activity of *Cytisus purgans*  methanol extract in relation to *Staphylococcus aureus*, *Escherichia coli* and *Pseudomonas* 

*Melilotus albus* extracts showed weaker antibacterial activity than other tested plants. Between extracts, the most active one was acetone extract, then ethyl acetate and ethanol extract which was confirmed by statistic analysis (*p*EtOH = 0,018; *p*EtAc=0,029). Action interval of extracts was from 1.25 mg/ml tо 20 mg/ml (Figure 10). In most of the cases ethanol

Fig. 10. Antibacterial activity of *Melilotus albus* extracts expressed as MIC values (mg/ml)

*Bacillus subtilis, Staphylococcus aureus* ATCC 25923 and *Pseudomonas aeruginosa* ATCC 27853 showed sensitivity according to tested extracts where *Bacillus subtilis* and *Staphylococcus aureus* ATCC 25923 were the most sensitive (MIC=1.25; 2.5 mg/ml). *Escherichia coli* showed resistance to all three extracts, while *Klebsiella pneumoniae*, *Staphylococcus aureus* and *Enterococcus faecalis* showed resistance to ethanol and ethyl acetate extract. *Pseudomonas aeruginosa*, *Proteus mirabilis* and *Escherichia coli* ATCC 25922 were resistant only to ethanol extract. Aćamović-Đoković et al., 2002 tested antibacterial activity of petrol ether and ethyl acetate extract of *Melilotus officinale, Melilotus albus* and *Melitis melissophyllum* in relation to

*aeruginosa*. The biggest zones of inhibition showed at 80 mg/ml.

**3.9 Antibacterial activity of** *Melilotus albus*

extracts did not act at tested concentrations.

Fig. 8. Antibacterial activity of *Cytisus nigricans* extracts expressed as MIC values (mg/ml)

Fig. 9. Antibacterial activity of *Cytisus capitatus* extracts expressed as MIC values (mg/ml)

*Staphylococcus aureus* and *Enterococcus faecalis* are unexpected since Gram-positive bacteria were more sensitive than Gram-negative bacteria, and in this case they showed resistance or lowered sensitivity. Growth of other bacteria was inhibited at approximately the same concentration. Antibacterial activity of *Cytisus capitatus* was tested for the first time in this study. Based on literary data and other species from *Cytisus* genus are less explored as potential antimicrobial agents. Benaiche, 2007 tested antibacterial activity of *Cytisus purgans*  methanol extract in relation to *Staphylococcus aureus*, *Escherichia coli* and *Pseudomonas aeruginosa*. The biggest zones of inhibition showed at 80 mg/ml.

#### **3.9 Antibacterial activity of** *Melilotus albus*

16 Antimicrobial Agents

Fig. 8. Antibacterial activity of *Cytisus nigricans* extracts expressed as MIC values (mg/ml)

Fig. 9. Antibacterial activity of *Cytisus capitatus* extracts expressed as MIC values (mg/ml)

*Melilotus albus* extracts showed weaker antibacterial activity than other tested plants. Between extracts, the most active one was acetone extract, then ethyl acetate and ethanol extract which was confirmed by statistic analysis (*p*EtOH = 0,018; *p*EtAc=0,029). Action interval of extracts was from 1.25 mg/ml tо 20 mg/ml (Figure 10). In most of the cases ethanol extracts did not act at tested concentrations.

Fig. 10. Antibacterial activity of *Melilotus albus* extracts expressed as MIC values (mg/ml)

*Bacillus subtilis, Staphylococcus aureus* ATCC 25923 and *Pseudomonas aeruginosa* ATCC 27853 showed sensitivity according to tested extracts where *Bacillus subtilis* and *Staphylococcus aureus* ATCC 25923 were the most sensitive (MIC=1.25; 2.5 mg/ml). *Escherichia coli* showed resistance to all three extracts, while *Klebsiella pneumoniae*, *Staphylococcus aureus* and *Enterococcus faecalis* showed resistance to ethanol and ethyl acetate extract. *Pseudomonas aeruginosa*, *Proteus mirabilis* and *Escherichia coli* ATCC 25922 were resistant only to ethanol extract. Aćamović-Đoković et al., 2002 tested antibacterial activity of petrol ether and ethyl acetate extract of *Melilotus officinale, Melilotus albus* and *Melitis melissophyllum* in relation to

Antibacterial Activity of Naturally Occurring Compounds from Selected Plants 19

confirm this statement. Significant activities of ethanol extract from *Aegopodium podagraria*  and ethyl acetate and acetone extract from *Cichorium intybus*, one of insufficiently explored plants, indicate their use as potential, new antibacterial agents. These results, also, offer a scientific basis for the traditional use of extracts of *Salvia officinalis* and *Melissa officinalis*. The extracts from *Clinopodium vulgare*, *Torilis anthriscus*, *Cytisus nigricans*, *Cytisus capitatus* and *Dorycnium pentaphyllum* showed interesting activity against certain pathogenic bacteria. Mostly, the most sensitive bacteria were *B. subtilis* and *S. aureus* ATCC 25923. Susceptibility of *E. cloaceae*, *Ent. faecalis*, *K. pneumoniae*, *S. aureus*, *Ps. aeruginosa* ATCC 27853 was moderate while *Ps. aeruginosa*, *P. mirabilis*, *E. coli* were resistant. This study represents the first preliminary report on antibacterial activity of the extracts from *Cytisus nigricans*, *Cytisus capitatus* and *Dorycnium pentaphyllum* and contributes to overall examine antibacterial activity of plant species. Since, the compounds and mechanisms of action responsible for the antibacterial activities of these extracts are currently unclear; the further work will be performed on the isolation and identification of the active compounds and understanding of

This work was supported by the Ministry of Science and Education of the Republic of Serbia

Alves, T.M., Silva, A.F., Brandão, M., Grandi, T.S., Smânia, E., Smânia Júnior, A. & Zani, C.

Oswaldo Cruz, Vol. 95, No. 3, (May-Jun 2000), pp. 367-373, ISSN 0074-0276 Ahmad, I. & Beg, A.Z. (2001). Antimicrobial and phytochemical studies on 45 Indian

Andrews, J.M. (2001). Determination of minimum inhibitory concentrations. *Journal of Antimicrobial Chemotherapy*, Vol. 48, Suppl. 1, pp. 5-16, ISSN 0305-7453. Akroum, S., Satta, D. & Lalaoui, K. (2009). Antimicrobial, Antioxidant, Cytotoxic Activities

Acamovic-Djokovic, G., Djukic, D., Mandic, L., Kalinic, S. & Boskovic, T. (2002).

Brkovic, D.L., Comic, Lj. & Solujic-Sukdolak, S. (2006). Antibacterial activity of some plants

Benaiche G. (2007). Extraction and GC/MS Analysis of major Alkaloids found in the Family

*Research,* Vol. 31, No. 2, pp. 289-295, ISSN 1450-216X

*Journal of Science*, Vol. 28, pp. 65-72, ISSN 1450-9636

and Engineer Sciences, University of M'sila, Algeria, Africa

*sirovine,* Vol. 22, pp. 59-63, ISSN 0455-6224

(2000). Biological Screening of Brazilian Medicinal Plants. Memórias do Instituto

medicinal plants against multi-drug resistant human pathogens. Journal of Ethnopharmacology, Vol. 74, No. 2, (February 2001), pp. 113–123, ISSN 0378-8741 Atindehou, K.K., Koné, M., Terreaux, C., Traore, D., Hostettmann, K. & Dosso, M. (2002).

Evaluation of the antimicrobial potential of medicinal plants from the Ivory Coast. Phytotherapy Research, Vol. 16, No. 5, (August 2002), pp. 497–502, ISSN 0951-418X

and Phytochemical Screening of Some Algerian Plants. *European Journal of Scientific* 

Antimicrobial activity of the petrol-ether and ethyl-acetate extracts of Melilotus officinalis (L.) Pall, Melilotus albus Medic. and Melitis melissophyllum L. *Lekovite* 

from family Apiaceae in relation to selected phytopathogenic bacteria*. Kragujevac* 

of Fabaceae. Academic Dissertation, Department of Chemistry, Faculty of Sciences

mechanisms of action.

**5. Acknowledgment** 

(grant number OI173032).

**6. References** 

*Escherichia coli*, *Proteus mirabilis*, *Salmonella enteritidis*, *Pseudomonas aeruginosa*, *Streptococcushaemoliticus* A, *Staphylococcus aureus* and *Candida albicans*. *Melilotus albus* extracts were less efficient than other tested plants.

#### **3.10 Antibacterial activity of** *Dorycnium penthaphyllum*

*Dorycnium penthaphyllum* extracts showed different level of antibacterial activity. Ethanol extract acted in the interval from 2.5 mg/ml tо 20 mg/ml, ethyl acetate from 1.25 mg/ml tо >20 mg/ml, and acetone extract from 1.25 mg/ml tо 20 mg/ml (Figure 11.). Between the extracts there was no statistically significant difference in action (*p*<0.05).

Fig. 11. Antibacterial activity of *Dorycnium penthaphylum* extracts expressed as MIC values (mg/ml)

The most significant results were obtained for *Bacillus subtilis, Staphylococcus aureus* ATCC 25923, *Pseudomonas aeruginosa* and *Proteus mirabilis*. MIC values were between 1.25 mg/ml - 10 mg/ml. Other bacteria showed sensitivity at approximately the same concentrations (20 mg/ml). The exception was *Escherichia coli* which was resistant to ethylaacetate extract. Sensitivity of tested bacteria to the extracts of *D. penthaphylum* was presented for the first time in this study. A group of scientists tested anti - *Helicobacter pilory* effect of medicinal plants of Greek traditional medicine, among which is also *D. penthaphylum,* although they did not note this plant's effect (Stamatis et al., 2003).
