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Antibacterial activity of extracts from Salvia offcinalis and Rosmarinus

officinalis obtained by sonication and maceration methods

Article  in  International Journal of Pharmacy and Pharmaceutical Sciences · February 2014

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Academic Sciences International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 6, Issue 2, 2014

Research Article
ANTIBACTERIAL ACTIVITY OF EXTRACTS FROM SALVIA OFFICINALIS AND ROSMARINUS
OFFICINALIS OBTAINED BY SONICATION AND MACERATION METHODS

MOUNYR BALOUIRI1,2, MOULAY SADIKI1,2, WESSAL OUEDRHIRI1,ABDELLAH FARAH1, SOUMYA EL ABED2, SAAD
IBNSOUDA KORAICHI2
1LPMASN, National Institute of Medicinal and Aromatic Plants, Sidi Mohamed Ben Abdellah University, BP 7048, 30007, Fez, Morocco,
2Department of biology, Faculty of sciences and techniques- Sidi Mohamed Ben Abdellah University, Fez, Morocco.
Email: [email protected]
Received: 02 Jan 2014, Revised and Accepted: 19 Feb 2014
ABSTRACT
Objective: Our study attempts to compare the phenolic contents and the antibacterial activity of the methanol extracts obtained by sonication and
classical maceration from Rosmarinus officinalis and Salvia officinalis.
Methods: The extracts were obtained from two plants belonging to Lamiaceae family; S. officinalis and R. officinalis. These plants were collected at
February 2011 in the National Institute of Medicinal and Aromatic Plants (Morocco). The total phenolic content was evaluated using Folin Ciocalteu
reagent. Furthermore, the antibacterial effect of the extract was tested against eight bacterial strains using the disk-diffusion method.
Results: Our results show that sonication was more effective for the extraction of phenolic compounds than the classical maceration. Moreover, all
extracts showed antibacterial activity against Gram-positive bacteria tested (Bacillus subtilis and Staphylococcus aureus). Gram-negative bacteria
(Pseudomonas aeruginosa and Escherichia coli) were found to be resistant.
Conclusion: The greatest antibacterial activity was exhibited by the extracts obtained using sonication method.
Keywords: Sonication, Maceration, Phenolic compounds, Antibacterial activity, Methanol extracts.

INTRODUCTION processed only by a single molecular entity [19]. Therefore, they

could have important implications for the development and
Salvia officinalis L. (common sage) and Rosmarinus officinalis L. implementation of therapeutic antimicrobial strategies and have a
(common rosemary) two important medicinal and aromatic plants potential to be used in the food industry as a preservative. The main
from Lamiaceae family. Traditionally leafs infusion of Salvia goal of our work is to evaluate in vitro the antibacterial activity of
officinalis is used by the Moroccan population against chill, methanol extracts from Salvia officinalis and Rosmarinus officinalis
rheumatism and cough [1]. Also, leaves decoction of Rosmarinus cultivated in the Garden of National Institute of Medicinal and
officinalisare used as remedies for intestinal parasites, rheumatism Aromatic Plants- Taounate (Morocco). Moreover, we determine the
and kidney. Rosemary (Rosmarinus officinalis) from plants that have most effective extraction method maceration or sonication, while
been extensively studied for their biological activities; comparing the total phenolic contents and the antibacterial activity
antioxidant[2,3], antimicrobial [4–6], anti-inflammatory [7], of crude extracts obtained.
antidepressant[8], anticancer [9], antiulcerogenic
[10],antinociceptive [11], Hepatoprotective [12], antidiabetic [13] MATERIAL AND METHODS
and spasmolytic [14].
Plants material
The plant secondary metabolites spans an extremely large and
diverse range of chemical compounds derived from plants. Several The aerial parts of Salvia officinalis and Rosmarinus officinalis were
methods are suggested for more or less selectively extracting collected from the garden of the National Institute of Medicinal and
specific classes of these compounds. For most applications, relatively aromatic Plants-Taounate (NIMAP) (Morocco) on 20 February 2011.
simple techniques, such as percolation and maceration are used. Plants were identified and deposited in the herbarium of the
However, some specific applications require more sophisticated and institute.
costly extraction techniques using specialized equipment such as
Preparation of Extracts
supercritical-fluid extraction[15].
Maceration
Nowadays, a lot of causes associated with resistance and toxicity of
microorganisms are present several problems to the Scientifics: The One hundred gram of dried and crushed aerial part of each plant was
nosocomial infections, the biofilms [16] and the emergence of macerated in first time using 300 ml of hexane during 2 hours at
multidrug-resistant microorganisms which become a major global room temperature. Extracts were filtered using Whatman paper n°1.
healthcare problem[17].The situation is particularly complicated in Second maceration was performed with 300 ml of methanol during
hospitals, with resistant Staphylococcus aureus, Streptococcus 10 hours. After filtration each mixture was evaporated under
pneumoniae and the Enterococcus strains [18]. Moreover, food is a vacuum to obtain crude extracts. Two repetitions were performed.
rich environment for most bacteria. Food-borne illnesses caused by
consumption of food contaminated with pathogenic bacteria and/or Ultrasonic Extraction
their toxins are a great problem in public health [19]. Extraction was carried out in an ultrasonic bath (Elma–Transsonic
TI-H-15) (Power: 100 W, Temperature: 30°C). Flasks containing 50 g
Among the solutions taken was the exploitation of natural resources, of air-dried and crushed plant material and 200 ml of methanol were
as plant natural products like flavonoids, tannins, coumarins, immersed in the ultrasonic bath. Sonication was performed with
alkaloids, terpenoids, lectins and polypeptides, which present an ultrasound frequency 35 KHz for 30 min (Two repetitions were
important antimicrobial activity [20]. This multiplicity of performed). After filtration each mixture was evaporated under
metabolites can inhibit the multi-resistance of bacteria which was vacuum to obtain crude extracts.
 Balouiri et al.
Int J Pharm Pharm Sci, Vol 6,Issue 2, 167-170

Phytochemical Screening for Flavonoids Staphylococcus aureus CIP 543154, Bacillus subtilis ILP 1428B,
Bacillus subtilis ILB142B, Escherichia coli 0128B12, Escherichia coli
This qualitative test was carried out according Shinoda reaction CIP 5412, Pseudomonas aeruginosa A22, and Pseudomonas
(cyanidin reaction)[21]. 1 ml of the methanol extract (crude extract aeruginosa ATCC 27853.
dissolved in 1ml of methanol) was mixed with 1 ml of hydrochloric
alcohol (1:1:1 HCl-Ethanol-Distilled water) and approximately 0.5 g Antibacterial Assay
of Mg metal. After 3 min of reaction the change of coloration was
watched. The presence of flavonoids was confirmed by the pink- The antibacterial activity of the methanol extracts was examined by
orange coloration (flavones), pink or purplish red (flavonones) or disk-diffusion method [23] with some modifications. Briefly,
red (flavonols and flavononols). bacterial strains were cultured overnight at 37 °C on Luria-Bertani
broth, then inoculum consisting of 0.5 McFarland was prepared in
Total Phenolic Content physiologic saline.
Folin-Ciocalteu reagent was used for the quantification of total Bacterial inoculum (100 µl) was inoculated in Petri dishes
phenolic content described by [22], with some modifications as containing a sterile Luria-Bertani Agar medium. Sterile filter paper
follows: 4 mg of methanol extract were added to 4 ml of distilled discs (5 mm diameter) were deposited on medium and impregnated
water, 20 µl of this mixture was then pipetted and added into a tube with 10 µl of extract solution (500 mg/ml of DMSO to 2%). The
containing 1.58 ml of distilled water and 100 µl of Folin-Ciocalteu control was performed with discs containing 10 µl of DMSO to 2%.
reagent. After 5 min, 300 µl of NaCO3 (25%) were added and the Each experiment was performed in duplicate
reaction mixture was allowed to stand for 2 hours at room
temperature, then the optical density at 765 nm was measured. RESULTS
Gallic acid was used as a standard, the results are given as mg gallic Yield of Methanol Extracts
acid equivalent per gram of methanol extract. Two repetitions were
performed. The methanol extract yield which corresponds to the maceration
extraction was slightly higher than that of sonication extraction for
Bacterial Strains both plants (Table 1). Common rosemary gave a maceration extract
with a higher yield than common sage. However, the extract of sage
The methanol extracts were tested for antibacterial activity against was obtained by sonication with a higher yield compared with
the following bacterial strains: Staphylococcus aureus ATCC 25922, rosemary.

Table 1: Yield of methanol extracts of S. officinalis and R. officinalis

Plant Extraction method Yield % (w/w)
S. officinalis Maceration 13.88 ± 1.24
Sonication 11.78 ± 0.42
R. officinalis Maceration 15.8± 0.56
Sonication 8.7± 0.70

Presence of flavonoids officinalis by sonication contain flavonoids compounds, particularly

the flavones. Of same the sage extract obtained by classical maceration
The results of phytochemical screening for flavonoids compounds are also contains flavones compounds. While, extract obtained by
represented in Table 2. The extracts obtained from R. officinalis and S. maceration from rosemary was negative for this test.

Table 2: Revelation of flavonoids of S. officinalis and R. officinalis extracts

Plant Extraction method Presence of flavonoides Type of flavonoides
S. officinalis Maceration + Flavones
Sonication + Flavones
R. officinalis Maceration - -
Sonication + Flavones

Table 3: Results of total phenolic contents of R. officinalis and S. officinalis

Plant Extraction method Concentration of total phenolic compounds (mg gallic acid equivalent/g of extract)
S. officinalis Maceration 116.5±9.19
Sonication 195.1±11.31
R. officinalis Maceration 181.4±9,89
Sonication 193.6±8,48

Table 4 : The growth-inhibitory diameters (mm) of methanol extracts against the tested bacteria
Bacterial strains S. officinalis R. officinalis
Maceration Sonication Maceration Sonication
S. aureus ATCC 25922 10 ± 0.7 13 ± 1,4 11,25 ± 0.35 13,5 ± 0,7
S. aureus CIP 543154 12 ± 0.35 15 ± 0.7 11 ± 0.35 13 ± 0.35
B. subtilis ILP1428B 12 ± 0.7 15 ± 0.35 11 ± 0.35 12 ± 0.7
B. subtilis ILB142B 8 ± 0.7 11 ± 0.7 10 ± 0.7 12 ± 0.35
E. coli 0128B12 - - - -
E. coli CIP 5412 - - - -
P. aeruginosa A22 - - - -
P. aeruginosa ATCC 27853 - - - -
 Balouiri et al.
Int J Pharm Pharm Sci, Vol 6,Issue 2, 167-170

Total phenolic content fight resistant microbes in the pharmaceutical, food and veterinary.
Moreover, our work shows likewise the effectiveness of sonication
As shown in Table 3. The total phenolic contents of the methanol as a method of extraction of bioactive compounds from plant
extracts of rosemary and sage varied from 116± 9.19 and 195.1 ± material.
11.31 (mg gallic acid eq/g of extract). Methanol was used in this
research because these phenolic compounds and most other Conflict of Interest statement
reported bioactive compounds are generally more soluble in polar
solvents[24]. Therefore, the methanol extracts obtained from both We certify that there is no conflict of interest with any financial
plants by sonication have the highest phenolic content than those organization regarding the material discussed in the manuscript
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