Potential of wild plant Artemisia judaica L. as sustainable source of antioxidant and antimicrobial compounds

Artemisia judaica L. (Shih Balady, family Asteraceae), is a medicinal plant that grows in the desert, commonly used as tea by population in Egypt Sinai and of Saudi Arabia. The present study aims to evaluate antimicrobial and antioxidant activities of aerial parts of A. judaica collected from the inland desert (Wadi Hagoul) of Egypt. Total phenolics, tannins, alkaloids, flavonoids, and saponins were measured in the plant of interest. According to the reduction of DPPH the anti-oxidant activity was measured. Different extracts of A. judaica were screened for their antimicrobial activity against nine pathogenic microorganisms using filter paper disc assay. The secondary metabolites of A. judaica were examined and proved that it was rich in tannins and phenolics (13.29 and 7.62 mg/g dw). The IC 50 values of A. judaica extract was 1.78 mg.ml -1 compared to standard catechol (0.15 mg.ml -1 ). In the present study, the extracts of A. judaica showed unusual activity depending on the type of bacteria and fungi. Also, ethyl acetate and methyl alcohol extracts showed considerable broad spectrum (77.78%, each) against both bacteria and fungi, followed by acetone, then methylene chloride and petroleum ether extracts. This study showed that A. judaica extracts could be used as antimicrobial substances in pharmaceutical and food preservation systems.


INTRODUCTION
Nearly all cultures in the world depends on medicinal plants medicinal purpose.Around 60% of population continue to use herbal extraction in health care [1,2].Many species of the genus Artemisia have been identified as aromatic and used in medicine [3].Artemisia judaica L. belongs to the family Asteraceae (Compositae) and known as wormwood (Arabic name, Shih Balady).It is a fragranced shrub that widely grows in the Arabian area [4]. A. judaica is used as a tea commonly by population in Egypt Sinai and of Saudi Arabia.In the Arabic area traditional medicine; A. judaica is used as a treatment for cardiovascular health, and many other diseases and dysfunctions [5,6].
Secondary plant metabolites are active chemical defense against pathogens as well as they play an essential role in plant defense against herbivory or other interspecies defenses [7][8][9][10].Phytochemical analysis of A. judaica which carried out by many authors [11][12][13][14] showed antioxidant activity [15,16], anti-malarial, antibacterial, anti-inflammatory [17,18], biopesticide [19].In the current paper, we investigate the potential antioxidant and antimicrobial properties of A. judaica collected from the inland desert (Wadi Hagoul) of Egypt, to evaluate their medicinal potentiality and their future industrial uses.

Determination of the Radical Scavenging Activity
The antioxidant activity by DPPH was determined as described by Miguel [25].

Preparation of the crude extracts
Plant extract was prepared by standard methods of Freedman et al. [26] and Su and Horvat [27].In each solvent, the treated filtrate evaporates, and the dried residue was dissolved in dimethyl sulfoxide (DMSO) and reserved at -20 o C for future use [28].

Antimicrobial Assay
The plant extracts were examined for the presence of antimicrobial bioactivity by the method of Cappuccino and Sherman [29] using different bacterial species (Bacillus subtilis, Escherichia coli, Klebsiella pneumonia, Staphylococcus aureus, and Streptococcus pyogenes) and fungi (Candida albicans, Aspergillus niger, Aspergillus fumigatus, and Mucor spp.).The tested samples were taken from the Laboratory of Bacteriology, Department of Botany, Faculty of Science, Mansoura University, Egypt.
The filter paper discs are prepared with a diameter of 5 mm and sterilized in the autoclave for 20-30 minutes.Then taken by sterile forceps, soaked in DMSO solution and then placed over the surface of the inoculated nutrient agar.In 37 o C the Petri dishes were incubated for 24 hours, While in fungi, A sterile disc set over the surface of the inoculated potato dextrose agar and incubated for 3-4 days at 28 o C.After incubation for period of time, the inhibition zones diameter (mm) was calculated to record the clear zone to determine the most active extract against pathogenic microorganisms.

Phytochemical Constituents
Plants are the basis of many natural products such as; alkaloids, amines, cyanogenic glycosides, flavonoids, glucosinolates, phenylpropanes, terpenes, saponins, steroids, etc.It is estimated that there are more than 200,000 chemical structures is synthesized by plants [30,31].Analysis of secondary metabolites of A. judaica showed high values of tannins and phenolics (13.29 and 7.62 mg/g dry weight, respectively).Besides, saponins, alkaloids, and flavonoids (3.55, 2.84 and 1.63 mg/g dry weight, respectively) showed comparable contents, (Figure 1).These results were consistent (compatible) with the results of Essiett and Akpan [32] on Aspilia africana and Tithonia diversifolia (Asteraceae) and Erol-Dayi et al. [33] on Centaurea species, but they were lower than some wild plants such as Anthemis arvensis, Artemisia campestris, Senecio glaucus and Urospermum picroides [34][35][36].It is not surprising to discover that secondary metabolites of the plant have medicinal activities, as most of the prescriptions have plant based origin [30].

Antioxidant Activity
Most organisms own antioxidant systems against ROS [37].Excessive ROS states have main action in diseases like idiopathic pulmonary fibrosis and respiratory distress syndrome [38].The evaluation of the antioxidant activity evaluation in plant extracts is shown in Figure 2. By increasing the concentration of plant extract, a continuous corresponding increase in the scavenging activity.In methanol extract and standard antioxidant, the scavenging activities were 55.43% and 82.72%, respectively at 4000 μg.ml -1 .
The IC 50 values of A. judaica extract was 1.78 mg.ml -1 compared to standard catechol (0.15 mg.ml -1 ).According to Al-Ismail et al. [39], A. judaica extract showed moderate scavenging activity (IC 50 ≥ 1 and ≤ 2 mg/ml).These results suggest that methanol extract of A. judaica has an obvious effect on scavenging of DPPH radical.Similar results were reported by Pandey and Singh [40] on the same genus Artemisia, El-Amier and Abdullah [41] on Calligonum comosum and Salem et al. [42] on Silybum marianum.
The plant extract potent antioxidant activity can be mainly attributed to phenolic content, as a result of heir hydroxyl group, and/or flavonoids that react with DPPH radical by donating hydrogen atom the free radicals [43], while a highly  positive correlation between total phenolic content and antioxidant activity was established in the case of many plant species [11,44,45].

Antimicrobial Activity Assessment
The plants showed well defined action agains microbes [46,47].In this investigation, the petroleum ether extract showed higher activity (Figure 3).The Stroptococcus pyogenes was the most potent inhibitor followed by Staphylococcus aureus and Bacillus subtilis.Methylene chloride does not act on Escherichia coli and S. aureus but inhibiting others.The ethyl acetate and methyl alcohol extracts inhibit all pathogenic bacteria with different rates.Previous reports supports these results [48,49].These results were similar to those of Ocimum sanctum Xanthoxylum armatum, Cinnamomum zeylanicum, and Origanum majorana as investigated by , Joshi, et al. [50] and Iranbakhsh et al. [51] in Datura stramonium (B.subtilis, E. coli, and S. aureus).
On the other hand, petroleum ether extract had no antifungal activities on all the pathogenic fungi except Candidia albicans (Figure 3).The methylene chloride extract inhibited the growth of Aspergillus fumigatus only, and ethyl acetate extract does not affect both A. fumigatus and Mucor spp.But hindered others.No antifungal activities were detected with acetone extract on all the pathogenic fungi except Mucor spp.Methyl alcohol extract inhibited the growth of C. albicans and A. fumigatus (Figure 3).Abdel-Sattar et al. [52] reported Achillea biebersteinii and Vernonia schimperii was active against the same tested fungi strain.Also, these results were compared with other investigators [53][54][55].
The extracts of A. judaica showed action (77.78%, each) against microbes (Figure 4), followed by acetone (55.56%) then methylene chloride and petroleum ether extracts (44.44%, each).The pathogen S. aureus and E. coli (13 and 11 mm, respectively) were the sensitive bacteria in ethyl acetate and methylene chloride extracts, respectively, whereas C. albicans was sensitive fungi in petroleum ether extract (10 mm).
There are many actions for flavonoids in medicinal field [56].Tannins are capable of inhibiting the digestive enzymes activity, and the nutritional effects of tannins are mainly linked with their interaction with protein [57].Tannin-protein complexes are insoluble, and the protein digestibility is decreased [58].Plant saponins are well known for their medicinal actions [59].Alkaloids have profound effects on diseases [60,61].

CONCLUSION
In the present study, A. judaica phytochemical analysis showed increase in the secondary compounds, and their extract showed moderate scavenging activity.The petroleum ether extract of A. judaica exhibited pronounced activity against bacteria than fungi.The methyl alcohol and ethyl acetate extracts showed inhibitory action on bacteria and fungi.The pathogen S. pyogenes was the most potent inhibitor followed by S. aureus and B. subtilis.Also, the pathogen S. aureus, E. coli and C. albicans were the most sensitive microorganism.This study revealed that A. judaica extracts could be an alternative in pharmaceutical and food preservation systems.

Figure 1 :
Figure 1: The concentration of the active constituents in mg/g dry weight for the Artimisa judaica

Figure 2 :
Figure 2: % of scavenging activity of Artimisa judaica extract and natural antioxidant catechol

Figure 3 :
Figure 3: Antimicrobial activity of a different extract of Artimisa judaica against tested pathogen

Figure 4 :
Figure 4: Percent of the antimicrobial spectrum of Artimisa judaica extracts against tested pathogen