الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
In view of continuously rising of healthcare issues, tremendously appearance of illness problems and the raising of drugs prices, the exploration of medicinal plants as potential replacements in the world of drugs holds great promise for both traditional and modern medicine. Throughout history, various cultures have relied on the healing properties of plants to treat a wide range of ailments. With advancements in scientific research, there is a renewed interest in identifying and harnessing the therapeutic potential of these natural sources. Neem as a medicinal plant has gained widespread attention due to its various bioactive compounds, particularly in the field of agriculture and medicine. The tree’s leaves, seeds, bark, and oil contain a range of bioactive compounds. When considering the interaction between neem and endophytic fungi, researchers have explored how endophytes residing in neem tissues can contribute to the production of bioactive compounds. Thus the isolation and purification of endophytic fungi producing active metabolites as antimicrobial, antioxidant and anticancer activities were the objective of our study.
Firstly, Sampling of Azadirachta indica (neem) medicinal plant from different places, (The faculty of women, Ain Shams University, Cairo; Qalyub agricultural unit, El-Qalyubia, and Nuclear Research Centre, Inshas, El-Sharkeya) was done and twenty-two endophytic fungal isolates were recovered from the leaves, bark, twigs and fruits of neem. They were found to be related to the following genera; Alternaria, Cladosporium, Penicillium, Aspergillus, Mucor, Fusarium and Trichoderma. Out of the 22 isolated endophytic fungi, the fungal isolate E3 was found to give positive results during antimicrobial activity. The E3 endophytic fungal isolate was further identified using morphological and molecular approaches based on the sequence of ITS regions and was deposited to the Gene bank as Aspergillus terreus under the accession number ON532722.1.
Optimization of the culture conditions was performed by the selected fungal strain and the results showed that the inhibition zone diameter was increased in case of the modified potato dextrose broth (MPDB) medium and glucose as carbon source. Further optimization using Response Surface Methdology (RSM) was assessed, in which Plackett-Burman statistical design was used to evaluate the important factors affecting the antimicrobial activity as four factors were showed to be significant which are temperature, medium volume, incubation period and inoculum size, Central Composite design was used to determine the optimum concentrations of the selected factors, which have a positive effect on the antimicrobial activity. The results showed that the maximum antimicrobial activity of 18.0 mm in diameter was achieved by the fungus when it was grown for ten days at 30 °C under shaking conditions (150 rpm) in the presence of glucose, KH2PO4 and MgSO4 at final concentrations of 20.0, 1.0 and 0.5 g/L, respectively. The GC-MS analysis was performed and ascertained the presence of a variety of bioactive compounds which had antimicrobial, antioxidant and anticancer activities like Orcinol (50.10%) which had antioxidant, anticancer and antifungal activities. Also FTIR was applied and exhibited many functional groups belonging to many compounds of alcohols, amines, amides, aldehydes, esters and others.
The antioxidant activity and antiproliferative activity against HEPG-2 and A-549 cell lines were assessed. The IC50 values were 85.5 ± 3.62 µg / ml, 20.74± 1.08 µg / ml and 26.89 ± 1.63 µg / ml, respectively confirming that the A. terreus ON532722 displayed significant antioxidant and anticancer activities.
In order to enhance the production of significant active compounds, the endophytic strain underwent mutagenesis through irradiation by gamma rays. Radiation induces chemical, metabolic or physiological changes to living cells and disturbs their organization. Exposure of microorganisms to sub-lethal doses of radiation produces mutations which may be desired in production of important products such as antibiotics, organic acids, amino acids, vitamins, alcohols and pigments. In the current study, the tested endophytic strain was subjected gamma radiation at different radiation doses in which the 1.5 KGy was found to give the highest antimicrobial activity GC-MS produced new active compounds in which2-Hexadecen-1-ol, 3,7,11,15-tetramethyl-, [R- [R*, R*-(E)]] (60.12 %) had antimicrobial, anti-inflammatory and anticancer effect, also FTIR had shown different important functional groups. The antioxidant activity was also improved after irradiation with 1.5 KGy where IC50 had become 37.5 ± 1.48 µg / ml. After irradiation the cytotoxicity against HEPG-2 and A-549 cell lines had IC50 values of 48.91 ± 3.85µg / ml and 30.95 ± 1.97 µg / ml, respectively.The previous results suggesting a promising alternative way for production of important bioactive compounds from neem endophytic fungi.