الفهرس 
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Abstract
Biosurfactants or microbial surfactants are surface active biomolecules that are produced by a variety of microorganisms. Biosurfactants have a broad range of commercial applications in various industries, and in the fields of enhanced oil recovery and environmental bioremediation, also have a considerable advantage over their chemical counterparts.
In the present study:
Part I: characterization of Gemsa Company and Adabiya Suez port crude oils.
The crude oil sample from Gemsa was mainly as a paraffinic oil, while the crude oil sample from Adabiya fundamentally has high molecular weight hydrocarbons. Additionally the result proved the petrogenic pollution type of these sites.
Part II: Isolation of Bacteria Producing Biosurfactants and evaluation of the production of biosurfactants by these isolates .
The total bacterial count of oil-polluted water Gemsa sample showed count at 4.5 x 106 CFU/mL using (LB) medium. On the other hand, the total bacterial population of oil-polluted water Adabiya sample at 3 x 108 CFU/mL using (LB) medium. Additionally, the bacterial count on MSM was 6 x1010 and 9 x109 CFU/mL of oil-polluted water Adabiya and Gemsa samples after 7 days of incubation, respectively.
The bacterial strains (B1-B3) were isolated from Gemsa water sample. from which the B3 isolate demonstrated abundant growth on the culture media. On the other hand, from the bacterial isolates (B4-B9) isolated from the Adabiya water sample B4 showed abundant growth on the culture media (MSM).
The bacterial isolates (B3 and B4) were selected for high potential of biosurfactant production after 72 h.
The bacterial isolates screened from the production of biosurfactants showed the highest oil displacement test at 45 and 60 mm with B3 and B4 bacterial strains respectively. Additionally, the highest percentage of the emulsification power was obtained at 40 and 60 % with the same bacterial strains. While, the lowest surface tension at 38 and 29 mN/m was obtained by the collected of culture supernatant from bacterial isolates B3 and B4 respectively after 72 h.
Therefore, the bacterial isolates (B3 and B4) were selected for further study due to their high potential of biosurfactant production after72 h.
The most potent bacterial isolates (B3 and B4) were identified by 16S rRNA gene sequence to Bacillus flexus strain NBRC 15715 and Gelidibacter japonicus strain Bio7-1 16S.
Part III: Optimum Conditions for Enhancing of Biosurfactants Production by the selected Bacterial Strains.
The increase of the inoculum concentration from 1.0 to 5.0 % raised the bacterial count, meanwhile the best surface tension was obtained at 39 and 30 mN/m of the supernatant of B3 and B4 culture media respectively. The produced biosurfactants have high emulsification power at 42 and 55 in culture media of B3 and B4 respectively.
The measurements of surface tension confirmed that the bacterium was able to produce biosurfactants having surface tension values at 38 and 28 mN/m for B3 and B4 respectively. But the emulsification power increased to 41 and 60 % for B3 and B4 respectively with optimum pH value at 7.5.
The sucrose as a carbon source was selected for the subsequent investigations for the production of biosurfactants. The bacterial isolates B3 and B4 utilized the sucrose for production of biosurfactants, since it favored substrate for both solubilization and emulsification.
In the present study, Sodium nitrate and ammonium sulfate as well, proved to be suitable nitrogen sources that have a remarkable capacity to increase the emulsification power and decreased the surface tension with B3 and B4 bacterial strains respectively.
Additionally, some surface properties of the produced biosurfactants were determined. The biosurfactant was obtained when the bacterial isolates B3 and B4 were grown under the optimum conditions (pH 7.5, at 30 C, shaking at 150 rpm, sucrose as a carbon source, and sodium nitrate as nitrogen source in the case of B3 isolate, and use ammonium sulphate with B4 isolate). The target biosurfactant was formed after 72 h with the best surface tension and emulsification power.
Part IV: Extraction of the produced biosurfactants and their characterization.
However, classical techniques are well suited for batch recovery; they include solvent extraction, precipitation and crystallization. The biosurfactants were recovered from cultural media of (B3 and B4) at 1.1 and 1.3 g/L respectively. The evaluation and characterization by FTIR analysis of biosurfactants were performed. The Bacillus flexus and Gelidibacter japonicus proved to have their potency in the production of polymeric biosurfactants with excellent bioemulsifier properties.
Part V: Applications of the produced biosurfactants .
1. Antimicrobial activity of the produced biosurfactants.
in this work, the antimicrobial potential of the produced biosurfactants (polymeric biosurfactant) from Bacillus flexus (B3) and Gelidibacter japonicus (B4) was determined against the tested microbial strains, including Gram-positive and Gram-negative rods and cocci bacteria, yeast strain, Candida albicans, and Aspergillus niger as fungi strains were evaluated. from the results obtained, the biosurfactants have significant antibacterial activity against all bacterial species examined (Gram-positive and negative bacterial strains) and fungi strains (fungi and yeast strains).
2. Biodegradation of different petroleum oil samples using the bacterial strains B3 and B4.
The results showed that the bacterial strains were able to degrade crude oil and its maltene fractions in the range from 72 % to 96 % with Suez crude oil. In addition, the percentages biodegradation were from 50 % to 96 % with Gemsa crude oil. It was found that addition of the petroleum oil to sucrose in the MSM media for production of biosurfactants with bacterial consortium (mixture of two bacterial strains) was the best choice for attaining a higher biodegradation capacity for two different crude oils and their maltene fractions after 7days from two different crude oil samples. The Gas chromatographic (GC) analysis of the crude oils and their matlenes that remained after incubation with the bacterial isolate B3 and B4 with/ without biosurfactants were measured. The percentage concentration of the total samples (paraffins and UCM) increased from 71 to 97 % with Suez crude oil. On the other hand, the microcosm containing the bacterial consortium (mixture of two bacterial strains) and biosurfactants attained a higher biodegradation capacity for Suez crude oil and its maltene fractions after 7 days about 95 % and 97 % respectively. Moreover, the percentage concentration of the total samples increased from 49 to 95 % with Gemsa crude oil. The microcosm with bacterial consortium (mixture of two bacterial strains) and biosurfactants was obtained a higher biodegradation capacity for crude oil and its maltene fractions after 7days about 93 % and 95 % respectively.
Biosurfactants or microbial surfactants are surface active biomolecules that are produced by a variety of microorganisms. Biosurfactants have a broad range of commercial applications in various industries, and in the fields of enhanced oil recovery and environmental bioremediation, also have a considerable advantages over their chemical counterparts.
In the present study, we can conclude that:
1. Selection of (Bacillus flexus and Gelidibacter japonicus) from the other bacterial species (the nine isolates from the sea water of Suez, Egypt according to the reduction of surface tension after shorter time 18 hrs).
2. The emulsification power (E24) of broth culture favored the final selection of bacterial isolate B3 and B4 for the sequence studies in this work.
3. Studying of the optimization conditions which controlling the ability producing of the biosurfactants by the most promising BP1 shown that,
a. The best inoculum size of bacterial isolate was 1.0 × 108 CFU/ml for production of biosurfactants.
b. The best pH for growth and production of biosurfactants was fixed at 7.5.
c. The water soluble Glucose was selected as carbon source for the subsequent investigations.
d. The best nitrogen source used for growth and production of biosurfactant was ammonium sulfate.
e. The best incubation period for growth and production of biosurfactants by bacterial isolate B3 and B4 was 72 hrs.