الفهرس 
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Abstract
There is a growing concern about the rate of oil spills in water currently experienced throughout the world today, much of it arising from the growing production and use of fossil fuels. In all countries, oil exploration and use threatens the health of the environment and living creatures including humans. An oil spill is the release of petroleum hydrocarbon into the environment. Oil spills may be due to the release of crude oil from tankers, off shore platforms, drilling rig, sand wells as well as spills of refined petroleum products ( such as gasoline, diesel oil) and their by-products, heavier fuels used by large ships, or the spill of only oily refuse or waste oil.
The study aims at treating oil spills in Fresh and Marine water by biological method.
Isolation of biosurfactant producing bacteria from fresh water, two fresh water samples, as well as sea water ones. One from Ismailia canal near mustard and the other one a little bit far by few meters. Eight microorganisms were isolated from the polluted samples with oil spills.
For the isolation of biosurfactant producing bacteria from sea water, one sea water sample polluted with oil spills were collected from Al Suez, near Al-Suez harbor. Six microorganisms were isolated and the purification procedure of crude oil utilizing Bacterial isolates was carried out by agar streak method. Determination of morphology of the isolates, count, forms of them was done.
Gram Staining was done that a smear is formed from each isolate. Growth on MacConkey Agar Medium was done, results were nine isolates are gram positive and five isolates are gram negative. All isolates were streaked on MacConky agar medium which is a selective and differential medium designed to isolate and differentiate enterics based on their ability to ferment lactose, results showed that five are gram negative and nine are gram positive. DROP Collapse Assay was done to assess the surfactant activity, results were all of them positive DROP collapse test. The collapse of oil DROP has been visualized and the less time taken indicates the higher activity of surfactant. Emulsification Index (E24%) was done by The emulsion index was calculated after 24 hours; hence it is also called E24. Emulsion index was measured, results were that six isolates gave emulsion.
Hemolytic Activity was done that a pure culture of each bacterial isolate was streaked on the freshly prepared blood agar and incubated at 37 oC for 48–72 Hrs. Results were recorded based on the type of clear zone observed, results showed seven isolates gave gamma hemolytic activity and seven isolates gave beta hemolytic activity.
Oil Spreading Method was done results were that the isolates gave oil spreading from 2cm to 7.5 cm.
Fourier Transform Infrared Spectroscopy was made that the biosurfactant extract recovered from the supernatant of the bacterial isolate was characterized FTIR spectra Surface Tension was done that the surface tension of the culture supernatants was measured using Theta Lite - surface tensiometer - Biolin Scientific giving results to the chemical composition of six isolates’ biosurfactants and how it reduce surface tension. Biosurfactant Stability Tests were done that Stability studies (optimization) were carried. The stability of the biosurfactants against pH, temperature, salinity was determined as following with some modifications, results were that the highest OD and E24 at temperature ranges from 20oC to 40oC and PH range from 6-7 and salinity range from 3% to 5 %.
Three isolates named O3, O11, H2 which were identified by biologists as Bacillus subtilis, Bacillus licheniformis, and Bacillus licheniformis (isolated from sea water) respectively, that were isolated from petroleum hydrocarbon contaminated fresh and sea water gave a high emulsifying activity 71%, 65% and 62% respectively.
Also, the biosurfactant was characterized by Fourier transform infrared spectroscopy (FTIR) and surface tension measurement. FTIR showed the production of biosurfactant from Bacillus subtilis similar to surfactin, while the biosurfactant from two isolates Bacillus licheniformis (one from fresh contaminated water and the other from contaminated sea water with oil spill) similar to lichenysins. The use of microbial biosurfactants significantly decreases the hydrophobicity and increases the rate of hydrocarbon biodegradation. Biosurfactants obtain demonstrated good surface tension reduction capacity by Bacillus subtilis and Bacillus licheniformis, and Bacillus licheniformis (isolated from sea water) up to 30, 36, and 35 Nm/m. we obtained a high yield of biosurfactants and bioemulsifiers at the same time, so we can use that in bioremediation of oil spill in both fresh and sea water.
CONCLUSION:
The Bacillus subtilis and Bacillus lichenformis strains, isolated from fresh water and sea water contaminated with oil spill, are capable of producing yield of biosurfactants and bioemulsfiers. The preliminary chemical characterization revealed that they belong to the lipopeptides. Bacillus subtilis strain are capable of producing a biosurfactant similar to surfactin and Bacillus lichenformis strains are capable of producing a biosurfactant similar to lichenysins. In terms of the surface activities, the crude biosurfactants showed comparable physicochemical properties.. Indeed, they are capable of forming stable emulsions with a variety of hydrophobic compounds, and their activities are not affected by exposure to exaggerated environmental conditions. The features of these biosurfactants and bioemulsfiers make them an interesting biotechnological product for many environmental and industrial applications such as bioremediation processes and Microbial enhanced oil recovery.
RECOMMENDATIONS:
Growing of bacterial strains that produces yield of biosurfactants and bioemulsifiers in situ to be used ex situ in bioremediation process.
Trying to Produce biosurfactants from bacterial strains on the industrial level to be used in bioremediation of oil spill in water.