Date of Award

1990

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biological Science

First Advisor

Dr. Taxler

Abstract

A microorganism capable of synthesizing biosurfactant from cheap and renewable water-soluble substrate such as carbohydrate was isolated' from the environment in our laboratory. It was selected from other isolates by screening for its ability to reduce the tensiometric properties of the culture broth. The isolated organism (JIZAN-1) produced significant extracellular surfactant activity during growth in batch culture on modified mineral salts medium (MMSM) containing 0.2% glucose as a carbon source and 0.02% yeast extract as a growth cofactor. This was demonstrated by lowering of the whole broth surface tension to less than 27.0 dynes/cm and the interfacial tension to less than 2.0 dynes/cm.

The isolated organism (JIZAN-1) was identified based on morphological, cultural and biochemical tests as an Arthrobacter species. Optimum culture conditions such as the amount of glucose as the carbon Source, amount of NH4NO3 or NH4C1 as the nitrogen Source, and the environmental conditions including initial medium pH and incubation temperature, for growth and yield of biosurfactant were determined through batch cultivation on MMSM.

Arthrobacter JIZAN-1 was used for the development of a continuous process for biosurfactant production under carbon limitation. The goal of such a process was to produce the surfactant in a cost efficient process.

Biosurfactant recovery procedure consisted of repeated extraction of the culture broth by chloroform at 35°C. The chloroform extract was run in various solvent systems on silica gel TLC plates to determine the mobility of the components in the chloroform extract. The first purification step consisted of fractionating the crude chloroform extract by silica gel open column, then the surfactant #1 which was obtained from the first step was further purified through fractionation by C18 silica (reverse phase) open column chromatography and the purified surface active compound was designated AL-Hazmi's surfactant.

After the purity of AL-Hazmi's surfactant was confirmed by TLC, followed by detection with sugar and lipid specific reagents, the structure was examined by chemical and physical methods. When AL-Hazmi's surfactant was hydrolyzed under alkaline and acidic conditions, the fatty acids released were determined by GC and the water soluble portion contents of sugars and sugar derivatives were determined by HPLC. The structure was also examined by means of 1H and 13C nuclear magentic resonance spectroscopy (NMR) and by infrared spectrophotometry (IR). The surfactant is characterized as a complex glycolipid consisting of a number of homologs from C10 to C20 saturated and unsaturated fatty acids including α-branched-β-hydroxy fatty acids. The water soluble portion obtained after acid hydrolysis yielded raffinose, galacturonic acid and four unidentified peaks.

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