Microbial bioremediation of olive mill wastewater

Abstract

Olive mill wastewater (OMWW) is the toxic, malodorous, recalcitrant waste from the three-phase decanter system used in the processing of olives for oil. The waste has a high organic load, and is high in polyphenols which largely account for its phytotoxicity, preventing its use in irrigation or its discharge into waterways. There have been many OMWW bioremediation studies; however, most require supplementation and sterilisation of OMWW to facilitate establishment of the bioremediating strain. The focus of this study was to discover a novel organism/s capable of reducing total phenols (TP) and phytotoxicity in unsterilised, minimally supplemented OMWW and to determine the means by which OMWW was degraded. From a collection of 223 environmental isolates that were screened for their ability to bioremediate OMWW, four fungi were identified and studied in greater detail for their ability to reduce TP, phytotoxicity and COD, and compared against the known bioremediator, Phanerochaete chrysosporium. All four performed well in unsterilised waste under the conditions used, achieving similar reductions in COD. Reductions of TP were between 65% and 82% within 15 d compared to a 62% reduction by P. chrysosporium. The isolate Cerrena consors, a white rot fungus, was of particular interest as it achieved a 75% phenol reduction within 2 h of addition of the inoculum to the OMWW, a rate which has not been previously reported in the OMWW literature. TP removal after 15 d was 80%. A 45% reduction in phytotoxicity was also noted within the 2 h period by this fungus, which increased to 84% by day 15. C. consors has not previously been reported in the bioremediation of OMWW or any other waste. After cultivation of C. consors in OMWW, the culture fluid demonstrated high laccase activity but neither MnP or LiP activity were present. This was confirmed using a syringaldazine agar overlay assay developed for use with the zymograms. Laccase was produced mainly during the stationary phase (idiophase), when carbon was limiting. The high laccase activity produced in the C. consors inoculum added to the fermentation flasks correlated with the removal of TP from OMWW. Subsequent experiments indicated that other OMWW-degrading enzymes were produced when the organism was grown on agar-solidified OMWW, but not in liquid OMWW. The laccase was shown to exist as isoforms and its isoform production varied with the type of OMWW used as the growth medium. Laccase activity produced by C. consors growing in OMWW reached 20,400 U L-1 after 30 d, indicating that this waste product represented an excellent production substrate for an industrially-relevant enzyme. When cultivated in malt extract broth laccase activity was increased 645 % within 14 d, in the presence of 0.75 mM copper. It was also reported for the first time that unspecified components in OMWW act as natural laccase mediators, since TP removal from a solution of mixed phenols was enhanced in the presence of 1% (v/v) added OMWW. The advantages offered by C. consors as a bioremediator of OMWW included its ability to grow vigorously in minimally supplemented unsterilised 50% (v/v) OMWW in the presence of a competitive background microbial population and to produce high levels of laccase and probably other as yet unknown enzyme activity capable of rapid dephenolisation and detoxification of OMWW. It was further concluded that OMWW-based medium represents an economical substrate for the production of laccase.

Date of Award	2011
Original language	English