EXTRACELLULAR REDOX ACTIVITY IN ACTIVATED-SLUDGE

The tetrazolium salts triphenyltetrazolium chloride (TTC) and -iodophe nyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride (INT) have been us ed extensively for the measurement of respiratory activity in natural waters, drinking water, sediments and activated sludge. Usually this i s done spectrophotometrically after extracting the reduced formazan sa lt. Other tetrazolium salts like 5-cyano-2,3-di-4-tolyl-tetrazolium ch loride (CTC) are reduced to formazan salts which are fluorescent This property has been used to microscopically determine the proportion of metabolically active cells in a sample. The observation that CTC is al so reduced extracellularly prompted an investigation of the role of th is activity in activated sludge. In order to test the assumption that extracellular reduction is stimulated by recalcitrant molecules a benc h-scale sequencing batch reactor (SBR) was fed with synthetic wastewat er containing either lignin or amylose as well as hexanoic acid, glyce rol and sucrose as organic constituents. About 4 times as much tetrazo lium salt '-(1-[(phenylamino-)carbonyl]-3,4-tetrazolium)-bis (4-methox y-6-nitro) benzene-sulfonic acid hydrate (XTT) was reduced by extracts containing extracellular polymeric substances (EPS) produced in the p resence of lignin than by extracts derived from amylose-fed cells. The se differences are statistically significant at P > 0.05. Extracellula r redox activity in lignin-containing activated sludge accounted for a pproximately 2% of total redox activity. In addition to experiments in volving reactors fed with different substrates, the extacellular redox activity was measured microscopically using a Zeiss confocal laser mi croscope. Planktonically grown cells of Escherichia coil or activated sludge flocs were treated with CTC and the nucleic acid stain PicoGree n which fluoresces in the red and green visible light regions, respect ively. In Escherichia coil preparations, CTC-formazan crystals were fo und exclusively inside cells. By contrast, activated sludge flocs reve aled crystals both inside and outside the cells. Formaldehyde-treated flocs did not show any reduction of CTC. The extracellular crystals ac counted for up to 50% of the total CTC reduction. It follows that extr acellular redox activity is associated with the production of EPS. Its role may be to help microbial systems to deal with organic materials which are difficult to utilize. (C) 1998 IAWQ. Published by Elsevier S cience Ltd.