A new process has been developed to reduce excess sludge production, in which both excess sludge digestion and wastewater treatment are conducted simultaneously in the same aeration tank. The ozonation enhances biological degradation of the activated sludge, which is decomposed in a subsequent biological treatment. A considerable amount of biomass is mineralized biologically in proportion to the amount of recirculated biomass from the ozonation stage to the biological stage. It was observed that the amount of excess sludge is reduced to nearly zero when 1.2 kg/m3-aeration tank volume of biomass is recirculated in a day from the biological stage to the ozonation stage at a BOD loading of 1.0 kg/m3/d. A biomass concentration of 4200 mg/L was maintained at 1.0 kg-BOD/m3/d without drawing excess sludge for 6 weeks of experimental period under ozone dose of 0.05 g-O3/g-SS and recirculation rate at 0.3 d−1. Only a limited difference in the effluent quality was observed between the new process and the conventional activated sludge process.

An activated sludge process which produces no excess sludge was developed. The process is very simple as a small amount of return sludge is ozonated and then returned to the aeration tank. The ozonation enhances biodegradability of activated sludge, which is biologically oxidized in the aeration tank. A full-scale plant for treating 450m3/d of municipal wastewater was constructed and has been operated successfully for 9 months. The amount of excess sludge eliminated is directly proportional to the amount of ozone dosed to the sludge. At the ozone dosing rate of 0.034 kg/kg-SS, complete elimination of excess sludge has been achieved when 4 times more amount of sludge is ozonated than that of the excess sludge expected in the treatment without ozonation. After 5 months of operation without any withdrawal of excess sludge, small amount of inorganic substances like sand and silt accumulated in the sludge. On the other hand, inert organic substances does not seem to accumulate. As for effluent quality, BOD and nitrogen were kept good. Although effluent SS was 2–15 mg/l higher compared to a control without ozonation, it has been well below the discharge limit.

A detailed study was conducted on biological removal of phosphorus by an activated sludge process composed of an anaerobic stage followed by an aerobic stage in order to clarify the role of the anaerobic stage of the process. Two distinctive sludges, one contained approximately 10 percent phosphorus, most of which existed in the form of polyphosphates, and the other contained 1.9 percent phosphorus, were obtained by changing BOD concentration of influent and hydraulic detention time of the process. Although the polyphosphate-containing sludge released phosphorus in proportion to the absorbed BOD at anaerobic conditions, the sludge which did not contain polyphosphates also absorbed an equal amount of BOD at the identical rate without any external sources of oxygen. Both sludges accumulated identical levels of poly-β-hydroxybutyrate within the sludges as the BOD sources were absorbed. These results suggest that the polyphosphate-containing sludges have no advantage over the sludges with no polyphosphates when they uptake BOD in the anaerobic stage.

Protein-encapsulated fibermats are an attractive platform for protein-based bioactive materials. However, the choice of methods is still limited and not applicable to a wide range of proteins. In this study, we studied new polymeric materials for constructing protein-encapsulated fibermats, in which protein molecules are encapsulated within the nanofibers of fibermats without causing deleterious changes to protein structure or function. We constructed a protein-encapsulated fibermat using the poly(γ-glutamate) (PGA)/(3-glycidyloxypropyl)-trimethoxysilane (GPTMS) hybrid as a precursor for electrospinning. Because the PGA/GPTMS hybrid is water-soluble, protein molecules can be added to the precursor in an aqueous solution, significantly enhancing protein stability. Polycondensation during electrospinning (in-flight polycondensation) makes the obtained fibermats water-insoluble, which stabilizes the fibermat structure such that it is resistant to degradation in aqueous buffer. The molecular structure of the PGA/GPTMS hybrid gives rise to unique molecular permeability, which alters the selectivity and specificity of biochemical reactions involving the encapsulated enzymes; lower molecular-weight (MW) substrates can permeate the nanofibers, promoting enzyme activity, but higher MW substrates such as inhibitor peptides cannot permeate the nanofibers, suppressing enzyme activity. We present an effective method of encapsulating bioactive molecules while maintaining their structure and function, increasing the versatility of electrospun fibermats for constructing various bioactive materials.

Peptide gemini-surfactant (PG-surfactant), a kind of lipopeptide, is composed of a short linker peptide (X) between two alkyl-chain-modified Cys residues and peripheral peptides at the N-terminal (Y) and the C-terminal (Z) sides, respectively, of the alkylated Cys residues. In this study, we developed and examined a series of PG-surfactants containing two C12 saturated alkanes and oligo-Lys, arranged at the X-, Y-, or Z-positions. To arrange oligo-Lys at the Y- or Z-positions, a repeat sequence of -Asp-Lys-Asp-Lys- was used at the X-position. All of the PG-surfactants exhibited high antimicrobial activity against both Gram-positive and -negative bacteria. In addition to high antimicrobial activity, a low hemolysis activity is prerequisite for efficient intravenous administration. Among the synthesized PG-surfactants, those having -(Lys)3- at the Y- or Z-positions, i.e. K 3 -DKDKC 12 and DKDKC 12 K 3 , showed reasonably low hemolytic activities. This combination of high antimicrobial activity along with low hemolytic activity is an essential and unique property and has not been previously reported for the synthesized lipopeptides. Further, using scanning electron microscopy (SEM) and N-phenyl-1-naphthylamine (NPN) uptake assay we showed that the antimicrobial activity of these PG-surfactants may be attributed to membrane disruptive mechanisms. Although the PG-surfactants with low hemolytic activity could interact and localize onto red blood cell surfaces and cause slight expansion of cell morphologies, no subsequent penetration occurred. In summary, we describe here the successful development of PG-surfactants having high antibacterial and low hemolytic activity, thus providing a significant molecular platform to develop novel antimicrobial agents.