Molecular weight distribution of hydrolysis products during biodegradation of model macromolecules in suspended and biofilm cultures I. Bovine serum albumin

Macromolecules can comprise a significant portion of dissolved organic carbon in wastewater and affect wastewater treatability by engineered systems, but little information is available about mechanisms of macromolecule degradation. The release of protein hydrolytic fragments was monitored during the degradation of the model protein bovine serum albumin (BSA) in batch and continuous suspended cultures and in fixed-film reactor systems. Three different inocula that represent a range of bacterial diversity were used: a protein degrading isolate, a commercial biological oxygen demand (BOD) test inoculum (limited diversity culture) and a wastewater inoculum. Molecular size distributions were monitored during degradation using membrane ultrafiltration techniques. Intermediate-molecular-weight protein hydrolytic fragments (2000–10,000 amu) were produced and released into solution by all cultures in all reactor types investigated. In batch suspended culture reactors with initial BSA concentrations of 100 mg liter−1, maximum concentrations of intermediate-molecular-weight hydrolytic fragments of 25, 10 and 6 mg liter−1 were found, respectively, in pure, limited diversity and wastewater cultures. Since material in the 2000–10,000 amu fraction decreased as culture diversity increased, it was concluded that the involvement of many microbial species during protein degradation limits the accumulation of hydrolytic fragments under conditions typical of wastewater treatment systems. These results lead to the proposal of a protein degradation mechanism that features cell-bound hydrolysis of protein and the subsequent release of hydrolytic fragments back into bulk solution. Hydrolysis and release is repeated by the same or different cells until fragments are small enough (< 1000 amu) to be directly assimilated by cells. Because the concentration of protein hydrolysis fragments released in high-diversity wastewater inoculated reactors was low and they were rapidly degraded, the release of hydrolysis fragments should have little impact on overall macromolecule degradation kinetics in domestic wastewater treatment systems.