Use of poly(ethylene glycol)s to regulate poly(3-hydroxybutyrate) molecular weight during Alcaligenes eutrophus cultivations

The ability of poly(ethylene glycol)s, PEGs, to control poly(3-hydroxybutyrate), P3HB, molecular weight in a microbial fermentation polymerization process was studied using Alcaligenes eutrophus with fructose as the sole carbon source. PEGs varying in molecular weight and end group functionality were added to the cultivation medium subsequent to cell growth, and their effects on polymer formation were evaluated. In general, A. eutrophus showed substantial tolerance for PEGs. This was illustrated by similar viable cell concentrations for the medium without PEG, 10% (w/v) PEG-IO 000 and 2% PEG-800. Furthermore, detrimental effects on polymer yields were not observed for concentrations of 5% PEG-106 and 10% PEG-10 000. The greatest reductions in molecular weight were obtained when relatively low molecular weight PEG was added to the medium. PEG-106 was most effective in that only 0.25% was required to reduce the number average molecular weight (M(n)) by 74%. The largest decrease in P3HB M(n) (from 455 000 to 19 400) was observed by adding 10% PEG-106 to the medium. The largest change in P3HB M(n) per incremental addition of PEG occurred in the 0-1% PEG concentration range. Supplementing the incubation medium with the monomethoxy ether CH3O-PEG-OH-350 and PEG-300 resulted in almost identical molecular weight reductions. However, the dimethoxy ether of tetraethylene glycol was not an effective agent for molecular weight reduction. Therefore, interaction between PEG and the PHA production system leading to molecular weight reduction was enhanced for lower molecular weight PEGs and required at least one PEG chain end functionality which may be a hydroxyl group. It is believed that PEG interacts with the A. eutrophus synthase in such a way to increase the rate of chain termination by water relative to chain propagation reactions.