Microbial Polyester Synthesis: Effects of Poly(ethylene glycol) on Product Composition, Repeat Unit Sequence, and End Group Structure

Poly(ethylene glycol) of Mn 200 g/mol (PEG-200) was added to cultivation media of Alcaligines eutrophus during polymer formation, and the carbon source used was 4-hydroxybutyrate. The addition of PEG-200 from 0 to 2 wt % resulted in increased 4-hydroxybutyrate (4HB) incorporation from 66 to 86 mol % into the poly(3-hydroxyalkanoate) (PHA), formed. When 4% PEG was used, the mole percent of 4HB in PHA decreased to 64 mol %. Thus, PEG in cultivation media resulted in polyester compositional changes. An unexpectedly large amount of the dry cell weight (∼10%) was found to be PEG-200 for cultivation media amended with 4% PEG-200. Analysis of this product by gel permeation chromatography (GPC) showed it to be complex, having multiple peaks. The repeat unit sequence distribution determined by 75 MHz 13C NMR spectroscopy indicated that the addition of PEG-200 to media resulted in the formation of a product with predominantly 3-hydroxybutyrate (3HB)−3HB and 4HB−4HB diads instead of random copolyesters which are formed in the absence of PEG addition to media. Two dimensional homonuclear (1H) correlated NMR analysis along with a number of control experiments indicates that PEG was covalently linked at the carboxy terminal position of PHA chains, forming natural−synthetic diblocks. Fractionation of this product into acetone soluble (AS, 84 mol % 4HB, Mn = 37 400 g/mol) and insoluble (AIS, 95 mol % 3HB, Mn = 130 000 g/mol) fractions was carried out which proved that the product formed is a mixture of polyesters differing in composition and molecular weight. Analysis of the unfractionated product as well as AS and AIS fractions by differential scanning calorimetry supported the above conclusions.