Poly[(R)-3-hydroxybutyrate] (P(3HB)) single crystals, which had different morphologies with and without chain-folding surfaces, were grown from dilute solutions of chloroform and ethanol. Two types of extracellular PHB depolymerases purified from Alcaligenes faecalis T1 and Comamonas acidovorans YM1609, defined as types I and II according to the position of the lipase box in the catalytic domain, were used in the enzymatic degradation and adsorption experiments. The enzymatic degradation of P(3HB) single crystals was investigated by means of transmission electron microscopy, atomic force microscopy, and gel-permeation chromatography. Adsorption of PHB depolymerase on P(3HB) single crystals was examined using an immuno-gold labeling technique. Enzymatic degradation of single crystals progressed from the edges and ends of crystals to yield the narrow cracks and the small crystal fragments along their long axis, independent of both surface morphologies of single crystals and types of extracellular PHB depolymerases. Lamellar thicknesses of single crystals and molecular weights of P(3HB) chains remained unchanged during the enzymatic hydrolysis. Adsorption of extracellular PHB depolymerase demonstrated a homogeneous distribution of enzyme molecules on the chain-folding surface of crystals. The above results suggest that the adsorption of enzyme increases with the mobility of P(3HB) chains of single crystals as a whole and that the endo − exo type attack by the active-site of PHB depolymerase takes place preferentially at the disordered chain-packing regions of crystal edges and ends rather than the chain-folding surfaces of single crystals, in spite of the difference in the structure of enzymes.

The relationships between the molecular weight (MW) and the chemical composition of components in copolymers and also the molecular weight distributions (MWD) of those copolymers were determined by gel permeation chromatography and compared with the calculations based on the classical theory of copolymerization kinetics. The samples were three low-conversion and one high-conversion copolymers of styrene (St) and methyl acrylate (MA) prepared by radical polymerization in bulk. Agreement between theory and experiments is satisfactory for the low-conversion samples. For the high-conversion copolymer, however, the MWD curve has a long tailing in the higher MW region and is remarkably broader than the curve calculated. Moreover, the MA content of components in the copolymer increases as the MW of the components increases, in contrast to theoretical prediction. The discrepancy between theory and experiments for the high-conversion sample may be due to the gel effect or the diffusion-controlled termination, which was neglected in the theoretical calculation.

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Enzymatic degradation of poly[(R)‐3‐hydroxybutyrate] (P(3HB)) film by the poly(hydroxybutyrate) (PHB) depolymerase from Ralstonia picketti T1 was studied in 0.01 M phosphate buffer solution (pH 7.4) at 37 °C by using a quartz crystal microbalance (QCM) technique. Enzymatic degradation of P(3HB) film was quantitatively followed by QCM as a positive frequency shift. While, the amount of depolymerases adsorbed on the film could be evaluated as a negative frequency shift by using a mutant enzyme which had no hydrolytic activity in a catalytic site. The degradation rate increased with enzyme concentration to reach a maximum value at 1.0 μg · mL−1, and then the rate decreased at higher enzyme concentration. This enzyme concentration dependence could be quantitatively explained in terms of a change of coverage of the film surface by the adsorbed enzyme. When the wild‐type enzyme solution in a QCM cell was replaced with the mutant enzyme solution in the middle of the reaction, the degradation rate was reduced markedly, indicating that the wild‐type enzyme adsorbed on the P(3HB) surface is easily substituted by the mutant enzyme in the solution. On the other hand, replacement of the wild‐type enzyme solution with other proteins or buffer solutions did not affect the degradation rate at all, suggesting that the adsorbed enzyme was not desorbed from the film surface. Thus, the adsorbed PHB depolymerase is released from the P(3HB) surface only by interaction with the same depolymerase in solution.

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