Modification of Polyhydroxyalkanoates (PHAs) - Polyhydroxyalkanoate (PHA) Based Blends, Composites and Nanocomposites - page 165

two-fold increase in the chondrocytes' attachment after two hours of incu-
bation when electrospun nanofiber was used as compared to the cast film. 123
Yu et al. 121 observed a decrease in the crystallinity and crystallization rate by
doping zinc oxide (ZnO)/PHBV electrospun nanofiber with ZnO nano-
particles. They attributed the observed effect to the interaction between the
hydroxyl groups on the ZnO nanoparticles' surface and the polymeric carb-
oxylic groups. Ying et al. 124 evaluated the biocompatibility and biosorption
characteristics of the electrospun scaffold of P3HB4HB through subcutane-
ous implantation of the fibers in rats. The researchers found a highly in-
creased tissue response with increasing content of 4HB monomer.

d n 2 r 4 n g | 1

7.4 Modification of PHAs with Enzymes
PHA modification via an enzyme-mediated process is seen as a mild, specific
and environmentally-friendly method. In this section, PHA modification
using enzymatic degradation and/or synthesis methods in both in vivo and
in vitro processes is discussed. Also included in the discussion is enzymatic
modification of PHA using the degradation products of PHA itself.

7.4.1 In Vivo Enzymatic Degradation of PHA
During the bacterial fermentation process, PHA granules are accumulated
inside the cells in the presence of excess carbon source(s) but limited es-
sential nutrients such as nitrogen, oxygen, phosphorous, potassium, or
sulfur. 125 In a reverse situation where the microbes are starved of a carbon
source in the presence of abundant nutrients, they will start producing
intracellular PHA depolymerase and dimer hydrolase to degrade the accu-
mulated PHA and continue growing. 126 The monomer produced from the
intracellular degradation is subsequently oxidized by the cell to form acet-
oacetate. In an attempt to bypass the accumulated PHA hydrolysis,
Lee et al. 127 proposed a process that incorporates continuous limitation of
the carbon source and nutrient(s) in an anaerobic condition. The absence of
oxygen inhibits the cells from metabolizing the 3HB monomers from
intracellular PHB degradation by lowering the concentration of R-3-hydro-
xybutyrate dehydrogenase, which in turn minimizes the conversion of 3HB
to acetoacetate.
It has been reported that a high yield of 3HB (96%) can be obtained within
a relatively short time (30 min) in Alcaligenes latus by using a fed-batch
culture system with sucrose as a carbon source. The cells with stored PHB
were collected and incubated at pH 4 and 37 1C to provide an environmental
condition in which cells exhibit high activity of intracellular PHA depoly-
merase and low activity of (R)-(-)-3-hydroxybutyric acid dehydrogenase. 127
Other identified factors that contribute in in vivo depolymerization are
substrate concentration and extracellular pH. Ren et al. 128 reported that the
optimal initial pH range for initiation of intracellular depolymerization of
PHA by Pseudomonas putida was 8-11, and pH 11 after commencing

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