fed with only 10 g/L jatropha oil as the carbon source showed to be able to
synthesize P(3HB-co-3 mol% 3HHx). Up to 12.5 g/L of jatropha oil was found to
be safe to be fed, and beyond that, it exerts toxicity to cells. On the other hand,
R. eutropha wild-type strain fed with a mixture of jatropha oil with 3HV precursors
could synthesize P(3HB-co-3HV) with 3HV fraction of 41 mol%. The results
discussed here using promising plant oils are tabulated in Table 3.2.
A fatty acid is a structurally related carbon source, and it is pivotal in synthesizing
PHA especially mcl-PHA using microorganisms. Generally, fatty acid will exert a
certain level of toxicity to microbes, thus utilizing efficient feeding strategy in terms
of the concentration, and feeding time will give a better output. It was shown that
intermittent feeding of low concentration of sodium decanoate (C10Na) or sodium
dodecanoate (C12Na) along with the low concentration of glucose, glycerol, or
xylose yielded better PHA production in E. coli strain with defective β-oxidation
(Fadzil et al. 2018). The efficient intermittent feeding strategy proposed in this study
has enabled the engineered E. coli to have better conversion yield of C10Na and
C12Na to P(3HD) and P(3HDD), respectively. A near homopolymer of 5.44 g/L P
(3HD) and 3.50 g/L P(3HDD) was obtained.
The addition of amino acid also plays an interesting role in PHA biosynthesis. For
instance, an obligate anaerobic bacterium Clostridium difficile can convert leucine to
4-methylvalerate (Saika et al. 2014). In the conversion process, 4-methyl-2pentenoyl-CoA (4M2PE-CoA) is intermediate, and this can also be converted to
3-hydroxy-4-methylvalerate (3H4MV) and eventually channeled for the synthesis of
PHA copolymer with 3H4MV unit (Table 3.3) (Saika et al. 2014). Besides, it has
confirmed that amino acid is a promising precursor for the synthesis of 2HA with
various side-chain structures when related enzymes are used to convert amino acids
to 2HA-CoA as the substrate for the lactate-polymerizing enzyme (LPE). For
supplementation of 1 g/L of valine, the fraction of 2H3MB, which has the same
carbon backbone as valine does, increased to 8.3 mol%. Meanwhile, for 1 g/L of
leucine supplementation, the fraction of 2-hydroxy-4-methylvalerate (2H4MV),
which has the same carbon backbone as leucine does (Fig. 3.2), was detected in
the amount of 23.6 mol%. On the other hand, phenylalanine supplementation at 1 g/
L increased the 2-hydroxy-3-phenylpropionate (2H3PhP) fraction to 17.2 mol%.
After the increase in the phenylalanine supplementation to 5 g/L, the 2H3PhP
fraction slightly increased to 20 mol% (Mizuno et al. 2018). However, usually, a
supply of amino acid can hamper cell growth due to the end product feedback
inhibition, and thus, the concentration of amino acid has to be adjusted for desirable
cell growth and PHA production.
Apart from all these carbon sources, agricultural and domestic waste materials also
were tested for PHA production. Such waste materials are rich in xylose, but
PHA-producing microorganisms are generally weak in assimilating them. However,
via anaerobic fermentation of xylose which is easier to assimilate, lactic acid and acetic
acid can be produced; thus, they can be utilized for PHA production (Tsuge 2002).
Rice husk is one of the notable agricultural wastes and it is rich in cellulose and
hemicellulose. The richness of rice husk has enabled it to be a potential carbon source
in PHA production. B.cepacia USM was shown to have a dry cell weight of 7.8 g/L
and PHA content of 50 wt% when it was cultivated in a 5 L fermenter supplemented
66
R. Sivashankari and T. Tsuge
synthesize P(3HB-co-3 mol% 3HHx). Up to 12.5 g/L of jatropha oil was found to
be safe to be fed, and beyond that, it exerts toxicity to cells. On the other hand,
R. eutropha wild-type strain fed with a mixture of jatropha oil with 3HV precursors
could synthesize P(3HB-co-3HV) with 3HV fraction of 41 mol%. The results
discussed here using promising plant oils are tabulated in Table 3.2.
A fatty acid is a structurally related carbon source, and it is pivotal in synthesizing
PHA especially mcl-PHA using microorganisms. Generally, fatty acid will exert a
certain level of toxicity to microbes, thus utilizing efficient feeding strategy in terms
of the concentration, and feeding time will give a better output. It was shown that
intermittent feeding of low concentration of sodium decanoate (C10Na) or sodium
dodecanoate (C12Na) along with the low concentration of glucose, glycerol, or
xylose yielded better PHA production in E. coli strain with defective β-oxidation
(Fadzil et al. 2018). The efficient intermittent feeding strategy proposed in this study
has enabled the engineered E. coli to have better conversion yield of C10Na and
C12Na to P(3HD) and P(3HDD), respectively. A near homopolymer of 5.44 g/L P
(3HD) and 3.50 g/L P(3HDD) was obtained.
The addition of amino acid also plays an interesting role in PHA biosynthesis. For
instance, an obligate anaerobic bacterium Clostridium difficile can convert leucine to
4-methylvalerate (Saika et al. 2014). In the conversion process, 4-methyl-2pentenoyl-CoA (4M2PE-CoA) is intermediate, and this can also be converted to
3-hydroxy-4-methylvalerate (3H4MV) and eventually channeled for the synthesis of
PHA copolymer with 3H4MV unit (Table 3.3) (Saika et al. 2014). Besides, it has
confirmed that amino acid is a promising precursor for the synthesis of 2HA with
various side-chain structures when related enzymes are used to convert amino acids
to 2HA-CoA as the substrate for the lactate-polymerizing enzyme (LPE). For
supplementation of 1 g/L of valine, the fraction of 2H3MB, which has the same
carbon backbone as valine does, increased to 8.3 mol%. Meanwhile, for 1 g/L of
leucine supplementation, the fraction of 2-hydroxy-4-methylvalerate (2H4MV),
which has the same carbon backbone as leucine does (Fig. 3.2), was detected in
the amount of 23.6 mol%. On the other hand, phenylalanine supplementation at 1 g/
L increased the 2-hydroxy-3-phenylpropionate (2H3PhP) fraction to 17.2 mol%.
After the increase in the phenylalanine supplementation to 5 g/L, the 2H3PhP
fraction slightly increased to 20 mol% (Mizuno et al. 2018). However, usually, a
supply of amino acid can hamper cell growth due to the end product feedback
inhibition, and thus, the concentration of amino acid has to be adjusted for desirable
cell growth and PHA production.
Apart from all these carbon sources, agricultural and domestic waste materials also
were tested for PHA production. Such waste materials are rich in xylose, but
PHA-producing microorganisms are generally weak in assimilating them. However,
via anaerobic fermentation of xylose which is easier to assimilate, lactic acid and acetic
acid can be produced; thus, they can be utilized for PHA production (Tsuge 2002).
Rice husk is one of the notable agricultural wastes and it is rich in cellulose and
hemicellulose. The richness of rice husk has enabled it to be a potential carbon source
in PHA production. B.cepacia USM was shown to have a dry cell weight of 7.8 g/L
and PHA content of 50 wt% when it was cultivated in a 5 L fermenter supplemented
66
R. Sivashankari and T. Tsuge