Isolation and Structure Determination of Complexed Poly(3‐hydroxyalkanoate) from Beet (Beta vulgaris L.)

Abstract

Complexed poly(3-hydroxyalkanoate)s (cPHAs), one of two types of natural PHAs, occur in both prokaryotes and eukaryotes as a complex with biomacromolecules and could be involved in various physiological functions. In this study, a cPHA-component derived from a complex with calcium polyphosphate was isolated from sugar beet (Beta vulgaris L.) and determined to be a homopolymer composed of 3-hydroxybutyrate. MALDI MS provided the number-average molecular weight (Mn = 9,124 Da) and polydispersity index (PDI = 1.01), showing that beet cPHA has a slightly lower molecular mass than the known Escherichia coli cPHA. In addition, the structural analysis of both end groups showed that (i) 100 mol-% of the carboxyl end is free, while about 30 mol-% of the hydroxyl end is free and about 70 mol-% masked and (ii) the end hydroxyl group is masked by at least six identified short-chain alkanoic and alkanedioic acids. Based on such end-group characteristics, the polymerization mechanism of beet cPHA is discussed.

Bioactive and Functional Oligomers Derived from Natural PHA and Their Synthetic Analogs

“…The detailed extraction of PHA oligomers from cells have been previously described by Reusch and Sadoff [47], which has also been modified by Seebach et al and Suzuki et al [48,49].…”

Characterization of short-chain poly3-hydroxybutyrate in baker’s yeast

“…M n corresponds to a chain length of n = 57.4. This result means that cPHA-1 is much shorter in chain length than beet cPHA (Suzuki et al, 2005). PDI is slightly broad compared with that of beet cPHA (PDI: 1.01), but is sufficiently narrow (Seebach et al, 1994).…”

“…2a, the three signals characteristic of the terminal 3-HB units having free 3-hydroxyl groups were observed at d 4.18, 3.10, and 1.22 ppm (Suzuki et al, 2005). These assignments are shown in 3-HB of Table 1.…”

“…However, Suzuki et al employed the drastic condition of a boiling CHCl 3 -MeOH (10:1) for 2 days to efficiently extract cPHA from beets, and showed that no partial methanolysis and/or no formation of the cyclic molecules via intramolecular transesterification did not occur in the purified beet cPHA. Also, it was shown that a change of extraction solvent (only CHCl 3 ) did not influence the chain length and 3-HV content (Suzuki et al, 2005). Therefore, the isolation method employed here was performed as we described previously for beet cPHA (see Experimental).…”

“…Reusch et al have isolated the native cPHApolyP complex from such cells and proposed its whole structure (Reusch and Sadoff, 1988;Seebach et al, 1994), the polyester part of which was short-chain PHB of degree of polymerization (n) = 140 with 14 mol% 3-HV comonomer. Very recently, Suzuki et al have characterized the polyP-complex-derived cPHA from the eukaryote of sugar beet (Beta vulgaris L.), designated as beet cPHA (Suzuki et al, 2005). It had the slightly shorter chain PHB of n = 106 and a much lower 3-HV content than E. coli cPHA.…”