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Thermally reversible physical gels of poly[(R)-3-hydroxybutyrate- co -(R)-3-

hydroxyhexanoate]: Part 1 gelation in dimethylformamide

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DOI: 10.1016/j.polymer.2017.10.015

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 Polymer 131 (2017) 217e223

Contents lists available at ScienceDirect

Polymer
journal homepage: www.elsevier.com/locate/polymer

Thermally reversible physical gels of poly[(R)-3-hydroxybutyrate-co-

(R)-3-hydroxyhexanoate]: Part 1 gelation in dimethylformamide
Brian J. Sobieski a, b, Liang Gong a, c, Steven R. Aubuchon a, d, Isao Noda a, e,
D. Bruce Chase a, John F. Rabolt a, *
a
Department of Materials Science and Engineering, University of Delaware, Newark, DE 19716, USA
b
FXI Inc., Aston, PA 19014, USA
c
3M, St. Paul, MN 55144, USA
d
TA Instruments, New Castle, DE 19720, USA
e
Danimer Scientific, Bainbridge, GA 39817, USA

a r t i c l e i n f o a b s t r a c t

Article history: A series of thermally reversible gels made from a solution of bio-based, biodegradable poly[(R)-3-
Received 25 July 2017 hydroxybutyrate-co-(R)-3-hydroxyhexanoate] (PHBHx) in dimethylformamide (DMF) were analyzed
Received in revised form using wide angle X-ray diffraction (WAXD), Fourier transform infrared spectroscopy (FTIR), and scanning
1 October 2017
electron microscopy (SEM). WAXD results demonstrate the existence of crystalline PHBHx in the polymer
Accepted 7 October 2017
gel by the presence of diffraction peaks characteristic of the a crystalline form. The results obtained from
Available online 8 October 2017
FTIR are consistent with the WAXD, all indicating that gelation is caused by the presence of polymer
crystals within the gel system. SEM images confirmed the phase separation of the solvent and the
Keywords:
P(HB-co-HHx) copolymers
polymer by revealing a highly interconnected, submicron-scale polymer network comprised of PHBHx
Polymer gels ribbons and fibrils.
Infrared spectroscopy © 2017 Published by Elsevier Ltd.
X-ray diffraction

1. Introduction
The interactions between polymer chains and solvents have
been and continue to be a complex and extremely important topic
for understanding solution behavior and properties, especially in
the case of polymer gel systems. Gelation is of key importance to
many industries, including pharmaceuticals and food, and is uti-
lized for controlled release and surface coating [1]. Additionally, the
high surface area of the cross-linked (either chemical or physical)
network inherent in the polymer gel state provides enhanced
function when used as a catalyst support, tissue growth scaffolding,
or even as a filtration medium. Polymeric gelation falls into two
categories: one which relies on chemical interactions and another
which is based primarily on physical interactions [1,2]. The first
requires a polymer to undergo a chemical reaction which cova-
lently links the polymer chains, forming a bound, interconnected
network. Formation of such a polymer gel occurs when a single
chain spans the entire system, yielding a macromolecule with
essentially infinite molecular weight. The resulting network
* Corresponding author.
E-mail address: rabolt@udel.edu (J.F. Rabolt).
becomes insoluble but can swell when a good solvent is introduced.
In contrast, physical gelation requires no chemical reaction and is
the product of non-covalently bonded chain-chain and solvent-
chain interactions.
Physical gelation has previously been reported for a variety of
biopolymers and synthetic polymers [1e27]. Gelatin and poly-
saccharide agar are two biopolymer gels widely used in the food
industry and as a cell culture medium, respectively [1]. Polystyrene
(PS) of varying tacticity forms physical gels in cyclohexanol, 1-
dodecanol, benzene, chloroform, decalin and toluene [2e5]. Pra-
sad et al. found that PS gelation is dependent on the tacticity of the
polymer and location of the solution concentration within the
polymer-solvent phase diagram. Within the coexistence curve,
gelation of isotactic PS occurred through liquid-liquid phase sepa-
ration, followed by crystallization. Outside of the coexistence curve,
the morphology suggested a different gelation method, though no
theory was proposed [3]. In two studies by Daniel et al., it was re-
ported that gelation of syndiotactic PS in chloroform, benzene, and
toluene occurs via stabilization of a nearly helical conformation of
the PS chain through interactions with the solvent, possibly fitting
into the voids between phenyl groups in the polymer [4,5]. A
separate set of studies indicate that gelation of atactic PS in carbon

https://doi.org/10.1016/j.polymer.2017.10.015
0032-3861/© 2017 Published by Elsevier Ltd.
218 B.J. Sobieski et al. / Polymer 131 (2017) 217e223
disulfide (CS2) is due to organization of highly syndiotatic se-
quences of the polymer chain using neutron scattering [6e10].
Furthermore, gelation of atactic PS in CS2 occurred around room
temperature for highly concentrated polymer solutions, which
suggests that the conclusions of Prasad et al. that gelation occurs
due to liquid-liquid phase separation may be incorrect. It was also
shown, as in Daniel et al., that gelation of isotactic PS results from
the formation of a complex between the polymer and the solvent.
Poly(vinylidene fluoride) (PVDF) also forms physical gels in
aliphatic ketones, cyclic ketones, aromatic diesters, and other
organic solvents [11e14]. Each study revealed that PVDF formed
crystalline regions within the gel system and that the crystalline
polymorph could be controlled by the solvent used. In solutions of
monoesters and diesters, poly(vinyl chloride) (PVC) forms physical
gels through crystallite formation of the polymer or solvent-
polymer complexes [15e17]. The results indicate a possibility that
specific interactions between solvent and polymer, such as
hydrogen bonding, stabilize the crystalline polymer within the
solvent.
Poly(methyl methacrylate) (PMMA), undergoes physical gela-
tion in the presence of bromobenzene, o-xylene, chlorobenzene,
and toluene [18e20]. PMMA gels appear to be one of the more
complex systems due to the formation of a double-helix structure
where the chains of different tacticity wrap around one another.
These structures, called stereocomplexes, form the linkages be-
tween PMMA chains, causing physical gelation to occur. Physical
gelation has also been reported for random copolymers of ethylene
and propylene [21]. When these copolymers were dissolved in
toluene and methylcylohexane they arranged into a fiber-like
structure within the solution. There is some indication that crys-
tallization of the ethylene segments yields the linkages for gelation,
though no theory was directly proposed or supported. Another
study on polyethylene with varying degrees of chlorination was
performed by Tan et al. to determine the relationship between bulk
polymer crystallinity and gelation in toluene and o-dichloroben-
zene [22]. The study revealed that the noncrystalline polymers
formed gels whose sol-gel and gel-sol transition temperatures were
equal and that gel formation occurred through chain association. In
the crystalline polymers, however, there was a difference of
10e20
C between the transitions implying that the polymer
network was stabilized through the formation of polymer
crystallites.
Physical gelation has also been reported for a class of bio-based,
biodegradable polymers, poly(hydroxyalkanoates) (PHAs) [23,24].
PHAs are chiral, aliphatic polyesters produced by a variety of bac-
teria which incorporate these polymers as granules to store carbon
and energy [25e27]. The homopolymer from this class of polyesters
is poly(hydroxybutyrate) (PHB) which has received extensive study
over the last few decades due to its biocompatibility and potential
to replace petroleum-base thermoplastics. PHB is known to crys-
tallize primarily as an a-form with polymer backbones adopting a
21 helical conformation which arranges into an orthorhombic unit
cell with a ¼ 5.76 Å, b ¼ 13.2 Å, and (fiber axis) c ¼ 5.96 Å [31,32].
However, as a result of this polymer's relatively high crystallinity
(>70%) [27] and the proximity of its melting point to its thermal
degradation temperature, the applicability of this material is
limited for use as a thermoplastic. In order to generate a polymer
with improved functional material properties, PHB has been
copolymerized by incorporating a small amount of a co-monomer,
3-hydroxyhexanoate (3HHx), shown in Fig. 1.
3HHx monomer units are similar to 3HB except that they
contain a longer side chain; a propyl rather than a methyl group.
The propyl group is what yields superior material properties in
terms of ductility and strength by interrupting the crystallization of
PHB segments. These branches along the polymer backbone act as
Fig. 1. Chemical structure of PHBHx, where the left monomer is hydroxybutyrate and
the right monomer 3-hydroxyhexanoate.
defects within the highly regular PHB sections, thus lowering the
crystallinity and melting point of the polymer [28]. Gelation of PHB
in dimethylformamide was reported by Turchetto et al. who sug-
gested causation through crystalline PHB [23]. However, the only
evidence of crystalline PHB revealed by this study was the presence
of melting endotherms and exothermic peaks on cooling the gel
samples. A separate study by Li et al. demonstrated physical gela-
tion of PHB and blends of PHB/PHBHx of 12 mol % 3HHx, as well as
blends of PHB/P(3HB-4HB) in a mixture of chloroform and dioxane
[24]. The focus of the article was on the use of these polymer net-
works as a scaffold for cell growth, rather than the determination of
the cause of gelation or the material properties of the system.
The focus of the current study is to understand the cause of
gelation in PHBHx random copolymers and to determine the
morphology and source of the gel system. Characterization of the
morphology and conformation of the PHBHx in the gel state was
accomplished using scanning electron microscopy (SEM) and
Fourier transform infrared spectroscopy (FTIR). The combined re-
sults are highly suggestive that crystalline PHBHx domains linked
by amorphous chains serve as the linkage forming the gel state in
these systems. Wide angle X-ray diffraction results confirmed this
hypothesis.
2. Experimental
2.1. Materials
Poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate]
(PHBHx) copolymers with 3.9, 5.8, 9.4, and 13 mol % 3HHx como-
nomer contents were supplied by the Procter & Gamble Company,
Cincinnati, Ohio with no further purification. The weight average
molecular weight of the copolymers with 3.9, 5.8, 9.4 and 13 mol %
3HHx content were 843, 1174, 455, and 792 kg/mole, respectively.
Dimethylformamide (DMF) and chloroform were purchased from
Sigma-Aldrich Co., Ltd and used without further purification.
2.2. Sample preparation
Polymer solutions of 2, 3, 4, 5, 7, 10, and 15 wt % PHBHx of each
comonomer content in DMF were prepared by dissolving PHBHx in
DMF at 85
C in a silicon oil bath with constant stirring for
approximately 30 min followed by quenching to 20
C in a water
bath. The resulting solutions were kept at room temperature, un-
disturbed for at least 1 week before any analysis was performed to
ensure gelation had occurred. An additional two sets of gel solu-
tions of 2, 5, 7, 10, 12, and 15 wt % PHBHx of 3.9, 5.8, 9.4 and 13 mol %
3HHx content in DMF were prepared using the same preparation
conditions except one set was slowly cooled to room temperature
and the other was quenched to 25
C in an ethanol bath. Slow
cooling was performed by leaving the samples in the silicon oil as
the oil returned to room temperature. Polymer films were prepared
 B.J. Sobieski et al. / Polymer 131 (2017) 217e223
by solution casting using 10 wt % PHBHx in chloroform. The films
were dried under vacuum for 4 h before annealing in a convection
oven at 70
C for 4 h.
2.3. Wide angle X-ray diffraction (WAXD)
The wide angle X-ray diffraction profile of the polymer gel was
measured using a Bruker D8 XRD with a Cu tube source operated at
40 kV and 40 mA producing a 1.5418 Å X-ray beam collected uti-
lizing the LYNXEYE-XE detector in OD mode. WAXD profiles were
recorded over a 2q range of 10e30
with a 0.025 2q increment and
each point collected for 1 s. A gel sample was prepared for analysis
by placing a section of gel on an amorphous glass slide, forming it
into a line across the slide, and covering and sealing with com-
mercial plastic wrap to prevent solvent evaporation. The plastic
wrap was insoluble in the solvent used, i.e. DMF. Data collection
was cycled and co-added until reasonable signal to noise level was
achieved. The same procedure was performed on a clean slide
covered in plastic wrap to confirm that the diffraction peaks arose
from the sample and neither the plastic wrap nor the slide
contributed to the scattered intensity.
2.4. Solvent exchange and lyophilization of polymer gels
Solvent exchange to water, followed by freeze drying of the
polymer gels, was implemented to generate samples that were
viable for imaging in the SEM in order to obtain morphological
information. Solvent exchange was performed by “quenching” a
portion of the gel in a larger volume of non-solvent, i.e., water,
stirring constantly for 10 min, replacing the water with fresh DI
water and repeating for a total of 3 exchanges. This treatment was
followed by an overnight exchange in fresh DI water with constant
stirring, and replaced one last time with fresh DI water the
following day. Lyophilization of the exchanged polymer gels was
performed using a Millrock Technology bench top freeze drier with
3Gen microprocessor control by freezing the samples in liquid ni-
trogen before adding to the sample chamber. The lyophilization
process was determined to be complete when the pressure of the
system was below 14 mTorr. This drying process was carried out to
protect the delicate morphology of the polymer gel and avoid
collapse of the microstructure.
2.5. Fourier transform infrared (FTIR) spectroscopy
The infrared absorbance spectra were recorded using a Specac
Golden Gate diamond attenuated total reflectance (ATR) attach-
ment with a Thermo Nicolet 670 Nexus FTIR spectrometer equip-
ped with a DTGS KBr detector, and a KBr beamsplitter. Spectra were
averaged over 128 scans from 4000 to 600 cm1 at 4 cm1 reso-
lution. A single 128 scan background was taken before the sample
was placed on the ATR at ambient temperature. Gel samples were
sealed with a solvent cap on the ATR prior to beginning data
collection to ensure constant solvent content. Spectra of the pure
solvent were recorded using the same methods. Dried gel samples
and annealed polymer films were measured without the solvent
cap and with the anvil fully engaged to ensure full contact between
the ATR crystal and the sample.
2.6. Scanning electron microscopy (SEM)
The SEM images of the freeze-dried gel samples were obtained
with a Jeol JSM-7400F with an accelerating voltage of 3 kV. Samples
were sputter coated with a Denton Vacuum Desk IV sputter coater
using AuPd for 45 s before imaging to increase the interaction with
the electron beam. All polymer gels were imaged at the same
219
magnification of 6,000X. Magnification was limited due to the
sensitivity of the sample to the electron beam, which resulted in
melting of the polymer at higher magnifications.
3. Results and discussion
PHBHx dissolved in DMF at elevated temperatures appears as a
translucent, colorless liquid. As the temperature returns to
ambient, the opacity of the solution and the viscosity increase.
Once gelation has concluded, the sample transitioned to a white
and opaque solid. An example of the thermalereversibility of these
polymer-solvent systems is depicted in Fig. 2.
At room temperature, after gelation, the PHBHx gel is a white
and opaque solid. Upon heating past the gel-sol transition tem-
perature to 85
C, the sample will return to the original clear, non-
viscous solution. The “degree” of gelation of a sample, based on the
opacity, homogeneity, and viscosity, varies with the comonomer
content and the polymer concentration, as illustrated in Fig. 3.
The 13 mol % 3HHx PHBHx group portrays the effect of polymer
concentration on the strength of the gel. As the concentration in-
creases, so does the opacity, homogeneity, and viscosity of the
sample. At the same polymer concentration, the lower 3HHx con-
tent polymer gels have a higher opacity and better retention of their
structure upon tilting compared to those of higher 3HHx content.
Because the 3HHx content of PHBHx is directly related to the mo-
lecular regularity of the polymer, which dictates the eventual
crystallinity, this trend indicates a relationship between physical
gelation and the presence of crystalline polymer for thermally
reversible PHBHx gels.
In order to visualize the morphology of the gel system, solvent
exchange to a non-solvent, water, followed by lyophilization was
performed. Solvent exchange was required to ensure the polymer
could no longer interact with the solvent and form a film upon
drying. Freeze drying was necessary to ensure that the delicate
structure of the polymer gel was retained and did not collapse. SEM
images of the freeze dried gel samples are shown in Fig. 4.
The SEM images reveal that microscopic phase separation is
occurring between the polymer and the solvent, resulting in a
network of PHBHx ribbons and fibrils, less than a micrometer in
width and thickness, suspended in the solvent. As the concentra-
tion of the polymer increases, the morphology of the system does
not change, but the polymer regions increase in density. It is this
polymer network and change in density that causes the opacity of
the gels and its variation with polymer concentration. The effect of
3HHx content, or the molecular regularity of PHBHx, on the gel
morphology is more pronounced; the lower the regularity of the
polymer, the less defined the ribbons and fibrils appear. At 13 mol %
3HHx, the polymer network has noticeable regions of smooth,
globular structure and the fibrils increase in diameter. If the
network is being formed by crystallization of PHBHx in solution,
this observation would explain the difference in the strength of the
gels at different 3HHx content. The higher the 3HHx content, the
lower the molecular regularity, the less well defined the subse-
quent polymer network, which results in less light scattering, lower
opacity, less connectivity throughout the sample, and lower
viscosity.
The opacity and viscosity trends observed in the gel systems as a
function of 3HHx content, which is known to affect polymer crys-
tallinity in the pure solid, suggest the possible existence of a crys-
talline phase in the gels. In order to obtain conclusive evidence of
the presence of crystalline PHBHx in the gel system, WAXD profiles
of i) a 10 wt % PHBHx 3.9 mol % 3HHx in DMF prepared on an
amorphous glass slide sealed with plastic wrap and ii) the substrate
covered in plastic wrap are shown in Fig. 5.
The profile observed for the polymer gel sample contains
220 B.J. Sobieski et al. / Polymer 131 (2017) 217e223

Fig. 2. PHBHx gel at room temperature (left) and solution at 85
C (right).

Fig. 3. PHBHx gels at various 3HHx content and polymer solution concentration. Both (a) and (b) groups are increasing in polymer concentration from left to right. Image illustrates
the viscosity of the gel.

Fig. 4. PHBHx in DMF freeze dried gels. Top: left to right, 2,5,7,15 wt % of 3.9 mol % 3HHx. Bottom: 10 wt % of, left to right, 3.9, 5.8, 9.4, and 13 mol % 3HHx.

diffraction peaks assigned to the a-form crystal, specifically the
a(020) peak at 2q ¼ 13.3
, the a(110) peak at 2q ¼ 16.7
, and the
a(101) peak at 2q ¼ 21.1
. The diffraction peaks were assigned by
using Bragg's Law to calculate the d-spacing and comparing with
the report by Wang and Tashiro [30]. There is a broad feature which
begins at 2q ¼ 17.5
and extends to 2q ¼ 23
. Though diffraction
peaks do occur in this region for the crystalline forms of PHB, it is
more likely that a broad feature such as this is due to the amor-
phous content of the sample. The lattice spacings for the polymer
gel are a little higher than those in the bulk crystalline polymer; on
the order of 0.1 Å. One possible explanation for the larger lattice
spacing is that the PHBHx in the gel system is still favorably
interacting with the solvent, which may allow the crystalline re-
gions of the polymer to swell slightly. It is possible that this larger
 B.J. Sobieski et al. / Polymer 131 (2017) 217e223
800
700
600
Arbitrary Units
500
400
300
200
100
0
2θ
Fig. 5. WAXD profiles of (top) 10 wt % PHBHx 3.9 mol % 3HHx in DMF and (bottom)
clean amorphous glass slide covered in plastic wrap.
lattices spacing is also due to the formation of a complex between
some of the polymer sequences and the solvent, as was observed in
PS [4e10] and PVC [15e17]. Alternatively a larger concentration of
the 3HHx comonomers might be incorporated into the lattice due
to the differing processing protocols used to prepare the gel.
Infrared spectra were obtained on the PHBHx gels to confirm the
existence of polymer crystals demonstrated by the WAXD mea-
surements. The IR spectrum of PHAs contain bands that are asso-
ciated with the crystalline and amorphous portion of the polymer
[29], making IR analysis a viable option for demonstrating the ex-
istence of PHBHx crystals in the gel phase. Specifically, the carbonyl
stretching region for PHBHx contains a band associated with a
crystals centered at 1722 cm1 and a broad amorphous band
centered at 1740 cm1 when measured using ATR-IR [29e33].
There is one major drawback for IR as a characterization method for
PHBHx and that is overlap of absorbance bands between the
polymer and the solvent, as shown in Fig. 6.
The overlap of DMF and PHBHx in the carbonyl region is not
major, though the tail end of the DMF carbonyl band on the high
wavenumber side does overlap slightly with the PHBHx carbonyl
band. Comparing the neat 3.9 mol % 3HHx PHBHx spectrum with
the 15 wt % gel of the same polymer in DMF one observes a similar
Fig. 6. Representative ATR-IR spectra of neat 3.9 mol % 3HHx PHBHx, neat DMF, and
15 wt % PHBHx 3.9 mol % 3HHx in DMF gel focused on the carbonyl region.
221
band shape and the presence of both amorphous and a crystalline
peaks, thus confirming the results obtained from WAXD.
To evaluate the IR spectra as a function of polymer concentration
and 3HHx content, the ratio of carbonyl absorption intensity of
1722e1740 cm1, reflecting the crystalline content, was measured.
Fig. 7 contains said ratios for each PHBHx gel in DMF.
The amount of crystalline PHBHx in the 3.9 and 5.8 mol % 3HHx
PHBHx gels remains nearly constant as the polymer concentration
increases. However, there is a slight decrease in the amount of
crystalline PHBHx for 9.4 mol % 3HHx and a more noticeable
decrease in 13 mol % 3HHx with increasing polymer concentration.
For the lower 3HHx content polymer gels, the crystallinity is higher
indicating that even at lower polymer concentrations, there is a
consistent amount of crystalline content in the gel network. The
reason for the negative trend for the higher 3HHx content polymers
with polymer concentration is that at lower concentrations the
solutions are most likely not forming continuous interconnected
polymer networks. Rather, they are forming small crystallites that
remain partially suspended in the solvent. This effect can be
observed by the phase separation of the lowest concentration gel of
the 13 mol % 3HHx PHBHx gel in Fig. 3. The effect of the molecular
regularity of the polymer backbone is more apparent when the
1722 to 1740 cm1 absorption intensity ratio is plotted as a function
of 3HHx content, yielding a distinct negative trend with decreasing
molecular regularity at each polymer concentration. This trend in
the intensity ratio between the amorphous and crystalline bands
confirms the results of the XRD analysis, in that there is a decrease
in the relative intensity of the crystalline band as the molecular
regularity decreases. A decrease in the relative intensity of the
crystalline band compared to the amorphous band strongly sug-
gests that the polymer is crystallizing in the solution.
For further confirmation of the results in the wet gel samples, IR
spectra of the freeze-dried polymer gels were collected. Spectral
figures are displayed with the y-axis on a common scale and pro-
cessed utilizing the method outlined in a previous manuscript31
Fig. 8 shows IR spectra of 15 wt % PHBHx 3.9, 5.8, 9.4, and 13 mol %
3HHx in DMF freeze-dried gels showing both the carbonyl and
backbone regions and focused on the carbonyl solely.
As 3HHx content increases the crystalline band intensities
decrease and the amorphous intensities increase. This trend is most
noticeable in the C-O-C stretching bands and the carbonyl band, the
second of which has a clear reversal of intensity between 1722 and
1740 cm1 as the molecular regularity varies. To once again simplify
the analysis, Fig. 9 contains intensity ratios of 1722e1740 cm1
plotted as a function of 3HHx content for PHBHx in DMF freeze-
dried gels prepared under various cooling methods.
Just like the wet polymer gels, there is a negative trend in the
intensity ratio with increasing 3HHx content for each cooling
method. This trend indicates that as the molecular regularity of the
copolymer decreases, so does the amount of crystalline polymer
generated during gelation. Notice that there is a slight difference in
the data between the three cooling methods. Slow cooling the
samples to room temperature yields both the most variation in the
data points at each 3HHx content but also a smaller slope. However,
quenching the samples to 25
C yields the least amount of vari-
ation and the largest slope. Such results help confirm that materials
in the samples were generated during the gelation step of sample
preparation and not the subsequent drying process, otherwise
there would be no difference between the three cooling methods.
Recall that the samples were first exchanged to a non-solvent
before the drying process occurred. This exchange should “freeze”
the high molecular weight polymer in its gel state. Therefore, if the
polymer is crystallizing during the drying process and not the
gelation process, one would expect only the 3HHx content to
determine the crystallinity, which is not the case. For comparison,
222 B.J. Sobieski et al. / Polymer 131 (2017) 217e223

5 5
4 4

1722/1740

1722/1740
3 3.9mol 3
5wt
2 5.8mol 2
10wt
1 9.4mol 1
15wt
0 13mol 0

Concentration PHBHx (wt %) 3HHx Content (mol %)

Fig. 7. Ratio of 1722 cm1 (a crystal) to 1740 cm1 (amorphous) intensity as a function of polymer concentration (left) and 3HHx content (right).

Fig. 8. IR spectra of 15 wt % PHBHx in DMF quenched to 25
C freeze-dried gels of various 3HHx contents. Left contains the backbone and the carbonyl regions of the spectra and
right focuses on the carbonyl only.

2.5 (a) 2.5 (b)

2wt 2wt
1722/1740

1722/1740

2 2
5wt 5wt
7wt 7wt
1.5 1.5
10wt 10wt
1 12wt 1 12wt
15wt 15wt
3HHx Content (mol %) 3HHx Content (mol %)

2.5 (c)
2wt
1722/1740

2
5wt
7wt
1.5
10wt
1 12wt
15wt
3HHx Content (mol %)

Fig. 9. Ratio of intensity at 1722:1740 cm1 for PHBHx freeze-dried gels originally in DMF as a function of 3HHx content for each polymer concentration. (a) slowly cooled gels. (b)
gels quenched to 20
C. (c) gels quenched to 25
C.

the intensity ratios of 1722e1740 cm1 for the neat PHBHx films
annealed at 70
C for 4 h are listed in Table 1.
Initially, it may be surprising to find that the crystallinity of the
freeze-dried gel samples is higher than the annealed films. How-
ever, one must realize that these are produced under two very
different crystallization conditions. In the annealed samples the
polymers were cast from solution as amorphous films and allowed
to crystallize partially at room temperature before annealing at a
slightly elevated temperature. The thermal history of the samples
never allowed them to reach their equilibrium crystallization
temperature, which must be higher than 70
C based on the ther-
mal analysis, especially for 3.9 mol % 3HHx. Conversely, the samples
 B.J. Sobieski et al. / Polymer 131 (2017) 217e223
Table 1
Ratio of 1722 cm1 (a crystal) to 1740 cm1 (amorphous)
as a function of 3HHx content for neat PHBHx films
annealed at 70
C for 4 hours.
3HHx Content 1722/1740
(mol %)
3.9 2.02
5.8 1.76
9.4 1.63
13 1.27
in solution have substantially depressed melting and crystallization
temperatures, due to the interactions with the solvent and greater
entropy and mobility of the solvated polymer chains. Therefore,
even though the polymers are crystallizing closer to room tem-
perature, even at temperatures well below 70
C, crystallization in
solution is allowing both more ordered crystals to form and a
greater number of crystallites. As the differences in the cooling
method data illustrated, comparison with the annealed polymer
films also indicates that crystallization occurred during gelation
rather than the drying process. If crystallization occurred during
the drying or solvent exchange processes, then the 1722 to
1740 cm1 intensity ratios should mimic those of the annealed
polymer or show less crystalline content. The reason is that the
solvent exchange process takes place at room temperature and the
freeze-drying process below Tg, which are both lower crystalliza-
tion temperatures than the annealed films and the second of which
would not allow crystallization to occur due to the immobility of
the polymer chains.
4. Conclusion
These results demonstrate that PHBHx random copolymers
form physical gels when dissolved in DMF. The nature of the
resulting gels is dependent on the amount of the 3HHx comonomer,
which alters the molecular regularity and crystallinity of the
polymer in neat, bulk PHBHx. Through solvent exchange, lyophili-
zation, and subsequent SEM imaging, the structure of the gel sys-
tem was revealed as an interconnected polymer network of sub-
micron ribbons and fibrils. WAXD and FTIR results confirm this
result, yielding convincing evidence indicating the presence of a
crystalline phase as the source of linkage throughout the gel sys-
tem. Not only were crystalline a-form diffraction peaks observed in
WAXD, but infrared absorbance bands related to the crystalline
phase of the polymer were recorded for the gels. An understanding
of the cause and morphology may allow gels of PHBHx to be utilized
in relevant applications and bring to bear its exceptional biode-
gradability and biocompatibility.
Acknowledgments
The authors would like to acknowledge the support from
Delaware NSF EPSCoR Grant # 1301765 and the NSF DMR Polymers
Program Grant #1407255. The authors would also like to
acknowledge Gerald Poirier for his valuable insight and contribu-
tions to this work.
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