Enzymatic Synthesis and Stereocomplex Formation of Chiral Polyester Containing Long-Chain Aliphatic Alcohol Backbone

Article abstract of DOI:10.1021/acs.biomac.9b00918

Herein we demonstrated a novel lipase-catalyzed synthesis of isotactic D-/L-poly(aspartate-octanediol) ester containing long chain alcohols backbone and discovered their stereocomplex feature with an increased T_m for the first time. Simple desi

Full text of DOI:10.1021/acs.biomac.9b00918

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Article
Enzymatic synthesis and stereocomplex formation of chiral
polyester containing long-chain aliphatic alcohol backbone
Bo Xia, Yu Zhang, Qiaoyan Zhu, Xianfu Lin, and Qi Wu
Biomacromolecules, Just Accepted Manuscript • DOI: 10.1021/acs.biomac.9b00918 • Publication Date (Web): 19 Aug 2019
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Enzymatic synthesis and stereocomplex formation of
chiral polyester containing long-chain aliphatic
alcohol backbone
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Bo Xia ‡, # Yu Zhang, †, # Qiaoyan Zhu, † Xianfu Lin, † Qi Wu *, †
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† Department of Chemistry, Zhejiang University, Hangzhou 310027, People’s Republic of China;
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‡ Jiyang College of Zhejiang A&F University, Zhuji 311800, People’s Republic of China;
KEYWORDS: Enzymatic polymerization, Chiral polyesters, Stereocomplex
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ABSTRACT
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Herein we demonstrated a novel lipase-catalyzed synthesis of isotactic D-/L-poly (aspartate-
octanediol) ester containing long chain alcohols backbone and discovered their stereocomplex
feature with an increased Tm for the first time. Simple design of monomer structures not only
overcomes the inherent selectivity limitation of enzyme used, but also achieves totally isotactic
polyester products. By crystallizing the mixed enantiopure isotactic polyesters in different
solvents, the formation of amorphous mixture, homocrystallites or stereocomplex crystallites were
observed, respectively. This study is expected to open up a new way to prepare various polyesters
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stereocomplex containing long-chain aliphatic alcohol backbone and a wide variety of functional
groups.
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Introduction
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Polymer stereocomplexation is the stereoselective interaction between two stereoisotactic
complementary polymers in the crystalline state, and it has become an attractive route to improve
thermal and mechanical properties of polymeric materials.1 To date, the most studied examples of
polymer stereocomplex are poly (methyl methacrylate) (PMMA) 2, 3 and poly (lactic acid) (PLA).
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Compared with the common homocrystallite (HC) composed of enantiopure D- or L-
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polylactide, melting temperature (Tm) of the stereocomplex increases about 50 C and the
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hydrolytic degradation rate can be reduced by more than 3 times. These improved mechanical
and thermal properties associated with the polylactide stereocomplexation accelerate its
application in biological medicine field greatly. 8
Besides polylactide stereocomplex, CO2-based stereoisotactic polycarbonates are another
important type of stereocomplex developed in recent years, such as poly (limonene carbonate)s,
poly (propylene carbonate)s and poly (cyclopentene carbonate), which were synthesized through
steric selective copolymerization between CO2 and enantiopure- or meso-epoxides under the
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catalysis of chiral complexes of cobalt (Co) or zinc (Zn). Significant improvement in thermal
properties was observed for these stereoisotactic CO2-based polycarbonates. 9b-9c What’s more, as
a pioneer in this field, Coates and coworkers first observed the formation of stereocomplex
between (S)- and (R)- poly (propylene succinate), which were synthesized via a cobalt complexes-
catalyzed alternating copolymerization of enantiopure propylene oxide and succinic anhydride. 10
More recently, Wang and coworkers reported a new crystalline stereocomplex by mixing isotactic
(D)- and (L)-poly(mandelic acid) (PMA), an aryl analogue of poly(lactic acid) (PLA), which were
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prepared via ring-opening polymerization of manOCA monomer (O-carboxyanhydrides from
mandelic acid) mediated by a bifunctional single molecule organocatalysts.11 However, only very
limited classes of polymer stereocomplexes have been reported so far. And to the best of our
knowledge, no example of the formation of stereocomplex between enantiocomplementary
polyesters containing long-chain alcohols backbone has been reported. 12 Therefore, the discovery
of more novel polymer stereocomplexes is highly desirable.
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On the other hand, developing more eco-friendly synthesis methods for polymer
stereocomplexes is another challenge because almost all polymer stereocomplexes reported to
date, except poly (mandelic acid) (PMA) 11, were synthesized by using specific chiral metal-
complexes, which usually are expensive and complicated, and have some unadmired
environmental disadvantages. Enzymatic polymerization is a highly environmentally friendly
pathway with high chemo-, regio- and stereo-selectivity in polymer synthesis.13-14 Chiral polyesters
can be enzymatically synthesized by either the enantioselective polycondensation of diacids and
diols (or hydroxy acids), or ring-opening polymerization (ROP) of lactones. For the last decades,
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some significant advances of enzymatic synthesis of chiral polyesters have been achieved,
however, there are still some problems to be solved. For example, the molecular weight of the
produced chiral polyesters is usually low because there is a contradiction between molecular
weight and optical purity of products in the kinetic resolution polymerization of racemic
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monomers.
Another problem is that lipases are usually highly selective towards one specific
configuration of the monomers, thus enantiocomplementary polyesters are difficult to be prepared.
16b, e
Moreover, one of the biggest challenges is that it usually cannot provide isotactic chiral
22 polyesters when using racemic diacids or diols with asymmetric terminals as monomers because
23 there are several possible connections of their monomers, such as tail-to-tail, head-to-head or head-
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to-tail. Thus, these enantiopure polyesters are difficult to form stereocomplex. Therefore, we
proposed to exploit a new strategy for enzymatic synthesis of enantiocomplementary isotactic
polyesters with controlled molecular weights (up to 24 kDa) and long-chain alcohols backbones
and hope to discover their stereocomplexation features.
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Herein, we reported a new class of chiral polyester stereocomplex of isotactic D- and L-poly
(aspartate-octanediol) ester containing long-chain aliphatic alcohol backbone and readily
commercially available amino acids, which was prepared via a facile and eco-friendly lipase-
catalyzed route (Scheme 1). By crystallizing the mixed enantiopure polyesters in different solvent,
we were able to obtain the amorphous mixture, homocrystallites or stereocomplexed crystallites,
respectively. Stereocomplex of stereoisotactic D- and L-poly (aspartate-octanediol) ester possesses
an increased Tm and a greater level of crystallinity.
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Cbz
Cbz
HN
O
HN
O
OH
O
OH
O
O
O
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O
O
L-2a
D-2a
CALB-catalyzed
polycondensation
Cbz
Cbz
HN
O
HN
O
O
O
O
O
n
n
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O
O
isotactic L-polyester
isotactic D-polyester
Mixed
Amorphous mixture
DCM/MeOH (1:2)
Homocrystallites
DCM or DCM/hexane (1:3)
Stereocomplex
DCM/MeOH (1:3)
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Scheme 1. Lipase-catalyzed synthesis of enantiocomplementary isotactic polyesters with long-
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chain alcohol backbone and their stereocomplex
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Experimental Section
Materials
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The L- or D- aspartic acid were purchased from Sigma-Aldrich. Immobilized Lipase B from
Candida antarctica (Novozym 435) was purchased from Sigma-Aldrich too. Toluene (99%) and
other reagents were purchased from Sinopharm Chemical Reagent Co. Ltd. Other regents and
compounds were purchased by J&K Scientific.
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Methods
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Nuclear Magnetic Resonance (NMR): H NMR and 13C NMR spectra were recorded with
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tetramethylsilane (TMS) as the internal standard using a Bruker AMX-400 MHz spectrometer
(Rheinstetten, Germany) using CDCl3 as solvents. 1H and 13C spectra were determined using the
standard pulse sequence provided by Bruker. The samples were analyzed at 25 oC. TMS (ppm: 0)
was used as the internal standard for NMR analysis.
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Differential scanning calorimetry (DSC): Thermal analysis was carried out with a DSCQ1000
TA Instrument, with the calorimeter under nitrogen (30 mL/min) connected to a cryostat from the
same manufacturer. DSC experiments were prepared in crimped aluminum pans and standard
experiments were conducted with a heating rate of 10 ºC/min from -50 ºC to 100ºC.
Gel Permeation Chromatography (GPC): Molecular weights of polyesters were measured by
GPC with a system equipped with a refractive-index detector (Waters 2414) and Waters Styragel
GPC columns (Massachusetts, USA). The GPC columns were standardized with narrow-dispersity
polystyrene in molecular weight ranging from 6×105 to 500. The mobile phase was THF at a flow
rate of 1.0 mL/min.
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Circular Dichroism (CD): CD spectra were recorded on a JASCO J-815 spectrometer. The light
path length of the quartz cell used was 10 mm. The concentration was about 0.01 mg/mL, and the
solvent was acetonitrile.
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X-ray Diffraction (XRD): XRD patterns were recorded with a PANalytical X’Pert PRO powder
diffractometer using Cu Kα radiation (λ= 0.1541 nm). The working voltage was 40 kV and the
working current was 40 mA. The patterns were collected with a 2θ range from 3◦ to 50◦at a step of
0.0501◦.
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All geometry optimizations were carried out using the semi-empirical GFN2-xTB method17.
Graphical representations of molecular structures were drawn using PyMOL.99rc6 program18.
Synthesis of N-Cbz- Asp diester
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2.0 mL Thionyl chloride was added into a solution of N-Cbz Asp (5.0 g) in 50 mL MeOH. Then
the reaction mixture was refluxed overnight and concentrated in vacuum. The obtained crude
product was used for next polymerization without any purification.
Dimethyl L-N-Cbz-aspartate, L-1: 1H-NMR (400 MHz, CDCl3) δ: 7.26 (m, 5H, -phenyl), 5.05
(s, 2H, -CH2-phenyl), 4.57 (m, 1H, -CH-NH-), 3.69 (s, 3H, β-OCH3), 3.61 (s, 3H, α-OCH3), 2.88
(m, -CH2-COOMe) ppm. 13C-NMR (100 MHz, CDCl3) δ: 171.4, 171.2, 155.9, 136.1, 128.6, 128.3,
128.1, 67.2, 52.9, 52.1, 50.3, 36.5 ppm. The NMR spectra of Dimethyl D-N-Cbz-aspartate, D-1
was as same as L-1.
Synthesis of D- and L-N-Cbz-α-octanediol-β-methyl-aspartate
A solution of D- or L- N-Cbz-β-methyl-aspartate (0.1 mmol) in 50 mL DCM was added 1, 8-
octanediol (0.15 mmol), DMAP (0.01 mmol) and DCC (0.1 mmol). Then the reaction mixture was
stirred overnight at R.T. and TLC was used to monitor the reaction progress. After the reaction
completed, the mixture was washed with 5% NaHCO3 and 5% citric acid. The organic phase was
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dried over anhydrous MgSO4 and concentrated in vacuum. The residue was purified with column
chromatography (petrol ether/EtOAc=3:1).
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N-Cbz-D-α-octanediol-β-methyl-aspartate, D-2a: 1H-NMR (400 MHz, CDCl3) δ: 7.37-7.32 (m,
5H, -phenyl), 5.81-5.79 (m, 1H, -NH-Cbz), 5.12 (s, 2H, -CH2-phenyl), 4.64-4.61 (m, 1H, -CH-
NH-), 4.17-4.13 (m, 2H, -CH2-O-CO-), 3.68 (s, 3H, -OCH3), 3.65-3.61 (m, 2H, -CH2-OH), 3.06-
2.83 (m, 2H, -CH2-COOMe), 1.62-1.47 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OH), 1.32-
1.26 (m, 8H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OH) ppm; 13C-NMR (100 MHz, CDCl3) δ: 171.3,
170.7, 156.0, 136.1, 128.6, 128.2, 128.1, 67.1, 66.0, 63.0, 52.1, 50.4, 36.5, 32.7, 29.2, 29.0, 28.4,
25.6 ppm.
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N-Cbz-L-α-octanediol-β-methyl-aspartate, L-2a: 1H-NMR (400 MHz, CDCl3) δ: 7.37-7.32 (m,
5H, -phenyl), 5.81-5.79 (m, 1H, -NH-Cbz), 5.12 (s, 2H, -CH2-phenyl), 4.64-4.61 (m, 1H, -CH-
NH-), 4.17-4.13 (m, 2H, -CH2-O-CO-), 3.68 (s, 3H, -OCH3), 3.65-3.61 (m, 2H, -CH2-OH), 3.06-
2.83 (m, 2H, -CH2-COOMe), 1.62-1.47 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OH), 1.32-
1.26 (m, 8H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OH) ppm; 13C-NMR (100 MHz, CDCl3) δ: 171.3,
170.7, 156.0, 136.1, 128.6, 128.2, 128.1, 67.1, 66.0, 63.0, 52.1, 50.4, 36.5, 32.7, 29.2, 29.0, 28.4,
25.6 ppm.
Preparation of regio-random L-poly (N-Cbz-Asp-octanediol) ester
CAL B (45 mg, 15% wt of diester) were added into the solution of diols (0.2 mmol) and diesters
(0.2 mmol) in 3.0 mL toluene. Then the flask was replaced with nitrogen and the reaction mixture
was pre-polymerized at 80 oC for 3 days. The mixture was concentrated in vacuum to remove the
solvent and then reacted in vacuum for another 2 days (0.01 MPa). The obtained polymers were
dissolved in 1.0 mL chloroform (EtOAc). Then 5.0 mL MeOH (60-90 oC) was added slowly. After
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10 minutes’ standing, the supernatant was discarded and the residue was treated with this
procedure for 3 times. The final obtained polymers were concentrated in vacuum.
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Regio-random L-poly (N-Cbz-Asp-octanediol) ester, L-poly-1a, 1H-NMR (400 MHz, CDCl3), δ:
7.29-7.19 (m, 5H, -phenyl), 5.73-5.71 (d, 1H, J=8 Hz, -NH-Cbz), 5.04 (s, 2H, -CH2-phenyl), 4.55-
4.53 (m, 1H, -CH-NH-), 4.06-3.97 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-), 3.68 (s,
0.02 H, CH3O-), 2.91-2.78 (m, 2H, -CH2-COOMe), 1.58-1.52 (m, 4H, -OCO-CH2-CH2-(CH2)4-
CH2-CH2-OCO-), 1.22 (s, 8 H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-) ppm. 13C-NMR (100
MHz, CDCl3) δ: 169.96, 169.73, 154.94, 135.16, 127.51, 127.18, 127.04, 66.04, 64.90, 64.15,
49.44, 35.64, 28.04, 28.01, 27.43, 27.36, 24.74, 24.64 ppm.
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Preparation of isotactic D- and L- poly (N-Cbz-Asp-octanediol) ester
CAL B (47 mg, 15%, wt% of monomer) were added into the solution of diester monomer (0.2
mmol) in 3.0 mL toluene. Then the flask was replaced with nitrogen and the reaction mixture was
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pre-polymerized at 80 C for 3 days. The mixture was concentrated in vacuum to remove the
solvent and then reacted in vacuum for another 2 days (0.01 MPa). The obtained polymers were
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dissolved in 1.0 mL chloroform. Then 5 mL petrol ether (60-90 C) was added slowly. After 10
minutes’ standing, the supernatant was discarded, and the residue was treated with this procedure
for 3 times. The final obtained polymers were concentrated in vacuum.
Isotactic D-poly (N-Cbz-Asp-octanediol) ester, D-poly-2a, 1H-NMR (400 MHz, CDCl3), δ: 7.29-
7.19 (m, 5H, -phenyl), 5.73-5.71 (d, 1H, J=8 Hz, -NH-), 5.04 (s, 2H, -CH2-phenyl), 4.55-4.53 (m,
1H, -CH-NH-), 4.06-3.97 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-), 3.68 (s, 0.06 H,
CH3O-), 2.91-2.78 (m, 2H, -CH2-COOMe), 1.58-1.52 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-
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CH2-OCO-), 1.22 (s, 8H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-) ppm. C-NMR (100 MHz,
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CDCl3) δ: 170.89, 170.77, 155.97, 136.19, 128.54, 128.21, 128.07, 67.07, 65.93, 65.18, 50.47,
36.67, 29.09, 29.05, 28.46, 28.39, 25.78, 25.67 ppm.
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Isotactic L-poly (N-Cbz-Asp-octanediol) ester, L-poly-2a, 1H-NMR (400 MHz, CDCl3), δ: 7.36-
7.31 (m, 5H, -phenyl), 5.80-5.78 (d, 1H, J=8 Hz, -NH-), 5.12 (s, 2H, -CH2-phenyl), 4.63-4.61 (m,
1H, -CH-NH-), 4.15-4.05 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-), 3.68 (s, 0.06 H,
CH3O-), 2.87-2.83 (m, 2H, -CH2-COOMe), 1.61-1.59 (m, 4H, -OCO-CH2-CH2-(CH2)4-CH2-
CH2-OCO-), 1.29 (s, 8H, -OCO-CH2-CH2-(CH2)4-CH2-CH2-OCO-) ppm. 13C-NMR (100 MHz,
CDCl3) δ: 170.88, 170.76, 155.97, 136.17, 128.53, 128.20, 128.06, 67.06, 65.92, 65.17, 50.46,
36.66, 29.08, 29.03, 28.45, 28.38, 25.76, 15.66 ppm.
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Results and Discussion
Considering the high commercial availability of D- and L-aspartic acid (Asp), and the
convenience for introducing functional groups into the -NH2 of Asp, we selected D-/L-Asp as an
alternative of racemic diacids for preparing chiral polyesters. Candida antarctica lipase B (CALB,
Novozym 435) was used as the catalyst due to its high stereo- and regioselectivity, and wide
applications in various organic transformations including kinetic resolution and
polymerization.14,19,20 We selected dimethyl N-Cbz-aspartate (N-Cbz-Asp (OMe)2) as an Asp
monomer to avoid the formation of polyamide by protecting -NH2 group with -Cbz.21 In order to
test the selectivity of Novozym 435 towards L- and D-Asp monomers, we compared the
polymerization results of D- and L-N-Cbz-Asp (OMe)2 (D-1 and L-1) and 1, 8-octanediol as shown
in Table 1 (entry 1, 2) and Scheme 2. The polycondensation of L-1 successfully provided L-
poly(N-Cbz-Asp-octanediol)) ester (L-poly-1a) with a molecular weight of 24.2 kDa (Mn) and
76% yield, while the polycondensation of D-1 was unsuccessful (entry 1 vs. 2, Table 1). The
structure of the targeting polyester was confirmed by NMR (Figure S4-6). These results indicated
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that Novozym 435 had higher preference for L-monomer than D-monomer in polymerization. It
was also confirmed by the Novozym 435-catalyzed transesterification resolution between 1-
octanol and D- or L-Asp monomers (D-/L-N-Cbz-Asp (OMe)2) (results were listed in Figure S11).
Novozym 435 presented high L-configuration stereoselectivity, and the ee value of obtained
dioctyl N-Cbz- aspartate reached up to 97%.
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Table 1. CALB-catalyzed Polycondensation for the Synthesis of Regio-random or Isotactic
Polyesters
entry monomer
polymer
Mn c (kDa)
1.4
Mw c (kDa)
1.4
PDI c
DP d
5
yield (%)
4
Regio-random D-
1 a
2 a
3 b
4 b
D-1 + octanediol
-
e
poly-1a
Regio-random L-
L-1 + octanediol
D-2a
24.2
12.0
12.3
39.5
21.7
19.4
1.64
1.81
1.57
141
78
76
72
70
poly-1a
Isotactic D-poly-
2a
Isotactic L-poly-
L-2a
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2a
a
o
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Reaction conditions: 0.1 mmol diester (D-1 or L-1), 0.1 mmol diol, pre-polymerization at 80 C for 3 days,
b
then polymerization in vacuum for 2 days. Reaction conditions: 0.1 mmol monomers (D-2a or L-2a), pre-
o
c
polymerization at 80 C for 3 days, then polymerization in vacuum for 2 days. The Mn, Mw and PDI were
determined by GPC using THF as solvent. d DP refer to average degree of polymerization. e Not detected.
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Concerning the failed polycondensation between D-1 and octanediol, we believe that steric
hindrance of D-Asp diester is perhaps the main reason. There are three acyl groups in D-1 (α-, β-
and N-acyl) (Scheme 2). The N-acyl group cannot react with diols because of the lower
nucleophilicity of hydroxyl group than amino group, and also the bulky benzyl group in -Cbz
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which prevents from binding into the active site of CALB. Only α- and β-acyl can participate in
the polycondensation with diols. The steric hindrance and non-preferential selectivity limitation of
the α-acyl in D-Asp diesters would be much larger than that of β-acyl due to its’ shorter distance
to the chiral center. Transesterification between α-acyl and hydroxyls was probably unfavorable,
thus hindering the chain propagation.
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Scheme 2. CALB-catalyzed Polycondensation of D-/L-N-Cbz Asp Diesters and Octanediol
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In order to remove the selectivity limitation of Novozym 435 for the slow reacting D-Asp
diesters, and to obtain the D- poly (N-Cbz-Asp-octanediol) ester, we designed N-Cbz-D-α-
octanediol-β-methyl-aspartate (D-2a, Scheme 1) as a starting monomer by hiding the chiral center
and unfavorable α-acyl in D-1. Interestingly, CALB-catalyzed polycondensation of D-2a
successfully provided the targeting D-configurational polyester (D-poly-2a) with a molecular
weight of 12.0 kDa (Mn) and 72% yield (entry 3, Table 1). In order to prove the efficiency of this
strategy, polycondensation of N-Cbz-L-α-octanediol-β-methyl-aspartate (L-2a) was tested, and a
good result was also obtained (entry 4, Table 1). It was consistent with our prediction that the
hided chiral center of D-2a was far away from the terminal reacting group, and then a successful
polycondensation can be achieved. This successful demonstration also offers a new option for
overcoming the selectivity limitation of enzymes through facile and simple substrate engineering,
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rather than protein engineering which is generally used to solve these types of problem, while is
relative difficult for chemists. 22 Another more important advantage of this novel synthetic strategy
is that the prepared chiral polyesters are totally isotactic, however lipase-catalyzed
polycondensation of the common N-Cbz-Asp diester and diols monomers only provided regio-
random polyesters (Scheme 2). Circular dichroism (CD) spectroscopy was applied to investigate
the conformation and chiroptical properties of obtained chiral polyesters. As shown in Figure 1,
it clearly showed that the L-poly-1a, L-poly-2a and D-poly-2a, respectively, produce vertically
mirrored negative and positive CD signals at 225 nm.
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L-poly-2a
D-poly-2a
L-poly-1a
4
2
0
-2
-4
200
250
300
350
Wavelength (nm)
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Figure 1. CD spectra of enantiopure regio-random and isotactic poly (N-Cbz-Asp-octanediol) ester
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Neither regio-random L-poly-1a itself nor the mixture of regio-random L-poly-1a and isotactic
D-poly-2a showed any crystallinity. Interestingly, the DSC of both isotactic D-poly-2a and L-
poly-2a showed obvious crystallinity in the first heat (Figure 2A and 2C). For D-poly-2a, a Tm
around 49 oC and a ΔH value of 12.2 J/g were observed along with a glass transition temperature
(Tg) of −6 °C. Tm, ΔH and Tg values of L-poly-2a were 56 oC, 23.8 J/g, and -3.0 oC respectively.
Figure 2B and 2D showed that both D-poly-2a and L-poly-2a parent polymers do slowly
crystallize from the melt. After about 24 days, crystallinity of D-poly-2a was first observed with
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a slightly low Tm and ΔH value (46 oC, 8.5 J/g). And, the crystallization process of L-poly-2a was
a little slower than its D-stereoisomer, with crystallinity first appearing after about 36 days. It is a
reason why no Tm was seen on the 2nd heat.
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A
B
1 days
nd
2
heat
3 days
st
1
heat
12 days
Cbz
O
HN
O
O
24 days
-20
n
6
O
D-poly-2a
-40
-20
0
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60
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0
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o
o
Temperature ( C)
Temperature ( C)
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C
1 day
D
3 days
nd
2
heat
12 days
24 days
st
1
heat
36 days
Cbz
HN
O
O
O
n
6
O
L-poly-2a
-40
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0
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60
-40
-20
0
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o
o
Temperature ( C)
Temperature ( C)
5
F
E
DCM/MeOH=1:2
DCM/MeOH=1:3
Sample 1
(Evaporated in DCM)
Sample 2
(Precipitated in DCM/hexane)
L-poly-2a + D-poly-2a
DCM/MeOH=1:5
DCM/MeOH=1:7
Sample 3
(Precipitated in DCM/MeOH)
L-poly-2a + D-poly-2a
-40
-20
0
20
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60
-40
-20
0
20
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60
o
o
Temperature( C)
Temperature ( C)
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Figure 2. DSC of (A) isotactic D-poly-2a, (B) crystallization of isotactic D-poly-2a over time,
(C) isotactic L-poly-2a, (D) crystallization of isotactic L-poly-2a, (E) isotactic D-poly-2a and L-
poly-2a mixture prepared by different methods, (F) isotactic D-poly-2a and L-poly-2a mixture
crystallized in DCM/MeOH with different ratio.
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To test the formation of stereocomplex, three samples were prepared with different methods,
and each sample consisted of equal amount of D-poly-2a and L-poly-2a. Thermal properties of
these samples were analyzed by DSC and the data were compared with their parent polyesters
(Figure 2E). No stereocomplexation was observed in the DSC test of sample 1 and 2. For these
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two samples, a same Tm around 51 C was observed. The Tm of sample 1 and 2 is very close to
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their parent polyesters (49 C, D-poly-2a; 56 C, L-poly-2a), indicating the formation of
homocrystallites. The increased Tm of sample 3 (67 °C with a ΔH value of 23.9 J/g) showed the
formation of stereocomplex. Compared with its parent polyesters, the Tm of stereocomplex
o
increased about 20 C. The influence of solvents on the stereocomplex formation might be
attributed to the solubility difference of D- or L-poly-2a in different mixed solvents. When n-
hexane was added to the DCM solution of D-poly-2a and L-poly-2a, the precipitate formed much
faster than when MeOH was added. We believe that the rapid precipitation would disturb
stereocomplexation between D-poly-2a and L-poly-2a. Therefore, the stereocomplex probably
only formed in the mixed solvent of DCM/MeOH with a suitable ratio.
According to the above test, stereocomplex formed only in a specific DCM/MeOH mixed
solvent. Thus, the ratio of DCM/MeOH (1:2, 1:3, 1:5 and 1:7) was further investigated (Figure
2F). For the sample prepared from the 1:2 DCM/MeOH solvent, no crystallization melting peak
o
but a low Tg (-11.7 C) was observed. Interestingly, polyester mixture precipitated in 1:5
DCM/MeOH solvent formed a polymorph which contained a homo-crystallite with a Tm of 50 oC
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and a stereocomplex with a Tm of 62 oC. However, as the coexistence of both homocrystallites and
stereocomplex, Tm of stereocomplex was 5.0 oC lower than the pure stereocomplex (67 oC vs 62
oC). With the increase of methanol content to 1:7, stereo-complexation disappeared and only
homocrystallites were observed with a Tm around 51 oC and a ΔH value of 43.1 J/g.
Powder X-ray diffraction (XRD) was also used to confirm the formation of stereocomplex
(Figure 3). The presence of different crystalline diffraction peaks in comparison with the
diffraction profiles of its parent polymers demonstrated the successful stereocomplexation. The
polymer stereocomplex shows four large peaks at 18.1o, 19.3o, 21.9o and 23.2o, and one small
shoulder at 21.0o. In contrast, both D-poly-2a and L-poly-2a show small crystalline diffraction
signals at 19.8 o and 21.9o. The degree of crystallinity of stereocomplex was calculated through the
multipeak fitting method, and the results are shown in Figure S12. An approximation of the
amorphous region of the polymers can be subtracted from an approximation of the crystalline peak
area. The stereocomplex shows a crystallinity of 85%, while the crystallinity of D-poly-2a and L-
poly-2a was 63% and 71%, respectively.
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C
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2 Theta (degree)
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Figure 3. Powder XRD profiles: (A) stereocomplex. (B) isotactic L-poly-2a. (C) isotactic D-poly-
2a.
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In order to make the formation of stereocomplex of D-poly-2a and L-poly-2a much clearer, a
preliminary structure model containing chains of opposite chirality was constructed and optimized
using the semi-empirical GFN2-xTB method17 that has shown to be particularly suitable and
accurate to highly efficient geometry optimization of very large molecules (Figure 4). The
minimum energy packing model implied that it probably benefited from some formed H-bonds
between the amino groups of Asp and carbonyls of Asp or Cbz in the enantiomeric polyester
chains, as well as pi-pi interactions between benzene rings (Figure 4A). Crystallization of the
stereocomplex of D-poly-2a and L-poly-2a might be driven by the tight interdigitation of the
backbone and side chains of enantiocomplementary polyesters belonging to adjacent layers,
similar as a “steric zipper” described by Coates and coworkers (Figure 4B).9d, 23
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Figure 4. Minimum energy packing model for enantiomeric polyester chains. A, side view, B,
front view. D-poly-2a is indicated as ball and stick in black and L-poly-2a as stick in green. Cbz
groups is indicated as lines in grey. Oxygen and nitrogen atoms are indicated in red and blue,
respectively.
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Conclusions
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In this work, we reported a novel lipase-catalyzed synthesis of isotactic D- and L-poly (aspartate-
octanediol) ester containing long chain diols and readily commercially available amino acids and
discovered the first example of a crystalline stereocomplex in this area. By designing a N-Cbz-α-
octanediol-β-methyl-aspartate as the starting monomer to hide the chiral center and create a ‘non-
chiral’ condensation environment artificially for enzymatic polymerization, isotactic D- and L-
poly (N-Cbz-Asp-octanediol) ester could be prepared under the catalysis of CALB. Generally
speaking, it is difficult to prepare the similar isotactic polyesters with high molecular weight and
optical purity by conventional catalysts, considering the eco-friendly reaction conditions and strict
demand of the residual metal ion.24 The stereocomplex formation between isotactic D- and L- poly
(N-Cbz-Asp-octanediol) ester was confirmed by DSC and XRD. The influence of solvent
compositions on the formation of stereocomplex was observed. The formation of stereocomplex
in-creased the crystallinity and their Tm about 20 oC. Based on the highly commercial availability
of amino acid and aliphatic diols, a series of biocompatible polyester stereocomplexes with
controlled stereoregularity and improved physical and chemical properties could be synthesized in
the future via eco-friendly enzymatic polymerizations.
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ASSOCIATED CONTENT
Supporting Information
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(Experimental procedures, characterization data and spectra of all new compounds. This material
is available free of charge via the Internet at http://pubs.acs.org.)
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AUTHOR INFORMATION
Corresponding Author
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* E-mail for Q. W.: wuqi1000@163.com
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ORCID
Qi Wu: 0000-0001-9386-8068
Bo Xia: 0000-0002-7791-2636
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Author Contributions
# These authors contributed equally to this work
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Funding Sources
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Financial support from the National Natural Science Foundation of China (21574113), and
Natural Science Foundation of Zhejiang province, China (LY19B020014, LY17B040002) is
gratefully acknowledged.
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Notes
The authors declare no competing financial interests.
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Enzymatic synthesis and stereocomplex formation of chiral polyester
containing long-chain aliphatic alcohol backbone
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Bo Xia ‡, # Yu Zhang, †, # Qiaoyan Zhu, † Xianfu Lin, † Qi Wu *, †
†
Department of Chemistry, Zhejiang University, Hangzhou 310027, People’s Republic of China;
‡
Jiyang College of Zhejiang A&F University, Zhuji 311800, People’s Republic of China;
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