New insights into poly(lactic- co -glycolic acid) microstructure: Using repeating sequence copolymers to decipher complex NMR and thermal behavior

Article abstract of DOI:10.1021/ja102670n

Sequence, which Nature uses to spectacular advantage, has not been fully exploited in synthetic copolymers. To investigate the effect of sequence and stereosequence on the physical properties of copolymers, a family of complex isotactic, syndiotactic, and atactic repeating sequence poly(lactic-co-glycolic acid) copolymers (RSC PLGAs) were prepared and their NMR and thermal behavior was studied. The unique suitability of polymers prepared from the bioassimilable lactic and glycolic acid monomers for biomedical applications makes them ideal candidates for this type of sequence engineering. Polymers with repeating units of LG, GLG and LLG (L = lactic, G = glycolic) with controlled and varied tacticities were synthesized by assembly of sequence-specific, stereopure dimeric, trimeric, and hexameric segmer units. Specifically labeled deuterated lactic and glycolic acid segmers were likewise prepared and polymerized. Molecular weights for the copolymers were in the range M_n = 12-40 kDa by size exclusion chromatography in THF. Although the effects of sequence-influenced solution conformation were visible in all resonances of the ¹H and ¹³C NMR spectra, the diastereotopic methylene resonances in the ¹H NMR (CDCl₃) for the glycolic units of the copolymers proved most sensitive. An octad level of resolution, which corresponds to an astounding 31-atom distance between the most separated stereocenters, was observed in some mixed sequence polymers. Importantly, the level of sensitivity of a particular NMR resonance to small differences in sequence was found to depend on the sequence itself. Thermal properties were also correlated with sequence.

Full text of DOI:10.1021/ja102670n

Published on Web 07/15/2010
New Insights into Poly(lactic-co-glycolic acid) Microstructure:
Using Repeating Sequence Copolymers To Decipher Complex
NMR and Thermal Behavior
Ryan M. Stayshich and Tara Y. Meyer*
Department of Chemistry, UniVersity of Pittsburgh, Pittsburgh, PennsylVania 15260
Received April 8, 2010; E-mail: tmeyer@pitt.edu
Abstract: Sequence, which Nature uses to spectacular advantage, has not been fully exploited in synthetic
copolymers. To investigate the effect of sequence and stereosequence on the physical properties of
copolymers, a family of complex isotactic, syndiotactic, and atactic repeating sequence poly(lactic-co-glycolic
acid) copolymers (RSC PLGAs) were prepared and their NMR and thermal behavior was studied. The
unique suitability of polymers prepared from the bioassimilable lactic and glycolic acid monomers for
biomedical applications makes them ideal candidates for this type of sequence engineering. Polymers
with repeating units of LG, GLG and LLG (L ) lactic, G ) glycolic) with controlled and varied tacticities
were synthesized by assembly of sequence-specific, stereopure dimeric, trimeric, and hexameric segmer
units. Specifically labeled deuterated lactic and glycolic acid segmers were likewise prepared and
polymerized. Molecular weights for the copolymers were in the range Mn ) 12-40 kDa by size exclusion
chromatography in THF. Although the effects of sequence-influenced solution conformation were visible in
1
1
all resonances of the H and 13C NMR spectra, the diastereotopic methylene resonances in the H NMR
(CDCl3) for the glycolic units of the copolymers proved most sensitive. An octad level of resolution, which
corresponds to an astounding 31-atom distance between the most separated stereocenters, was observed
in some mixed sequence polymers. Importantly, the level of sensitivity of a particular NMR resonance to
small differences in sequence was found to depend on the sequence itself. Thermal properties were also
correlated with sequence.
foldamers12 and in the benefits already derived from the
relatively modest degrees of sequence control present in
Introduction
The creation of polymers with a high degree of sequence and/
or stereocontrol represents an exciting frontier in materials
science as a result of both the paucity of examples of polymers
with high values in these two categories and the potential
benefits to be realized from the creation of new polymers from
readily available and well-understood monomers.1-11 A valuable
lesson on the key role of sequence/stereochemical control can
be learned from Nature, which exploits stereospecific sequences
of amino acid monomers to form proteins with a myriad of
properties and functions. This lesson is further echoed in the
recent reports describing the unique behavior of sequenced
conventional polymer architectures, e.g., block copolymers, and
stereocontrol, e.g., isotactic polypropylenes.13-16 Finally, there
are clear advantages to be realized from the rearrangement of
“old” monomers to create new polymers: economic, in that the
infrastructure that already exists for the large-scale synthesis
of these old monomers translates into low cost and high
availability, and pragmatic, in that the knowledge base that
already exists regarding the suitability of these monomers for
particular applications translates into an efficient path to
application.
The extensive investigation of poly(lactic-co-glycolic acid)s
(PLGAs) for in ViVo applications requiring slow polymer
degradation renders them ideal candidates for the investigation
of the role of sequence in polymer properties. The popularity
of these polymers for applications such as stem cell scaffolding
and drug delivery vehicles stems from the fact that bulk
structures made from PLGAs hydrolyze at a moderate rate in
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10920 J. AM. CHEM. SOC. 2010, 132, 10920–10934
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Poly(lactic-co-glycolic acid) Microstructure
A R T I C L E S
Table 1. Naming Conventions for Segmers and Polymers
Symbol
Definition
L
LR
Lrac
Ld,rac
G
L-Lactic unit (S configuration)
D-Lactic unit (R configuration)
rac-Lactic unit
R-d-rac-Lactic unit
Glycolic unit
Gd2
Bn
SiR3
R-d,d-Glycolic unit
Benzyl protecting group
Silyl protecting group
structural and stereosequences more complex than those previ-
ously prepared. Selected examples are presented in Figure 1.
Previous efforts to prepare sequenced PLAs and PLGAs have
relied on the ring-opening polymerization (ROP) of the cyclic
lactide, glycolide, and methyl glycolide dimers.26-40 Despite
the laudable success achieved though the sophisticated design
of selective catalysts and the exploitation of chain-end control,
the number and types of sequences that can be prepared is
limited by the dimeric form of the ROP monomer and by
challenges inherent in programming catalysts to deliver a pattern
more complex than alternation. While the strategy we employ
in this paper of preassembling a sequenced segmer by a
condensation mechanism is arguably less efficient than ROP, it
is convergent and molecular weights >20 kDa are routinely
achievable. Most importantly this approach is general: any RSC
PLGA enVisioned can be prepared.
The first benefit realized from our creation of a family of
PLGA RSCs and the primary focus of this paper is the creation
of a partial “Rosetta Stone” for the interpretation of NMR data
of polymers with complex stereochemical patterns in general
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Figure 1. Comparison of complex sequences prepared by segmer assembly
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the body and the products, lactic and glycolic acid, are
bioassimilable.17-25 Although other polymers are employed for
these applications, studies on PLGAs and poly(lactic acid)s
(PLAs) represent a significant proportion of all work in the area.
The ubiquity of the materials and the special suitability of the
monomers to bioengineering applications make the idea of
creating repeating sequence copolymers (RSCs) from these
monomers particularly attractive.
Our approach to the investigation of the role of sequence on
the properties of PLGAs is to create and then analyze a family
of RSCs. Our methodology, which involves the condensation
of preformed segmers, allows for the synthesis of polymers with
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Stayshich and Meyer
Scheme 1
and for PLGAs in particular. The most relevant precedents for
this work are the extensive investigations of the NMR for PLA
of varying tacticities.29-31,33,41-53 It is worth noting that in the
PLA system the 13C NMR spectroscopic shifts have been found
to be most sensitive to the relationships between distant
stereocenters; differences in relative stereochemistries up to 5-6
units away from the nucleus under observation can be
detected.41,43,44,47,49,51,54 Although the NMR data for PLGAs
has likewise been studied, the added complexity introduced
by having two variables, sequence and stereochemistry, as
well as the fairly modest control of these variables achievable
using the common ROP synthetic approach, has inhibited
progress. These studies have primarily been limited to the
partial assignment of local sequence within otherwise random
copolymers.34,36,39,40,45,55-58 Our approach to preparing
PLGAs, which allows for nearly perfect sequence and
stereocontrol, greatly expands the database and offers thereby
a significant advance in the understanding of PLGA NMR
data, as well as some interesting new conclusions of a more
general nature pertaining to sequenced copolymers.
Starting from the previously reported dimer, Bn-LG-SiR3,59
catalytic hydrogenation was used to remove the benzyl group
and generate the partially deprotected LG-SiR3 unit in a 95%
yield (Scheme 1). Benzyl (S)-lactate (Bn-L) or benzyl (R)-lactate
(Bn-LR) were coupled to LG-SiR3 to generate diprotected
trimers Bn-LLG-SiR3 and Bn-LRLG-SiR3 in 84% and 72%
yields, respectively. Removal of both protecting groups gave
segmers LLG and LRLG in 83% and 67% yields. Using the
same approach, the segmers LG, LracG, GLG, GLracG,
LracLracG, LracLG, LLracG, and LLRG were prepared. Hexam-
eric units such as LRLGLLG were synthesized by coupling
partially deprotected trimeric segmers Bn-LRLG and LLG-SiR3
followed by subsequent deprotections in 75% yield overall.
LLGLLRG and GLGLR were prepared similarly from the
appropriate trimeric and dimeric precursors. It should be noted
that in the preparation of LracLracG from the racemic precursors,
there is a slight preference, 60:40, for the formation of LLG
units with different stereocenters, i.e., LLRG and LRLG are
(41) Zell, M. T.; Padden, B. E.; Paterick, A. J.; Thakur, K. A. M.; Kean,
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Results
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T. A.; Doscotch, M. A.; Siepmann, J. I.; Munson, E. J. Macromolecules
1997, 30, 2422–2428.
Naming Conventions. Segmers are named by listing the
monomers in sequence order from the C-side to the O-side using
the abbreviations in Table 1. BnLLracG is, therefore, a trimer
of stereopure L-lactic acid, rac-lactic acid, and glycolic acid
that bears a benzyl protecting group on the carboxylic acid (C-
side) terminus. Additionally, the deuterium-labeled lactic and
glycolic acids are labeled Ld,rac and Gd2, respectively. Polymers
are named from the exact segmer used. Thus, the polymer
prepared from LLG is named poly LLG rather than poly GLL
despite the homology of the two sequences after polymerization,
i.e., ... LLGLLGLLGLLGLLGLLG....
(44) Thakur, K. A. M.; Kean, R. T.; Hall, E. S.; Doscotch, M. A.; Munson,
E. J. Anal. Chem. 1997, 69, 4303–4309.
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2287–2296.
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2817–2824.
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53–59.
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Wade, C. G. J. Am. Chem. Soc. 2007, 129, 12610–12611.
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Chem. Phys. 1996, 197, 2627–2637.
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(53) Espartero, J. L.; Rashkov, I.; Li, S. M.; Manolova, N.; Vert, M.
Macromolecules 1996, 29, 3535–3539.
Synthesis. A series of PLGA RSCs based on dimeric, trimeric,
tetrameric, and hexameric repeating units have been prepared
by condensation of the preformed segmers. Segmers were
assembled in a convergent fashion by reacting partly protected
subunits to form completely protected products, which after
subsequent deprotection of both termini yielded the desired
segmers as R-hydroxy carboxylic acids. The synthesis of the
LLG and LRLG segmers are presented as representative
examples.
(54) Kasperczyk, J.; Bero, M. Makromol. Chem. 1993, 194, 913–925.
(55) Dobrzynski, P.; Kasperczyk, J.; Janeczek, H.; Bero, M. Macromol-
ecules 2001, 34, 5090–5098.
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Table 2. PLGA Repeating Sequence Copolymer Characterization Data
Scheme 2
tion of the fully deprotected LLG segmer. All polymers were
isolated as colorless solids by precipitation into methanol and
purified by reprecipitation from methylene chloride into metha-
nol to give yields ranging from 50% to 99% (Table 2). It should
be noted that we did not observe significant sequence prefer-
ences when assembling segmers with racemic units into
polymers. We have found that the inherent but slight preferences
for alternation of stereochemical centers that were observed in
selected segmer preparations can be minimized in the polymer
preparation by coupling segmers bearing L groups on one
terminus and G groups on the other.
Molecular weights for the polymers were determined by both
size exclusion chromatography (SEC) and multiangle laser light
scattering (MALLS). Relative molecular weights were obtained
by SEC in both THF and CHCl3 versus polystyrene standards
with the average molecular weights (Mn) ranging from 12.0 to
41.2 kDa in THF and 19.3 to 42.3 kDa in CHCl3. Absolute
favored over LLG and LRLRG. This preference can be clearly
seen in the copolymer spectrum as well. The synthesis of the
oligomer LracLG also results in a slight bias toward LRLG over
LLG, but it is difficult to detect by NMR once polymerized.
Complete synthetic schemes for all segmers and deuterium-
labeled compounds can be found in the Supporting Information.
Polymerization conditions, utilizing mild esterification re-
agents 1,3 diispropylcarbodiimide (DIC) and 4-(dimethylami-
no)pyridinium p-toluenesulfonate (DPTS), were adapted from
Stupp and Akutsu (Scheme 2).60,61 In this way, for example,
poly LLG was prepared by the DIC/DPTS-mediated condensa-
(61) Akutsu, F.; Inoki, M.; Uei, H.; Sueyoshi, M.; Kasashima, Y.; Naruchi,
K.; Yamaguchi, Y.; Sunahara, M. Polym. J. (Tokyo) 1998, 30, 421–
423.
(60) Moore, J. S.; Stupp, S. I. Macromolecules 1990, 23, 65–70.
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1
Figure 2. (Top) Full spectrum of poly LG. (Bottom) Expansions of selected regions for poly LG, LracG, and GLGLR. H NMR spectra at 600 MHz in
CDCl3.
molecular weights were determined for poly LG, GLG, LLG,
and LracLG using SEC-MALLS. The dramatic lack of correla-
tion between the SEC molecular weights in the two solvents
and the differences between these molecular weights and the
absolute molecular weights determined by MALLS makes it
clear that the Rg of the polymers is extremely sequence- and
solvent-dependent. The SEC molecular weights must therefore
be regarded with special care. By interpolation of the SEC and
SEC-MALLS data obtained, however, we can say with a high
Figure 3. Example of tetrad stereosequence encoding for poly LG (all
degree of confidence that the majority of the polymers prepared
have an absolute molecular weight >15 kDa, which corresponds
to a DP > 200 (based on the count of glycolic and lactic units).
This conclusion is further substantiated by the lack of end groups
observed in the NMRs of these polymers. Previous studies by
others on related polymers have shown microstructure-dependent
SEC behavior.62,63 A more detailed study of Rg versus sequence
is underway but is not the focus of this paper.
isotactic) and GLGLR (all syndiotactic).
Table 3. Tetrad Assignments for Stereosequences of Polymers
from the poly LG Series
The molecular weights of the RSCs, although lower than those
routinely achieved by ring opening of lactides and glycolides,
are respectable for a condensation polymerization on the 1-2
g scale that we are currently employing. Moreover, it worth
noting again that polymers with the sequence complexity of those
reported herein cannot be produced by any known ROP catalytic
system.
Microstructural Analysis of RSCs Prepared from Dimeric
Segmers. Although this article focuses on the polymers with
trimer-based sequences, poly LLG and poly GLG, we will
begin with a brief discussion of the simpler, dimeric RSCs based
on poly LG. Although a portion of this work was reported in
a previous article,59 we have since prepared a key member of
the series, poly GLGLR, that was not available at the time of
that publication.
surprisingly high sensitivity of the glycolic methylene protons
for the relative stereochemistry of the neighboring lactic units
(Figure 2). The difference in the methylene signals of isotactic
poly LG (δ 4.86 and 4.63, ∆ ) 0.23 ppm) and syndiotactic
poly GLGLR (δ 4.81 and 4.69, ∆ ) 0.12 ppm) are particularly
illustrative. Although each spectrum contains a pair of doublets,
as would be predicted for the clearly diastereotopic methylene
protons, the chemical shifts are dramatically different.
Our major discovery in the study of the alternating poly LxG
series polymers for poly LG, LracG and GLGLR was the
Each of these alternating copolymers possesses a unique
stereosequence that can be encoded using the widely accepted
convention of assigning relative stereochemistries in polymers
as either i for neighboring units with the same absolute
stereochemistry or s for the opposite. Using this coding system
and focusing on a tetrad level of resolution, the stereochemistry
(62) Kang, S.; Zhang, G.; Aou, K.; Hsu, S. L.; Stidham, H. D.; Yang, X.
J. Chem. Phys. 2003, 118, 3430–3436.
(63) Penco, M.; Donetti, R.; Mendichi, R.; Ferruti, P. Macromol. Chem.
Phys. 1998, 199, 1737–1745.
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A R T I C L E S
1
Figure 4. (Top) Full spectrum of poly GLG. (Bottom) Expansions of selected regions for poly GLG, GLracG, and GLGd2. H NMR spectra at 600 MHz
in CDCl3.
Microstructural Analysis of RSCs Prepared from Trimeric
GLxG Segmers. In moving from a dimer-based LG copolymer
to a trimer-based copolymer, additional complexity was intro-
duced both in the architecture and in the resulting NMR spectra.
The third monomer presents the possibility of two different
structural sequences, GLG or LLG, and multiplies the number
of possible stereoisomers. It should be noted again that the
choice to refer to a particular sequence as GLG rather than GGL
was based on the unit actually used in the synthesis. Once
polymerized, of course, such differences are irrelevant since poly
GLG would necessarily be the same in all respects as poly GGL
except in the identity of the end groups; both would have a
repeating sequence of -GGLGGLGGLGGLGGLGGL- in the
backbone.
of poly LG can, for example, be expressed as iii and that of
poly GLGLR as sss (Figure 3 and Table 3).
We focus on the tetrad level because we can clearly see this
level of resolution for some sequences present in the more
complex spectrum of atactic poly LracG (Figure 2). Using the
data from the stereopure polymers and from a sequence-
weighted copolymer prepared by mixing LG and LracG in a
60:40 ratio (see ref 59 for details on this copolymer; see Table
3 for the expected/observed tetrads), we were able to make a
nearly complete assignment of the signals. We see that the
methylene resonances divide into two regions based on the
relative stereochemistry of the closest neighboring lactic units:
i-centered (outer signals) and s-centered (inner signals). The
resonances for each of the s-centered tetrads, sss, ssi, iss, and
isi, can be clearly differentiated, while those for the i-centered
tetrads, iii, iis, sii, and sis, overlap such that the effective
resolution is expressed at a lesser diad or triad level.
Analysis of the NMR spectra of poly GLG in both the
stereopure and racemic forms required that we first assign
the resonances for the two chemically distinct G monomers.
The G units are inequivalent due to the intrinsic lack of
symmetry of the polyester chain, which has a distinct C- and
O-terminus for each polymer and for each sequence within the
polymer. We can differentially label the Gs then as either being
connected to the C-terminus of the lactic residue (GC) or the
O-terminus (GO). This inequivalence was expressed both in the
1H NMR spectrum (two pairs of doublets, Figure 4) and in
the 13C NMR spectrum (CdO, δ 166.5 and 166.4; δ 60.9 and
60.7, see Supporting Information for 13C NMR spectra). The
assignment of the resonances of poly GLG was facilitated by
comparison with the spectrum of the polymer that was selec-
tively deuterated at GO, poly GLGd2. The inner pair of doublets
The other resonances in the spectra of the dimeric polymers
also show stereosequence-dependent chemical shifts, though
none exhibit as high a level of sensitivity/interpretability as the
methylene protons. A clear chemical shift difference was noted,
for example, between the methine and methyl groups from
syndiotactic poly GLGLR and isotactic poly LG. The smaller
chemical shift range, however, limits the resolution of the
stereosequences present in poly LracG to the triad level. In the
13C NMR spectra (see Supporting Information) the L-carbonyl
resonances were clearly resolved to the triad level while the
G-carbonyl, L-methyl, and L-methine resonances were nearly
resolved triads. The s-carbonyl resonances for both L and G
appeared upfield relative to the i-stereoisomer. In contrast, the
s-methine and s-methyl resonances were downfield of the
i-versions. Interestingly, the methylene resonance, which was
1
were absent from the H NMR spectrum, and the downfield
methylene resonance was not present in the 13C NMR spectrum.
Further confirmation of the assignment came from the 2D
heteronuclear multiple bond coherence (HMBC) NMR spectrum
of poly GLG in which a 3-bond correlation of the L-carbonyl
with the outer pair of doublets from the methylene protons of
1
so sensitive in the H NMR data, was only a broad singlet in
the 13C NMR spectrum.
9
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1
1
Figure 5. 2D HMBC H-13C correlation NMR spectrum for poly GLG in CDCl3 (700 MHz, H; 175 MHz 13C). The detailed cross peaks correspond to
3-bond correlations between the L-carbonyl with the GC methylenes and the GO carbonyl with the L-methine.
GC was observed along with the correlation between the GO
carbonyl and the L-methine (Figure 5).41,46,64,65
Spectra of poly GLracG established that the chemical shifts
of the G resonances were only modestly sensitive to the tacticity
of the polymer. More sensitive than the methyl and methine
resonances, which exhibited only a slight broadening, the
methylene region exhibited 8 pairs of doublets, which given
the inequivalence of the two G units is consistent with a dyad
level of resolution. In other words the chemical shifts of the G
resonances depend only upon the relative stereochemistries of
the two closest L units. The peaks labeled as i-centered
resonances corresponded well with the isotactic poly GLG
standard. The slightly shifted pattern of two pairs of doublets
was assigned to the s-dyad. Partially resolved dyads were
likewise visible in the 13C NMR spectrum where the lactic
carbonyl appeared to broaden while 3 resonances, two at ca. δ
166.5 and a broadened resonance about δ 166.4, were observed
for the glycolic carbonyls.
Microstructural Analysis of RSCs Prepared from Trimeric
LxLxG Segmers. In moving from the poly GLG series to the
other trimeric series, poly LLG, we must again address the
nomenclature and consider the stereochemical implications of
the second L unit in the series. The two L units are necessarily
inequivalent and are designated as LO or LC in relation to the G
unit. LO is connected to the G through the O-terminus, and the
LC is connected through the C-terminus. Since the monomer
used to prepare these polymers is LLG rather than LGL,
however, the LO unit will be listed first in the series, i.e., LOLCG.
With these designations in mind, the stereochemical variants
of poly LxLxG can be considered. In Figure 6, our approach to
assigning tacticity patterns is illustrated. In the case of poly
LRLGLLG, for example, the sequence yields two types of
G-centered octads, one of which has an issiiss pattern. A subset
tetrad pattern of sii is recognized from the larger pattern if the
relationships from only the nearest four neighbor Ls are
considered. Interestingly, the same pattern can be reasonably
used as a reference for either LO or LC despite the shift in the
“center” of the sequence. The lack of symmetry in the number
Figure 6. Example of octad-level stereosequence encoding for poly
LRLSGLSLSG. The central tetrads for each type of unit are defined as
including one distant and two adjacent relationships.
of relationships considered relevant on each side of the L center
is countered by the fact that the relationships chosen for the
polyad maximize the number of adjacent versus distant interac-
tions, on the assumption (substantiated by the data) that the
adjacent relationships will be more influential than the distant
ones.
In considering the stereochemical possibilities for the poly
LxLxG series we have found it useful to divide the polymers
into three families: LO-variable (poly LxLG); LC-variable (poly
LLxG); and (LO + LC)-variable (poly LxLxG). Table 4 shows
the stereochemical possibilities for the first two families at the
tetrad, hexad, and octad levels. It is important to note that poly
LLG, LRLG, and LRLGLLG exhibit all of the stereochemical
variations possible in poly LracLG at the tetrad level. Likewise
poly LLG, LLRG, and LLGLLRG exhibit the possible tetrad
level variations in poly LLracG. Stereosequence-specific shading
in Table 4 illustrates the hierarchical relationship of the hexads
and octads to their parent tetrads.
(64) Bovey, F. A.; Mirau, P. A. Acc. Chem. Res. 1988, 21, 37–43.
(65) Agarwal, S.; Naumann, N.; Xie, X. Macromolecules 2002, 35, 7713–
7717.
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Poly(lactic-co-glycolic acid) Microstructure
A R T I C L E S
Table 4. Listing of Select Polyads for Specific poly LxLxG Repeating Sequence Copolymersa
a Tetrads subsets are identified by color within the hexad and octad tacticity patterns.
1
The H NMR spectra of the glycolic methylene regions of
these trimeric series are spectacularly informative: octad-leVel
resolution of indiVidual stereosequences is obserVed in some
cases. By comparison with the spectra of poly LLG, LRLG,
LRLGLLG, and LracLG for the LO-variable family (Figure 7)
and poly LLG, LLRG, LLGLLRG, and LLracG for the LC-
variable family (Figure 8), it was determined that the chemical
shifts of these resonances were inherently hierarchical and facile
to interpret.
controlled shift regions, the “fine” chemical shifts are then
determined by the relative stereochemistries of L units beyond
the central four.
The degree of resolution of longer range relationships in the
methylene region is clearly sequence-dependent. By focusing
on the upfield proton of the G methylene of poly LracLG, several
levels of resolution can be identified (Figure 9). In the case of
the iii set, for example, there are four nearly resolved doublets,
the number that would be expected if there was a doublet present
for each of the four possible octad level sequences. Although
we do not have standards with sufficient complexity to label
the individual sequences, the extremely high level of resolution
is striking. The other three sets of doublets in the same region
of this spectrum exhibit varying sensitivities ranging from the
tetrad to hexad to octad level.
A similar hierarchical pattern is observed in the methylene
region of poly LLracG (Figure 8). Sets of pairs of doublets
corresponding to the expected tetrads, iii, sss, ssi, and iis, can
be easily assigned. Within these sets, further resolution of certain
sequences is observed. It should be noted that the spectra of
poly LRLG and LLRG (Figures 7 and 8) are identical, as would
be predicted from their enantiomeric relationship.
Changing the NMR solvent from CDCl3 to d6-DMSO
dramatically decreases the resolution of the spectra (see Sup-
porting Information for figure depicting the difference). The
chemical shift range of the methylene (and other regions) is
decreased, and sequence resolution is reduced.
We were also able to make significant progress in assigning
the methyl and methine regions of the trimeric series of
polymers. These regions are inherently more difficult to interpret
because of the presence of the two inequivalent lactic units,
Beginning by focusing on the LO-variable family (Figure 7),
we can assign the resonances from the stereopure standards.
The spectrum of the simplest sequence, isotactic poly LLG,
exhibits a pair of well-separated doublets for the diastereotopic
G methylene protons. Poly LRLG, the fully syndiotactic
sequence, also exhibits a single pair of doublets, and poly
LRLGLLG exhibits two pairs of doublets, which is consistent
with expectations for this more complex sequence (see Table
4). Although these are stereopure standards with an “infinite”
pattern, the resonances are labeled in the figures at the tetrad
level for reasons that become clear in the analysis of the racemic
variants (Vide infra).
Analysis of the spectrum of poly LracLG reveals the useful
hierarchical nature of the shift pattern in the methylene region:
the gross shifts are determined by the central tetrad relationships,
while the fine shifts are determined by more distant ones. The
methylene region of this polymer manifests as well-separated
sets of pairs of doublets, one pair for each of the resolved
stereosequences. By comparison of the stereosequence standards
and the spectrum of poly LracLG, it is clear that the resonances
are grouped by the central tetrad relationships, iii, sss, iss, and
sii, into four well-separated regions. Within these tetrad-
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Stayshich and Meyer
Figure 7. 1H NMR spectra of LO-variable LLG polymers at 700 MHz in CDCl3. Comparisons of the expansions of selected regions for poly LLG, LracLG,
Ld,racLG, LRLGLLG, and LRLG.
the relatively small chemical shift range, and the lack of an
hierarchical shift pattern. Primary differentiation of LO versus
LC in the fully sequenced polymers poly LLG, LRLG, and
LRLGLLG was accomplished using a combination of 2D NMR
techniques. Methine resonances were assigned by 2D HMBC
NMR spectra where a correlation between the G carbonyl and
the LC methine was observed. Frustrating facile assignment, the
relative chemical shifts of the LO and LC methines were not
consistent throughout the stereochemical series. As Figure 10
illustrates, in isotactic poly LLG the LC resonance is upfield of
the LO resonance, but the order is reversed in syndiotactic poly
LRLG. In polymers that contain mixed stereochemical relation-
ships, poly LRLGLLG, and multiple mixed relationships, poly
LracLG and LracLracG, the 2D correlations showed that the
chemical shifts of the LO and LC methines were extremely
sequence-dependent and likely to invert chemical shift order
from sequence to sequence. By coupling the assignments of the
methine region with 2D COSY NMR spectroscopy, however,
the methyl region could be assigned. Unlike the methine
resonances, the LO methyl resonances were always downfield
of the corresponding LC resonances.
homonuclear decoupling of the regions impractical, while the
small chemical shift range and sequence-dependent shift inver-
sions (Vide supra) hindered the interpretation of 2D spectra.
The relatively modest simplification of the spectra for the
selectively deuterated polymers poly Ld,racLG, and LLd,racG,
however, establishes the sensitivity of these signals for the
stereochemical relationships of lactic units beyond the tetrad
level in the chain. Moreover, by simple counting of the
resonances and comparison of the spectra with the stereose-
quence standards, it is clear that some sequences are actually
resolved to a hexad or even octad level.
Chemical shifts in the 13C NMR spectra of poly LracLG and
LLracG showed primarily a tetrad level of resolution. Partial
assignment was accomplished by comparison with the stereopure
and deuterated RSCs (Figures 11 and 12) and by 2D HMBC
NMR as described earlier. Analysis of the stereopure RSCs
established that the LO carbonyl resonance was generally
downfield of the LC resonance and the L carbonyl resonances
were more sensitive to stereochemistry than the 13C NMR
resonances for other groups. The presence of >8 lactic carbonyl
signals in the spectra of both poly LracLG and LLracG was
consistent with a higher than tetrad (mixed hexad/octad) level
of resolution, although it was not possible to make individual
assignments due to overlap. It is also of note that, in both the
L- and G-carbonyl regions, the i-centered resonances were
downfield of the s-centered resonances. Methine LO and LC
resonances were assigned by comparison to the deuterium-
labeled polymers, as the deuterium-substituted carbons exhibited
Although complete interpretation of the methine and methyl
regions for poly LracLG and LLracG could not be accomplished,
it was clear that the sequence resolution of both regions was
greater than a tetrad level. It should be noted that neither
homonuclear decoupling nor 2D NMR was of much use in
interpreting these spectra further. The chemical shift envelope
for each type of signal was large enough to render the complete
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Poly(lactic-co-glycolic acid) Microstructure
A R T I C L E S
Figure 8. 1H NMR spectra of LC-variable LLG polymers at 700 MHz in CDCl3. Comparisons of the expansions of selected regions for poly LLG, LLracG,
LLd,racG, LLGLLRG, and LLRG.
Figure 9. 1H NMR expansion for the upfield diastereotopic proton of the glycolic methylenes of poly LracLG (full spectrum in Figure 7). The level of
sensitivity for sequence ranges from tetrad (sii, iii) to hexad (sss) to octad (iss) depending on the core tetrad sequence.
much lower intensities due to slow relaxation. These signals
clearly resolved to a slightly more than tetrad level for RSCs
incorporating racemic units. The methyl region also showed a
greater than tetrad level of resolution, but the small chemical
shift range made individual assignments in this region impractical.
The chemical shifts for stereopure polymers are compiled in
Tables S1 and S2, which are located in the Supporting
Information. It should be emphasized that these are stereopure
samples, and exact matches to unknowns will not be expected
unless the sequences of the spectra compared are homologous
at or beyond the level of resolution observed. For convenience,
however, the polyads with more complex sequences have been
labeled at the tetrad level.
Using these spectral tables as a guide and focusing on the
methylene region, it is possible to interpret the spectra of
samples that contain more than one structural sequence and of
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is some degradation of the samples in the experiment. The full
spectra for all samples can be found in the Supporting
Information.
Thermal Properties. Differential scanning calorimetry (DSC)
was performed on polymer samples and annealed films. The
glass transition temperature (Tg) varied slightly due to changes
in the sequence composition (Table 5). When the L content
increased from 50% in poly LG to 67% in poly LLG the Tg
remained constant around 57 °C. However, increasing the G
content to 67% in poly GLG the Tg decreased slightly to 50
°C. Within the LG and LLG series the Tg was dependent on
stereochemistry. Isotactic polymers possessed the highest Tg’s
(57 °C), the atactic polymers possessed intermediate Tg’s
(55-51 °C) and the syndiotactic polymers possessed the lowest
Tg’s (48-50 °C). For the higher G content GLG series, changes
in stereochemistry did not change Tg’s; poly GLG and GLrac
G
both exhibit a Tg of 50 °C. Interestingly, melting transitions
were only found for the syndiotactic LLG polymers, poly LRLG
andLLRG, at 154 and 158 °C, respectively.
Polymer films were prepared by drop-casting from methylene
chloride into DSC pans, drying under vacuum and annealing at
85 °C for 3 h. Annealing had little effect on the transitions for
poly LLRG and LRLG, but the Tg’s for all of the remaining
annealed samples dropped and became less sequence-sensitive.
A new melting transition appeared for poly LLG at 114 °C,
which is much lower than that observed for poly LLRG and
LRLG.
Although direct homologues of these RSCs have not been
previously reported, the behavior of these sequences can be
examined relative to the known PLA sequences. Coates and
co-workers have reported the Tm for syndiotactic PLA as 153
°C, which is close to those of poly LLRG and LRLG. The Tm’s
for isotactic and stereocomplexed PLA are much higher at 175
and 230 °C, respectively.30,31
Given the regularity of the remaining stereopure polymers,
the lack of crystallization is somewhat surprising. Although
annealing did promote crystallization in the case of poly LLG,
other sequences did not exhibit melting points despite repeated
efforts. We do not rule out the potential for crystallinity,
however, as we suspect that we may not have discovered the
proper thermal conditions to promote the longer range organiza-
tion of the chains. Notably, Coates and co-workers did not
observe crystal formation for the predominantly heterotactic
sequenced PLA.31
1
Figure 10. 2D HMBC H-13C correlation NMR spectra at 700 MHz in
CDCl3 for poly LLG (A), LRLGLLG (B), LRLG (C), LracLracG (D), and
LracLG (E).
samples that contain multiple stereosequences. In a sample
comprising a mixture of different structural sequences, in this
case a mixture of poly LG, GLG, and LLG, resonances for
each sequence can easily be distinguished (Figure 13).
A more stringent test is the assignment of the methylene
region of poly LracLracG. This spectrum is complicated because
of the number of possible polyads (8 at the tetrad level) and
the presence of multiple levels of resolution for each polyad.
Nevertheless, it is possible to make a fairly complete assignment
of the sequences present at the tetrad level (Figure 14).
MALDI-TOF Mass Spectroscopy. MALDI TOF analysis
confirms the sequence fidelity of the PLGA RSCs. Using
conditions optimized for PLGAs,66,67 the analysis of selected
copolymers was carried out. The masses of the chains present
in poly LG, for example, differ in molecular weight by
increments of exactly 58 amu, corresponding with the glycolic
unit, and 72 amu, corresponding to the lactic unit (Figure 15).
Moreover, chains differing by exactly one segmer weight (L +
G ) 130 amu) dominate, which is consistent with synthesis by
assembly of segmers. Analogously, the envelope of masses
present in the spectrum of poly GLG differ in molecular weight
by a pattern of G, L, and G molecular weights with a majority
of chains differing by exactly a segmer weight. Unfortunately,
an analysis of the absolute molecular weights did not correspond
to chemically reasonable end groups. It seems likely that there
Discussion
We are able routinely to prepare RSC PLGAs with DPs >
200 on a per monomer basis with a greater than 95% sequence
fidelity. Previously, we were successful in preparing RSC
PLGAs using DCC/DMAP but the DPs were significantly
lower.59 The newly adopted DIC/DPTS method, first developed
by Stupp and Moore, utilizes DPTS instead of DMAP in order
to suppress chain terminating N-acylurea byproducts, neutralize
the reaction pH, and minimize depolymerization.60,68 Similar
methods were used by Hawker and co-workers in the preparation
of well-defined oligomers of lactic acid and caprolactone.69,70
Examination of the NMR spectra establishes that under the
(68) Nederberg, F.; Connor, E. F.; Glausser, T.; Hedrick, J. L. Chem.
Commun. 2001, 2066–2067.
(66) Huijser, S.; Staal, B. B. P.; Huang, J.; Duchateau, R.; Koning, C. E.
Angew. Chem., Int. Ed. 2006, 45, 4104–4108.
(69) Takizawa, K.; Nulwala, H.; Hu, J.; Yoshinaga, K.; Hawker, C. J. J.
Polym. Sci., Part A: Polym. Chem. 2008, 46, 5977–5990.
(70) Takizawa, K.; Tang, C. B.; Hawker, C. J. J. Am. Chem. Soc. 2008,
130, 1718–1726.
(67) Huijser, S.; Staal, B. B. P.; Huang, J.; Duchateau, R.; Koning, C. E.
Biomacromolecules 2006, 7, 2465–2469.
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Poly(lactic-co-glycolic acid) Microstructure
A R T I C L E S
Figure 11. 13C NMR spectra of LO-variable LLG polymers at 175 MHz in CDCl3. Comparisons of the expansions of selected regions for poly LLG,
LracLG, Ld,racLG, LRLGLLG, and LRLG.
Figure 12. 13C NMR spectra of LC-variable LLG polymers at 175 MHz in CDCl3. Comparisons of the expansions of selected regions for poly LLG,
LLracG, LLd,racG, LLGLLRG, and LLRG.
specified reaction conditions, neither sequence scrambling nor
epimerization are significant problems, although in select
samples mistakes (<5%) can be observed. The spectrum of poly
LG (Figure 2, methylene region), for example, shows contami-
nation by syndiotactic sequences.
generate preferential packing domains.39 Others have reported
that as glycolic content increases, the Tg drops to as low as 36
°C at a 50% glycolic unit incorporation.39,56,71 RSC PLGAs,
on the other hand, maintained Tg’s at or above 50 °C even when
the glycolic unit content exceeded 60%. As long as the
Sequence was found to have a dramatic effect on the thermal
properties of PLGA. In random PLGAs the material properties
are primarily dependent on the length of the lactic blocks that
(71) Park, P. I. P.; Jonnalagadda, S. J. Appl. Polym. Sci. 2006, 100, 1983–
1987.
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1
Figure 13. Glycolic methylene region of a mixed H NMR spectrum for mixed sample (1:1:1) of poly LG, GLG, and LLG at 600 MHz in CDCl3.
Figure 14. Glycolic methylene region of poly LracLracG at 700 MHz in CDCl3.
uniformity of the structural sequence remained, altering the
stereosequence only slightly lowered the Tg. Most of the
polymers remained amorphous even after annealing. Poly LLG,
LRLG, and LLRG were the only polymers that displayed a
melting transition, and poly LLG only crystallized after
annealing. Although sequence and stereoregularity should
promote crystallinity, it is clear that it is challenging for these
polymers to exhibit long-range order based on these relatively
short sequences.
family of RSCs, not only to make assignments but also to
develop a deeper understanding of the issues involved in the
interpretation of NMRs of RSCs.
Although the complexity of PLGAs makes the assignment
of NMR resonances more challenging than in PLAs, our studies
have also shown that the copolymer structure offers compensa-
tory advantages that facilitate interpretation. The most important
advantage offered by the copolymer is the exquisite sensitivity
of the methylenic protons of the glycolic units to sequence and
stereochemistry. The chemical shift range is relatively large and
the shifts appear to depend on stereosequence in a hierarchical
fashion: the gross chemical shift is determined by the central
polyad, while the fine chemical shift depends on longer range
relationships. The sensitivity of this signal to stereosequence is
much greater than any other 1H NMR resonance or the
classically more informative 13C NMR resonances. The second
compensatory advantage of PLGAs over PLAs (and other well-
studied vinyl polymers) is a function of the fact that the
resonances for the two monomers are necessarily distributed
over a larger chemical shift range than the resonances for any
single monomer and the fact that the polymers are unsymmetric;
each monomer and each polymer have distinct C and O termini.
Using 2D NMR, these differences were exploited to allow for
the definitive assignment of resonances.
Sequence and NMR. Our synthetic approach, which allowed
for the creation of an unprecedented array of polymer micro-
structures, facilitates the understanding of the NMR assignments
for specific PLGA sequences. The polymers prepared serve as
a partial Rosetta Stone for the assignment of PLGA NMR data.
Prior assignments of the NMR resonances for PLGA have been
based on random copolymers (and a very small set of easily
prepared well-defined homo- and copolymers). While this
approach has led to significant understanding for a variety of
polymers, it is worth noting that controversies over assignments
of stereochemistry in the closely related but obviously simpler
PLA system have taken years and the input of several groups
to resolve.41,43,46,72 The fact that PLGA has the potential for
both structural and stereosequence variability complicates the
issue significantly. However, we have been able, using our
The most intriguing single result of the NMR studies of these
RSC PLGAs, one that is applicable beyond the narrow scope
of these polymers, is the unambiguous demonstration that
(72) Chisholm, M. H.; Iyer, S. S.; Matison, M. E.; McCollum, D. G.; Pagel,
M. Chem. Commun. 1997, 1999–2000.
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Poly(lactic-co-glycolic acid) Microstructure
A R T I C L E S
Figure 16. Simulated NMR spectra highlighting the challenges inherent
in comparing a standard with perfect stereosequence control to a sample
with multiple resolved sequences. Although all sequences share the same
“iii” central tetrad and exhibit similar chemical shifts, the shifts of the nearly
hexad-resolved sequences create a pattern that does not show an easily
interpretable correspondence with the standard.
the iss tetrad is a simple doublet with no fine structure; this
signal is resolved only to the tetrad level. The next group is a
barely resolved pair of doublets that corresponds to the gross
shift of the sss tetrad but which is resolved to a hexad level.
The final groups, which exhibit gross chemical shifts consistent
with iii and sii tetrads, exhibit four nearly resolved pairs of
doublets that correspond to the much higher octad level of
resolution. An octad level of resolution maps to an impressive
distance along the backbone of 31 atoms between the most
separated stereocenters. Such sequence-dependent resolution
occurs throughout these spectra, although individual signals
exhibit varying levels of sensitivity.
Figure 15. MALDI TOF patterns for poly LG (top) and poly GLG
(bottom).
Table 5. Thermal Properties for RSC PLGAs
precipitated
annealeda
polymer
Tg (°C)
Tm (°C)
Tg (°C)
Tm (°C)
LG
racG
GLGLR
GLG
GLracG
LLG
LRLG
LLRG
LLGLLRG
LRLGLLG
57b
55b
50b
50b
50b
57b
50b
48b
52b
52b
51b
51b
53b
ND
ND
ND
ND
ND
ND
154c
158c
ND
ND
ND
ND
ND
49c
ND
ND
L
48c
Another important lesson learned from the study of RSC
PLGAs is that there are some inherent limitations in our ability
to make accurate assignments, limitations that are general to
the use of exact sequence RSCs as a key to the interpretation
of the NMR spectra of samples with mixed sequence chains,
no matter what polymer is involved. The first order approach
to assigning spectra, one that works in some cases, is to assign
the resonances for specific sequences by comparison of the
chemical shifts with those of a stereopure standard. Our analysis
of the extensive database of shifts for the polymers described
herein, however, highlights the potential for misinterpretation
in systems where the sensitivity of a resonance to stereochem-
istry is high and not yet understood. The difficulties are
illustrated in the simulated spectrum depicted in Figure 16. If a
stereosequence with only “i” relationships, for example, is
compared to a sample that contains several stereosequences,
no peak with a 1:1 shift correspondence to the standard will be
found despite the fact that the sample contains a significant (25%
in this case) proportion of the “i” sequence in the form of “iiiii”
at a hexad level of resolution. A similar problem would occur
if the mixed sample contained only iiiis and siiis sequences.
Despite the fact that both share the same iii central tetrad, the
shift of neither peak would correspond with that of the “all i”
standard.
43c
50c
ND
114c
48c
48c
48c
47c
48c
48c
155c
ND
ND
ND
ND
ND
L
L
racLrac
racLG
G
LLracG
a Polymer films were drop-cast into DSC pans, dried under vacuum,
and annealed at 85 °C for 3 h. b Transitions were measured in the
second heating cycle. c Transitions were measured in the first cycle.
chemical shift resolution in a polymer bearing multiple se-
quences depends on the exact sequence. Although investigators
who have previously analyzed polymers with complex stereo-
sequences have proposed shift assignments based on different
levels of resolution in some cases,42,47 the range of sequence-
specific polymers available in our system gives a particularly
dramatic validation of the hypothesis. The clearest illustration
in our system was found in the methylene region of poly
LracLG. Examining the upfield proton, four groups of doublets
are visible (Figure 9). The leftmost group that corresponds to
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The lack of correlation between stereopure standards and the
resonances for individual sequences can thus be attributed to a
combination of two phenomena: (1) a full set of standards is
not available, i.e., sequences longer than those prepared as
standards can be distinguished in mixtures and (2) the ultimate
sensitivity of the chemical shift for a particular sequence in a
mixture, e.g., tetrad, hexad, etc., is determined by the inherent
resolution of the NMR spectrum. We see the effects of both of
these phenomena in our spectra in that the shifts of our standards
do not exactly match those observed in our mixed sequence
polymers. To achieve a 100% correlation, it would have been
necessary in the case of poly LracLracG, for example, to prepare
all 32 possible octad sequences. As the synthesis of all of these
sequenced polymers is impractical, it was necessary to extrapo-
late from the observed trends from the available standards in
assigning certain sequences. The second phenomenon that results
in chemical shift miscorrelation, the fact that the certain chemical
shift differences are on the same order of magnitude as the peak
widths, is a universally recognized spectroscopic challenge and
could be overcome by a combination of expanding the database
of sequences and “fitting” the mixed spectra. Although the lack
of long-sequence standards prevented an analysis of this depth,
these phenomena were taken into account when assignments
were made.
of an amino acid into a peptide to determine the importance of
a particular amino acid to the tertiary structure of a protein.73,74
Although we cannot yet correlate our NMR spectra with
specific conformational preferences the data suggests that there
is much to be learned. The fact that we observe a sequence-
specific sensitivity to stereochemistry is, for example, intriguing.
Such behavior could arise either because specific sequences
create conformations that place the diastereotopic methylene
protons in better positions to “observe” distant stereochemical
relationships, or it could arise because certain sequences simply
have stronger conformational preferences (or both). Also
relevant to the understanding of the rules governing the preferred
conformations in this system is the observation that the chemical
shift difference between the two methylenic protons on a
particular carbon was consistently smaller for s-centered polyads
than i-centered polyads. An analogous trend has been well-
documented in vinyl polymers such as polypropylene.75,76
Conclusions
The solution-phase conformations for RSC PLGAs were
found to be extremely sequence- and stereosequence-dependent,
analogous to peptides. To access these RSC PLGAs, we have
developed a methodology that allows the preparation of complex
sequenced copolymers of lactic and glycolic acids from simple
monoprotected building blocks. The method is general and
convergent and allows for the synthesis of sequences unavailable
by other methods such as ROP. Although RSC synthesis can
be used to make stereopure NMR standards, the inherent
resolution of specific sequences must be taken into account as
the level of resolution for a particular nucleus may be sequence-
dependent. Improved thermal properties were also a direct result
of the uniformity of polymer sequence. We plan to extend this
work by undertaking conformational modeling and bulk char-
acterization studies.
The broad manifestation of these chemical shift correlation
phenomena in our data can readily be observed in the stacked
1
plots of H NMR spectra for the PLGA RSCs (Figures 8 and
9). The fact that peaks labeled as arising from the same tetrad
do not always “line up” is due to the peak matching problems
just described, not to poor chemical shift calibration. It should
be noted that these issues disproportionally complicate the
interpretation of the methine and methyl regions because those
regions span a smaller chemical shift range and are not as
hierarchical as the methylene region.
The analysis of the NMR data for these polymers has also
given us some insight into the specific interpretation of the
spectra of PLGAs. In particular, we note that the shift range
observed for differing stereosequences within the same structural
sequence overlaps exactly with the chemical shift range observed
for differing structural sequences. Given the similarity of the
monomers involved, it is perhaps not surprising that structural
sequence does not introduce a larger perturbation. The bottom
line is that, for PLGAs, stereosequence is extremely important
in determining the NMR spectral pattern.
Acknowledgment. This research was supported by NSF
(0809289), Research Corp (RA0356), and the University of
Pittsburgh. The authors would like to thank Drs. Robbert Duchateau
and Saskia Huijser for MALDI-TOF analysis, Dr. Krishnan
Damodaran for NMR consultation, Dr. Joel Gillespie for materials
characterization, Jian Li for synthetic contributions, and Drs. David
J. Earl, Megan M. Spence, W. Seth Horne, and Mr. Benjamin N.
Norris for helpful discussions. The authors also acknowledge NIH
grant 1S10RR017977-01 for support of TOF MS.
Sequence and Conformation. One inescapable conclusion to
be drawn from the solution-phase behavior of these PLGA
RSCs, both NMR and SEC, is that the conformations of these
polyesters are sequence-dependent. Conformational differences
must, for example, be responsible for the differences in chemical
environment exhibited by the diastereotopic methylene protons
imbedded in different stereosequences. Conformation is also
likely to be responsible for the sequence dependence of the SEC;
the Rg is determined by the shape assumed by the polymer in
solution.
Supporting Information Available: Detailed procedures and
full characterization of all synthetic intermediates and polymers
are provided, along with figures for the 13C spectra of poly GLG
and LG series, the 1H spectra for selected polymers in DMSO,
and full MADLI-TOF spectra for poly LG and GLG. This
material is available free of charge via the Internet at http://
pubs.acs.org.
JA102670N
Although the homology of the monomers in these polymers
with those in amino acids, specifically alanine and glycine,
renders the sequence-dependence of the conformations unsur-
prising, the fact that the effect is so strong, given the lack of
amide-mediated hydrogen bonding, is perhaps less expected.
Indeed, it is common practice to substitute the ester analogue
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10934 J. AM. CHEM. SOC. VOL. 132, NO. 31, 2010