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1995 Synthesis, Characterization, Liquid Crystals and Crosslinking of Poly(gamma-Alkyl-Alpha,L- Glutamates). Javier Nakamatsu Louisiana State University and Agricultural & Mechanical College
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Recommended Citation Nakamatsu, Javier, "Synthesis, Characterization, Liquid Crystals and Crosslinking of Poly(gamma-Alkyl-Alpha,L-Glutamates)." (1995). LSU Historical Dissertations and Theses. 6123. https://digitalcommons.lsu.edu/gradschool_disstheses/6123
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A Bell & Howell Information Company 300 North Zeeb fload. Ann Arbor. Ml 48106-1346 USA 313/761-4700 800/521-0600 SYNTHESIS, CHARACTERIZATION, LIQUID CRYSTALS AND CROSSLINKING OF POLY(y-ALKYL-oc,L-GLUTAMATES)
A Dissertation
Submitted to the Graduate Faculty of the Louisiana State University and Agricultural and Mechanical College in partial fulfillment of the requirements for the degree of Doctor of Philosophy
in
The Department of Chemistry
by Javier Nakamatsu .S., Pontificia Universidad Catolica del Peru, 1989 December 1995 UMI Number: 9618313
UMI Microfonn 9618313 Copyright 1996, by UMI Company. All rights reserved.
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UMI 300 North Zeeb Road Ann Arbor, MI 48103 ... facts are meaningless, unless meanings are put into them...
from “Dr. Zhivago” B. P astern ak ACKNOWLEDGMENTS
I would like to express my appreciation to my major Professor, Dr.
William H. Daly. He is an interminable source of knowledge and imagination. Thanks also to Dr. Paul S. Russo for sharing his time and effort in the characterization and physical chemistry studies done in this research work.
I want to thank Dr. loan I. Negulescu for his invaluable help, especially at the beginning of my research. I would also want to thank Dr.
Mark McLaughlin and Dr. Kevin Robbins for serving as members of my com m ittee.
1 would like to acknowledge the financial support of the Louisiana
State University Chemistry Department and the NSF.
I will never forget the support and friendship of the members of the
Daly and R usso groups: Dr. Cecil G eiger, Dr. Z haoyao Qiu, Dr. Doris
C ulberson, Dr. Larry Morris, Dr. Daewon Sohn, Dr. Jav ier M acossay,
Xiaoian Wang, Jack Davies, Tim Evenson, Melissa Manuszack, Michael
Baylis, Lucille Smith, Keunok Yu, and Brian Fong. I am very grateful to the undergraduate students who also helped to the completion of this work,
Christian Negulescu, Donnell Ward, Hal Traxler, and very special thanks to
Michael Clark and Holly Ricks. Living far away from family and friends is not easy. However, it is
very satisfying to realize that there are people that one can call friends
everywhere, they make adaptation easier. I will always be in debt to all the
people I have befriended, they have become part of me. No doubt this is the most important thing I will take with me from my years in graduate school. Among them I would like to thank Carl Snyder and Dr. Marcelo
Saraiva for their friendship. A very special thanks to Kevin Lacour and his family, for all these years of support and thoughtfulness.
Finally, I would like to thank my family, who has always supported me unconditionally and across the distance.
iv TABLE OF CONTENTS
EPIGRAPH ...... ii
ACKNOWLEDGMENTS...... iii
LIST OF TABLES...... vii
LIST OF FIGURES ...... viii
ABBREVIATIONS...... xi
ABSTRACT...... xiv
CHAPTER 1: LITERATURE REV IEW ...... 1 1.1 High M odulus P o ly m ers ...... 2 1.2 Poly(y-alkyl-a,L-glutamates) ...... 4 1.2.1 Poly(y-benzyl-a,L-glutamate) ...... 8 1.2.2 Polymerization of NCAs...... 9 1.2.3 Applications and Potentials of PALGs...... 15
CHAPTER 2: SYNTHESIS AND CHARACTERIZATION OF PSLG 21 2.1 Introduction ...... 22 2.2 Synthesis ...... 23 2.2.1 Synthesis of PSLG ...... 23 2.2.2 Synthesis of Poly{y-benzyl-a,L-gliitamate)-co- poly(y-stearyl-a,L-glutamate) ...... 28 2.2.3 Silylated Amines as Initiators...... 28 2.2.4 Study of Coinitiators ...... 30 2.2.5 Synthesis of Labeled PSLG ...... 34 2.3 Characterization ...... 36 2.3.1 Fractionation ...... 36 2.3.2 Gel Permeation Chromatography...... 38 2.3.3 Intrinsic V iscosity...... 42 2.3.4 Static Light Scattering ...... 46 2.4 Experimental...... 50
CHAPTER 3: LIQUID CRYSTALS...... 59 3.1 Introduction...... 60 3.1.1 Theory of Liquid Crystals ...... 63 3.1.2 Cholesteric Superstructure ...... 68 3.2 Liquid Crystals of PSLG ...... 74 3.2.1 Phase Boundary Studies ...... 74 3.2.2 Effect of Concentration and Temperature on the Pitch ...... 81 3.3 Experimental...... 84
CHAPTER 4: MODIFICATION AND CROSSLINKING OF PBLG ...... 86 4.1 Introduction ...... 87 4.2 Reaction on the Side Chains of PBLG ...... 88 4.2.1 Cleavage of Small Esters with Trimethylsilyl Iodide ...... 92 4.2.2 Activation of PBLG Side C hains ...... 93 4.2.3 Reaction of Activated PBLG with /?-Butyl Isocyanate ...... 95 4.3 Crosslinking of PBLG ...... 97 4.3.1 PBLG Isotropic G els...... 101 4.3.2 PBLG Liquid Crystal G els ...... 108 4.4 Experimental...... 111
BIBLIOGRAPHY...... 116
VITA...... 121 LIST OF TABLES
2.1 Results from the polymerizations by different silylated amine initiators...... 31
2.2 Results from fractionation of PSLG ...... 40
2.3 Results of characterization of monodisperse samples of PSLG 49
2.4 Study of silylated amine initiators ...... 52
2.5 Polymer recovered from fractionation ...... 56
3.1 PSLG of Mw=239,000 (MWD=1.13) in THF at 25°C ...... 76
3.2 PSLG of Mw=208,000 (MWD=1.08) in THF at 25°C...... 76
3.3 PSLG of Mw=129,000 (MWD=1.12) in THF at25°C ...... 76
3.4 Comparison of experimental results and theoretical predictions for the biphasic region (concentrations in v/v% )...... 80
vii LIST OF FIGURES
1.1 PMLG, PBLG and PSLG ...... 7
1.2 Fuzzy rod ...... 8
1.3 N-carboxyanhydrides: methyl, benzyl and stearyl glutamates 10
1.4 Primary amine (i) and carbamate (ii) mechanisms...... 12
1.5 Activated monomer mechanism ...... 14
2.1 IR spectra of SLG, NCA-SLG and PSLG...... 2 5
2.2 1H-NMR spectrum of PSLG in CDCU ...... 27
2.3 1H-NMR spectrum of poly(y-benzyl-a,L-glutamate)-co- poly(Y-stearyl-a,L-glutamate) in CDCI3 ...... 29
2.4 GPC chromatograms of polymerizations by different silylated amine initiators...... 31
2.5 Propagation via the activated monomer mechanism...... 32
2.6 Initiation steps via the activated monomer mechanism ...... 33
2.7 PDMS spectra of (A) 15:1 monomer:coinitiator ratio, chlorosulfonyl isocyanate coinitiator; (B) 10:1 monomer:coinitiator ratio, 4-chlorophenyl isocyanate...... 35
2.8 Dye fluorescein isothiocyanate, isomer I ...... 36
2.9 GPC chromatograms of PSLG after fractionation...... 41
2.10 Viscosity plot of PSLG of Mw=239,000 (MWD=1.13) in THF at 25°C ...... 44
2.11 Viscosity plot of PSLG of Mw-208,000 (MWD=1.08) in THF at 25°C ...... 4 4
2.12 Viscosity plot of PSLG of Mw=129,000 (MWD-1.12) in THF at 25°C ...... 45
viii 2.13 Mark-Houwink plot of PSLG in THF at 2 5 ° C ...... 46
2.14 Zimm plot of PSLG (Mw-208,000; MWD=1.08) in THF...... 49
3.1 Liquid crystalline phases ...... 61
3.2 Schematic phase diagram of lyotropic liquid crystals...... 67
3.3 Cholesteric liquid crystal superstructure ...... 70
3.4 Cholesteric liquid crystal PSLG in THF at 25.0°C (Mw=239,000; MWD=1.13;
3.5 Spherulites with maltase-crosses of PSLG in THF at 25.0°C (Mw=129,000 ; MWD=1.12 ; <(>=21.0%)...... 79
3.6 Distance between striations vs. temperature, PSLG liquid crystal solution (Mw=208,000), 28.4% (v/v) in TH F ...... 82
3.7 Distance between striations vs. concentration, PSLG (Mw=208,000) in THF at 25°C ...... 84
4.1 Reaction of an ester with trimethylsilyl iodide ...... 91
4.2 Reaction of benzyl acetate with trimethylsilyl iodide ...... 93
4.3 1H-NMR of reaction of benzyl acetate with trimethylsilyl iodide in CDCI3 ...... 94
4.4 Activation of the side chains of PBLG with trimethylsilyl iodide ...... 95
4.5 1H-NMR of PBLG and PBLG reacted with trimethylsilyl iodide, in CDCI3 ...... 96
4.6 Reaction of n-butyl isocyanate with the partially silylated PBLG ...... 97
4.7 ’H-NMR of PBLG reacted with n-butyl isocyanate, in CDCI3 ...... 98
4.8 Scheme of the crosslinking of activated PBLG with 1,6-diisocyanato hexane ...... 102
ix 4.9 Change in hydrodynamic radius and apparent diffusion coefficient as crosslinking of PBLG occurs ...... 105
4.10 Change in hydrodynamic radius and apparent diffusion coefficient as crosslinking of poly(BLG)-co-poly(SLG) occurs 105
4.11 Diffusion coefficient distribution for the crosslinking of (A) PBLG and (B) poly(BLG)-co-poly(SLG) (see text for explanation)...... 106
4.12 (A) Logarithmic plot and (B) semi-log plot of the crosslinking of poly(BLG)-co-poly(SLG)...... 109
4.13 (A) Isotropic network of rod-like polymers, and (B) Anisotropic network of rod-like polym ers...... 110
4.14 Setup for crosslinking under electric field ...... 111 ABBREVIATIONS
A Angstrom s a 2 osmotic second virial coefficient
BLG y-benzyl-a, L-glutamate c concentration
D«pP apparent diffusion coefficient
DCM dichloromethane
Df fractal dimension dL deciliter
DMF N.N-dimethylformamide
DMSO deuterated dimethylsulfoxide
DP degree of polymerization
Fig. Figure g gram s
GPC gel permeation chromatography
I.D. internal diameter
IPN interpenetrating polymer network
ITO indium-tin oxide
LB Langmuir-Blodgett h hours
1H-NMR proton nuclear magnetic resonant
xi IR infra-red kG kilo G au ss m multiplet min m inutes mL milliliter
Mw weight-average molecular weight
MWD molecular weight distribution
NCA N-carboxyanhydride nm nanometer
P pitch
PALG poly(y-alkyl-a, L-glutamate)
PBLG poly(y-benzyl-oc, L-glutamate)
PDMS plasma desorption mass spectroscopy
PMLG poly(y-methyl-a,L-glutamate) ppm parts per million
PSLG poly(y-stearyl-a, L-glutamate)
PTFE polytetrafluoroethylene
Rg radius of gyration A V 73
N z-average radius of gyration (O
S periodicity (1/2 pitch) s singlet
SEC size exclusion chromatography
xii SLG y-stearyl-a, L-glutamate
t time
TFA trifluoroacetic acid
THF tetrahydrofuran
UV ultraviolet
V volts
v volume w weight pm m icrom eter
fo] intrinsic viscosity
T)*P specific viscosity
Tirol relative viscosity
* volume fraction (v/v%)
xiii ABSTRACT
As most polypeptides, poly(Y-alkyl-a,L-glutamates) exhibit the ability to exist in well defined chain conformations of extensive order such as the a-helix. This helical conformation imparts a rod-like character to the polymer which is maintained in the melt and in certain solvents at particular concentrations.
This dissertation reports the synthesis and characterization of poly(y- alkyl-a.L-glutamates), in particular, of poly(y-stearyl-a,L-glutamate) (PSLG), poly(y-benzyl-a,L-glutamate) (PBLG), and poly(y-stearyl-a,L-glutamate)-co- poly(y-benzyl-a,L-glutamate). Polymerization studies of PSLG with primary, tertiary, and silylated amines as initiators; and PBLG with coinitiators are described. The labeling of PSLG with a fluorescein dye is also explained.
All the polymers were obtained from the corresponding N-carboxyanhydride monomers. Special focus was given to nearly monodispersed samples of
PSLG. Polydisperse samples of PSLG were fractionated using a tetrahydrofuran-methanol system to obtain fractions with polydispersities of
1.1 to 1.7. The nearly monodispersed PSLG samples were characterized by ’H-NMR, gel permeation chromatography, static light scattering, and intrinsic viscosity studies.
xiv Formation of lyotropic liquid crystalline phases of the nearly monodispersed samples of PSLG in tetrahydrofuran were studied by optical microscopy. The isotropic and cholesteric liquid crystalline phases are detected using cross polarizers. The biphasic region is manifested by the presence of spherulites. The phase boundaries for PSLG of different molecular weights were determined.
The reactivity of trimethylsilyl iodide with ester groups was used for the modification of the side chains of PBLG. The trimethylsilyl ester formed reacts with isocyanate groups under neutral conditions at room temperature to form amides. Formation of crosslinks between polymer chains using 1,6- diisocyanatohexane produces isotropic gels made of rigid rods. The gelation process can be followed by dynamic light scattering. The formation of anisotropic, highly ordered gels (crosslinking of the polymer in the liquid crystalline phase) is accomplished by applying an electric field while the gelation occurs.
xv CHAPTER 1
LITERATURE REVIEW
1 2
1.1 HIGH MODULUS POLYMERS
Since the mid 1800s, when the first modifications of natural polymers were attempted and were followed later by the first synthetic polymers obtained at the beginning of the 20th century, scientists have been constantly searching for new materials. One of the driving forces that has propelled these efforts is to mimic the physical properties of macromolecules found in nature, like rubber, wood, cotton and silk among others. In addition to this, many ventures have been directed toward new materials that combine properties of different natural resources such as m etals, ceram ics, glass, etc. Exam ples of th e se abound in today’s life, some examples are plastic films that are clear (like glass) and at the same time, shock resistant, or a strong (like a metal) and also light weight composites. The majority of the synthetic polymers commonly used are replacing, in some way, other natural materials that were used in the past.
In the majority of the cases, these have not been just substitutions but they have come with improvement in performance or costs (or both) over the materials they have replaced.
After decades of development of industrial processes to produce the incredible array of polymers that are now available, enormous efforts have been focused on the search for even higher performance materials. One of the areas of research that has received increased attention during the last years is the development of materials with high performance properties. 3
For applications that require extremely good mechanical properties and are, at the same time, light weight, a whole class of polymers have been developed. These polymers are frequently called high modulus polymers, and possess a common characteristic, rigidity, either in their backbone or in the pendant side chain. Examples of this class of polymers include: aromatic polyamides and esters, poly(benzobisoxazoles), polypeptides and carbohydrates.
Thus, high modulus polymers find their most important applications in areas that require a material with very high specifications but in which the cost is not critical. Some of the areas where these products have found to be especially attractive are the aerospace industry, high quality sports equipment, ballistic protection devices, and reinforcement for tires, ropes and cables. Newer uses of these polymers are envisioned in electronics, telecommunication, and applications in hostile environments1.
Even though some of these new polymers have been known for more than a decade, only few of them are been exploited commercially today.
Some examples are: the aramid fibers, Kevlar (DuPont) and Arenka (Akso), thermotropic copolyesters, Vectra (Celanese) and Wydar (Dart) and high modulus polyethylene fibers, Spectra 900 (Allied)1. This is because the high performance qualities of these polymers come with a drawback, i.e., with the exception of the thermotropic copolyesters, these high performance polymers cannot be processed by commonly utilized techniques. Many of 4
these polymers are available only as fibers and hence must be used in composite form to produce complex three-dimensional parts. Because of the intrinsic characteristics of these polymers, traditional processing techniques used for normal polymers cannot be applied, therefore, new processes have to be discovered. Hence, a mastering of their synthetic processes and a better understanding of their behavior in solution and in the melt is clearly a necessity.
Although the unique properties of these polymers are based on their high anisotropy, for this same reason, the high modulus is achieved only in the direction of the molecular chain orientation. Properties perpendicular to the molecular axis are usually an order of magnitude or more lower than the values parallel to the main chain1. However, new studies of molecular architecture involving the stacking of monolayers of these polymers in different orientations are part of the solution to the problem.
1.2 Poly(y-alkyl-a,L-glutamates)
In an attempt to simulate natural macromolecules with very specific physical properties and possibly striking biological effects, during the last decades, efforts have been directed toward the synthesis and physical studies of polypeptides, polynucleic acids and simple polysaccharides.
This investigation focuses on the study of polypeptides. Like proteins, synthetic polypeptides can assume different conformations, such as a-helix, p-sheet or random coil. Naturally occurring proteins can, on the other hand, 5
not only adopt these conformations, but any combination of them. These conformation domains depend on the amino acid sequence and external conditions like solvent and temperature. This is the basis for the high specificity of most proteins in enzymatic reactions, but for the same reason, it makes them very complex systems to study. A simpler model for proteins are shorter polypeptides. And homopeptides (only one amino acid component) are still an easier model to study, but nevertheless, with plenty of fascinating properties.
Polypeptides are unique among synthetic polymeric materials. A reason for their very singular characteristics and properties is the high density of inter and intramolecular hydrogen bonds formed in the backbone by the amide functions. These hydrogen bonds allow polypeptides to assume secondary and tertiary structures that depend on the amino acids involved. And similarly to proteins, they can adopt different conformations,
Of special interest is the formation of the a-helix which is allowed in certain solvents (helicogenic solvents) and under certain conditions (such as temperature, pH, etc.)2.
Poly(y-alkyl-a,L-gIutamates), PALGs, are polypeptides composed of a single monomer unit derived from an ester of a.L-glutamic acid. PALGs have been subjects of numerous studies for the last 40 years. One of the reasons for such a special attention is that glutamic acid is the most 6
inexpensive optically active a-amino acid available, its use as sodium L- glutamate in the food industry guarantees this. And although PALGs are still far from being commercially exploited they are unique among synthetic polymers because of their ability to exist in well defined conformations of extensive order such as the a-helix and to retain it in solution. Moreover, this ordered conformation can undergo a transition into a random coil, making these macromolecules very versatile. Another reason for their popularity among researchers is their good solubility in several common solvents.
Most poly(y-alkyl-a,L-glutamates) adopt the a-helical conformation in a variety of solvents and in the melt. The solubility and thermal characteristics of this class of polymers are dictated by the nature of the ester group. The pendant chain can give the polymer very distinct properties. W atanabe et al3 has reported that the structure in the solid state for PALGs with small side chains, with less than 10 carbons in the alkyl ester group, is an hexagonal lattice, in which the side chains are in a glassy state and surround each helix backbone symmetrically. When the side chains are longer (Cio or more), the side chains form a crystalline phase composed of paraffin-like crystallites. This crystallization forces the a- helices to pack into the a layer structure, and the crystallites are located between the layers. Figure 1.1 shows the structures of the repeat units for 7
some of the most common PALGs: polyfr-methyl-a.L-glutamate), PMLG, poly(y-benzyl-a, L-glutamate), PBLG, and poly(y-stearyl-a, L-glutamate),
PSLG.
c h 2 (CH2)|7—CH3
R
Fig. 1.1: PMLG, PBLG and PSLG
In helicogenic solvents and in the melt, PALG’s a-helical conformation provides a cylindrical rigid rod backbone surrounded by the side chains which extend out radially. When the pendant chains are long aliphatic moieties, they are also called fuzzy rods, hairy rods, or bottle brushes. Fig. 1.2 shows a representation for a polymer like PSLG. The side chains provide a "solvent skin” that is responsible for the solubility and tractability of the polymer. 8
Lateral View
Cross Section
Fig. 1.2: Fuzzy rod
1.2.1 PoIy(Y-benzyl-a,L-glutamate)
PBLG was first synthesized by Courtaulds Ltd. in the early 1950s as a potential silk replacement. It has been reported that PBLG can assume four possible conformations: a-helix, extended p-sheet, cross p-sheet or random coil. The conformation depends mainly in the strength and ordering of hydrogen bonds, and this changes with varying molecular weight, the solvent used, and the chirality of the monomer units4. Due to the nature of the benzyl side chain, PBLG is soluble in many organic solvents and shows various conformation and aggregation structures with varying polarity g
and/or acidity of the solutions. PBLG dissolves with no aggregation and adopts an a-helical conformation in DMF and m-cresol, and a random coil conformation in dichloroacetic acid and trifluoroacetic acid. It has been reported that a-helices of PBLG aggregate in an anti-parallel and side-by- side fashion in nonpolar solvents such as benzene, benzyl alcohol and dioxane; and head-to-tail in chloroform, 1,2-dichloroethane and THF. This aggregation phenomenon is molecular weight dependent, lower molecular weight polymers aggregate more than the larger molecular weight in chloroform solutions. And it is well known that high concentration solutions of PBLG in dichloromethane, chloroform, 1,2-dichloroethane and DMF, form liquid crystalline phases5.
1.2.2 Polymerization of NCAs
N-carboxyanhydrides (NCA) of amino acids (also referred to as 4- substituted oxazolidine-2,5-diones) are the most common monomers used to produce synthetic homopolypeptides. The first report of a polymerization of an NCA dates to 1906 when Leuchs observed the evolution of CO 2 w hen water reacted with glycine N-carboxyanhydride to form glycine polypeptide1.
Although polymerization of a-amino acid NCAs has been used for decades for the synthesis of high molecular weight polypeptides, the mechanism through which the polymer is formed cannot be controlled completely. There have been numerous review articles on this subject2,6'10 10
showing the existence of polymerization mechanisms that are constantly competing. One mechanism is favored over another depending on the polymerization conditions and on the nature of the initiator employed, however, independently of the initiation characteristics, all the mechanisms involve ring opening polymerization11.
a-Amino acid NCAs possess four reactive sites (two electrophilic and two nucleophilic centers), as Fig. 1.3 shows. The two electrophilic centers are the carbamoyl group (C-2) and the carbonyl group (C-5). The two nucleophilic sites are the NH and the a-CH groups, which after deprotonation yield highly nucleophilic amide anions and carbanions, respectively.
o R: — CH3 R
Fig. 1.3: N-carboxyanhydrides: methyl, benzyl and stearyl glutamates
This multiple reactivity and their capability to polymerize make the polymerization mechanisms very complex and difficult to control. Another reason is the low solubility of most oligopeptides in organic solvents. The 11
precipitation of growing oligopeptides diminishes the reactivity of the active end-groups and changes the kinetic rate of the polymerization, which usually translates into polydisperse products12.
Besides heat, the polymerization reactions are most commonly initiated by protic nucleophiles or by strong bases. Among the protic nucleophiles the most commonly studied are water, alcohols, and primary and secondary amines. The reaction proceeds through a nucleophilic attack to the carbonyl group C-5 of the NCA ring forming a carbamate end- group (studies with 14C-labeled NCAs show that the evolved C02 comes from the C-2 carbonyl exclusively), as shown in Fig. 1.4. If this is followed by decarboxylation, an amine end-group is produced. Since this amine end-group is a stronger nucleophile than water, alcohols and aromatic amines, this becomes the growing end of the polymer. In this case, the initiation step is slower than the propagation rate (this is called the amine mechanism). This accounts for the usually higher degrees of polymerization (DP) than monomer:ratio used. Primary aliphatic amines react faster than the end-groups producing a more uniform molecular weight distribution. However, in spite of the living character of the polymerization, it does not produce narrow molecular weight distributions (MWD). Initiators that react in this manner remain covalently attached to one end of the polypeptide. On the other hand, if reaction conditions favor the stability of the carbamate end-group, such as low temperatures, high C02 pressures 12
H R O. NH . + ,R' nn / H2N—R' Y
U H’N-Y ^ w K H. VN-V . R
- COj O ^ o ° * - COj
O R O
NH .« NH. -AO' NH' Y * ^ H,N'NH' O R H. R
-002
polymer polymer
Fig. 1.4: Primary amine (i) and carbamate (ii) mechanisms 13
and the presence of metal ions, the carbamate is the nucleophile that attacks the next available monomer. This mechanism is known as the carbamate mechanism and is shown also in Fig. 1.4.
When secondary amines are used, the initiation step depends upon the competition between the nucleophilicity and the basicity of the amine.
This competition is determined by the nature and size of the substituents in the amine. Good nucleophiles (smaller amines) react just like the primary amines. On the contrary, secondary amines that are better bases than nucleophiles (bulkier groups) react through a completely different reaction mechanism, the so called activated monomer mechanism. Some secondary amines can react through both mechanisms producing a bimodal MWD.
When strong bases are used, the initiation step consists of the abstraction of the NH proton of an NCA monomer by the strong base to produce an NCA anion. This NCA anion is the nucleophile that attacks the next available NCA to form a dimer with an N-acyl NCA on one end and a carbamate group on the other (Fig. 1.5). Chain growth can occur through any end, via a carbamate mechanism or via an activated monomer mechanism. In the latter case, there is a proton transfer from another NCA monomer to the dimer to produce another NCA anion which attacks the N- acyl NCA since it is more electrophilic than an unreacted monomer. In this mechanism there is no incorporation of the basic initiator to the polymer chain. This polymerization mechanism yields higher molecular weight H R N—^
oA 0 A 0
■ o . NH.
4 I - CQz
R N_ ( 0 ^ 0 °
polymer
Fig. 1.5: Activated monomer mechanism 15
polypeptides than the other initiators but with broader MWD. This phenomenon is explained by the occurrence of coupling reactions of the growing chains. The reaction time is normally much shorter than in other mechanisms.
It has been observed that when aprotic solvents (such as THF or dioxane) are used, the polymerization reaction exhibits an increase in the rate once dodecamer chains are formed8. This autoacceleration is attributed to an increase in the reactivity of the active chain end when the growing chain adopts the a-helical conformation.
1.2.3 Applications and Potentials of PALGs
Most of the applications and prospective research interests of PALGs relies in their ability to form highly organized structures in solution and in the melt. The rigidity of the backbone, the properties that the nature of the side chains confer to the polymer, and the chirality of the a-helical backbone are the main characteristics that make these polymers so unique and attractive.
The first application for a PALG was reported in 1961, when a patent for fabrication of PMLG fibers was issued13. Cloth materials made of PBLG or the D-isomer, possess similar characteristics as silk, they show excellent water permeation and are more temperature resistant than natural silk.
Films of PMLG have been reported to have high strength and excellent abrasion resistance. Improvement in dyeability is obtained when PMDG is 16
used as a coating on poly(propylene), polyamide, rayon or glass fiber
fabrics. The Japanese literature shows numerous uses of films of PMLG as
leather substitutes13. When used as a coating, this polymer also improves
the weather and abrasion resistance, heat stability and protects against fungal attacks on natural leathers.
A number of applications others than in the textile or leather
industries have been also reported for PMLG. Some of them are:
permeable membranes for oxygen monitoring, support for photographic films, use to disperse chemicals in photocopying paper and pressure and
electrolytic sensitive papers13. Also, in the area of medical applications
PMLG have been used for slow drug release systems, highly permeable
membranes for bandaging, among many others.
During the last decades the area of molecular architecture has been
attracting enormous attention. It was in 1925 when Gorter and Grendel14 first observed the formation of a monomolecular layer of proteins on the water surface. Since then, Langmuir-Blodgett (LB) films have become of
much interest because they provide a convenient technique to assemble two and eventually three-dimensional ordered structures at the molecular
level. With this technique, the alignment of the transferred molecules parallel to the dipping direction is determined by the geometric conditions of the monolayer flow on the water subphase15. This technique has been applied in nanoengineering for the construction of highly organized 17
molecular devices with applications in optics, electronics and sensor
devices. It has been reported that multilayers of PALGs LB films possess excellent optical properties due to their very minimal loss in wave guiding, patterning and information storage16.
Among these applications, W egner17 and other investigators16,18 have studied nanostructures built with PALGs. Perfectly oriented monolayers of
PALGs formed at the air-water interface produce anisotropic films that can be easily transferred onto a solid support. When PALGs of long hydrocarbon side chains are used, the polymer molecules lie flat at the interface adopting a two dimension liquid crystalline state (nematic-like) with low defect sites due to a lack of crystallization. Tsukruk et al1s have reported that only small optical losses occur from layer interfaces and refractive index changes at domain boundaries in multilayer devices used as wave guides. The polymer rods align with the helical axes along the dipping direction. It is believed that the side chains are partly in a glassy state (somewhat fluid). Molecular devices with potential applications in optics and electronics could be obtained by the stacking of these LB monolayers with the same or different orientations, or by alternating monolayers of rigid rod polymers with monolayers of a completely different composition. Moreover, multilayer assemblies of PALGs show improved thermal stability compared to common LB systems. W egner has reported on studies incorporating dye molecules into mono- and multilayer 18
assemblies for potential applications in non-linear optics, energy transfer systems, and photoelectric effects17. In addition, these systems have been envisaged as optical information storage devices20. According to these investigators, there are different ways to “write" the information, one is to optically heat an oriented polymer film into the isotropic phase and quenching this “bit" by rapid cooling. Another mechanism to store information on a anisotropic LB film involves the optical induction of a birefringence pattern by disturbing the orientation of the polymer chains which translates to a difference in the index of refraction. Both mechanisms increase their sensitivity when dyes are used as optical labels. Menzel et al21 attempted to store information by incorporating azobenzene chromophores and isomerizing it with UV irradiation.
In a completely different type of application, Canet and Cortieu22 have reported that liquid crystalline solutions of PBLG in dichloromethane can be used as the solvent for enantiomeric excess analysis of molecules containing several chiral centers. According to the researchers, the discrimination occurs because in such an anisotropic chiral medium, two enantiomers exhibit different averaged molecular ordering parameters.
Yoda et al23 have reported the formation of microheterophase structures that are auto-assembled by copolymers containing a PALG block.
A tri-block copolymer of the type A-B-A (where A is the rigid PALG block and B is a flexible polymer) shows the formation of micelles. Films cast 19
from solutions of these copolymers promise to have interesting optical
characteristics24. Tests for biomedical applications are exciting due to their
antithrombogenic activity, and the polypeptide block in these synthetic
copolymers could improve the biocompatibility of materials used for
implants.
PALGs have also been proposed as promising drug delivery
systems. Studies by Cho et al2S of AB block copolymers of PBLG and
poly(ethylene oxide), to which [0-p-D-galactopyranosyl-(1-4)]-D-
gluconamide or [0-a-D-glucopyranosyl-(1-4)]-D-gluconamide were attached
to the end, are intended to avoid the side-effects of drugs by delivering the
active components at the receptor sites. It has also been reported that
modified PMLG (usually with ethylene diamine) has antiviral and
bacteriostatic activities13.
Also, the addition of small amounts of PALGs to poly(vinyl chloride),
poly(vinyl acetate) or their copolymers improves the shatter resistance of
the products made of these materials13.
The number of ventures aimed for applications of PALGs are proof of their great potential as a specialty polymer. However, for a study of
physical properties of polymers to have significance, the samples have to
be well characterized. In polymers in particular, the relationship of the
molecular weight, and the polydispersity of the sample to the properties of the material are of major importance. Obtaining monodisperse and pure 2 0
homopolymers of PSLG is, thus, one of the objectives of this research work.
Synthesis of the homopotymer, fractionation of the product and
characterization of the molecular weight and polydispersity are the steps
necessary to achieve the goal. Only after monodisperse samples are
available, the physical properties measured can provide true information on the attributes of this polymer. Chapter 2 describes the synthesis, fractionation and molecular weight characterization of different PSLG samples. Chapter 3 accounts for studies of the liquid crystalline properties of PSLG using polarizing optical microscopy. Finally, chapter 4 explains the work in the modification of PBLG through a new approach for its side chain reactivity under mild and neutral conditions. This is followed by the utilization of this methodology for the formation of crosslinked gels of rigid rods of PBLG in solution. CHAPTER 2
SYNTHESIS AND CHARACTERIZATION OF PSLG
21 2 2
2.1 INTRODUCTION
Like other PALGs. PSLG can adopt an a-helix conformation in
solution and in the melt. Most rod like polymers show very poor solubility
and their melting points are usually higher than their decomposition
temperature. Nevertheless, in PSLG, the nature of the long and flexible
paraffinic side chain (Cie) provides this polymer with very unique properties.
PSLG is soluble in ordinary organic solvents, such as chloroform,
dichloromethane, THF and dioxane. In addition to this, the a-helix can
undergo a transition to the random coil conformation in solution by addition of a strong acid, like trifluoroacetic acid, TFA, to break intramolecular hydrogen bonds. In the solid state, when the side chain crystallites melt, the side chains act as solvent molecules surrounding the rigid helical backbone, thus lowering the temperature at which the polymer flows.
Microscopically, the melt is a two phase system, one of them consists of the rigid backbone (a-helix), and the other of the flexible side chains. Large molecular weight PSLG forms lyotropic liquid crystals in THF at appropriate concentrations, their study is described in chapter 3.
This chapter describes the synthesis of homopolymer PSLG from L- glutamic acid and stearyl alcohol (1-octadecanol). One of the objectives is the polymerization of the NCA monomer to produce monodisperse products, and for this different initiators were studied. Among them, primary and 2 3
tertiary amines, sodium methoxide, and a group of silylated amines. In an
attempt to synthesize monodisperse oligomers of PBLG, we studied the
polymer formation via the activated monomer mechanism in the presence of
highly electrophilic molecules acting as coinitiators. The synthesis of a
copolymer, poly(Y-benzyl-a,L-glutamate)-co-poly(y-stearyl-a,L-glutamate),
and the labeling of PSLG with a fluorescence dye are also explained. The
second part of this chapter accounts for the fractionation of polydisperse
PSLG and the characterization of nearly monodispersed samples by a
variety of methodologies such as intrinsic viscosity, gel permeation
chromatography (GPC), and light scattering.
2.2 SYNTHESIS
2.2.1 Synthesis of PSLG
Homopolymer PSLG is usually synthesized from the corresponding
N-carboxy-anhydride of stearyl-L-glutamate (NCA-SLG) as reported by Daly
and Poche26. A different approach to obtain PSLG is the transesterification
of PMLG or PBLG, which are commercially available. Studies of the
transesterification reaction using p-toluene sulfonic acid at high
temperatures show the incomplete removal of all the original ester groups27.
Transesterification occurs on more than 80% of the side chains, but is not totally complete; therefore, a copolymer (which possesses different
properties from the homopolymer), is the final product. Moreover, acid 2 4
catalyzed scission of the main chain also occurs during the
transesterification, lowering the molecular weight of the polymer and
changing its molecular weight distribution.
The synthesis of the ester, y-stearyl-a.L-glutamate, from L-glutamic
acid and the corresponding alcohol was developed by W asserman28 in
1966. In this procedure, the use of sulfuric acid as the catalyst and tert-
butanol as the solvent forces the esterification reaction to occur selectively
at the y-carboxylic acid. The reaction procedure for the formation of NCA-
SLG using triphosgene is described by Daly and Poche26. It should be
pointed out that NCA-SLG is sensitive to heat and to nucleophiles,
especially water (moisture is able to start the polymerization). Therefore,
great precautions have to be taken in its handling. Solvents and glassware
need to be carefully dried. Identification of the NCA is easily accomplished
b y IR spectroscopy12, the characteristic frequencies at 1855 cm'1 and 1780
cm'1 due to the stretching of the carbonyl groups in the ring are evident (Fig.
2. 1).
When the NCA route is followed, the initiator for the polymerization is
of great importance. As stated previously, the nature of the initiator
determines the mechanism of polymerization. Primary amine initiators give
predictive monodisperse samples when used in low monomer to initiator ratios (<100)To obtain high molecular weight polymers, on the other hand, a strong base initiator is preferred. The extent of the polymerization PSLG
NCA-SLG
SLG
3000 2000 1000
Wavenumber (cm-1)
Fig. 2.1: IR spectra of SLG, NCA-SLG and PSLG 2 6
reaction can be easily followed by IR spectroscopy, the disappearance of
the peaks that are characteristic of the NCA shows the consumption of the
monomer in the reaction (Fig. 2.1). Prystupa and Donald4 have reported that
the frequencies in IR spectroscopy due to the carbonyl groups in the polymer
backbone can also reflect the conformation of the polymer chain. The a-helix
and p-sheet conformation present frequencies for the ester C=0 group at 1735
cm"1 while the random coil conformation does not. The amide I band for a-helix occurs at 1652 cm"1, for p-sheet at 1704 cm"1, and for random coil at 1660 cm'1.
A ^-NM R spectrum of PSLG shows the different kinds of protons present in the polymer, as shown in Fig. 2.2. Peaks at 5 between 2.0 and 2.8 ppm correspond to the H in p andy positions. The peak corresponding to the a-H overlaps the 0-CH2- hydrogens (a in the figure) at around 4.0 ppm. The hydrogens from the next two -CH2- groups (b and c in the figure) appear at
6=1.8 ppm and 6=1.6 ppm, respectively. The remaining -CH2- hydrogens in the side chain appear at 5= 1.2-1.3 ppm and the hydrogens in the side chain methyl group appear at 6=0.8 ppm. But ^-NM R spectroscopy also provides information about the conformation of the polymer. It has been reported29 that the a-H peak in in the 1H-NMR spectrum appears at around 4.0 ppm when the polymer adopts the a-helix conformation in solution. When the helicity is destroyed by addition of TFA (breaking of intramolecular hydrogen bonds) the > NHt ^ P ^ h 2 " Y C H ,
9.0 8.0 7.0 6.0 5.0 4.0 3.0 2.0 1.0 PPM
Fig. 2.2:1H-NMR spectrum of PSLG in CDCI3 2 8
peak for the a-H shifts to about 4.5 ppm, which is an indication of the random coil conformation.
2.2.2 Synthesis of Poly(y-benzyl-a,L-glutamate)-co-poly(y-stearyl-a,L-
glutamate)
NCAs of different ester groups can also be polymerized together to obtain a copolymer. A copolymer containing benzyl-L-glutamate and stearyl-L-glutamate units, is produced when a mixture of the corresponding
NCAs are polymerized with a primary amine. The reaction takes place in the same manner as a normal homopolymerization since both monomers are soluble in THF. The copolymer obtained produced the 1H-NMR spectrum showed in Fig. 2.3. From the integrals of the peaks at 5 - 5.0 ppm
(corresponding to the benzylic hydrogens in the benzyl ester units) and 5 =
0.9 ppm (methyl group in stearyl units) of the 1H-NMR spectrum it is apparent that the proportion of benzyl to stearyl units in the copolymer is
30:70. The copolymer is soluble in organic solvents such as THF, chloroform, DCM. This copolymer is used to investigate the reactions at the side chains (chapter 4).
2.2.3 Silylated Amines as Initiators
According to the mechanism proposed for primary amines, good nucleophiles should improve the rate of the initiation step (nucleophilic attack of the initiator to the NCA monomer). Should the rate of the initiation 0 o N H -C H -C -)— f NH“ CH—c4- | h \ | h
(c h 2) CH CH 5 = 5.0 ppm 5 = 0.9 ppm
9.0 8.0 7.0 6.0 5.0 4.0 3.0 2.0 1.0 PPM
Fig. 2.3:1H-NMR spectrum of poly(y-benzyl-a,L-glutamate)-co- polyfy-stearyl-a.L-glutamate) in CDCl3 3 0
step increase in comparison with the propagation steps, the molecular
weight distribution would become narrower. Silylated primary amines are
expected to be stronger nucleophiles than common primary amines due to
the nature of the trimethyl silyl group. Their reactivity as initiators for the
polymerization was studied as an attempt to improve the polydispersity of
the polymer. The initiators used were ferf-butyl trimethylsilyl amine, and
N.N-diethyl trimethylsilyl amine. These initiators were tested in different
monomerinitiator ratios. All the tested silylated amines produced polymers,
however, GPC analysis of the products of the different initiators and
different monomer:initiator ratios showed no clear evidence of an
improvement of the initiation rate, or molecular weight distribution (Fig 2.4
and Table 2.1). It is apparent from the GPC chromatograms that there is no
correlation between the amount of initiator added and the degree of
polymerization obtained. The polymerization with low monomer to initiator
ratio produced a larger molecular weight polymer than the high
monomer: initiator reaction. Moreover, there is no improvement in the MWD compared to a normal primary amine polymerization for low monomer to
initiator ratios.
2.2.4 Study of Coinitiators
Kricheldorf30,31 and Hashimoto et al32 have reported the use of a coinitiator molecule in the synthesis of monodisperse oligomers of different oligopeptides. These coinitiator molecules are more electrophilic than the 31
PSLG-D Q 0) T3 C PSLG-C o >
PSLG-A Elution Volume (mL) Fig. 2.4: GPC chromatograms of polymerizations by different silylated amine initiators Table 2.1: Results from the polymerizations by different ______silylated amine initiators ______Monomer: Initiator Polym er Initiator Mwgpc MWD ratio PSLG-A ferf-Butyl 50:1 15,500 1.7 trimethylsilyl amine PSLG-B ferf-Butyl 100:1 9,500 4.5 trimethylsilyl amine N.N-diethyl PSLG-C 50:1 11,000 1.7 trimethylsilyl amine PSLG-D N.N-diethyl 100:1 9,000 1.5 trimethylsilyl amine 32 NCA monomer. As explained before, in NCA polymerization, when a tertiary amine is used to initiate the NCA polymerization, the reaction occurs via the activated monomer mechanism. The propagation step of this mechanism involves the attack of a NCA anion (activated monomer) to the NCA-end of the growing chain. This reactive end of the polymer is a N-acyl NCA, which is more electrophilic than a normal NCA monomer, thus the growth of the chain is preferred over the initiation of a new one (Fig 2.5). Fig. 2.5: Propagation via the activated monomer mechanism On the other hand, in the initiation step, the activated monomer can only attack a normal NCA, which has a slower rate than the propagation steps (NCA is less electrophilic than the growing end of a chain, N-acyl NCA), resulting in a broad molecular weight distribution. It is in this step where the presence of a coinitiator can improve the distribution. The acceleration effect of the coinitiator results from the replacement of the slow initiation step (reaction i) in which a NCA anion attacks another NCA, by a faster initiation step involving the faster attack of the activated monomer (NCA anion) to the coinitiator (reaction //), which is more electrophilic, as shown in Fig. 2.6. 3 3 HN 1 / N------1 HN------C H 2-U -N ------1 o^o^o + o^o^o ^ o^o- o^cr^o Ruction i O /" — 'N 0 0 H3O “- 11 1?----- 1 f + ^ ----- 1 ^ CH3-U-N----CH2-U-N------1 O ^ O - ^ O cr'cr^o 0 ^ 0 ' cr'or>to Reaction ii Fig. 2.6: Initiation steps via the activated monomer mechanism This faster initiation reaction would produce more narrowly disperse polymers, since the propagation steps will be of slower or comparable rates as the initiation step rate. When the polymerization proceeds through this mechanism a derivative of the coinitiator remains attached to one end of the polymer while the growing end possesses an NCA ring which is where the addition of new monomers occurs. It should be noted that after the first NCA anion attacks the coinitiator, it becomes an N-acyl NCA, which is more electrophilic than any other free NCA monomers (thus, it becomes the preferred site of reaction). Kricheldorf30,31 and Hasihimoto et al32 have reported the formation of monodisperse oligopeptides when coinitiators were used. They characterized their products by 1H-NMR analysis. In our studies of the 3 4 effect of different coinitiators in the oligomerization of NCA benzyl-L- glutamate, we used: 4-chloro phenylisocyanate and chlorosulfonyl isocyanate, which are more electrophilic than the NCA monomer. The basic initiator used was triethylamine, in four times the amount of coinitiator. Analysis by plasma desorption mass spectroscopy (PDMS) showed the formation of oligomers, however, the products were not monodisperse, the mass-peaks of a molecular weight distribution is apparent from the spectra, as shown in Fig. 2.7. It can be seen in the spectra the periodicity of the signals, which corresponds to the monomer unit weight of benzyl glutamate (219). The spectrum shown in Fig. 2.7 (A) corresponds to a 15:1 monomer to coinitiator ratio, the coinitiator was chlorosulfonyl isocyanate; peaks corresponding to DPs of 5, 6 and 7 are dominant. Fig. 2.7 (B) shows the spectrum for the oligomer obtained using 4-chlorophenyl isocyanate as the coinitiator in a ratio of 10:1 (monomer to coinitiator); there are no dominant peaks. 2.2.5 Synthesis of Labeled PSLG A new method to attach a dye to the polymer utilizing the mechanism of the polymerization itself was developed. During the polymerization (primary amine mechanism) a primary amine group is present at the growing end of the chain, and it attacks nucleophilically the next monomer to increase the polymer chain. It is this active primary amine end-group that is used to attach the dye molecule to the polymer at the end of the 3 5 800 700 600 z- 500 o £ 400 h- co O 300 Q. 200 100 1000 1400 1800 2200 2600 3000(M/Z) 1800 1600 1400 co O 1200 Ui > H 1000 co O 800 400 200 500 1000 1500 2000 2500 3000 3500 (M/2) Fig. 2.7: PDMS spectra of (A) 15:1 monomercoinitiator ratio, chlorosulfonyl isocyanate coinitiator; (B) 10:1 monomercoinitiator ratio, 4-chlorophenyl isocyanate 3 6 polymerization, when there are no more monomers present. The dye used is fluorescein isothiocyanate, isomer I (Fig. 2.8), which possesses the isothiocyanate electrophilic group. This functional group is attacked nucleophilically by the amine group at the end of the polymer. This methodology produces a polymer chain with only one dye attached to it, and the position of the dye guarantees that the helical conformation of the polymer remains undisturbed. Furthermore, the capping of the amine terminus may inhibit end to end aggregation. s = c = n HO' OH Fig. 2.8: Dye fluorescein isothiocyanate, isomer I 2.3 CHARACTERIZATION 2.3.1 Fractionation Polymers are usually mixtures of homologues differing in their molecular weight. A parameter of the heterogeneity in molecular weights is the molecular weight distribution, which is the ratio of the weight-average and the number-average molecular weights ( ^ ^ n). The physical 3 7 properties of polymers are intimately related to the distribution, since the molecular weight is of major importance to most physical characteristics of the polymer sample. It is therefore, of great importance to obtain monodisperse samples prior to any study of physical properties or characterization of any kind of polymer. Fractionation is one method to separate polymer molecules quantitatively according to their size. It is based on the partition of the molecules in two phases. The separation of the two phases occurs when a nonsolvent is added to a homogeneous polymer solution. The partition of the polymer into the two phases depends upon its molecular weight or chemical composition (in case of copolymers). Usually, the polymer is first dissolved in a solvent at sufficiently dilute concentration. A nonsolvent that is miscible with the solvent, is then added to form the two phases. Phase separation is accelerated by centrifugation and the polymer-rich phase is isolated as the first fraction (high molecular weight molecules). More nonsolvent is then added to the solvent-rich phase until the next fraction appears, and the same procedure is repeated to obtain more fractions, until all the polymer is separated. In some cases, depending on the system been used, elevating the temperature after the phases separate and then allowing the system to phase separate slowly by decreasing the temperature improves the fractionation. 3 8 Two solvent/nonsolvent systems were tested for PSLGs, the first consisted of hexane/acetone and the second system of THF/methanol. PSLG is soluble in hexane at 30°C, and precipitates when acetone is added. In some situations a gel was obtained, which made the separation of the phases almost impossible. With the second system no gel was observed, and the polymer is readily soluble in THF at room temperature. This was the preferred system for fractionation of PSLG. The fractionation is more effective when the sample is heated when phase separation occurs to redissolve the sample. The sample is then allowed to cool down slowly to room temperature, at this point a fraction of the polymer reprecipitates again and can be separated. 2.3.2 Gel Permeation Chromatography GPC, gel permeation chromatography, also known as SEC, size exclusion chromatography, is another technique to separate macromolecules according to their size. This method is widely used for polymer characterization and molecular weight determination. The principle for the separation of the different sizes of molecules lies in the high and controlled porosity of the particles or gel inside the GPC column. As a mixture of different polymer sizes flows through the column, these pores delay the smaller molecules which can diffuse in and out of them, while the larger polymer molecules just flow with the solvent through the column in less time. A detector connected to the end of the column, usually a refractive index detector, responds to the concentration of 39 polymer in the solution eluting out of the column by measuring the change in refractive index of the solution with respect to the pure solvent (the refractive index changes proportionally to the concentration of the polymer in the solution). The capacity of the column to separate the macromolecules according to their size does not indicate the absolute molecular weight of the polymer. This process of separation by the size of the molecules provides with information about the molecular weight distribution of the sample. The system needs to be calibrated with standard samples of known molecular weights to obtain their elution volumes prior to any unknown sample can be analyzed. And these results are not absolute, they are relative to the standards used in the calibration curve, this means, standards of the same polymer been tested should be available. It is also important that all the parameters in the system remain the same (solvent, columns, flow rate). When a multi-angle light scattering detector is connected to the GPC system, it is possible to measure absolute molecular weights without the need of any calibration curves. The DAWN F (from Wyatt Technology Corp.) is such a detector, it can measure scattered light at 18 different angles simultaneously as the polymer solution flows out of the GPC column. The intensity of the light that a polymer scatters is used to determine its weight-average molecular weight. In addition to this, from the angular dependence of the scattered light, information about the physical size of the molecule can be obtained. This instrument uses high-gain hybrid transimpedance photodiode detectors that 4 0 measure the scattered light over a wide range of angles, from 5° to 175°. The software package that manipulates the data from the detectors (both light scattering and refractive index) is called ASTRA, and determines the number, weight and z-average molecular weights and sizes. Analysis of the different polymer sizes obtained from the fractionation of PSLG by GPC, as described above, produced the chromatograms (refractive index detector response) shown in Fig. 2.9 and Table 2.2. It can be observed that a good separation of the different molecular weight polymers was accomplished by the fractionation procedure, however, not all the fractions have a unimodal distribution. These fractions would probably require subsequent fractionations. Table 2.2: Results from fractionation of PSLG Fraction Mw MWD 1 1.6 x104 1.08 2 1.3 x 1 0 4 1.18 3 8.1 x 10® 1.21 4 4.0x10® 1.45 5 2.7x10® 1.69 6 1.4x10® 12.93 Fig. 2.9: GPC chrom atogram s of PSLG after fractionation after PSLG of s atogram chrom GPC 2.9: Fig. Refractive Index Detector 16.0 'PSLG -Second fraction -Second •fraction Fifth Unfractionated fraction First Third fraction Third Fourth fraction Fourth 18.0 uti ml ) l_ (m e m u l o V n io t lu E 20.0 22.0 24.0 4 2 2.3.3 Intrinsic Viscosity In 1930 Staudinger discovered that the viscosity of a solvent changed dramatically when a polymer was dissolved in it, even at very low concentrations33. Measurements of viscosities of dilute solutions of polymers can provide information on the polymer molecular weight, branching, polymer dimensions, and conformation. According to Newton’s law of viscous flow, the friction force, F, that resists the flow of any two adjacent layers of liquid is given by: Ft= = tiA A —dV dx where A is the area of contact between the layers, d%Jx is the velocity gradient between them (shear rate), and the proportionality constant, r\, is the viscosity. In polymer science, however, the relative viscosity, which is the change in viscosity for the solution with respect to the pure solvent, is more important than the absolute viscosity. Assuming that the polymer solution (dilute) has the same density as the pure solvent, the relative viscosity, ti„i, is defined by: n t Tirol = — = — T)o to w here t\ is the viscosity of the polymer solution, t)0 is the viscosity of the pure solvent, t is the flow time of the polymer solution and to is the flow time for the pure solvent. Flow times are measured in capillary viscosities, and they have 4 3 to be determined under the same conditions. Another term, the specific viscosity, risp, is defined by: (T| — T|o) ( t - t o ) T|*P = ------= ------Ho to Both the relative and the specific viscosities depend on the concentration of the polymer in solution, they increase with increasing concentration. The intrinsic viscosity, [r|], is defined as: [n] = lim ^ = lin,l5^ e-tO C e-*0 C The Mark-Houwink equation relates the intrinsic viscosity to the molecular weight of the polymer, M: fo] = KM" where K and a are constants for a given polymer, solvent and temperature system. The value of a contains information on the conformation of the polymer in solution. Values of a between 0.5 and 0.8 correspond to flexible polymers, and stiff polymers have values of a from 0.8 to 1.8. It should be noted that viscosity measurements to obtain the values of K and a should be done on monodisperse samples. Intrinsic viscosities studies were performed on different monodisperse PSLG samples in THF at constant temperature using a Ubbelohde viscometer. The resulting plots of ijjpfcvs. cand (In ri»i)/c vs. c are presented in Fig. 2.10 to Fig. 2.12. The intrinsic viscosities obtained for the different molecular 1.4- 0 3 -=* 1 .0 - 0.8- 0.0 0.1 0.2 0.3 0.4 c (g/dL) Fig. 2.10: Viscosity plot of PSLG of Mw=239,000 (MWD=1.13) in THF a t 25°C 1.4- 0 3 0.8- P 0 .6 - 0.0 0.2 0.4 0.6 0.8 c (g/dL) Fig. 2.11: Viscosity plot of PSLG of Mw=208,000 (MWD=1.08) in THF at 25°C 4 5 0.8 0.7- XI p 0.6- £ c 0.5- r(0 0.4 0.0 0.2 0.4 0.6 c (g/dL) Fig. 2.12: Viscosity plot of PSLG of Mw=129,000 (MWD=1.12) in THF at 25°C weight polymers are presented in Table 2.3, including the weight-average molecular weight obtained from light scattering measurements described in the next section. According to the Mark-Houwink equation, from the plot of log(Mw) vs. log[ri] the exponent a for PSLG in THF results to be 1.17±0.005, and the value for K 5.94±0.71 x 10'7 dL/g, for the molecular weight range from 129.000 to 239,000 (Fig. 2.13). Daly et ai27 have previously reported values for K=1.29±0.35 x10’7 dL/g and for a= 1.29±0.09, for PSLG samples of higher polydispersity samples with molecular weights in the range from 38.000 to 250,000. 4 6 0.1 0.0 - j=! -o.i - O) - 0.2 - -0.3 5.1 5.2 5.3 5.4 log(Mw) Fig. 2.13: Mark-Houwink plot for PSLG in THF at 25°C 2.3.4 Static Light Scattering The molecular weight of a macromolecule can be obtained from the intensity of the light scattered by different solution concentrations at different angles. The scattered intensity is directly proportional to the molecular weight of the polymer, and there are no calibrations necessary, it is an absolute method. Evaluation of the angular dependence of static light scattering allows one to determine the molecular weight of the polymer, the radius of gyration and the second virial coefficient. All this information is normally compiled graphically by plotting the Rayleigh ratio, R{q) , against the scattered angle and 4 7 the concentration. The resultant family of curves is commonly referred to as a Zimm plot. The Rayleigh ratio is given by: VI, 4tHL 0 w here q = ------sin(—), a reciprocal dimension; h is the intensity of the Xo 2 incident light on the sample; Xo is the wavelength of the beam; I(q) is the intensity of the scattered light that is detected at an angle 6 to the incident beam direction and at a distance r from the center of the sample; n is the refractive index of the solution and V is the scattering volume. The Rayleigh ratio is equal to the difference of the Rayleigh ratios for the solution and the pure solvent. The equation that produces the Zimm plot is: K c 1 q2D 2R 2 ( 1+ ^ - ^ - ) + 242c R(Q) Mw' 3 In this expression M» is the weight-average molecular weight; Rg is the radius of gyration; A% is the second virial coefficient (which depends on polymer-polymer and polymer-solvent interactions);c is the concentration of the polymer solution; and K is and optical constant that contains the differential index of refraction for the system. To obtain the molecular weight, radius of gyration and second virial coefficient of a polymer in solution a plot of vs- sin2(% ) + Ae is needed. In this expression, k is just a scaling constant. Extrapolation of the graph to zero concentration and zero angle yields the value for the weight-average molecular weight (y- intercept). The value for the radius of gyration is obtained at the limit of concentration equal to zero, Rt = 3(slopeof c-0) And thQ second ^(intercept ofc = 0) virial coefficient is given by Ai = (s *°Pe of 6 - 0) |.n e s for c _ 0 ancj 0 = 0 intercept the y-axis at the same point For the case of PSLG, the value for is 0.08 ± 0.002 at X* = 488.0 nm as reported by Pochd29. Fig. 2.14 shows the Zimm plot for a monodisperse PSLG sample (Mw=208,000; MWD 1.08). Table 2.3 summarizes the results of the characterization of the monodisperse PSLG samples. Molecular weights, radii of gyration and second virial coefficients were obtained from static light scattering experiments; molecular weight distributions were obtained from GPC. The corresponding molecular weights calculated from the GPC experiments were 225,000, 212,000 and 125,000, respectively. The difference of these values with the ones obtained from static tight experiments can be attributed to calibration problems on the light scattering detector connected to the GPC system. Calculated values for the radii of gyration, Rgcic, were obtained from the equation 4 9 where L is the length of the rod, and is given by L=1.5(Mw/382) A; and R is the radius of the rod (37 A). 80 - 7 0- 60 - c/R 5 0- 4 0 - 0.0 0.2 0.4 0.6 0.8 sin2(0/2) + 10c Fig. 2.14: Zimm plot of PSLG (Mw=208,000; MWD=1.08) in THF Table 2.3: Results of characterization of monodisperse samples of PSLG Mw MWD M (dL/g) 239,000 1.13 1.13 284.5±15.7 272.5 1.61 x 10-4 208,000 1.08 0.95 275.4±16.2 237.5 2.71 x 10“* 129,000 1.12 0.55 163.5±14.6 146.5 1.23 x 10"4 5 0 2.4 EXPERIMENTAL NMR spectra were recorded on an IBM Bruker 200 MHz or Broker 250 MHz instrument. IR spectra were collected on a Perkin Elmer 1760 X FT-IR spectrometer. Synthesis of v-Stearvl-ot. L-alutamate28. 90.0 g of stearyl alcohol (1- octadecanol) and 13.0 g of a.L-glutamic acid are stirred in 130 mL of tert- butanol and heated. When the temperature of the suspension reaches 40°C, 9 mL of concentrated sulfuric acid are added dropwise. The mixture is then heated to 65°C under reflux until a homogeneous solution is obtained. After this, the source of heat is removed while keeping the stirring and the solution is neutralized with 9 mL of triethylamine, followed by 15 mL of water, 220 mL of ethanol and 51 mL more of triethylamine. The slurry produced is let to stand until it reaches room temperature. Recrystallization of the product is accomplished by addition of 500 mL of water and 500 mL of n-butanol, the mixture is heated to 93°C, at this temperature a clear solution is obtained. The solution is allowed to reach room temperature slowly. The solid obtained is washed with hot methanol and diethyl ether. The crystals are dried in a vacuum oven overnight, producing 12.3 g (35% yield). Synthesis of v-stearvl-g.L-alutamate N-carboxvanhvdride. 2.00 g of stearyl glutamate are suspended in 50 mL of THF and heated to 50°C in a two-neck 51 flask adapted with a nitrogen inlet and a condenser connected to a base trap to collect any HCI or phosgene that escapes the reaction. At this temperature 0.50 g (1/3 equivalents) of triphosgene are added all at once and a small current of dried nitrogen is allowed to purge the reaction mixture to carry away any HCI being produced. The reaction becomes homogeneous in approximately 20 to 30 minutes. After 2 hours of reaction the clear solution is precipitated in dried hexanes (500 mL) and refrigerated overnight. The product is filtered under argon and dried in a vacuum oven. The NCA-SLG is redissolved in dried THF and filtered through celite under a dry atmosphere, and then precipitated in hexanes again. 1.92 g (90% yield) are obtained. ’H-NMR: 0.8 ppm (t), 1.3 ppm (s), 2.2 ppm (m), 2.6 ppm (t), 4.1 ppm (t), 4.4 ppm (t), 6.8 ppm (s). Synthesis of PolWv-stearvl-a.L-alutamatel. To a solution of the NCA monomer in THF (approximately 10%) a desired amount of initiator is added. Primary amines include benzylamine and n-butylamine, among the bases, triethylamine and a 25% (v/v) solution of sodium methoxide in methanol were used. The reaction takes 3 to 4 days at room temperature depending on the initiator used. The reaction is kept dry with a freshly prepared CaCt2 drying tube. When the reaction is finished, the solution is concentrated under vacuum and precipitated in methanol, the polymer is then dried under vacuum overnight (yield -90%). 5 2 Synthesis of PolvCv-benzvl-a.. L-alutamatel-co-fv -stearvi-a. L-alutamatel. A mixture of NCA-SLG (0.70 g, 0.0027 mol) and NCA-BLG (0.50 g, 0.0012 mol) in 15 mL of THF is reacted with benzylamine (0.0042 g, 3.9 x 10"6 mol) as the initiator. The flask is capped with a CaCI2 drying tube, stirred for 4 days, after which the solution is precipitated in methanol. After filtration, the copolymer is dried in a vacuum oven overnight. Yield is 0.93g (-90%). Studies of Silvlated Amines as Initiators. A solution containing 7.20 g of NCA-SLG monomer in THF (5 to 10%) was divided into four equal portions and reacted with the different initiators and with two different monomer to initiator ratios (100:1 and 50:1), as shown in Table 2.4. The polymerization Table 2.4: Study of silylated amine initiators Monomer: Initiator Polym er Initiator Monomer Initiator (mol:mol) [Yield] ferf-Butyl 1.41g 1.80 g 16.4 pL 50:1 trimethylsilyl amine [86%] te/f-Butyl 1.36 g 1.80 g 8.2 pL 100:1 trimethylsilyl amine [81%] Diethyl 1.35 g 1.80 g 16.1pL 50:1 trimethylsilyl amine [81%] 1.40 g Diethyl 1.80 g 8.1 pL 100:1 [86%] trimethylsilyl amine 5 3 reactions and later precipitations were carried out in the same manner as described in the previous paragraph. The initiators used were ferf-butyl trimethylsilyl amine, and N.N-diethyl trimethylsilyl amine. The silylated amines were obtained from Huls America, Inc. Synthesis of v-benzvl-«.L-alutamate. For the synthesis of y-benzyl-a.L- glutamate, a slightly modified procedure from Block13 was used. A suspension of 35 g of a.L-glutamic acid in a mixture of 125 mL of benzyl alcohol and 35 mL of concentrated hydrochloric acid is heated to 65°C with stirring. When the mixture becomes homogeneous the source of heat is removed and the solution is kept stirring until it reaches room temperature. 750 mL of acetone are added to the slurry formed and the mixture is refrigerated overnight. After the product is filtered and dried under vacuum,the hydrochloride is dissolved in 200 mL of ice-cold water. Small volumes of a saturated aqueous solution of sodium bicarbonate are added to neutralize the solution. The precipitate formed after neutralization is removed by filtration and the filtrate is continued to be neutralized until no more precipitate forms (~pH 7). The crude y-benzyl- a.L-glutamate, the precipitate, is washed with ice water and then recrystallized from hot water and dried under vacuum. Yield ca. 12 g (-22%). The synthesis of NCA benzyl glutamate from the ester and triphosgene is performed in the same manner as the procedure for NCA stearyl glutamate described above. 5 4 Coinitiator Studies. The coinitiators chlorosulfonyl isocyanate and 4-chloro phenyl isocyanate were purchased from Aldrich. Triethylamine was distilled under vacuum and dried with molecular sieves (4 A). THF was distilled over calcium hydride and DMF was dried over molecular sieves (4 A). To a solution of 10% (w/w) NCA benzyl glutamate in a mixture of THF:DMF (10:1) the desired'amount of coinitiator was added. To a solution of 0.20 g of NCA-BLG, 4.4 pL of chlorosulfonyl isocyanate were added; which corresponds to a monomercoinitiator ratio of 15:1. To this, 28.3 pL of triethylamine (which is the equivalent to four times the number of moles of coinitiator) were added. When 4-chlorophenyl isocyanate was utilized as the coinitiator, a monomer to coinitiator ratio of 10:1 was used. To 0.20 g of NCA-BLG monomer dissolved in the same solvent system mentioned above, 0.012 g of 4-chlorophenyl isocyanate were added. To start the polymerization, 10.6 pL of triethylamine were added. After 30 min of reaction, the mixture is precipitated in ethyl ether, then the polymer is filtered and dried under vacuum. For the reaction involving chlorosulfonyl isocyanate as the coinitiator, 0.11 g ('66%) of polymer were recovered. From the second reaction with 4-chlorophenyl isocyanate, 0.09 g (~54%) were recovered. Synthesis of Labeled PSLG. Fluorescein isothiocyanate, isomer I, 90%, was purchased from Aldrich. 1.5 g of NCA-SLG monomer is dissolved in 5 5 THF (between 5 to 10%). The polymerization is initiated with 3.9 pL of benzyl amine (monomer to initiator ratio of 100) as described previously. After the reaction is completed (about 4 days) 27 mg of the dye, fluorescein isothiocyanate, isomer I, are added (twice the number of moles of initiator). The mixture is allowed to react for two more days to ensure completion and then it is precipitated in methanol. After filtration, the product possesses a light orange coloration and the filtrate is yellow. The polymer is dried under vacuum overnight. The excess free dye in the dry polymer is extracted with methanol using a Soxhlet apparatus until the washings are colorless. After drying the polymer, it is redissolved in THF and reprecipitated in methanol. The extraction and reprecipitation are repeated until the washings are free of dye (washings are tested by UV-Visible spectroscopy). Fractionation of PSLG. The polymer to be fractionated is dissolved in THF to a concentration of approximately 0.05 g/mL. The solution is stirred and once the polymer is completely dissolved the non-solvent, methanol, is added dropwise. The addition of the non-solvent is stopped when turbidity appears. The heterogeneous mixture is then warmed to 40°C, at which it becomes homogeneous again. After this, the solution is allowed to cool down to room temperature and the turbidity reappears. The sample is then centrifuged. The supernatant is poured into a clean flask and more non-solvent is added to it in the same way. The operation is repeated until no more polymer precipitates out of the solution. The centrifugates are dried under vacuum and then 5 6 characterized individually by GPC. For the fractionation of the polydisperse sample of PSLG discussed in previously in this chapter, six fractions were obtained from 1.00 g of polymer (see tables 2.2 and 2.5 for results). Table 2.5: Polymer recovered from fractionation Fraction Weight recovered 1 0.12 g 2 0.10 g 3 0.32 g 4 0.18 g 5 0.04 g 6 0.05 g Total 0.81 g Gel Permeation Chromatography. The GPC system consists on a programmable solvent delivery unit, a W aters 590 (by Millipore Corp.), which pumps the solvent through the columns. The columns are protected by a guard column (Phenogel 50x7.8 mm, 10|im from Phenomenex). The separation is achieved by two columns connected sequentially, a Phenogel 10MXH (NFO, 300x7.8 mm, 10 urn) and a Phenogel 10 (NFO, 300x7.8 mm, 10-5A), both of them manufactured by Phenomenex. The first detector 5 7 connected to the end of the columns is a multi-angle light scattering detector, the DAWN F (from Wyatt Technology Corp.), which uses vertically polarized light provided by a He-Ne 5 mV laser (A,=632.8 nm), the read head contains 18 high-gain hybrid transimpedance photodiode detectors arranged around the flow cell (from 5° to 175°). The DAWN F light scattering detector is provided with an amplifier booster board to enhance the sensitivity (up to 100 times). The second detector connected in-line is a differential refractometer unit, the W aters 410 (from Millipore Corp.) and is provided with a LED light source. The solvent flowing through the system is THF filtered through a 0.1 pm pore size filter (from Nucleopore) at a flow rate of 0.9 mL/min. The concentration of the samples are usually between 3 and 5 mg/mL 0.3 - 0.5% w/v). Samples are prefiltered through a 0.2 pm PTFE filters (from Whatman), before their injection into a 500 pL injector loop. The system is connected to a personal computer running the software package called ASTRA (from Wyatt Technologies). Intrinsic Viscosities. Intrinsic viscosities were measured using a Ubbelohde capillary viscometer in a temperature controlled water bath at 30.0°C. The solvent was THF. Polymer solutions in the range of 0.8 to 0.5 g/dL were used. Plots of vs c and ln vs c were extrapolated to zero concentration to obtain the intrinsic viscosities. 5 8 Static Light Scattering. For the static light scattering determinations an "in- house” machine was used, equipped with a Lexel Model 95 Argon ion laser (X„= 514.5 nm). An EMI-9863 photomultiplier detector was used. The correlator was a Langley-Ford Model 1096. Dust free polymer solutions in the range of 0.1 % to 0.5% (w/v) were prepared in THF. The value for dn/dc for PSLG in THF is 0.08±0.002 mL/g (at A«=488.0 nm)29. Measurements were made for at least four different concentrations at different angles (no less than four). CHAPTER 3 LIQUID CRYSTALS 5 9 6 0 3.1 INTRODUCTION Peculiar melting behaviors of some organic molecules were first observed in the late 1800s by Reinitzer34. The molecules were cholesterol esters, and the crystals melted to form a nontransparent melt whose opacity disappeared later at a higher temperature. Reinitzer concluded that the opacity was due to some order in this intermediate molten state. The term “liquid crystal” was introduced for this stage between the crystalline and the real isotropic fluid state. Lyotropic liquid crystals are those that form in solution, and thermotropic liquid crystals are formed in the melt. In liquid crystals, molecules are partially ordered, they are arranged in an intermediate phase between the random distribution (disorder) of a liquid and the regular three dimensional packing (order) of a crystal. There are different ordered structures the molecules can adopt in the liquid crystalline phase, in the nematic phase, there is partial orientation of the long axes of the molecules parallel to a preferred axis (Fig. 3.1). In smectic liquid crystals, there is both a partial orientation of the axes and a partial positional ordering of the centers of mass of the molecules into layered structures (Fig. 3.1). A third liquid crystalline phase, called cholesteric, is described later in this chapter. 61 / y y y i'1 ’.'I'll!1 ////// mini ////// mini ////// mini n'liV'i y ////// / mini Nematic Smectic C Sm ectic A Fig. 3.1: Liquid crystalline phases Even though small organic molecules that form liquid crystals have been known for about 100 years, only for the last 20 years have polymers, that make liquid crystals, received increased attention. This is because traditionally, research on polymer melts and solutions have dealt with macromolecules that can assume a variety of conformations whose only restrictions are due to the covalent bonds and valence angles of its primary structure. The secondary structure of such polymers is not defined since it is characterized by a dynamic sequence of rapid changes in the polymer's internal degrees of freedom as the polymer structure yields to shear stresses and density fluctuations in its environment (random coil conformation). Almost all synthetic polymers exhibit a random coil conformation in solution and in the melt. On the other extreme, there are some macromolecules that possess a very well defined 6 2 secondary structure and even a tertiary structure (three-dimensional spatial arrangements of secondary structures) in solution. This is the class of polymers that usually develops liquid crystalline states. Elliot and Ambrose35 were the first to discover a polymeric liquid crystal. They were studying films of PBLG from solution when they discovered a birefringent solution phase. Later, Robinson38,37 studied this birefringent solution in detail and observed that it behaved like low molar mass cholesteric liquid crystals. W hen liquid crystalline polymers were first introduced commercially, they seemed very promising for the manufacture of high strength and light-weight materials. Since the late 1970s the Air Force Wright Laboratory and Air Force Office of Scientific Research38 have had very strong interests in these polymers. Their focus was not only in processing, mechanics and morphology of liquid crystalline polymers, but also in theoretical aspects, synthesis, and solution properties. The objective of these studies was to develop the technology for a plastic material that possesses the same or improved mechanical properties of high performance metals but with the economics and ductility of a nonmetallic material. In the early 1980s, ICI investigated the potential application of liquid crystal polymers as additives to aid in the processing of high viscosity conventional polymers, due to their high molecular weight and low 6 3 viscosity39. A few years later, even though these products have some commercial applications such as rubber reinforcement, rigid composites for boat hulls and aerospace applications, protective clothing, ballistic vests, and for asbestos replacement in brake linings, they are not been as widely used as expected40. And the reason for this is mainly due to difficulties in the processing technology (designed for random coil polymers) and their complicated physics in solutions and blends. Therefore, in order to maximize the utilization of this class of polymers in the commercial area, a better understanding of their rheology, processing, structure- property relationship, and fundamental physics is needed. 3.1.1 Theory of Liquid Crystals The formation of liquid crystals is a consequence of molecular asymmetry, and is based in the fact that two molecules cannot occupy the same space. Polymeric liquid crystals are formed due to the limited number of rodlike chains that can be accommodated in a random arrangement in solution at high concentrations, or to the axial ratio of chains that can exist randomly in the melt. When this critical concentration or axial ratio is exceeded, a liquid crystalline phase starts to appear. In the liquid crystalline phase, the molecules align parallel to each other in uniform domains in order to accommodate the higher concentration or molecular weight. As the 6 4 total concentration increases, an additional ordered phase is formed at the expense of the isotropic phase until a certain concentration is reached, at which the whole solution becomes anisotropic. In melts, the anisotropic phase formation depends only on the molecular weight of the polymers (axial ratio). It was before the first experimental studies of polymer liquid crystal systems were done when in 1947, Onsager41 formulated a theory predicting the formation of liquid crystalline solutions of polymeric rods. This theory is based on the density dependence of the free energy of a gas (or suspension) of long rods in terms of cluster integrals and excluded volume. The driving force for a transition from an isotropic to an anisotropic phase is the competition between the orientational entropy (minimum in disordered phases) and the translational entropy of the rods (minimum when they adopt a parallel array). This transition is of the first order, and at the transition point the volume fraction, ffo, occupied by the rod polymers in the ordered phase (liquid crystal) is: 08-= ------4 -5 x where x is the axial ratio, d is the diameter and L is the length of the rod. For the isotropic phase in equilibrium with the liquid crystalline phase the volume fraction, ^A.of rods is smaller: 6 5 x Onsager developed this theory for an athermal system of impenetrable rods; therefore, and ^ a are independent of temperature. In 1956, Flory42 published his theory for liquid crystal polymer solutions based on statistical thermodynamics. Flory's lattice model deals with the probabilities of arranging a rod polymer (treated as a number of connected monomer segments) and solvent molecules. The predicted concentration at which an equilibrium of isotropic and anisotropic phases coexist is given by the volume fraction, ^ a: XXX According to Flory, the phase separation is due to the asymmetry of the molecules; there are no attractive forces required43. In the melt, an axial ratio of only 6.42 (higher values are needed for solution liquid crystals) is required to obtain a liquid crystalline phase. Some experimental results utilizing polypeptides in solutions of different solvents have suggested that the Onsager theory agrees with experimental results at low values of (low molecular weights) while the Flory predictions appear closer for samples with high ^ (high molecular weights)44. 6 6 As the rigidity of the polymer decreases, the critical concentration increases, therefore, the value of is an indicator of the stiffness of the polymer. Reported values of 0.25 to 0.39. On the other hand, stiffer polymers like the aramids show lower values for comparable or lower molecular weights ($a< 0.1). The values for PALGs are believed to fall in between them44. More contemporary theories take into account enthalpic factors in addition to the entropic terms. Flory introduced %, the polymer-solvent interaction parameter, which considers the enthalpy of mixing of the polymer in a solvent. A phase diagram for solutions of rod polymers shows three regimes: an isotropic solution, a biphasic region (coexistence of isotropic and liquid crystalline solution), and a liquid crystalline solution. The transition from isotropic to anisotropic phases is dominated by entropic considerations since it appears that the enthalpy favors the isotropic solution (the transition is endothermic)44. Flory’s theory predicts the phase diagram shown in Fig. 3.2. According to the phase diagram, there is an isotropic region, a region where two phases coexist, and a third region which is uniformly birefringent. The concentration at which the birefringent 6 7 x Isotropic Liquid Crystal Biphasic + Fig. 3.2: Schematic phase diagram of lyotropic liquid crystals phase first appears (left side of the "chimney") was denoted by Robinson as the A point36. And the concentration at which the solution starts to become one uniform birefringent phase (right side of the “chimney") was denoted the B point. The A and B points depend on the molecular weight of the polymer and the temperature of the solution. The phase transitions can be followed by polarizing optical microscopy. When the polymer concentration exceeds point A, the solution separates into two phases. A birefringent polymer rich phase separates from a more dilute phase, initially in the form of liquid droplets or spherulites. This biphasic region prevails increasing the concentration, the spherulites grow in size until the B 6 8 point is reached and the spherulites coalesce forming a continuously birefringent phase. The rings inside the spherulites are equidistant and correspond to the arrangement of cholesteric planes warped into a spherical surface. The size of the spherulites depends on the molecular weight of the polymer and the density of the solvent. High molecular weight polymers form generally smaller spherulites, and if the density of both polymer and solvent are similar, the spherulites tend to grow to large sizes37. The lower portion of the phase diagram (low temperatures) indicates a wide biphasic region. However, if an isotropic, biphasic, or uniformly ordered solution are brought into this wide biphasic region, a transparent gel forms. The gel is concentration and temperature dependent and reversible. There is no general agreement on the formation of the gel; however, it is believed it is formed due to the tendency of some polypeptides to aggregate in a parallel or side-by-side fashion. 3.1.2 Cholesteric Superstructure Cholesteric liquid crystals show a parallel packing of rods at the microscopic level, but on a more macroscopic scale, a twisted superstructure is apparent. In cholesteric phases, also called "twisted nematics”, formed by rod polymers, the molecules lie in a 6 9 layer with one-dimensional nematic order and the direction of orientation of the molecules rotates by a small constant angle, from one layer to the next. The displacement occurs about an axis of torsion, or cholesteric axis Z, normal to the layer planes, as shown in Fig. 3.3. This periodic superstructure (super-helix) is characterized by the distance between the two layers with molecular orientation differing by 360°, which is known as the pitch, P, and is given by: where D is the distance between two rods (along the Z direction) and can be determined from X-ray diffraction studies of polypeptide liquid crystals. $ is also called the helical twisting power of the liquid crystalline solution44. When cholesteric liquid crystals are seen at right angles to the cholesteric axis through a polarizing optical microscope, the solutions show regions of regular striations arranged in a swirl-like, or fingerprint pattern. These patterns are the maxima and minima in the major refractive index and birefringence of the cholesteric superstructure. The maxima appear when the molecules are at right angles to the direction of observation, and the minima when the 7 0 Fig. 3.3: Cholesteric liquid crystal superstructure 71 molecules are parallel to the direction of observation. Robinson36,37 has noted that for PBLG solutions in dioxane, the distance between striations, or the periodicity, S (which is one-half the pitch of the cholesteric structure), is of the order of 3 to 100 pm, and it depends linearly on the temperature. The periodicity of the striations also depends on the concentration of the samples. Robinson found that for PBLG in dioxane the pitch is inversely proportional to the square of the concentration, while DuPre et al4S reported a dependence of c'18 for the same system. The choice of solvent has a remarkable effect in the striation separation in PBLG lyotropic liquid crystals, it has been found that the periodicity of the pattern can change as much as three times depending on the solvent (with the same polymer, concentration and temperature)45. In contrast to a small number of reports of studies of PSLG in liquid crystalline state, there is abundant literature of investigation of the liquid crystalline behavior for PBLG. Studies of the liquid crystalline phase diagram of PBLG in different systems have been reported by several investigators4,46. In molecular dynamic simulation studies, Helfrich et al47 calculated the range of the induced solvent structure for dilute solutions of PBLG in DMF to be 30 A, which corresponds to the effective diameter obtained from light scattering experiments. This indicates the importance of including 7 2 the solvation zone in the molecular excluded volume for experimental and theoretical considerations that depend on the lateral dimension of the particles. It was also found that at the border of the polymer rods, the solvent was highly structured. In studies on the rheology of a-helical polypeptides, it has been shown that liquid crystals of PBLG in benzyl alcohol produced three regimes in the shear dependence of the viscosity40. This dependence can be explained with respect to the response of the domain structure to the shear flow. At low shear rates, the first regime, the shear flow is not flow aligning and the director tumbles under flow. In this regime the domain structure is not altered, the domains just flow over each other, resulting in a shear-thinning regime. As the shear rate increases, second regime, the flow dampens the tumbling of the director and the domains start to break. Although the overall order of the system decreases dramatically in this regime, the molecular order of the solution is not greatly affected by the shear flow. In the last regime, the shear rate becomes so strong that it breaks up all the domains (the flow becomes shear aligning) creating a single domain. And the order increases with the increase of shear rate. It has been reported that cholesteric liquid crystals are highly non-Newtonian in their flow properties. A tremendous increase in the 7 3 apparent viscosity (r|app) is observed when the shear rate drops to a low value. Helfrich47 has also suggested that at low shear rates, flow occurs along the helical axis without the helical structure itself moving. Assuming that the velocity profile is flat, the energy gained by the translational motion of the fluid in the pressure gradient should be equal to that dissipated by the rotational motion of the director. For shear flow normal to the helical axis, the cholesteric is expected to behave more or less like a nematic. An interesting prediction is that for a given shear rate and sample thickness the apparent viscosity vs. pitch becomes comparable to the sample thickness48. Polymer melts of nonracemic mixtures of chiral (optically active) molecules often produce cholesteric liquid crystals. This is the case for most polypeptides and for PSLG in particular. But it has been reported44 that the chiral a-helical conformation of polypeptides produces a cholesteric twist that is related to the achiral solvent. The right- or left-handedness of the cholesteric twist is related to the form optical rotation. For example, the form optical rotation of PBLG liquid crystals is positive when prepared in dioxane, negative in methylene chloride, while PBLG itself is a right-handed a-helix in both solvents. In an 8:2 methylene chloride:dioxane mixture, the PBLG liquid crystal 7 4 is compensated (nematic). This is believed to be produced by the influence of the dielectric properties of the medium. 3.2 LIQUID CRYSTALS OF PSLG As mentioned before, in helicogenic solvents, such as THF, PSLG adopts an a-helical conformation. This conformation provides the polymer with a rigid backbone. Concentrated solutions of high molecular weight polymer form liquid crystals. The formation of a cholesteric liquid crystalline phase is due to the chiral nature of PSLG. 3.2.1 Phase Boundary Studies As shown in the theories of Onsager and Flory, the axial ratio is an important indicator of the potential of the polymer to develop liquid crystalline order. Since PSLG forms a-helices, the ratio ^ can be readily calculated from the side chain of the repeat unit and the degree of polymerization (DP). In a-helices, each peptide residue translates a distance h=1.5 A along the helix axis. Hence, for an ideal a-helix, L=DP xh, in Angstroms. The appropriate value for d is somewhat ambiguous. The diameter of the helical core of the polypeptide backbone chain itself is approximately 6 A. The side chains of PSLG extend out radially from this core and mix with solvent molecules. A lower limit for d may be calculated from the 7 5 density of the solid polypeptide and the value for L. Molecular modeling may also give some insight into estimates of d; however, a solvation layer should also be considered44. It is expected that for PSLG a correct value for d is 37 A Assuming the values for PSLG are 37 A for its diameter when it adopts an a-helix conformation, and the length for each monomer unit equal to 1 .5 A , the minimum degree of polymerization to form liquid crystals according to the Onsager theory is 82, which corresponds to a molecular weight of 31,000, for the appearance of the biphasic-isotropic phase boundary, ^a. The same theory predicts that for PSLG, a molecular weight of at least 42,000 (DP = 111) is required to reach the biphase-liquid crystal transition point, Flory’s theory calls for a degree of polymerization of 198, which corresponds to a molecular weight of more than 75,000, to form liquid crystals. A range of concentrations of solutions of nearly monodisperse PSLG of molecular weights 239,000, 208,000 and 129,000 in THF were studied using polarizing optical microscopy. At sufficiently high concentrations of polymer, all these samples formed cholesteric liquid crystals, as is evidenced by the presence of fingerprint patterns when observed with an optical microscope provided with cross 7 6 polarizers (Fig. 3.4). Tables 3.1, 3.2 and 3.3 show the characteristics of the different PSLG samples in THF at different concentrations. For conversion to volume fractions, the specific volume of PSLG was taken as 1.024 mL/g and the density of THF as 0.889 g/mL. It is assumed that the specific volume of PSLG does not change with concentration. Table 3.1: PSLG of Mw=239,000 (MWD=1.13) in THF at 25°C <|> % (v/v) 20.3 17.5 15.5 11.6 P h a se LC LC Bi-ph Iso Table 3.2: PSLG of Mw=208,000 (MWD=1.08) in THF at 25°C P h a se LC LC LC LC Bi-ph Iso Table 3.3: PSLG of Mw=129,000 (MWD=1.12) in THF at 25°C <|> % (v/v) 26.6 23.9 21.0 18.3 14.1 P h a se LC Bi-ph Bi-ph Bi-ph Iso LC: liquid crystalline phase; Bi-ph: biphase; Iso: isotropic phase It is evident that the critical concentration for the different PSLG samples varies with the molecular weight, the higher the molecular weight the lower the critical concentration. When viewed Fig. 3.4: Cholesteric liquid crystal PSLG in THF at 25.0°C (Mw= 239,000; MWD=1.13; <{>=20.3%) 7 8 through a microscope, the isotropic phase does not allow light to pass through the cross polarizers (black). Inside the biphasic region, the coexistence of the isotropic and the anisotropic phases is manifested by the presence of spherulites possessing black maltase- crosses as shown in Fig. 3.5. The spherulites are surrounded by a dark (isotropic) background. An 11.6% (v/v) solution of PSLG (Mw=239,000) in THF is in the isotropic phase. At 15.5% volume fraction of the same polymer, the isotropic phase coexists with the liquid crystalline phase. At concentrations of 17.5% or higher, only the liquid crystalline phase is present. Therefore, the observed 0 a for this molecular weight polymer is 13.6±2.0%, and 0b is 16.0±1.0%. PSLG of Mw=208,000 shows an isotropic phase at concentrations of 13.8% or lower. A solution with a volume fraction of 16.5% of this polymer lies in the biphasic region. At a concentration of 19.2% the solution is homogeneous, and the striation pattern of the cholesteric liquid crystalline phase can be observed throughout the entire solution. Consequently, for PSLG of Mw=208,000, the values for 0 a and 0 b are 15.2±1.4% and 17.9±1.4%, respectively. 7 9 % 0 100 pm Fig. 3.5: Spherulites with maltase-crosses of PSLG in THF at 25.0°C (Mw= 129,000; MWD=1.12; <(>=21.0%) 8 0 For PSLG of lower molecular weight (Mw=129,000) at 25°C, the coexistence of the isotropic and liquid crystalline phases is broader. Both phases coexist already at 18.2% volume fraction and this biphasic region continues to be present at 23.9%. A solution with concentration of 26.6% displays a uniform (iquid crystalline cholesteric phase. On the other hand, volume fractions of 14.1% or below show only isotropic phase. As a result, for PSLG of Mw=129,000, ^ a=16.2±2.1% and *}B=25.3±1.4%. Table 3.4: Comparison of experimental results and theoretical predictions for the biphasic region (concentrations in v/v%) Mw L/d Experimental Onsager Flory 239,000 25.4 13.6-16.0 13.0-17.7 29.0 208,000 22.1 1 5 .2 -1 7 .9 14.9-20.4 32.9 129,000 13.5 16.2-25.3 24.4 - 33.3 50.5 Table 3.4 shows the concentrations calculated for the boundaries of the biphasic region for the different polymers and the calculated values from the Onsager and Flory theories. It is clear that the concentrations at which these samples present the coexistence of the two phases more closely correspond to the former than to the latter theory. For example, for PSLG of Mw=239,000, we 81 observe the biphasic region from 13.6% to 16.0% (v/v). The Onsager theory predicts the limits of this region to be 13.0% to 17.7%, while the Flory theory calls for a volume fraction of 31.5% for the biphase boundary. The Onsager theory predicts the presence of the biphasic region for PSLG of Mw=208,000, to be between 14.9 - 20.4%; and the observed region apears within those limits (15.2% to 17.9%). For PSLG of Mw=129,000, the biphasic region is present from 16.2% to 25.3%, this region is broader than the calculated from the Onsager theory, 24.4 - 33.3%. This broadening of the biphasic region may be due to end-to-end aggregation of the polymers. Aggregation occurs more frequently in low molecular polymers. The Flory theory requires a volume fraction of 50.5% to observe the formation of the biphase region for this polymer. 3.2.2 Effect of Concentration and Temperature on the Pitch The distance between striations, S, corresponds to half the pitch of the cholesteric helix. It is usually used as a measure of the twisting power of the cholesteric super helix. It is also common to represent this characteristic as the reciprocal of half the value of the pitch (S'1). From studies on poly(y-methyl-a,L-glutamate), poly(y- ethykx.L-glutamate) and poly(y-propyl-a.L-glutamate), Uematsu et al49 have reported that there is a temperature and concentration 8 2 dependence of the pitch, P. Keating50 proposed a theory for the linear temperature dependence of the pitch, given by: where < is the average twisting angle and is directly proportional to the inverse of one half the pitch (S'1), k is the Boltzmann constant, a is the frequency of the excited twisted mode, / is the moment of inertia of the molecule, and A is a constant derived from the anharmonic equation of motion. Fig. 3.6 shows the dependence of the distance between striations with temperature for a sample of PSLG (Mw= 208,000) in THF, 28.4% (v/v). It is apparent that the 0.030- 0.025- i0 0.020- 0.015- 25 30 35 45 Temperature (°C) Fig. 3.6: Distance between striations vs. temperature, PSLG liquid crystal solution (Mw=208,000), 28.4% (v/v) in THF 8 3 inverse of half the pitch of the cholesteric helix is directly proportional to the temperature (slope= -8.2 ± 0.2 x 10"4 pm‘1/°C). Increasing the temperature increases the distance between the striations. Therefore, there is a temperature at which the pitch becomes infinity. This corresponds to the change from the cholesteric to the nematic liquid crystalline phase. This temperature can be obtained from the graph by extrapolating S — oo. T he temperature obtained is T=62.4°C. There is also a dependence of the inverse of half the distance of the cholesteric pitch with the concentration of the cholesteric liquid crystalline solution. However, this dependence is not linear, and varies with temperature. It has been speculated that the relationship is S '1 oc c" where the exponent, n, is a function of temperature and lies normally in betw een o n e and two49. For PSLG of molecular weight 208,000 different concentration solutions in THF were measured at 25°C. As described previously, at concentrations higher than 19.2%, solutions of this polymer show uniform cholesteric liquid crystalline phases. There is a clear decrease in the pitch of the liquid crystalline pattern as the concentration increases (for solutions more concentrated than the 8 4 interface boundary). Fig. 3.7 shows a plot of log(S_1) vs. Iog{volume fraction). The value for the slope calculated from the plotted data is 1.04 ±0.10 for 25°C. -1.50- -1.55- -1.60- I J . -1.65- t -1.75- -1.80 1.25 1.30 1.35 1.40 1.45 1.50 1.55 logft) Fig. 3.7: Distance between striations vs. concentration, PSLG (Mw=208,000) in THF at 25°C 3.3 EXPERIMENTAL An Olympus BH optical microscope with cross polarizers equipped with a 35 mm camera was used, the objective utilized was 10X. The temperature control system used was a Mettler FP82 Hot Stage. The magnification was obtained directly by photographing a micrometer scale (100 x 0.05 = 5.0 mm). 8 5 Procedure. PSLG of molecular weights 239,000, 208,000 and 129.000 daltons, with MWD of 1.13, 1.08 and 1.12 respectively, synthesized as described in chapter 2 were used for these studies. Reagent grade THF was distilled over calcium hydride and kept under Argon and 4A-molecular sieves. Microslides from Vitro Dynamics Inc. with a path length I.D. of 0.6 mm and a width I.D. of 6.0 mm were used. The cells were flame sealed on one end, then filled with a known amount of solid polymer (usually in the range of 5 to 10 mg, depending on the desired solution concentration) followed by the needed amount of THF. The cell is then centrifuged and the open end is flame sealed rapidly. The concentration of the solution is obtained gravimetrically. For conversion to volume fractions, the value of the specific volume of PSLG was taken as 1.024 cm3/g and the density of THF as 0.889 g/cm3. At room temperature, the solutions were visually clear, and on the polarizing microscope the high concentration samples showed the typical striation pattern produced when cholesteric liquid crystals are present. CHAPTER 4 MODIFICATION AND CROSSLINKING OF PBLG 8 6 8 7 4.1 INTRODUCTION Polymer chains that are connected together form a crosslinked or network polymer. The polymer chains can be chemically crosslinked by means of covalent bonds (creating what is known as thermosets) while others can be linked physically by weak intermolecular interactions such as hydrogen bonds, ionic interactions or van der W aals forces. The term gel is commonly used to refer to a network polymer swollen in a liquid medium. However, there are some ambiguous definitions for gels, like a system with no fluidity (failure of an air bubble to raise in it), or a system with infinite viscosity. These networks do not dissolve, they are just swelled by the solvent. The polymer network retains the solvent, and at the same time the solvent prevents the network from collapsing. The amount of swelling of the polymer depends on the density of crosslinking; high number of crosslinks account for little swelling. Few crosslinks in a polymer network usually correlates with rubber-like properties, while a high density of crosslinks imparts more rigidity to the system (the extreme example is diam ond). This insolubility of gels makes crosslinking reactions commercially important, since crosslinked plastics are very stable at elevated temperatures and under physical stress. Normally, these crosslinked systems do not flow when heated and do not change their dimensions 8 8 drastically over a variety of conditions making them very useful for structural applications. 4.2 REACTION ON THE SIDE CHAINS OF PBLG The presence of the ester linkage in the side chain of PALGs provides a site for the binding of different moieties to the rigid rod polymer. This reactive site is the common target for usual chemical modifications of these polymers. And since the ester group is not part of the helical backbone, generally, these reactions do not disrupt the conformation and therefore, the rigidity of the polymer, unless the group attached interacts with the intramolecular hydrogen bonds in the helix. Three types of reactions are commonly performed at this particular functional group, conversion of PALGs to poly(a-glutamic acid), conversion to poly(y- glutamine) derivatives by amidation, and ester interchange reactions. These modifications are usually performed when there are problems with the preparation of the targeted polymers directly from the corresponding NCA monomer. In some cases it is difficult to prepare the required y-ester of glutamic acid, or the synthesis of the NCA monomer is unfavorable because of side reactions. In many cases, even though the chemical modification of the side chains is the only way to obtain certain polymers, problems such as incomplete reactions or by-product formation are very common, and do not 8 9 permit a complete transformation of the polymer. In such cases, the products usually have impurities bound to the polymer chain (producing a copolymer instead of a homopolymer) that cannot be eliminated by normal purification techniques. In other cases, side reactions that result in cleavage of the peptide bond in the polymer backbone result in a decrease in the molecular weight of the polymer, and change in its molecular weight distribution. Amidation of PALGs and poly(glutamic acid esters) have been reported previously13 by reacting the polymer with ammonia or an appropriate amine. One application of these amidation reactions is the formation of water soluble polymers or copolymers due to the presence of glutamyl residues when amino alcohols are reacted. These modifications also have an effect on the dyeability of fibers of PMLG among other properties. However, in this particular case, one complication that is not uncommon is the occurrence of some transamidation of the peptide bonds in the polymer backbone originating a reduction in the molecular weight of the product. It is a common practice in organic chemistry to cleave molecular alkyl carboxylic esters under acidic or basic conditions. Some procedures allow cleavage to occur under neutral conditions but they usually require the use of strong nucleophiles and high temperatures to effect dealkylation. It has been reported that these reaction conditions also react with the peptide 90 bonds in the backbone of the PALGs consequently lowering its molecular weight51. Transesterification reactions on the side chains of PALGs are usually carried out in dioxane or a chlorinated hydrocarbon with the desired alcohol in the presence of a strong acid catalyst, such as p~toluene sulfonic acid. The reactions usually require high temperatures and usually proceed to more than 80% conversion; however, the transesterification is never totally complete, and there are always some residual groups from the original polymer27. A different approach for the dealkylation or debenzylation of carboxylic esters involves the reaction of trimethylsilyl iodide with the ester. It has been previously reported52'56 that this is an extremely efficient alternative, and the reaction occurs under very mild conditions. The reaction proceeds in essentially quantitative yields under neutral conditions. Trimethylsilyl iodide cleaves the carboxylic ester to produce the corresponding alkyl iodide and a reactive trimethylsilyl carboxylate. The reaction conditions depend on the ester that is been cleaved; benzyl esters are one of the most reactive54'56. Subsequent treatment of the silyl ester with an alcohol produces a transesterification (Fig. 4.1), a reaction with an amine forms an amide as the product. The solvent choice is. important, and trimethylsilyl iodide is usually used in chlorinated hydrocarbons, such as carbon tetrachloride or chloroform. Trimethylsilyl iodide reacts with THF. 91 R. O— R' + CHi— Si— I Yo / J R. .0—Si—CH3 + R' 1 o Fig. 4.1: Reaction of an ester with trimethylsilyl iodide Our effort has been directed towards the application of this procedure used for small organic molecules to polymers containing the same functional groups in the pendant chain. This methodology activates the ester linkages in the side chain of poly(glutamic acid esters) in a very controlled manner, and at the same time under mild conditions. For these studies PBLG was chosen because the benzyl group forms benzyl iodide readily after addition of trimethylsilyl iodide, permitting a very controlled activation of the sites in the polymer under neutral conditions and at room temperature. These active sites can react later with molecules that have reactive functional groups and which can become attached to the polymer 9 2 replacing the benzyl group. Molecules that have amino and isocyanate functional groups have been successfully bonded to the polymer at the activated sites. This chapter describes the reaction of trimethylsilyl iodide with carboxylic esters. The first part accounts for cleavage of small esters as a model reaction. Later in the chapter, the description of reactions at the side chains of PBLG are reported, and using the same methodology, the crosslinking of PBLG is explained. 4.2,1 Cleavage of Small Esters with Trimethylsilyl Iodide As a model reaction, two small esters were tested in their reactivity with trimethylsilyl iodide, benzyl acetate and methyl benzoate. A difference in reactivity is observed, benzyl acetate reacts faster than methyl benzoate. This is in agreement with reported results that state the good leaving nature of the benzyl group. The reaction of benzyl acetate and trimethylsilyl iodide produces trimethylsilyl acetate and benzyl iodide (Fig. 4.2). The cleavage of the ester group is distinctly followed by 1H-NMR spectroscopy. The disappearance of the benzylic hydrogens of the ester (5 - 5.0 ppm) and the appearance of the benzylic hydrogens of benzyl iodide, which appear at 5 - 4.5 ppm, prove the cleavage of the ester (Fig. 4.3). Trimethylsilyl acetate reacts with amines and isocyanates at room temperature. Addition of benzyl amine to the trimethylsilyl acetate produced N-benzyl acetamide. Reaction in chloroform of phenyl isocyanate with 93 Ou A Si(CH3)3 + (CH3)3S i - I ------► + Fig. 4.2: Reaction of benzyl acetate with trimethylsilyl iodide trimethylsilyl acetate formed N-phenyl acetamide. The reaction of a trimethylsilyl carboxylate ester with isocyanates has not been reported previously. 4.2.2 Activation of PBLG Side Chains PBLG was used to test the formation of the silyl ester derivatives in polymers. The reaction of PBLG with a small amount of trimethylsilyl iodide produces a partially silylated derivative of PBLG and benzyl iodide (Fig. 4.4). Similarly to the small ester molecules, this reaction can also be followed by 1H-NMR spectroscopy. The broad peak of benzylic hydrogens in the polymer appears at 5 - 5.0 ppm and shifts to a narrow peak at 6 - 4.5 ppm once benzyl iodide has been formed (Fig. 4.5). This reaction is performed also at room temperature. Trimethylsilyl iodide is added in the desired amount, usually corresponding to 10 to 20 mole% with respect to the monomer units of PBLG. 94 10 h of reaction J 1 10min of reaction 8.0 7.06.0 5.0 4.0 3.0 2.0 1.0 PPM Fig. 4.3: ’H-NMR of reaction of benzyl acetate with trimethylsilyl iodide in CDCb 9 5 O jO ojO Si(CH3)3 l-SKCH,), r 0 ^ 0 ° ^ o n o Fig. 4.4: Activation of the side chains of PBLG with trimethylsilyl iodide 4.2.3 Reaction of Activated PBLG with n-Butyl Isocyanate At this point, a portion of the side chains in PBLG have been cleaved and, the reacted PBLG has some active sites (trimethylsilyl esters). n-Butyl isocyanate reacts with the partially silylated PBLG at the active sites, at room temperature, to produce n-butyl amides in these side chains (Fig. 4.6). After the reaction is complete, the polymer is recovered from the solution by precipitation in methanol and the presence of the n-butyl hydrocarbon chain is evident in its ’H-NMR spectrum (Fig. 4.7). 96 20 h of reaction time r 8.0 r.o 6.0 s . o 4.0 3.0 2.0 1.0 PPM Fig. 4 .5 :1H-NMR of PBLG and PBLG reacted with trimethylsilyl iodide, in CDCI3 97 s Fig. 4.6: Reaction of n-butyl isocyanate with the partially silylated PBLG 4.3 CROSSLINKING OF PBLG Usually, crosslinked gels of synthetic polymers are isotropic and show no microscopic and macroscopic order. And correspondingly, the mechanical and optical properties of these gels are also isotropic. However, during the last years, many research efforts have been directed to study the potential to stabilize mesomorphic structures of polymers in the solid state (and the accompanying physical properties of these materials). The retention of ordered structures in the solid state offers immense possibilities in materials for non linear optical and electronic devices. This is because the anisotropy of the polymer (microscopic) is translated into a 8.0 7.0 6 0 5.0 4.0 3.0 2.0 1.0 PPM Fig. 4.7:1H-NMR of PBLG reacted with n-butyl isocyanate, in CDCI3 ID GO 99 is because the anisotropy of the polymer (microscopic) is translated into a anisotropy of the gel (macroscopic) conferring the gel material with unusual properties. Such an anisotropic gel possesses asymmetric swelling properties or asymmetric response to external stimuli, like temperature, pH, solvent interaction, etc. Furthermore, the use of these organized materials in the formation of composites with other random polymers or as part of interpenetrating polymer networks (IPNs) is very promising. Control of the density of the crosslinks and the distance between the rigid polymer chains in a network is promising for separation techniques; very specific filtration membranes can be tailored in this fashion. The first attempts to retain specific mesomorphic structures in polymers started in the 1930s with the quenching of some thermotropic phases to low temperatures to form a brittle glassy state in which the liquid crystalline structure persisted44. In the case of lyotropic liquid crystals, similar results could be obtained by techniques such as freeze-drying or simply by slow evaporation of the solvent. However, these methodologies do not produce crosslinked networks, the mesomorphic state, although transferred in the solid state, is only momentarily maintained, increasing the temperature near the melting point of the polymer or the addition of solvent destroys the order. A truly permanent ordered state that remains under a variety of different conditions requires the bonding of the polymer chains chemically while they are in an ordered state. 100 Immobilization of the cholesteric liquid crystalline structure of PBLG has been attempted by y-irradiation of a dry film of PBLG or a film of mixture of PBLG and a plasticizer57. A mixture of PBLG and polyethylene glycol in dimethylformamide cast to form a film possesses cholesteric liquid crystalline order. In this case the cholesteric liquid crystalline order is stabilized not by covalent bonds, but by hydrogen bond networks. In another report58, the cholesteric liquid crystalline structure of a concentrated polypeptide solution is stabilized by using a vinyl monomer as a solvent and then immobilize it by polymerizing the vinyl monomer in the presence of a small amount of divinyl monomer as a crosslinker. Nonetheless, in all these studies there is no formation of a gel that possesses the cholesteric order, and there are no covalent crosslinks between the rigid polymer chains. Kishi et al58 have reported the fixation of PBLG in the lyotropic liquid crystalline state using diamines as crosslinking reagents, in one of the crosslinking reactions, the concentrated polymer solution was mixed with the corresponding diamine and kept at 25°C for 10 days to develop the cholesteric liquid crystalline order. Once the liquid crystal solution was formed, the sample was heated to 70°C in a sealed glass cell for 10 more days to speed the crosslinking reaction. The gel produced retained the high ordered state as the original solution. Aviram59 and Kishi et al60 have also reported the crosslinking of a concentrated solution of PBLG in dioxane using triethylenetetramine and 101 diethylene glycol bis(3-:.aminopropyl) ether as the crosslinking reagents. The crosslinking reaction was performed under the influence of an external magnetic field to prepare polymer gels with nematic liquid crystalline order. The reaction was carried out at 55-70°C for 7-10 days under a magnetic field of 21 kG perpendicular to the glass cell. In the nematic liquid crystal state the polypeptide helices are known to be oriented in the direction of the magnetic field. These authors also reported the loss of the anisotropy in the gel when dissolved in a non-helicogenic solvent, but its recovery when the solvent was changed back to the original one. 4.3.1 PBLG Isotropic Gels The methodology used to react the side chains of PBLG with trimethylsilyl iodide under mild conditions described earlier in this chapter is also effective to produce covalent crosslinks between polymer chains when the reagent that reacts with the activated sites is difunctional. Once more, the reaction occurs under neutral conditions at room temperature. The activated polymer chains (trimethylsilyl carboxylate esters) were treated with a crosslinker molecule with isocyanate groups at both ends. Dilute solutions of PBLG in chloroform (between 0.5 and 2.5 % in weight) activated with trimethylsilyl iodide produced transparent gels when 1,6-diisocyanato hexane was added as the crosslinker (Fig. 4.8). The amount of activated sites (trimethylcarboxylate esters) in the polymer corresponds to 10 to 20 mole % (with respect to the monomer unit ojD © s - o O SKCHih sN=C=0 Fig. 4.8: Scheme of the crosslinking of activated PBLG with 1,6-diisocyanato hexane 103 concentration). One half of that number of moles of the crosslinking reagent was added to form the gel. The gelation takes from 4 to 7 days at room temperature, depending on the concentration of the polymer solution and the amount of activated sites. Crosslinked networks were also obtained using a copolymer of benzyl and stearyl glutamates, 30:70 ratio of benzyl to stearyl units (its synthesis is described in chapter 2). The better solubility of poly(y-benzyl- a,L-glutamate)-co-poly(y-stearyl-a,L-glutamate) allows a higher concentration of the polymer in chloroform, making the crosslinking reaction easier to detect. The reaction conditions are the same for the crosslinking of PBLG. However, in this case, the gel formation takes a longer time, some reactions requiring as long as 22 days. This could be explained by steric factors introduced by the long paraffinic stearyl chains. The formation of a microgel of PBLG and a copolymer of PSLG and PBLG has been followed by light scattering techniques. Dynamic light scattering is a powerful tool to measure the motion of particles. When light is scattered by particles in a solution, there is a fluctuation of the intensity of the scattered light with time. This fluctuation, which is produced by the interference of the scattered light from different points in the measured volume, is therefore, dependent on the velocity at which the particles move in the solution. These fluctuations are not random however, there is a 104 correlation between them at appropriate time scales. From these fluctuations, a correlation function, is obtained: g® (x)= 1 + f lg (1)(x)l2 w here g (1)(x) = e 'rt ; r = q2D and q = (4jin/Xo) sin(0/2); x is the correlation time, f is an instrumental parameter (0 < f < 1), D is the mutual diffusion coefficient, Xo is the vacuum wavelength of the laser, n is the refractive index of the solution, and 0 Is the scattering angle. From cumulant analysis61,62 of the raw data obtained at different stages of the crosslinking process, the variation in the decay rate r and the hydrodynamic radius of the network can be observed. The results show a decrease of the decay rate (which translates into a decrease of the diffusion coefficient or increase in the hydrodynamic radius) as the gelation progresses. Figs. 4.9 and 4.10 show the change in the apparent diffusion coefficient and the hydrodynamic radius as a function of time for the crosslinking of PBLG and the copolymer, respectively. Fig. 4.11 shows the change in the distribution of the apparent diffusion coefficient of the growing nework as a function of time for PBLG and the copolymer, respectively. In these graphs, the distribution shifts to the left (lower apparent diffusion coefficient) as the network increases in size, which is equivalent to a change in size distribution towards larger particles. From these data collected from dynamic light scattering it is evident that crosslinking reactions are taking place. A Fig. 4.10: C hange in hydrodynam ic radius and ap p aren t diffusion coefficient coefficient diffusion t aren p ap and radius ic hydrodynam in hange C 4.10: Fig. Fig. 4.9: C hange in hydrodynam ic radius an d ap p aren t diffusion coefficient diffusion t aren p ap d an radius ic hydrodynam in hange C 4.9: Fig. Apparent Diffusion Coefficient (cm2/s) 5.0x10 1.0x10' 2.0x10' 1.5x10' 0.0 0.0 cosikn fpl(L)c-oySG occurs poly(BLG)-co-poly(SLG) of crosslinking s a 0 a s crosslinking of PBLG occurs PBLG of crosslinking s a 0 0 2 50 Rnm Time (hr) Time Dapp ie (hr) Time 400 100 a D — Radius app 150 800600 2000 4000 6000 8000 10000 12000 60 80 » Z) ais (nm) Radius 105 Fig. 4.11: Diffusion coefficient distribution for th e crosslinking of (A) PBLG PBLG (A) of crosslinking e th for distribution coefficient Diffusion 4.11: Fig. Distribution 0 0 0 0 0 10 10 10 10 10 10 -16 an d (B) poly(BLG)-co-poly(SLG) (se e text for explanation) for text e (se poly(BLG)-co-poly(SLG) (B) d an 120 h 120 177 h 177 20 h 20 70 h 70 h 82 ifso (m/) ifso (cm2/s) Diffusion (cm2/s) Diffusion A (B) (A) -14 -12 310 h 310 -16 134 h 134 597 h 597 78 h 78 -14 -12 106 107 limitation of the extent Qf network size produced in this experiment is the high dilution of the polymer concentration. A relatively low concentration is required to filter the sample before crosslinking to remove any dust particles prior to measurements by light scattering. This low concentration limits the size of the networks produced, however, microgelation is apparent. Fractal geometry provides a quantitative description of complex structures. A fractal is an object that is self-similar and possesses no intrinsic length scale (it is scale invariant to an isotropic change of length scale). An intuitive approach to characterize quantitatively a fractal is to observe the change of a physical property with its size. For example, the change of the mass, M, with the radius of the object, R, are related by M ~ R Dr where the exponent Df, the fractal dimension, characterizes the long-range dilation symmetry. The concept of fractal structures can be applied to studies of the aggregation of particles to form larger clusters. Weitz et al6^65 have studied the aggregation regimes of colloids, and have found two distinct regimes. One regime of cluster aggregation is diffusion-limited, in which clusters aggregate immediately upon contact with each other. In this case, the aggregation kinetics are determined by the diffusion of the clusters. The second regime, called reaction-limited aggregation, is slower than the first and is characterized by an exponential growth of the aggregates. 108 Fractal dimensions can be obtained from the slopes of logarithmic plots of aggregation rate with colloid concentration, mass vs. size of the clusters, or radius of the cluster vs. reaction time. The fractal dimension is the inverse of the slope of these plots. Weitz et al have found that a fractal dimension of ~1.75 corresponds to diffusion-limited cluster aggregation while a fractal dimension of -2.0 corresponds to a slower reaction-limited aggregation process. Similar fractal analysis of the crosslinking of poly(BLG)-co-poly(SLG) results in plots presented in Fig. 4.12. Fig. 4.12-A shows a logarithmic plot of the growth of the hydrodynamic radius as the crosslinking reaction progresses. Even though the data presents a linear behavior, the linear fit produces a slope of 1.82638±0.14314, corresponding to a fractal dimension of 0.5475, which has no physical significance. On the other side, the semi log plot (Fig. 4.12-B) shows an exponential growth of the gel (slope= 0.0028±0.00013), similar to results obtained by Weitz et65 al for reaction- limited cluster aggregation. 4.3.2 PBLG Liquid Crystal Gels The gels obtained in the previous section are composed of rigid rod polymers connected covalently. Even though the rod polymers are anisotropic, since there is no macroscopic order among the rods, the gel is isotropic (Fig. 4.13-A). To produce an anisotropic gel the rods must be Fig 4.12: (A) Logarithmic plot and (B) sem i-log plot of th e crosslinking e th of plot i-log sem (B) and plot Logarithmic (A) 4.12: Fig R/nm 10000 1000 100 - 10 - - -6.5 9 7 . . . 0 5 0 - - 4 1.6 - - - 0 20 0 40 0 60 0 800 700 600 500 400 300 200 100 2.0 of poly(BLG)-co-poly(SLG) of . 2.4 2.2 me/ ) /h e im t { g o I time/h 2.6 2.8 3.0 109 110 ordered, and the crosslinking reaction has to occur after alignment is achieved (Fig. 4.13-B). One way to accomplish this is to apply an external electric field to the polymer solution to create a nematic liquid crystal phase prior to the gelation. In this fashion the highest possible induced anisotropy of the network may be obtained. This process of aligning the polymer chains under an external field is often called poling. B Fig. 4.13: (A) Isotropic network of rod-like polymers, and (B) Anisotropic network of rod-like polymers For this procedure the setup shown in Fig. 4.14 was used. It consisted of two one-sided indium-tin oxide (ITO) coated glasses (conductive faces in the inside, in contact with the sample) connected to a power source. In a preliminary test of the effectiveness of the setup, a solution of PSLG (Mw= 208,000; MWD=1.08) in THF (20% v/v) was placed between the electrodes. Upon observation under the microscope with cross polarizers the expected cholesteric liquid crystal pattern is evident. Once the electric field is connected to the electrodes, the pattern vanishes very 111 rapidly showing the destruction of the cholesteric superstructure. This effect is due to the orientation of the polymer chains with the electric field. For the crosslinking of PBLG under the effect of the electric field, a chloroformic solution of the activated PBLG mixed with the crosslinker (1,6- diisocyanatohexane) is injected in between the conductive glasses and inside the O-ring. After the interior of the O-ring is filled and no air bubbles are present, the plates are held in place by clamps and an electric field of 40 V is applied. The reaction mixture is allowed to react for 7 days at room temperature, after which a gel is obtained. The gel is insoluble in chloroform and retains its shape. When viewed through an optical microscope provided with cross polarizers, the gel transmits light and appears to show the presence of the liquid crystalline phase. SAMPLE VOLTAGE ITO g la ss Fig. 4.14: Setup for crosslinking under electric field 4.4 EXPERIMENTAL Trimethylsilyl iodide was purchased from Aldrich, it needs to be stored under anhydrous conditions in the freezer. It was handled under N2 112 or Ar atmosphere. The crosslinking reagent, 1,6-diisocyanatohexane, was also purchased from Aldrich, also required anhydrous conditions. The solvent used in all the reactions is dry deuterated chloroform, also from Aldrioh. All the following reactions were carried out at room temperature and neutral conditions. To produce an electric field a Hewlett Packard, 6515A DC power supply unit was connected to the cell described in the previous section. 1H-NMR spectra was recorded on an IBM Bruker200 MHz or 250 MHz instrument. Formation of liquid crystals was detected by a polarizing optical microscope. Dynamic Light Scattering. Samples for dynamic light scattering studies were reaction mixtures of 1-2% concentration in chloroform of the activated- PBLG. The solutions were filtered through 0.2 pm PTFE filters (from Whatman) into dust free glass cells. The dynamic light scattering data was collected at 25°C, at an angle of 90°. The apparatus consisted of a He-Ne laser (A= 6328 A), and a photomultiplier detector. The data is collected and analyzed by an ALV-5000 digital autocorrelator software. Synthesis of Trimethvlsilvl Acetate and Trimethvlsilvl Benzoate. 100 pL (0.69 mmol) of benzyl acetate were dissolved in 2 mL of chloroform. 90 pL (0.63 mmol) of trimethylsilyl iodide were added at room temperature (~25°C). The reaction was followed by 1H-NMR (formation of benzyl 113 iodide). After 10 hours, the reaction was completed, and the presence of trimethylsilyl acetate was confirmed by GC-MS. Methyl benzoate (100 pL, 0.80 mmol) was reacted with trimethylsilyl iodide (110pL, 0.77 mmol) in chloroform at room temperature. There was still some unreacted methyl benzoate after 24 hours of reaction. The formation of trimethylsilyl benzoate is also followed by ’H-NMR (methyl iodide formation), and confirmed by GC-MS. Synthesis of Acetanilide via Trimethvlsilvl Acetate and Phenvl Isocyanate. The reaction of trimethylsilyl acetate with phenyl isocyanate occurs in 4 hours. Phenyl isocyanate (65 pL, 0.60 mmol) was added to the reaction mixture of benzyl acetate and trimethylsilyl iodide described above. Crystals of acetanilide (n-phenyl acetamide) were obtained. The product was identified by IR (KBr, cm'1: 3328, N-H stretching; 1650, C=0 amide I band; 1595 and 1556, N-H bending amide II band; 754 and 698, out of plane N-H wagging) and ^-NM R (DMSO, 5 in ppm: 2.15, s, CH3; 7-7.6, m, aromatic ring; 8.5, s, NH) spectroscopy. Synthesis of Activated-PBLG. When trimethylsilyl iodide reacts with the side chains of PBLG it forms an activated-PBLG. The activation of the ester groups to trimethylsilyl carboxylate esters was completed in 24 hours. The reaction was carried out also in chloroform and at room temperature. 114 In a typical reaction, 0.030 g of PBLG were allowed to dissolve in 1.5 mL of chloroform overnight. 2.0pL of trimethylsilyl iodide were added at once; the amount corresponding to 10 moie% of the monomer units. The mixture reacted for 24 hours. The formation of benzyl iodide was followed by 1H-NMR. Reaction of Activated-PBLG with /7-Butvl Isocyanate. n-Butyl isocyanate was added to the activated-PBLG obtained previously. The amount of n- butyl isocyanate corresponded to the number of activated sites. The reaction of activated-PBLG with n-butyl isocyanate is completed in 2 days (no further change in ^-NM R spectrum). The modified polymer is then precipitated in methanol, filtered and dried under vacuum. The product, which is a copolymer containing benzyl glutamate and n-butylamide of glutamic acid side chains, presented 1H-NMR peaks at 5= 0.9 ppm (methyl group in side chains), 5= 1.3 to 1.5 ppm (methylene groups in n-butyl side chains) corresponding to the 4-carbon linear chain, in addition to the peaks of the regular PBLG. Crosslinkina of Activated PBLG. For the crosslinking reactions, PBLG, with 10-15 mole % activated units (activated with trimethylsilyl iodide, as described above) was reacted with the crosslinker, 1,6-diisocyanatohexane at room temperature, until the solution gelled, which usually occurs after 4 days. Once the gel was formed, it was washed with chloroform several times to remove any by-products. 115 In a typical preparation, 0.030 g of PBLG dissolved in chloroform (~2% w/w) were reacted with 2.0 pL of trimethylsilyl iodide as described previously. After the activation was completed, the crosslinker, 1,6- diisocyanatohexane (1.1 pL, 0.007 mmol) was added. The amount of crosslinker corresponds to one half the number of activated sites in the polymer. The reaction was stirred at room temperature until a gel was formed. Crosslinkina of Activated-PBLG under an Electric Field. For the crosslinking of the aligned PBLG in an electric field, a procedure similar to the one described in the previous section was performed. Immediately after the crosslinker was added, the reaction was transferred to a cell. The cell was composed of two electrodes made of one-sided ITO coated glass (conductive sides facing each other). A 3/8” ID x 1/2"ID x 1/16" width Viton O-ring (from Small Parts Inc.) was placed in between the two conductive surfaces. These O-rings are resistant to chloroform. Once the reaction mixture was in the cell, the glass plates were compressed to avoid any leakage, and the electrodes were connected to the power source. The distance between the electrodes is ca. 1.0 mm. A differential potential of 40 V was applied for 7 days, at room temperature. The gel was removed from the cell and washed with chloroform to remove any by-products. BIBLIOGRAPHY 117 1. Kroschwitz, J. I. Concise Encyclopedia of Polymer Science and Engineering; John Wiley & Sons, Inc., New York, 1990; pp 449-451. 2. D esstpri, E.; Tirrell, D. A. Macromolecules 1994,27, 5463. 3. Watanabe, J.; Ono, H.; Uematsu, I.; Abe, A. 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A.; Huang, J. S.; Lin, M. Y.; Sung, J. Physical Review Letters 1985, 54, 1416. 65. W eitz, D. A.; Lin, M. Y.; Huang, J. S.; W itten, T. A.; Sinha, S. K.; Gethner, J. S.; Ball, R. C. Scaling Phenomena in Disordered Systems; Plenum Publishing Corp., New York, 1985; pp 171-186. VITA Javier Nakamatsu was born on May 25,1965, in Lima, Perti. After graduating from high school in 1981, he did his undergraduate studies at the Pontificia Universidad Catdlica del PerO in Lima. In 1989 he received his Bachelor in Science degree in Chemistry. He came to Baton Rouge, Louisiana, in 1990, to enroll in the graduate school at Louisiana State University. He is currently a candidate for the Doctor of Philosophy degree in Chemistry. His research area is in the field of Macromolecular Chemistry. 121 DOCTORAL EXAMINATION AND DISSERTATION REPORT Candidate: Javier Nakamatsu Major Field: Chemistry Title of Dissertation: Synthesis, Characterization, Liquid Crystals and Crosslinking ofPoIy(y-Alkyl-a,L-Glutamates) Approved: Major Professor Chairman Deahof the Graduate School EXAMINING COMMITTEE: i Date of Bxamlnat ion: November 1, 1995