Fundamental Aspects of the Chemical Applications of Cross-linked

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Downloaded by UNIV OF CALIFORNIA SANTA BARBARA on May 27, 2018 | https://pubs.acs.org Publication Date: July 19, 1982 | doi: 10.1021/bk-1982-0193.ch027

Fundamental Aspects of the Chemical Applications of Cross-linked Polymers 1

DAVID LIVE and STEPHEN Β. H. KENT

The Rockefeller University, New York, NY 10021

High resolution NMR spectroscopy has been used to study in detail the properties of swollen copoly (styrene-1%divinylbenzene) with an emphasis on fac tors relating to i t s use as a support in novel solid-phase methods for chemical synthesis. 1% cross-linked and linear polystyrene i n chloroform and dimethylformamide have been compared. We have been able to demonstrate a high degree of mobility in chain segments at frequencies > 10 /s uniformly throughout the resin comparable with linear poly styrene. This implies good solvation of the chain segments even though the polymer beads are macroscopically insoluble. Pendant chains show an even greater degree of freedom, and their effects on the properties of the system have been investigated. The environment i n the swollen 1% cross-linked bead is substantially l i k e a solution of linear polymer with no evidence for a fundamental resin-caused physical limitation to the solid-phase synthesis method. 8

The novel concept of synthesizing a molecule while attached to a swollen cross-linked resin bead was introduced and demon strated by R. B. Merrifield with the solid-phase peptide synthesis method about 20 years ago(1,2). The procedure involves the covalent attachment of an amino-acid residue to the polymer bead followed by the addition of subsequent amino-acid units i n a step wise manner under conditions that do not disrupt the attachment to the support. At the completion of the assembly of the peptide, the product i s cleaved from the resin and recovered. The macroscopically insoluble support provides convenient containment of the desired product so that isolation and purification from soluble co-products i n the synthesis can be achieved by simple

1

Current address: Molecular Genetics, Incorporated, Minnetonka, Minnesota 55343.

0097-6156/82/0193-0501$06.00/0 © 1982 American Chemical Society Mark and Lal; Elastomers and Rubber Elasticity ACS Symposium Series; American Chemical Society: Washington, DC, 1982.


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washing and f i l t r a t i o n procedures. The speed and s i m p l i c i t y of the manipulations are an important advantage compared to convent i o n a l solution synthesis methods. This allows for the standardization of reaction conditions i n the solid-phase method. Recently i t has been realized that there are also i n t r i n s i c chemical advantages which derive from the solvation effects imparted to the pendant molecule by the supporting polymer(3). This has the potential for extending the range of environments i n which a particular chemical transformation can take place which has important implications for the general usefulness of the solid-phase method. Although a variety of supports have been employed for s o l i d phase syntheses i n general and the peptide version i n particular(4), polymerization have been by far the most widely and successfully employed. In particular co-poly(styrene-divinylbenzene), the support o r i g i n a l l y u s e d ( 2 ) , i s the most popular, t y p i c a l l y 1% cross-linked. Among i t s advantages are favorably physical and chemical properties and a v a i l a b i l i t y . From time to time, however, questions have been raised about limitations on the use of this methodology purportedly arising from inadequacies i n the physical properties of this support. Generally the hypotheses have been based on r a t i o n a l i z a t i o n of chemical d i f f i c u l t i e s and have not been substantiated by direct physical i n v e s t i g a t i o n s ( 5 , 6). I t i s our goal to investigate the physical properties of the swollen, cross-linked resin-pendant molecule system to develop a fundamental understanding of the behavior of components i n the system and their interplay. This has been accomplished by examining the molecular structure and dynamics of the polymer and pendant chain at a molecular l e v e l using high-resolution NMR spectroscopy. The study has been carried out i n the context of the solid-phase peptide synthesis method because i t offers a means of correlating chemical to physical effects, p r a c t i c a l a p p l i c a b i l i t y of the results to a widely used technique, and proven chemical procedures for manipulating the system. The relevance of the results i s not limited to improving the use of these supports i n chemistry, but also to provide a better understanding of the phenomena of rubber e l a s t i c i t y . Approach Although high-resolution NMR has been widely used i n studying polymers(7, Cl /CD>Cl . Shifts relative to TMS. 2

2

2

Mark and Lal; Elastomers and Rubber Elasticity ACS Symposium Series; American Chemical Society: Washington, DC, 1982.

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a

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Y

C , the solvent, the C , the C , the C , and the C respect ively. Weaker l i n e s deriving from other portions of the peptide are also v i s i b l e i n this region. The peak immediately downfield of the C resonance i s from the methine of the PS. The aromatic region around 129 ppm i s made up of resonances from PS phenyl carbons and the aromatic carbons of the ΡAM group and the benzyl group. Some of the l a t t e r ones are probably responsible for the sharp component of the peak. In relaxation studies i t was clearly observed that there was a sharp central component that relaxed more slowly than the rest. These non-styrene carbons should ac count for about 20% of the intensity i n the region and they w i l l tend to bias the width measurement, T , and Overhauser enhancement values since the peak i n that region was considered a single unit. Table I I I 13 C Linewidth(Hz), Nuclear Overhauser Enhancement Factor(η) and T ( s ) of H N-IleAlaGlu(Y-0-Bzl)-PAM-(S-DVB) i n Chloroform and Dimethylf ormamide. x

2

a

Aromatic* He

C

a

5

LW

η

125.9 (87. 4)

O. 82 (O. 98)

O.37 (O. 67)

29.6 (25. 2)

1. 32 (1. 32)

O.46 (O. 42)

28.6 (23. 1)

1. 21 (1. 38)

O.38 (O.45)

29.4 (28. 0)

1. 44 (1. 46)

O.33 (O. 34)

h

1. 70 (1. 34) O.57 ( O . 67) c O.64 (O. 86) 24.9 (24. 4) 1. 67 (1. 62) DimethyIformamide value follows the chloroform value i n parentheses. Measured at 75 MHz. 24.3 (23. 6)

&

a

Includes contirgutions from a l l aromatic carbons. 0 Ν C'Y Labeling of isoleucyl reisidue i s Λ J.B. Γ γ Γ δ 2

The widths of resonances a r i s i n g from the l i e residue(Table III) are f a i r l y uniform, and do not show much solvent dependence. Those numbers probably r e f l e c t a significant contribution from the f i e l d inhomogeneity i n this heterogeneous sample since the normally s h a r p ( l e s s than 1 Hz) solvent l i n e i s 20 Hz wide.



27.

LIVE

Chemical Applications of Cross-linked Polymers

ANDKENT

513

The aromatic region provides the only PS related l i n e that i s readily measurable, although i t can only provide an approximation for the reasons mentioned above. The width of this combination i s somewhat greater than found i n the resin above, which i s con sistent with the proton results. The values of T- and η of the l i e carbons point clearly to an increase of mobility over that for chain segments i n the crosslinked samples. In general a linear chain attached to a polymer shows greater m o b i l i t y ( 2 2 ) from the additional degrees of rota tional freedom about i t s bonds * I t i s of great interest from the chemical point of view that these T values are comparable with those for small free peptides i n s o l u t i o n ( 2 3 ) . The use of the approximation of isotropic motion improves as the mobility becomes greater, and using t h i s , a correlation time on the order of 10 s i s estimated for the l i e moiety. Even allowing for unavoidable bias, the results for the aromatic protons are sugges tive of an increase of motional freedom for the resin. The s i m i l a r i t y of results for peptide resins i n both solvents f i t s nicely with the estimate that this resin sample with about 50 weight % peptide should have similar swelling characteristics i n the two solvents C3) . In these samples we have measured the integrals of the aromatic region r e l a t i v e to an unenriched peptide peak, and the results are consistent with the observation of a l l the carbon n u c l e i , and hence the results are t y p i c a l of a l l por tions of the cross-linked matrix. 1 0

Conclusions The aim of this work has been to develop an understanding of the environment at a molecular l e v e l within the cross-linked PS resin matrix and to determine if there were any fundamental phys i c a l limitations to the use of such supports i n chemical syn thesis. The physical evidence has shown that to a good approxima tion the properties of chain segments i n swollen copoly(styrenel%divinylbenzene) closely r e f l e c t those of linear PS molecules i n solution under conditions t y p i c a l of those used i n solid-phase synthesis. The dynamics of both chain segments and free molecules are characterized by rapid fluctuations of substantial magnitude at frequencies of 10 /s or greater arising from segmental motions i n the chain. The high degree of mobility i n the cross-linked material indicates that for the samples examined PS chain segments are w e l l solvated. From the i n t e n s i t i e s of the NMR signals, our data show that these characteristics are present uniformly throughout the matrix. This i s the contrast with an analysis of results reported by others(14) on a cross-linked, 25% chloromethylated resin. From the chemical point of view, the rapid fluctuations mean that diffusion of solvents and reagents should be rapid, a l l sites on the resin readily accessible,and pendant chains easily accom modated throughout the matrix. The uniformity of physical 8



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properties should rule out any dispersion i n reactivity from site to site on the resin. The pendant chains appear to have even greater rotational freedom than the PS chain segments, approaching that typical of such a free molecule i n solution. From the data presented here there i s no evidence for a fundamental resin-caused physical limitation on the application of the solid phase method. These results indicate that older notions of pore-like cavities in a semi-rigid matrix were incorrect(24); on the contrary the interior of the resin bead can be visualized as a solution of intermingled polymer and pendant chains. An additional point i l l u s t r a t e d i n this paper i s the useful ness of high resolution NMR techniques i n examining elastomer gels. This provides a relatively simple approach to dynamic and conformational information at a basic molecular level. The interpretation of the data i n terms of a complete picture of molecular motion may be complicated by the nature of the distribu tion of motional modes, but by sampling at a variety of fre quencies of different nuclei and at different magnetic fields i t should be possible to develop a more accurate picture than we have been able to present here. Acknowledgements

The NT-300 NMR spectrometer at Rockefeller University was purchased i n part from funds from the National ScienceFoundation(PC The WM-500 spectrometer i s part of the Southern California Re gional NMR F a c i l i t y at Caltech and i s supported by National Science Foundation Grant CHE-7916324.

Literature Cited 1. Merrifield, R. B. Fed. Am. Soc. Exp. B i o l . 1962, 21, 412. 2. Merrifield, R. B. J. Am. Chem. Soc. 1963, 85, 2149. 3. Sarin, V. K.; Kent, S. Β. H.; Merrifield, R. B. J. Am. Chem. Soc. 1980, 102, 5463. 4. Barany, G.; Merrifield, R. B. "The Peptides"; Gross, E.; Meienhofer, J., Eds.; Academic Press: New York, 1980; Vol I I , Chap. 1. 5. Sheppard, R. C. "Peptides 1971"; Nesvadba, H., Ed.; NorthHolland: Amsterdam, 1973; p. 111. 6. Stahl, G. L.; Walter, R.; Smith, C. W. J. Am. Chem. Soc. 1979, 101, 5383. 7. Bovey, F. A. "High Resolution NMR of Macromolecules"; Academic Press: New York, 1972. 8. Schaefer, J. "Topics i n Carbon-13 NMR Spectroscopy"; Levy, G. C., Ed., J. Wiley: New York, 1974; Vol. I, Chap. 4. 9. Heatley, F. "Progress i n NMR Spectroscopy"; Emsley, J. W.; Feeney, J.; S u t c l i f f e , L. Η., Eds., Pergamon Press: Oxford, 1979, Vol. 13, p. 47.


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Liu, K.-J.; Burlant, W. J. Polymer Sci, Pt. A-1 1967, 5, 1407. Doskocilova, D.; Schneider, B.; Jakes, J. J. Mag. Res. 1978, 29, 79. Manatt, S. L.; Horowitz, D.; Horowitz, R.; Pinnell, R. P. Anal. Chem. 1980, 52, 1532. Doskocilova, D.; Schneider, B.; Jakes, J. Polymer 1980, 21, 1185. Ford, W. T.; Balakrishnan, T. Macromolecules 1981, 14, 284. Ford, W. T.; Yacoub, S. Α.; J. Org. Chem. 1981, 46, 819. Flory, P. J. "Principles of Polymer Chemistry"; Cornell University Press: Ithaca, N.Y. 1953. Regen, S. L. J. Am. Chem. Soc. 1975,97,3108. "NTCFT-1180 Manual"; Nicolet Magnetics, Mountain View, C a l i f . 1980. Bovey, F. Α.; Hood III, F. P.; Anderson, E. W.; Snyder, L. C. J. Chem. Phys. 1965, 42, 3900. Inoue, Y.; Nishioka, Α.; Chujo, R. Die Macromolecular Chemie 1972, 156, 207. Schaefer, J.; Natusch, D. F. S. Macromolecules 1973, 5, 416. Levy, G. C.; Rinaldi, P. L.; Dechter, J. J.; Axelson, P. E.; Mandelkern, L. "Polymer Characterization by ESR and NMR", Woodward, A. E,; Bovey, F. Α., Eds. ACS Symposium Series No. 142, American Chemical Society: Washington, D.C., 1980, p. 119. Deslauries, R. Smith, I. C. P. "Topics i n Carbon-13 NMR Spectroscopy"; Levy, G. C., Ed., J. Wiley and Sons; New York, 1976; Vol. I I , Chap. 2. Heitz, W. Adv. Polymer Sci. 1977, 23, 1.

RECEIVED February 15,

1982.


Fundamental Aspects of the Chemical Applications of Cross-linked

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