Sequence analysis of synthetic polymers

The methods for sequence analysis of synthetic polymers differ from the sequence analysis of biopolymers (e. g. DNA or proteins). Synthetic polymers are produced by chain-growth or step-growth polymerization and show thereby polydispersity, whereas biopolymers are synthesized by complex template-based mechanisms and are sequence-defined and monodisperse. Synthetic polymers are a mixture of macromolecules of different length and sequence and are analysed via statistical measures (e. g. the degree of polymerization, comonomer composition or dyad and triad fractions).[1]

## NMR-based sequencing

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Nuclear magnetic resonance (NMR) spectroscopy is known as the most widely applied and “one of the most powerful techniques” for the sequence analysis of synthetic copolymers.[1][2]⁠ NMR spectroscopy allows determination of the relative abundance of comonomer sequences at the level of dyads and in cases of small repeat units even triads or more. It also allows the detection and quantification of chain defects and chain end groups, cyclic oligomers and by-products.[2]⁠ However, limitations of NMR spectroscopy are that it cannot, so far, provide information about the sequence distribution along the chain, like gradients, clusters or a long-range order.[1]

### Example: Copolymer of PET and PEN

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Monitoring the relative abundance of comonomer sequences is a common technique and is used, for example, to observe the progress of transesterification reactions between polyethylene terephthalate (PET) and polyethylene naphthalate (PEN) in their blends.

During such a transesterification reaction, three resonances representing four diads can be distinguished via 1H NMR spectroscopy by different chemical shifts of the oxyethylene units: The diads -terephthalate-oxyethylene-terephthalate- (TET) and -naphthalate-oxyethylene-naphthalate- (NEN), which are also present in the homopolymers polyethylene naphthalate und polyethylene terephthalate, as well as the (indistinguishable) diads -terephthalate-oxyethylene-naphthalate- (TEN) and -naphthalate-oxyethylene-terephthalate- (NET), which are exclusively present in the copolymer. In the spectrum of a 1:1 physical PET/PEN mixture, only the resonances corresponding to the diads TET and NEN are present at 4.90 and 5.00 ppm, respectively. Once a transesterification reaction occurs, a new resonance at 4.95 ppm emerges that increases in intensity with the reaction time, corresponding to the TEN / NET sequences.[2]

The example of polyethylene naphthalate and polyethylene terephthalate is relatively simple, as only the aromatic part of the polymers differ (naphthalate vs. terephthalate). In a blend of polyethylene naphthalate and polytrimethylene terephthalate, already six resonances can be distinguished, since both, oxyethylene and oxypropylene, form three resonances.[3] The sequence patterns can become even more complex, when triads can be distinguished spectroscopically.[2]⁠ The extractable information is limited by the difference in chemical shift and the resonance width. In addition to 1H NMR spectroscopy, also 13C NMR spectroscopy is a common method for the sequencing shown above, which is characterized in particular by a very narrow resonance width.

Deconvolution and assignment of these triad-based resonances allows a quantitative determination of the degree of randomness and the average block length via integration of the distinguishable resonances. In a 1:1 mixture of two linear two-component 1:1 polycondensates (A1B1)n and (A2B2)n (with molecular weight high enough to neglected chain-ends), the following two equations are valid:

[ Ai] = [Bi], wherein (i = 1,2) (1)

[ A1B2 ] = [ A2B1] (2)

Equation 1 states that the molar ratio of all four repeat units is identical and equation 2 states that both types of copolymer are of identical concentration. In this case, the degree of randomness χ is calculated as given by equation 3:

χ = [ A i B j A 1 A 2 ] {\displaystyle \chi =[{\frac {A_{i}B_{j}}{A_{1}A_{2}}}]} , wherein (i, j = 1, 2) (3)

In the beginning of a transreaction process (e. g. transesterification or transamidation), the degree of randomness χ ≈ 0 as the system comprises a physical mixture of homopolymers or block copolymers. During the transreaction process χ increases up to χ = 1 for a fully random copolymer. If χ > 1 it indicates a tendency of the monomers to form alternating structure, up to χ = 2 for a completely alternating copolymer.[4]⁠ The degree of randomness χ gives thereby statistical information about the polymer sequence. The calculation can be modified for three-component[5]⁠ and four-component[6]⁠ polycondensates.

### Application

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NMR spectroscopy is used in industrially relevant systems to study the sequence distribution of copolymers or the occurrence of transesterification in polyester blends. A change in sequence distribution can effect the crystallinity, and transesterification can affect the compatibility of two otherwise incompatible polyesters. Depending on their degree of randomness, copolyesters can show different thermal transitions and behaviours.[7]

## Other sequencing

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Other options besides traditional NMR spectroscopy for sequence analysis are listed here;[8] these include Kerr-effect for characterization of polymer microstructures, MALDI-TOF mass spectrometry, depolymerization (controlled chemical degradation of macromolecules) via chain-end depolymerization (i.e., unzipping) and nanopore analysis (most of such reported studies, however, have focused on poly(ethylene glycol), PEG).

## References

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This article incorporates text by Marcus Knappert available under the CC BY-SA 3.0 license.

1. ^ a b c Mutlu, Hatice; Lutz, Jean-François (2014). "Reading Polymers: Sequencing of Natural and Synthetic Macromolecules". Angewandte Chemie International Edition. 53 (48): 13010–13019\. doi:10.1002/anie.201406766. ISSN 1521-3773. PMID 25283068.
2. ^ a b c d Ilarduya, Antxon Martínez de; Muñoz‐Guerra, Sebastián (2014). "Chemical Structure and Microstructure of Poly(alkylene terephthalate)s, their Copolyesters, and their Blends as Studied by NMR". Macromolecular Chemistry and Physics. 215 (22): 2138–2160\. doi:10.1002/macp.201400239. hdl:2117/24793. ISSN 1521-3935.
3. ^ Huang, Doan-Ho; Woo, E. M.; Lee, Li-Ting (May 2006). "A comparative study on transreactions induced phase changes in blends of poly(trimethylene terephthalate) and poly(ethylene naphthalate) upon annealing". Colloid and Polymer Science. 284 (8): 843–852\. doi:10.1007/s00396-005-1443-x. ISSN 0303-402X. S2CID 93649623.
4. ^ Eersels, K. L. L.; Aerdts, A. M.; Groeninckx, G. (1999-04-09), Fakirov, Stoyko (ed.), "Reactive Melt Processing of Aliphatic/Aromatic Polyamide Blends: Effect on Molecular Structure, Semicrystalline Morphology, and Thermal Properties", Transreactions in Condensation Polymers, Weinheim, Germany: Wiley-VCH Verlag GmbH, pp. 267–317, doi:10.1002/9783527613847.ch7, ISBN 978-3-527-61384-7, retrieved 2021-01-28
5. ^ Yamadera, Reizo; Murano, Masao (September 1967). "The determination of randomness in copolyesters by high resolution nuclear magnetic resonance". Journal of Polymer Science Part A-1: Polymer Chemistry. 5 (9): 2259–2268\. doi:10.1002/pol.1967.150050905.
6. ^ Devaux, J.; Godard, P.; Mercier, J. P.; Touillaux, R.; Dereppe, J. M. (October 1982). "Bisphenol-A polycarbonate–poly(butylene terephthalate) transesterification. II. Structure analysis of the reaction products by IR and 1H and 13C NMR". Journal of Polymer Science: Polymer Physics Edition. 20 (10): 1881–1894\. doi:10.1002/pol.1982.180201011.
7. ^ Li, Wen-Da; Zeng, Jian-Bing; Lou, Xiao-Jie; Zhang, Jing-Jing; Wang, Yu-Zhong (2012). "Aromatic-aliphatic random and block copolyesters: synthesis, sequence distribution and thermal properties". Polymer Chemistry. 3 (5): 1344. doi:10.1039/c2py20068f. ISSN 1759-9954.
8. ^ Mutlu, Hatice; Lutz, Jean-François (2014-11-24). "Reading Polymers: Sequencing of Natural and Synthetic Macromolecules". Angewandte Chemie International Edition. 53 (48): 13010–13019\. doi:10.1002/anie.201406766. PMID 25283068.