The methods for sequence analysis of synthetic polymers differ from the sequence analysis of biopolymers (e. g.
DNA or
protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respo ...
s).
Synthetic polymers
Some familiar household synthetic polymers include: Nylons in textiles and fabrics, Teflon in non-stick pans, Bakelite for electrical switches, polyvinyl chloride (PVC) in pipes, etc. The common PET bottles are made of a synthetic polymer, polye ...
are produced by
chain-growth or
step-growth polymerization
Step-growth polymerization refers to a type of polymerization mechanism in which bi-functional or multifunctional monomers react to form first dimers, then trimers, longer oligomers and eventually long chain polymers. Many naturally occurring a ...
and show thereby
polydispersity
In chemistry, the dispersity is a measure of the heterogeneity of sizes of molecules or particles in a mixture. A collection of objects is called uniform if the objects have the same size, shape, or mass. A sample of objects that have an incons ...
, 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).
NMR-based sequencing
Nuclear magnetic resonance (NMR) spectroscopy
Nuclear magnetic resonance spectroscopy, most commonly known as NMR spectroscopy or magnetic resonance spectroscopy (MRS), is a spectroscopic technique to observe local magnetic fields around atomic nuclei. The sample is placed in a magnetic fie ...
is known as the most widely applied and “one of the most powerful techniques” for the sequence analysis of synthetic copolymers.
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.
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.
Example: Copolymer of PET and PEN
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
Polyethylene terephthalate (or poly(ethylene terephthalate), PET, PETE, or the obsolete PETP or PET-P), is the most common thermoplastic polymer resin of the polyester family and is used in fibres for clothing, containers for liquids and foods ...
(PET) and
polyethylene naphthalate
Polyethylene naphthalate (poly(ethylene 2,6-naphthalate) or PEN) is a polyester derived from naphthalene-2,6-dicarboxylic acid and ethylene glycol. As such it is related to poly(ethylene terephthalate), but with superior barrier properties.
Pro ...
(PEN) in their blends.
During such a
transesterification
In organic chemistry, transesterification is the process of exchanging the organic group R″ of an ester with the organic group R' of an alcohol. These reactions are often catalyzed by the addition of an acid or base catalyst. The reaction can ...
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.
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. The sequence patterns can become even more complex, when triads can be distinguished spectroscopically.
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 (A
1B
1)
n and (A
2B
2)
n (with molecular weight high enough to neglected chain-ends), the following two equations are valid:
i">Ai=
i">i wherein (i = 1,2) (1)
1B2 ">A1B2 =
2B1">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: