2D chromatography

The separation of interactions by 2D spectroscopy can be compared with 2D chromatography. In a onedimensional thin layer or paper chromatogram, the separation of the constituents by elution with a given solvent is often incomplete. Elution with a second solvent in a perpendicular direction may then achieve full separation. In NMR spectroscopy, the choice of two solvents is replaced by the choice of two suitable (effective) Hamiltonians for the evolution and detection periods which allow unique characterisation of each line. 

Figure 12 Schematic representation of 2D chromatography using an eight-port injection valve and two storage loops. (A) In the first position of the valve the storage loop 1 is loaded with the HPLC eluent, while the content of the storage loop 2 is analyzed by SEC. (B) In the second valve position storage loop 2 is loaded with HPLC eluent and the content of loop 1 is analyzed according the molecular size of the solute.

D chromatography with silica rod columns for the separation of polar aromatic compounds was also employed by Ikegami et al. [197]. 

Such a system has been used for the comprehensive 2D chromatography of proteins [9,14], synthetic polymers [16], oxygen heterocyclic fraction of cold-pressed citrus oils [22,29], carotenoids [39], triglycerides in fats and oils [18-21], pharmaceuticals [29], and acidic and phenolic compounds [27,28]. 

Fig. 15. Schematic representation of 2D chromatography using an eight-port injection valve and two storage loops...

The mapping of ethoxylated fatty alcohols and ethylene oxide-propylene oxide block copolymers by 2D chromatography was discussed by Trathnigg et al. [95]. They combined LAC and SEC and were able to determine the chemical composition distribution and the molar mass distribution of the polyethers. 

The versatility of the 2D approach is illustrated with a four-arm starshaped block copolymer (a mixture of 16 components), which was synthesized in our laboratories to understand and demonstrate the advantages of 2D chromatography. These 16 components are a mixture of four different styrene-butadiene (St-Bd) copolymer compositions, each consisting of four molar masses (the St-Bd precursor with one to four arms). Another polymer investigated is a real-world application the deformulation of a telomeric aliphatic polyester. 

D chromatography Two chromatographs (special software) Methods for each dimension This work This work 11, 12, 14, this work... 

We tried to combine the advantages of HPLC and SEC by using a fully automated, software controlled, 2D chromatography system for the on-line analysis of composition, end-group functionality, and MMD. It consists of two chromatographs, one that separates by chemical composition (e.g., a gradient HPLC) and a SEC instrument for subsequent separation by size (cf. Figure 2). 

Figure 2. Experimental setup of the 2D chromatography system (horizontal, LC components vertical, SEC components).

The preparations were carried out in a way to give all structures that are theoretically possible St-Bd precursor (of molar mass Marm) linear (St-Bd)2 (M = 2Marm), the three-arm product (St-Bd)3 (M == 3Marm), and, finally, the four-arm compound (St-Bd)4 (M == 4M ). Four samples with varying Bd content (20, 40, 60, and 80%) were prepared in this way. A mixture of these samples was used to check the capabilities of the 2D chromatography system. 

The deformulation of a polyester was done with liquid-adsorption chromatography under critical conditions (LACCC) that was coupled to an SEC system. The dimension was separated by the end-group functionality to study byproducts formed during polycondensation. By 2D chromatography a number of species (e.g., cycles, ethers, alkyl terminated chains, and so on) could be identified. Some of them influenced the mechanical and thermal properties of the polyesters significantly. 

The high resolution of LC-SEC separations and the full automation using 2D-CHROM software enable the reliable and comprehensive characterization and deformulation of complex analytes like copolymers, polymer blends, and additives. 2D chromatography will become a powerful tool with flexible and easy-to-use software. Basically, all types of LC methods can be combined to give superior resolution and reproducibility. 

Identification and quantitation can be done with model compounds or with specific detection, like Fourier transform infrared or other kinds of multiple detection. However, 2D chromatography is also extremely useful for fingerprinting and visual comparison. 

Complex polymer topologies, polymer blends, and multicomponent formulations require a different approach to perform a proper molecular characterization. In two-dimensional (2D) chromatography, different separation techniques are used to avoid co-elution of species and to measure molar mass and chemical composition in a truly independent way [5]. 

In 2D chromatography separations, an aliquot from a first column (method) is transferred into the next separation method in a sequential and repetitive manner using automated sample transfer valves which are equipped with one or more sample loops. Alternatively, as a simpler and not quite as useful transfer technique, heart cuts from peaks in the first separation... 

Fig. 2 Various data presentation views in 2D chromatography showing the 2D analysis of a 16-component star block copolymer (see text for details).

Further data analysis allows the determination of molar mass, chemical composition, and relative concentration for each component. The 2D chromatography data can be displayed in various forms (cf. Fig. 2) ... 

Additional certainty in using fj values to assign peak identity is obtained when retention times match on two columns with significantly different stationary phases, or a single planar stationary phase eluted by orthogonal, sequential application of two mobile phases. This is the principle and application of 2D chromatography discussed earlier in Section... 

Comprehensive two-dimensional (2D) chromatography is a very powerful tool for the separation of complex samples because it offers a significant increase of peak capacity as a result of the expansion of the available separation space. Co-eluting peaks in one-dimensional (ID) chromatography might be separated in the second dimension, meaning that more peaks can be individually detected in comprehensively interfaced two-dimensional chromatography. 

Off-line SEC-NMR is certainly more labour-intensive and time-consuming than on-line SEC-NMR. Nevertheless, this difference can be minimized by reducing the number of fractions. Clearly, the reduction cannot go beyond a certain limit, otherwise it will cause a loss of accuracy in the measurement of copolymer properties. The hypothesis that it is possible to reduce the time for copolymer analysis without the cited loss of accuracy was put forward by Murphy et al. (24) in another context (2D-chromatography). It would be interesting to check whether the hypothesis is valid also in SEC-NMR and to develop a methodology which allows the determination of the optimal conditions, viz., those where the cited loss of accuracy is still negligible. 

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