In recent years, new developments in the area of Size Exclusion Chromatography (SEC) have resulted in the introduction of Ultra High-Performance Liquid Chromatography (UHPLC or UPLC®) for polymer applications. Waters ACQUITY Advanced Polymer Chromatography (APC™) is a breakthrough technology that is able to provide SEC analyses of polymeric materials with higher-resolution and higher-speed than conventional SEC.
The new APC technology involves the use of high efficiency separation columns and a low dispersion flow path, resulting in:
- High resolution analysis
- Reduced time
- Reduced solvent consumption
- Reduced sample concentration
- Quick mobile phase changeover
However, the APC system is limited to conventional calibration, meaning it can only generate relative molecular weight information. In order to obtain comprehensive knowledge of the polymers under analysis, it is necessary to combine the high-resolution APC system with additional advanced detectors.
Figure 1: Waters APC and Malvern Panalytical OMNISEC-REVEAL
The combination of the Waters APC system with the Malvern Panalytical OMNISEC REVEAL (Figure 1) greatly widens the information attainable from a SEC analysis. OMNISEC REVEAL is an advanced detector system combining refractive index (RI), ultraviolet PDA (UV), light scattering (LS) and differential viscometry (IV). Using OMNISEC REVEAL the sample under analysis can now be fully characterized in terms of:
- Absolute molecular weight: without the need for column calibration
- Molecular weight distribution
- Size: hydrodynamic radius across the samples entire molecular weight range
- Structure: Mark-Houwink parameters and branching
Combining the high-resolution separation obtained using an APC with a multi-detection system brings a number of challenges, most notable of which is band broadening. As a sample moves through the detector flow path it is stretched, essentially losing the resolution obtained on the column. OMNISEC REVEAL already benefits from a unique flow path with all the detectors connected in series and the RI detector, which typically introduces most band broadening, is the first detector in line. To fully realise the potential of the APC a new OMNISEC REVEAL has been developed with a significantly reduced internal volume designed specifically to maintain the APC resolution.
In this study, the analyses of several polymers by Waters APC combined with the Malvern Panalytical OMNISEC REVEAL multi-detector system are presented and discussed.
Methods and Materials
The APC and OMNISEC REVEAL were set up as follows:
Samples were separated using three Waters APC XT column, 900Å, 450Å and 200Å connected in series. The mobile phase was THF. Samples were allowed to dissolve overnight to ensure full dissolution. All detectors and columns were maintained at 35oC to ensure good separation and maximise baseline stability.
The samples analysed in this note are two NIST (National Institute of Standards and Technology)polystyrene (PS) standards, PS1478 and PS706a, a broad polyvinylchloride (PVC) and a broad polycarbonate (PC). These materials were dissolved in THF and injected onto the column using the APC system. After separation on the APC columns the sample flowed into the OMNISEC-REVEAL where the macromolecular properties were measured.
NIST Polystyrene Standards
A chromatogram for the NIST PS1478 polystyrene narrow molecular weight standard is shown in Figure 2. Also shown in this figure is the Log molecular weight (Mw), Log intrinsic viscosity (IV) and Log hydrodynamic radius (Rh). Figure 3 shows the chromatogram for the NIST PS706a polystyrene broad molecular weight standard. Also shown in this figure is the Log Mw, Log IV and Log Rh.
Both chromatograms show excellent detector response with low sample loadings.
Figure 2: RI, RALS and DP chromatograms obtained for PS1478 NIST standard.10µL injection volume, 2.5 mg/mL concentration. Log Mw, IV and Rh also shown
Figure 3: RI, RALS, LALS and DP chromatograms obtained for PS706a NIST standard. 5µL injection volume, 2 mg/mL concentration. Log Mw, IV and Rh also shown
Table 1 shows the quantitative results obtained from the OMNISEC-REVEAL. The absolute molecular weights are calculated directly from the right angle light scattering (RALS) and the low angle light scattering (LALS). In addition, the presence of the viscometer detector gives the possibility to calculate the size and structural information of the polymers. The values of molecular weight, dispersity (Mw/Mn), intrinsic viscosity (IV), hydrodynamic radius (Rh) and the Mark-Houwink a and log K parameters are all consistent with the values expected. The calculated values of PS706a and PS1478 match the NIST certified Mw values of 285±0.23K g/mol and 37.4±0.26K g/mol respectively, illustrating excellent precision and accuracy. These results primarily demonstrate how an OMNISEC-REVEAL linked to an APC can allow full characterization of polymers.
Table 1: Molecular weight, size and structure results for PS1478 and PS706a
Polycarbonate and polyvinylchloride
Samples of PC and PVC were dissolved in the mobile phase and injected into the APC-REVEAL. Figure 4 shows the chromatogram obtained for PC and Figure 5 that for PVC. As with the NIST samples both chromatograms show excellent detector response with low sample loadings.
Figure 4: Representative RI, RALS, LALS and DP chromatograms and molecular weight, IV and Rh distributions of PC. 10µL injection volume, 3 mg/mL concentration. Log Mw, IV and Rh also shown
Figure 5: Representative RI, RALS, LALS and DP chromatograms and molecular weight, IV and Rh distributions of PVC. 10µL injection volume, 3 mg/mL concentration. Log Mw, IV and Rh also shown
Table 2 shows the results obtained for PC and PVC by the OMNISEC REVEAL. The quantitative data obtained align with the expected results obtained previously using a coventional OMNISEC chromatography system.
Table 2: Molecular weight, size and structure results for PC and PVC samples
An overlay of the molecular weight distributions for duplicate injections of PC, PVC and PS706a is shown in Figure 6. The figure shows that the three samples PC, PVC and PS have very different molecular weight distribution plots, with the PS sample shifted toward the high Mw region and PC toward the low Mw region.
The results in Table 1 and 2 show the average values for the IV and Rh for the different polymers. These values can be used to make broad structural comparisons between samples, but to get a complete picture of the structure it is necessary to use a Mark-Houwink plot to look at the IV distribution across the entire Mw range.
Figure 6: Overlay of duplicate molecular weight distributions of PC, PVC and PS
The Mark-Houwink plot in Figure 7 shows the structural comparison between the three different samples, PC, PVC and PS706a. This plot shows the Log Mw plotted against the Log IV and allows structural differences between the samples to be clearly identified. PS is lowest on the plot indicating it has a lower IV than PC and PVC at comparable Mws. A lower IV is indicative of a higher molecular density and suggests a more compact configuration. PC has the highest IV value indicating it has the most open configuration.
PVC has a density falling between PC and PS. In addition to this the IV of PVC deviates from the a linear behaviour, with a decreasing IV values in the high Mw region. This indicates the presence of branching at higher Mws. It would have not been possible to obtain this insight into the polymer’s architecture without the use of a viscometer.
Figure 7: Mark-Houwink plots of PC, PVC, and PS overlaid and compared to show structural differences
The combination of the OMNISEC-REVEAL multi-detector system with the Waters ACQUITY® APC System provides high quality, high-resolution and rapid sample analysis. The presence of the highly sensitive LS and RI detectors and the viscometer in the OMNISEC REVEAL system allowed for the calculation of absolute molecular weight, hydrodynamic radius and structure of the materials. Two NIST polystyrene standards were correctly characterised using the system as well as PC and PVC samples. A structural comparison was performed using the Mark-Houwink plot with differences between samples observed and regions of branching within PVC identified.
A comparison of the polymer characteristics was performed which would not have been possible using only a conventional calibration system. Accurate molecular weights and structural information can only be attained using a multi-detection platform, such as OMNISEC-REVEAL, which combines not only RI and light scattering but also viscometry. The unique combination of Malvern Panalytical’s advanced detectors with APC separations offers users greater resolving power with full sample characterisation.