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Online Size Exclusion Chromatography-NMR for the Determination of Molar Mass Distributions of Copolymers Wolf Hiller,*,† Mathias Hehn,† Thorsten Hofe,‡ and Kirsten Oleschko‡ TU Dortmund, Faculty of Chemistry, Otto-Hahn-Strasse 6, 44227 Dortmund, Germany, and PSS Polymer Standards Service GmbH, In der Dalheimer Wiese 5, 55120 Mainz, Germany A general approach of size exclusion chromatography (SEC)-NMR is introduced for the determination of the classical molar mass parameters MW, MN, and MP. It can be used for the determination of molar mass distributions of homopolymers and copolymers. The main advantage of SEC-NMR of copolymers is the possibility of detecting each monomer unit simultaneously with NMR as a quantitative concentration detector. Therefore, it is possible to provide the chemical compositions of copolymers at any elution volume without calibrations. In this respect, a new method will be presented for getting correct signal quantities of onflow data with sufficient NMR sensitivities. As the consequence, the chemical composition of copolymers can be correctly quantified under typical chromatographic conditions with respect to sample concentration and flow rate. Finally, the molar mass calibrations of the copolymers can be easily adjusted according to their chemical compositions. The methods were applied to polystyrene-b-poly(methyl methacrylate) block copolymers of different molar masses. The results of the molar mass distributions and the chemical composition distributions obtained by SEC-NMR are in very good agreement with the complex SEC multidetector analysis. The online coupling of liquid chromatography and NMR spectroscopy is known since the end of the 1970s.1 Meanwhile, various applications in pharmaceutical, food, and fuel chemistry have been reported. Korhammer and Bernreuther2 have given a short and Albert a general overview of LC-NMR developments and different applications.3 Only a few applications are related to the hyphenation of liquid adsorption chromatography (LAC) to NMR of polymers.4-6 Other polymer applications are focused on the online coupling of liquid * To whom correspondence should be addressed. Fax: +49 231 755 3771. E-mail:
[email protected]. † TU Dortmund. ‡ PSS Polymer Standards Service GmbH. (1) Watanabe, N.; Niki, E. Proc. Jpn. Acad., Ser. B 1978, 54, 194–199. (2) Korhammer, S. A.; Bernreuther, A. Fresenius J. Anal. Chem. 1996, 354, 131–135. (3) Albert, K. On-line LC-NMR and Related Techniques; John Wiley & Sons Ltd.: Chichester, U.K., 2002. (4) Pasch, H.; Hiller, W. Macromolecules 1996, 29, 6556–6559. (5) Pasch, H.; Hiller, W.; Haner, R. Polymer 1998, 39, 1515–1523.
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chromatography at critical conditions (LCCC) and NMR, e.g., the separation of poly(ethyl methacrylate) regarding its tacticity7 or the analysis of the end groups of poly(ethylene oxides).8 Recent applications of LCCC-NMR allowed the determination of the true molar mass and the true chemical composition of block copolymers.9-11 This method is capable to separate the homopolymer precursor from the block copolymer. Further important polymer applications are related to the coupling of size exclusion chromatography (SEC) with NMR. SEC-NMR has been used to determine MN of poly(methyl methacrylate) (PMMA) homopolymers via end groups12,13 or the dependence of tacticity on molecular weight.13,14 It has been shown that SEC-NMR can deliver the chemical composition distribution (CCD) of block copolymers15 or give information about random copolymers regarding CCD15 and chemical heterogeneity.16 Blends of isotactic PMMA and syndiotactic PMMA could be investigated.17 Further developments allow even the performance of SEC-NMR at high temperatures.18 However, there is only one application of SEC-NMR for the determination of the molar mass parameters MW and MN. Hatada et al. have determined MW and MN for low molar mass PMMA via the end group.19 Most of the SEC-NMR studies analyze only MN via end groups or compare these data with refractive (6) Kra¨mer, I.; Hiller, W.; Pasch, H. Macromol. Chem. Phys. 2000, 201, 1662– 1666. (7) Kitayama, T.; Janco, M.; Ute, K.; Niimi, R.; Hatada, K.; Berek, D. Anal. Chem. 2000, 72, 1518–1522. (8) Hiller, W.; Bru ¨ll, A.; Argyropoulos, D.; Hoffmann, E.; Pasch, H. Magn. Reson. Chem. 2005, 43, 729. (9) Hiller, W.; Sinha, P.; Pasch, H. Macromol. Chem. Phys. 2007, 208, 1965– 1978. (10) Hiller, W.; Sinha, P.; Pasch, H. Macromol. Chem. Phys. 2009, 210, 605– 613. (11) Hiller, W.; Pasch, H.; Sinha, P.; Wagner, T.; Thiel, J.; Wagner, M.; Mu ¨ llen, K. Macromolecules 2010, 43, 4853–4863. (12) Hatada, K.; Ute, K.; Okamoto, Y.; Imanari, M.; Fujii, N. Polym. Bull. 1988, 20, 317–321. (13) Ute, K.; Hatada, K. Anal. Sci. 1991, 7, 1629–1632. (14) Hatada, K.; Ute, K.; Kitayama, T.; Nishimura, T.; Kashiyama, M.; Fujimoto, N. Polym. Bull. 1990, 23, 549–554. (15) Hatada, K.; Ute, K.; Kitayama, T.; Yamamoto, M.; Nishimura, T.; Kashiyama, M. Polym. Bull. 1989, 21, 489–495. (16) Kra¨mer, I.; Pasch, H.; Ha¨ndel, H.; Albert, K. Macromol. Chem. Phys. 1999, 200, 1734–1744. (17) Ute, K.; Niimi, R.; Matsunaga, M.; Hatada, K.; Kitayama, T. Macromol. Chem. Phys. 2001, 202, 3081–3086. (18) Hiller, W.; Pasch, H.; Macko, T.; Hofmann, M.; Ganz, J.; Spraul, M.; Braumann, U.; Streck, R.; Mason, J.; van Damme, F. J. Magn. Reson. 2006, 183, 290–302. (19) Ute, K.; Niimi, R.; Hongo, S.-Y.; Hatada, K. Polym. J. 1998, 30, 439–443. 10.1021/ac1013095 2010 American Chemical Society Published on Web 09/01/2010
Table 1. Molar Masses of the Block Copolymers As Given by the Supplier; Chemical Composition (S/MMA) and Tacticity Determined by 1H-NMR and SEC-NMR 1
H NMR
SEC (RI-UV)
SEC-NMR
sample
MW [kg mol-1]
S/MMA [mol %]
mm/mr/rr [mol %]
S/MMA (corrected) [mol %]
S/MMA (original)) [mol %]
S/MMA (corrected) [mol %]
mm/mr/rra [mol %]
1 2 3 4 5
20.5 65 108 158 610
47.0/53.0 45.1/54.9 48.1/51.9 47.4/52.6 64.8/35.2
2.4/23.0/74.6 1.7/21.1/77.2 3.0/26.1/70.9 1.4/20.7/77.9 2.3/24.3/73.4
49.2/50.8 47.5/52.5 51.9/48.1 51.0/49.0 70.4/29.6
38.8/61.2 35.3/64.7 38.5/61.5 36.3/63.7 55.7/44.3
50.2/49.8 46.3/53.7 49.8/50.2 47.6/52.4 66.6/33.4
2.4/22.0/75.6 1.7/22.0/76.3 3.0/27.6/69.4 1.4/20.9/77.7 2.3/21.8/75.9
a The content of mm was taken from 1H NMR due to the low intensity of the mm triad and the constant microstructure distribution of the mr and rr triads as shown in Figure S-11 in the Supporting Information.
index data.20 There is no comprehensive analysis of the molar mass distributions (MMD) using SEC-NMR. It is the aim of this publication to demonstrate the strength of SEC-NMR for the determination of MW, MN, and MP. A general approach for obtaining these parameters will be shown with respect to common chromatographic conditions. In particular, the benefits of the NMR as a concentration detector will be demonstrated for the analysis of copolymers. These results will be compared with conventional SEC using the multidetector method. PS-b-PMMA block copolymers are characterized regarding their distributions of molar masses, chemical compositions, and microstructures. In particular, the distributions of molar masses and chemical compositions will be combined for a comprehensive analysis. EXPERIMENTAL SECTION Samples. Polystyrene (PS) and PMMA calibration standards as well as the PS-b-PMMA copolymers were produced by PSS GmbH (Mainz, Germany). The copolymers were mainly anionically polymerized. Sample 3 of Table 1 was produced via controlled radical polymerization. The copolymers are summarized in Table 1. The molar masses of the calibration standards for SEC-NMR are shown in Table S-1 in the Supporting Information. In addition, sample 6 (PS-b-PMMA, total MW ) 50 kg mol-1; PS, MW ) 22 kg mol-1; PMMA, MW ) 28 kg mol-1) (42.5/57.5 mol % PS/ PMMA) was used for several preparation experiments. SEC Copolymer Analysis (Multidetector Analysis). A PSS “Agilent 1200 SECcurity system” (PSS GmbH, Mainz, Germany) and the PSS-WINGPC-Unity for data acquisition and processing were used. It consisted of a UV (λ ) 254 nm) and RI detector and four columns (PSS SDV 5 µm precolumn, 103, 105, and 106 Å with 300 mm × 8 mm inner diameter). The sample concentration was 1.0 mg mL-1, and the injection volume was 20 µL. The mobile phase was THF, and the flow rate 1.0 mL min-1. PS and PMMA standards were used in the range of 0.484 up to 2530 kg mol-1 (ReadyCal Kit, PSS GmbH, Mainz, Germany). The comonomer content of each copolymer was calculated based on the response factors for each comonomer and each detector (see Table S-2 in the Supporting Information). SEC-NMR. The SEC-1H/NMR experiments were performed on a 500 MHz spectrometer DRX (Bruker BioSpin GmbH, Germany) coupled to an Agilent 1100 series HPLC (20) Hatada, K.; Ute, K.; Kashiyama, M.; Imanari, M. Polym. J. 1990, 22, 218– 222.
system (Agilent Technologies GmbH, Böblingen, Germany). Three columns (PSS SDV 5 µm precolumn, 103 and 105 Å with 300 mm × 8 mm inner diameter) were used. The onflow measurements were performed with a flow probe containing a 60 µL flow cell (90° 1H pulse was 7 µs, 16 scans per increment, recycle delay of 0.69 and 0.32 s acquisition time, 3 kb data, 4.7 kHz spectral width). The data were processed as 2D contour plots (zero filling to 16 kb with a line broadening of lb ) 5 Hz). The accuracy of the NMR retention time was ±2 s. The sample concentration for the SEC-NMR measurements was 2.7 mg mL-1. A volume of 100 µL was injected. HPLC grade THF was used as mobile phase with a flow rate of 0.8 mL min-1. WET solvent suppression was applied to THF.21 RESULTS AND DISCUSSION Preparation of SEC-NMR. First, the SEC-NMR system has to be evaluated regarding the highest concentration of injection. A polystyrene standard was measured with SEC and UV (see Figure S-1 in the Supporting Information). It was found that 3 mg mL-1 and a 100 µL injection was the highest possible concentration of injection. Higher concentrations deliver smaller molar masses. This behavior is already described by Yau et al.22 Figure 1 shows an SEC-NMR onflow experiment of sample 1 with the elution of the aromatics of PS and the R-CH3 group of PMMA. The methoxy group of the PMMA block is overlapping with the low field signal of THF and is suppressed. However, the R-CH3 group of PMMA reveals the signals of isotactic (mm), atactic (mr), and syndiotactic (rr) triads. The NMR chromatogram of PS is determined as the sum of the ortho, meta, and para protons and the NMR chromatogram of PMMA as the sum of the mr and rr triads of the R-CH3 group. The mm triad is neglected for further quantifications due to the low signal-to-noise. The simultaneous detection of PS and PMMA allows the calculation of the CCD of the copolymer vs retention time. Quantification of SEC-NMR Data. To evaluate the chemical compositions of the onflow data, a closed system consisting of pump, flow probe, and solvent bottle was set up. The bottle contained 40 mg of PS-b-PMMA (sample 6) dissolved in 200 mL of THF. This setup allowed continuous flow of a homogeneous polymer solution through the NMR probe. It enables the mea(21) Smallcombe, S. H.; Patt, S. L.; Keifer, P. A. J. Magn. Res. A 1995, 117, 295–303. (22) Yau, W. W.; Kirkland, J. J.; Bly, D. D. Modern Size-Exclusion Liquid Chromatography; John Wiley & Sons: New York, 1979.
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Figure 1. SEC-NMR onflow experiment of sample 1: solid line ) vertical projection of PMMA, dashed line ) vertical projection of PS, top spectrum ) most intense trace.
surement of T1 as well as the intensities of PS and PMMA in dependence of the recycle delay, pulse angle, and flow rates. All experiments required solvent suppression. T1 was measured with a WET modified inversion recovery experiment (see Figure S-2 in the Supporting Information). It is decreasing with increasing flow rates (Figure S-3 in the Supporting Information) and obeys eq S-1 in the Supporting Information.3 The τ values of the aromatic protons of PS are closer to the theoretical ones than for the syndiotactic species of PMMA (Figure S-4 in the Supporting Information). To prove the quantification of the NMR onflow data, normalized NMR intensities of PS and PMMA were measured at different flow rates, recycle delays, and pulse angles. It is important to use short recycle delays for optimal chromatographic resolution without losing much sensitivity. According to Figure 2, the use of 90° pulses provides good NMR sensitivities even for a recycle delay of 0.69 s where the highest NMR sensitivity is obtained at the flow rate of 1.2 mL min-1. The complete flow diagram for different recycle delays is shown in Figure S-5 in the Supporting Information. The curves present maxima which are shifted toward higher flow rates with decreasing recycle delays. According to Figure S-5 in the Supporting Information, the intensities of the aromatic protons decrease with decreasing recycle delays. The intensities of PMMA show only small effects due to the much shorter relaxation times. For example, the meta-para protons reduce their intensity from 90% at a recycle delay of 2.48 s to 48% 8246
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at 0.69 s and the rr triad of PMMA from 100% at 2.48 s to 88% at 0.69 s (measured at a flow rate of 0.8 mL/min). The raw data give correct quantification of the CCD only for short pulse angles (
