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Boletín de la Sociedad Chilena de Química

versión impresa ISSN 0366-1644

Bol. Soc. Chil. Quím. v.44 n.3 Concepción set. 1999

http://dx.doi.org/10.4067/S0366-16441999000300011 

SEPARATION OF COPOLYMERS OF VARIABLE COMPOSITIONS
BY CAPILLARY ZONE ELECTROPHORESIS

IVAN M. PERIC1*, BERNABE L. RIVAS2, AMALIA POOLEY2,
ELIZARDO RIFFFO1 AND LUIS A. BASAEZ1

1Departamento de Química Analítica e Inorgánica. Fax: 56 41 245974;
e-mail: iperic@udec.cl
2Departamento de Polímeros, Facultad de Ciencias Químicas, Universidad de Concepción,
Casilla 160-C, Concepción, Chile.
(Received: April 6, 1999 - Accepted: June 22, 1999)

ABSTRACT

Capillary zone electrophoresis (CZE) analysis of several copolymers obtained by radical copolymerization of acrylamide and N,N'-dimethylacrylamide using acrylic acid and 1-vinyl-2-pyrrolidone as comonomers under different feed molar ratios had been carried out. Results demonstrate that absolute value of the electrophoretic mobility is strongly dependent of the linear charge density of the copolymer in agrement with the Manning's counterion condensation theory. Analyses were performed in under 15 minutes using a phosphate or borate buffer.

KEY WORDS: Capillary zone electrophoresis, polymers, polyelectrolytes, acrylamide, acrylic acid.

RESUMEN

Se ha realizado el análisis mediante electroforesis capilar de zona (ECZ) de diversos copolímeros obtenidos por copolimerización radical de acrilamida y N,N'-dimetilacrilamida empleando como comonómeros ácido acrílico y 1-vinil-2-pirrolidona en diferentes relaciones comonoméricas molares en la alimentación. Los resultados demuestran que el valor absoluto de la movilidad electroforética depende fuertemente de la densidad de carga lineal del copolímero en concordancia con la teoría de la condensación de contraiones de Manning. Los análisis se realizan en menos de 15 minutos usando un tampón fosfato o borato.

PALABRAS CLAVES: Electroforesis capilar de zona, polímeros, polielectrolitos, acrilamida, ácido acrílico.

INTRODUCTION

Polymeric materials are among materials that are widely used both in industry and in households today. In fact, it is difficult to imagine our everyday existence without these materials. These products are most often formulated to consist of mixtures of isomers, homologues, and even oligomers having a wide distribution of molecular weight in order to meet specific physical and chemical requirements. The complexity of othe chemical composition of these materials has increased the need for more reliable analytical methodologies for both process and quality control of these materials.

Since the pioneering work of Jörgenson and Lukacs1) capillary electrophoresis (CE) has rapidly emerged as a fast high-resolution separation technique applicable to a wide variety of charged and uncharged compounds2-6). The advantages of CE such as high efficiency, small sample size and unique selectivities compared to HPLC have frequently been discussed but the significant reduction in running costs and the avoidance of the use of large volumes of expensive and hazardous solvents also render CE an attractive alternative to HPLC. However, the application of CE to the study of synthetic water-soluble polymers is relatively new but shows considerable promise. Sulfonated polystyrene has been separated over a wide range of molecular weight, by using a carrier solution containing hydroxyethyl cellulose7). A baseline separation of two polyanions, polyphosphates and polycarboxylates was obtained8). Recently, the separation of a series of the random copolymers consisting of the ionic monomer sodium 2-acrylamido-2-methyl-propanesulfonate and the nonionic monomer acrylamide, by free-zone CE, was described9). These research studies indicate the potential of CE for separating and analyzing polyelectrolytes in aqueous solution.

Polyelectrolytes in aqueous solution are typically modeled as charged polyion chains tightly surrounded by an atmosphere of counterions10-12). According to Manning's counterion condensation theory10), when the dimensionless linear charge density (x) is higher than |Z|-1, where Z is the valence of the counterion, counterions will condense on the polyion until x is |Z|-1. For x £ |Z|-1, no counterion condensation takes place at all. Here, the dimensionless linear charge density, x, is defined in S.I. units as:

where e is the elementary charge of a proton (C), D is the dimensionless dielectric constant of othe solvent, kB is the Boltzmann constant (J/K), e0 is the permitivity of vaccuum (F/m). T is the temperature (K), and b is the average charge spacing along the polyion chain (m). Encouraged by earlier results9), and in order to investigate this phenomenon and evaluate the corresponding theories, CE in the open tube format with conventional UV detection seems to be the most adequate method to investigate and characterize water-soluble copolymers obtained by radical copolymerization of acrylamide (Am) and N,N'-dimethylacrylamide (DMAm) using acrylic acid (AA) and 1-vinyl-pyrrolidone (VPyr) as comonomers in different molar feed ratios13-15).

 

EXPERIMENTAL

Synthesized copolymers, with different comonomeric feed ratios, consisting of (Am/AA), (DMAm/AA), and (Am/VPyr) were studied. Their structures are depicted as follows:

It is known that the average charge spacing (b) for vinyl polymers, such as polyacrylic acid, carrying a charge on each monomeric unit is 2.55 Å corresponding to a linear charge density of x = 2.8 at room temperature9,12). The linear charge densities of the copolymers were therefore obtained by multiplying the linear charge density of the homopolymer, poly(AA), by the mole fraction of AA monomer in the copolymers. Copolymer composition of charged and neutral polymers as well as the corresponding linear charge densities calculated for charged polymers are shown in Table I. Copolymer composition was determined from elemental analysis and also from 1H-NMR spectra13-15).

TABLE I. Copolymer composition and linear charge density (x).

Polymer
samplea
 
Feed monomer ratio
Am or DMAm:AA,
or Am : VPyr
Copolymer composition
Am or DMAm:AA,
or Am : VPyr
Ref.
xb
   
   

1
3:1
2.73 : 1.00
 
0.75
2
3:2
2.04 : 1.00
 
0.92
3
1:1
1.15 : 1.00
13
1.30
4
2:3
0.94 : 1.00
 
1.52
5
1:3
0.58 : 1.00
 
1.77
6
3:1
33.3 : 1.00
 
0.08
7
3:2
5.26 : 1.00
 
0.45
8
1:1
2.13 : 1.00
14
0.89
9
2:3
1.56 : 1.00
 
1.09
10
1:3
0.69 : 1.00
 
1.66
11
3:1
4.71 : 1.00
 
-
12
3:2
3.00 : 1.00
 
-
13
1:1
2.60 : 1.00
15
-
14
2:3
2.34 : 1.00
 
-
15
1:3
1.54 : 1.00
 
-

a) Copolymers 1-5, 6-10 and 11-15 correspond to poly(Am-co-AA), poly(DMAm-co-AA) and poly(Am-co-VPyr) respectively.
b) x cannot be determined for neutral copolymers 11-15.

Separations were conducted using a PrinCE 450 Capillary Electrophoresis System connected with a Lambda 1010 UV-Vis detector (Bishoff, Leonberg, Germany). System DAx software was used for data acquisition and analysis. Polymicro capillaries (Phoenix, AZ) were used with an inner diameter of 50 µm, an outer diameter of 375 µm and a total length of 70 cm (55 cm to the detector). The experiments were performed at 30 kV and 25°C. Detection was by direct UV absorbance at 200 nm. Samples were injected hydrodynamically at 50 mbar for 15 s.

The buffers were prepared using reagent sodium hydrogen phosphate (Aldrich) and boric acid (Aldrich). The buffer was adjusted to basic pH using NaOH (Merck). Samples were prepared by dissolving the copolymers (1.0 mg/mL) in 50 mM phosphate or borate buffer solution containing at 0.05% (w/w) of mesityl oxide, as neutral marker. All solutions were filtered through 0.45 µm nylon filters prior to use. Quintuplicate runs of each sample showed that the electrophoretic mobility, µep calculated by subtracting the electroosmotic flow (EOF) of the run buffer from the apparent electrophoretic mobility of samples, was reproducible to within 2%.

RESULTS AND DISCUSSION

Conclusions derived from literature9) show that polymers of strongly ionized and uncharged monomers can be studied in a straightforward way than carboxylic acid polymers due to the adsorption of unionized acrylic units onto the negative charged wall of the bare fused silica capillary, as it has been reported elsewhere16). Consequently, in order to avoid adsorption, the electrophoretic runs were performed in a basic pH range. So, the change as a result of electrostatic induction effects on the pK of acrylic acid was minimized as it is seen in Figure 1 which clearly shows breaks in the slope of the best-fit lines at x =1.1 for data obtained for different copolymers of Am/AA in phosphate and borate buffers indistinctively. As expected, these results are in agreement with the predictions of Manning's theory for systems with monovalent counterions as well as supporting to the evidence for the physical reality of counterion condensation when x >1. Thus, counterion condensation explains very well the strong dependence of polymer composition on the electrophoretic mobility of the copolymers. Therefore, free-zone CE should be effective in separating polyelectrolytes as long as x < 1. On the other hand, due to counterion condensation where counterions condense until the net value of x is lowered to 1, it could not distinguish different polymers with x > 1. Figures 2 and 3 show the electrophrograms of a series of copolymers of Am/AA and DMAm/AA synthesized according to the different feed molar ratios shown in Table I. Figure 2 shows clearly the linear charge density dependence on migration time. The width of the peaks may also reveal the extent of polydispersity in terms of monomer compositions. In Figure 3 the migration time differences between copolymers a and b are higher because of their x values, 0.08 and 0.45 respectively, whereas c and d have x > 1. Moreover, the width of the peaks may also reveal the extent of polydispersity according to comonomer composition and the meaning of shoulders should be the coexistence of two fractions, one of them with a slightly higher acrylic acid content.

 

 

FIG. 1. Electrophoretic mobility of a series of copolymers
of poly(Am/AA) with different linear charge densities (see Table I).

 

 

 

 

 

 

FIG. 2. Electropherograms of a series of copolymers of poly(Am-co-AA).
Feed monomer ratio (Am:AA): a (3:1); b(3:2); c(2:3) and d (1:3).
Carrier electrolyte: 50 mM borate buffer pH 11.5.
For other experimental conditions, see Experimental.

 

 

 

 

 

 

 

FIG. 3. Electropherograms of a series of copolymers of poly(DMAm-co-AA).
Feed monomer ratio (DmAm:AA): a(3:1); b(3:2); c(1:1) and d(2:3).
Carrier electrolyte: 50 mM phosphate buffer pH 11.5.
For other experimental conditions, see Experimental.

 

 

 

 

In order to verify that neutral copolymers should migrate with the electroosmotic flow, the poly(Am-co-VPyr) series and the homopolymer poly(VPyr) were analyzed. As expected the electropherograms in Figure 4 show, within experimental error, the certainty of this.

Figure 5 demonstrates the separation of a mixture of copolymers of poly(Am-co-AA) consisting of two mole composition, 33% AA and 47% AA. Since the linear charge densities of these copolymers are closer to x = 1, two components of the mixture are not well resolved by CE.

 

FIG. 4. Electropherograms of a series of copolymers of poly(Am-co-VPyr). Feed monomer ratio (Am:VPyr): a(1:3); b(1:1); c(3:2); d(3:1). Homopolymer poly(VPyr) (e). Conditions: 50 mM phosphate buffer pH 11.5. For other experimental conditions, see Experimental.

 

 

 

 

 

 

 

FIG. 5. Electropherogram of a mixture of copolymers of poly(Am-co-AA). Feed monomer ratio (Am:AA); a(3:2) and b(1:1). Conditions: 30 mM phosphate buffer pH 9.5. For other experimental conditions, see Experimental.

 

 

 

 

 

 

CONCLUSIONS

Accordingly, CE has been shown to be a useful tool for analysis and separation of charged copolymers based on their difference in linear charge density, as long as x < 1. Then, CE can be used to characterize the composition distribution of copolymers made up of charge and uncharged monomers. On the other hand, CZE is not able to separate neutral polymers and, due to counterion condensation, polyelectrolytes with higher linear charge density.

 

ACKNOWLEDGEMENTS

Financial support for this research was provided by the Dirección de Investigación of the Universidad de Concepción, DIUC (Project N 97.021.007-1.2). Dr. I. Peric thanks to the Deutscher Akademischer Austauschdienst DAAD program of the Federal Republic of Germany for financing his stay at University of Tübingen and to the Deutsche Gesellschaft für Technische Zusammenarbeit (GTZ) for the provision of the capillary electrophoresis device.

______________________

*To whom correspondence should be addressed.

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