Liquid–Liquid Equilibrium Data for Aqueous Two-Phase Systems

Article pubs.acs.org/jced

Liquid−Liquid Equilibrium Data for Aqueous Two-Phase Systems Containing PPG725 and Salts at Various pH Values Mohammad-Kazem Shahbazinasab and Farshad Rahimpour* Biotechnology Research Lab, Chemical Engineering Department, Faculty of Engineering, Razi University, Kermanshah 67149-67346, Iran ABSTRACT: The phase diagrams and compositions of coexisting phases have been determined for three aqueous two-phase systems (ATPS's) containing poly(propylene glycol) (PPG) with an average molecular weight of 725 and sodium citrate, potassium phosphate, or magnesium sulfate at 25 °C and several pH values. The effect of pH on the salting-out effect of salts in the polymer phase by sodium citrate, potassium phosphate, and magnesium sulfate has been studied. It was found that an increase in salt composition at constant pH increases the concentration of PPG in the top phase and an increase in pH caused the expansion of the two-phase region for these systems. Increasing the pH also increases the concentration of PPG in the PPG-rich phase for systems containing sodium citrate and magnesium sulfate and decreases it for systems containing potassium phosphate, while the salt-rich phase for sodium citrate and magnesium sulfate systems will be more diluted and for potassium phosphate system will be somewhat concentrated. Furthermore, the Merchuk equation has been successfully used to correlate the experimental binodal data of mentioned systems (high R2 and low SD). Also, the dependence of Merchuk equation coefficients with pH were presented by two empirical equations.

■

INTRODUCTION An aqueous two-phase system (ATPS) is produced by adding either two chemical structurally different hydrophilic polymers, such as dextran and poly(ethylene glycol) (PEG), or a polymer and a salt, such as PEG and K2HPO4 + KH2PO4, to water, causing the system to separate into two immiscible water-rich phases.1 ATPS's provide a mild nondenaturing environment for biomolecules, and they are easy to scale-up. ATPS's have been extensively exploited to process different biological sources for the recovery and purification of biological products and in the extractive bioconversion of enzymes.1,2 The separation technique is also becoming important in nonbiotechnology areas such as industrial waste remediation.3−5 Data on the composition and properties of phase systems are necessary for the design and optimization of extraction processes, the understanding of general factors determining the partition of solutes and particles in such ATPS's, and the development and testing of both thermodynamic and mass transfer models of ATPS's. Poly(ethylene glycol) (PEG), which is a hydrophilic polymer, is often used in aqueous two-phase partitioning studies. In laboratory-scale separations, the most commonly used systems are comprised of the polymers PEG and dextran, while for large scale bioprocesses, PEG−salt systems provide an attractive alternative to other process, because of the inexpensive chemicals that compose these systems, their greater selectivity, lower viscosity, lower cost, rapid phase disengagement, and the availability of commercial separators, which allow a faster and continuous protein separation.2,4 Poly(propylene glycol) (PPG) is a polymer that is structurally closely related to PEG. Low molecular weights of PPG are completely soluble in water, while high molecular weights are © 2012 American Chemical Society

only partially soluble. This polymer can also be used for the separation of biomolecules, since its aqueous solutions with a suitable polymer or a salt form a two-phase system. However, liquid−liquid equilibrium (LLE) data of the aqueous PPG−polymer and PPG− salt systems are relatively scarce. In regard to PPG−salt systems, the liquid−liquid equilibrium data for PPG425 + sodium chloride,6 PPG425 + magnesium sulfate,7 PPG425 + ammonium sulfate,8 PPG425 + sodium sulfate,8 PPG400 + sodium sulfate,9 PPG400 + sodium carbonate,9 PPG400 + sodium nitrate,9 PPG400 + tripotassium citrate,10 and PPG425 + sodium citrate11 ATPS's have been reported. This work is devoted to obtaining phase equilibrium data and the correlation of binodal data for ATPS's containing PPG725 + sodium citrate at pH 8.22, 7.05, 6.38, 5.5, 4.81, and 3.81, PPG725 + potassium phosphate at pH 9.5, 8.4, 7.3, 6.35, 5.3, 4.54, and 3.45, and PPG725 + magnesium sulfate at pH 5.56, 4.05, 3, and 2.43. As far as we know there are no reports on the phase diagrams of these systems at these wide ranges of pH. These results can be used to obtaining experimental data for choosing suitable systems for the purification of biomolecules.

■

EXPERIMENTAL SECTION Materials. PPG (average molecular weight Mr = 725, containing (130 to 190) ppm proprietary phenolic antioxidant), trisodium citrate (anhydrous GR for analysis, > 99 %), citric acid (anhydrous powder extra pure, > 99.5 %), dipotassium hydrogen phosphate (anhydrous GR for analysis, > 99.0 %), potassium Received: April 30, 2011 Accepted: May 31, 2012 Published: June 14, 2012 1867

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

Table 1. Binodal Data for the PPG725 (1) + Sodium Citrate (2) + H2O (3) System at Different pH Values pH = 8.22

pH = 7.05

pH = 6.38

pH = 5.5

pH = 4.81

pH = 3.81

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

16.65 13.85 11.83 9.46 7.99 7.24 5.92 5.39 5.13 4.49 4.13 3.91 3.62 3.45 3.22 3.03 2.87 2.74

1.07 1.71 2.2 2.8 2.98 3.37 3.79 4.17 4.2 4.42 5.36 5.6 5.88 6.21 6.33 6.38 6.49 6.67

19.29 16.52 13.05 11.46 10.39 9.08 7.85 6.91 6.14 5.23 4.42 4.01 3.56 3.15 2.87 2.27

0.88 1.62 1.87 2.27 2.59 2.92 3.35 3.34 3.48 4.11 6.02 6.13 6.08 6.18 6.93 7.82

15.68 13.36 12.12 11.32 10.03 9.48 8.66 8.03 7.29 6.78 6.48 5.97 5.57 5.09 4.78 4.44 3.88 3.71 3.37 3.24 2.95 2.87 2.76 2.58 2.37 2.17

2.2 3.05 3.45 3.53 3.74 4.05 4.22 4.41 4.49 4.35 4.47 4.58 4.83 4.99 5.29 5.97 5.51 5.97 6.17 6.61 6.79 6.94 7.68 7.83 8.95 9.22

19.49 15.64 12.06 10.23 8.73 7.829 7.195 6.109 5.672 5.139 4.6 4.116 3.791 3.316 2.98 2.668 2.351 1.916

1.49 1.99 2.99 3.57 4.21 4.73 4.84 5.39 5.66 5.89 6.04 6.81 7.15 7.66 8.02 8.71 8.96 9.17

17.7 14.9 13.6 11.6 10.6 9.58 8.92 8.35 7.51 6.89 6.46 5.89 5.26 4.92 4.58 4.16 4.13 3.91 3.6 3.37 3.14 2.92 2.76 2.58

2.68 3.34 3.95 4.88 5.27 5.88 6.17 6.58 6.93 6.81 7.13 7.69 8.08 8.29 8.69 9.87 10.16 10.26 10.51 11.08 11.13 11.76 12.18 12.33

16.56 13.48 11.51 10.33 8.98 8.08 7.12 6.02 5.25 4.77 4.38 3.78 3.37 3.02 2.83 2.54 2.35 2.09

4.262 6.458 7.635 8.384 9.493 10.14 10.84 11.78 12.34 12.25 12.76 13.19 14.23 14.94 15.57 15.9 16.45 16.58

Table 2. Binodal Data for the PPG725 (1) + Potassium Phosphate (2) + H2O (3) System at Different pH Values pH = 9.5

pH = 8.4

pH = 7.3

pH = 6.35

pH = 5.3

pH = 4.54

pH = 3.45

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

14.91 12.4 10.34 8.632 6.254 5.957 5.58 5.196 5.007 4.741

0.75 1.25 1.99 2.21 3.08 3.2 3.25 3.45 3.75 3.82

13.8 10.3 9.3 8.1 7.31 6.4 5.98 5.42 5.01 4.29 3.88 3.48 3.27 2.81 2.51 2.32 2.21 2.03 1.74 1.57 1.4

1.27 2.81 3.42 3.6 3.92 4.02 4.38 4.68 5.16 5.34 5.78 5.67 6.12 6.69 7.12 7.26 7.54 7.75 8.14 8.28 9.05

24 20.1 16.2 14.4 12.7 11.3 10.3 9.15 8.48 7.63 6.98 6.56 6.09 5.76 5.4 5.03 4.73 4.53 4.25 3.87 3.56 3.31 3.01

0.97 1.8 2.46 3.02 4.04 4.07 4.48 4.57 4.81 5.29 5.2 5.71 5.86 5.9 6.19 6.38 6.6 6.85 7.15 7.51 7.72 8.04 8.34

21 17.3 15.4 13.1 11.8 10.8 10.3 9.48 8.55 7.96 7.55 7 6.36 5.87 5.61 5.2 4.87 4.57 4.2 3.88 3.57

1.21 2.14 2.78 3.1 4.23 4.52 4.6 4.75 5.2 5.22 5.24 5.42 5.97 6.12 6.35 6.48 6.53 6.8 6.99 6.98 7.15

22.8 17.8 15.7 14 12.3 10.7 9 8.42 7.86 7.4 7.02 6.63 5.84 5.26 4.91 4.67 4.48 4.26 4.06 3.72 3.49 3.29

1.69 2.43 3.36 3.46 3.82 4.34 4.27 4.6 4.84 5.06 5.25 5.51 7.98 8.12 8.58 8.59 8.43 8.85 9.23 9.13 9.62 9.66

22 18.9 16.3 14.3 12.3 10.6 9.47 8.26 7.4 6.8 6.41 5.67 5.21 4.81 4.54 4.2 3.93 3.59 3.24

1.56 2.37 3.26 4 4.67 6.05 6.34 6.76 6.81 7.31 7.28 7.8 8.2 8.5 8.64 9 9.13 9.45 9.98

12.1 9.81 9.03 8.33 7.59 6.99 6.25 5.9 5.54 4.9 4.48 3.95 3.71

3.65 3.99 4.25 4.7 4.9 5.46 6.24 6.38 6.96 7.86 8.1 8.28 9.02

dihydrogen phosphate (GR for analysis, > 99.5 %), magnesium sulfate (anhydrous GR for analysis, > 99.0 %), and sulfuric acid ((95 to 97) % H2SO4, GR for analysis, > 95.0 %) were obtained from Merck (Darmstadt, Germany). The polymer and salts were used

without further purification, and double-distilled−deionized water was used. All of the other reagents were of analytical grade. Apparatus and Procedure. Aqueous two-phase systems (ATPS's ) were prepared by mixing PPG725 and sodium citrate, 1868

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

Table 3. Binodal Data for the PPG725 (1) + Magnesium Sulfate (2) + H2O (3) System at Different pH Values pH = 5.56

pH = 4.05

pH = 3

pH = 2.43

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

100 w2

9.44 7.293 6.772 5.954 5.464 4.922 4.004 3.655 3.166

0.538 1.681 2.332 3.221 3.423 4.098 4.957 5.645 6.138

14.19 11.31 8.511 7.341 6.061 5.507 4.208 3.699 3.049 2.385 1.851 1.469

0.946 2.088 5.368 5.685 5.812 5.789 6.96 7.92 9.443 11.72 13.99 13.87

9.19 6.23 4.86 3.679 3.384 3.12 2.634 2.184 1.721 1.519

2.16 3.83 5.601 6.344 6.815 7.368 8.304 9.828 9.706 10.72

10.9 9.471 7.74 6.443 5.806 4.775 4.037 3.48 2.749 2.491 2.142 1.748 1.614 1.429

0.515 1.318 2.173 3.633 4.01 4.887 6.309 7.618 9.283 9.68 11.14 12.00 12.64 12.44

Table 4. Phase Compositions for the PPG725 (1) + Sodium Citrate (2) + H2O (3) System at Different pH Values total composition

bottom phase

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

STLa

TLLb

8.22

15 20 25 30 15 20 25 30 15 20 25 30 15 20 25 30 15 20 25 30

10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10

0.004 0.003 0.002 0.002 0.006 0.004 0.003 0.003 0.008 0.007 0.007 0.005 0.007 0.006 0.006 0.005 0.006 0.005 0.005 0.004

84.7 89.25 90.95 93.38 77.23 82.6 85.83 88.33 69.17 75.83 79.67 81.42 62.42 65.19 69.08 71.84 51.75 55.33 63.8 65.83

16.96 22.49 28.07 33.57 17.2 22.72 28.27 33.81 17.48 23.01 28.57 34.19 17.78 23.55 29.17 34.85 18.22 24.23 29.26 34.99

0.2 0.15 0.1 0.05 0.15 0.1 0.05 0.04 0.2 0.1 0.05 0.03 0.3 0.2 0.1 0.04 0.8 0.4 0.25 0.15

−4.98 −3.962 −3.237 −2.779 −4.482 −3.63 −3.034 −2.611 −3.944 −3.291 −2.786 −2.38 −3.493 −2.759 −2.364 −2.06 −2.792 −2.267 −2.165 −1.871

86.184 91.894 95.087 99.183 78.974 85.570 90.317 94.541 71.149 79.147 84.588 88.278 64.612 69.123 74.892 79.809 54.108 60.035 69.960 74.417

7.05

6.38

5.5

4.81

a

top phase

pH

Slope of the tie line. bTie line length.

by this method is about 0.2 (wt %). The concentration of PPG was determined by refractive index measurements at 25 °C using an ATAGO-DTM1 refractometer with a precision of 0.002. Since the refractive index of phase samples depends on PPG, and salt concentration, calibration plots of refractive index versus polymer concentration were prepared for different concentrations of salt. The binodal curves were determined by a titration method. A polymer solution of known concentration was titrated with the salt solution or vice versa, until the solution turned turbid, which indicated the formation of two liquid phases. In accordance with the amount of titrant added until turbidity was observed, the composition of the mixture for each point on the binodal curve was calculated by mass using an A&D GF-300 analytical balance with a precision of ± 1·10−4 g. To determine the compositions of coexisting phases, feed samples (10 g) were prepared by mixing appropriate amounts of polymer, salt, and water in the tube. After the separation of the two transparent phases, the concentrations

potassium phosphate, or magnesium sulfate solution at the required pH in 15 mL graduated tubes. The final weight of the systems was adjusted to 10 g by the addition of double-distilled− deionized water. The pH of the salt solutions were adjusted by mixing the appropriate ratio of trisodium citrate and citric acid, dipotassium hydrogen phosphate, and potassium dihydrogen phosphate, magnesium sulfate, and sulfuric acid, respectively. All experiments were carried out at 25 °C. The pH values of the solutions were measured precisely with a pH meter JENWAY 3345. The resulted solution was mixed by rigorous vortexing the test tube for 2 min. The tubes were placed in 25 °C water bath for 24 h and then were centrifuged at 1500 rpm for 10 min; the solution reached equilibrium, and the samples of the top and bottom phase were carefully withdrawn, with care being taken to leave a layer of solution at least 0.2 cm thick above the interface. The concentration of salt were determined by atomic absorption spectroscopy (AAS), using Shimatzu AA-6300 equipment. The average relative deviation of salt concentration 1869

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

Table 5. Phase Compositions for the PPG725 (1) + Potassium Phosphate (2) + H2O (3) System at Different pH Values total composition

bottom phase

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

STLa

TLLb

9.5

10 15 20 25 10 15 20 25 10 15 20 25 10 15 20 25 10 15 20 25 10 15 20 25 10 15 20 25

10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10

0.005 0.003 0.003 0.003 0.006 0.006 0.004 0.003 0.007 0.006 0.003 0.003 0.006 0.005 0.004 0.004 0.008 0.006 0.006 0.005 0.008 0.008 0.006 0.005 0.009 0.009 0.008 0.007

42.5 54 68.25 79.09 43.25 56.3 69.75 81.98 44.92 58.67 73.5 85.12 45.24 61.5 74.42 89.11 57.29 73.42 79.88 91.5 63.52 75.75 83.92 94.08 82.01 84.92 87.32 96.07

13.03 18.16 23.21 28.15 12.86 17.95 23.16 28.12 12.58 17.35 23.11 28.02 12.47 17.27 23.08 27.99 11.63 17.19 22.99 27.98 11.38 17.01 22.9 27.84 11.01 16.8 22.53 27.81

0.145 0.02 0.012 0.01 0.506 0.311 0.21 0.106 1 0.4 0.1 0.05 1.3 0.55 0.1 0.05 2.3 0.7 0.1 0.05 2.6 0.9 0.12 0.07 2.7 1 0.2 0.1

−3.25 −2.96 −2.929 −2.79 −3.325 −3.108 −2.997 −2.898 −3.492 −3.317 −3.176 −3.023 −3.524 −3.494 −3.22 −3.175 −4.729 −4.229 −3.478 −3.265 −5.351 −4.391 −3.676 −3.371 −7.203 −4.992 −3.866 −3.448

44.312 56.952 72.076 83.940 44.635 58.794 73.294 86.567 45.684 60.796 76.951 89.565 45.674 63.348 77.820 93.354 56.205 74.723 83.025 95.633 61.972 76.757 86.871 98.044 80.069 85.583 89.984 99.916

8.4

7.3

6.35

5.3

4.54

3.45

a

top phase

pH

Slope of the tie line. bTie line length.

Table 6. Phase Compositions for the PPG725 (1) + Magnesium Sulfate (2) + H2O (3) System at Different pH Values total composition

bottom phase

100 w2

100 w1

100 w2

100 w1

100 w2

100 w1

STLa

TLLb

5.56

15 20 25 10 15 20 25 10 15 20 25 10 15 20 25

10 10 10 10 10 10 10 10 10 10 10 10 10 10 10

0.007 0.006 0.006 0.008 0.007 0.006 0.006 0.006 0.006 0.005 0.003 0.010 0.008 0.007 0.005

93 97.17 99.17 87.38 91.25 95 97.5 80.87 82 85.8 91.67 57.57 66.25 75 85.25

16.72 22.27 27.81 10.96 16.67 22.27 27.81 11.21 16.98 22.58 28.01 11.73 17.45 22.99 28.27

0.47 0.15 0.05 2.6 0.95 0.34 0.2 1.4 0.5 0.15 0.1 1.8 0.8 0.25 0.15

−5.533 −4.357 −3.565 −7.737 −5.416 −4.25 −3.499 −7.087 −4.799 −3.792 −3.267 −4.756 −3.75 −3.251 −3.01

94.02726 99.54178 102.9458 85.48447 91.82454 97.243 101.1946 80.25591 83.24882 88.57513 95.75728 56.98817 67.73423 78.20346 89.67118

4.05

3

2.43

a

top phase

pH

Slope of the tie line. bTie line length.

takes a sample with a known composition above the binodal, this mixture becomes a two-phase system where the composition of resulted top and bottom phases can be related by tie lines. Binodal data obtained from turbidimetric titrations for ATPS's containing sodium citrate, potassium phosphate, and magnesium sulfate at various pH are presented in Tables 1, 2, and 3, and equilibrium phase compositions are shown in Tables 4, 5, and 6, respectively.

of the salts and PPG in the top and bottom phases were determined. Each experiment was done two times, and the results were the average of them.

■

RESULTS AND DISCUSSION

The region below the binodal represents homogeneous solutions, and above it represents the two-phase region. If one 1870

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

where y = a or b.

For the correlation of binodal data of some aqueous PEG + salt systems the following nonlinear expression was developed by Merchuk et al.:12 w1 = a exp(bw2 0.5 − cw2 3)

c = A exp[− ((pH − B)/pH)2 ] + c exp[− ((pH − D)/pH)2 ]

(1)

The obtained fitted coefficients are given in Tables 10, 11, and 12, respectively. On the basis of the obtained standard deviations and coefficient of determination (R2), we conclude that eq 1 can be satisfactorily used to reproduce the binodal curves of the investigated system. Experimental and correlated binodals curves of investigated systems with above Sequence are shown in Figures 1, 2, and 3, respectively. The effect of pH on the phase-forming ability of biphasic systems containing sodium citrate, potassium phosphate, and magnesium sulfate systems is illustrated in Figures 1, 2, and 3, respectively. It can be seen that the two-phase area is expanded and the binodal curve shifts downward as the pH of the medium increased, indicating that the smaller concentration of the phase polymers is required to form ATPS's. In other words, if we take a sample with a known composition on the binodal curve, this mixture becomes a two-phase system at increasing pH as has been observed experimentally. This behavior was also seen in the case of study of the temperature effect on phase equilibrium of PEG-salt ATPS's.18 The pH could affect the binodal location, either by changing the charge of the solute or by altering the ratio of the charged species present. It was found that, at higher pH, hydrogen-bond interactions of PPG are weakened.7 The depression of the cloud point by increasing pH may be related to the salting-out phenomenon resulting from the weakening of the PPG−solvent interaction. This may be explained based on the salt's ability to promote the water structure.19 Based on the concept of the water structure-promoting capability (kosmotropicity) of salts, when a kosmotropic salt like sodium citrate, potassium phosphate, or magnesium sulfate is dissolved in an aqueous solution, the ionic hydration process will occurs, and salt ions are surrounded by a layer of water molecules. These water molecules are structured and immobilized, so that their function as solvent to other molecules is reduced. In this case, when a kosmotropic salt is added to an aqueous solution of a hydrophilic PPG, they compete with each other for the water molecules. Due to the stronger affinity of salt ions for the water relative to PPG, a decrease in the hydration and the solubility of PPG occurs. As a result, at certain concentrations, the hydrophilic PPG is salted-out and excluded from the rest of the solution as a separated phase. This may be due to the fact that the degree of protonation of the salt anions is changed as medium pH changes.19 The anions are less protonated and hence have a higher valency, as the aqueous medium pH increases. Anions with a higher valence are better salting-out agents than anions with a lower valence. This is because the anion with a higher valence hydrates more water molecules than the anions with lower valence and also in the case of anions with higher valence, the repulsive interaction between the anions and the anionic-like polyether functionality of PPG is larger. Thus, the anions with a higher valency that exist in the basic medium pH are more efficient to promote the phase separation, compared to those with minor valency found in the acidic medium pH. This observation that an increase in aqueous medium pH increases the water-structure-promoting capability of the salt ions is consistent with the fact that the water structuring is accompanied by a decrease of the water acidity in pure water.19 From this point of view, the hydronium ion, H3O+, which is present in the acidic medium pH, seems to block the water structure.

where a, b, and c represent the fitting parameters and w1 and w2 represent the concentrations (in weight percent) of polymer and salt, respectively. Recently, the above equation has been successfully used for the correlation of binodal data of some aqueous PEG + salt,13−15 PPG + salt,8,16 and UCON + (sodium or potassium) phosphate salt systems.17 The binodal data of examined systems were correlated by least-squares regression to the above expression. Using eq 1, the fitting parameters a, b, and c obtained from the correlation of experimental binodal data along with the corresponding standard deviations for sodium citrate, potassium phosphate, and magnesium sulfate ATPS's at various pH are given in Tables 7, 8, and 9, respectively. Also, the relations between pH and the coefficients a and b were presented by eq 2, while the relation between coefficient c and pH was fitted by eq 3. Table 7. Values of Parameters of Equation 1 for PPG725 (1) + Sodium Citrate (2) + H2O (3) at Different pH Values pH

a

b

c·104

SDa

R2

8.22 7.05 6.38 5.5 4.81 3.81

61.99167 62.17792 368.7062 42.94843 88.23467 62.80282

−1.18 −1.15 −1.93 −0.774 −0.942 −0.594

0.84 0.93 9.17 8.97 1.14 2.14

0.5707 0.8777 1.7478 0.4338 0.5073 0.8237

98.8 98.8 99.1 96.4 97.4 98.7

2 0.5 SD = (∑Ni=1(wcalcd − wexptl 1 1 ) /N) , where w1 and N represent the concentration (in weight percent) of polymer and the number of binodal data, respectively.

a

Table 8. Values of Parameters of Equation 1 for PPG725 (1) + Potassium Phosphate (2) + H2O (3) at Different pH Values pH

a

b

c·104

SD

R2

9.5 8.4 7.3 6.35 5.3 4.54 3.45

30.56942 47.46535 61.55924 34.81332 103.5443 45.15044 79.83803

−0.799 −0.953 −0.841 −0.46 −1.14 −0.552 −1.04

59.6 10.7 11.3 27.6 2.54 9.91 1.07

0.3417 0.701 0.8417 0.4284 0.9332 0.43 0.3963

98.9 98.4 98.5 99.1 96.2 99.2 98

Table 9. Values of Parameters of Equation 1 for PPG725 (1) + Magnesium Sulfate (2) + H2O (3) at Different pH Values pH

a

b

c·104

SD

R2

5.5 4.05 3.0 2.43

12.8071 29.37077 27.93834 16.60992

−0.419 −0.652 −0.754 −0.525

15.8 1.73 3.52 2.6

0.1158 0.907 0.1936 0.2284

99.6 95.7 98.3 99.5

(3)

y = (p1 · pH3 + p2 · pH2 + p3 · pH + p4 )/(pH2 + q1· pH + q2) (2) 1871

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

Table 10. Values of Parameters of Equation 2 between Coefficient a and pH system

p1

p2

p3

p4

q1

q2

SD

R2

PPG725 (1) + sodium citrate (2) + H2O (3) PPG725 (1) + potassium phosphate (2) + H2O (3) PPG725 (1) + magnesium sulfate (2) + H2O (3)

0.9546 −10.38 0

43.89 244.9 −22.07

−597.6 −1759 224.4

1783 3947 −344

−11.4 −11.45 0

32.03 32.16 0.05816

1.514 2.667 0.039

99.1 96.0 100

Table 11. Values of Parameters of Equation 2 between Coefficient b and pH system

p1

p2

p3

p4

q1

q2

SD

R2

PPG725 (1) + sodium citrate (2) + H2O (3) PPG725 (1) + potassium phosphate (2) + H2O (3) PPG725 (1) + magnesium sulfate (2) + H2O (3)

0 0.03077 0

−1.021 −1.487 0.1822

10.09 13.41 1.854

−23.71 −33.17 3.285

−10.06 −10.82 0

23.96 28.63 2.154

0.0513 0.0498 0.0013

98.9 97.0 100

Table 12. Values of Parameters of Equation 3 between Coefficient c and pH system PPG725 (1) + sodium citrate (2) + H2O (3) PPG725 (1) + potassium phosphate (2) + H2O (3) PPG725 (1) + magnesium sulfate (2) + H2O (3)

A −0.05567 0.008092 0.0002395

B

C

−6.717 −2.403 2.638

0.002183 387.9 0.4852

D 9.466 −41.6 −18.8

R2

SD −4

3.42·10 7.02·10−4 62.35·10−4

95 98.5 99

Figure 3. Experimental and correlated binodal curves of ATPS's of PPG725 (1) + magnesium sulfate (2) + H2O (3) at different pH values. The solid curves represent correlated binodals.

Figure 1. Experimental and correlated binodal curves of ATPS's of PPG725 (1) + sodium citrate (2) + H2O (3) at different pH values. The solid curves represent correlated binodals.

Also it can be concluded from the data of Tables 4 to 7 that an increase in salt composition at constant pH increases the concentration of PPG in the top phase. This might be explained by the hydration effect of salt. The slope of the tie line (STL) is given by the ratio of the difference between the concentration of the polymer (CP) and of the salt (CS) in the top and bottom phases, as presented in eq 4: STL = (C Ptop − C Pbottom)/(CSbottom − CStop)

(4)

The tie-line length (TLL) is an empirical measure of the compositions of the two phases, which can be calculated using eq 5: TLL = [(C Ptop − C Pbottom)2 + (CSbottom − CStop)2 ]0.5

(5)

The tie lines are determined by connecting each corresponding set of total, bottom, and top-phase points. All of the samples with feed compositions on a tie-line have the same top and bottom phase composition. On the basis of the lever rule, the relative amounts of the phases are in inverse proportion to the distances of the respective phase boundary lines (binodal) from the point of overall composition.21 As an example, the tie lines for the systems containing sodium citrate with pH 8.22, 5.5, and 4.81, potassium phosphate with pH 9.5, 5.3, and 3.45 and magnesium sulfate with pH 5.56, 3, and 2.43 are shown in Figures 4, 5, and 6, respectively. As shown in Figure 5, the slope and the length of the equilibrium

Figure 2. Experimental and correlated binodal curves of ATPS's of PPG725 (1) + potassium phosphate (2) + H2O (3) at different pH values. The solid curves represent correlated binodals.

However for potassium phosphate, the breakdown of structured water molecules around the PPG-chains associated with an increase in pH is probably the other cause of phase separation since binodal curves corresponding to the some different pH's overlap.20 1872

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

varying pH. Increasing the pH of the aqueous PPG−salt two-phase system will cause an increase of the concentration in the PPG-rich phase and a decrease of the concentration in the salt-rich phase. As mentioned above, the attraction between PPG and water will decrease with an increase in pH and therefore by increasing pH of the aqueous PPG−salt two-phase system water is driven from the PPG-rich phase to the salt-rich phase, so the PPG concentration of the PPG-rich phase increases, while the salt-rich phase will be somewhat more diluted (i.e., the salt concentration will be decreased), and the volume of the salt-rich phase increases at expense of the PPG-rich phase. The STL values are resulting from interactions between all ATPS forming components, which promote the partitioning of each of them between the phases. The phase separation of a mixture of aqueous salt and polymer systems becomes more favorable by the addition of the electrolyte until a saturation point. It is important to stress that this saturation does not mean physical saturation of the binding sites around the polymer, but a significant amount of electrolyte is left in solution, without interacting with polymer, destabilizing the system, and hence, leading to phase separation. The weakness of citrate or sulfate + PPG interaction might be attributed to the complete dissociation of the citrate and sulfate forming an anion that interacts strongly with water (ion−dipole interaction) and cationic species (ion− ion interaction). On the other hand, phosphate is not completely ionized, producing anions that interact with water and cationic species more weakly when compared to citrate and sulfate.22

tie-lines for the PPG + potassium phosphate ATPS decreased with increasing pH. Figures 4 and 6 show that the slope and the length of the equilibrium tie-lines for biphasic systems containing sodium citrate and magnesium sulfate increased with increasing pH. Also, by increasing pH the volume of salt-rich phase increases at the expense

Figure 4. Effect of pH on the equilibrium phase compositions and on the slope and length of tie lines for the PPG725 (1) + sodium citrate (2) + H2O (3) systems: ■, total composition; ●, bottom phase composition; ×, top phase composition.

■

CONCLUSIONS Liquid−liquid equilibrium data for the PPG725 (1) + sodium citrate (2) + H2O (3), PPG725 (1) + potassium phosphate (2) + H2O (3), and PPG725 (1) + magnesium sulfate (2) + H2O (3) systems have been determined experimentally at various pH values of 8.22, 7.05, 6.38, 5.5, 4.81, and 3.81; 9.5, 8.4, 7.3, 6.35, 5.3, 4.54, and 3.45; and 5.56, 4.05, 3, and 2.43, respectively. The experimental binodal data for all of investigated systems were satisfactorily correlated with the Merchuk equation. It was found that the two-phase area is expanded with increasing pH. It was also observed that with increasing pH the slope and length of equilibrium tie-lines for biphasic systems containing sodium citrate and magnesium sulfate increased and for the PPG + potassium phosphate ATPS decreased. An empirical model was developed to describe the coexistence curve of the PPG−salts ATPS's and pH dependence of its model parameters. The model reproduces the experimental data accurately.

Figure 5. Effect of pH on the equilibrium phase compositions and on the slope and length of tie lines for the PPG725 (1) + potassium phosphate (2) + H2O (3) systems: ■, total composition; ●, bottom phase composition; ×, top phase composition.

■

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected], [email protected]. Phone: +98-831-427 45 30; fax: +98-831-427 45 42. Funding

This work was supported in part by the research vice department of Razi University. Notes

The authors declare no competing financial interest.

Figure 6. Effect of pH on the equilibrium phase compositions and on the slope and length of tie lines for the PPG725 (1) + magnesium sulfate (2) + H2O (3) systems: ■, total composition; ●, bottom phase composition; ×, top phase composition.

■

REFERENCES

(1) Albertsson, P. A. Partitioning of cell particles and macromolecules, 3rd ed.; Wiley: New York, 1986. (2) Walter, H.; Brooks, D. E.; Fisher, D. Partitioning in aqueous two phase systems; Academic Press: New York, 1985.

of the PPG-rich phase, as we observed experimentally. This is because the compositions of the equilibrium phases change with 1873

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874

Journal of Chemical & Engineering Data

Article

(3) Huddleston, J. G.; Willauer, H. D.; Boaz, K. R.; Rogers, R. B. Separation and recovery of food coloring dyes using aqueous biphasic extraction chromatographic resins. J. Chromatogr., B 1998, 711, 237− 244. (4) Hatti-Kaul, R. Aqueous Two Phase Systems: Methods and Protocols; Humana Press: Totowa, NJ, 2000. (5) Rogers, R. B.; Willauer, H. D.; Griffin, S. T.; Huddleston, J. G. Partitioning of small organic molecules in aqueous biphasic systems. J. Chromatogr., B 1998, 711, 255−263. (6) Cheluget, E. L.; Gelinas, S.; Vera, J. H.; Weber, M. E. Liquid-liquid Equilibrium of Aqueous Mixtures Poly(propylene glycol) with NaCl. J. Chem. Eng. Data 1994, 39, 127−130. (7) Zafarani-Moattar, M. T.; Salabat, A. Thermodynamics of Magnesium Sulfate-Polypropylene glycol Aqueous Two-Phase system. Experiment and Correlation. Fluid Phase Equilib. 1998, 152, 57−65. (8) Salabat, A.; Dashti, H. Phase Composition, Viscosities and Densities of Systems PPG425 + Na2SO4 + H2O and PPG425 + (NH4)2SO4 + H2O at 298.15 K. Fluid Phase Equilib. 2004, 216, 153−157. (9) Zafarani-Moattar, M. T.; Sadeghi, R. Phase Diagram Data for Several PPG + Salt Aqueous Biphasic Systems at 25 °C. J. Chem. Eng. Data 2005, 50, 947−950. (10) Zafarani-Moattar, M. T.; Emamian, S.; Hamzehzadeh, S. Effect of Temperature on the Phase Equilibrium of the Aqueous Two-Phase Poly(propylene glycol) + Tripotassium Citrate System. J. Chem. Eng. Data 2008, 53, 456−461. (11) Rahimpour, F.; Baharvand, A. Phase equilibrium in aqueous twophase systems containing poly (propylene glycol)425 and sodium citrate at different pH. International Conference on Biotechnology and Bioengineering (ICBB2009), Bali, Indonesia, Nov 25−27, 2009. (12) Merchuk, J. C.; Andrews, B. A.; Asenjo, J. A. Aqueous Two-Phase Systems for Protein Separation: Studies on Phase Inversion. J. Chromatogr., B 1998, 711, 285−293. (13) Huddleston, J. G.; Willauer, H. D.; Rogers, R. D. Phase Diagram Data for Several PEG + Salt Aqueous Biphasic Systems at 25°C. J. Chem. Eng. Data 2003, 48, 1230−1236. (14) Zafarani-Moattar, M. T.; Tolouei, S. Liquid−liquid equilibria of aqueous two-phase systems containing polyethylene glycol 4000 and dipotassium tartrate, potassium sodium tartrate, or di-potassium oxalate: Experiment and correlation. CALPHAD: Comput. Coupling Phase Diagrams Thermochem. 2008, 32, 655−660. (15) Ferreira, L. A.; Teixeira, J. A. Salt Effect on the Aqueous TwoPhase System PEG 8000-Sodium Sulfate. J. Chem. Eng. Data 2011, 56, 133−137. (16) Zhao, X.; Xie, X.; Yan, Y. Liquid−liquid equilibrium of aqueous two-phase systems containing poly(propylene glycol) and salt ((NH4)2SO4, MgSO4, KCl, and KAc): experiment and correlation. Thermochim. Acta 2011, 516, 46−51. (17) Silvério, S. C.; Rodríguez, O.; Teixeira, J. A.; Macedo, E. A. LiquidLiquid Equilibria of UCON + (Sodium or Potassium) Phosphate Salt Aqueous Two-Phase Systems at 23 °C. J. Chem. Eng. Data 2010, 55, 1285−1288. (18) Sadeghi, R.; Jamehbozorg, B. The salting-out effect and phase separation in aqueous solutions of sodium phosphate salts and poly(propylene glycol). Fluid Phase Equilib. 2009, 280, 68−75. (19) Zafarani-Moattar, M. T.; Hamzehzadeh, S. Effect of pH on the phase separation in the ternary aqueous system containing the hydrophilic ionic liquid 1-butyl-3-methylimidazolium bromide and the kosmotropic salt potassium citrate at T = 298.15 K. Fluid Phase Equilib. 2011, 304, 110−120. (20) Gisela Tubio, G.; Bibiana, B.; Nerli, B. B.; Pićo, G. A.; Ven̂ancio, A.; Teixeira, J. Liquid−liquid equilibrium of the Ucon 50-HB5100/sodium citrate aqueous two-phase systems. Sep. Purif. Technol. 2009, 65, 3−8. (21) Sadeghi, R.; Golabiazar, R. Thermodynamics of Phase Equilibria of Aqueous Poly(ethylene glycol) + Sodium Tungstate Two-Phase Systems. J. Chem. Eng. Data 2010, 55, 74−79. (22) Da Silva, L. H. M.; Loh, W. Calorimetric investigation of the formation of aqueous two-phase systems in ternary mixtures of water, poly(ethylene oxide) and electrolytes (or dextran). J. Phys. Chem. B 2000, 104, 10069−10073. 1874

dx.doi.org/10.1021/je300266r | J. Chem. Eng. Data 2012, 57, 1867−1874