construction of aqueous two-phase systems of the cyclopolymer of (diallylamino)propylphosphonate and...

Construction of Aqueous Two-Phase Systems of the Cyclopolymer of(Diallylamino)propylphosphonate and Its Sulfur Dioxide Copolymerwith Polyoxyethylene Using 1H NMR SpectroscopyShaikh A. Ali,* Izzat W. Kazi, and Nisar Ullah

Chemistry Department, King Fahd University of Petroleum and Minerals, Dhahran 31261, Saudi Arabia

ABSTRACT: A pH-responsive polyzwitterionic acid [PZA)],poly[3-(N,N-diallylammonio)propanephosphonic acid], and analternate copolymer PZA·SO2, poly[3-(N,N-diallylammonio)-propanephosphonic acid-alt-SO2] have been prepared viacypolymerization. Aqueous two phase systems (ATPSs) ofpolyoxyethylene (POE)−(NaOH-treated PZA or PZA·SO2)−water (NaCl) have been investigated. To construct the tielines, the compositions of the phases were determined by 1HNMR spectroscopy. The addition of 1, 1.5, and 2 equiv NaOHtransform the (±) zwitterionic polymers to their corresponding (±−) zwitterionic−anionic, (±−)/(=) zwitterionic−anionic/dianionic, and (=) dianionic polyelectrolyte. The influence of NaOH and salt (NaCl) on the binodals have been examined.Increasing the amount of added NaOH pushes the binodals toward lower concentrations, thereby requiring a lesser amount ofpolymers for phase separation. The NH+ and the OH functional groups in (±) PZA and (±) PZA·SO2 provide the requiredmotifs for the base-induced change of the charge densities in the macromolecule, whose size and shape can thus be manipulatedto facilitate potential separation of proteins in the ATPSs. A combined total mass fraction of less than 0.10 for the componentpolymers makes these ATPSs viable for industrial applications. The water-insolubility of PZA·SO2 at lower pH values makes it asuitable component for recycling ATPS.

1. INTRODUCTION

Since its introduction in the 1950s, liquid−liquid extraction(LLE) in aqueous two-phase systems (ATPSs)1 has become anattractive downstream biotechnology process for efficientseparation and purification of high value biomolecules suchas proteins,2−5 nucleic acid,6 antibiotics,7 nanomaterials,8 andbiopharmaceuticals,9 etc. The combination of appropriateconcentrations of two hydrophilic polymers or a polymer andsalt or other alternative components including green ionicliquids10 is used to construct biphasic systems known as ATPSs,which have favorable low interfacial tension and can be readilyscaled-up. The high water content makes it economical,environmentally friendly, benign to biomaterials, and biocom-patible. The binodal and tie-line data of ATPSs are exhibited asphase diagrams, where the binodal curves are obtained by aturbidimetric titrationmethod11,12 while tie-lines can be achievedfrom different concentration detection methods (such asrefractive index measurements,13 spectrophotometry,11 atomicabsorption spectrometry,12 NMR spectroscopy,14,15 etc.).A green polymer−organic salt ATPS has been developed for

bioseparation to avoid environmental problems associated withpolymer−inorganic salt ATPSs.16 Another kind of polymer−surfactant ATPS utilizes the influence of the hydrophobicdomains formed by the surfactant molecules for effectiveseparation of a variety of biomolecules.17 ATPSs formingpolymers have been modified with hydrophobic groups18 andbiospecific ligands for the affinity partitioning of biomolecules.19

ATPSs containing polymers with a pH-responsive aminoacetate

ligand have been developed for separation of some proteins.20

Recently, various ATPSs containing hydrophilic organic solventshave been employed to separate various natural products.21 Notethat in nonbiotechnology areas like industrial waste remediation,ATPSs have been utilized for the removal of coloring dyes,22

metal ions,23 and organic pollutants.24

The effective and environmentally friendly use of ATPSsrequires a way to recycling them after the back-extraction of thebiomacromolecules.25 The most commonly used polymercomponent in many an ATPS is dextran which is quite expensiveand biodegradable,26 thereby imposing some limitations on itsuse. While the use of nonionic polymers as components ofATPSs is well documented, the application of ionic polymers isscarce.14 For environmental reasons, we are in search of ATPSscontaining pH-responsive zwitterionic polymers that can beeasily recycled at their isoelectric points. For this work, we havechosen biomimicking polyzwitterion acid (PZA) 1 (Scheme 1)and PZA·SO2 5 (Scheme 2) for the construction of ATPSscontaining poly(oxyethylene) (POE) as the second polymercomponent. To our knowledge, the only other polyamino-phosphonate used in an ATPS is a counterpart of 5 havinga single methylene group separating the zwitterions.14 Theeffect of pH-induced change of the (±) zwitterionic polymersto (±−) zwitterionic−anionic polyelectrolyte (ZAPE), (±=)

Received: August 18, 2014Accepted: October 16, 2014

Article

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pubs.acs.org/jced

zwitterionic-anionic/dianionic polyelectrolyte (ZAPE/DAPE),and (=) dianionic polyelectrolyte (DAPE) on the binodal curveand tie-lines has been studied using the turbidimetric titration/dilution method and NMR spectroscopy. The consistency of thetie-lines has been checked using a correlation model.27

2. EXPERIMENTAL SECTION2.1. Materials. Table 1 includes the description of the

materials used for the current work. The pH responsive PZA 1and PZA·SO2 5 have been prepared as described (Schemes 1and 2).28,29 POE has a Mn of 35.0 kg·mol−1. DAPE 4 has anapproximate MW of 27.3 kg·mol−1 and a polydispersity index of2.2, while the corresponding values for DAPE·SO2 8 were foundto be 65.5 kg·mol−1 and 2.3, respectively.28,29 Table 2 sum-marizes the intrinsic viscosities of ZAPE 3, DAPE 4, ZAPE·SO25, and DAPE·SO2 8 in 0.1 NaCl.2.2. Stock Solutions.The stock solutions of PZA 1, PZA·SO2

5, and POE of polymer mass fraction (w) of 0.20 were made in

mNaCl of 0.3 or 0.6 mol·kg−1. The stock solutions of PZA 1 and

PZA·SO2 5 were obtained after treating the polymer/watermixture with 1, 1.5, or 2 equiv NaOH so as to give thecorresponding ZAPE (2 or 6), ZAPE/DAPE (3 or 7), and DAPE(4 or 8), respectively.A Precisa balance with an accuracy of 10−4 g was used to

prepare solutions. The temperature in phase equilibration has anuncertainty of 0.2 K. A Sartorius pH meter PB 11 was used tomeasure the pH of the solutions. The 1H NMR spectra of thepolymers have been measured in D2O (using HOD signal atδ = 4.65 ppm as internal standard) on a JEOL LA 500 MHzspectrometer. Polymer compositions were obtained by carefulintegration of appropriate 1HNMR signals with an uncertainty of1.89% (vide inf ra).

2.3. Phase Compositions of PZA 1 (or PZA·SO2 5)−POE−H2O (NaCl) Systems. 2.3.1. The Tie-Lines by 1H NMRSpectroscopy. The stock solutions of PZA 1 (treated with 1, 1.5,and 2.0 equiv NaOH) and POE having w of 0.20 were made inmNaCl of 0.6 mol·kg

−1. Several known total-systems (∼7 cm3) ofcompositions (Atotal) were made in calibrated cylinders (10 mLwith an uncertainty of 0.1 mL) (Tables 3 to 8). After thoroughmixing, themixtures were centrifuged to ensure a complete phaseseparation. The phases were equilibrated (296.0 K, 24 h), and thevolumes of the phases were measured. After obtaining the massof a solution in a volumetric pipet (1.00 mL with an uncertaintyof 0.01 mL), the densities of the top and bottom layers weredetermined to be∼1020 kg·m−3 and∼1045 kg·m−3, respectively.After removal of H2O from a small portion of each layer, the 1HNMR spectra were taken in D2O. Note that the polymersolutions in D2O (especially the ones treated with 1 and 1.5 equivNaOH) were treated with K2CO3 to avoid any overlap of theproton signals of the DAPEs (4 or 8) with that of POE. Thetreatment with K2CO3 makes the positive nitrogens neutralthereby causing an upfield shift of the proton signals from thepeak of POE at δ = 3.6 ppm. Figure 1 panels a and b show that the1H NMR spectra of bottom and top layers of system 1 (Table 5)are overwhelmingly rich in DAPE 4 (derived from PZA 1) andPOE, respectively.The area (A) of the 1H NMR signals appearing in the range

0.5≤ δ≥ 3.2 ppm belonged to the 16 protons of each repeat unitof DAPE 4, while the area (B) under the singlet at δ = 3.6 ppmaccounted for the four protons of POE. The area ratio of single Hof POE and PZA gave their mole ratio nPOE/nPZA (eq 1).

=nn

BA

/4/16

POE

PZA (1)

[PZAb] describing the polymer in the bottom phase in termsof mol·m−3 is calculated using eq 2.30

Scheme 1. Reaction of Homopolymers with NaOH

Scheme 2. Reaction of Copolymers with NaOH

Table 1. Purities of Materials

chemical supplier puritya

sodium chloride JT Baker > 0.995sodium hydroxide Fluka > 0.98water deionized waterPOEb MERCK-SchuchardtPZA 1c ref 41c > 0.99c

PZA·SO2 5d ref 42d > 0.99d

aPurities refer to the mass fraction. bPoly(oxyethylene) of molar mass35.0 kg mol−1. cSynthesized as described in ref 28 and supported byelemental with an uncertainty of 0.3%. dSynthesized as described in ref29 and supported by elemental analysis with an uncertainty of 0.3%.

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=−

−M M n n

V n n n n[PZA ]

POE / (PZA / )( / ){( / ) ( / ) }b

o POE o PZA POE PZA t

b POE PZA b POE PZA t(2)

where top and bottom phases are described by subscript t and b,respectively, while PZAo and POEo represent the mass inkilograms of PZA 1 and POE, respectively, while V stands forvolume in m3. The molar mass of the repeat unit of PZA 1 andPOE, as described by MPZA and MPOE, are taken as 0.2192 and0.04405 kg·mol−1, respectively. The mole ratio nPOE/nPZA isobtained by analysis of the 1H NMR integration. PZAbrepresenting the mass of the polymer in kilograms in the bottomphase was then calculated using eq 3.30

= ×V MPZA [PZA ]b b b PZA (3)

Then the rest of the polymer concentration in the top phase iscalculated from various other known quantities.30 After deter-mining the weight fraction (w) in each phase (Tables 3−5), thetie lines (Figure 2) were constructed.

Likewise, the experiments were carried out for PZA·SO2 5using the molar mass of the repeat unit as (i.e., 0.2833 kg·mol−1)in eq 2. Figure 1 panels c and d show that the 1H NMR spectraof the bottom and top layers of system 3 (Table 8) were found tobe rich in PZA·SO2 5 and POE, respectively. The area (A) underthe four-proton signals for 6 or 7 or 8 in the range δ 1.1 to 1.6 andthe area (B) for the four-proton singlet at δ 3.6 ppm attributed tothe POE gave the molar ratio [POE]/[PZA·SO2] as B/A. Themass of polymer PZA·SO2 5 in the bottom phase is calculatedusing eqs 2 and 3. The tie lines (Figure 3) were constructed asbefore (vide supra).For the purpose of validation of the molar ratios obtained from

NMR measurements, a total of eight mixtures each of PZA1/POE and PZA.SO2 5/POE with known mol ratios of 30:1,20:1, 10:1, 5:1, 1:5, 1:10, 1:20, and 1:30 in the presence of 2 equivNaOH, were prepared using the Precisa balance. The mol ratioswere then determined experimentally by 1H NMR integrationsas discussed above. The measured ratios have an uncertainty of1.89 % (with 95 percentile lower and upper values of 1.29 % and2.48 %, respectively) of the known ratios, and are empiricallycorrelated as measured ratio =−0.024966 (±0.184) + 1.0039415

Table 2. Intrinsic Viscosity ([η]) and Solubility Behaviours of Ionic Polymers

solubilitya

salt-addedc HCl-added [η]b/m3 kg−1

polymer salt-free water mNaCl/mol kg−1 mHCl/mol kg−1 mNaCl/mol kg−1c

PZA 1 − + (0.03) + (0.1) NDd

ZAPE 2 + + NA 0.0117 (0.1)DAPE 4 + + NA 0.0129 (0.1)PZA·SO2 5 − − + (0.8) NDd

ZAPE·SO2 6 + + NA 0.0640 (0.1)DAPE·SO2 8 + + NA 0.0621 (0.1)

a Symbols: +, soluble; −, insoluble. bDetermined by Huggins viscosity relationship using viscosity of 1 % to 0.125 % polymer solution in m/kg·mol−1

NaCl at 303 K (accurate to 0.1 K) as measured with an Ubbelohde viscometer (having viscometer constant of 0.005718 cSt/s at all temperatures);the flow time was measured using a digital stopwatch with an uncertainty of 0.01 s. cConcentration of NaCl is given in the parentheses. dNotdetermined.

Table 3. Mass Fraction (w) Compositionsa of the Phasesof the [POEb + PZAc] System (1.0 equiv NaOH, mNaCl of0.6 mol·kg−1) at 296.0 Kd Shown in Figure 2a

NMR method

total system top phase bottom phase

(100·w)/(w/w)

system POE PZA POE PZA POE PZA volume ratioe

1 4.62 4.63 8.96 0.867 0.435 8.25 0.972 4.20 4.20 7.93 1.03 0.546 7.28 0.953 4.73 3.00 6.83 1.23 0.678 6.48 1.94 3.43 3.32 5.65 1.33 0.887 5.64 1.1

dilution method

total system (two-phase) total system (one-phase)

(100·w)/(w/w)

system POE PZA POE PZA

a 6.34 3.45 3.69 2.01b 3.28 3.80 2.43 2.82c 3.35 5.87 1.95 3.41

aThe uncertainty in the determination of mol ratios required to calculate w inthe separated phases (using eqs 1 to 3) is considered to be within ± 1.89 %(as 95 % confidence). bPoly(oxyethylene) of molar mass 35.0 kg·mol−1.cPoly[hydrogen 3-(diallylammonio)propylphosphonate] 1. dStandarduncertainty u is u(T) = 0.2 K. eVolume ratio of top and bottom phaseas determined by a calibrated cyclinder with an uncertainty of 0.1 mL.

Figure 1. 1H NMR spectra in D2O of (a) bottom phase, system 1(Figure 2c, Table 5): PZA 1−POE; (b) top phase, system 1 (Figure 2c,Table 5): PZA 1−POE; (c) bottom phase, system 3 (Figure 3c, Table 8):PZA·SO2 5−POE; (d) top phase, system 3 (Figure 3c, Table 8).


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(±0.0096) × known ratio, where the values in the parenthesesrepresent the ranges for 95 percentile confidence intervals of theparameters.2.3.3. Binodals by Turbidity Method.The binodal data for the

PZA·SO2 5/POE systems (Figure 3) were obtained by theturbidimetric method at 296 K using a procedure describedelsewhere.15

2.3.4. Binodals by Dilution Method. For the PZA 1/POEsystems, it was difficult to visualize a clear-cut phase transitionfrom a turbid mixture to homogeneous solution duringattempted turbidimetric titration. In this case, the binodalswere constructed by the dilution method. Thus, NaCl solutionwas added in portions to several two-phase systems with knowncompositions (Atotal), and after centrifugation more NaClsolution is added dropwise until the systems become homo-geneous; the compositions before the last drop added gave thebinodal points (Figure 2).

3. RESULTS AND DISCUSSION

3.1. Solution Properties. Both PZA 1 and PZA·SO2 5 arepH responsive polymers, which under the influence of pH can beconverted to ZAPE and DAPE polymers (Schemes 1 and 2). Inthe presence of 1, 1.5, and 2 equiv NaOH, PZA 1 is expected toremain in the dominant form of (±−) ZAPE 2, (ZAPE/DAPE)3, and (=) DAPE 4, respectively, while for the PZA·SO2 5 thecorresponding forms would be (±−) ZAPE·SO2 6, (ZAPE/DAPE)·SO2 7, and DAPE·SO2 8 as described in Schemes 1 and2. The intrinsic viscosities of some of the ionic polymers arerecorded in Table 2.

For the (PZA 1-POE) ATPSs, the pH of the top and bottomphases (written, respectively, under parentheses) were measuredto be as follows: 1 equiv NaOH, Table 3, system 2, (7.09 and7.12); 1.5 equiv NaOH, Table 4, system 2, (10.20 and 10.31); 2equiv NaOH, Table 5, system 1, (11.43 and 11.61). The pH forthe corresponding (PZA·SO2 5-POE) ATPSs were 1 equivNaOH, Table 6, system 1, (6.97 and 7.08); 1.5 equiv NaOH,Table 7, system 3, (8.96 and 9.14); 2 equiv NaOH, Table 8,system 3, (10.19 and 10.40). Note that the polyaminophoph-onate-rich bottom phases have slightly higher pH values than thePOE-rich top phases because the basic aminophophonate motifson hydrolysis generate OH− ions. The presence of electron-withdrawing SO2 in the copolymers weakens the basicity of theaminophophonate and thus lowers the pH values as presentedabove (cf. Tables 6−8 vs 3−5).pH-induced changes in the backbone charge types [(±) to

(±−) to (=)] and their densities are shown in the upper halfof Scheme 3. Cationic (+) and anionic (−) polyelectrolytesare soluble in water while PZA 1 is insoluble in salt-free waterbut soluble in mNaCl of 0.03 mol·kg

−1 or in mHCl of 0.1 mol·kg−1.

Its corresponding water-soluble (±−) ZAPE 2 and (=) DAPE4 were found to have intrinsic viscosities of 0.0117 and0.0129 m3/kg in mNaCl of 0.1 mol·kg−1 at 30 °C, respectively.DAPE 4 has an approximate MW of 27 300 g mol−1 and apolydispersity index of 2.2.28 Note that the solubility of PZA 1in HCl is a result of transformation of the zwitterionic (±) motifto cationic (+) motif upon protonation of the negatively chargedoxygen.

Figure 2. Phase diagram [■ and□ represent mass fraction (w) data obtained by respective NMR and dilution method] at 296 K ofmNaCl of 0.6 mol·kg−1

containing (a) PZA 1 (treated with 1 equiv NaOH)−POE; (b) PZA 1 (treated with 1.5 equiv NaOH)−POE; (c) PZA 1 (treated with 2 equiv NaOH)−POE; and (d) phase diagram of PZA 1 (treated with■, 1 equiv;□, 1.5 equiv; and▲, 2.0 equiv NaOH)−POE−water (mNaCl of 0.6 mol·kg

−1) at 296 K.


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(±) PZA·SO2 5 remained insoluble in salt-free water.However, it became soluble in mHCl of 0.8 mol·kg−1 as a result

of conversion of the zwitterionic motif to a cationic motif uponprotonation of O−. Even though polyzwitterions are known to be

Figure 3. Phase diagram [■ and □ represent mass fraction (w) data obtained by respective NMR and turbidimetric method] at 296 K of mNaCl of 0.3mol·kg−1 containing (a) PZA·SO2 5 (treated with 1 equiv NaOH)−POE; (b) PZA·SO2 5 (treated with 1.5 equiv NaOH)−POE; (c) PZA·SO2 5(treated with 2 equiv NaOH)−POE; and (d) phase diagram of PZA·SO2 5 (treated with ■, 1 equiv; □, 1.5 equiv; and ▲, 2.0 equiv NaOH)−POE−water (mNaCl of 0.3 mol·kg

−1) at 296 K.

Table 4. Mass Fraction (w) Compositiona of the Phasesof the [POEb + PZAc] System (1.5 equiv NaOH, mNaCl of0.6 mol·kg−1) at 296.0 Kd Shown in Figure 2b

NMR method


(100·w)/(w/w)


1 4.70 4.53 8.48 0.254 0.111 9.73 1.22 4.06 4.08 7.37 0.312 0.188 8.43 1.23 3.53 3.53 5.61 0.890 0.466 7.38 1.54 3.00 3.13 4.67 1.02 0.580 6.35 1.6

dilution method


(100·w)/(w/w)


a 4.12 3.45 2.42 2.03b 3.02 4.87 1.79 2.89c 2.85 6.01 1.58 3.34

aThe uncertainty in the determination of mol ratios required to calculate w inthe separated phases (using eqs 1 to 3) is considered to be within ± 1.89 %(as 95 % confidence). bPoly(oxyethylene) of molar mass 35.0 kg mol−1.cPoly[hydrogen 3-(diallylammonio)propylphosphonate] 1. dStandarduncertainty u is u(T) = 0.2 K. eVolume ratio of top and bottom phaseas determined by a calibrated cyclinder with an uncertainty of 0.1 mL.

Table 5. Mass Fraction (w) Compositiona of the Phasesof the [POEb + PZAc] System (2 equiv NaOH, mNaCl of0.6 mol·kg−1) at 296.0 Kd Shown in Figure 2c

NMR method


(100·w)/(w/w)


1 4.33 4.26 7.77 0.372 0.448 8.62 1.12 3.65 3.76 6.43 0.522 0.483 7.37 1.13 2.94 3.01 5.05 0.873 0.499 5.50 1.14 2.51 2.56 4.01 1.11 0.551 4.49 1.3

dilution method


(100·w)/(w/w)


a 4.12 3.45 2.42 2.03b 3.02 4.87 1.79 2.89c 2.85 6.01 1.58 3.34

aThe uncertainty in the determination of mol ratios required to calculate win the separated phases (using eqs 1 to 3) is considered to be within 1.89 %(as 95 % confidence). Composition is defined in terms of weight fraction w.bPoly(oxyethylene) of molar mass 35.0 kg mol−1. cPoly[hydrogen3-(diallylammonio)propylphosphonate] 1. dStandard uncertainty u isu(T) = 0.2 K. eVolume ratio of top and bottom phase as determinedby a calibrated cyclinder with an uncertainty of 0.1 mL.


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soluble in the presence of added salts, a sample of aheterogeneous mixture of PZA·SO2 5 having a mass fraction(w) of 0.01 remained insoluble in any concentration of NaCl,KCl, KBr, KI, and CaCl2.

29 Polyzwitterions are usually insoluble

in salt-free water,31,32 but soluble in the presence of lowmolecular weight added salts (e.g., NaCl). The addition of a smallamounts of electrolytes (NaCl, etc.) to linear polyelectrolytesleads to effective shielding of the backbone charges by chloride

Table 6. Mass Fraction (w) Compositiona of the Phasesof the [POEb + PZA·SO2

c] System (1.0 equiv NaOH, mNaClof 0.3 mol·kg−1) at 296.0 Kd Shown in Figure 3a

NMR method


(100·w)/(w/w)

system POE PZA·SO2 POEPZA·SO2 POE

PZA·SO2

volumeratioe

1 4.45 4.87 8.81 0.383 0.183 9.80 1.062 4.05 4.13 7.40 0.456 0.307 8.57 1.193 3.71 3.70 6.00 0.803 0.685 7.46 1.334 3.11 3.13 4.78 0.854 0.840 6.00 1.33

turbidity method

binodal data

(100·w)/(w/w) (100·w)/(w/w)

system POE PZA·SO2 system POE PZA·SO2

a 0.501 7.03 g 3.58 1.36b 0.832 5.59 h 4.11 1.01c 1.34 4.42 i 4.69 0.720d 1.86 3.53 j 5.66 0.507e 2.33 2.81 k 7.04 0.326f 2.86 2.24 l 10.1 0.219

aThe uncertainty in the determination of mol ratios required tocalculate w in the separated phases (using eqs 1 to 3) is considered tobe within 1.89 % (as 95 % confidence). bPoly(oxyethylene) of molarmass 35.0 kg mol−1. cPoly[hydrogen 3-(diallylammonio)propyl-phosphonate-alt-sulfur dioxide] 5. dStandard uncertainties u is u(T) =0.2 K. eVolume ratio of top and bottom phase as determined by acalibrated cyclinder with an uncertainty of 0.1 mL.

Table 7. Mass Fraction (w) Compositiona of the Phases of the [POEb + PZA·SO2c] System (1.5 equiv NaOH, mNaCl of

0.3 mol·kg−1) at 296.0 Kd Shown in Figure 3b

NMR method


(100·w)/(w/w)

system POE PZA·SO2 POE PZA·SO2 POE PZA·SO2 volume ratioe

1 4.62 4.62 9.79 0.246 0.111 8.95 0.9442 4.10 4.10 7.94 0.305 0.181 8.33 1.093 3.67 3.57 7.10 0.430 0.410 6.76 1.004 3.12 3.12 5.89 0.470 0.561 5.72 0.971

turbidity method

binodal data

(100·w)/(w/w) (100·w)/(w/w)


a 0.251 7.56 g 2.67 2.11b 0.619 5.51 h 3.54 1.45c 1.03 4.39 i 4.21 1.12d 1.46 3.62 j 6.10 0.47e 1.87 3.08 k 8.72 0.233f 2.25 2.59

aThe uncertainty in the determination of mol ratios required to calculate w in the separated phases (using eqs 1 to 3) is considered to be within 1.89 %(as 95 % confidence). bPoly(oxyethylene) of molar mass 35.0 kg mol−1. cPoly[hydrogen 3-(diallylammonio)propylphosphonate-alt-sulfur dioxide] 5.dStandard uncertainties u is u(T) = 0.2 K. eVolume ratio of top and bottom phase as determined by a calibrated cyclinder with an uncertainty of 0.1 mL.

Table 8. Mass Fraction (w) Compositiona of the Phasesof the [POEb + PZA·SO2

c] System (2.0 equiv NaOH, mNaClof 0.3 mol·kg−1) at 296 Kd Shown in Figure 3c

NMR method


(100·w)/(w/w)

system POE PZA·SO2 POEPZA·SO2 POE

PZA·SO2

volumeratioe

1 4.65 4.65 10.7 0.137 0.0910 8.36 0.8002 4.22 4.10 9.37 0.132 0.172 7.38 0.8213 3.75 3.72 8.04 0.323 0.192 6.87 0.8894 3.01 3.02 5.90 0.366 0.472 5.42 0.917

turbidity method

binodal data

(100·w)/(w/w) (100·w)/(w/w)


a 0.251 6.01 g 3.03 1.32b 0.652 4.39 h 3.69 0.821c 1.11 3.36 i 4.33 0.612d 1.73 2.66 j 4.98 0.460e 2.17 2.15 k 6.53 0.313f 2.55 1.76 l 7.38 0.242

aThe uncertainty in the determination of mol ratios required tocalculate w in the separated phases (using eqs 1 to 3) is considered tobe ur (w) = 1.89 % (as 95 % confidence). bPoly(oxyethylene) of molarmass 35.0 kg mol−1. cPoly[hydrogen 3-(diallylammonio)propyl-phosphonate-alt-sulfur dioxide] 5. dStandard uncertainty u is u(T) =0.2 K. eVolume ratio of top and bottom phase as determined by acalibrated cyclinder with an uncertainty of 0.1 mL.


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ions, thereby inducing the polymer chain to adopt a morecompact conformation (polyelectrolyte effect).33 The poly-zwitterions or polyampholytes, however, exhibit the oppositebehavior where intragroup and intra- and interchain associa-tions34 involving charges of opposite algebraic signs lead toglobular as well as network conformations. These zwitterionic/ampholytic interactions are disrupted by small electrolytes; theefficacy of Cl− ion in neutralizing the cationic charges ismuch higher than the shielding of O− by Na+. As a result, eachzwitterionic dipole inherits an excess negative charge, and therepulsion among the dipoles induces chain expansion leading to aglobule-to-coil transition (antipolyelectrolyte effect) (inside thebox in Scheme 3).35 It is surprising to note that even the readilypolarizable iodide anions were ineffective in neutralizing thecationic charges so as to disrupt the zwitterionic interactionsand thus failed to promote water-solubility of PZA·SO2 5. Itscorresponding water-soluble (±−) ZAPE·SO2 6 and (=) DAPE·SO2 8 were found to have intrinsic viscosities of 0.0640 and0.0621 mNaCl of 0.1 mol·kg−1 at 30 °C, respectively. DAPE 6 hasan approximate MW of 65 500 g mol−1 and a polydispersity indexof 2.3.Water-soluble 7 and 8 have the important advantage that these

can be recycled by precipitation at lower pH as water-insolublePZA·SO2 5. The relationship between the solubility behavior ofthe polymers and the hydrodynamic volume of a macromolecularchain with or without charge symmetry can be rationalized usingeq 4.36

πκ

πκ

* = − +Δ

vfI I f( ) 4B

2

S

B2

S2

(4)

where IB represents Bjerrum length, f and Δf describe the totalfraction of charge of a repeat unit and imbalance in the charge,respectively, whereas κS stands for the Debye−Huckel screeningparameter. A negative value for the electrostatic excluded volumev* affirms contraction, while a positive value ascertains chain

expansion. For (±) PZA 1 or PZA·SO2 5 being electroneutral, v*becomes negative as the second term of eq 4 vanishes as aconsequence of Δf = 0. This leads to a collapsed polymer chainand insolubility in salt-free water (Scheme 3). The water-solubility of (±−) ZAPE 2, (±−) ZAPE·SO2 6, (±=) DAPE 4and (±=) DAPE·SO2 8 is a consequence of expanded polymerchains where the increased charge imbalance Δf raises theimportance of the second term in eq 4, thereby leading to a lessnegative or even a positive value for the v*.

3.2. Phase Diagrams Using [PZA 1 or PZA·SO2 5 +xNaOH]−POE−H2O (NaCl) Systems. Several systems ofknown composition of POE and PZA 1 or PZA·SO2 5 (treatedwith x equiv NaOH) were prepared (Tables 3 to 8). In thepresence of 1, 1.5, and 2 equiv NaOH, PZA 1 was transformed toZAPE 2 (Table 3), ZAPE/DAPE 3 (Table 4), and DAPE 4(Table 5), respectively, while PZA·SO2 5 was converted intoZAPE 6 (Table 6), ZAPE/DAPE 7 (Table 7), and DAPE 8(Table 8). Equilibrium polymer concentrations in the phases,determined by 1H NMR spectroscopy, are connected by tie line(Figures 2 and 3). The composition of each total system, top andbottom phases, are described as Atotal, Atop, and Abottom,respectively. While the binodals for the systems containingPZA·SO2 5 were constructed using data (marked by unshadedsquares□) from the turbidimetric method (Figure 3a−c), it wasdifficult to visualize a clear phase transition (i.e., the appearanceor disappearance of turbidity) with systems having PZA 1.Therefore, a part of the binodals for PZA 1-derived polymerswere constructed by the dilution method which caused the threesystems, indicated by the unshaded squares (□), to change viathe dashed lines from heterogeneous (two-phase) to homoge-neous (one-phase) systems (Figure 2a−c) (see ExperimentalSection). The binodal curves were then constructed usingcomposition of Atop and Abottom of several systems marked asshaded squares (■) along with compositions obtained by thedilution methods as indicated by the three unshaded squares(□) (Figures 2a−c). The top and bottom layers were found tobe rich in POE and the ionic polymers, respectively.In all the phase diagrams, w of POE (rich in top phase) is

assigned to the y-axis. The ratio of the tie line lengths of Atotal−Abot and Atotal−Atop represents the volume ratio of the two phases(Tables 3 to 8). The binodals are fairly symmetrical and confirmthat the polymers segregate during phase separation (Figures 2and 3).1 A total w of < 0.10 required for phase separation is awelcome situation for industrial application.Figures 2d and 3d show the effect of increasing NaOH on the

binodals, where the shifts toward lower concentration ofpolymers are attributed to the greater negative charge densityon the chains. The increased electrostatic repulsion23−25 leads toa transition from a compact to an expanded coil as depicted inScheme 3. Noteworthy is the (=) DAPE 4 or 8 (∼ 100% negativecharge), DAPE/ZAPE (=)/(±−) 3 or 7, and ZAPE (±−) 2 or 6with a respective Δf of 1.0, 0.60, and 0.33 favoring the negativecharges. As a result, lesser amounts of polymers are required forphase separation. In the absence of NaCl, no phase separa-tion occurred in the concentration ranges used in this work.H-bonding among the neutral POE, water, and the negativeoxygens of the ionic polymers leads to an association between thepolymers thus preventing phase separation. In the presenceof NaCl, the H-bonding structure is broken because water wouldbe energetically more favorable to hydrate the Na+ ions therebyforcing relatively more hydrophobic POE to opt for com-partmentalization i.e. phase separation.37,38 The Na+ ions wouldalso neutralize the negative charges on the oxygens. The highest

Scheme 3. pH-Induced Change of Backbone Charges andZwitterionic Interaction


dx.doi.org/10.1021/je500767z | J. Chem. Eng. Data XXXX, XXX, XXX−XXXG

charge imbalance between the neutral POE and (=) DAPE 4or (=) 8 leads to the highest shift of the binodal toward lowerconcentration of the polymers.With the increase of mNaCl from 0.3 mol·kg−1 to 0.6 mol·kg−1

the shift of the binodal toward lower concentration of polymersindicates that lesser amounts of PZA·SO2 5/POE are needed for

phase separation (Figure 4a). At a higher mNaCl, the increasedcontraction of the ionic backbone makes it less compatible withthe POE. As mentioned earlier, the higher concentration of NaClalso forces the neutral POE to compartmentalize. For both thepolymer systems at the samemNaCl of 0.6 mol·kg

−1, the PZA·SO2

5/POE system required lesser polymer concentrations than the

Figure 4. (a) Phase diagram of POE−PZA·SO2 5 (treated with ■, 1 equiv;●, 1.5 equiv; and▲, 2.0 equiv NaOH) in mNaCl of 0.3 mol·kg−1 and POE−

PZA·SO2 5 (treated with□, 1 equiv;○ 1.5, equiv; and△, 2.0 equiv NaOH) inmNaCl of 0.6 mol·kg−1 at 296 K. (b) Phase diagram inmNaCl of 0.6 mol·kg

−1

of POE−PZA 1 (treated with■, 1 equiv;●, 1.5 equiv; and▲, 2.0 equiv NaOH) and POE−PZA·SO2 5 (treated with□, 1 equiv;○, 1.5 equiv; and△, 2.0equiv NaOH) at 296 K.

Figure 5. Correlation of phase diagram of PZA 1 (NaOH)−POE−H2O (mNaCl of 0.6 mol·kg−1) and PZA·SO2 5 (NaOH)−POE−H2O (mNaCl of 0.3

mol·kg−1) systems using the method of Diamond and Hsu, using the tie line data of (a) Figure 2a (Table 3), (b) Figure 2b (Table 4), (c) Figure 2c(Table 5), (d) Figure 3a (Table 6), (e) Figure 3b (Table 7), and (f) Figure 3c (Table 8), respectively. (The trendlines having zero intercept values aredrawn on each plot).


dx.doi.org/10.1021/je500767z | J. Chem. Eng. Data XXXX, XXX, XXX−XXXH

PZA 1/POE system for phase separation to occur (Figure 4b) asa result of a greater mismatch in the sizes of the polymercomponents in the former system.3.3. The Correlation of the Phase Diagrams. A Florey−

Huggins theory-based correlation developed by Diamond andHsu39 of the phase diagrams to check the consistency of the tie-linesof 2 or 3 or 4-POE−water (NaCl) systems is given by eqs 5 and 6

= ″ − ′K Aln (W W )1 1 1 1 (5)

and

= ″ − ′K Aln (W W )2 1 1 2 (6)

where the subscripted 1 and 2 represent polymer 1 (PZA 1 or PZA·SO2 5 + NaOH) and polymer 2 (POE). The polymer weightpercent in the top and bottom phase are denoted as w″ and w′,respectively. The polymer molar masses and their interactions withwater dictate themagnitude of the slopesA1 andA2, whileK1 andK2represent the partition coefficient (Ct/Cb) of the polymer. The verygood least linear square fits of the data in Figure 5 indicate that thephase behavior can be reasonably well described with the model byDiamond and Hsu. The simple model thus requires only a singlephase composition to calculate A1 and A2.Using eqs 5 and 6, a linear regression of ln K versus wi″ − wi′,

with zero intercept value afforded the parameters A1 and A2. Theroot mean-square deviation (rmsd) was calculated using the ex-perimentalKexp and the calculated partition constantsKcalc by eq 7:

40

=∑ −

−= K K

Nrmsd

( )

1iN

i1 exp calc2

(7)

where the parameters Ai and rmsd values of the correlation modelfor the partition coefficients using N number of tie lines (Table 9)

ascertain that the Diamond and Hsu eqs 5 and 6 are helpful incorrelating the experimental data of the current work.

4. CONCLUSIONSSeveral ATPSs were constructed using POE-pH-responsiveaminopolyphosphonates−water (NaCl). The behavior of thephase diagrams have been studied in terms of charge densities onthe polymer chain and mNaCl. The shift of the binodals towardlower concentration of polymers with increasing mNaCl leads tothe requirement of lesser concentration of the polymers for thephase separation. Almost zero solubility of PZA·SO2 5 in water in

the presence of any amount of added NaCl is the most gratifyingaspect for the use of 6, 7, and 8 since these polymers could beprecipitated and thus recycled at a lower pH value by theirconversions into PZA·SO2 5. The solubility behavior of thesebiomimicking polymers makes 6, 7, and 8 suitable forapplications involving bioseparations. Judicious choice of thecharge types and their densities on the polymer backbone isexpected to impart selectivity in the separation involvingbiomolecules like proteins. The charge imbalance in favor ofanionic fractions makes the polymers less compatible with POE.The ATPSs have great potential for the removal of toxic metalions.

■ AUTHOR INFORMATION

Corresponding Author*Tel.: +966-13-860-3830. Fax: +966-13-860-4277. E-mail:[email protected].

NotesThe authors declare no competing financial interest.The authors gratefully acknowledge King Abdulaziz City forScience and Technology (KACST) through the Science &Technology Unit at King Fahd University of Petroleum &Minerals (KFUPM) for funding this work through Project No.11-ADV2132-04 as part of the National Science, Technology andInnovation Plan.

■ ACKNOWLEDGMENTS

The authors are grateful for the facilities provided by KFUPM.

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Table 9. Values for Parameter Aa in eqs 5 and 6 along with thermsdb of the Model from the Experimental Data of thePartition Coefficients

ionic polymerc POEd

entry system A1a rmsd A2

a rmsd

1 POE + PZA + 1.0 equiv NaOH −0.314 0.0139 0.365 1.242 POE + PZA + 1.5 equiv NaOH −0.365 0.0382 0.511 2.833 POE + PZA + 2.0 equiv NaOH −0.388 0.0138 0.441 5.104 POE + PZA·SO2 + 1.0 equiv NaOH −0.352 0.0143 0.441 2.155 POE + PZA·SO2 + 1.5 equiv NaOH −0.425 0.0149 0.459 5.806 POE + PZA·SO2 + 2.0 equiv NaOH −0.505 0.00652 0.449 6.20

aAi is the slope of the linear regression of ln Ki versus wi″ − wi′, withzero intercept value (eqs 5 and 6). bRoot mean-square deviation. cPZA1 for entries 1 to 3 and PZA·SO2 5 for entries 4 to 6. dPoly-(oxyethylene) of molar mass 35.0 kg·mol−1.


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mailto:[email protected]

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