Download pdf - Separation of parent homopolymers from polystyrene-b-poly(ethylene oxide)-b-polystyrene triblock copolymers by means of liquid chromatography: 1. comparison of different methods

Separation of Parent Homopolymers fromPolystyrene‑b‑poly(ethylene oxide)‑b‑polystyrene TriblockCopolymers by Means of Liquid Chromatography: 1. Comparison ofDifferent MethodsMarion Rollet,*,† Berengere Pelletier,† Anaïs Altounian,† Dusan Berek,‡ Sebastien Maria,†

Emmanuel Beaudoin,† and Didier Gigmes*,†

†Aix-Marseille Universite, CNRS, Institut de Chimie Radicalaire, UMR7273, 13397 Marseille Cedex 20, France‡Polymer Institute of the Slovak Academy of Sciences, Dubravska Cesta 9, 84541 Bratislava, Slovakia

ABSTRACT: Separation of parent homopolymers, polystyreneand poly(ethylene oxide), from the triblock copolymerpolystyrene-b-poly(ethylene oxide)-b-polystyrene was investi-gated by means of liquid chromatography techniques. Overallsuitability was evaluated and compared for size exclusionchromatography, (SEC), liquid chromatography under criticalconditions of enthalpic interactions (LC CC), and liquidchromatography under limiting conditions of desorption (LCLCD). Among these techniques, LC LCD was the only one ableto fully separate block copolymers from both their parenthomopolymers in one single run. The efficiency of the separationwas proven by 1H NMR analysis of previously collected fractions.

Poly(ethylene oxide) (PEO) is a widely employed andstudied hydrophilic polymer. Besides its good biocompat-

ibility and biodegradability, interest in PEO is also related tothe fact that the polyether backbone is fairly inert to mostchemical reactions. These favorable chemical propertiesstimulated strong efforts to develop PEO-based blockcopolymers by means of various polymerization techniquesand macromolecular engineering strategies including couplingreaction or click chemistry approaches. Among the differentpossible compositions and architectures, amphiphilic diblockand triblock PEO copolymers represent a fascinating class ofpolymers. Indeed, thanks to their capacity to self-assemble insolution or bulk, these materials find a large range ofcommodity and/or specific applications. Typically, diblockand triblock PEO copolymers are particularly attractive assurfactants, stabilizers, compatibilizers,1,2 structuring agents formesoporous silica preparation but also for elaboration offiltration membranes3−5 and solid polymer electrolytes for Limetal/polymer batteries.6−11 Like most of amphiphilic blockcopolymers, diblock and triblock PEO copolymers propertiesare strongly affected by their molecular characteristics, such asmolar masses and dispersity,7 chemical structure (composi-tion), macromolecular architecture, functionality, as well as bytheir contamination by residual parent homopolymers.12,13 Tooptimize both the synthesis procedure and the macroscopicproperties of block copolymers, a comprehensive molecularcharacterization of polymer sample is highly needed but oftendifficult to obtain. For example, NMR, which is one of the mostwidely used molecular characterization tool for synthetic

polymers is limited to provide specific information such asend group functionalities or presence of residual parenthomopolymers. In this context, currently, three liquidchromatography techniques are mainly employed for molecularcharacterization of block copolymers, namely, size exclusionchromatography (SEC), gradient elution-liquid chromatogra-phy (GE-LC), and liquid chromatography under criticalconditions of enthalpic interactions (LC CC). Recently,another interesting technique so-called LC LCD has beendeveloped for efficient block copolymer characterization.SEC is the most common method for characterization of

synthetic and natural polymers. When employed correctly, itcan give valuable information on a polymer sample, such asmolar mass distribution and dispersity, chemical composition,or even branching distribution. Moreover, SEC determinationof exact molar mass and molar mass dispersity (distribution) isfast, simple, and relatively economic. It has to be mentionedthat SEC is often used improperly and even misused14,15

leading to strong errors on determined molar masses anddispersity. However, for block copolymers characterization,SEC exhibits some limitations. Indeed, because of both poordetector sensitivity and separation selectivity,16 usually parenthomopolymers are hardly separated from the block copolymersor are sometimes not even detected.

Received: December 16, 2013Accepted: January 27, 2014Published: January 27, 2014

Article

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© 2014 American Chemical Society 2694 dx.doi.org/10.1021/ac4040746 | Anal. Chem. 2014, 86, 2694−2702

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GE-LC is a widely used method to separate polymersaccording to their chemical compositions. Numerous mecha-nisms can rule the separation (adsorption, enthalpic partition,precipitation/redissolution...). Briefly, macromolecules areinjected into a low strength or quality eluent, leading toretention or precipitation of macromolecules near the columninlet. Eluent strength or quality is then increased during the runtime of the analysis. When it reaches a specific value, polymersare preferentially desorbed or dissolved and they will be elutedfrom the column packing dependent on molar mass and/orchemical composition.17

GE-LC can be used for the characterization of complexpolymer mixtures and the separation of block copolymers fromtheir parent homopolymers.18−22 Regarding block copolymers,Brun showed that statistical copolymers are eluted at theircritical adsorption point (CAP) in GE-LC, but blockcopolymers A-b-B elute at a mobile phase composition betweenthe CAP of homopolymer A and the CAP of homopolymer B,and when homopolymer B is more adsorptive thanhomopolymer A, the retention time of block copolymerincreases with the length of the B-block.19 So, despite the B-block length, the block copolymer could not be separated fromhomopolymer B. On top of that, it is an adsorption-basedelution mechanism technique, which could exhibit low samplerecovery.23−25

Moreover, GE-LC needs at least a binary pump. In thispaper, GE-LC will not be investigated because we want to focuson isocratic techniques because the apparatus is cheaper andmore widespread in laboratories.LC CC is a powerful method, which enables separation of

block copolymers according to the block length.26 In LC CC,exclusion and adsorption effects counterbalance each other anda linear homopolymer chain transported along the columnbecomes chromatographically “invisible”, which means that itselution becomes independent of the molar mass. Therefore, inthe case of a block copolymer, one block can be eluted undercritical conditions while another block elutes in the exclusionmode. In this way, one of the parent homopolymers can beefficiently separated from the block copolymer. The secondparent homopolymer elutes together with the block copolymer.Similarly, LC CC is efficient to separate oligomers according totheir functionality. This technique has been employed withvarious block copolymers and functional polymers.26−28

Recently, several authors reported successful LC CCprocedures to characterize PEO-b-PS diblock copolymer.Critical conditions for PEO29,30 have been established, whilePS eluted in the SEC mode. Conversely, at critical conditionsfor PS, the exclusion eluted block was PEO.31,32 The importantissue concerning critical conditions for PEO is that mostreported studies were carried out with a mobile phasecomposed of water and water miscible organic solvents suchas acetone, acetonitrile, methanol, etc. Such mixtures arerestricted to the PEO-based block copolymers, in which thesecond block exhibits similar solubility26,33,34 or its molar massis sufficiently low35 to avoid precipitation. Moreover, the highsensitivity of the critical conditions to both eluent compositionand temperature may result in the limited interlaboratoryreproducibility and reduced intralaboratory repeatability.36,37

Furthermore, the sample recovery for higher molar masses isoften insufficient24,38,39 leading to misinterpretation, especiallywhen hyphenated LC techniques are used. On top of that, twoindependent LC CC systems are needed to separate bothparent homopolymers from the corresponding block copoly-mer.Recently, a novel separation method was introduced, namely,

liquid chromatography under limiting condition of desorption,LC LCD.40−43 The strength of this method is to simultaneouslydiscriminate several distinct kind of polymers in one single run,including both parent homopolymers from the diblockcopolymers. The principle of LC LCD lies on the combinationof two different retention mechanisms within a column packedwith active, porous particles: (i) exclusion mechanism wheremacromolecules are eluted faster than low-molecular eluentcomponents and (ii) adsorption retention mechanism, which ispromoted by an appropriate low-molecular substance. Becauseof their adsorption, macromolecules may be retained within thestationary phase.Usually, LC LCD eluent is formed with two distinct solvents.

One of them prevents adsorption of sample constituents (adesorli), while another is an adsorption-promoting solvent (anadsorli). Eluent composition is adjusted so that the desorlieffect prevails. In other words, if a sample is dissolved in eluentand injected into the LC LCD column, it would elute in theexclusion mode. To allow fine-tuning of the adsorption ofsample constituents and to secure solubility of all sampleconstituents, a mixture of several solvents can also be employed

Figure 1. Principle of LC LCD applied to a three polymer blend.

Analytical Chemistry Article

dx.doi.org/10.1021/ac4040746 | Anal. Chem. 2014, 86, 2694−27022695

as eluent. In LC LCD, sample is preceded by a narrow zone of aliquid, which promotes adsorption of at least one of the sampleconstituents, namely, the adsorli. This zone forms a sort of porepermeating and therefore a slowly transported barrier, which isimpenetrable for adsorbing macromolecules and thus signifi-cantly decelerates their elution. At the same time, thenonadsorbed macromolecules elute fast in the exclusionmode. In this way macromolecules that possess distinctadsorptivity can be efficiently separated. For practical reasons,barrier composition is made from the same solvents as eluent.However the concentration of adsorption promoting liquid inbarrier is appropriately increased in comparison with eluent. Itwas shown that LC LCD enables simultaneous separation ofseveral chemically distinct macromolecules. In the case ofsample, which contains n constituents, at least n − 1 barrierswith different compositions must be employed so that eachbarrier decelerates only one sample constituent. When thenumber of barriers and their compositions are properly chosen,one can separate a complex mixture of polymer and especially amixture of homopolymer X, homopolymer Y, and blockcopolymers X-b-Y or Y-b-X-b-Y. To realize such a separation,two barriers are needed. Figure 1 illustrated LC LCD applied toa three polymer blend. In this figure, homopolymer X (red) isnonadsorptive within the chromatographic conditions em-ployed. As a consequence, it will not be retained by any barrierand it will elute in SEC. Homopolymer Y (blue) is adsorptiveand it will be decelerated by barrier 2 (B2). Block copolymer X-b-Y or Y-b-X-b-Y (purple) will not be slowed-down by B2,which is not efficient enough. It will break-through B2 and itwill elute in SEC until it will reach barrier 1 (B1). Since B1 ismore efficient than B2, block copolymer X-b-Y or Y-b-X-b-Ywill be retained by B1. The retained polymers will accumulateon the barriers edge and will be detected as focused peaks.Moreover, with this method, no upper limit for molar mass

was observed.44 The LC LCD method possesses severaladvantages including its high sample capacity, overallexperimental simplicity (isocratic elution mode), fair robustnessto eluent composition and to temperature variation, andacceptable sample recovery.45 LC LCD method showed itsability to discriminate polystyrene-block-poly(methyl methacry-late) (PS-b-PMMA) diblock copolymers from their parenthomopolymers40 as well as low-solubility polymers such asaromatic polyesters.42 Block copolymers containing a highlyadsorptive block as PEO chains can also be successfully

separated by LC LCD with the help of high-strength solvents,like dimethylformamide.43 In this previous study, separations ofPEO-b-PS and poly(propylene oxide)-b-poly(ethylene oxide)(PPO-b-PEO) from their parent homopolymers were alsoachieved. Nevertheless, triblock copolymers have never beencharacterized by LC LCD.Herein, we report original conditions for LC LCD allowing a

detailed characterization of previously prepared triblockcopolymer PS-b-PEO-b-PS sample. Even if the principle ofLC LCD separation is the same whereas the block copolymer isa diblock or a triblock copolymer, it is the first time that LCLCD on triblock copolymers is reported. Moreover, a carefulcomparison of SEC, LC CC, and LC LCD method potential foranalysis of a complex polymer mixture obtained upon PS-b-PEO-b-PS triblock copolymers synthesis chosen as a model isalso provided. On top of that, for the first time, LC LCD off-line coupling with 1H NMR is applied to validate the separationefficiency.

■ EXPERIMENTAL SECTIONMaterials. PEO of molar mass (Mn) 35 kg mol −1 was

purchased from Sigma-Aldrich. BlocBuilder MA (>99%)alkoxyamine based on the nitroxide SG1 (N-tert-butyl-N-[1-diethylphospono-(2,2-dimethylpropyl)]nitroxide) and the 1-carboxy-1-methylethylalkyl moiety was kindly provided byArkema (France). Acryloyl chloride and triethylamine werepurchased from Sigma-Aldrich and were used as received.PEO standards with a molar mass from 0.615 to 114 kg

mol−1 and PS standards (2.17−124 kg mol−1) were purchasedfrom both PSS (Germany) and Agilent. 1-Chlorobutane (CLB)of HPLC-grade was purchased from Sigma-Aldrich and wasused without further purification. Dimethylformamide (DMF)of analytical grade was filtered with a 0.2 μm Nylon Alltechmembrane before use. Tetrahydrofuran (THF) of analyticalgrade was filtered with a 0.2 μm PTFE Alltech membranebefore use. DMF and THF were purchased from Carlo Erba.The mixed eluent and barrier composition is always given inweight parts.

PS-b-PEO-b-PS Triblock Copolymer Synthesis. PS-b-PEO-b-PS triblock copolymer synthesis has been alreadydescribed by our group46 following the procedure depicted inScheme 1.After the synthesis, the material is precipitated in cold

diethylether and then filtered and dried out under vacuum.

Scheme 1. Synthesis Procedure of PS-b-PEO-b-PS Triblock Copolymers



From this synthesis procedure, we can assume that residualfunctionalized PEO and self-initiated PS can be present in theobtained complex polymer mixture. Therefore, the aim of ourwork is to characterize such a polymer blend and highlight thepossible presence of residual parent homopolymers. On thebasis of 1H NMR analysis, the complex polymer mixtureinvestigated contains 56.5 wt % of PS and it is named PS-b-PEO-b-PS in this paper.

1H NMR Analysis. 1H NMR measurements were performedwith a Bruker Avance III 400 MHz Nanobay spectrometer. 1HNMR spectra were recorded at 300 K with a 12.7 μs 30° pulse,a repetition time of 2 and 64 scans. The 1H NMR spectrum ofthe final PS-b-PEO-b-PS copolymer in CDCl3 shows peaks (δ =6.5−7.2 ppm) corresponding to the phenyl protons of the PSblocks and peaks (δ = 3.6−3.7 ppm) corresponding to the−CH2−CH2−O− protons of the PEO blocks. The final overallaverage copolymer composition was thus determined from theratio between the integrals of PS-phenyl 1H signals and thePEO 1H signals. Considering the molar mass values given bythe supplier for the PEO block, the 1H NMR analysis led to thedetermination of the PS block average molar mass given inTable 1.

Size Exclusion Chromatography. SEC experiments wereperformed on an EcoSEC apparatus from PSS, equipped with adual flow cell refractive index detector. Eluent was THF at aflow rate of 0.3 mL min−1 for the sample pump and 0.15 mLmin−1 for the reference pump. The stationary phase was acombination of one PL Resipore (50 mm × 4.6 mm) guardcolumn and two PL Resipore (250 mm × 4.6 mm) columnsthermostated at 40 °C. Samples were prepared at aconcentration of 0.25 wt % in THF containing 0.25 vol % oftoluene, as a flowmarker. The injection volume was 20 μL.Polystyrene equivalent number-average molar masses (Mn) anddispersities Đ were calculated by means of PS calibration curve

using PS-M Easivial standards (Agilent). Results are shown inTable 1 and Figure 2.

LC Measurements. PEO LC CC Analysis. Because of thehigh molar mass (35 kg mol−1) of the PEO block in thecopolymer, we have used the LC CC system described by Maliket al.30 Two Nucleosil columns (4000 Å + 1000 Å)−7 μm (250mm × 4.6 mm) at 30 °C were employed. The eluent was amixture of DMF 90 vol % and THF 10 vol % at 0.5 mL min−1.Sample injection volume and concentration were 20 μL and 2.5mg mL−1, respectively. The evaporative light scattering detector(ELSD) from Polymer Laboratories/Agilent, PL-ELS 2100 wasset at an evaporation temperature of 90 °C and at anebulization temperature of 40 °C. Nitrogen was used as acarrier gas at a flow rate of 1.4 mL min−1.

PS LC CC Analysis. As already described,47 LC CCconditions for PS was neat DMF at 80 °C with the silicaC18 bonded phase (two Nucleodure columns 110 Å−(3 μm +5 μm) (250 mm × 4.6 mm)).

LC LCD Analysis. LC LCD analyses were performed on aWaters 600 chromatography system equipped with a Waters600E pump, a Waters 600 controller, and a Waters 717plusautosampler. Manual Rheodyne valve with a loop of 1000 μLwas employed for barrier injection. A home-packed column 7.5mm × 300 mm that contained bare silica gel Kromasil 60 Å−10μm was thermostated in the Crococil oven (PolymerLaboratories/Agilent). The temperature of measurements was30 °C. The mobile phase was a mixture of dimethylformamide30 wt % and 1-chlorobutane 70 wt %, and the flow rate was setto 1 mL min−1. Samples were dissolved in eluent at aconcentration of 0.25 wt % and filtered with a 0.2 μm PTFESodipro filter. Sample injections were done by the autosamplerat a volume of 50 μL. Two barriers were employed to achieveseparation of all three constituents. They are denoted Barrier 1,B1, and Barrier 2, B2. Their actual composition is indicated inrelated figures. Accurate time delays between the injections ofbarriers and sample are necessary to adjust the peak retentionvolumes and to obtain a well-defined separation. Here, thedelays are noted “0-3-5” which means that B1 is injected at 0min, B2 is injected 3 min after B1, and the sample is injected at5 min after B1. Data recording starts with the injection ofsample. A Polymer Laboratories/Agilent, PL-ELS 2100detector worked at an evaporation temperature of 70 °C and

Table 1. PS-b-PEO-b-PS Average Composition andPolystyrene Equivalent Number Average Molar Mass AsDetermined by 1H NMR and Dispersity (Đ) Provided bySEC

sample PS wt % PS vol % Mn (kg/mol) Đ

PS-b-PEO-b-PS 56.5 58.3 80.5 1.20

Figure 2. SEC chromatogram of PS-b-PEO-b-PS.



at nebulization temperature of 40 °C. Nitrogen was used as acarrier gas at a flow rate of 1.2 mL min−1.

■ RESULTS AND DISCUSSION1H NMR, SEC, and LC CC Characterization. Block length,

number-average molar mass (Mn), and dispersity Đ wereobtained using 1H NMR spectroscopy and size exclusionchromatography. Results are given in Table 1, and thechromatogram obtained by SEC is presented in Figure 2.

1H NMR provides information on the proportions of PS andPEO in the sample but, due to signal overlapping betweenhomo and block copolymers, cannot highlight the presence ofparent homopolymers in the block copolymer sample.The SEC chromatogram is shown in Figure 2. As expected,

resolution of SEC is not high enough to separate blockcopolymers from their parent homopolymers. The SEC profileis slightly fronting in the area of low retention volume, and thepeak also shows a shoulder at high elution volume, which mayindicate the presence of PEO and/or PS homopolymer but theseparation is highly insufficient to establish the presence orabsence of this homopolymer.Then, PS and PEO LC CC analysis were performed to

highlight the presence of parent homopolymers in the blockcopolymer sample. Results are given in Figure 3 for PS LC CCanalysis and in Figure 4 for PEO LC CC analysis.First LC CC at PS critical conditions was performed. The

log(Mp) vs elution volume curve of PS standards is presented inFigure 3a. PS from 2.17 to 28.77 kg mol−1 are eluted in LC CC

and PS with molar masses above 28.77 kg mol−1 are eluted inSEC or enthalpic partition assisted SEC,48 Indeed, thestationary phase used in these conditions is an octadecylgrafted silica. The C18 groups are solvated by the mobile phaseso the stationary phase is considered as a liquid. Separation isbased on enthalpic partition, which means that it depends oninteractions between analytes and mobile phase, analytes andstationary phase, and mobile phase and stationary phase. In thecase of polymers, the elution mechanism will be ruled bysolubility of macromolecules in both stationary and mobilephase. It is well-known from Martin’s rule that the solubility ofmacromolecules decreases with their molar mass. If thesolubility of macromolecules is more influenced by theirmolar mass in the solvated stationary phase than in the mobilephase, macromolecules will be retained by the stationary phase,leading to a rise of elution volume with molar mass. On thecontrary, if solubility of macromolecules decreases with theirmolar mass faster in the stationary phase than in the mobilephase, macromolecules will stay preferentially in the mobilephase and their elution volumes diminished with molar mass.Consequently, under enthalpic partition effects, elution volumeincreases for low molar mass polymer but it will rapidlydecrease with molar mass. These reduction of elution volumeswith molar mass can be more prononced if macromolecules areexcluded from the pores volume.48 In the present case,enthalpic partition and entropic forces compensate for eachother for PS from 2.17 to 28.77 kg mol−1 so they are eluted inLC CC. PS with molar mass higher than 28.77 kg mol−1 aremore soluble in the mobile phase than in the stationary phaseso they are less submitted to enthalpic partition than low molarmass PS and their elution volumes drop with molar mass rising.This is why high molar mass PS elute in enthalpic partitionassisted SEC. LC CC chromatogram of triblock copolymer andPS homopolymers are in Figure 3b. We can notice the presenceof a system peak, probably due to an impurity in DMF. Thepeak at about 3 mL corresponds to PS-b-PEO-b-PS. There is nopeak at about 6 mL, which is the elution volume of PSstandards at critical conditions. Nevertheless, one can notice abroad peak between 3.5 and 4.5 mL, which could be PSoriginated from thermal self-initiation. Indeed, the presence of asmall amount of self-initiated PS is expected in regards to

Figure 3. log(Mp) vs elution volume of PS standards (a) and LC CCchromatograms of a PS standard and PS-b-PEO-b-PS (b). Stationaryphase: 2 Nucleodure C18 columns 3 μm + 5 μm and 250 mm × 4.6mm at 80 °C. Eluent: DMF at 0.8 mL min−1; injection volume, 20 μL;concentration, 2.5 mg mL−1; RI detection.

Figure 4. LC CC chromatograms of a narrow PEO as well as of PS-b-PEO-b-PS. Stationary phase: 2 Nucleosil columns (4000 Å + 1000Å)−7 μm and 250 mm × 4.6 mm at 30 °C; eluent, DMF90 vol%/THF10 vol % at 0.5 mL min−1; injection volume, 20 μL;concentration, 2.5 mg mL−1; ELS detection.



synthesis conditions.46 Because of its high molar mass, it wouldelute in exclusion, as confirmed by Figure 3a. Correlationbetween parts a and b of Figure 3 suggests that the molar massof this self-initiated PS is higher than 28 kg mol−1. We can alsonotice that the peak of triblock copolymer again exhibitsmarked fronting. However, no obvious conclusion can be madefrom this fronting. Particularly, PEO homopolymer, which canbe present in the sample could not be identified.LC CC at PEO critical conditions was also carried out. The

chromatograms of triblock copolymer and PEO homopolymersare depicted in Figure 4. The peak with a maximum at about6.1 mL belongs to PS-b-PEO-b-PS. It exhibits a shoulder at thehigh elution volume, which could correspond to homopolymerPEO. Yet, resolution is too poor to conclude about thepresence of unreacted PEO.From these results, one can conclude that even if 1H NMR,

SEC, and LC CC are widely used for polymer characterization,in the present case, selectivity of separation is too poor tounambiguously identify presence of parent homopolymers inthe investigated PS-b-PEO-b-PS triblock copolymer sample. Inorder to overcome these drawbacks, liquid chromatographyunder limiting conditions of desorption was also explored.Setup and Validation of LC LCD. 1-Chlorobutane was

chosen as the adsorli component of the eluent, because itpromotes adsorption of PEO and PS on bare silica gel. Indeed,its eluent strength (ε°) is about 0.26 whereas ε° of THF, adesorli for PS on bare silica gel, is about 0.45. Dimethylforma-mide (ε° > 0.7) was employed as the efficient desorli whichallows PS and PEO to be eluted in the unambiguous exclusionmode from bare silica gel.30,49

The first step was to identify the eluent composition, whichwould exhibit a rather weak eluent strength but would stillallow SEC elution of PS and PEO. Initially, we chose an eluentcomposition made from 30 wt % DMF and 70 wt % CLB.Under these conditions, PS is eluted in SEC whereas elutionvolumes of PEO are markedly enhanced compared to elutionvolume of PS (cf. Figure 5). This situation is denoted

adsorption assisted SEC, ADA SEC.50 In ADA SEC, retentionvolumes of lower molar mass polymers are increased incomparison with real SEC. Indeed, low molar mass PEO canenter the pores of the column packing and so they interact withattractive sites, which are mainly located in the pores volume.16

Depending on the amount of adsorli in the mobile phase, lowmolar mass PEO will interact more or less strongly with thestationary phase. On the contrary, high molar mass PEO are

totally excluded from the pores and they elute in the interstitialvolume where attractive sites are less present. So high molarmass PEO are submitted to less attractive forces than low molarmass PEO. This is why high molar mass PEO are eluting in theexcluded volume at about 5.8 mL while low molar mass PEOare eluted in ADA-SEC at an elution volume about 16 mL forPEO 0.615 kg mol−1, which is far higher than the total volumeof liquid in the column, determined at 9.7 mL from low molarmass PS eluted in SEC. As a result, selectivity of separation maybe enhanced. On the other hand, the sample recovery of highermolar masses may decrease.The next step was to find appropriate barrier compositions

and the ideal injection delays between injection of barrier 1(B1), barrier 2 (B2), and sample. Our chromatographic systemis close to one already used to separate PMMA-b-PS blockcopolymers from their PMMA and PS parents homopol-ymers.50 As a consequence, we decided to use the sameinjection delays, which are 0-3-5 (see the ExperimentalSection). B2 should separate the block copolymers from theirPEO parent homopolymer, by retaining the PEO and allowingthe copolymer to pass through. We performed a B2composition screening until it fully retained the PEO standardof 35 kg mol−1, corresponding to PEO used for the triblocksynthesis, without affecting elution of block copolymers. Theappropriate B2 composition was DMF24/CLB76. The B1barrier has to decelerate the block copolymers, withoutaffecting PS. In this way, block copolymers would bediscriminated from their PS parent homopolymers. The B1composition screening revealed that the appropriate composi-tion was DMF5/CLB95. Such a barrier retained blockcopolymers but did not affect elution of PS 28.77 kg mol−1

(center of the PS molar mass range studied here). Under theseconditions, the separation of PS-b-PEO-b-PS from both parenthomopolymers in a single run was investigated. Results, shownin Figure 6, demonstrate clearly the obtained chromatograms

with baseline-separated peaks. PS standard 28.77 kg mol−1,PEO standard 35 kg mol−1, and PS-b-PEO-b-PS are eluted at5.8 mL, 9.7 mL, and 7.7 mL, respectively. Interestingly, itappears that the block copolymer sample PS-b-PEO-b-PScontains a non-negligible amount of self-initiated PS and alsosome unreacted PEO. PS is not retained by any barrier andelutes in the exclusion mode. As we can see in Figure 5, thetotal exclusion volume of the column is about 5.8 mL;

Figure 5. log(Mp) vs elution volume for PS and PEO standards.Stationary phase: column Kromasil 60 Å−10 μm 300 mm × 7.8 mm;eluent, 70 wt % CLB/30 wt % DMF; injection volume, 50 μL;concentration, 2.5 mg mL−1; ELS detection.

Figure 6. LC LCD chromatograms of block copolymer PS-b-PEO-b-PS (blue) as well as of PS standard 28.77 kg mol−1(black) and PEOstandard 35 kg mol−1 (red). See the Experimental Section forchromatographic conditions. B1, DMF5/CLB95; B2, DMF24/CLB76.



therefore, PS standard 28.77 kg mol−1 is near to its fullexclusion from the column packing pores. PS present in theblock copolymer sample PS-b-PEO-b-PS also elutes at about5.8 mL, so it is totally excluded from the pores of the columnpacking and no reasonable direct estimation of its molar mass ispossible. However, it appears from Figure 5 that this lattershould be equal or higher than 28 kg mol−1, which confirmsvery nicely the hypothesis of self-initiated PS, already suggestedfrom PS LC CC experiments. PEO is retained by B2, and it iseluted independently of its molar mass at about 9.7 mL.Consequently, we cannot estimate the molar mass of the PEOpresent in the block copolymer sample PS-b-PEO-b-PS.Nevertheless, this block copolymer was synthesized from acommercially available PEO with molar mass of 35 kg mol−1

and a low dispersity. Consequently, residual PEO molar massshould be 35 kg mol−1 too. It is also important to notice thatPEO with a molar mass below 12 kg mol−1 elutes too slowly tocatch B2 and be retained. However, because of their elution inADA SEC, this low-molar-mass PEO still can be separated fromtriblock copolymer (results not shown here).To verify the efficiency of separation, we have inspected the

presence of PS in the “PEO fraction” and absence of PEO inthe “PS fractions”. Indeed, PS-b-PEO-b-PS with short PEOblocks and long PS blocks could break-through B1 and on thecontrary, some PS-b-PEO-b-PS with long PEO blocks and shortPS blocks could be retained by B2. Therefore, we have repeatedthe above separation 40 times and collected both PS fractionsfrom 5.5 to 7.0 mL and PEO fractions from 9.4 to 10.5 mL.The solutions were dried out with the help of a rotaryevaporator under vacuum. Next, 700 μL of CDCl3 were addedand the obtained samples were analyzed by 1H NMR(conditions described in the Experimental Section). We alsorealized a blank by evaporating 1.5 mL of eluent with a rotaryevaporator, added 700 μL of CDCl3, and analyzed it by 1HNMR. Results are shown in Figure 7 for the blank sample, inFigure 8 for the PS fractions and the block copolymer, and inFigure 9 for the PEO fractions.On the 1H NMR spectrum of the blank sample, we notice

the presence of residual DMF (singlet at δ = 8 ppm and adoublet at δ = 2.9 ppm). As indicated by the provider, DMFcontains traces of methanol (about 100 ppm per liter) observedas a small singlet at δ = 3.65 ppm.

The 1H NMR spectrum of the PS-b-PEO-b-PS samplepresented Figure 8 shows peaks (δ = 6.5−7.2 ppm)corresponding to the phenyl protons of the PS block and asinglet at δ = 3.65 ppm corresponding to the −CH2−CH2−O−protons of the PEO block. When the integration of the phenylprotons of the PS block is set to 1.00, the obtained value for theintegration of the methylene protons of the PEO block is thenequal to 1.06. On the same figure, the 1H NMR spectrumcorresponding to the PS fractions shows peaks between δ =Figure 7. 1H NMR spectrum of blank sample.

Figure 8. 1H NMR spectrum of PS-b-PEO-b-PS sample and PSfractions.



6.5−7.2 ppm still corresponding to the aromatic protons of PSand also a small singlet at δ = 3.65 ppm. In this case, when theintegration of the phenyl protons of PS is set to 1.00, theobtained value for this singlet equals 0.03. Interestingly, thisresult highlights an almost complete decrease of the PEOintegration value. Unfortunately, because MeOH and PEO givesignal at the same position, it is difficult to confirm whether ornot this residual peak is assigned to PEO or methanol tracesarising from DMF. Typically, from the study of the blanksample 1H NMR spectrum and the integration values for thesinglet at δ = 3.65 ppm in the PS-b-PEO-b-PS 1H NMRspectrum as well as in the PS fractions 1H NMR spectrum, wecan reasonably conclude that there is no or a negligible amountof block copolymer in the PS fractions. We then conclude thatB1 is an efficient barrier.The 1H NMR spectrum of the PEO fractions in Figure 9

shows a singlet at δ = 3.65 ppm. The intensity of this singlet ismuch higher than in Figure 7, which indicates the presence ofthe −CH2−CH2−O− protons of PEO. No peaks between δ =6.5−7.2 ppm corresponding to the phenyl protons of the PSblock is found in the PEO fraction. So B2 is proved to be wellefficient as well.

■ CONCLUSIONTo conclude, we demonstrated that SEC cannot discriminate asmall amount of parent homopolymers present in the triblockcopolymer PS-b-PEO-b-PS. LC CC separates only one parenthomopolymer from the block copolymer, while the secondhomopolymer elutes together with the block copolymer.Interestingly, LC LCD is able to separate diblock or triblockcopolymers from their parent homopolymers in one single run.LC LCD separation is very efficient and particularly convenient.The method also shows high sensitivity and resolutioncompared to both SEC and LC CC. Minor macromolecularadmixtures present in the block copolymer matrix can beidentified. 1H NMR analysis confirmed the high efficiency ofLC LCD separation, although it seems important to verify if thefraction of block copolymer is not contaminated withhomopolymers too. On top of that, we highlighted a limitconcerning low molar mass PEO. Indeed, because of theirelution in ADA SEC, they are too slow to catch B2 and beretained. Although the LC LCD separation of the triblockcopolymer studied here is not affected by this phenomenon, thelimits of this method have to be investigated deeply. This is thesubject of our future work.

■ AUTHOR INFORMATIONCorresponding Authors*E-mail: [email protected].*E-mail: [email protected] authors declare no competing financial interest.

■ ACKNOWLEDGMENTSThis work was done in the frame of Interacademic Agreementon Cooperation between Centre National de la RechercheScientifique, France and the Slovak Academy of Sciences,Slovakia, Project “Synthesis, molecular characterization andpurification of block copolymers”. The financial support fromAix-Marseille University and CNRS and from the Slovak GrantAgencies VEGA (Project 2/0001/12) and APVV (Projects0109-10 and 0125-011) is acknowledged.

■ REFERENCES(1) Dixit, S. G.; Mahadeshwar, A. R.; Haram, S. K. Colloids Surf.1998, 133, 69.(2) Sheiko, S. S. Adv. Polym. Sci. 2000, 151, 61.(3) Lee, J. S.; Hirao, A.; Nakahama, S. Macromolecules 1988, 21, 274.(4) Widawski, G.; Rawiso, M.; Francois, B. Nature 1994, 369, 387.(5) Patel, N. P.; Spontak, R. J. Macromolecules 2004, 37, 1394.(6) Singh, M.; Odusanya, L.; Balsara, N. Polym. Mater. Sci. Eng. 2005,93, 549.(7) Panday, A.; Mullin, S.; Gomez, E. D.; Wanakule, N.; Chen, V. L.;Hexemer, A.; Pople, J.; Balsara, N. P. Macromolecules 2009, 42, 4632.(8) Sadoway, D. R. Power Sources 2004, 129, 1.(9) Trapa, P. E.; Won, Y. Y.; Mui, S. C.; Olivetti, E. A.; Huang, B.;Sadoway, D. R.; Mayes, A. M.; Dallek, S. J. Electrochem. Soc. 2005, 152,A1.(10) Ryu, S.-W.; Trapa, P. E.; Olugebefola, S. C.; Gonzalez-Leon, J.A.; Sadoway, D. R.; Mayes, A. M. J. Electrochem. Soc. 2005, 152, A158.(11) Bouchet, R.; Maria, S.; Meziane, R.; Aboulaich, A.; Liefana, L.;Bonnet, J.-P.; Phan, T. N. T.; Bertin, D.; Gigmes, D.; Devaux, D.;Denoyel, R.; Armand, M. Nat. Mater. 2013, 12, 452.(12) Riess, G. Prog. Polym. Sci. 2003, 28, 1107.(13) Hamley, I. W. The Physics of Block Copolymers; OxfordUniversity Press: Oxford, U.K., 1998.(14) Moad, C. L.; Moad, G.; Rizzardo, E.; Thang, S. H.Macromolecules 1996, 29, 7717.(15) Guillaneuf, Y.; Castignolles, P. J. Polym. Sci., Part A: Polym.Chem. 2008, 46, 897.(16) Philipsen, H. J. A. J. Chromatogr., A 2004, 1037, 329.(17) Brun, Y.; Alden, P. J. Chromatogr., A 2002, 966, 25.(18) Jandera, P.; Holcapek, M.; Kolarova, L. J. Chromatogr., A 2000,869, 65.(19) Brun, Y.; Foster, P. J. Sep. Sci. 2010, 33, 3501.(20) Ryu, C. Y.; Han, J.; Lyoo, W. S. J. Polym. Sci., Part B: Polym.Phys. 2010, 48, 2561.(21) Chagneux, N.; Trimaille, T.; Rollet, M.; Beaudoin, E.; Gerard,P.; Bertin, D.; Gigmes, D. Macromolecules 2009, 42, 9435.(22) Philipsen, H. J. A.; Wubbe, F. P. C.; Klumperman, B.; German,A. L. J. Appl. Polym. Sci. 1999, 72, 183.(23) Chan, H. C., Chia, C. H., Thomas, S., Eds. Physical Chemistry ofMacromolecules: Macro to Nanoscales; Apple Academic Press, Inc.:Waretown, NJ, 2014; Chapter 11.(24) Berek, D.; Russ, A. Chem. Pap. 2006, 60, 249.(25) Teramachi, S.; Hasegawa, A.; Matsumoto, T. J . Chromatogr.1991, 547, 429.(26) Lee, H.; Chang, T.; Lee, D.; Shim, M. S.; Ji, H.; Nonidez, W. K.;Mays, J. W. Anal. Chem. 2001, 73, 1726.(27) Batsberg, W.; Ndoni, S.; Trandum, C.; Hvidt, S. Macromolecules2004, 37, 2965.(28) Mass, V.; Bellas, V.; Pasch, H. Macromol. Chem. Phys. 2008, 209,2026.

Figure 9. 1H NMR spectrum of PEO fractions.



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(29) Rollet, M.; Gle, D.; Phan, T. N. T.; Guillaneuf, Y.; Bertin, D.;Gigmes, D. Macromolecules 2012, 45, 7171.(30) Malik, M. I.; Harding, G. W.; Grabowsky, M. E.; Pasch, H. J.Chromatogr., A 2012, 1244, 77.(31) Beaudoin, E.; Dufils, P. E.; Gigmes, D.; Marque, S.; Petit, C.;Tordo, P.; Bertin, D. Polymer 2006, 47, 98.(32) Sinha, P.; Grabowsky, M. E.; Malik, M. I.; Harding, G. W.;Pasch, H. Macromol. Symp. 2012, 313−314, 162.(33) Gorbunov, A. A.; Trathnigg, B. J. J. Chromatogr., A 2002, 955, 9.(34) Rappel, C.; Trathnigg, B.; Gorbunov, A. A. J. Chromatogr., A2003, 984, 29.(35) Girod, M.; Phan, T.; Charles, L. Rapid Commun. Mass Spectrom.2008, 22, 3767.(36) Berek, D. Macromol. Symp. 1996, 110, 33.(37) Philipsen, H. J. A.; Klumperman, B.; German, A. L. J.Chromatogr., A 1996, 746, 211.(38) Beaudoin, E.; Favier, A.; Galindo, C.; Lapp, A.; Petit, C.;Gigmes, D.; Marque, S.; Bertin, D. Eur. Polym. J. 2008, 44, 514.(39) Siskova, A.; Macova, E.; Corradini, D.; Berek, D. J. Sep. Sci.2013, 36, 2979.(40) Berek, D. Macromol. Chem. Phys. 2008, 209, 695.(41) Berek, D. Prog. Polym. Sci. 2000, 25, 873.(42) Siskova, A.; Macova, E.; Berek, D. Eur. Polym. J. 2012, 48, 155.(43) Berek, D. J. Sep. Sci. 2010, 33, 3476.(44) Snauko, M.; Berek, D. J. Sep. Sci. 2005, 28, 2094.(45) Snauko, M.; Berek, D. Chromatographia 2003, 57 (Suppl), S55.(46) Perrin, L.; Phan, T. N. T.; Querelle, S.; Deratani, A.; Bertin, D.Macromolecules 2008, 41, 6942.(47) Petit, C.; Luneau, B.; Beaudoin, E.; Gigmes, D.; Bertin, D. J.Chromatogr., A 2007, 1163, 128.(48) Berek, D. Macromol. Symp. 2004, 216, 145.(49) Berek, D. Chromatographia 2003, 57 (Supplement), S45.(50) Berek, D. Macromol. Chem. Phys. 2008, 209, 2213.

