Polímeros biocompatíveis baseados em poli oxido de etileno

8/12/2019 Polmeros biocompatveis baseados em poli oxido de etileno

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http://localhost/var/www/apps/conversion/tmp/scratch_3/dx.doi.org/10.1016/j.progpolymsci.2011.10.002mailto:[email protected]:[email protected]://www.elsevier.com/locate/ppolyscihttp://www.sciencedirect.com/science/journal/00796700http://localhost/var/www/apps/conversion/tmp/scratch_3/dx.doi.org/10.1016/j.progpolymsci.2011.10.002


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688 C.Weber et al. / Progress in Polymer Science 37 (2012) 686714

particles [1] . Thermo-responsive polymers that undergo asolubility transition in water have received major inter-est in recent decades based on their broad applicationpotential in various interesting elds, such as protein chro-matography [2] , sensing devices [3] , protein adsorptionand tissue engineering [4] , temperature triggered drugdelivery and regenerative medicine [58] . Nowadays, liv-ing and controlled polymerization mechanisms are widelyapplied for the synthesis of thermo-responsive polymersproviding a perfect tool for the construction of advancedpolymer architectures and enabling the combination of thermo-responsiveness with a second response towardsother environmental triggers, such as pH value, magneticeld or light, within one molecule [9] .

Several polymers that respond with a solubility tran-sition to temperature changes in aqueous solution areknown from literature among whose the most well-known is poly( N -iso-propylacrylamide) (PNiPAm) sinceits lower critical solution temperature (LCST) in water,i.e. the temperature at which the polymer switches fromhydrophilic to hydrophobic, is close to body tempera-ture [10] . The progress in the design and applicationof new PNiPAm based thermo-responsive copolymersduring the last years was already covered in a rangeof review articles [5,6,11] . On the other hand, besidespolymers where the polyacrylamide structure is main-tained, there are other interesting polymers that exhibitLCST behavior in water based on various structuralmotifs, such as methyl cellulose, polyethers, poly(2-oxazoline)s, poly( N,N -dimethyl aminoethyl methacrylate)(PDMAEMA), poly(vinyl caprolactone), certain polypep-tides or poly(methyl vinyl ether) [7,12] . Apart from thelatter polymer class, which was recently reviewed byAoshima and Kanaoka [11] the thermo sensitive prop-erties of these polymers were only briey mentionedin recent reviews. In addition, Lutz discussed the syn-thesis [13] and application [14] of poly(oligo[ethyleneoxide]methacrylate) based thermo-responsive materials intwo recent highlights.

The scope of the current review is to provide aninsight into the structural and environmental factorsthat can inuence the thermo-responsive properties of certain biocompatible polymers. For this purpose, twospecic polymer classes that are both interesting candi-dates for biomedical applications will be discussed [15] :poly(2-oxazoline)s (POx) and systems whose thermo-responsiveness is based on their structural similarity topoly(ethylene oxide) (PEO). Therefore, experimental datathat were obtained by different groups will be com-pared in detail, although this is complicated by thedifferent measurements conditions that were applied dur-ing the investigation of the LCST behavior. Nonetheless,as we will demonstrate, it is possible to draw con-clusions about the inuence of the polymer structureon its LCST behavior. Most of the polymers that willbe discussed were synthesized by living or controlledpolymerization techniques, but the applied synthesismethods will not be mentioned in detail since the focusof this review is on the detailed evaluation of thethermo-responsive properties of the synthesized materi-als.

2. Methods to investigate thermo-responsivepolymers in solution

When a polymer is molecularly dissolved in a suit-able solvent, it may become insoluble upon increase ordecrease in temperature and, thus, precipitate from thesolution. In other words, the binary polymer/solvent mix-ture undergoes a temperature induced phase separationfrom a one-phasic towards a bi-phasic system due to theexistence of a miscibility gap in the phase diagram [16,17] .If elevation of temperature results in phase separation thesystems exhibits lower critical solution temperature (LCST)behavior. To be more precise, the polymer does not simplyprecipitate from the solution, but two phases are formedin equilibrium, whereby one phase has a high polymerconcentration and the other one has a low polymer con-centration. As shown in Fig. 1 , the LCST is dened as thetemperature at the minimum of the binodal (or the coex-istence curve) of the phase diagram. The correspondingconcentration is the lower critical solution concentration(LCSC). The reverse case, where phase separation occursupon decreasing temperature, is called upper critical solu-tion temperature (UCST) behavior.

Due to the numerous possible applications in biomedi-cal science there is an increasing interest in polymers thatexhibit LCST behavior in water. Apparently, such a poly-mer behaves hydrophilic at low temperatures and turnshydrophobic at elevated temperature. Below the demix-ing temperature of the solution the polymer is capable toform hydrogen bonds with surrounding water moleculesresulting in hydration. With increasing temperature thosehydrogen bonds are weakened and it is more favorablefor the water molecules to be expelled from the polymerstructure into the bulk water. As a result, the polymerchains are (partially) de-hydrated and agglomerate. Sincethe phase separation is accompanied by conformationalchanges of the polymer, such an effect is often referredto as coil to globule transition of the responsive polymer(Fig. 2). From the thermodynamic point of view, hydro-gen bonding between polymer chainsandwatermoleculesgives a favorable enthalpy contribution to the free energyof mixing whereas the binding of the water molecules tothe polymer chain results in an enhanced ordering. There-fore, it contributes negatively (unfavorably) to the entropyof mixing. At higher temperatures, the entropy term T S becomes predominant and the free energy of mixing getspositive, which is manifested in phase separation.

The easiest and most widely used method to obtaininformation about the coil to globule transition temper-ature of a polymer in solution is turbidimetry. A solutionwith a dened concentration of polymer is subjected toa variable temperature program, whereby the transmit-tance of light through the solution is constantly measured.As soon as the phase separation takes place, the transmit-tance will rapidly decrease due to agglomeration of thecollapsed polymerglobules forming aggregates thatscattertheirradiated light.The temperatureat which this happensis called the cloud point temperature ( T cp ). If the inves-tigated mixture is cooled down, the polymer re-dissolvesandthetransmittanceincreases again. In addition,dynamiclight scattering (DLS) measurements are often applied in


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C. Weber et al. / Progress in Polymer Science 37 (2012) 686714 689

Fig. 1. Phase diagram for a binary polymersolvent mixture exhibiting (a) LCST behavior and (b) UCST behavior.

order to gain information about the size and stability of the collapsed polymer globules. Since the coil to globuletransition of a thermo-responsive polymer represents anendothermic process, it can also be followed by calori-metric methods, such as differential scanning calorimetry(DSC) and pressure pertubation calorimetry (PPC). In addi-tion, (modulated) DSC allows the determination of theenthalpy of the transition ( H ) and the glass transitiontemperature of the collapsed globules, whereas PPC canbe used to determine the volume change V /V during thephase transition.

In a range of applications, the thermo responsive poly-mer is exposed to a large amount of varying cosolutes in its

aqueous solution. All those additives might alter the T cp of theaqueous polymer solution. In themost simplecase,oneadditive changes the binary system into a ternary system.Besides theeffect of other solventsor surfactants, in partic-ular theinuenceof salts hasbeen investigated in detailforthepolymers that arediscussed withinthis review. Inorder

to understand the effect of salts on the solubility behaviorof synthetic polymers it is helpful to apply the knowledgethat has been achieved in protein chemistry: different saltsincrease or decrease the solubility of proteins since theyinduce changes in thesecondaryand tertiarystructure. Thisobservation is referred to as salting in or salting out effect,respectively.Salts thatenhancethesolubility (elevated T cp )are called kosmotropes they decrease the ordering of water. In contrast, salts that have a salting out effect (low-ered T cp ) arechaotropes they strengthen thehydrophobicinteractions. Both, anions and cations exhibit such effects,but the inuence of the anion is more pronounced. In theso-calledHofmeister series, anions and cations are ordered

according to their ability to strengthen the hydrophobicinteraction:

Anions : SO 4 2 > OAc > Cl > I > ClO4 > SCN

Cations : NH 4 + > K+ > Na + > Li+ > Mg 2+ > Ca2+

Fig. 2. Coil to globule transition of a thermo-responsive polymer in aqueous solution.


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Fig. 3. Schematic representation of the structures of poly(ethylene oxide) (PEO), poly(propylene oxide) (PPO), poly(1,3-dioxolane) (PDXL), poly(1,3,6-trioxacyclooctane) (PTCO) and poly(1,3,6,9-tetraoxacycloundecane) (PTCU).

3. Linear polyethers

3.1. Poly(ethylene oxide)

Poly(ethylene oxide) (PEO, Fig. 3) is a hydrophilic poly-mer that retains its solubility in water upon heating atambient pressure. However, increaseof temperatureabove100 C results in phase separation and further heating sol-ubilizes the PEO again. Thus, the phase diagram of thebinary mixture PEO/water represents a closed loop coex-istence type that shows both, an LCST as well as an UCST[18] . As shown in Fig. 4 , increasing molar mass dimin-ishes thesolubility of thePEO in water resulting in loweredLCST but elevated UCST, as typical for FloryHuggins Type

Fig. 4. Temperature/weight fraction phase diagram for the systemPEO/water. Molar masses are provided as M .

Reproduced from Saeki et al. [19] by permission of Elsevier Science Ltd.,UK.

I behavior [19] . The observed cloud point temperatureswere found to be independent of the pressure applied tothe system, which could not be explained by simple vander Waals interactions between water and PEO. Instead,hydrogen bonds that are formed between the PEO chainsand water are supposed to enhance the structuring of sol-vent molecules around thepolymer coil [20] . This behavioris entropically unfavorable but the favorableenthalpytermis dominant at low temperatures resulting in miscibility.In contrast, the entropy contribution becomes predomi-nant at higher temperatures causing a disruption of thewater structurearound thePEO and, therefore, a phase sep-aration. The peculiar properties of the system PEO/water

have attracted many theoretical scientists who tried todescribe its phase behavior, mainly by modication of the FloryHuggins theory [2126] . Already in 1959 Bai-ley and Callard investigated the inuence of inorganicsalts as additives upon the phase transition temperature of PEO aqueous solutions ( c = 0.5 wt%) and found salting outeffects of both anions and cations to follow the Hofmeis-ter series for proteins [27] . In addition, the salting outeffects of sodium chloride and sodium propionate werefurther studied by Saeki and co-workers and the resultscould be related to the thermodynamic equation of state[28] .

3.2. Poly(propylene oxide)

Although the LCST behavior of PEO in combination withits rather simple polymer structureis certainlyof academicinterest for the understanding of the LCST phenomenon,the high phase transition temperature is of little practi-cal importance. However, when the hydrophobicity of thepolyether is increasedby additionofonecarbonatom to therepeating unit, as in the case of poly(1,2-propylene oxide)(PPO), the phase transition temperature of the binary mix-ture with water decreases below the boiling point of thesolvent under ambient pressure. Already in 1957, Mal-colm and Rowlinson developed the phase diagram for thissystem and found a LCST of around 50 C for PPO with amolar mass of M n =400gmol 1 [18] . Saito determined thecloud point curves for PPO in the concentration range upto 80mM [29] . The T cp s of aqueous solutions were foundto decrease from around 38 C to 20 C and 17 C withincreasing molar masses of the PPO ( M n =1000gmol 1 ,2000gmol 1 and 3000g mol 1 , respectively). It should benoted that a large range of block copolymers of PEO andPPO are commercially available under the trade namePluronic and have also been investigated with respect totheir solubility behavior in aqueoussystems,which willnotbe covered here.


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3.3. Statistical copolymers of ethylene oxide and propylene oxide

Statistical copolymerization of EO with PO results incopolymers that exhibit LCST behavior in a temperaturerange in between the LCST of both homopolymers. Thiswas already demonstrated in 1959 by Bailey and Callardwho found a linear decrease of the cloud point tempera-tures of aqueous copolymer solutions ( c =0.1 and 1wt%)with increasing content of PO in the statistical copolymer[27] . A further example is given by Tjerneld andco-workerswho investigated a copolymer containing equal amountsof EO and PO with a molar mass of 4000g mol 1 [30] . Theexperimentally determined cloud point curve revealed aminimum at 50 C and 1020 wt% concentration. Based onthis knowledge, the theoretical phase diagram was alsoconstructed for degrees of polymerization from 39 to 156.The calculations using the FloryHuggins theory providehints towards a similar closed loop coexistence for the sta-tistical copolymers as for PEO. With respect to applicationsof PEOPPO copolymers for enzyme purication, Perssonet al. determined the coexistence curves of three poly-mers with PO content of 50%, 70% and 80%, respectively[31] . The LCST was found to decrease with increasing con-tent of hydrophobic PO from around 50 to 40 and 30 C,respectively. Copolymers with a lower content of PO from12 to 26% but similar M w of around 30,000g mol 1 wereinvestigated by Louai et al. [32] . The LCSTs of their binarymixtures with water were determined as minima of thecloud point curves and were found to increase in a linearfashion from 74 to 87 C with increasing EO content of thecopolymer. Thephasediagrams were calculated in analogyto PEO. Determination of the heat of fusion of water andof the excess mixing volume revealed rather similar val-ues as for PEO. This led to the conclusion that, despite theattachment of an additional methyl group at some repeat-ing units, the structure of water around the copolymer isvery similar to that in PEO solutions. Further experimentswiththesepolymerswere conductedin orderto investigatethe effect of various additives upon the phase transitiontemperature of the aqueous solutions [33] . The salting outeffect of inorganicsalts wasshown to follow theHofmeisterseries: KI< NaCl< Na 2 CO3 < Na3 PO4 < Na2 B4 O7 . It is inter-esting to note that the more polar copolymers with higherEO content were affected in a stronger manner than themore hydrophobic ones, which is most likely a result of theenhanced dehydration of the more hydrophilic polymersin the presence of salts. In addition, also organic additives,such as carboxylic acids, alcohols, diols and amides, withvaryinghydrophilicityeitherincreased or decreasedthe T cpof the copolymer solutions ( c polymer =4wt%).

3.4. Other linear polyethers

Benkhira et al. performed comprehensive studies onthe solubility behavior of linear polyethers that are com-posed ofbothEOandmethylene oxide(MO)repeating units(Fig. 3). The cationic polymerization of cyclic monomers,1,3-dioxolane, [34] 1,3,6-trioxacyclooctane and 1,3,6,9-tetraoxacycloundecane, [35] resulted in polyethers thatcontain varying molar fractions of EO and MO in an

alternating fashion. Since PMO is insoluble in waterincreasing EO content of the polymer led to an increasedLCST, as shown by comparison of the phase diagrams,which were developed by turbidimetry (102 C for PTCU,95 C for PTCOand 70 C for PDXL, respectively). In contrastto PTCU and PTCO, whose demixig curves had simi-lar shapes as PEO, the solubility behavior of PDXL wasmore complex: in the concentration range between 20and 40wt% the binodal reached a plateau and stronglydecreased at higher concentrations, where the demixingcurve only represent a metastable state. When higher con-centratedmixtures of PDXL andwater were kept forseveraldays, PDXL was found to crystallize even under ambientconditions. Analysis of the polymer series in aqueous solu-tion by DSC revealed that PDXL forms a stable hydrateand that mainly the EO segments of the polymer chainwere hydrated. In addition, the inuence of anionic [36]and cationic surfactants [37] on the phase separation of polymer/water mixtureswas wellstudied for the completepolyether series and compared to PEO.

Although the LCST behavior of PEO has been wellinvestigated throughout the last decades, it is difcultto incorporate other functionalities to the polymer itself because of the rather complicated polymerization of theEO monomer, which is gaseous and highly toxic. There-fore, most of the studies relied on commercially available(co)polymers for their investigations. Thisdrawbackcan beovercome by the end functionalization of short PEO chainswith polymerizable units resulting in macromonomers.With this approach, the properties of PEO can be main-tained and a straightforward access to a larger quantityof well-dened copolymers is provided using a variety of polymerization methods.

4. Systems containing oligomeric ethylene oxideside chains

By attachment of short hydrophilic PEO chains to vari-ous kinds of polymeric backbones, the advantages of PEOwith respect to its biomedical applications can be main-tained while the phase transition temperature is lowereddue to thehydrophobicityof thepolymer backbone.A suit-able way for the synthesis of such comb shaped or bottlebrush polymers is the macromonomer method where theshort PEO chain is end functionalized with a moiety thatcan be polymerized in a subsequent step. Since the usedabbreviations for the resulting polymers are not consistentin the literature, they will be systematically named dur-ing this review as illustrated in Fig. 4 (right). The degree of polymerization of the pendant EO side chains will be givenby EOn and the type of polymeric backbone will be abbre-viated as follows: V for vinyl ether, St for styrene, A foracrylate, MA for methacrylate and LA for lactide. The endgroup of the pendant EO n side chains will be indicated by:m for methyl, Et for ethyl and H for hydrogen.

4.1. Polymacromonomers with varying side chain length

Besides the nature of the used backbone, the thermo-responsive properties of the polymer will be stronglyinuenced by the length of the pendant PEO chains.


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the aqueous solution by 5 C (entrance and in Fig. 6 ).In addition, a small effect of the polymer tacticity uponthe thermo-responsiveness was observed. 1H NMR mea-surements in D 2 O at elevated temperatures showed thecompletedisappearanceof signals for PmEO 2 MA above thecloudpointof thesolution whereasthe longer side chainsof PmEO 3 MA remained mobileeven upon precipitation of thepolymer above T

cp. The corresponding PmEO

4MA, which

was synthesized from a monodisperse macromonomer aswell, was found to precipitate from its aqueous solutionat 68 C under similar measurement conditions reectinga further increase of hydrophilicity by the longer EO 4 sidechain [40] .

PmEO 2 MA as well as PmEO 3 MA were also synthesizedby ATRP by Yamamoto et al.who extended theinvestigatedmolar mass range of PmEO 2 MA to M n =60,000gmol 1

[45] . The T cp values observed by turbidity measurements(c =3mg mL 1 ) are in excellent agreement with resultsobtainedfor thepolymersthatweresynthesizedby anionicpolymerization as demonstrated in Fig. 6 (entrance ). Thepolymer concentration wasshownto have a stronger inu-ence on the phase separation temperature for PmEO 3 MAthan forPmEO 2 MA. In theinvestigatedconcentrationrangefrom 1mg mL 1 to 10mg ml 1 , T cp decreased from 26to24 C for PmEO 2 MA and from 51 to 47 C for PmEO 3 MAupon increase of concentration. Next to turbidity mea-surements, the thermal properties of the aqueous polymersolutionswereinvestigatedby DLSrevealingthe formationof very large aggregates of 2.7 m size upon precipitationat T cp for PmEO 2 MA. In addition, PmEO 4.5 MA was synthe-sized by RAFT polymerization of a commercially available(but not monodisperse) macromonomer [47] . Turbiditymeasurements in buffered solutions at pH 4, 7 and 10,respectively, and a polymer concentration of 5 mgmL 1

revealed a T cp of 64 C, which was unaffected by the pHof the solution.

An additional increase of the hydrophilicity of thePmEO n MA polymers can be achieved when the EO chain iselongated to approximately, i.e. not monodisperse, 8 unitsas demonstrated by Laschewsky who applied RAFT poly-merization of mEO 8 MA [41] . Turbidity measurementsrevealed a T cp of 85 C of a 1wt% aqueous solution of acomb polymer with a DP of 16 in the polymer backbone.Further increase of the EO side chain length towards 22repeating units resulted in polymers that are water solublein the entire temperature range of liquid water at ambientpressure [48] .

Theutilization of monodispersemacromonomers basedon acrylates for NMP yielded the corresponding PmEO n Awith 2 as well as 3 repeating units of EO as side chains,as was demonstrated by the Zhao group (entrance inFig. 5 ) [42] . Turbidity measurements of aqueous polymersolutions were carried out in a concentration range from0.05 wt% to 5 wt% and revealed T cp s from 35 C to 47 Cfor PmEO 2 A and from 57 C to 65 C for PmEO 3 A, respec-tively. Thecloud point curves of both polymers were foundto decrease steeply at lower concentrations (up to 1 wt%)whereas they became rather at at higher polymer con-centrations. 1H NMR spectroscopy in D 2 O revealed linebroadening of the peaks above the cloud point, but thesignals were still visible indicating a high hydrophilicity

of the polymers even after dehydration. In contrast to thePmEO n MA discussed above, an increase of the molar massof the PmEO n A resulted in elevated T cp s (by around 5 C)inthe M n range from 8000g mol 1 to 16,000g mol 1 , whichwas attributed to the relatively large hydrophobic poly-mer end group resulting from NMP synthesis that has alarger impact on the solubility of shorter polymer chains(entrance and in Fig. 6 ).

A similar study was carried out applying NMP of monomers that are based on styrene comprising monodis-perse oligomeric EO units (entrance in Fig. 5) [43] . Theaqueous solutions ( c =0.5wt%) of the resulting PmEO n Stcontaining 3, 4 and 5 repeating units of EO as side chainsdisplayed T cp s of 13 C, 39 C and 53 C, respectively. ForPmEO 3 St, the aromatic signals of the polymer backbonecompletely disappeared from the 1 HNMR spectrum in D 2 Oas soon as the cloud point of the polymer solution wasreached while the PEO signals remained visible, althoughbroadened. However, the increasing hydrophilicity of thestructures obtained with higher EO content (PmEO 4 St andPmEO

5St) was reected in the fact that even the sig-

nals deriving from the highly hydrophobic PS backboneremained visible above T cp .

The use of this strategy was also applied in order tosynthesize thermo-responsive polyglycolides (entrancein Fig.5) f rom thecorresponding EO containingcyclic dilac-tides [44] . Dueto therelatively long and, thus, hydrophobicalkyl chains that were used to connect the oligomeric EOunits to the lactide, the polymers containing one as well astwoEO units were found to be insolublein water.However,turbidity measurements of aqueous solutions at polymerconcentrations of 15wt% revealed cloud point temper-atures at 20 C and 38 C for PmEO 3 LA and PmEO 4 LA,respectively.Thepolymer signals in 1 HNMR spectra inD

2O

were broadened above the cloud point, and DLS measure-ments at 3mg mL 1 conrmed the agglomeration of thepolymer chains intolarge aggregates above thecloudpoint.

4.2. Effect of side chain end groups

Allvalues discussedin theprevious section arebasedonpolymer systems with methylgroups at theend of thepen-dant PEO chains. However, since a comb polymer containsasmanyoftheseendgroupsassidechains,theyhavealargeimpact on the solubility properties of the entire polymer.In addition, those end groups point towards the outside of the macromolecular bottle brush structure in solution asdemonstrated in Fig. 7 . This further enhances their effecton the phase transition temperature since the surround-ing water molecules will strongly interact with these endgroups upon hydration of the polymer.

This effect has already been observed by Aoshimaet al. in the 1990s for their vinyl ether based poly-macromonomers [38] . Replacementof themethyl moietiesat the end of the oligomeric EO side chains with ethylgroups resulted in T cp s that were decreased by 4050 C(entrance and in Fig. 7 ). Similar results were found forthe methacrylate based systems of Ishizone et al. althoughthe T cp s were only decreased by around 2025 C whencompared to the corresponding values from polymers car-rying methyl end groups (entrance and in Fig. 7 ) [40] .


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Fig. 7. Effect of the hydrophilicity of the side chain end groups on the cloud point temperatures of aqueous solutions of various PEO based poly-macromonomers and schematic representation of the polymer structures. Data taken from Ref. [38] ( , ), [39] ( ), [40] ( ) . Lines are added toguide the eye.

PEtEO3 MA was also synthesized via RAFT polymerization,and the T cp s of buffered polymer solutions ( c =5mg mL 1 )at pH 4, 7 and 10 are in agreement with values obtained forthe anionically synthesized polymers [48] . A similar inu-ence can be observed when PEtEO 2 A, which has recentlybeen obtained by NMP [49] , is compared to PmEO 2 A [42] :the T cp of a 0.5 wt% aqueous solution of PEtEO 2 A was foundto be 13 C, which is 15 C lower than the T cp of PmEO 2 Aat the same concentration. In addition, the T cp of PEtEO2 Asolutions was slightly increased to 16 C when the poly-mer was synthesized by FRP, which is a result of the largehydrophobic backbone end group derived from NMP syn-thesis.

On the other hand, replacement of the side chain endgroups by a proton results in hydroxyl end groups thatsignicantly enhance the hydrophilicity of the macro-molecules so that the corresponding PHEO n MA andPHEOn A are water soluble in the entire temperature rangeof liquid water at ambient pressure [39] . In agreement tothat the T cp of an aqueous solution of PmEO 4 St is increasedfrom 39 Cto64 CforPHEO 4 St [42] . Since this type of poly-mers was synthesized by NMP, the large hydrophobic endgroup of the polymer backbone caused an increase in T cpwith increasing molar mass (entrance and in Fig. 6 ).

4.3. Effect of backbone end group

The application of living and controlled polymeriza-tion techniques during the synthesis of thermo-responsivepolymers provides the opportunity to obtain polymericstructures withwell-dened endgroups.Thefact thatsomeof those end groups can be used for further modicationof the obtained polymer with hydrophilic or hydropho-bic moieties enables the investigation of their effect uponthe LCST behavior. Since there are only two end groups atthe polymer backbone (at the and at the chain end) itseems to be quite obvious that the impact of the backboneend groups on the solubility behavior of the entire macro-molecule will be much less pronounced than the effectof the side chain end groups. However, the effect of end

groups will be more signicant for polymers with lowermolar masses [50] .

Theato and co-workers recently applied the RAFTpolymerization technique to obtain PmEO 4 MA from acommercially available methacrylate monomer havingapproximately 4 repeating units of EO [51] . The obtainedpolymer (DP 10) was subsequently modied at both as well as end groups resulting in a library of PmEO 4 MA having the same DP but a large variety of end groups (see Table 1 ). T cp of the aqueous polymersolutions ( c =10mg mL 1 ) having the same end groupwas found to decrease when the hydrophobicity of the end group was increased following the order: CH 3 ,C16 H33 , CH2 CH2 C6 F13 . When CH 3 was kept constant as end group, the same trend could be followed when thehydrophobicity of the end group was increased in thefollowingorder: ammonium, PEG, C 3 H7 , C16 H33 , C8 F17 CH2 ,(C18 H37 )2 . However,by introductionof two veryhydropho-bic moieties at both as well as end group deviationsfrom this series could be observed, which was attributedto the formation of micellar structures below the cloudpoint as could be conrmed by DLS measurements. Aro-matic end groups, such as C 6 F5 O, SCSPh and an azodye, were found to decrease T cp more than long aliphaticchains, which was proposed to be a result of the rigid-ity of those structures making them even more difcultto hydrate by the solvent molecules. In particular, the azodye (last entrance in Table 1 ) represents a very interestingend functionality since irradiation with UV light induces acistrans isomerization of the azo moiety that is accompa-nied by an increased dipole moment and, thus, elevatedT cp (by 211 C, c =10mg mL 1 ) [50] . This UV triggeredsolubility change was found to be fully reversible after re-isomerization of the azo functionality.

Soeriyadi et al. functionalized the vinylic polymer endgroup of PmEO 2 MA, which was synthesized via catalyticchain transfer polymerization, with 2-mercaptoethanol, 1-dodecanethiol and benzyl mercaptan and found a similarinuence of the hydrophobicity of the polymer end group[52] .


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Table 1Effect of various end groups upon the cloud point temperature of aqueous solutions of PmEO 4 MA [51] .

S

OO

O

4

SR 2

CNO

R 1

R1 R2 T cp [ C]c =1wt%

C6 F5 O SCSPh 42.8CH3 (OCH2 CH2 )11 NH CH3 62.7

C16 H33 62.4CH2 CH2 C6 F13 62.1

C3 H7 NH CH3 58.3C16 H33 50.1CH2 CH2 C6 F13 49.7

C16 H33 NH CH3 53.6C16 H33 45.8CH2 CH2 C6 F13 44.7

C8 F17 CH2 NH CH3 50.9C16 H33 51.7

CH2 CH2 C6 F13 50.3(C18 H37 )2 NH CH3 48.9

CH2 CH2 C6 F13 48.9

HN

N +

CH3 66.3

C16 H33 62.5CH2 CH2 C6 F13 62.1

HN

HN

O

NN

CH3 49.1

4.4. Block copolymers composed of twothermo-responsive parts

In cases where both blocks of a block copolymer con-sist of thermo-responsive polymers, the aqueous solutionmight display a single cloud point at an intermediate tem-perature, or bothblocks might collapseindependently fromone another. Both behaviors have been observed for poly-mers based on EO macromonomers and will be discussedin the following section.

Ishizone and co-workers reported that the aqueoussolution of a block copolymer of PmEO 2 MA and PmEO 3 MAdisplays a single cloud point at 39 C, which is in betweenthe T cp s of both independent blocks (at 26 C and 52 C,respectively) [39] . Yamamoto et al. also observed sin-gle cloud points of 0.1wt% aqueous solutions of twoPmEO 2 MA-b-PmEO 3 MA block copolymers that were syn-thesizedby ATRP [45] . Althoughboth T cp s were in betweenthe cloud point temperatures of the homopolymers, theexact value for the block copolymer containing a longerPmEO 3 MA block was found to be higher than the expectedvalue for a statistical copolymer of both monomers. Thisdeviation is most likely causedby theformation of micellesthat consist of the collapsed PmEO 2 MA block in the coreand still water soluble PmEO 3 MA in the shell below T cp .Indeed,DLSconrmedthe presenceof aggregates thatwereabout 100 nm in size already before T cp (which was deter-mined by turbidimetry) was reached. Further increase intemperature resulted in the formation of larger aggregates

(>1 m) above the cloud point indicative of a full collapseof the block copolymer.

Similar observations were made by Zhao et al. for blockcopolymers of PmEO n St having EO side chains of vary-ing DP from 3 to 5 [43] . The aqueous solutions of 5 blockcopolymers (PmEO 3 St- b-PmEO 4 St, PmEO 3 St- b-PmEO 5 St,PmEO 4 St- b-PmEO 5 St) displayed phase transition temper-atures that coincide with values that were calculated fromthe T cp s of the corresponding homopolymers using a for-mula that takes into account the degree of polymerization(DP) of each block:

T cp =DP 1

DP 1 + DP 2 T cp , 1 +

DP 2DP 1 + DP 2

T cp ,2 (1)

In contrast, aqueous solutionsof various mixtures of thehomopolymers showed phase separation at the T cp sof thehomopolymer with the lowest phase transition tempera-ture.

Alexander and co-workers prepared hybrid blockcopolymers that consist of a statistical copolymerof EtEO 3 MA with mEO 8.5 MA as rst block and of PmEO 8.5 MA homopolymersas secondshorter block usingAGET ATRP [53] . Aqueous solutions of the polymers(c =1.5mg mL 1 ) revealed T cp s that were increased by47 C when compared to those of the rst block. The dif-ference was found to beenlargedupon increaseof theblocklengthof thesecond, more hydrophilic block dueto thefor-mation of micelle-like assemblies of 40100nm radii uponcollapse of the rst thermo-responsive block, as could be


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conrmed by DLS, TEM as well as uorescent probing withpyrene. The addition of sodium sulfate as kosmotrope tothose systems resulted in changes of the micellar structureleading to a sharp burst release of carboxyuorescein thatwas encapsulated into the core of the micelles.

On the other hand, block copolymers where the cloudpoints of the individual blocks, PmEO 3 A (T cp = 62 C) andPmEO

3St ( T

cp= 13 C), are further apart underwent a series

of phase transitions in water, as demonstratedin Fig.8 [54] .Heating of a 1 wt% aqueous solutionofPmEO 3 A-b-PmEO 3 Stled rst to the collapse of the more hydrophobic PmEO 3 Stblock at transition temperatures that were up to 20 Chigher when compared to the homopolymer PmEO 3 St.This rst phase transition temperature was found to bedecreasedwith increasinglengthof thePmEO 3 Stblockand,thus, came closer to the T cp of the PmEO 3 St homopolymer.Upon further increase in temperature to around 2539 C,the turbid solutions became clear, and micelles consist-ing of the collapsed PmEO 3 St block in the core and thestill water soluble PmEO 3 A block in the shell were formed.The hydrodynamic diameter of these spherical micelleswith narrow size distributions was increased from 27nmto 58nm with increasing length of the PmEO 3 St block, ascould be shown by DLS measurements. Further increasein temperature towards 4555 C resulted in a collapse of the PmEO 3 A blocks and, therefore, the formation of largeaggregates. Thissecond cloud pointwas lowered comparedto the PmEO 3 A homopolymer and, again, increasing lengthof the PmEO 3 St block resulted in a shift to lower tem-peratures. This fact can nicely be explained by the largerhydrophobicityof thelonger, collapsed PmEO 3 Stblock. Theindependent collapse of both blockscould also be followedby 1 H NMR spectroscopy in D 2 O revealing a broadening of the aromatic signals of the PmEO

3St around the rst tran-

sition temperature and a broadening of the peak derivedfrom the methyl side chain end group of PmEO 3 A at thesecond cloud point.

4.5. Statistical copolymers of monomers with different length of oligomeric EO

The homopolymers described above already cover awidetemperaturerange in their responsive behavior.How-ever, sometimes the monomers have to be synthesizedsince not all of them are commercially available. In addi-tion, specic monomers result in polymers with a denedT

cp and intermediate temperatures are not accessibleusing

homopolymers. An easy and straightforward method tomore accurately tune the phase transition temperature of PmEO n MA is therefore the statistical copolymerization of two monomers with different EO chain length. The result-ing copolymers will display thermo-responsive propertiesin a temperature interval ranging in between the T cp s of both homopolymers. Most studies involve the copolymer-ization of mEO 2 MA with monomers having a longer EOchain so that the rather low phase transition temperatureof PmEO 2 MA is increased. The increased PEO chain lengthof the second monomer enhances the hydrophilicity of thecopolymer, which is reectedby thesteeper decreaseof theT

cp values with increasing mEO

2MA content, as displayed

in Fig. 9 .

Kitanoet al. used small amounts (310%) of mEO 8.5 MAandmEO 22 MA as comonomers during free radical copoly-merization of mEO 2 MA in order to investigate the effectof added macrocycles that might be able to incorporate afew longer pendant PEO chains [55] . Turbidity measure-ments of 0.1wt%aqueoussolutionsrevealed increased T cp swith increasing comonomer content as well as increas-ing length of the oligomeric EO chain of the comonomer(entrance and in Fig. 9 ). Addition of SCX6, a macrocy-cle that contains six anionically charged sulfonate groups,was found to raise the phase transition temperature byup to 12 C due to the fact that the pendant PEO chainswere enclosed into the macrocyle obstructing the coil toglobule transition of the polymer. -Cyclodextrin had asimilar effect, although much less pronounced, whereashydrogen bonding of neighbouring hydroxyl groups of -cyclodextrin caused a slight decrease of T cp .

A study that covered a wider range of copolymercompositions was carried out by Lutz and Hoth whocopolymerizedmEO 2 MA withmEO 8.5 MA using ATRP [56] .Turbidity measurements ( c =3mg mL 1 ) i n H

2O revealed a

linear increase of T cp with increasing mEO 8.5 MA contentfor polymers that contained up to 30% mEO 8.5 MA so thatthe phase transition temperature could be precisely tunedbetween 28 C and 59 C (entrance in Fig. 9). The poly-mer concentration was found to have only a small effectsince T cp only varied by a few degrees in a concentrationrange from 1mgmL 1 to 20mg mL 1 . Subsequently, thecopolymer containing 5% mEO 8.5 MA was further inves-tigated due to the fact that it exhibited LCST behavior ata similar temperature as PNiPAm [59] . The turbidity curveof the aqueous solution was found to be sharper with lesshysteresis between heating and cooling compared to PNi-PAm. Addition of sodiumchloride( c 0.2mol L 1 ) causedasimilar small salting out effect of about 2 C for both poly-mers. In addition, the dependence of T cp of the polymersolutionson theconcentrationwas foundtobeverysimilar,decreasing by 3 C for concentration between 1 mgmL 1

and 10mg mL 1 . Theeffect of thedegreeof polymerizationupon T cp was found to be even smaller for the PmEO n MAcopolymer than for PNiPAm when varying the DP from 25to 100. DLS experiments of 1mg mL 1 aqueous solutionswere applied for thePmEO n MA copolymerscontaining 10%and 20% mEO 8.5 MA, respectively [60] . Below the cloudpoint of the solution, both polymers showed the coexis-tence of particles having a hydrodynamic radius of around4nm, which most likely represent individually solvatedpolymer coils, with larger particles of about 150 nm size,which contain less than 0.01% of the polymer chains andwere attributed to loose aggregates. Increase of temper-ature resulted in a slight shrinkage of the particles, whichwas attributed to thepartial loss of hydrationwater, beforethe individual polymer chains formed larger aggregates of 300 nm and 200nm upon their coil to globule transitionat T cp . The reason for the fast reversibility of the phasetransition was proposed to be the fact that the polymeraggregates are only held together by weak van der Waalsinteractions instead of intramolecular hydrogen bonds asbeing the case for PNiPAm. With respect to further use of this type of polymer in biological systems, the biodegrad-ability of the polymer was enhanced by copolymerization


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Fig. 8. Conformational changes of a copolymer consisting of two thermo-responsive parts in aqueous solution upon temperature variation.

with BMDO [57] . Since BMDO was incorporated mainly viaa ring opening mechanism, the resulting ester function-alities inside the methacrylate backbone could either becleaved enzymatically or under basic conditions. As dis-played in Fig. 9 (entrance ) inclusion of the hydrophobicBMDO resulted in a slight decrease of the observed T cpvalues ( c =3mg mL 1 ) when compared to the copolymerwithout BMDO.

Copolymerization of mEO 2 MA with a monomer con-taining a shorter oligomeric EO chain, mEO 4.5 MA, resultsin decreased hydrophilicity of the copolymer at the samemEO 2 MA content as depicted in Fig. 9 (entrance ) [58] .Nevertheless, turbidity measurements at a polymer con-centration of 3 mg mL 1 revealed that T cp can be tunedin a similar way in between the cloud point tempera-tures of PmEO 2 MA and PmEO 4.5 MA at 28 C and 65 C,respectively. PBS was shown to have a small salting outeffect since the observed T cp s were 34 C lower than inwater. In addition, DLS measurements revealed the pres-ence of free hydrated polymer chainswith a hydrodynamic

diameterof3.5nm belowthe cloudpointand theformationof mesoglobules of sizes around 250nm upon the tem-perature induced collapse of the polymer. A comparisonbetween ATRP and free radical polymerization as synthe-sis techniques revealed a T cp that was lowered by 2 C forthe conventionally synthesized copolymer, which may bea result of its higher molar mass.

Yamamoto et al. applied ATRP in order to obtainwell-dened copolymers of mEO 2 MA with mEO 3 MA [45] .Investigation of the thermal properties of their aqueoussolutions ( c = 0.1wt%) by turbidity measurements showeda linear increase of T cp with increasing content of mEO 3 MAfrom 26 Cto53 C. As demonstrated in Fig. 9 (entrance ),the shorter oligomeric EO chain of mEO 3 MA results in adecreased hydrophilicity of the statistical copolymers and,thus, lower cloudpointswhencomparedto thecopolymersdescribed above.

Besides the copolymerization of mEO 2 MA, also thecopolymerization of other oligoEO based monomers withone another was used in order to netune the LCST of

Fig. 9. Cloud point temperatures observed for aqueous solutions of statistical copolymers of mEO 2 MA with other monomers containing oligomeric EOchains (mEO n MA). Data taken from Ref. [55] ( , ), [56] ( ), [57] ( ), [58] ( ), [45] ( ).


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the corresponding homopolymers. The AGET ATR copoly-merization of EtEO 3 MA with mEO 8.5 MA revealed a seriesof statistical copolymers with mEO 8.5 MA content of upto 20mol% whose aqueous solutions ( c =1.5mg mL 1 ) dis-played thermo-responsivebehavior in a temperature rangefrom 24 C to 42.5 C [53] . The T cp s measured by tur-bidimetry increased linearly with the content of the morehydrophilic mEO

8.5MA in the polymer. In addition, the

effect of varioussaltadditives on thedemixingtemperatureof aqueous polymer solutionswas investigatedfor salt con-centrations up to 0.5mol L 1 . The observed effects for bothsodium as well as potassium salts followed the Hofmeis-ter series of anions: SCN >ClO4 > I > Cl >OAc > SO4 2 ,the rst 3 (kosmotropes) causing a salting-in effect andthe latter 3 (chaotropes) causing a salting-out effect. Theadditives altered the T cp of PEOn MA solution (33 C) overa broad temperature range from 3 C for Na 2 SO4 to 46 Cfor NaSCN, causing stronger effects than for aqueous solu-tions of PNiPAm, which might be due to the highly orderedwater structure around PEO or crownether type interac-tions between PEO and the cations.

The approach has been extended to acrylate basedmonomers, mEO 2 A and mEO 8.5 A, which were copoly-merized using ATRP by Laschewsky and co-workers [61] .Aqueous solutions ( c =3mg mL 1 ) of the obtained sta-tistical copolymers containing up to 30 mol% mEO 8.5 Aexhibited phase transitions in a temperature range from9 C to 50 C, as was followed by turbidity measurements.Also in this case increasing the mole fraction of themore hydrophilic mEO 8.5 A was found to increase thehydrophilicity of the copolymer resulting in elevated T cpof the aqueous solution. Sodium chloride was shown tocause a salting out effect of up to 4 C for salt concentra-tions of up to 12gL 1 . The molar mass of the copolymerhad only a very small effect on its coil to globule tran-sition since T cp s only deviated by 1 C in the M n regionof 10,00030,000 g mol 1 . Recently, Cornelissen and co-workers reported copolymers of mEO 1 A and mEO 2 A withan equal content of both monomers that were preparedby ATRP [62] . Turbidimetry in PBS ( c = 0.5 wt%) revealedT cp s between 25 C and 35 C that decreased with increas-ingmolar mass of thepolymer( M n = 900030,000g mol 1 ).Increasing concentration of polymer and added amountof NaCl resulted in decreased T cp s. Coupling of thethermo-responsive polymers to enhanced green uores-cent protein (EGFP) induced the formation of micellarstructures above the coil to gobule transition temperatureof the thermo-responsive copolymer, as shown by DLS aswell as TEM measurements. In addition, random copoly-mers of mEO 1 A and HEO 1 A with varying composition weresynthesized by RAFT polymerization [63] as well as NMP[64] . The T cp of aqueous polymer solutions ( c =0.5wt%)could be tuned from 2 Cto60 C and was found to increasewith increasing content of the more hydrophilic HEO 1 Amonomer. Increasing the molar mass resulted in lower T cpvalues, whereas removal of one benzylic end group of thepolymer signicantly increased the coil to globule transi-tion temperature of the polymers. Hoogenboom, Keul andMoeller recently reported the copolymerization of EtEO 2 Awith HEO

1A by FRP as well as NMP [49] . The T

cps of aque-

ous polymer solutions ( c = 0.5 wt%) were found to increase

from 13 C for PEtEO2 A to 48 C for a copolymer with 75%HEO1 A content with increasing mole fraction of the morehydrophilic HEO 1 A in the copolymer, as determined byturbidimetry. Comparison of the polymers that were syn-thesized by NMP and FRP revealed 3 C higher T cp valuesfor the free radically polymerized copolymers, which wasascribed to the hydrophobicity of the nitroxide end groupof the polymers obtained by NMP. Increasing amount of HEO1 A in the copolymer resulted in a larger heating cool-ing hysteresis, whereas the PDI of the polymers had nosignicant effect on the phase transition.

4.6. Copolymers with other monomers

Besides the copolymerization of oligoEO basedmonomers with one another, these monomers mayalso be copolymerized with other monomers that areable to provide additional interesting features to theresulting thermo-responsive copolymer. Various groupscombined PEO with the most well-known LCST polymer,PNiPAm, either by grafting PEO onto PNiPAm [65] , bycopolymerization of mEO n MA [66] or HEOn MA [67] withthe NiPAm monomer, or by terpolymerization with dode-cyl methacrylate [68] . In these works EO n MA is mainlyregarded as comonomer in order to investigate its effecton the phase transition temperature of PNiPAm sinceincreasing length of the PEO chain as well as increasingmole fraction of EO n MA in the copolymers resulted in anincreased T cp of the aqueous PNiPAm solutions due to thehydrophilicity of the pendant PEO chains. In contrast, alsoa wide range of other monomers has been copolymerizedwith EO n MA in order to investigate their effect uponthe phase transition temperatures of PEO n MA aqueoussolutions ( Fig. 10 ).

4.6.1. Copolymers with hydrophobic monomersAli and Stover applied ATRP for the synthesis of

PHEO 7.3 MA [69] . Due to thehydrophilicity of thehydroxylend groups of the pendant side chains, the PHEO 7.3 MAhomopolymers were water soluble in the investigatedtemperature range up to 80 C. However, copolymeriza-tion with MMA rendered them more hydrophobic sothat the resulting copolymers displayed LCST behavior inaqueous solutions of 1 wt% concentration. Turbidity mea-surements revealed increasing T cp from 43 Cto56 C withincreasing mole fraction of PHEO 7.3 MA in the copoly-mers (2440%). Vincent and co-workers reported a rangeof copolymers of mEO 5.5 MA with BuMA that were syn-thesized by free radical polymerization to show LCSTbehavior in 1 wt% aqueous solution at temperatures rang-ing from 45.8 C to 53.6 C, but their research was focusedon the investigation of the formation of micellar struc-tures at room temperaturerather than on LCST phenomena[70] .

4.6.2. Copolymers with ionizable monomers4.6.2.1. Cationically charged comonomers. As already dis-cussed in Section 4.5 , it is possible to tune the phaseseparation temperature of aqueous solutions of copoly-mers that arecomposedof twodifferent monomers (whosehomopolymers also display LCST behavior) between the


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Fig. 10. Schematic representation of monomers that have been applied for copolymerization with EO n MA orEO n A.

T cp s of the homopolymers. This approach has been fol-lowed by Fournier et al. who prepared a series of gradientcopolymers of DMAEMA with mEO 4.5 MA using the RAFTpolymerization technique ( r DMAEMA = 0.93; r mEO

4 .5 MA =

0.66) [47] . Turbidity measurements of aqueous solutions(c =5mg mL 1 ) at pH 4 only revealed LCST behavior for acopolymer with a mEO 4.5 MA content of 90% as well asthe PmEO 4.5 MA homopolymer due to the fact that theamine function of DMAEMA was protonated and, there-fore,chargedresultingin a highhydrophilicity. However,atpH 7 theDMAEMA moieties were only partially protonatedand the T cp of the aqueous solutions was found to increaselinearly from 47 C (PDMAEMA) to 71 C (PmEO 4.5 MA)with the mole fraction of mEO 4.5 MA in the copolymer.UnderbasicconditionsatpH10, allamine functionsexistedin an unprotonated state resulting in a decrease of theT cp values by 10 C. A similar study was performed byYamamoto et al. who applied turbidity measurements of aqueous solutions ( c = 0.3 wt%) of copolymers of mEO 2 MAwith DMAEMA that were synthesized by ATRP [71] . Forthis copolymer system, the coil to globule transition tem-perature of PDMAEMA is higher than that of PmEO 2 MAand, thus increasing content of DMAEMA (up to 26mol%)was found to increase the T cp of the aqueous copoly-mer solution. This effect was more pronounced at pH7(T cp = 22.548 C) than at pH 9 ( T cp = 22.529 C) since theamino functions of the DMAEMA are partially protonatedat pH 7 resulting in a further increase of hydrophilicity.

Tenhu and co-workers investigated the formation of complexes between two oppositely charged polyions,namely PEO- b-poly(sodium methacrylate) as anionand poly(methacryl oxyethyl trimethylammoniumchloride) (PTMAEMA +) as polycation [72] . Turbiditymeasurements as well as DLS revealed increasing T cp(555 C) of the aqueous solutions of the polymer com-plexes ( c =0.251mg mL 1 ) with increasing content of sodium chloride ( c = 0.50.6 mol L 1 ) and sodium nitrate(c = 0.50.6 mol L 1 ). When a statistical copolymer of TMAEMA+ with mEO 4.5 MA was used as polycation, thehydrophilicity of the resulting complex was increasedresulting in higher T cp than for aqueous solutions of complexes without grafted oligoEO.

4.6.2.2. Anionically charged comonomers. The copolymer-ization of mEO n MA with MAA results in polymers whosesolubility in water is not only dependent on the temper-ature but also on the pH of the solution since MAA asBrnsted acid will be deprotonatedunder basic conditions.

Under acidic conditions, theprotonated MAA moieties willserve as hydrogen bond donorsandcanoccupy hydrogenbond accepting sites of the oligoEO chains. These compet-ing interactions result in the formation of less hydrogenbonds with surrounding water molecules that keep thepolymer in solution and, thus, LCST transitions at lowertemperatures, as illustrated in Fig. 11 . In addition, thedeprotonated ionic form is much more hydrophilic com-pared to the protonated form.

Jones et al. varied the comonomer ratio between 0and 100% during the free radical copolymerization of mEO 4.5 MA and MAA (Fig. 11 ) [73] . The aqueous solu-tions ( c = 3 wt%) of the resulting copolymers containingup to 5% mEO 4.5 MA displayed T cp s that were decreasingfrom 68 Cto12 C with increasing content of mEO 4.5 MA.Polymers that contained 624% mEO 4.5 MA were foundto be insoluble in water due to the formation of hydro-gen bonds between the protonated MAA moieties withether functions of the oligoEO side chains. Further increaseof the mole fraction of mEO 4.5 MA rendered them watersoluble again, and the T cp was found to increase withthe mEO 4.5 MA content up to 61 C for PmEO 4.5 MA. Asimilar behavior could be observed for copolymers of MAA with longer oligoEO side chains, mEO 9 MA andmEO 22.7 MA, but the phase transition temperatures wereelevated at constant ether to acid ratios due to the increas-ing hydrophilicity of the longer oligimeric EO chains. Inaddition, the demixing temperatures of aqueous solutionswere found to be increased by 15 C with increasing pHvalue from 2 to 5 because of a partial deprotonation of theMAA moieties. Increasing polymer concentration in a rangefrom 0.5 to 3 wt% was found to lower the T cp by 6 C, whichwaspartiallyattributed to a decreasedpH valueof themoreconcentrated solutions. The addition of urea (0.8 mol L 1 ),which disrupts the intramolecular hydrogen bonding,resulted in T cp s that were elevated by 4 C. Introductionof more hydrophobic comonomers, such as BuMA or lau-ryl methacrylate, to the copolymers resulted in an almostlinear decrease of the T cp values with increasing content of hydrophobic comonomer (0.8 C per mol%). Surprisingly,copolymers with more hydrophilic glycidyl methacrylateshowed a similar trend, although less pronounced (0.5 Cper mol%) presumably also due to intramolecular hydro-genbonding.Schubert andco-workers performed a similarstudy applying copolymers of MAA with PmEO 8.5 MAand PmEO 22 MA, respectively, that were synthesizedby RAFT polymerization [48] . Turbidity measurements(c =5mg mL 1 ) at pH2 and pH 4 revealed T cp s that were


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Fig.11. Inuence of theionization of MAA (left) andthe copolymercomposition (right) upon intramolecular hydrogen bonding in copolymers of MAA andEOn MA.Reproduced from Jones et al. [73] by permission of John Wiley & Sons Ltd., UK.

increasing with the mEO n MA content of the copolymersfrom 30 to 90 C for P(mEO 8.5 MA-stat -MAA) and from38 to 95 C for P(mEO 22 MA-stat -MAA), respectively. Poly-mers with mole fractions of mEO 22 MA larger than 40%were found to be water soluble up to 100 C. Increase of the pH value to pH7 and pH10 resulted in deprotonationof the MAA moieties, which rendered the polymers morehydrophilic and, thus, water soluble.

Instead of direct copolymerization with MAA, simi-lar copolymers were obtained by hydrolysis of statisticalcopolymers of mEO 2 MA and tert -butyl methacrylate thatwere synthesized by ATRP [71] . The thermal behavior of thecopolymer solutionswas affected in a similar fashion asdescribed above.As long as theMAA moietiesof thecopoly-mer were protonated in de-ionized water, increasing themole fraction of MAA up to 28% resulted in decreased T cp s(2715 C). Partial deprotonation resulted in the oppositeeffect, as demonstrated by turbidity measurements at pH7 and 9. The tremendous inuence of MAA was revealedduring investigation of the solubility properties of a ter-polymer that consisted of 89% mEO 2 MA, 6% MAA and 5%DMAEMA. Its aqueous solution exhibited a T cp of 34 C atpH4 and of 40 C at pH 7, whereas it was water soluble upto 80 C at pH 9. The lowered T cp under acidic conditionshints towards the fact that the hydrogen bonding, whichis caused by the protonation of the MAA, dominates overthe increased hydrophilicity of the protonated DMAEMAmoieties.

Jiang and Zhao investigated in detail the effect of pH upon the phase transition temperatures of aqueoussolutions ( c =0.2wt%) of a mEO 2 MA based copolymer con-taining 13mol% MAA [74] . The T cp s were in the range of 2460 C and increased with increasing pH value from 4 to6.8. A block copolymer with linear PEO was found to formmicelles thatconsistedof the collapsed thermo-responsiveblock in the core and hydrophilic PEO in the shell when theaqueous solution was heated above its cloud point. Thesemicelles could be disrupted by increase of thepH value andreformed either upon further elevation of the temperature

or by decrease of the pH value, as was conrmed by DLSas well as uorescence spectroscopy and is illustrated inFig. 12 .

In addition, a similar system that undergoes reversiblemicellization at two different temperatures and is trig-gered by UV irradiation was developed by Zhao andco-workers [75] . A statistical copolymer of mEO 3 A with13mol% of o-nitrobenzyl acrylate (NBA) exhibited a T cpin 0.2wt% aqueous solution at 18.5 C, which is low-ered by 17.5 C when compared to PmEO 3 A due to thehydrophobicity of NBA. Upon irradiation with UV light, o-nitrosobenzaldehyde is cleaved and an acrylic acid moietyis formed. The aqueous solution of the resulting copoly-mer displayed a higher T cp at 30 C due to the absenceof the hydrophobic aromatic moieties. Utilization of aPEO macroinitiator for the synthesis of the mEO 3 A basedpolymer resulted in a block copolymer that undergoesreversible formation of micelles above the T cp of thethermo-responsive block, as demonstrated by DLS anduorescence spectroscopy. Cleavage of the NBA moietycaused disruption of the micelles since the resulting acidcontaining PmEO 3 A block was still below its phase transi-tion temperature. Upon increase of temperature towards36 C, the coil to globule transition of this newly formedthermo-responsive block was reached and the micelleswere re-formed.

4.6.3. Copolymers with dye containing monomersPietsch et al. developed a dual polymeric sensor for

temperature and pH value based on the LCST behavior of PmEO 2 MA in water [76] . After detailed kinetic studies onthe copolymerization of mEO 2 MA and mEO 22 MA [77] ,RAFTpolymerizationwas applied for the copolymerizationof mEO 2 MA with 5mol% of a monomer that is based on asolvatochromic azo dye, disperse red 1. Below the T cp ofanaqueous solution (pH 1, c =1mg mL 1 ) of the copolymer at17 C, the dye is in contact with water and, thus, in a polarenvironment. Upon precipitation of the PmEO

2MA, the

hydrating water molecules are expelled from the polymer


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Fig. 12. Reversible formation of micellar structures from pH and temperature responsive block copolymers.

and the azo dye is directly protonated in its nonpolarenvironment, which induces a color change ( Fig. 13 ).Since the protonation of DR1 is not possible under basicconditions, this color change could not be observed atpH> 7, as quantied by UV/vis spectroscopy of the aque-

ous solutions in a temperature range around the cloudpoint. Unfortunately, the high hydrophilicity, even aboveits coil to globule transition at 92 C, of a tercopolymerthat contained 45% mEO 2 MA, 50% mEO 22 MA and 5%DR1 prevented the extension of this interesting concepttowards an elevated temperature range.

A similar sensor that is based on the uorescence of pyrene wasrealized by RAFT copolymerization of mEO 2 MAwith 5 mol% pyrene-1-ylmethyl methacrylate [78] . Belowthe T cp of the aqueous polymer solution at 18 C the

hydrophilic polymeric structure facilitates the formationof excimers between the pyrene moieties. Upon the coilto globule transition of the thermo-responsive polymer,the hydrophobic surrounding of the pyrene decreased theexcimer formation, which was manifested in a decrease of

the excimer uorescence band between 11

C and 21

C.DLS measurements at varying temperatures conrmed theformation of particles with sizes about 180 nm in this tem-perature range.

4.7. Grafted structures

The possibility to use functional initiators for the ATRPof mEO n MA has been applied for the synthesis of combandgraft copolymers that contain PmEO n MA as side chains

Fig. 13. Color change of an aqueous solution containing a PmEO 2 MA

based temperature sensor upon coil to globule transition of the polymeric sensor.Reproduced from Pietsch et al. [76] by permission of John Wiley & Sons Ltd., UK.


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Fig. 14. Schematic representation of grafted structures with PmEO n MA side chains.

via the grafting from approach. As depicted in Fig. 14 ,methacrylate [45,71] as well as ethyl cellulose [79] basedpolymeric backbones were functionalized with an ATRPinitiator that wassubsequently used to graft thePmEO n MAside chains from these polymeric backbones.

Matyjaszewski and co-workers prepared a series of comb polymers with PmEO 2 MA side chains of varying DPof 25, 50 and 115 from the same polymethacrylate back-bone ( Fig. 14a) [80] . The aqueous solutions of the combpolymers ( c = 0.3wt%) were investigated viaturbidity mea-surements and revealed similar T cp values of 22 C duringthe heating run, which is slightly lower compared to linearPmEO 2 MA. The heating-cooling hysteresis of the turbid-ity curves was signicantly smaller for the polymers withlonger side chains hinting towards a delayed collapse of the PmEO 2 MA parts close to the polymeric backbone dueto sterichindrance. Thegraftingdensity wasshownto haveno effect on the phase separation temperature of aqueouspolymer solutions since copolymers with varying amountsof MMA (2470 mol%) in the polymer backbone exhib-ited similar T cp in water. However, DLSrevealed increasingagglomerate sizes of the precipitated collapsed polymerglobules above T cp with increasing grafting density. Whenthe side chains of the polymeric structure consisted of sta-tistical copolymers of mEO 2 MA and mEO 3 MA, the T cp of the aqueous polymer solutions was found to increase withthe content of the more hydrophilic mEO 3 MA in a simi-lar fashion to their linear analogues. Again, the demixingtemperature of a solution of comb shaped PmEO 3 MA of 45 C was found to be lowered by 6 C when comparedto linear PmEO 3 MA. Comb polymers that contained sidechains consisting of block copolymers with PmEO 2 MA inthe inner sphere and PmEO 3 MA on the outside exhib-ited thermo-responsive behavior at 50 C, which is higherthan expected for a statistical copolymer of the same com-position, because the polymer chains were still held insolution by the more hydrophilic PmEO 3 MA parts. DLSmeasurements indeed revealeda shrinkingof thedissolvedmacromolecules at temperatures lower than T cp hintingtowards a collapse of the inner parts (PmEO 2 MA) of the

polymer structure. In contrast, such comb polymers werefound to aggregate at 26 C when the more hydrophobicPmEO 2 MA was located in the shell of the structure. In thiscase, DLS revealed a shrinking of the polymer globules attemperatureshigher than T cp duetothecoiltoglobuletran-sitionof thePmEO 3 MA segmentsat elevated temperatures.

In addition, the composition of the side chains wasfurther varied by copolymerization with pH respon-sive monomers, such as MAA and DMAEMA ( Fig. 14 b)[71] . In general, the thermo-responsive properties of thePmEO 2 MA were affected in a similar fashion as the lin-ear anologues (see Section 4.6 ), but some additionaleffectsof the comb structure will be discussed here. The inu-ence of the intramolecular hydrogen bonding that ispresent in the MAA containing copolymers was foundto be enhanced when compared to linear copolymers of PmEO 2 MA andMAA dueto theproximityof hydrogenbondacceptingether moietiesand hydrogenbond donating pro-tonated MAA moieties at pH 7 in the dense bottle brushlike structure. On the other hand, T cp s of aqueous solu-tions of comb like copolymers of mEO 2 MA and DMAEMAshowed the same trend as their linear analogues at pH 7,but the copolymer composition did not affect the phasetransition temperature at pH 9. In addition, the thermo-responsiveness of a comb polymer, whose side chainsrepresented a terpolymer of mEO 2 MA, MAA and DMAEMA,was less affected by changes in pH than its linear ana-log. These observations were proposed to be a result of theadditional electrostatic association of theside chains inthe complex structure of the comb polymer that made itsthermo-responsive behavior less sensitive to composition.

Liu and co-workers grafted PmEO 4.5 MA from a func-tionalized ethyl cellulose backbone in grafting densitiesof 2% and 20% ( Fig. 14 c) [79] . Turbidity measurementsof aqueous graft copolymer solutions ( c =1mg mL 1 )revealed T cp s of around 64 and 67 C for 20% and 2%grafting density, respectively. TEM as well as DLS investi-gations showed the presence of micellar structures withthe hydrophilic PmEO 4.5 MA as shell already below thecloud point (which explains the rather small difference of


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the observed T cp s). These micelles (100nm and 65nm)further agglomerated upon coil to globule transition of thePmEO 4.5 MA segments and formed particlesof slightlylarger size (175and 120 nm, respectively).

4.8. Towards bio-medical applications

Theexcellent thermo-responsiveproperties gaverise tomany initial studies aiming for applications of in particularPmEO n MA based polymers that were recently reviewed ina highlight by Lutz [14] and, thus, are only briey summa-rized here.

PmEO n MA based polymers have been applied for thefabrication of functionalized surfaces: functionalization of silica monoliths resulted in suitable column materials fortheseparationof steroids andproteins [81] . Functionalizedgold or glass surfaces allowed switching of cell adhesion:above the phase transition temperature of the polymer inPBS at 35 C, the surface behaved hydrophobic and couldbe used to grow L929 mouse broblast [82,83] and MCF-7breast cancer cells [84] . Afterwards, the cell sheet could beeasily removed from thesurface by decreasing thetemper-ature to 25 C since the surface behaved hydrophilic belowthe coil to globule transition temperature of the copoly-mer. Very recently, this approach could even be applied tomouse embryonic stem cells [85] . Attachment of a bacte-ricidal protein mangain-I to the thermo responsive surfaceresulted in a coating that is bactericidal below and cell-repellent above the temperature at which the PmEO n MAcollapses [86,87] .

In addition, PmEO n MA based polymers have beenapplied for the preparation of smart particulate sys-tems: grafting to magnetite nanoparticles resulted inpromising MRI contrast agents [88] , while PLGA [89] andPCL [90] microparticles were coated with PmEO n MA toform thermo-responsive dispersions that may be appliedto support cell growth. Nanoparticles were preparedfrom PLGA block copolymers with PmEO n MA coating forpotential bio-medical applications [91] . Other approachesinvolve the application of mEO n MA copolymers withDMAEMA as nonviral gene transfection agents [92] , thecomplex formation of P(DMAEMA- b-mEO 2 MA) with coq-pea chlorotic mottle virus by electrostatic interaction [93]or the synthesis of protein hybrids [94,95] and polymerconjugates with other biologically active molecules, suchas biotin and thyroxin [96,97] .

With respect to future biological application of PmEO n MAs their LCST behavior wasalso studied in a rangeof buffers that are typically applied for biomedical studies,cellculturemedia [98] aswellasbovineserum [99] . Thefactthat T cp differed by up to 5 C in some of these media com-paredto pure waterstressesthe necessityto tailorthe com-position of the thermo-responsive polymer specically forthe environmental conditions of the targeted application.

5. Poly(2-oxazoline)s

Various 2-oxazoline-based monomers can be polymer-ized via a living cationic ring-opening polymerization(CROP) mechanism ( Fig. 15 ). Due to the livingness of the polymerization, the use of functional electrophilic

initiating as well as nucleophilic end-capping agentsenables thesynthesisof both - and - end-functionalizedpolymers. The nature of the substituent R1 in 2-positionof the oxazoline monomer determines the hydrophilicityof the resulting poly(2-oxazoline) (POx): poly(2-methyl-2-oxazoline) (PMeOx) is water soluble in the entiretemperature range of liquid water under atmosphericpressure, poly(2-ethyl-2-oxazoline) (PEtOx) and poly(2-propyl-2-oxazoline) (PPrOx) exhibit LCST behavior inaqueous solution whereas longer substituents result inwater insoluble hydrophobic polymers.

Several studies on the investigations of the bio-compatibility of POx in comparison to the widely appliedPEO have been carried out revealing similar protein repel-lentproperties of PMeOx functionalized surfaces [100,101]and rapid blood clearance rates of 111 In and 125 I radioac-tively labeled PMeOx and PEtOx when injected to micewithout signicant accumulation in the body [102,103] .PEtOx and PMeOx grafted liposomes showed similarenhanced blood circulation times and tissue distributionas the corresponding PEO conjugates [104,105] . In addi-tion, cytotoxicity studies of PEtOx [106] and a wide rangeof amphiphilic di- and tri-block copolymers (compris-ing PMeOx, PEtOx, PPrOx and poly(2-butyl-2-oxazoline)(PBuOx) [107] revealed a fast endocytosis of especiallymore hydrophilic polymers without affecting the viabilityor activity of the cells. These encouraging ndings alreadyled to initial application studies of POx for drug deliv-ery purposes: PMeOx- b-PBuOx block copolymer micelleshave been successfully applied as carrier systems for thedelivery of the anticancer drug Palitaxel into mice [108] .In addition, PEtOx conjugates with enzymes (i.e. trypsin,RNAse, uricase, catalase), insulin and the anticancer drugAra-C revealed similar enzyme activity and cytotoxicity asPEO conjugates [109,110] . Despite these very promisingstudies demonstrating the biocompatibility of POx, thereare almost no reports regarding biomedical applications of thermoresponsive POx yet.

5.1. Homopolymers

Lin et al. were the rst who reported the LCST behav-ior of three high molar mass PEtOx homopolymers in1988 [111] . The performed cloud point measurementsrevealed T cp s in the range of 6169 C for concentrationsof 0.520wt% PEtOx in aqueous solution. The cloud pointcurves showed minimaat a concentration of about 23wt%and the corresponding temperatures were 61 C, 63 Cand 63.5 C for PEtOx having a molar mass of 20kgmol 1 ,50kgmol 1 and 500 kgmol 1 , respectively. NaCl induceda salting out-effect, whereas addition of tetrabutyl ammo-nium bromide resulted in a salting-in effect. In addition,the T cp s were found to increase by the addition of dioxaneas co-solvent to theaqueous solutions. Theinvestigation of the molar mass dependence of the thermo-responsivenesswas extended to lower molar mass PEtOx by Du Prezand co-workers [112] showing that the LCST behavior of PEtOx ceases to exist in the range of 0100 C for PEtOxwith molar masses lower than M n =10,000gmol 1 (whichcorresponds to a DP of approximately 100, entranceFig. 16 ). Theauthors also proposed the PEtOx/water binary


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Fig. 15. Schematic representation of the CROP of 2-oxazolines.

mixture to follow FloryHuggins Type I behavior due tothe lowered T cp for higher molar mass polymers. Similarresults were found by Hoogenboom and Schubert whoinvestigated PEtOx with DP from 50 to 500 using turbiditymeasurements in 0.5 wt% aqueous solutions (entrancein Fig. 16 ) [113] .

Recently, three investigations describing the thermo-responsive behavior of star shaped PEtOx were published.In accordance to linear PEtOx, an increasing molar mass of star shaped PEtOx with a hyperbranched polyglycerol coreresulted in lowered T cp s from 75 to 62 C (c =5mg mL 1 ),whichisloweredby710 Ccomparedtothe T cp of aqueoussolutionsoflinearPEtOxofsimilarmolarmass [114] . Ontheother hand, aqueous solutions of star shaped PEtOx withpoly(propylene imine) dendrimers as core revealed T cp sof around 90 C (c =5mg mL 1 ) that were not dependenton the molar mass of the polymer and elevated com-pared to linear PEtOx [115] . In addition, star shaped PEtOx,which was prepared by cross-linking of micelles with apoly(2-(3-butynyl)-2-oxazoline) core, showed T cp s in therange of 6064 C [116] in aqueous media at varying pHvalues ( c = 0.1wt%). pH sensitivity was introduced by func-tionalization of the core with amino- and carboxylic acidmoieties, respectively, resulting in elevated T cp when thepH sensitive groups were in their charged state (i.e. at

Fig. 16. Cloud point temperatures observed for aqueous solutions of

PEtOx,P iPrOx and P nPrOx with varying molar mass. Data taken from Ref.[112] ( ), [113] ( , ), [117] ( ), [118] ( ), [119] ( ).

pH= 3 for the amine containing star and at pH= 11 for theacid containing star). The phase transition temperature of the acid containing star could be shifted from 48 to 82 Cby addition of 0.1M Na 2 SO4 or NaSCN as salts from theHofmeister series to the solution.

Next to the simple observation of T cp by turbiditymeasurements, detailed thermodynamic investigations of the demixing process in aqueous solutions have beencarried out already in the 1990s using light scatteringexperiments as well as osmometry [120,121] . The LCSTbehavior of high molar mass PEtOx with broad PDI value(M w =116kgmol 1 , PDI= 2.4) was reected in a decreaseof the second virial coefcient with increasing tempera-ture, and the temperature (at which the solution displaysthe properties of an ideal solution) was determined to be56 C [120] . Since the second virial coefcients describe thenon-ideality of the mixture, excess thermodynamic dilu-tion functions, such as chemical potential 1 E, enthalpy

h1 E and entropy s1 E of mixing could be derived. At30 C, 1 E is negative due to the fact that it is dom-inated by the negative enthalpy term h1 E, whereas at60 C, 1 E is positive due to the fact that it is dominatedby the unfavorable entropy term T s1 E. As a result, theoverall chemical potential 1 = 1 id + 1 E becomespositive at 60 C resulting in demixing. In other words, thephase separation at elevated temperatures is a resultof theincreased entropy of the mixture when water moleculesare released from interactions with the polymer. The factthat hydrogen bonding of water molecules with the car-bonyl oxygen of the PEtOx amide structure contributes tothose polymerwater interactions was shown by a shift of the carbonyl vibration of PEtOx to lower wavenumbers inD2 O when compared to acetonitrile as a non hydrogen-bonding solvent. In addition, the free energy of mixing asa function of the concentration was calculated for severaltemperatures. The cloud point and the coexistence curvewere determined up to a PEtOx concentration of 0.2g mL 1

revealing a critical concentration of 0.109 g mL 1 and acritical temperature of 65.3 C from the minimum of thecoexistence curve, which was found to be inconsistentwith the minimum of the cloud point curve ( ca. 63 Cand 0.025g mL 1 ). This large deviation may result fromthe polydispersity of the utilized PEtOx. DLS as well asSLS were performed at several concentrations and tem-peratures revealing long-range concentration uctuationsthat reect partial organization even below the LCST. Theapproach to determine thermodynamic values from SLSmeasurements below theLCST was further followed by thecharacterization of morewell-dened PEtOxwith differentmolar masses (PDI= 1.3, M w =20, 30, 60kgmol 1 , respec-tively) [121] . Second virial coefcients as well as h1 E and

s1 E were found to decrease with increasing molar mass


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of the polymer, which was suggested to be a result of theincreasing availability of the carbonyl groups in the PEtOxchain to form hydrogen bonds with water with increasingmolar mass.

In 1992, Uyama and Kobayashi reported that P iPrOx(M n =16,700gmol 1 , PDI = 1.13) exhibits LCST behavior inthe range of the human body temperature using turbid-ity measurements of aqueous solutions. The T

cp was found

to decrease from 39 C to 35 C with increasing polymerconcentration (0.11wt%). A salting-out effect of NaCl wasreported whereas the addition of ionic surfactants such assodium dodecylsulfate and dodecyltrimethylammoniumchloride led to an increase of T cp [122] . A detailed inves-tigation of the LCST behavior of four P iPrOx with very lowPDI values of 1.031.05 was performed by Winnik and co-workers applying not only turbidity measurements, butalso high sensitivity DSC (HS-DSC) as well as PPC [117] .T cp as well as T max and T onset of the endothermic peaksfrom DSC of aqueous solutions of P iPrOx ( c =1mg mL 1 )were found to decrease from 73 to 48 C with increasingmolar mass (DP= 1750, entrance in Fig. 16 ). Both tech-niques veried the salting-out effect of NaCl and showedlowered phase transition temperatures in D 2 O comparedto H 2 O due to the difference in hydrogen-bond strengthin both solvents. Increasingconcentration(0.55 mg mL 1 )resulted in decreased T cp s (5649 C) for P iPrOx with a DPof 41, as could be veried by HS-DSC, too. Additionally, theenthalpyof transition H could be obtained from thelattertechnique.Interestingly, H increasedalmost linearlywiththe molar mass of the polymer, which might hint towardsan increasing availability of the polymers carbonyl groupsto form hydrogen bonds with water. This explanation wasconrmed by PPC measurements that revealed a strongincrease of thevolume change V/V during thephasetran-sition with increasing molar mass. V/V was also found tobe increased with increasing NaCl content of the solutionsas well as in D 2 O when compared to H 2 O. Subsequently,Park et al. found that the concentration dependence of T cpbecomes less pronounced with increasing molar mass of quite similar P iPrOx ( M n = 36009700g mol 1 , entrancein Fig. 16 ) [118] . The LCST behavior of four P iPrOx sam-ples with M n from 3000 to 13,000g mol 1 was recentlyinvestigated in detail by Van Mele and co-workers whoapplied modulated DSC in order to establish the phasediagrams [123] . Since both LCST as well as LCSC weredecreasing with increasing molar mass of P iPrOx(33.8 C at29.8wt% for M

n=3000gmol 1 and 26.2 C at 19.8wt% for

M n =13,000gmol 1 ) the binary system P iPrOx/water wasshown to follow TypeI FloryHuggins miscibility behavior.The reversible enthalpy of demixing H dm rev decreasedwith the molar mass of the polymer but remained almostconstant up to concentrations of 20 wt%. At higher con-centrations, H dm rev decreased because too few watermolecules were available to build up the full hydrationshell around the polymer coils. In addition, the (de)mixingkinetics during the phase transition could be followed byquasi-isothermal modulated DSC measurements showingthat the de-mixing is a faster process than the re-mixing.The equilibrium was reached in all cases after 180min,which is much faster than for PNiPAm, since the molec-ular structure of P iPrOx without hydrogen bond donating

Fig. 17. Transmission electron micrographs of coagulate particles.Reproducedfrom Meyer et al. [124] by permission of The Royal Society of Chemistry.

moieties only requires the disruption of polymer/waterhydrogen bonds, and vitrication effects are excluded dueto the fact that the T g of the precipitated polymer phase iswell below the de-mixing temperature.

All the cited references have described the phasetransition to occur abruptly and reversibly with smallheating-cooling hysteresis of the determined values. How-ever, Schlaad discovered that P iPrOx carrying either oneor two positively charged amino-functions at the end of the chain form so-called cotton balls when kept in aque-ous dispersion far above the cloud point (65 C for 24 h)[124] . These coagulates are built from brillar aggregatesthat are formed by crystallization of P iPrOx (see Fig. 17 )[125] . Most likely, non-specic hydrophobic interactiondraws the i-propyl groups together resulting in interactionand alignment of the amide-dipoles of the, thus, stretchedpolymer backbone. Interestingly, the LCST transition isfully reversible during continuous heating-cooling cycleswhile crystallization occurs upon prolonged heating abovethe phase transition temperature. This behavior indicatesthat the initial liquidliquid phase transition is entropydriven while upon isothermal treatment above the T cp , theconcentrated phase undergoes a second irreversible crys-tallization induced phase transition.

After theLCSTbehavior of PEtOxandP iPrOxwasalreadyreported almost 20 years ago, it is astonishing that itwas only recently reported that P nPrOx also exhibits LCSTbehavior by Park and Kataoka [126] . Due to fact that thelinear n-propyl group reduces the area that is accessiblefor water molecules, T cp was found to be almost 15 Clower than for a comparable P iPrOx solution (23.8 C forPnPrOx and 38.7 C for P iPrOx, in 1wt% PBS solution).Subsequently, Hoogenboom and Schubert investigated theeffect of the molar mass of P nPrOx (DP = 10300) uponthe cloud points of aqueous solutions using turbidity mea-surements ( c =0.5wt%) [113] . The T

cps were found to

decreasefrom43to24 C with increasing molar mass of the


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polymer (entrance in Fig. 16 ). In 2011, the propyl sub-stituent of PPrOx was further varied towards a cyclic struc-ture. Theresulting aqueous P c PrOx (DP = 20112) solutions(c = 0.13wt%) exhibited thermo-responsive properties atintermediate temperatures when compared to P nPrOx andPiPrOx (entrance in Fig. 16 ) [119] . In addition, the poly-mer was found to be amorphous and the LCST behaviorof the system P c PrOx/water was investigated by means of viscosimetry revealing increased viscosities shortly belowT cp due to deterioration of the solvent quality and morefavorable interactions between the polymer chains. DLSmeasurements showed the formation of large aggregatesupon coil to globule transition of the polymer.

The effect of various salts on the cloud point of aque-ous solutions of PEtOx, P iPrOx and P nPrOx was recentlyinvestigated by Hoogenboom and Schubert using turbiditymeasurements in 0.5wt% solutions with a systematic vari-ation of the salt concentration (00.5mol L 1 ) [127] . Theeffects of the investigated additives were found to followthe Hofmeister series of anions in an analog fashion as forPEtEO

3MA (compare Section 4.5 ). Also for POx the effects

were more pronounced for the more hydrophilic polymers(PEtOx>P iPrOx>P nPrOx) covering a temperature range of 90 C, whereas the polymer architecture seemed to causeno signicant effects as was shown by comparison of tur-bidity measurements of linear PEtOx and a PEtOx combpolymer.

5.2. Effect of the polymer end group and blockcopolymers

The livingness of the CROP of 2-oxazolines offers thepossibility to functionalize both - as wellas -chain endsof the POx with end groups of varying hydrophilicity andthestraightforward synthesisof block copolymers. Theuseof a functional initiator will attach the desired end groupto the beginning of the polymer chain, whereas varia-tion of the polymer terminal end can either be achievedby the direct attack of nucleophiles on the cationic poly-oxazolinium species or by subsequent modication of terminal OHgroups. Schematic representations of theendgroups that will be discussed in the following part aredepicted in Table 2 .

It is quite obvious that the impact of the end groups onthe LCST behavior of the polymer will be stronger for poly-mers with lower molar mass than for those with highermolar mass. In this context, a 3,3-diethoxy-propyl moietyat the -chain end of P iPrOx ( M n =9600gmol 1 , corre-sponding to DP= 85) did not affect the thermo-responsiveproperties of the P iPrOx when compared to the most fre-quently used CH 3 end group of a polymer with similarmolar mass [118] . In contrast to that, the attachmentof a hydrophobic acrylate function at the terminal endof a P iPrOx with a DP of 40 resulted in a T cp that isdecreased by 5 C when compared to OH terminatedPiPrOx ( c =110gL 1 ) [128] . Besides this, the transmit-tance curves were found to be much broader, and thesalting-outeffect of NaCl( c = 00.15mol L 1 )wasenhancedfor the acrylate oligomer solutions due to the stronghydrating contribution of the OH terminal group. In addi-tion, polyion complex (PIC) micelles that contain two

oppositely charged poly(amino acids) as the inner coreand P iPrOx as outer shells revealed T cp s that remainedremarkably unaffected by changes in concentration or NaClcontent but were signicantly lowered when compared tothe free outer P iPrOx blocks [128] . This could be eitherattributed to a high local concentration of P iPrOx in theshell of the PIC micelles or to the fact that the hydropho-bic methyl groups at the chain ends point towards theoutside of the micelles.

A very detailed investigation on the effect of octade-cyl - and -end groups on the phase transition of PiPrOx (DP = 57, 85, 111) solutions was carried outby Obeid et al. applying turbidity as well as HS-DSCand PPC measurements in dilute concentration regimes(c =0.110mg mL 1 ) [129] . As expected, the hydrophobicend group(s) had the largest inuence on the solubilitybehaviorof theshortestP iPrOx: T cp ofa 1mg mL 1 solutionwas decreased by 12 C and 16 C for P iPrOx of DP 57 withoneand twohydrophobic endgroups, respectively. HS-DSCand PPC revealed the existence of two separate transitionsduring heating of the aqueous solutions that contain thehydrophobically modied P iPrOx in micellar structures.Thetransition at lower temperatures could be correlated tothecloudpoint of thesolutionandwas assigned to changesof the packing of the C 18 chains in the micellar core since itwas hardly affected by the polymer concentration. On theother hand, the second endothermic peak in t

Polímeros biocompatíveis baseados em poli oxido de etileno

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