Separation of peptides with an aqueous mobile phase by temperature-responsive chromatographic column

Peptide separation technology is significant and is still an analytical challenge in proteomic studies. We report a simple preparation method for poly(N-isopropylacrylamide) grafted silica through the copolymerization of N-isopropylacrylamide with acetyl moieties immobilized on the silica surfaces. Differential scanning calorimetry results indicated that the prepared silica exhibited a sharp phase transition at 35.03°C. Silica grafted with poly(N-isopropylacrylamide) was evaluated as a temperature-responsive chromatography medium for the separation of peptides using 0.2 M NaCl solution as a mobile phase. Results indicated that at 10°C, the peptides were not resolved, but baseline separation with prolonged retention time at 50°C was attained. Particularly, a mixture of four peptides was efficiently separated within 8 min. The theoretical plate number of every peptide was more than 2500, and the resolutions were more than 3.40. The increased selectivity of the temperature-responsive column resulted from the temperature-modulated hydrophobic interaction with peptides. The retention times of these peptides were related to their hydrophobicities. This protocol provided a reliable set of chromatographic tool usable across all research and development applications that required isolation and analysis of peptides. It may represent a step forward in the complex analysis of hydrophobic and other proteins.     

Evaluation of peptide-peptide interactions using reversed-phase high-performance liquid chromatography.

The separation of peptides during RP-HPLC depends mainly upon differential hydrophobic interactions of the individual peptides being separated with the C18 group of the stationary phase. We have examined the behavior of dimeric disulfide-linked model peptides during RP-HPLC in order to study self-induced conformational effects. A set of 18 analogues of the amphipathic alpha-helical sequence Ac-LKLLKKLLKKLKKLLKKL-NH2 was used for this study. These analogues differed only by the successive replacement of each position with a cysteine. Strong peptide-peptide interactions, occurring through interchain hydrophobic forces, resulted in a presenting face to the C18 group, consisting primarily of lysine residues and, in turn, in early retention times. Three homo-dimers were also found to be strongly alpha-helical in water as determined by circular dichroism spectroscopy.     

Effect of solute hydrophobicity on phase behaviour in solutions of thermoseparating polymers

 Two-phase systems consisting of a polymer rich phase and polymer depleted phase, where the polymer is either ethyl(hydroxy ethyl)cellulose (EHEC) or Ucon (a random copolymer of ethylene oxide and propylene oxide), have been studied. Both of these polymers can be separated from an aqueous solution by either temperature increase or addition of cosolutes. The polymers are thermoseparating and phase separate in water solutions at the cloud point temperature. Two types of EHEC have been studied: one with a cloud point at 60 °C and the other at 37 °C. The Ucon polymer used in this study has a cloud point at 50 °C. Ternary phase diagrams of polymer/water/cosolute systems have been investigated. When a strongly hydrophilic or hydrophobic cosolute is added to an EHEC- or Ucon–water solution, a phase separation occurs already at, or below, room temperature. As cosolutes, hydrophobic molecules like phenol, butyric and propionic acid, and hydrophilic molecules like glycine, ammonium acetate, sodium carboxylates (acetate to valerate), were studied. The polymer rich phase formed when mixing polymer, water and cosolute was strongly enriched or depleted with hydrophobic or hydrophilic cosolutes, respectively. The two phase region increased for propionic acid, butyric acid and phenol as a result of increased cosolute hydrophobicity. The opposite occurred in the series sodium acetate, sodium butyrate and sodium valerate. The effect of temperature on the phase behaviour has also been investigated. Model calculations based on Flory–Huggins theory of polymer solutions are presented, in form of a phase diagram, which semiquantitatively reproduce some experimental results. Received: 5 July 1996 Accepted: 4 November 1996     

Dramatic thermal stability of virus-polymer conjugates in hydrophobic solvents

We have developed a method for integrating the self-assembling tobacco mosaic virus capsid into hydrophobic solvents and hydrophobic polymers. The capsid was modified at tyrosine residues to display an array of linear poly(ethylene glycol) chains, allowing it to be transferred into chloroform. In a subsequent step, the capsids could be transferred to a variety of hydrophobic solvents, including benzyl alcohol, o-dichlorobenzene, and diglyme. The thermal stability of the material against denaturation increased from 70 °C in water to at least 160 °C in hydrophobic solvents. With a view toward material fabrication, the polymer-coated TMV rods were also incorporated into solid polystyrene and thermally cast at 110 °C. Overall, this process significantly expands the range of processing conditions for TMV-based materials, with the goal of incorporating these templated nanoscale systems into conductive polymer matrices.     

Temperature-induced self-assembly of triple-responsive triblock copolymers in aqueous solutions

A series of triple-thermoresponsive triblock copolymers from poly(N-n-propylacrylamide) (PNPAM, A), poly(methoxydiethylene glycol acrylate) (PMDEGA, B), and poly(N-ethylacrylamide) (PNEAM, C) was synthesized by sequential reversible addition-fragmentation chain transfer polymerizations. Polymers of differing block sequences, ABC, BAC, and ACB, with increasing phase transition temperatures in the order A < B < C were prepared. Their aggregation behavior in dilute aqueous solution was investigated using dynamic light scattering, turbidimetry, and NMR spectroscopy. The self-organization of such polymers was found to dependent strongly on the block sequence. While polymers with a terminal low-LCST (lower critical solution temperature) block undergo aggregation above the first phase transition temperature at 20-25 °C, triblock copolymers with the low-LCST block in the middle show aggregation only above the second phase transition. The collapse of the middle block is not sufficient to induce aggregation but produces instead stable, unimolecular micelles with a collapsed middle block, as supported by NMR and fluorescence probe data. Continued heating of all copolymers led to two additional thermal transitions at 40-55 and 70-80 °C, which could be correlated to the phase transitions of the B and C blocks, respectively. All polymers show a high tendency for cluster formation, once aggregation is induced. The carrier abilities of the triple responsive triblock copolymers for hydrophobic agents were probed with the solvatochromic fluorescence dye Nile Red. With passing through the first thermal transition, the block copolymers are capable of solubilizing Nile Red. In the case of block copolymers with sequences ABC or ACB, which bear the low-LCST block at one terminus, notable amounts of dye are solubilized already at this stage. In contrast, the hydrophobic probe is much less efficiently incorporated by the BAC triblock copolymer, which forms unimolecular micelles. Only after the collapse of the B block, when reaching the second phase transition at about 45 °C, does aggregation occur and solubilization becomes efficient. In the case of ABC and ACB polymers, the hydrophobic probe seems to partition between the originally collapsed A chains and the additional hydrophobic chains formed after the collapse of the less hydrophobic B block.     

Comparison of silica-based cyanopropyl and octyl reversed-phase packings for the separation of peptides and proteins.

The performance of a silica-based C8 packing was compared with that of a less hydrophobic, silica-based cyanopropyl (CN) packing during their application to reversed-phase high-performance liquid chromatography (linear trifluoroacetic acid-water to trifluoroacetic acid-acetonitrile gradients) of peptides and proteins. It was found that: (1) the CN column showed excellent selectivity for peptides which varied widely in hydrophobicity and peptide chain length; (2) peptides which could not be resolved easily on the C8 column were widely separated on the CN column; (3) certain mixtures of peptides and small organic molecules which could not be resolved on the C8 column were completely separated on the CN column; (4) impurities arising from solid-phase peptide synthesis were resolved by a wide margin on the CN column, unlike on the C8 column, where these compounds were eluted very close to the peptide product of interest: and (5) specific protein mixtures exhibited superior resolution and peak shape on the CN column compared with the C8 column. The results clearly demonstrate the effectiveness of employing stationary phases of different selectivities (as opposed to the more common optimization protocol of manipulating the mobile phase) for specific peptide and protein applications, an approach underestimated in the past.     

Polyethylene as a nonvolatile solid cosolvent phase for catalyst separation and recovery

The studies described here show that a relatively low molecular weight, narrow polydispersity polyethylene (PE) wax (Polywax) can serve as a nontoxic and nonvolatile alternative to alkane solvents in monophasic catalytic organic reactions where catalysts and products are separated under biphasic conditions. In this application, a polymer that is a solid at room temperature substitutes for a conventional alkane solvent at ca. 80 °C. In addition to the advantages of being a nonvolatile, nontoxic, reusable solvent, this hydrocarbon polymer solvent, like heptane, can sequester nonpolar soluble polymer-bound catalysts after a reaction and separate them from products. The extent of this separation and its generality were studied using polyisobutylene (PIB)- and poly(4-dodecylstyrene)-bound dyes and PE-bound Pd allylic substitution catalysts, PIB-bound Pd cross-coupling catalysts, and PE- and PIB-bound metathesis catalysts. Catalytic reactions were effected using single-phase reaction mixtures containing Polywax with toluene, THF, or THF/DMF at ca. 80 °C. These solutions either separate into two liquid phases on addition of a perturbing agent or separate as a solid/liquid mixture on cooling. The hydrocarbon polymer-bound dyes or catalysts either separate into the hot liquid Polywax phase or coprecipitate with Polywax and are subsequently isolated as a nonvolatile Polywax solid phase that contains the dye or the recyclable catalyst.     

Spontaneous phase transfer of thermosensitive hairy particles between water and an ionic liquid

This article describes the temperature-induced phase transfer behavior of a series of thermosensitive polymer brush-grafted particles between water and a hydrophobic ionic liquid, 1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([EMIM][TFSI]). Six samples were made by surface-initiated atom transfer radical polymerization: silica particles grafted with poly(methoxypoly(ethylene glycol) methacrylate) (PPEGMMA) with two different molecular weights, poly(methoxytri(ethylene glycol) methacrylate) (PTEGMMA), poly(methoxydi(ethylene glycol) methacrylate) (PDEGMMA), and two copolymers of PEGMMA and TEGMMA with different compositions (P(PEGMMA-co-TEGMMA)-82 and P(PEGMMA-co-TEGMMA)-74). The cloud points of free PPEGMMA with M(n,SEC) of 23 and 40 kDa, P(PEGMMA-co-TEGMMA)-82, P(PEGMMA-co-TEGMMA)-74, and PTEGMMA in [EMIM][TFSI]-saturated water were 95, 94, 80, 72, and 43 °C, respectively. PDEGMMA was not soluble in the ionic liquid-saturated water. PPEGMMA brush-grafted particles moved spontaneously and completely from water to the [EMIM][TFSI] phase upon heating at 80 °C. When cooled to 22 °C, all particles returned to the water layer. From UV-vis absorbance measurements, the transfer temperature (T(tr)) of PPEGMMA-grafted particles from water to the ionic liquid was 42 °C. Thermodynamic analysis showed that the particle transfer was an entropically driven process. P(PEGMMA-co-TEGMMA)-82, P(PEGMMA-co-TEGMMA)-74, and PTEGMMA brush-grafted particles also underwent reversible and quantitative transfer between the two phases upon heating at 70 °C and cooling at 0 °C; their transfer temperatures from water to [EMIM][TFSI] were 36, 30, and 16 °C, respectively. T(tr) was a linear function of the cloud point of the corresponding free polymer in ionic liquid-saturated water. In contrast, PDEGMMA-grafted particles moved spontaneously to the ionic liquid layer upon heating but did not return to water even after prolonged stirring at 0 °C.     

Aqueous two-phase systems containing self-associating block copolymers. Partitioning of hydrophilic and hydrophobic biomolecules.

A series of proteins and one membrane-bound peptide have been partitioned in aqueous two-phase systems consisting of micelle-forming block copolymers from the family of Pluronic block copolymers as one polymer component and dextran T500 as the other component. The Pluronic molecule is a triblock copolymer of the type PEO-PPO-PEO, where PEO and PPO are poly(ethylene oxide) and poly(propylene oxide), respectively. Two different Pluronic copolymers were used, P105 and F68, and the phase diagrams were determined at 30 degrees C for these polymer systems. Since the temperature is an important parameter in Pluronic systems (the block copolymers form micellar-like aggregates at higher temperatures) the partitioning experiments were performed at 5 and 30 degrees C, to explore the effect of temperature-triggered micellization on the partitioning behaviour. The temperatures correspond to the unimeric (single Pluronic chain) and the micellar states of the P105 polymer at the concentrations used. The degree of micellization in the F68 system was lower than that in the P105 system, as revealed by the phase behaviour. A membrane-bound peptide, gramicidin D, and five different proteins were partitioned in the above systems. The proteins were lysozyme, bovine serum albumin, cytochrome c, bacteriorhodopsin and the engineered B domain of staphylococcal protein A, named Z. The Z domain was modified with tryptophan-rich peptide chains in the C-terminal end. It was found that effects of salt dominated over the temperature effect for the water-soluble proteins lysozyme, bovine serum albumin and cytochrome c. A strong temperature effect was observed in the partitioning of the integral membrane protein bacteriorhodopsin, where partitioning towards the more hydrophobic Pluronic phase was higher at 30 degrees C than at 5 degrees C. The membrane-bound peptide gramicidin D partitioned exclusively to the Pluronic phase at both temperatures. The following trends were observed in the partitioning of the Z protein. (i) At the higher temperature, insertion of tryptophan-rich peptides increased the partitioning to the Pluronic phase. (ii) At the lower temperature, lower values of K were observed for ZT2 than for ZT1.     

Properties of gastroretentive sustained release tablets prepared by combination of melt/sublimation actions of L-menthol and penetration of molten polymers into tablets

A novel floating sustained release tablet having a cavity in the center was developed by utilizing the physicochemical properties of L-menthol and the penetration of molten hydrophobic polymer into tablets. A dry-coated tablet containing famotidine as a model drug in outer layer was prepared with a L-menthol core by direct compression. The tablet was placed in an oven at 80°C to remove the L-menthol core from tablet. The resulting tablet was then immersed in the molten hydrophobic polymers at 90°C. The buoyancy and drug release properties of tablets were investigated using United States Pharmacopeia (USP) 32 Apparatus 2 (paddle 100 rpm) and 900 ml of 0.01 N HCl. The L-menthol core in tablets disappeared completely through pathways in the outer layer with no drug outflows when placed in an oven for 90 min, resulting in a formation of a hollow tablet. The hollow tablets floated on the dissolution media for a short time and the drug release was rapid due to the disintegration of tablet. When the hollow tablets were immersed in molten hydrophobic polymers for 1 min, the rapid drug release was drastically retarded due to a formation of wax matrices within the shell of tablets and the tablets floated on the media for at least 6 h. When Lubri wax? was used as a polymer, the tablets showed the slowest sustained release. On the other hand, faster sustained release properties were obtained by using glyceryl monostearate (GMS) due to its low hydrophobic nature. The results obtained in this study suggested that the drug release rate from floating tablets could be controlled by both the choice of hydrophobic polymer and the combined use of hydrophobic polymers.     

Displacement chromatography of biomolecules

Displacement chromatography was used for the preparative-scale separation of peptides, antibiotics, and proteins. The feed components were both purified and concentrated during the separation processes. The components of a peptide mixture were separated on a reverse-phase analytical column using 2-(2-butoxyethoxy) ethanol as the displacer. The use of organic modifiers in the carrier along with an elevated column temperature of 45 degrees C enabled the efficient separation of relatively hydrophobic peptides by displacement chromatography. In addition, the throughput of the process was significantly increased by carrying out the separation at an elevated flow-rate with no adverse effect on product purity. The antibiotic cephalosporin C was isolated from impurities in a fermentation broth using 2-(2-butoxyethoxy)ethanol as the displacer along with a step change in column temperature. The proteins cytochrome c and lysozyme were purified on a weak cation-exchanger column using cationic polymers as the displacers. While polymers of 60 and 20 kilodaltons were both found to be good displacers for these proteins, only the lower molecular weight polymer was readily removed from the column by standard regeneration techniques.     

Nanopore formation by self-assembly of the model genetically engineered elastin-like polymer [(VPGVG)2(VPGEG)(VPGVG)2]15

The self-assembly characteristics of the model genetically engineered elastin-like polymer [(VPGVG)2(VPGEG)(VPGVG)2]15 have been studied in this work. An AFM study of the topology of polymer films deposited from acid and basic solutions on a hydrophobic silicon substrate has been carried out. Under acidic conditions, polymer deposition results in a flat surface with no particular topological features. However, from basic solutions, polymer deposition clearly shows an aperiodic pattern of nanopores ( approximately 70 nm width and separated about 150 nm). This dramatic dependence of film topology on pH is explained in terms of the different polarity of the free gamma-carboxyl group of the glutamic acid. In the carboxylate form, this moiety shows a markedly higher polarity than the rest of the polymer domains and the substrate itself. Under these conditions, the charged carboxylates impede hydrophobic contact with their surroundings, which is the predominant assembly pathway for this type of polymer. The charged domains, along with their hydration sphere, are then segregated from the hydrophobic surroundings giving rise to nanopores.     

Temperature-responsive solid-liquid separations with charged block-copolymers of poly(N-isopropyl acryamide)

Temperature responsive charged block-copolymers of poly(N-isopropylacrylamide) (PNIPAM) have been used in the solid-liquid separation of alumina mineral particles from aqueous solution. The effects of temperature, polymer charge-sign and fraction of charged segment have been investigated. Batch settling and adsorption studies showed that rapid sedimentation results for suspensions with polymers of opposite charge-sign to the particle surface-charge (counterionic) at 50 °C. Cooling the suspensions after flocculation at 50 °C was found to increase the final solids volume fraction of the sediment beds formed through a mechanism related to partial desorption of polymer and the reduction of the hydrophobic attraction. Suspension stability results after dosing with polymers of similar charge-sign to the particle surface-charge (co-ionic) at both 25 and 50 °C. Increasing the amount of polymer charge increased the influence of polymer charge-sign on the adsorption and solid-liquid separation behavior. The performance of the charged block copolymers are compared to that of the random charged copolymer and neutral homopolymer PNIPAM structures.     

Stereoregulation of thermoresponsive polymer brushes by surface-initiated living radical polymerization and the effect of tacticity on surface wettability

In this study, stereocontrolled poly(N-isopropylacrylamide) (PIPAAm) brushes were grafted from surfaces by atom transfer radical polymerization (ATRP) in the presence of a Lewis acid, and the effect of PIPAAm brush tacticity on the thermoresponsive wettabiliy was investigated. PIPAAm grafted by ATRP in the presence of Y(OTf)(3) showed high isotacticity, while the control brush polymerized in the absence of Y(OTf)(3) was clearly atactic. The isotacticity and molecular weight of PIPAAm brushes were controlled by polymerization conditions. The wettability of isotactic PIPAAm-grafted surfaces decreased slightly below 10 °C, although the phase transition temperature of atactic surface was 30 °C, and the bulk isotactic polymer was water-insoluble between 5 and 45 °C.     

Time-dependent shrinkage of polymeric micelles of amphiphilic block copolymers containing semirigid oligocholate hydrophobes

Alkyne-derivatized poly(ethylene glycol) (M.W. 5000) was coupled to several azide-terminated oligocholates by the click reaction to form amphiphilic block copolymers. A copolymer with a cholate hexamer as the hydrophobic block formed polymeric micelles that shrank by ~50% over a period of 10 h at 25°C. Shrinkage was faster and more dramatic at 35°C. Shortening the oligocholate by two units or inserting a 4-aminobutyroyl spacer in the hexacholate eliminated or diminished the shrinkage. Metastable aggregates were proposed to form when the block copolymers began to aggregate in water. The large hydrophobic surface, awkward shape, rigidity, and facial amphiphilicity of the cholate repeat unit and the long chain made it difficult for the oligocholates to adjust within the micellar core. As the oligocholates rearranged to maximize hydrophobic interactions and hydrogen-bonding while minimizing conformational strain, the polymeric micelles became more compact over time.     

水杨酸分子印迹分离介质的水相制备及色谱行为

以水杨酸为模板分子,2-乙烯基吡啶为功能单体,在甲醇/水体系中制备了水杨酸分子印迹聚合物,并将其作为高效液色谱分离介质研究了其对水杨酸及水杨酸的位置异物体———对羟基苯甲酸的色谱行为。结果表明,该聚合物固定相表现出良好的分离选择性,能够使水杨酸与对羟基苯甲酸快速基线分离。通过对流动相组成与色谱分离效果关系的探讨,证实溶质与印迹固定相的疏水相互作用是分子识别的主要作用力。     

3D interconnected ionic nano-channels formed in polymer films: self-organization and polymerization of thermotropic bicontinuous cubic liquid crystals

Thermotropic bicontinuous cubic (Cub(bi)) liquid-crystalline (LC) compounds based on a polymerizable ammonium moiety complexed with a lithium salt have been designed to obtain lithium ion-conductive all solid polymeric films having 3D interconnected ionic channels. The monomer shows a Cub(bi) phase from -5 to 19 °C on heating. The complexes retain the ability to form the Cub(bi) LC phase. They also form hexagonal columnar (Col(h)) LC phases at temperatures higher than those of the Cub(bi) phases. The complex of the monomer and LiBF(4) at the molar ratio of 4:1 exhibits the Cub(bi) and Col(h) phases between -6 to 19 °C and 19 to 56 °C, respectively, on heating. The Cub(bi) LC structure formed by the complex has been successfully preserved by in situ photopolymerization through UV irradiation in the presence of a photoinitiator. The resultant nanostructured film is optically transparent and free-standing. The X-ray analysis of the film confirms the preservation of the self-assembled nanostructure. The polymer film with the Cub(bi) LC nanostructure exhibits higher ionic conductivities than the polymer films obtained by photopolymerization of the complex in the Col(h) and isotropic phases. It is found that the 3D interconnected ionic channels derived from the Cub(bi) phase function as efficient ion-conductive pathways.     

Microstructure and lateral diffusion in monolayers of polymerizable amphiphiles

Lipid analogue amphiphilic molecules containing polymerizable units were investigated in monolayers at the air/water interface by using film balance measurements, fluorescence microscopy, and photobleaching techniques. The polymerizable groups (diene-, diyne-, and methacrylate units) were introduced into the hydrophobic alkyl chains or into the polar head of the amphiphilic molecules.In the case of the diene- and diyne-containing compounds the polymerizable units are incorporated into the hydrophobic alkyl chains, enabling them to form a two-dimensional network. Due to the free chain flexibility of the monomers the lateral mobility was comparable to that of saturated lipid analogues and decreases upon polymerization proportionally to the dose of UV irradiation. In addition, fluid/solid phase transitions of compounds with polymerizable groups in the hydrophobic part tend to vanish during the formation of the polymers. However, the direct observation of the growth of polymeric crystalline domains can be followed by using diacetylene lipid analogues.In the case of the methacrylate derivatives the polymerizable unit was coupled to the polar part via a flexible spacer. For these systems the characteristics of the monomeric phase transition are retained after polymerization. However, it shows a significant, strong decrease of the in-plane mobility already in the fluid-expanded phase of the polymer. The quantitative measurements of the lateral diffusion in the monolayers can be correlated with fluorescence microscopic images of their structure.     

Peptide‐binding specificity of the prosurfactant protein C Brichos domain analyzed by electrospray ionization mass spectrometry

The C‐terminal domain of lung surfactant protein C (CTC) precursor (proSP‐C) is involved in folding of the transmembrane segment of proSP‐C. CTC includes a Brichos domain with homologs in cancer‐ and dementia‐associated proteins. Mutations in the Brichos domain cause misfolding of proSP‐C and hence amyloid fibril formation in interstitial lung disease. Electrospray ionization mass spectrometry (ESI‐MS) with collision‐induced dissociation (CID) experiments was applied to study non‐covalent interactions between human recombinant CTC or its Brichos domain, and SP‐C analogs, homotripeptides and peptides designed to model amyloid fibril formation. The results show that the Brichos domain contains the peptide‐binding function of CTC. In titration experiments, apparent dissociation constants (K_D) were in the micromolar range where triple‐valine showed the lowest K_D and triple‐tyrosine the highest. Non‐hydrophobic peptides failed to form complexes with Brichos. CID revealed that complexes with aromatic peptide ligands are more stable in the gas phase than complexes with non‐aromatic ligands. The Brichos domain was also shown to bind fibril‐forming peptides containing aromatic/hydrophobic residues. Copyright © 2009 John Wiley & Sons, Ltd.     

Continuous synthesis of CdSe(x)Te(1-x) nanocrystals: chemical composition gradient and single-step capping

Thermosensitive poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO) triblock copolymer, Pluronic F68, containing a hydrophobic unit, oligo-(lactic acid)(oligo-LA) or oligo-caprolactone (oligo-CL), 2-META and RGD as side groups was successfully synthesized and characterized by (1)H NMR, FTIR, and elemental analysis. Their aqueous solution displayed special gel-sol-gel phase transition behavior with increasing temperature from 10 to 70°C, when the polymer concentration was above critical micelle concentration (CMC). The gel-sol phase diagram was investigated using tube inversion method, rheological measurement, and dynamic light scattering. Based on these results, the gelation properties of modified F68 were affected by several factors such as the composition of the substituents, chain length of oligo L-LA or oligo ε-CL, and the concentration of the polymer solutions. The unique phase transition behavior with temperature was observed by modified F68 triblock copolymer, composed of the PPO blocks core and the PEO blocks shell in aqueous solution. This phenomenon was elucidated using (1)H NMR data; the alteration of hydrophobic interaction and chain mobility led to the formation of transparent gel, coexistence of gel-sol, and opaque gel. These hydrogels may be useful in drug delivery and tissue engineering.