Synthesizing a Trefoil Knotted Block Copolymer via Ring-Expansion

Feb 7, 2017 - A synthetic trefoil knotted poly(ε-caprolatone)-block-poly(l-lactide) (TK-PLA-b-PCL) is synthesized via a ring-expansion strategy from ...
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Synthesizing a Trefoil Knotted Block Copolymer via Ring-Expansion Strategy Peng-Fei Cao,†,§ Li-Han Rong,† Joey Dacula Mangadlao,†,‡ and Rigoberto C. Advincula*,† †

Department of Macromolecular Science and Engineering and ‡Department of Radiology, Case Western Reserve University, Cleveland, Ohio 44106, United States § Chemical Sciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37830, United States S Supporting Information *

ABSTRACT: A synthetic trefoil knotted poly(ε-caprolatone)block-poly(L-lactide) (TK-PLA-b-PCL) is synthesized via a ringexpansion strategy from a trefoil knotted tin (Sn) initiator. Ringclosing reaction between the bis-copper(I) templated phenanthroline complex and dibutyldimethoxytin results in a templated trefoil knotted initiator. The bis-copper(I) templated trefoil knotted poly(L-lactide) (TK-PLA) can be synthesized by ring-opening polymerization of L-lactide monomer, and decomplexation reaction of the templated TK-PLA will result in a geniune TK-PLA without constraint from the copper template. Subsequent insertion of εcaprolactone in the bis-copper(I) templated TK-PLA forms the templated trefoil knotted block copolymer, i.e., TK-PLA-b-PCL, and the copper-free TK-PLA-b-PCL can be obtained by decomplexation reaction. Both TK-PLA and TK-PLA-b-PCL are analyzed by the 1H NMR, FT-IR, UV−vis, DLS, and GPC.

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introduced to the synthetic polymers by several groups.26−31 However, rational access to the trefoil knotted polymer remains a formidable challenge although observation of the trefoil knotted polymer was claimed by several groups, which were typically formed by extremely slim chances of self-entanglement during the end-to-end coupling process and were usually done in poor solvents.32−34 Theoretical studies were performed on the “knot” effect on the physical properties of polymers, and initial studies also demonstrated the unique thermal, mechanical, and dynamic properties of the knotted polymers, which may potentially open wider applications of this polymeric material.35,36 Recent studies on the effect of local flexibility on the position and size of knots along the polymer chain may support the understanding of topological entanglement in natural polymers, such as DNA and proteins.37 To achieve the experimental study of the “knot” effect on the polymer materials, we have first reported the rational synthesis of trefoil knotted poly(εcaprolactone) (PCL) by ring-expansion strategy from a biscopper(I) templated phenanthroline tin (Sn) initiator.38 Comparative property study of the trefoil knotted polymer with their linear analogue confirmed their expected properties, such as reduced hydrodynamic radius and lower intrinsic viscosity. Moreover, the transformation of the C2 symmetry of

INTRODUCTION Knots are interesting structures not only in arts but also in mathematics. They are commonly encountered in modern society for different purposes such as knitting, fishing, and surgery.1,2 Beyond aesthetic reasons, i.e., attractive and fascinating architectures, natural biomacromolecules including DNA, RNA, and proteins also exude their fair share of knotty motif.3−7 The knotted structure in proteins appears to bury the active regions against degradation, therefore increasing stability and enhancing functionality.8 Over the past two decades, several efforts have been exhausted to the synthesis of molecular knots.2,9,10 Utilization of the metal template,11−15 hydrogen bonding,16 and hydrophobic interaction17 were demonstrated useful in generating the crossing points of these molecules with entwined structure. New bond formation methods, such as azide−alkyne cycloaddition18 and ring-closing olefin metathesis,12 were utilized to the covalent linkage of preorganized ligands. The topology effect on the physical properties of macromolecules is significant, which was experimentally demonstrated by the unique properties of cyclic polymers compared with their linear analogues.19−22 Initial studies of the cyclic polymers also demonstrated their unique biophysical properties, such as retarded degradation profile, unique biodistribution, and enhanced transfection efficiency, which may provide great advantages in drug/gene delivery and bioimaging applications.23−25 Aside from the “endless” structure, mechanically interlocked architecture, i.e., catenation, was also © XXXX American Chemical Society

Received: September 17, 2016 Revised: December 12, 2016

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DOI: 10.1021/acs.macromol.6b02029 Macromolecules XXXX, XXX, XXX−XXX

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Macromolecules Scheme 1. Synthesis of Bis-Copper(I) Templated Trefoil Knotted Initiator 5

bis-copper(I) templated trefoil knotted polymer to the C3 symmetry of the template-free trefoil knotted polymer (not absolute C3 symmetry due to the presence of phenanthroline group) was also first observed by atomic force microscopy (AFM) imaging. As a more advanced topologically interesting macromolecules compared to cyclic and catenated polymers, the synthetic trefoil knotted polymers are expected to be a unique polymeric material due to their self-entanglement and intrinsic chiral structure.9,35,36 Synthetic polyesters with knotted architectures are particularly interesting due to their biodegradability and biocompatibility and hence great potential in biomaterial fabrication and other environmentally friendly material development.39−42 Compared to poly(ε-caprolactone), poly(lactide) exhibits different physical properties, such as faster degradation rate, better mechanical strength, and lower permeability to most drugs.43−45 Forming the block copolymer of PLA and PCL will enable preparation of the polymer material with tunable properties for various applications, especially in biomedical fields.43,46−48 To enrich the variety of the synthetic trefoil knotted polyesters and contribute to both fundamental study of “knot” effect on polymer properties and their corresponding potential application, we report the synthesis of trefoil knotted poly(L-lactide) by ring-opening polymerization of L-lactide monomer from a trefoil knotted initiator, and subsequent insertion of ε-caprolactone affords a synthetic trefoil knotted block copolymer poly(L-lactide)-bpoly(ε-caprolactone). To the best of our knowledge, this is the first report on the synthetic trefoil knotted poly(lactide) and one of the first attempts to synthesize trefoil knotted block copolymers. The unique physical properties derived from the topologically interesting architectures will make these trefoil

knotted polyesters a unique polymeric material for different applications.

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EXPERIMENTAL SECTION

Materials. Chemical reagents were purchased from Sigma-Aldrich and used without further purification unless otherwise indicated. εCaprolactone (CL, 99%, Alfa) was distilled from CaH2 under reduced pressure. L-Lactide was purchased from Alfa Aesar and was purified by recrystallization from toluene before use. Tri(ethylene glycol) was purchased from Alfa Aesar. 1,10-Phenanthroline (99%) was purchased from Acros Organics. Benzolylated cellulose tubing (MWCO 2000) was purchased from Sigma and was used directly. Herein, DPn was defined as the sum of the two polymer chains from one tin atom, so it will be consistent with our previous report.29,38 Characterization. 1H nuclear magnetic resonance (NMR) spectra were recorded on a Varian Inova 600 MHz NMR spectrometer using chloroform-d. Gel permeation chromatography (GPC) measurements were carried out on a Viscotek 270 instrument with a triple detector array (RALS, IV, RI, or UV) equipped with two GMHHR-M and one GMHHR-L mixed bed ViscoGel columns (eluent: THF; flow rate: 1 mL min−1). Mn was obtained by Universal Calibration with linear polystyrene as the standard. UV−vis spectra were recorded on a UV− vis−NIR spectrometer (StellarNet. Inc.), and the scanning range was 200−1000 nm. Infrared (IR) spectra were recorded on a Cary 600 Series FT-IR spectrometer (Agilent Technologies), and the scanning range was 4000−400 cm−1. Dynamic light scattering (DLS) and static light scattering (SLS) measurements were performed using a DynaPro NanoStar (Wyatt). A Bruker AUTOFLEX III MALDI TOF/TOF mass spectrometer was operated using HABA [2-(4-hydroxyphenylazo)benzoic acid] as matrix and sodium trifluoroacetate, Na(CF3COO), as a doping salt. X-ray photoelectron (XPS) spectroscopy measurement was performed on a PHI 5700 X-ray photoelectron spectrometer. A monochromatic Al Kα X-ray source was employed with 90° relative to the axis of hemispherical energy analyzer. Synthesis. Synthesis of EG-Terminated Phenanthroline Ligand (3). Synthesis of compound 1 and monotosylated tri(ethylene glycol) B

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(2.8 mmol) of ε-caprolactone were combined in a 25 mL Schlenk flask. After three freeze−pump−thaw cycles, the polymerization was conducted in an oil bath at 80 °C under a nitrogen atmosphere for 18 h. Further purification was performed by dialysis with DCM as solvent. 1 H NMR spectra of Cu(I)-Knot-PLA91 and Cu(I)-Knot-PLA91-bPCL31 are shown in Figures 4 and 7. Decomplexation of Trefoil Knotted Polymer (8 and 9). A typical synthesis of Knot-PLA91 follows. 100 mg of Cu(I)-Knot-PLA91 was dissolved in 10 mL of acetonitrile with magnetic stirring, and 20 mg of KCN was added to the solution. The mixture was stirred overnight, and successful decomplexation was evidenced by the color change (from reddish to colorless). Solvent was removed under reduced pressure, and DCM was added to dissolve the resulting polymer. Extra KCN was filtered, and the polymer was purified by dialysis against DCM. 1H NMR spectra of Knot-PLA91 and Knot-PLA91-b-PCL31 are shown in Figures 4 and 7.

(TEG) (2) was performed according to our previous publications (see Supporting Information for details).38 0.309 g (0.5 mmol) of compound 1 was dissolved in 30 mL of dimethylformide (DMF). After 30 min stirring, 0.609 g (2.0 mmol) of compound 2 and 0.690 g (5.0 mmol) of K2CO3 were subsequently added. The mixture was stirred overnight at 90 °C under the protection of a nitrogen (N2) atomsphere. The solvent was removed by a vacuum. 30 mL of dichloromethane (DCM) was added to dissolve the solid, and the precipitate was filtered out. The obtained solution was washed by DI water (15 mL × 3), and the solvent was removed to afford a yellow solid. Further purification was achieved by column chromatography with methanol/chloroform (8:100) as eluent. Pure solid product can be obtained by recrystallization in methanol. 1H NMR (600 MHz, CDCl3), δ ppm: 9.68 (s, 1H, Ha), 8.70 (d, J = 7.5 Hz, 2H, Hb,), 8.50− 8.40 (m, 8H, He + Hd + Hj), 8.30 (d, 2H, Hh, J = 8.6 Hz), 8.10 (d, 2H, Hi, J = 8.0 Hz), 7.86−7.79 (m, 5H, Hc + Hf + Hg), 7.07 (d, 4H, Hk, J = 9.2 Hz), 4.24 (t, 4H, Hl, J = 4.9 Hz), 3.94 (t, 4H, Hm, J = 4.9 Hz), 3.80−3.70 (m, 12H, Hn + Ho + Hq), 3.64 (4H, t, Hp, J = 4.6 Hz). 13C NMR (600 MHz, CDCl3), δ ppm: 61.8, 67.5, 69.8, 70.5, 70.9, 72.5, 114.8, 119.4, 120.0, 125.7, 126.1, 127.0, 127.5, 128.1, 128.7, 129.1, 129.5, 132.4, 136.8, 136.9, 137.7, 146.0, 146.1, 156.4, 156.4, 160.0. MALDI-TOF found 883.24 [M + H+], calculated: 882.44 for C54H50O8N4. Synthesis of Bis-Copper(I) Templated Knotted Complex (4).38 Compound 3 (0.088 g, 0.10 mmol) in 25 mL of DCM and Cu(CH3CN)4PF6 (0.045 g, 0.12 mmol) in 25 mL of acetonitrile (CH3CN) were prepared separately, and both of the solutions were bubbled with nitrogen for 20 min. The CH3CN solution of Cu(CH3CN)4PF6 was slowly transferred to the DCM solution of compound 3 to afford a reddish solution. After another 20 min stirring under a N2 atmosphere, the solvent was removed to obtain the complex as reddish solid. 1H NMR (CDCl3, 600 MHz), δ ppm: 9.65 (s, 2H, Ha), 8.33 (d, 4H, Hh, J = 8.6 Hz), 8.05−7.96 (m, 12H, He+f+g), 7.59 (d, 4H, Hi, J = 8.6 Hz), 7.06 (t, 2H, Hc, J = 8.0 Hz), 7.00 (d, 8H, Hj, J = 8.6 Hz), 6.87 (d, 4H, Hb, J = 8.0 Hz), 6.50 (d, 4H, Hd, J = 8.0 Hz), 5.71 (d, 8H, Hk, J = 8.6 Hz), 3.87−3.67 (m, 48H). 13C NMR (DMSO, 600 MHz), δ ppm: 61.01, 67.44, 69.24, 70.44, 70.60, 73.10, 104.99, 112.81, 121.45, 124.65, 126.58, 126.77, 127.88, 128.03, 128.26, 129.87, 131.24, 136.73, 137.01, 138.06, 142.33, 142.52, 153.16, 156.52, 159.20. MALDI-TOF found 945.12 [M/2] + and 1828.83 [M−Cu]+, calculated: 1891.98 for C108H100O16N8Cu2. Synthesis of Bis-Copper(I) Templated Trefoil Knotted Initiator (5).38 Knot complex (4) (0.095 g, 0.050 mmol) was dissolved in 400 mL of dried chloroform (CHCl3). The solution in a round-bottom flask was bubbled with N2 for 30 min, and then it was heated to reflux. 0.031 g of dibutyldimethoxytin (0.11 mmol) in 5 mL of CHCl3 was then added to the chloroform solution of the knot complex. The mixture was refluxed overnight under the protection of a N2 atmosphere. The solvent was removed by vacuum to obtain the product as a reddish solid with 96% yield. 1H NMR (CDCl3, 600 MHz), δ ppm: 9.65 (s, 2H, Ha), 8.33 (d, 4H, Hh, J = 8.6 Hz), 8.05− 7.96 (m, 12H, He+f+g), 7.59 (d, 4H, Hi, J = 8.6 Hz), 7.06 (t, 2H, Hc, J = 8.0 Hz), 7.00 (d, 8H, Hj, J = 8.6 Hz), 6.87 (d, 4H, Hb, J = 8.0 Hz), 6.50 (d, 4H, Hd, J = 8.0 Hz), 5.71 (d, 8H, Hk, J = 8.6 Hz) 3.87−3.60 (m, 48H), 1.85−1.33 (m, 24H), 0.96−0.87 (m, 12H). 13C NMR (DMSO, 600 MHz), δ ppm: 13.7, 26.7, 27.4, 29.8, 61.00, 67.43, 69.23, 70.56, 70.58, 73.07, 112.80, 121.44, 124.64, 126.58, 126.76, 127.87, 128.02, 128.24, 129.85, 131.24, 136.73, 136.99, 138.06, 142.32, 142.51, 153.15, 156.51, 159.19. Molecular peak was not found in the MALDI-TOF spectra due to the sensitivity of the Sn−O bonds.49 Synthesis of Cu(I)-Knot-PLAm and Cu(I)-Knot-PLAm-b-PCLn (6 and 7). A typical synthesis of Cu(I)-Knot-PLA91 was performed as follows. Bis-copper(I) templated trefoil knot initiator (5) (45 mg, 0.020 mmol) and L-lactide monomers (577 mg, 4.0 mmol) were combined in a 25 mL Schlenk flask. The mixture was degassed by three freeze−pump− thaw cycles, backfilled with N2, and then subjected to polymerization in an oil bath at 120 °C for 18 h. The polymer was purified by dialysis against DCM before analysis. A synthetic block copolymer Cu(I)Knot-PLA91-b-PCL31 was performed similarly from a purified Cu(I)Knot-PLA91. 200 mg (0.007 mmol) of Cu(I)-Knot-PLA91 and 320 mg

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RESULTS AND DISCUSSION Bis-Copper(I) Templated Trefoil Kotted Initiator. The synthesis of the bis-copper(I) templated trefoil knotted initiator was previously reported by our group.38 Herein, a more detailed information is provided. Various methods were reported on the formation of molecular knots, and utilization of the metal template was proven very successful to preorganize the desired architecture.10 Different metal ions such as Cu(I), Fe(II), Zn(II), and Ln(III) can be employed to the synthesis of molecular knot.12−15 With Ln(III) ion as template, a molecular trefoil knot with single handedness was recently fabricated by Leigh and co-workers.13 As active-metal template, Cu(I) ions can not only organize the formation of open knot but also induce the covalent bond formation via an alkyne−azide “click” reaction.50 Copper(I)-templated phanothroline complex popularized by Sauvage and co-workers can theoretically form different kinds of topological interesting molecules, and until now, synthetic molecular catenane and trefoil knot were reported.51 Herein, the copper(I)-templated phenanthroline complex was selected for the synthesis of trefoil knotted initiator due to its quantitative formation of trefoil knotted complex and modifiable terminal groups. The synthesized trefoil knotted initiator is one of the first synthetic functionalized molecular knot which can be used as the initiator for further polymerization.38 Compound 1 was obtained by a multistep synthesis starting with the commercially available 1,10-N,N′-phenanthroline (see Supporting Information and our previous publication for details).38 Pure monotosylated tri(ethylene glycol) (TEG) (2) can be obtained by a simple purification method (see Supporting Information). The column purification on the obtained EG-terminated phenanthroline ligand (3) results a viscous yellow solid although the 1H NMR spectrum shows acceptable purity. Further purification by recrystallization in methanol will afford a colorless solid powder, and successful modification on the both sides of compound 1 by tri(ethylene glycol) was confirmed by the 1H NMR spectrum as shown in Figure 1. The obtained compound 3 showed better solubility in most of the organic solvents, such as methylene dicholoride (DCM), chloroform (CHCl3), and tetrahydrofuran (THF), than compound 1. The complexation reaction was performed by slowly transferring the acetonitrile solution of Cu(CH3CN)4PF6 to the DCM solution of compound 3 by cannula in a nitrogen atmosphere. Our previous reports demonstrated the construction of the catenated and knotted complex by a visible color change and UV−vis spectroscopy analysis.38,52 From the C

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knotted complex compared with free knotted ligand also confirmed the secondary interactions. The two hydroxyl groups positioned together in its 3D conformation were reacted with the two methoxyl groups of one dibutyldimethoxytin to form a bis-copper(I) templated trefoil knotted initiator. The XPS spectrum of the trefoil knotted initiator revealed the retention of the copper ion to hold the tetrahedral template and the insertion of the tin initiator to close the two end of the bis-copper(I) templated open knot (see Supporting Information Figure S1). As shown in Figure S2, the lower retention volume of bis-copper(I) templated trefoil knotted initiator (5) compared with linear knot ligand (3) demonstrated its increased hydrodynamic volume. The complete disappearance of the absorption peak corresponding to the O−H stretching (Figure 2, complex 4 and initiator 5) also verified the successful formation of the Sn−O bond at both sides of the knotted complex.

Figure 1. Comparative 1H NMR spectra of EG-terminated phenanthroline ligand (3) and bis-copper(I) templated knotted complex (4).

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H NMR spectra of knotted ligand (3) and bis-copper(I) templated knotted complex (4) as shown in Figure 1, the formation of the open knot complex can be verified. Significant chemical shift was observed to the aromatic segment of the knot ligand due to the coordination interaction between the copper(I) ions and phenanthroline groups and the π−π interaction between phenyl rings (π donors) and phenanthroline moieties (π acceptors).51 The influence of the phenanthroline core on the phenyl rings leads to the significant shielding of protons j and k in Figure 1.14 Moreover, the complete disappearance of the signals (such as peak b) derived from the free knot ligand (3) confirmed the quantitative transformation of the knotted complex. The rational design of the knotted ligand does not only help the preorganization of a copper(I)-templated open knot but also favors the covalent bond formation of the desired terminal groups. Previous reports proved that the secondary interactions in the complex can promote the ring closure reaction by positioning the desired reactive groups of the ligand.51 The introduction of rigid groups, i.e., phenyl groups in this design, will provide structural restriction for the desired conformation.9 Aromatic interaction between the hydrogen of the aromatic segment and oxygen of the EG moieties plays a significant role in favoring the formation of trefoil knot than other isomers. Our previous molecular modeling and theoretical calculation also confirm the 3D conformation of trefoil knot complex with the two desired terminal groups positioning next to each other.38 Moreover, the weaker IR signals at 3200−3600 cm−1 arising from the O−H stretching of the bis-copper(I) templated

Figure 2. Comparative IR spectra of knot ligand (3), bis-copper(I) templated knotted complex (4), and bis-copper(I) templated knotted initiator (5).

Bis-Copper(I) Templated Trefoil Knotted Poly(L-Lactide). Different kinds of monomers, mostly cyclic esters, can be inserted inside the Sn initiator and form various kinds of polyesters. From a cyclic dibutyltin initiator, Kricheldorf et al. have demonstrated the polymerization of β-butyrolactone, εcaprolactone, and L-lactide under kinetic control with no side reactions observed.49,53 On the basis of an organic spirocyclic initiatior, Waymouth and co-workers also reported the synthesis of cyclic polylactide and poly(ε-caprolactone) via zwitterionic polymerization.54,55 Recently, our group also reported the first synthesis of catenated poly(L-lactide) by ring-expansion strategy.31 Herein, the synthesis of the trefoil knotted poly(L-lactide) was also attempted as shown in Scheme 2. Bis-copper(I) templated trefoil knotted poly(L-lactide) was synthesized by the ring-opening polymerization of L-lactide monomers. Further dialysis against DCM was performed under a nitrogen atmosphere to remove the unreacted monomers and other impurities. The reddish color of the final product demonstrated the stability of the bis-copper(I) templated phenanthroline complex of the templated trefoil knotted polymer. Moreover, as shown in Figure 4A, the proton signals of phenanthroline segment in the templated TK-PLA were completely consistent with the freshly prepared bis-copper(I) D

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Macromolecules Scheme 2. Synthesis of Trefoil Knotted Poly(L-lactide) and Trefoil Knotted Poly(L-lactide)-block-poly(ε-caprolactone)

Figure 3. Comparative DLS analysis of Cu(I)-Knot initiator (5), Cu(I)-Knot-PLA91 (6), and Cu(I)-Knot-PLA91-b-PCL31 (7).

templated phenanthroline complex, which confirmed the intact tetrahedral metal−ligand conformation in the center of the templated polymer.38 The ring-expansion growth of the molecular knot by coordination−insertion polymerization of the L-lactide monomers was demonstrated by the increased hydrodynamic radius (Rh) of the Cu(I)-Knot-PLA91 (6, Rh = 7.2 ± 0.3 nm) compared with that of the Cu(I)-Knot initiator (5, Rh = 1.5 ± 0.3 nm). As claimed earlier, the DPn was defined as the sum of the two polymer chains derived from one tin atom. For clarity,

Figure 4. Comparative 1H NMR spetra of (A) Cu(I)-Knot-PLA91 (6) and (B) Knot-PLA91 (8).

the feed ratio was also defined as the ratio of monomers to tin atoms. Therefore, the monomer number in one trefoil knotted polymer is twice the DPn due to the presence of the two tin E

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Macromolecules initiating sites per trefoil knotted initiator. The two broad peaks at 5.2 and 1.6 ppm (peak r and s in Figure 4A), which are corresponding to the proton between the two ester bonds and three protons of the methyl moiety next to it, also suggested the successful polymerization of the L-lactide. Based on the comparative integrations of the peak r in the PLA segment and peak d from the phenanthroline segment, the DPn of the templated trefoil knotted PLA was calculated. Together with the initial feed ratio of 100, the monomer conversion was calculated to be 91.5%. The two strong absorption peaks at 1753 and 1178 cm−1 in the FT-IR spectrum of the templated trefoil knot PLA91 are corresponding to the stretching vibrations of CO and C−O bonds in the poly(L-lactide) segment, respectively (Figure 5).

value of the templated trefoil knotted polymer for the templatefree trefoil knotted polymer.

Figure 6. Comparative GPC curves of Cu(I)-Knot-Initiator (5), KnotPLA91 (8, Mn = 21.14 kDa, Mw = 30.95 kDa, PDI = 1.46), and KnotPLA91-b-PCL31 (9, Mn = 28.36 kDa, Mw = 43.84 kDa, PDI = 1.55).

The lower elution volume of the Knot-PLA91 compared with the trefoil knotted initiator corroborated the increased hydrodynamic radius of the formed trefoil knotted polymer. The relatively lower hydrodynamic volume and broader polymer peak of Knot-PLA91 as compared to Cu(I)-KnotPLA91 demonstrated the high conformation freedom and more compact architecture, which is consistent with our previous report (Figure S4).38 Moreover, no significant peak or shoulder at lower retention volume was observed after decomplexation, suggesting the absence of single macrocycle or linear polymer. The relatively high polydispersity of the obtained Knot-PLA91 stemmed from the comparatively higher propagation rate compared to the initiation rate.57 Initial attempts to reduce the viscosity of the reaction system by polymerization in chloroform failed, and further studies on reducing the polydispersity are underway. Trefoil Knotted Block Copolymer. The poly(ε-caprolactone) and poly(L-lactide) are two well-known aliphatic polyesters that exhibit inherent biodegradability and biocompatibility. They exhibit different properties, such as morphology, mechanical strength, and biodegradation rate, and these differences are somehow complementary.58 Block copolymerization is usually adopted to achieve materials with optimized properties compared to both of the parent homopolymers.48,59 Because of the presence of the active Sn−O bond, the purified bis-copper(I) templated TK-PLA can further initiate the ring-opening polymerization of the εcaprolactone. Initial attempts were conducted in chloroform and toluene solution of the caprolactone. However, only the polymers with negligible PCL blocks can be obtained (DPn < 5). Bulk polymerization with feed ratio (monomer: Sn atom) of 200:1 was performed at 80 °C, and after 18 h polymerization, the obtained polymer was purified by dialysis against DCM. The dramatic increase in hydrodynamic radius of the purified bis-copper(I) templated trefoil knotted block copolymer (7, Rh = 10.1 ± 0.4 nm) compared with the homopolymer (6, Rh = 7.3 ± 0.3 nm) confirmed the further ring expansion of the

Figure 5. IR spectra of bis-copper(I) templated trefoil knotted PLA91 and bis-copper(I) templated trefoil knotted block PLA91-b-PCL31.

Template-Free Trefoil Knotted Poly(L-lactide). To obtain a geniune trefoil knotted polymer, the copper template was removed by the addition of potassium cyanide. The decomplexation process was monitored by the UV−vis spectroscopy. Upon decomplexation, the ligand-centered π−π* and n−π* transitions (absorptions below 380 nm) underwent a hyperchromic effect, and the metal-to-ligand charge transfer bands (absorptions at visible region) disappeared (Figure S3).56 The distinct chemical shift of the phenanthroline moieties in the Knot-PLA91 confirmed the complete removal of the copper template from the phenanthroline ligand. The well-resolved proton signals of phenanthroline segments in the Knot-PLA91 compared with that of the Cu(I)-Knot-PLA91 revealed the well separation of the two phenanthroline segments, and the proton correlation interactions between the two phenathroline groups was hardly observed from the NMR spectra. Also, the comparatively well-resolved feature of the template-free trefoil knotted polymer compared to the molecular trefoil knot with low molecular weights stemmed from the long flexible chain of the trefoil knotted polymer and hence faster intramolecular reptation process.11,38 It can be seen that the relative integration of peaks r and d in the 1H NMR spectrum of Knot-PLA91 was slightly different than that of the Cu(I)-KnotPLA91, which may be due to the well-known integration error of 1H NMR spectra. For consistency, we still used the DPn F

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significant chemical shift in the 1H NMR spectrum of KnotPLA91-b-PCL31 (9) compared with that of Cu(I)-Knot-PLA91b-PCL31 (7) confirmed the successful decomplexation. The reduced retention volume of the formed block copolymer (Knot-PLA91-b-PCL31) compared with the homopolymer (Knot-PLA91) (see GPC analysis, Figure 6) confirmed the increase in hydrodynamic diameter of the trefoil knotted polymer. Linear block copolymer PLA158-b-PCL29 was also synthesized for comparative property studies with NMR spectrum shown in Figure S6. With even higher absolute molecular weight (Mn,NMR = 35.66 kDa for Knot-PLA91-bPCL31 vs Mn,NMR = 26.38 kDa for block copolymer PLA158-bPCL29), the trefoil knotted block copolymer exhibited lower hydrodynamic volume due to its compact architectures, which is also consistent with our previous report.38 Similar to KnotPLA91, the obtained block copolymer Knot-PLA91-b-PCL31 also exhibited the well-resolved proton peaks. The fast intramolecular reptation process is also a unique property of the synthetic trefoil knotted polyester compared with the trefoil knot with low molecular weight.

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CONCLUSION

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ASSOCIATED CONTENT

By ring-expansion strategy, a templated trefoil knotted poly(Llactide) can be synthesized from a bis-copper(I) templated trefoil knotted tin initiator which was formed by the ringclosing reaction of bis-copper(I) templated phenanthroline complex with dibutyldimethoxytin. With initial monomer-toinitiator feed ratio of 100, Cu(I)-Knot-PLA with DPn of 91 can be obtained, and following decomplexation will render a geniune trefoil knotted PLA without the restriction of the copper template. The bis-copper(I) templated trefoil knotted block copolymer was obtained by subsequent insertion of εcaprolactone monomers from a Cu(I)-Knot-PLA. The 1H NMR spectrum analysis of the Cu(I)-Knot-PLA-b-PCL revealed a rather stable bis-copper(I) templated phenanthroline complex during the polymerization and purification process. GPC, IR, and DLS were also utilized to confirm the formation of the trefoil knotted block copolymer. The formation of the trefoil knotted block copolymer can bring some complementary properties to this topologically interesting polyester, such as adjustable degradation rate, which may be important for certain applications, especially in biomedicine. The capability of subsequent copolymerization of another monomer may provide a method to remove the tin atom by post-intramolecular cross-linking strategy.60,61 Studies on the synthesis of Sn-free trefoil knotted polyester as well as the unique properties of these topologically interesting macromolecules such as crystallinity and degradation rate are currently underway.

Figure 7. Comparative 1H NMR spetra of Cu(I)-Knot-PLA91-b-PCL31 (7) and Knot-PLA91-b-PCL31 (9).

trefoil knotted polymer. With well-controlled polymerization and purification conditions, the purified bis-copper(I) templated trefoil knotted block copolymer also exhibited stable biscopper(I) templated phenanthroline complex. The 1H NMR spectrum of compound 7 revealed the same aromatic signals with the initial bis-copper(I) templated knotted complex (4). The appearance of peaks x and t lying at 4.1 and 2.3 ppm, which are typical signals for poly(ε-caprolactone), confirmed the insertion of the ε-caprolactone monomers between the Sn− O bond of the Cu(I)-Knot-PLA91. Fixing the integration of the peak k in the phenanthroline segment, the relative integration of peak x can be obtained, and the DPn (related to one Sn atom, 2n in Figure 7) was calculated to be 31. Moreover, the similar DPn, i.e., 92, of the PLA segments in block copolymer (according to the integration from the 1H NMR spectrum of Cu(I)-Knot-PLA91-b-PCL31) compared with the one before copolymerization (DPn(PLA) = 91) demonstrate the stability of the PLA segments during the polymerization process of the εcaprolactone monomers. When the absorption intensity of the ester bond (1750 cm−1, CO stretching) is fixed, the absorption intensity of the CH2 bond (2800−2950 cm−1, stretching vibration of C−H bond) in the Cu(I)-Knot-PLA91-bPCL31 (7) increased compared with that of the Cu(I)-KnotPLA91 (6) as shown in Figure 5, which demonstrated the reduced density of ester bond in the block copolymer. A similar decomplexation method was performed to remove the restriction of copper template in order to obtain a genuine trefoil knotted block copolymer. The disappearance of the absorption peaks in the visible region of the UV−vis spectrum of the Knot-PLA91-b-PCL31(Figure S5) along with the

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.macromol.6b02029. Synthesis of compound 2 and synthesis scheme of compound 1; XPS spectra of complex 4 and initiator 5; UV−vis spectra of bis-copper(I) templated and templatefree trefoil knotted polymer (PDF) G

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AUTHOR INFORMATION

Corresponding Author

*(R.C.A.) E-mail [email protected]. ORCID

Rigoberto C. Advincula: 0000-0002-2899-4778 Author Contributions

P.-F.C. and L.-H.R. contributed equally to this work. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS We acknowledge funding from the National Science Foundation (NSF): NSF-1608457 and NSF-1333651. P.-F. Cao also acknowledge partial financial support by the U.S. Department of Energy, Office of Science, Basic Energy Science, Material Science and Engineering Division.

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NOMENCLATURE Cu(I)-Knot-PLAm: bis-copper(I) templated trefoil knotted poly(L-lactide) with degree of polymerization (DPn) to be m; Cu(I)-Knot-PLAm-b-PCLn: bis-copper(I) templated trefoil knotted block copolymer poly(L-lactide)-block-poly(ε-caprolactone); degree of polymerization (DPns) of poly(L-lactide) block and poly(ε-caprolactone) block are m and n, respectively; KnotPLAm: trefoil knotted poly(L-lactide) after the removal of the copper template; Knot-PLAm-b-PCLn: trefoil knotted block copolymer poly(L-lactide)-block-poly(ε-caprolactone) after the removal of the Cu template; degree of polymerization (DPn) of poly(L-lactide) block and poly(ε-caprolactone) block are m and n, respectively.

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