Letter pubs.acs.org/macroletters
Pure Anisotropic Hydrogel with an Inherent Chiral Internal Structure Based on the Chiral Nematic Liquid Crystal Phase of Rodlike Viruses Xiaodong Pei,† Tingting Zan,‡ Hengming Li,† Yingjun Chen,† Linqi Shi,† and Zhenkun Zhang*,† †
Key Laboratory of Functional Polymer Materials of Ministry of Education, Institute of Polymer Chemistry, College of Chemistry, Nankai University, Tianjin 300071, China ‡ School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China S Supporting Information *
ABSTRACT: Imparting ordered structures into otherwise amorphous hydrogels is expected to endow these popular materials with novel multiple-stimuli responsiveness that promises many applications. The current contribution reports a method to fabricate pure polymeric hydrogels with an inherent chiral internal structure by templating on the chiral nematic liquid crystal phase of a rodlike virus. A method was developed to form macroscopically homogeneous chiral templates by confinement induced self-assembly in the presence of monomers, cross-linkers and initiators. Polymerization induced gelation was performed without perturbing the elegant 3D chiral organization of the rodlike virus bearing double bonds. Furthermore, a suitable method was found to remove the organic virus template while keeping the desired polymeric replica intact, resulting in a pure polymeric hydrogel with a unique internal chiral feature that originates from the 3D chiral ordering of the cylindrical pores left by the virus. Multiple-stimuli responsiveness has been demonstrated and can be quantified by the change of the pitch of the chiral feature. The chiral structure endows the otherwise featureless hydrogel with a unique material property that might be used as a readout signal for sensing and acts as the basis for responsive, biomimetic nanostructured materials.
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sensitive to pH and ionic strength.9 In addition, while structure homogeneity is critical for sharp and ambiguous responsiveness, hydrogels with a homogeneously ordered internal structure up to the macroscopic scale are difficult to be achieved.10 Furthermore, the unique properties of anisotropic hybrid hydrogels reported so far are largely determined by the embedded structure components of the templates.1a,d Removal of the templates from the hybrid gels often leads to loss of the structure feature and accompanying functionalities. Therefore, pure polymeric hydrogels with inherent structural features independent of the guiding templates are rare. In the current work, we shall report a strategy to imprint the chiral nematic liquid crystal (CLC) phase of a monodisperse rodlike virus into polymeric hydrogels (Scheme 1). Etching away the virus template results in a pure polymeric hydrogel with a unique internal chiral feature that originates from the 3D chiral ordering of the cylindrical nanopores left by the viruses. The chiral feature distributes homogeneously along the whole macroscopic hydrogel at the centimeter scale and manifests as visible stripe-like fingerprints. Discernable change of such a chiral feature can be induced by multiple stimuli such as the
mparting ordered internal structures into otherwise amorphous hydrogels to create structured hydrogels is expected to produce novel multiple-stimuli responsiveness such as deformation induced structure color changes, tunable photonic bandgaps, unidirectional (de)swelling, and so on.1 These properties have further extended the application of the hydrogel into many other fields, including but not limited to mechanochromic sensors,2 deformation-based display devices,3 soft actuators,4 artificial muscles,5 and so on. To create structured hydrogels, the most versatile method is direct polymerization of gel precursors in the presence of preformed structure guiding templates.1a,d,3,6 Typical examples include photonic hydrogels based on colloidal crystals1b and anisotropic hydrogels templated by anisotropic structures of disc-like clay particles,1d lamellar structure of surfactants,1a the hexagonal lyotropic phase of the wormlike micelles,6 and so on. Even with such significant progresses, the types of the internal structure imparted by the templates are very limited.7 In contrast, many biological molecules or nanoobejcts, such as DNA, cellulose nanocrystals, and rodlike viruses, can self-assemble into intriguing three-dimensional (3D) hierarchical structures,8 which can be exploited as templates to structured hydrogels. However, it is highly challenging to replicate with high fidelity such subtle templates by in situ polymerization without perturbing their elegant 3D structures, which are extremely © XXXX American Chemical Society
Received: September 17, 2015 Accepted: October 20, 2015
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DOI: 10.1021/acsmacrolett.5b00677 ACS Macro Lett. 2015, 4, 1215−1219
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ACS Macro Letters
terminal or lysine residues at the position of 6 of pVIII (Scheme S1, see SI for the detailed chemistry). The virus bearing about 1000 double bonds per virus was used for the current work. As a proof of concept, the gel matrix is the conventional polyacrylamide hydrogel. In order to imprint the CLC phase into the hydrogel, M13-AM was mixed with acrylamide (AM), N,N-methylenebis(acrylamide) (BIS) as the cross-linker, and ammonium persulfate (APS) as the thermal initiator, resulting in a mixture containing 30 mg mL−1 M13-AM. Under such condition, the virus suspension is in the concentration range for the nematic LC phase.13 The mixture was loaded into a glass capillary with a diameter of 1 mm. After being stored at 4 °C for 5 h, the CLC phase formed in the whole samples, resulting in fingerprints that consist of black and white stripes (Figure 1A). The distance between two dark and bright strips corresponds to the pitch (P), the periodicity of the 3D chiral ordering (inset of Figure 1A). By carefully avoiding air bubbles and cleaning the internal surface of the glass capillary (see SI), the fingerprints develop along the long axis of the capillary, resulting in a homogeneous internal structure inside the whole samples. This confinement induced self-assembly (CISA) can be attributed to the alignment of the rodlike virus with the long axis of the capillary tube to minimize the interaction energy of the semirigid rod with the curved internal surface.14 The homogeneous yellow color is also a sign of a monodomain CLC phase, in contrast to the polydomain structure formed by many templates, as reported previously.10 The capillary was incubated at 40 °C to initiate the polymerization, resulting in a cylindrical gel (Figure 1B). It is noted here there was no phase separation and aggregation of the virus during polymerization, as monitored by POM. The hybrid gel was then taken out by breaking the glass capillary. The diameter of the final cylindrical hybrid gel is set by the internal diameter of the glass capillary while its’ length can be varied to more than several centimeters by changing the volume of the prepolymerization mixture. The gel can be knotted and stretched (Figure 1B). Due to the imprinted CLC phase of the virus, the gel has strong birefringence between cross polarizers (Figure 1B). In the cylindrical gel, the typical fingerprints of the CLC phase are identical to that formed before the polymerization, and both have a pitch of 36 μm, suggesting accurate replication of the viral template during polymerization (Figure 1C). Intriguingly, it is possible to observe the radial interface perpendicular to the long axis of the gel cylinder, which manifests as elegant concentric fingerprints with a center similar to the Chinese Yin-Yang figure under POM (Figure 1D). For comparison, other two kinds of gels were also prepared under the same conditions (Figure S3, as detailed in SI). The results indicate that the double bonds covalently coupled to the surface of the virus and the external cross-linkers-BIS play important roles, and only the above mixture can achieve high-fidelity replication of the template structure.6 Although the CLC phase of the rodlike virus has been intensively investigated and implemented in different containers,8d,13 we believe this is the first case that such structure can exist out of the containers by being imprinted into a polymeric gel. To remove the virus template by in-gel chemical etching, the hybrid gel was immersed into a 1 M NaOH for some time, and then rinsed with a solution containing 2.5 (w/v)% NaOH and 10 (w/v)% SDS (see SI). The etching process finished in less than 10 min (Movie S1 in SI). The viruses were disrupted into
Scheme 1. Illustration of the Preparation Procedure of Pure Polymeric Hydrogels with an Inherent Chiral Structurea
a Step I: Confinement-induced formation of the CLC phase of the M13 virus in the presence of the gel precursors; Step II: hybrid hydrogel formation resulted from thermal-initiated polymerization; Step III: In-gel chemical etching to remove the virus template.
mechanical force, swelling/deswelling, environmental humility, and so on, and can be quantified by the pitch. While there are emerging examples of liquid crystal elastomers that are based on thermotropic LC polymers,4b the current result represents one of the first examples of such materials based on colloidal rods. The virus recruited herein is the rodlike M13 bacteriophage that has a length of 880 nm and a diameter of only 6.6 nm.11 Besides its highly monodisperse size and nanorodlike shape, this virus can form a CLC phase, in which the long axis of the rodlike virus arranges in a helical way around a director, resulting in stripe-like fingerprints with a periodicity (pitch, P) in the micrometer range that can be conveniently observed by polarized optical microscope (POM; Figure 1A).8c M13
Figure 1. Hybrid PAAm hydrogels with the imprinted CLC phase of the M13 virus. (A) Chiral fingerprints of the CLC phase of the M13 virus in the presence of hydrogel precursors before polymerization. (B) Hybrid (bright colored) and pure PAAm (dark) hydrogels as observed between cross polarizers. Inset: a knotted hybrid hydrogel. (C, D) Chiral fingerprints along (C) and perpendicular to the long axis (D) of the cylindrical hybrid gel. Insets in (C) and (D) illustrate the plane of observation. Scale bar: 500 μm (A, C, and D) and 1 mm (B).
consists of about 2700 major coat proteins-pVIIIs, which assemble in a helical way into a cylindrical protein capsid. The surface of the virus has many functional groups that offer rich chances for chemical modifications.12 For the current work, in order to introduce double bonds onto the virus surface, we first modified the virus with N-hydroxysuccinimide methacrylate (HSAM) through the NHS and the amino groups of the N1216
DOI: 10.1021/acsmacrolett.5b00677 ACS Macro Lett. 2015, 4, 1215−1219
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ACS Macro Letters single coat proteins by the strong basicity since M13 is basically a protein assembly held together by noncovalent interactions.15 SDS can bind to the disassembled coat proteins to facilitate their complete removal from the gel matrix. The complete remove of the virus template was confirmed by coomassie brilliant blue staining of the coat proteins. While the original hybrid virus gel is stained and has the classic blue color, the gel without the virus appears as colorless as the pure PAAm gel (Figure 2A). Ethidium bromide staining of the viral DNA with enhanced sensitivity down to ng further confirms the complete removal of the virus (Figure 2B).
Figure 3. Internal structure of the hydrogels by SEM (A) and (B) hybrid hydrogel. (C, D) Etched hydrogel. Scale bar: 10 μm (A), 100 nm (B, D), and 500 nm (C).
direction which rotates in a way relative to each other in certain angle (Figure 3B). After etching away the virus templates, many channels-like structures appear, the long axis of which also rotates in a helical way (Figures 3C,D and S8C,D in SI). The channels must be left by the viruses. In the next, SAXS investigations were performed with the hydrated samples. For the virus suspension before polymerization, there is a peak in the scattered intensity (I) versus the scattering vector (q), due to the positional ordering of the rodlike virus in the CLC nematic phase.16 After formation of the gel, a scattering peak also exists in the similar positon of q, suggesting the positional ordering of the viruses is fixed in the gel state. Most importantly, there is still a clear peak in the case of the pure chiral hydrogel after etching away the virus. Therefore, the cylindrical voids left by the disassembled viruses still possess positional ordering. Finally, the N2 adsorption−desorption isotherm of the etched gel measured at 77 K appears as a reversible sorption profile with a hysteresis loop in the whole relative pressure range (inset of Figure 4B), supporting the presence of abundant mesopore structure in the chiral gel after etching away the virus template. In contrast, no pore-like feature can be detected in the template-included hydrogel (Figure S10 in SI). Pore size distributions of the etched gel based on Barrett−Joyner−Halenda (BJH) analysis of the desorption branch gave a mean pore diameter of 6 nm (Figure 4B). This size is surprisingly consistent with the 6.6 nm of the virus diameter, suggesting the pores are mainly the cylindrical voids left by the etched-away virus. The current method has unprecedented advantage of structure tailorability in creating hydrogels with versatile structure features in a single hydrogel body.17 The pitch of the CLC phase of the original virus template can be changed by the ionic strength of buffer and concentration of the virus, leading to final pure chiral gels with a pitch in the range of 80 to 30 μm (Figure S7A,B in SI). Combining different concentrations of the viruses into the same glass capillary successively results in a gel that consists of segments with a chiral feature of different pitches (Figure S7C in SI). Alternatively, a gel with one segment of a normal nonchiral nematic feature and another one a chiral feature can be also created (Figure S7D in SI). Compared to the normal gel, the best advantage of the current hydrogel is that the chiral structure endows the otherwise featureless hydrogel with a unique material property. Especially, the visible fingerprints of the internal chiral feature
Figure 2. Hydrogels after etching away the virus template. (A) Coomassie brilliant blue staining of the hydrogels. From top to bottom: hybrid hydrogel containing the virus template, etched hydrogel and pure PAAm hydrogel. (B) Ethidium bromide staining of hydrogels: the black and transparent gels are those before and after etching away the virus, respectively. (C, D) Chiral fingerprints along (C) and perpendicular to the long axis (D) of the etched hydrogel. (E) Complete dried hydrogel. Scale bar: 1 mm (A, B), 500 μm (C, D), and 200 μm (E).
During etching, the strong vivid color of the original hybrid gel due to birefringence faded out under POM (Figure S2 and Movie S1 in SI). However, in the etched gel, stripe-like fingerprints characterizing the chiral feature are still observed (Figure 2C). After removing the virus template, the gel swelled and the pitch of the fingerprints increased to 55 μm (Figure 2B). Again, concentric fingerprints can be observed under POM in the radial interface perpendicular to the long axis of the gel cylinder (Figure 2D). Even in the completely dried gel state, the fingerprints still exist and can be visualized by POM (Figures 2E and S6 in SI). The gel can be stored in the hydrated or dried state without losing the chiral feature for at least six months. These results suggest the chiral feature is robust and can be regarded as an inherent property of the gel. This is probably the first report of such centimeter-scale ordered structured feature developed in a pure polymeric hydrogel. The elastic modulus (E) was obtained from uniaxial elongation tests (Figure S9, and detail in SI) and 0.11, 5.60, and 0.03 MPa was obtained for the pure PAAm hydrogel, hybrid virus chiral gel, and etched chiral hydrogel, respectively, revealing their reasonable mechanical strength. The hydrated gels were turned into aerogels via frozen drying and the cracked aerogels were checked by SEM. While the normal PAAm aerogel is featureless (Figure S8A in SI), the hybrid virus gel has a lamellar structure typical of the materials originated from the CLC phase (Figure 3A and S8B).9b The rodlike feature exists in each layer and points to one general 1217
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Figure 5. Multiple stimuli responsiveness of the chiral hydrogel. (A) Swelling/deswelling behavior as characterized by the change of the pitch of the chiral fingerprint. Inset: on−off switch of the pitch during swelling/deswelling cycles. (B) Change of the size and pitch versus time during swelling. (C) Mechanical pressing induced reversible and fast change of the pitch. (D) Humility responsiveness. Insets: on−off switch of the pitch during humility switching cycles.
external stimuli are unambiguous and sharp, which can be used as a readout signal for sensing. In summary, a straightforward method has been devised to imprint the precious chiral LC phase of a rodlike virus into a common hydrogel made from the polymerization of AAm. To enhance the fidelity of the replication of the chiral structure, double bonds were introduced onto the surface of the virus. The virus can be removed by chemical etching under strong basic conditions, and the hydrogel still own the chiral structure left by the virus. Such properties endow the otherwise featureless PAAm hydrogel with a unique material property. The swelling and contraction of the current pure polymer hydrogel are accompanied by predictable internal structure changes on the molecular and nanometer length scale which manifest as the visible and quantitative change of the pitch of the chiral fingerprints. Therefore, the current “all-organic” hydrogel owns an inherent parameter that might be used as a readout signal for sensing and acts as the basis for responsive, biomimetic nanostructured materials.
Figure 4. SAXS of hydrogels at varied stages (A) and pore size distribution of the chiral hydrogel (B). The curves labeled with a, b, and c in (A) correspond to the virus mixtures before polymerization, hybrid gel, and etched gel, respectively. Inset in (B): N2 adsorption− desorption isotherm measured at 77 K.
and its quantitative parameter- the pitch (P) can be used as useful diagnostic parameters. For this, the swelling/deswelling behaviors of the chiral gel were first investigated (Movie S2). The chiral gel exhibits a superabsorbent capability and can absorb water with an amount of 270× its original dried weight (Table S1 in SI). The swelling behavior can be directly quantified by the change of the pitch of the chiral feature which reversibly increases and decreases during swelling and deswelling, respectively (Figure 5A). Such a procedure is highly reversible and can be performed for unlimited times without deteriorating the chiral feature. By monitoring the pitch versus time during swelling, it is found that the system approaches the swelling equilibrium state after >20 min, while the physical dimensions of the gel become constant during less than 15 min. This can be ascribed to radial diffusion of water into the internal core of the gel. Therefore, the microscopic pitch offers a more sensitive parameter to confirm whether or not a gel approaches a true swelling equilibrium. During swelling of the thoroughly dried gel, anisotropic swelling behavior occurred (Figure 5B). The swelling ratio (D/D0) in the diameter direction of the cylindrical gel is about 3, while 1.6 is found along the long axis (L/L0). In the next, the response of the chiral gel to mechanical forces was investigated. Under compression (inset of Figure 5C), the pitch increases from 45 to 60 μm with increasing applied stress up to the limited value without destroying the gel. This pitch change was reversible and fast: the gel quickly responses to an applied mechanical stress/strain and then returns to the initial pitch after removing the stress/strain. Finally, the pitch of the chiral gels at various humilities was monitored. A nearly linear relationship between the pitch and humility are found (Figure 5D). It is noted here that the changes of the chiral fingerprint during aforementioned
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsmacrolett.5b00677. Experimental details of the synthetic procedures, characterization, multiple stimuli responsiveness investigation and supplementary figures (PDF). Chemical etching of the virus template from the hybrid hydrogels. Speed: 60× (AVI). Swelling behavior of the etched hydrogel. Speed: 90× (AVI).
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AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest. 1218
DOI: 10.1021/acsmacrolett.5b00677 ACS Macro Lett. 2015, 4, 1215−1219
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ACKNOWLEDGMENTS This work was supported by the National Natural Science Foundation of China (Nos. 21274067, 91127045, 51390483), the Fundamental Research Funds for the Central Universities, Natural Science Foundation of Tianjin, China (No. 12JCQNJC01800), and PCSIRT (IRT1257).
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