Supplementary Figure S1. Rheological characterisation of hydrogels. (a) Frequency sweeps and (b) strain sweeps for gels formed from 1 alone (blue

Transcription

1 Supplementary Figure S1. Rheological characterisation of hydrogels. (a) Frequency sweeps and (b) strain sweeps for gels formed from 1 alone (blue data), 2 alone (red data) and a mixed system of 1 and 2 (black data). In all cases, closed symbols represent G. Open symbols represent G. For all gels, a GdL concentration of 12.6 mg/ml was used to adjust the ph. The total peptide concentration was 5 mg/ml in all cases. 1

2 Supplementary Figure S2. ph data on ph adjustment with GdL. ph data with time for a mixture of 1 and 2 with different amounts of added GdL. Left, the time-axis is linear. Right, the time-axis is on a log scale to highlight the early time points. For both graphs, 5.4 mg/ml GdL (blue), 12.6 mg/ml GdL (red) and 43 mg/ml GdL (black). The dashed lines represent the pka of (top) 1 and (bottom) 2. 2

3 Supplementary Figure S3. Scanning electron microscopy (SEM) of gels. (a) SEM of 1, (b) SEM of 2, (c) SEM of a mixture of 1 and 2. Scale bar represents 200 nm in each case. 3

4 Supplementary Figure S4. Statistical analysis of SEM data. Statistical analysis of the fibril diameters of 1, 2 and a mixture of these. Each analysis is based on 283 (1), 127 (2) and 180 (Mix) unique fibril measurements. 4

5 Supplementary Figure S5. Small angle neutron scattering data. SANS data for 1 ( ), 2 ( ) and d-2 ( ). For 1 alone, there is a peak in the data at Q max 0.1 Å -1 indicating a discontinuity in the scattering length density of the radial structure at around, thus, R = 63 Å. The fit parameters for 1 using the Kholodenko-Dirac worm-like chain model suggest that the fibrous structures are rather rigid, with relatively few (m = 10) short links (l = 150Å) and hence L = 1500 Å. For 2 (Fig. S4), the structures are more flexible (m = 70), with longer links (l = 360 Å) compared to 1 and hence L = Å. 5

6 Supplementary Figure S6. Small angle neutron scattering data for deuterated and nondeuterated gelator. Comparison between the SANS data for 2 ( )and d-2 ( ).The scattering from 2 and d-2 effectively collapsed onto a common curve once the relative scattering length densities and incoherent backgrounds had been accounted for, indicating the presence of a homogeneous structure with no local segregation (nanoscale ordering) of deuterated and nondeuterated sections of the molecule. 6

7 Supplementary Figure S7. Small angle neutron scattering data for deuterated and nondeuterated gelator in mixed systems. Comparison of the SANS data for ( ) 1 and 2, ( ) 1 and d-2. 7

8 Supplementary Figure S8. Fiber X-ray diffraction data for gels formed using different triggers. Overlay of fxrd for GdL triggered self-sorting (top) and HCl triggered gelation (bottom). 8

9 Supplementary Table S1. The final ph of gels formed from mixing 1 and 2 as determined by the amount of GdL added. GdL concentration (mg/ml) Final pd

10 Supplementary Methods Materials. Dipeptide gelators 1 and 2 were prepared as described previously. 13,14 The deuterated analogue of 2 was prepared in an analogous manner from 2-naphthol-d 7, purchased from Qmx Laboratories, UK. During the first step of the synthesis, one of the deuteriums on the aromatic ring exchanged for a proton, presumably due to the strongly basic conditions. After this step, no further H-D exchange occurred. Hence, all compounds are assigned as d 6. Tert-butyl 2-(naphthalene-d 6-2-yloxy)acetate (S1): 1 H NMR (400 MHz, CDCl 3 ) δ 7.07 (s, 1H), 4.63 (s, 2H), 1.50 (s, 9H). 13 C NMR (100 MHz, CDCl 3 ) δ 167.9, 155.8, 134.2, 129.2, 107.0, 82.4, 65.8, The crude material was directly deprotected using trifluoroacetic acid as described elsewhere. 1,2 2-(Naphthalen-d 6-2-yloxy)acetic acid (S2): 1 H NMR (400 MHz, DMSO) δ 7.27 (s, ArH, 1H), 4.80 (s, OCH 2, 2H). 13 C NMR (100 MHz, DMSO) δ 170.0, 155.5, 133.9, 128.5, 106.9, 64.5 ppm. 13 C NMR (DMSO) 170.0, 155.5, 133.9, 129.7, 106.9, MS (m/z) [M+NH 4 ] Methyl 2-(2-(naphthalene-d 6-2-yloxy)acetamido)propanoate (S3): 1 H NMR (400 MHz, CDCl 3 ) δ 7.19 (bs, 1H), 7.17 (s,1h), 4.72 (m, 1H), 4.63 (s, 2H), 3.76 (s, 3H), 1.48 (d, J = 7.2 Hz, 2H,). 13 C NMR (100 MHz, CDCl 3 ) δ 172.9, 167.7, 166.6, 134.2, 129.4, (t), (m), (t), (t), 107.7, 167.4, 52.6, 47.7, 30.9, The crude material was directly deprotected using lithium hydroxide as described elsewhere. 1,2 2-(2-(Naphthalen-d 6-2-yloxy)acetamido)propanoic acid (S4): 1 H NMR (400 MHz, DMSO) δ 8.45 (d, J = 8.0 Hz, 1H), 7.30 (s, 1H), 4.65 (d, J = 14.7 Hz, 1H), 4.61 (d, J = 14.7 Hz, 1H), 4.33 (m, 1H), 1.34 (d, J = 7.3 Hz, 3H). 13 C NMR (100 MHz, DMSO) 173.8, 167.5, 155.4, 133.9, 128.8, (m), (m), (t), (t), 107.3, 66.6, 47.3, HRMS (m/z) [M+Na] + calcd. for C 15 H 9 D 6 NO 4 Na: ; found: Methyl 2-(2-(2-(naphthalen-d 6-2-yloxy)acetamido)propanamido)propanoate (S5): NMR (400 MHz, CDCl 3 ) δ 7.23 (bd, J = 7.2 Hz, 1H), 7.16 (s, 1H), 6.57 (bd, J = 7.0 Hz, 1H), 4.64 (m, 3H), 4.56 (m, 1H), 3.76 (s, 3H), 1.45 (d, J HH = 7.2 Hz, 3H), 1.38 (d, J = 7.2 Hz, 3H). 13 C NMR (100 MHz, CDCl 3 ) δ 173.1, 171.2, 166.0, 154.9, 134.2, 129.4, 107.5, 67.2, 52.6, 48.4, 48.2, HRMS (m/z) [M+Na] + calcd. for C 19 H 16 D 6 N 2 O 5 Na: ; found: (2-(2-(Naphthalen-d 6-2-yloxy)acetamido)propanamido)propanoic acid (d-2): 1 H NMR (400 MHz, DMSO) δ 8.17 (d, J = 7.3 Hz, 1H), 8.18 (d, J = 17.8 Hz, 1H), 7.31 (s, 1H), 4.66 (d, J = 15.2 Hz, 1H), 4.62 (d, J = 15.2 Hz, 1H), 4.44 (m, 1H), 4.20 (m, 1H), 1.27 (m, 6H). 13 C NMR (100 MHz, DMSO) δ 173.9, 171.7, 166.9, 155.5, 133.9, 128.5, 107.3, 66.8, 47.5, 18.4, HRMS (m/z) calcd. for C 18 H 18 D 6 N 2 O 5 Na: ; found: Rheology. All rheological experiments were performed using an Anton Paar Physica MCR101 rheometer. Parallel plates (a sandblasted parallel top plate with a 50 mm diameter 1 H 10

11 was used with a 1.0 mm gap distance) were used to perform the frequency and strain sweeps. All gels were formed directly on the rheometer and left overnight at room temperature to gel before the measurements. Evaporation of water from the gel was minimized by covering the sides of the plates with low-viscosity mineral oil. Frequency scans were performed from 0.1 rad/s to 100 rad/s under a strain of 0.5%. At this strain, the frequency sweeps measurements were performed within the linear viscoelastic region, where the storage modulus (G ) and loss modulus (G ) are independent of the strain amplitude. Small-Angle Neutron Scattering (SANS). 1 and 2 were dissolved in D 2 O with equimolar amounts of NaOD. The final pd of the solutions were measured to be 10.5 after fine adjustment of the pd with dilute NaOD. Individual gelation of the systems was initiated by thorough but gentle mixing of GdL with a solution containing either 1 or 2. Mixed gels containing both systems were prepared by mixing GdL with a solution of 1 and 2. Smallangle neutron scattering (SANS) measurements were performed on the fixed-geometry, timeof-flight LOQ diffractometer (ISIS Spallation Neutron Source, Oxfordshire, UK). A white beam of radiation with neutron wavelengths spanning 2.2 to 10 Å was used to access a Q [Q = 4πsin(θ/2)/λ] range of to 0.25 Å -1 (25Hz), with a fixed sample-detector distance of 4.1 m. Samples of volume ~0.4 ml were contained in 2 mm path length, UVspectrophotometer grade, quartz cuvettes (Hellma) and mounted in aluminium holders on top of an enclosed, computer-controlled, sample chamber with temperature controlled to 25 ± 0.5 o C by use of a thermostatted circulating bath pumping fluid through the base of the sample chamber. Experimental measuring times were approximately 40 minutes. All scattering data were (a) normalized for the sample transmission, (b) background corrected using an empty quartz cell or one filled with the appropriate solvent (this also removes the inherent instrumental background arising from vacuum windows etc.) and (c) corrected for the linearity and efficiency of the detector response using the instrument-specific software package. The data were put onto an absolute scale by reference to the scattering from a partially deuterated polystyrene blend. Data were fitted using the Kholodenko-Dirac worm-like chain model, which analyses the data as a Gaussian distribution of m connected cylindrical elements of statistical length l and radius R ax, such that the contour length, L, is L = m.l. This model therefore interpolates between the expected Q -1 dependence for the rod-like character of the cylindrical elements, the Q -2 associated with the cross-section of the cylinder and a limiting Q -4 associated with the globular nature over large distances. TEM. Samples of the gels were prepared for inspection by TEM by adsorbing material in situ to a grid surface. Gelation of the individual and mixed systems were initiated as described for fibre X-ray diffraction. 10 µl aliquots were placed in a humid chamber and allowed to gel with an inverted 400-mesh copper TEM grid (Agar scientific) suspended on the solution for 24 hrs. These were prepared for visualisation by a 1 minute water wash and two 1 minute negative stains using 2% w/v uranyl acetate. Grids were dried and viewed using a Hitachi TEM operated at 100 kv. Images were collected digitally using an axially mounted 2 megapixel Gatan Ultrascan 1000 CCD camera (Gatan, Oxford, UK). 11

12 Fibril measurements were made using ImageJ 31 and characteristic morphologies selected and cropped using scripts and the in-built Fast Fourier transform bandpass filter. The fibril diameters for discrete morphologies were first confirmed by manual measurement and subsequently to produce an unbiased statistical analysis of fibril morphology and diameter 283, 127 and 180 unique fibril and fibril interactions were measured respectively for 1, 2 and the mixture. These were plotted on histograms and a 5-point Savitzky and Golay filter 32 applied to represent the range of fibril sizes. Mean fibril diameters and standard deviations were calculated from the histogram data for the peaks corresponding to each fibril morphology. SEM. For SEM imaging, 400 mesh copper TEM grids with lacy carbon films were purchased from Agar and graphene oxide (GO) films were deposited on them as described elsewhere. 33 Samples of the gels were prepared by lightly touching the GO grid surface onto the gel, followed by drying overnight in a dessicator. The GO grids were directly mounted onto an aluminium stub without the need for coating. The samples were examined with a ZEISS SUPRA55VP FEGSEM electron microscope using a 3, 5 or 10kV accelerating voltage and an in-lens or standard SE detector. 12

13 Supplementary References 31 Abramoff, M. D. et al. Image Processing with ImageJ. Biophotonics International 11, (2004). 32 Savitzky, A. & Golay, M. J. E. Smoothing and Differentiation of Data by Simplified Least Squares Procedures. Anal. Chem. 36, (1964). 33 Patterson, J. P et al. A Simple Approach to Characterizing block Copolymer Assemblies: Graphene Oxide Supports for High Contrast Multi-Technique Imaging. Soft Matter 8, (2012). 13