Hydrogen bonding asymmetric star-shape derivative of bile acid leads to supramolecular fibrillar aggregates that wrap into micrometer spheres

Star-shaped asymmetrically grafted low molecular weight polymers with hydrogen bonding end-groups undergo aggregation to nanofibers, which subsequently wrap into micrometer spherical aggregates.


Fourier Transform Infra-Red Spectroscopy (FT-IR)
FT-IR spectra of solid samples were measured on a Nicolet 380 spectrometer (Thermo Fisher Scientific) in transmission mode with an ATR (diamond), 2 cm -1 resolution and averaged over 64 scans.

Small-Angle X-ray Scattering (SAXS)
Samples with varying DMSO/H 2 O ratio were prepared by adding ultrapure water to CA(AGE 6 -C 6 H 12 -UPy) 4 dissolved in DMSO. Addition of water caused formation of larger assemblies which was visually observed as a milky appearance of the samples. The samples were sealed between Kapton foils during the SAXS measurement. The sample environment was evacuated to reduce scattering from air. The SAXS was measured using a rotating anode Bruker Microstar microfocus X-ray source (Cu Kα radiation, λ = 1.54 Å). The beam was monochromated and focused by a Montel multilayer focusing monochromator (Incoatec).
The X-ray beam was further collimated by a set of four slits (JJ X-ray). The size of the X-ray beam at the sample position was < 1mm. The scattered intensity was collected using a Hi-Star 2D area detector (Bruker). Sample-to-detector distance was 1.59 m, and silver behenate standard sample was used for calibration of the length of the scattering vector q. Onedimensional SAXS data were obtained by azimuthally averaging the 2D scattering data. The magnitude of the scattering vector q is given by q = 4π sinθ/ λ, where 2θ is the scattering angle.

DLS measurements
DLS data was achieved with Zetasizer Nano ZS instrument (model no. ZEN3600) with two different concentrations: 1 mg/mL and 0.1 mg/mL of sample in the used solvent mixtures. were obtained from Sigma-Aldrich and used as received.

Dialysis
Dialyses were performed using Spectra/Por 7 Standard Regenerated Cellulose tubing (Mw cut-off 2000 Da). Ultrapure deionized water with resistivity of 18 MΩ/cm was used. In a typical dialysis procedure, the sample was dissolved in DMSO at 1 mL/mg concentration. 1 mL of the solution was transferred to a dialysis tube, air was extracted from the tube and it was sealed tightly, but with extra tube length so that the volume inside the tube can increase without increase in pressure.

Synthesis of CA(AGE 6 -C 6 H 13 ) 4
CA(AGE 6 -C 6 H 13 ) 4 (Fig. S11, 7) was prepared by adding 39.3 mg hexyl isocyanate (6) to a dry 10 mL round-bottom flask. Also 5 mL of dry chloroform, 100 µL dibutyltin dilaurate (DBTDL) and 0.2 g of 1 were added. The flask was equipped with a condenser and a CaCl 2 tube. The mixture was stirred and refluxed for 24 h. After the reaction was finished, 1 mL of water was added and the mixture was dried with rotary evaporator. The product was dissolved in hexane and purified with dialysis against hexane. Yield was 0.17g. FT-IR (ATR, Fig. S13) υ cm -1 : 3430, 2995, 2912, 2030, 1644, 1438, 1404, 1309, 1023, 948.7, 693.6, 668.4. S17  water, in the case of CA(AGE 6 -C 6 H 12 -UPy) 4 , are roughly spherical and therefore the presented data for hydrodynamic range from Zetasizer Nano ZS is valid. SAXS of CA(AGE 6 -C 6 H 12 -UPy) 4 , as dissolved in DMSO, shows a broad correlation peak at the scattering vector magnitude q ~ 0.035-0.045 Å -1 corresponding to structural features of 14-18 nm, and is interpreted as the form factor of micelles of CA(AGE 6 -C 6 H 12 -UPy) 4 . For S19 samples with increasing amount of water the form factor gradually disappears, which agrees that the structurally less-defined larger aggregates start forming.

Particle size analysis
Particles for analysis were picked by hand due to poor contrast in cryo-TEM measurements.
The diameter of the particles were measured with Gatan Microraph software using contrast difference tool. Particles in same focus plane were used to prevent error due to over or under focus.  In DMSO, the particle sizes appear larger using SEM than using TEM. The reason for the different particle size might be in sample preparation, as the SEM samples were prepared by dropcasting 10 μL of solution on SEM stub followed by evaporation to dryness. During the evaporation the concentration of CA(AGE 6 -C 6 H 12 -UPy) 4 gradually increases, potentially resulting changes in the self-assembly. Also, upon drying the DMSO-water mixtures, water evaporates first, resulting in concentrated DMSO. Therefore SEM studies are to be considered only qualitatively.