Supramolecular Chirality: Vesicle-to-Chiral Helix Transition of the

Subscriber access provided by UNIV LAVAL

Article

Supramolecular Chirality: Vesicle-to-Chiral Helix Transition of the Micelles Consisting of a Sugar-Bearing Calix[4]arene Surfactant Shunsuke Sakamoto, Shota Fujii, Kenta Yoshida, and Kazuo Sakurai Langmuir, Just Accepted Manuscript • DOI: 10.1021/acs.langmuir.6b01671 • Publication Date (Web): 07 Jul 2016 Downloaded from http://pubs.acs.org on July 9, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Langmuir is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

Supramolecular Chirality: Vesicle-to-Chiral Helix Transition of the Micelles Consisting of a Sugar-Bearing Calix[4]arene Surfactant

Shunsuke Sakamoto, Shota Fujii, Kenta Yoshida, and Kazuo Sakurai* Department of Chemistry and Biochemistry, University of Kitakyushu, Fukuoka 808-0132, Japan

KEYWORDS: calix[4]arene, surfactant, circular dichroism, induced circular dichroism * Corresponding author; E-mail: [email protected] (KS)

ACS Paragon Plus Environment

Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Table of Contents


Page 2 of 21

Page 3 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

Abstract Supramolecular self-assembly and the resulting chiral transfer from the molecular level to nanoscale is a major topic in modern supramolecular chemistry. We synthesized a galactose-bearing calix[4]arene surfactant (chiral) and mixed it with a primary amine-bearing analogue (achiral). The mixture showed strong induced circular dichroism (ICD) at an almost 3:2 molar ratio of the two surfactants and exothermic heat was observed upon mixing. The magnitude of ∆ was comparable with that of van der Waals interactions. This phenomenon indicated that the ICD can be ascribed to formation of a new supramolecular assembly in which the stoichiometric interaction between the two molecules leads to complexation and the resultant complex has a chiral morphology. Transmission electron microscopy and small-angle X-ray scattering showed that the galactose-bearing surfactant forms vesicles, and the mixing induces transition from the vesicles to thread-like cylinders with a diameter of ~ 3.0 nm. We presume that these cylinder are twisted due to chiral transfer from the chiral galactose moiety and show ICD.


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 4 of 21

Introduction Supramolecular self-assembly that produces sophisticated structures and useful functions has attracted significant attention in terms of both fundamental interests and practical applications. Driving forces behind such supramolecular self-assembly consist of multiple inter- and intramolecular interactions and their combination, and the balance essentially determines the final structure. Even though each interaction is subtle and insignificant, their combination eventually induces a dramatic change, and its influence can spread over several orders of scale. One of such examples is chiral amplification, i.e., the phenomenon where local chirality of a small number of chiral molecules determines chirality of the entire assembly, otherwise it is achiral.1, 2, 3, 4 For example, when a small amount of a chiral disc-shaped molecule is added to the solution containing a large amount of its achiral analogue that forms cylindrical structures with π − π stacking, the circular dichroism (CD) show a nonlinear response in terms of the concentration of the added chiral molecules.2 The induced CD (ICD) in this system is ascribed to the induced helical assembly of the cylindrical architecture composed of the disc-shaped molecules. In most cases, the cylinder is twisted in one direction by the addition of the chiral molecule. This is an example of supramolecular chirality: the supramolecular assembly itself has chirality based on the chiral spatial arrangement of constituent molecules. An interesting feature of supramolecular chirality is that because of chirality of the entire assembly, achiral chemical bonds enter a chiral environment and thus the achiral chemical bonds can start to show ICD. Normally, this change is manifested in CD intensity because all molecules and bonds including achiral ones are in a chiral environment. Such a phenomenon is called “chirality transfer” and has been observed in many chiral supramolecules as well as proteins and other biological molecules.1 Chirality transfer can be achieved through hydrogen bonding, π − π stacking, and by means of solvent environments.1 Calixarenes are an attractive building block for well-controlled self-assembly.

5, 6, 7

Shinkai et al8 showed

that a cone-shaped p-sulfonatocalixarene bearing appropriate alkyl groups forms spherical micelles in aqueous solutions. The nature of the ionic headgroup is a key parameter determining the structure of calixarene amphiphiles. Recently, Fujii et al extensively studied a series of calix[4]arene derivatives, having different hydrophilic headgroups such as primary anime,9 cysteine,10 and choline phosphate11 and found that in the case of a suitable combination of the headgroup and the alkyl chain length as well as solvent conditions, the molecules assembled into a completely uniform spherical micelle, i.e., monodisperse in the aggregation number and structurally precise micelle. This shape-persistent micelle is quite unique and novel and this nature may be related to the rather rigid building block and symmetry of calix[4]arene. An attempt to achieve supramolecular chirality in a calix[4]arene system was made by introducing cysteine as a chiral headgroup, denoted as CCaL3.10 CCaL3 forms a monodisperse dodecamer at pH 3. This is because the amino group in the cysteine is protonated and creates repulsive interactions among the four headgroups; thus, the interfacial area between the hydrophobic and hydrophilic domains of the surfactant becomes large enough to fulfill the conditions for spherical-micelle formation according to the packing parameter principle.12 When AuCl ions are added to the CCaL3 spherical-micelle solution, strong ICD appears, indicating emergence of supramolecular chirality. A combination of electron microscopy and small-angle X-ray scattering (SAXS) showed that helically coiled bilayer sheets are formed after addition of AuCl . This is another example of supramolecular chirality and chirality transfer.10


Page 5 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

In the series of these studies, Fujii et al found that the micellar structure of the primary-amine-bearing calix[4]arene surfactant (PACaL3) depends on pH. At low pH, spherical shape is formed due to a rather large electrostatic repulsion among the four-protonated amines. The micellar shape transforms to a cylinder at neutral pHs, which is attributed to the conformational change of the surfactant due to the deprotonation of the amines. We presume that this cylindrical micelle may exhibit a chiral amplification or transfer by the addition of a chiral molecule. This is a chief motive for the present study, in which we attached four galactoses to the upper rim of calix[4]arene as a chiral hydrophilic headgroup and examined supramolecular chirality induced by the galactose moieties.


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 6 of 21

Experimental Section Materials We synthesized two type of calix[4]arene surfactants as shown in Figure 1: PACaL3 with primary amine as a headgroup and DGalCaL3 and LGalCaL3 with D- and L-galactose as a headgroup, respectively. The synthesis procedure for PACaL3 was described previously.9 DGalCaL3 was newly synthesized for this work and the details are as follows (the synthetic scheme is presented in Figure 1). In a similar manner except for preparation of 1-azido-1-deoxy-β-L-galactopyranoside acetate, we prepared LGalCaL3, as described in Supplementary Information [SI] file (Figure S1).

Synthesis of DGalCaL3 Synthesis of propynes bearing calix[4]arene derivative with propyl tails (II). The compound I were synthesized according to the reported method.9, 13 Sodium hydrate (0.305 g, 12.7 mmol) was dispersed in dry THF (5mL). 2-propyne-1-ol (0.738 g, 12.7 mmol) and compound I (1.0 g, 1.27 mmol) dissolved in dry THF (10mL) were added in the mixture at room temperature. The reaction mixture was stirred for 36 h at room temperature. Water was then added to quench the reaction, and the reactant was extracted with EtOAc. The organic layer was washed three times with saturated NaCl solution and dried over MgSO4. The solution was evaporated to dryness, and the residue was purified by flash chromatography (2:1 hexane/dichloromethane), which afforded a brown solid (1.02 g, 1.18 mmol, 93%). 1H NMR (500 MHz, CDCl3): δ (ppm) = 6.65 (s, 8H), 4.42 (d, J = 15.0 Hz, 4H), 4.26 (s, 8H), 4.02 (m, 8H), 3.82 (t, J = 7.5 Hz, 8H), 3.14 (d, J = 15.0 Hz, 4H), 2.46 (s, 4H), 1.92 (m, 8H), 0.98 (t, J = 7.5 Hz, 12H).

Synthesis of protected galactoses bearing calix[4]arene derivative with propyl tails (III). 1-Azido-1-deoxy-β-D-galactopyranoside (2.20 g, 5.90 mmol), copper (II) sulfate pentahydrate (15.1 mg, 94.4

µmol), and sodium ascorbate (0.187 g, 0.944 mmol) were dissolved in dry DMF (10 mL), and then a solution of compound II (1.02 g, 1.18 mmol) in dry DMF (5 mL) was added to the mixture. The mixture was stirred at 90 °C for 48 h, and then cooled to room temperature. Water was then added, and the reactant was extracted with EtOAc. The organic layer was washed three times with saturated NaCl solution and dried over MgSO4. The solution was evaporated to dryness, and the residue was purified by flash chromatography (5:1 EtOAc/dichloromethane), which afforded a brown solid (0.890 g, 0.377 mmol, 37%). 1H NMR (500 MHz, CDCl3): δ (ppm) = 7.85 (s, 4H), 6.63 (s, 8H), 5.87 (d, J = 10.0 Hz, 4H), 5.57 (m, 8H), 5.27 (m, 4H), 4.54 (s, 8H), 4.42 (d, J = 10.0 Hz, 4H), 4.27–4.10 (m, 20H), 3.82 (t, J = 7.5 Hz, 8H), 3.14 (d, J = 15.0 Hz, 4H), 2.22–1.88 (m, 48H), 1.66 (m, 8H), 0.97 (t, J = 7.50 Hz, 12H).

Synthesis of galactoses bearing calix[4]arene surfactant with propyl tails (DGalCaL3). Compound III (0.890 g, 0.377 mmol) was dissolved in dry methanol (5mL), then 1M sodium methoxide (10mL) was added into the mixture. HCl aqueous solution (10mL) was added to quench the reaction after stirring for 3 hours at room temperature. The solvent was evaporated, and the residue was washed with CH2Cl2. DGaLCaL3 was obtained as a white solid (0.634 g, 0.369 mmol, 98 %). 1H NMR (500 MHz, methanol-d4): δ = 8.17 (s, 4H), 6.70 (s, 8H), 5.59 (d, J = 10.0 Hz, 4H), 4.44 (d, J = 10.0 Hz, 4H), 4.43 (s, 8H), 4.23–4.19 (m, 4H), 4.16 (s, 8H), 3.85 (t, J = 5.00 Hz, 8H), 3.99–3.71 (m, 20H), 3.18 (d, J = 15.0 Hz, 4H), 1.96 (m, 8H), 1.03 (t, J = 7.50 Hz, 12H).


Page 7 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

I

II

~2.0 nm

~1.6 nm

DGalCaL3

PACaL3

Figure 1. Chemical structures of DGalCaL3 and PACaL3, and the synthesis procedure for DGalCaL3.

Sample preparation and critical micellar concentration measurements DGalCaL3, LGalCaL3 and their mixtures with PACaL3 in various ratios were adjusted to 5.0 mM with ultrapure water, and then further diluted to 2.0 mM with 50 mM NaCl / Tris-HCl (pH = 8.0) aqueous solution. The resulting samples were left for at least one day to equilibrate at room temperature. Critical micellar concentration was determined by means of 8-anilinonaphthalene-1-sulfonate as a probe molecule9 for PACaL3 and D- and LGalCaL3; this concentration was 0.11, 0.13, and 0.10 mM, respectively (for example, DGalCaL3 is shown in Figure S2). Based on these values, all measurements were carried out at concentrations more than 1.0 mM.

SAXS DGalCaL3, LGalCaL3, and their mixtures with PACaL3 in the [GalCaL3]:[PACaL3] ratio 1:1 (1.0 mM/mL, 50 mM NaCl / Tris-HCl) were used for the measurements. The resulting samples were left to equilibrate for at least 1 day at room temperature. SAXS measurements were conducted on the BL-40B2 beamline at the SPring-8 facility, Hyogo Prefecture, Japan. We used a 30 × 30 cm imaging plate (Rigaku R-AXIS VII) detector placed at 0.70 and 1.5 m from the sample. The wavelength of the incident beam (λ) was adjusted to 0.10 or 0.071 nm. A combination of these two sample-to-detector distances made the q range of 0.1−4.0 nm−1 available, where q is the magnitude of the scattering vector, defined as = 4⁄sin with the scattering angle of 2θ. X-ray transmittance of the samples was determined by means of ion chambers located in front of and behind the sample. The scattering intensity was determined by subtracting the scattering intensity of the solvent solution from that of the sample. The detailed experimental procedures are described elsewhere.9


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 8 of 21

CD and transmission electron microscopy (TEM): CD was measured on a JASCO J-720 WI CD spectrometer. The 1.0 mM solutions were placed into a quartz cell with the thickness of 0.2 mm, and the spectrum was recorded at 25 ºC in the range 190 – 400 nm at the scanning speed of 50 nm/min and a response of 8 s. For TEM, a 10-μL droplet of the sample solution was placed on a copper grid coated with an elastic carbon film, and after 1 min the solution was wiped with a filter paper and then dried in vacuum. Next, the grid was loaded into a JEOL JEM-3013 electron microscope operated at 200 kV in the magnification range 50K – 250K.

Isothermal titration calorimetry (ITC) Heat flow during isothermal titration was measured with a VP-ITC MicroCal microcalorimeter (Northampton, MA). The titration data were processed in the software provided by the manufacturer. A 0.1 mM solution of GalCaL3 was maintained in the sample cell (1.4301 mL) at 25 °C in a 50 mM NaCl/Tris-HCl (pH = 8.0) solution, and the whole cell was stirred at 300 rpm. PACaL3 at 0.75 mM with the same salt and buffer was introduced into the sample cell by means of a syringe via 30 individual injections. The injections were 20 s in duration, and individual injections were programmed at intervals of 300 s. The injection intervals as well as the volume and number of injections were adjusted such that the binding is completed toward the end of the titration. After the peak area was integrated and the heat values of dilution were subtracted, a thermogram for the binding was obtained.


Page 9 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

Results and Discussion Dependence of CD on the mixing ratio DGalCaL3 completely dissolves in methanol, and its CD spectrum in methanol was recorded (Figure 2a). There was a negative CD band at = 220 nm. The only chiral chemical moiety in DGalCaL3 is galactose, but it has no UV absorption at > 180 nm. The triazole is adjacently connected to the galactose, and its UV absorbance is at = 200 to 220 nm; therefore, this CD band can be assigned to the triazole moiety under the chiral influence of D-galactose. When we dispersed DGalCaL3 in water at pH = 7 to 8, dynamic light scattering showed the presence of rather large particles, and their size ranged from 50 to 200 nm (data not shown), suggestive of the presence of small aggregate or self-assembly. When we recorded CD spectrum of DGalCaL3 in water, there was no difference in the spectra between methanol and water as shown in Figure 2a. This means that the DGalCaL3 aggregate or self-assembly had no chirality at all, in stark contrast to cysteine-bearing calix[4]arene lipid after neutralization with AuCl . Furthermore, the same CD bands in both solvents indicated that the molecular conformation around the galactose and triazole did not change after formation of the aggregate or self-assembly.


Langmuir

a: DGalCaL3 itself

40

water 20

0

10-3[θ] / degree cm3 dmol-1

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 10 of 21

MeOH -20

-40

c: Region B

b: Region A

20 10

Increasing PACaL3

0 -10

DGalCaL3 : PACaL3 6:4 6.5 : 3.5 7:3 7.5 : 2.5 8 :2 9:1 10 : 0

-20 -30 -40 200

250

300

350

Increasing PACaL3

400 200

DGalCaL3 : PACaL3 5.5 : 4.5 1 : 10 5:5 1 : 25 4:6 1 : 50 1:5 1 : 100 250

300

350

400

Wavelength / nm Figure 2. Dependence of circular dichroism (CD) on the mixing ratio DGalCaL3/ PACaL3 measured at 25 ºC in panels b and c, and its comparison between DGalCaL3 in water (micelles) and in methanol (stand-alone molecules) in panel a. As we changed the mixing molar proportion of the achiral surfactant PACaL3 in the mixture, we recorded CD (Figure 2b and 2c). With an increase in the PACaL3 proportion, the 220-nm negative band decreased, and a new band at 236 nm appeared. At 6:4 (hereinafter [DGalCaL3]:[PACaL3] in mole), the intensity of the 236-nm band reached the maximum, where the molar ellipticity was calculated from the total amount of the surfactants. With a further increase of PACaL3, the 236-nm band decreased, while the band shape remained almost the same. The band shape at 6:4 showed a positive Cotton effect, suggesting that only a single-electron transition mode may be involved in chirality. Furthermore, there was a weak broad positive


Page 11 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

band near 250–320 nm. Its origin can be ascribed to the calix[4]arene aromatic moieties, judging by the UV absorbance.10 The Cotton band and the broad aromatic band increased in the same manner, i.e., their intensity increased almost proportionally to an increase in the molar fraction of PACaL3. Figure 3 plots the molar

ellipticity

at

= 236 nm

(

!"#$

)

against

the

molar

fraction

defined

as

[PACaL3]/([PACaL3]+[DGalCaL3]). The CD change indicates that two different phenomena occur between 10:0 and 6:4 and between 6:4 and 0:10, denoted as regions A and B. The CD responses to the composition change in both regions were almost linear relative to the molar fraction, indicating that the observed phenomena are not chiral amplification. This is because one of the criteria of the chiral amplification is significant nonlinearity in the CD response.

Figure 3. The molar ellipticity at = 236 nm: !"#$ plotted against the PACaL3 molar fraction in the mixture. The CD changes were identified into two regions denoted as A and B, and the upper illustrations briefly summarize the results of isothermal titration calorimetry (ITC), circular dichroism (CD), and


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 12 of 21

morphological examination by transmission electron microscopy (TEM) and small-angle X-ray scattering (SAXS). The middle illustration shows chiral cylinders bundled together. Here, for sake of simplicity, 1:1 well-defined stoichiometry between the two components is drawn to show helix (which gives ICD), but the distribution of the two components may be aleatory in reality and we do not know how they are distributed. In region A, the data suggested that there are only two chiral species in this system; one is DGalCaL3 itself, which yields the 220 nm negative band, and the other yields both the positive Cotton effect and the broad band at = 250–350. This suggestion is consistent the following three facts: the CD spectra could be fitted a linear combination of these two species, the former specie decreased and the latter one increased proportionally to a molar fraction of PACaL3, and an isosbestic point was observed at = 205 nm. Judging by the chemical structure, it is clear that there are many freely rotating bonds between the galactose and the calix[4]arene; thus the CD due to the calix[4]arene is not oriented to the galactose but is related to supramolecular chirality. To explain CD in this region, we assumed that DGalCaL3 and PACaL3 interact to construct a new aggregate form or assembly. More likely, DGalCaL3 and PACaL3 formed a complex and the complexes self-assembled to give a new supramolecular structure, and its stoichiometry is seemingly rich in DGalCal3; [D-GalCaL3]:[PACaL3] = 3:2 or 1:1 according to the composition to give the maximum CD intensity. The resultant new structure has chirality to yield the new CD band. Because the calix[4]arene moiety produced the CD, this chirality is ascribed to supramolecular origin. Due to experimental error, it is difficult to conclude which is a more reliable stoichiometric value. In B region, CD intensity decreased only with an increase of the PACaL3 proportion without any change in the band shape. This finding indicates that further addition of PACaL3 did not change the chirality; it decreased the population of the chiral species. According to our another work,9 PACaL3 forms cylindrical micelles in neutral conditions. Therefore, additional PACaL3 molecules in region B may form cylinders by themselves without involvement of the preformed chiral aggregates, and the population of the CD origin is simply decreased. In the above discussion, we assumed that there are two kinds chiral species of DGalCaL3 (molecular chiral) and GalCaL3/PACaL3 complex at the same time, which may not be conclusive. However, we believe the assumption because the ICD demonstrates the maximum point at an almost 3:2 molar ration of the two surfactants. If the ICD is induced with the confinement of the calixarene moiety of GalCaL3 by the interaction with PACaL3, the ICD value should be constant after the stoichiometric point.

ITC for the complex formation. Figure 4 shows heat flow during the titration, where we added PACaL3 to DGalCaL3. We observed a simple exothermic reaction, and the isothermal curve could be fitted by a single site model14 with thermodynamic parameters

Δ = −7.2 Kcal mol -

and

.Δ/ = −590 cal mol -

and

the

stoichiometry

was

PACaL3]:[DGalCaL3] = 1.2:10. It is interesting that the simple mixing of PACaL3 and DGalCaL3, which seem to have no reacting functional groups, leads to an exothermic reaction. The magnitude of Δ is comparable with that of hydrogen bond formation or van der Waals interactions.15 We happened to have a galactose-bearing surfactant (denoted as Gal-C12) that does not have calix[4]arene as a building block but contains a triazole linker connecting galactose and the aromatic ring (Figure S3).16 When we carried out a


Page 13 of 21

similar ITC measurement, we did not observe any heat flow. According to this comparison, we can conclude that the negative Δ for the mixing may be related to new interactions, probably − interactions between the calix[4]arene moieties of both PACaL3 and DGalCaL3.

Time (min) -20 0

20 40 60 80 100 120 140 160

A µcal/sec

0.0

-0.2

-0.4

kcal/mole of injectant

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

B 0

-2

-4 0.0

0.5

1.0

1.5

2.0

2.5

Molar Ratio Figure 4. A titration calorimetry experiment on DGalCaL3 with PACaL3. In panel A, the titration thermogram is presented as heat released per unit of time after each injection. In panel B, the dependence of the released heat in each injection versus the molar ratio of DGalCaL3 to PACaL3. Filled squares represent experimental data, and the line corresponds to the best fitting be means of a single site model. The obtained thermodynamic parameters are listed in panel B. The entropy loss is relatively large in comparison with the magnitude of Δ. This is the reason for an exponential change in the heat flow. Thermodynamic events that lead to an entropy loss may include (1) a conformational change in both surfactants, (2) enhancement in the hydrophobic interactions, and (3) a morphological change in aggregates. When the main reason is related to (1), the one-site model is suitable to calculate the stoichiometric number14, 17. On the other hand, in the case of (3), especially in conjunction with


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 14 of 21

(2), Δ/ changes are not directly related to the reaction to produce Δ. As shown below, addition of PACaL3 led to morphological changes in the preceding assembly of DGalCaL3 itself, and this is a transition from vesicles to cylinders, which increases the hydrophobic unfavorable interactions. We presume that this structural transition is the main reason for the entropy loss, because cylindrical micelles have larger interface-area between hydrophobic and hydrophilic domains than vesicular micelles. The larger interface is more entropically unfavorable; thus the plate-to-cylinder transition decreases entropy. There is a rather large discrepancy in stoichiometry between CD and ITC. Although the ICD is maximized at [GalCaL3]:[PACaL3] = 3:2, the stoichiometry demonstrated by ITC study is much lower than that seen in ICD. This is presumably because that the stoichiometry of ITC relates not the optimal composition demonstrating maximum ICD but the morphological transition from plates to cylinders. After the formation of the cylindrical complexes, the helical structure gradually grows with the addition of PACaL3. Another possible explanation is that the one-site model to analyze ITC and thus give the stoichiometry may not be appropriate for the present system, because the origin of entropy loss may not be directly related to the binding between two components, but be relate to the morphological changes.

Morphological studies with TEM and SAXS Figure 5 shows TEM images for DGalCaL3 and mixtures 1:1 and 3:2 (stoichiometry in CD). For comparison, we also carried out TEM for LGalCaL3 and its mixture at the same ratios. DGalCal3 and LGalCaL3 showed no difference, and we always observed a circular image of approximately 20–100 nm in diameter, indicating that both GalCaL3 enantiomers yields a single-layer vesicle. As shown in panels B, C, and E (Figure 5), the mixtures at 1:1 and 3:2 produced images completely different from that of GalCaL3 itself, and there seemed to be no difference between the ratios 1:1 and 3:2. With both, many thread-like images without branching were observed and these threads seem to associate to form a flat tape-like bundle. The diameter of an individual thread was 1~2 nm. As mentioned above, the mixture showed a strong ICD, which is associated with structural chirality, such as a twisted ribbon or cylinder. We looked carefully at the images but could not find such a twisted image. The ribbon or type -like bundle showed some twisted structures as indicated with arrows, but they should be more ordered and regular to yield a strong CD signal.1 Furthermore, it is possibility that this tape-like bundle may form owing to aggregation during the drying process, i.e., because of increasing the concentration when we prepared the TEM specimens.


Page 15 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

Figure 5. Transmission electron microscopy (TEM) images of DGalCaL3 (A) and its mixtures DGalCaL3 & PACaL3 at [DGalCaL3]:[PACaL3] = 1:1 and 3:2 (B and C), in comparison with L-GalCaL3 (D) and its mixture at 1:1 (E). Figure 6 compares the SAXS profiles of DGalCaL3, and the equimolar mixtures of DGalCaL3 & PACaL3 and of LGalCaL3 & PACaL3. As expected, DGalCaL3 and LGalCaL3 showed exactly the same profiles, with the first intensity minimum at = 1.5 nm - and a continuous decrease in with an increase in at the low region ( 1 1.0 nm - ). In this region, confirmed to the relation of ∝ 3 with 4 5 −2.0 for 0.4 nm - 1 1 1.0 nm - and 4 5 −1.0 around = 0.2 nm - . The dependence of 4 5 −2.0 indicates the presence of a plate-like scattering object.18 Nonetheless, the size of the plate would be small enough to yield 4 5 −1.0 at a lower of 0.2 nm - . This = 0.2 nm - corresponds to about 30 nm in length in the real space, suggesting that the upper limit of the plate may be a few dozen nanometers. In this sense, the plate observation by SAXS may be consistent with the vesicle observation by TEM, because a zoomed up image of bilayer structures of vesicles can be regarded as plates, and the size of the vesicles was 20–100 nm. We have to admit that the discussion of the low-q data is somewhat ambiguous because of the presence of a broad distribution in vesicle size and, probably, secondary aggregation, but these factors do not influence the high-q region. In fact, a clear peaks and intensity minima were observed at > 1.0 nm - . According to the scattering theory, the positon of to give the first minimum (denoted as ∗) in a vesicle or plate is related to the thickness 7 by the equation ∗ 7 = 3.13 ∙∙. From this relation, we can evaluate that 7~2.0 nm, which is a quite reasonable size, assuming that two molecules are constituting the cross-section of the vesicle; this value is consistent with the TEM observation.


Langmuir

7

α = –1

10

6

10

5

10

α = –1 α = –2

4

10

PACaL3

3

I(q) / (a.u.)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 16 of 21

10

α = –1 2

10

α = –2

1

DGalCaL3

10

0

10

α = –3 α = –1

-1

10

DGalCaL3/PACaL3

-2

10

-3

10

LGalCaL3/PACaL3

-4

10

0.2

0.1

0.3 0.4 0.5 0.6 0.8

2

3

4

5

6 7

1

q / nm-1 Figure 6. Comparison of small-angle X-ray scattering (SAXS) profiles of DGalCaL3, mixtures DGalCaL3+PACaL3 (1:1) and LGalCaL3+PACaL3, and PACaL3. The scaling exponent 4 in ∝ 3 in the region of low is indicated by a line in each point. SAXS from the mixtures showed the same high- behavior as in GalCaL3, showing the same shape of the second maximum at = 2.3 nm - and the first minimum at = 1.5 nm - . This indicates that the cross-sectional size and structures are almost identical between GalCaL3 and the mixtures. The low-

behavior was complicated. It showed 4 = −2.0~ − 1.5 in ∝ 3 for all ranges less than 0.1 nm - as shown in the figure. Nevertheless, sometimes, we observed a different behavior: 4 = −1.0 near =


Page 17 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

0.4 nm - then 4 1 −4.0 at 1 0.2 nm - . According to the theory of SAXS, cylindrical objects yield 4 = −1.0, plate-like ones yield 4 = −2.0, and random aggregates yield 4 1 −3.0. These relations, together with SAXS and TEM observations, lead us to speculate the following morphological features. The mixture forms thread-like cylinders with the size ~2 nm (yields 4 = −1.0), and these cylinders tend to aggregate to form a plate (yields 4 = −2.0) composed of parallel cylinders observed by means of TEM. The aggregation power is not strong enough to hold the plate together; therefore, they are dynamically switching between a plate bundle and isolated thread.

Supramolecular structures and chiral objects to provide ICD There are two questions about supramolecular chirality. First, why did the vesicle made of DGalCaL3 or LGalCaL3 show the same CD band as s single molecule? Second, what structure dose provide the rather strong ICD in Figure 2? The answer to the first question is easy. Spherical vesicles and their mirror images are identical, meaning that spherical objects are achiral in structure; therefore, there is no ICD. As for the second question, we propose formation of a twisted cylinder that is created by the weak exothermic interaction between GalCaL3 and PACaL3, and the chirality of the cylinder is transferred from the chiral galactose moiety in the headgroup. TEM showed formation of a thread cylinder and its diameter of 1.5 nm indicates that the cylinder is indeed cylindrical micelles composed of DGalCaL3 and PACaL3. If DGalCaL3 is involved in formation of cylinder micelles, we can expect that the chirality may get transferred to the cylinder helix. To test this idea, we compared CD between DGalCaL3 and LGalCaL3 (Figure 7). The 1:1 LGalCaL3/PACaL3 mixture showed the CD band with the exact axial symmetry of the DGalCaL3/PACal3 mixture at the 1:1 ratio.


Langmuir

20

A

10

DGalCaL3 LGalCaL3

0

-10

10-3[θ] / degree cm3 dmol-1

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 18 of 21

-20

B

40

DGalCaL3 / PACaL3 LGalCaL3 / PACaL3

20

0

-20

-40 200

250

300

350

400

Wavelength / nm Figure 7. A comparison of circular dichroism (CD) between the enantiomers: DGalCaL3 and LGalCaL3 in water (A) and their mixture with PACaL3 (B).


Page 19 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

Figure 8 summarizes the observed morphologies and their relationship with CD. DGalCaL3 and LGalCaL3 form a vesicle (A and H). These two molecules are inherently chiral and they are enantiomeric with each other. Vesicles are not chiral; thus the vesicles made from DGalCaL3 or LGalCaL3 only have an intrinsic CD originated from the constituting molecules. When achiral PACaL3 is added, PACaL3 and DGalCaL3 (or LGalCaL3) interact to form a chiral cylinder (B1 in B and I1 in I), presumably a twisted cylinder, and they exhibit strong ICD. These cylinders are enantiomers, and the former and the later give the positive and negative Cotton effects, respectively (Figures 2 and 7). Further addition of PACaL3 seems not to change the chiral sense such as a helix pitch (B1 and G, where α ; β), rather simply increases the concentration of the cylinders (from B to C, then D). Close to the stoichiometric composition, the majority is the DGalCaL3/PACaL3 cylinders and they form a tape-like flat bundles to yield ∝ " in SAXS (D). In the region B, the added PACal3 molecules assemble by themselves to form an achiral cylinder and the concentration of the DGalCaL3/PACal3 cylinders is decreased (from D to F).

Figure 8. Observed morphologies and their relationship with CD. Here, for sake of simplicity, 1:1 well-defined stoichiometry is presented, but their distribution may be aleatory in reality.


Langmuir

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 20 of 21

Conclusions Strong ICD was observed in the mixture of chiral GalCaL3 and achiral PACaL3, while there was no ICD in the vesicle made of GalCaL3. The response was almost proportional to the molar fraction of added PACaL3, indicating that ICD is ascribed to formation of a stoichiometric complex rather than chiral amplification. ITC revealed an exothermic reaction occurred between GalCaL3 and PACaL3 with the magnitude of enthalpy in the same range with van der Waals interactions and a relatively large entropy loss. The former phenomenon may be related to favorable interactions between the calix[4]arene moieties of GalCaL3 and PACaL3, and the latter may be interpreted as increased hydrophobic interaction upon mixing. This may be related to increase in interfacial area upon structural transition. SAXS and TEM showed that vesicle formation in GalCaL3 itself and addition of PACaL3 induced morphological transition from vesicles to thread-like cylinders and the cylinders bundled together to form a tape-like shape. The molecular origin of the ICD in the mixture can be attributed to the twisted cylinder although TEM did not show well-pronounced chirality. Chiral transfer from the galactose moiety to the supramolecular assembly was confirmed in a comparison experiment on the enantiomers.

Acknowledgment We appreciate JST/CREST program for financial support and all SAXS measurements were carried out at SPring-8 40B2 [2014A1268, 2014A1268, 2013B1203, 2013A1564].


Page 21 of 21

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Langmuir

References 1.

Liu, M.; Zhang, L.; Wang, T., Supramolecular Chirality in Self-Assembled Systems. Chem. Rev.

2015, 115 (15), 7304-7397. 2.

Palmans, A. R.; Meijer, E. e. W., Amplification of chirality in dynamic supramolecular aggregates.

Angew. Chem. Int. Ed. 2007, 46 (47), 8948-8968. 3.

Green, M. M.; Park, J.-W.; Sato, T.; Teramoto, A.; Lifson, S.; Selinger, R. L. B.; Selinger, J. V., The

Macromolecular Route to Chiral Amplification. Angew. Chem. Int. Ed. 1999, 38 (21), 3138-3154. 4.

Mateos-Timoneda, M. A.; Crego-Calama, M.; Reinhoudt, D. N., Supramolecular chirality of

self-assembled systems in solution. Chem. Soc. Rev. 2004, 33 (6), 363-372. 5.

Kellermann, M.; Bauer, W.; Hirsch, A.; Schade, B.; Ludwig, K.; Böttcher, C., The First Account of a

Structurally Persistent Micelle. Angew. Chem Int. Ed. 2004, 43 (22), 2959-2962. 6.

Lee, M.; Lee, S.-J.; Jiang, L.-H., Stimuli-Responsive Supramolecular Nanocapsules from

Amphiphilic Calixarene Assembly. J. Am. Chem. Soc. 2004, 126 (40), 12724-12725. 7.

Ikeda, A.; Shinkai, S., Novel Cavity Design Using Calix[n]arene Skeletons: Toward Molecular

Recognition and Metal Binding. Chem. Rev. 1997, 97 (5), 1713-1734. 8.

Shinkai, S.; Mori, S.; Koreishi, H.; Tsubaki, T.; Manabe, O., Hexasulfonated calix[6]arene

derivatives: a new class of catalysts, surfactants, and host molecules. J. Am. Chem. Soc. 1986, 108 (9), 2409-2416. 9.

Fujii, S.; Sanada, Y.; Nishimura, T.; Akiba, I.; Sakurai, K.; Yagi, N.; Mylonas, E., A

stimulus-responsive

shape-persistent

micelle

bearing

a

calix[4]arene

building

block:

reversible

pH-dependent transition between spherical and cylindrical forms. Langmuir 2012, 28 (6), 3092-101. 10.

Fujii, S.; Sakurai, K.; Okobira, T.; Ohta, N.; Takahara, A., Synthesis and Characterization of a

Calix[4]arene Amphiphilie Bearing Cysteine and Uniform Au Nanoparticle Formation Templated by its Four Cysteine Moieties. Langmuir 2013, 29(45), 13666-13675. 11.

Fujii, S.; Nishina, K.; Yamada, S.; Mochizuki, S.; Ohta, N.; Takahara, A.; Sakurai, K., Micelles

consisting of choline phosphate-bearing Calix [4] arene lipids. Soft matter 2014, 10 (41), 8216-8223. 12.

Israelachvili, J. N., Intermolecular and surface forces. Academic press London: 1992; Vol. 450.

13.

Bew, S. P.; Brimage, R. A.; L'Hermit, N.; Sharma, S. V., Upper Rim Appended Hybrid Calixarenes

via Click Chemistry. Org. Lett. 2007, 9 (19), 3713-3716. 14.

Wiseman, T.; Williston, S.; Brandts, J. F.; Lin, L. N., Rapid measurement of binding constants and

heats of binding using a new titration calorimeter. Anal. Biochem. 1989, 179 (1), 131-7. 15.

Atkins, P.; De Paula, J., Elements of physical chemistry. Oxford University Press, USA: 2013.

16.

Sakashita, M.; Mochizuki, S.; Sakurai, K., Hepatocyte-targeting gene delivery using a lipoplex

composed of galactose-modified aromatic lipid synthesized with click chemistry. Bioorg. Med. Chem. 2014,

22 (19), 5212-9. 17.

Ladbury, J. E.; Chowdhry, B. Z., Sensing the heat: the application of isothermal titration

calorimetry to thermodynamic studies of biomolecular interactions. Chem. & bio. 1996, 3 (10), 791-801. 18.

Roe, R. J., Methods of X-ray and Neutron Scattering in Polymer Science. 2000.


Supramolecular Chirality: Vesicle-to-Chiral Helix Transition of the

Recommend Documents