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Modifying Silica with
Corannulene Derivatives
Preparing stationary phase using click reaction to separate fullerene mixtures
Thuy-Tran, Ho-Thi
Master thesis, 30 hp Examiner: Fredrik Almqvist
Passed: XX June 2020
I
Abstract
The immobilization of corannulene derivatives onto porous silica substrate was
performed by the copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC) “click”
reaction in order to prepare stationary phases for HPLC. The synthesis was
accomplished in four steps. In the first step, the silica surface was activated in order
to form hydroxyl groups for further modifications with (3-azido-propyl) trimethoxy-
silane. This latter material was formed through a substitution reaction between
sodium azide and (3-chloro-propyl)-trimethoxy-silane. In the final step, the azide
groups available on silica surface were allowed to react with terminal alkyne of
corannulene derivatives, forming triazole rings. Furthermore, a variety of solvents
were tested for C60-molecular imprinting simultaneously with the click reaction. C60
was used as a template molecule, which has π-π interactions with corannulene. After
removal of the template, the shape of the cavity resembles that of C60, which was
believed to increase the selectivity as well as efficiency of the material. The
characterization of the synthesized materials was carried out by several analytical
techniques such as FT-IR, 1H NMR,
13C solid state NMR spectroscopies and XPS.
Fullerenes and fullerane isomers were tested on chromatography columns filled with
two different corannulene-modified silica.
II
III
List of abbreviations
APPI Atmospheric photoionization
C4O2 Corannulene with oxygenated linker containing 4 carbons and 2
oxygens
C6O3 Corannulene with oxygenated linker containing 6 carbons and 3
oxygens
C6 Corannulene attached with a six carbons linker
CuAAC Copper (I)- Catalyzed Alkyne-Azide Cycloaddition
DMF Dimethylformamide
EDTA Ethylenediaminetetraacetic acid
FT-IR Fourier Transform Infrared Spectroscopy
FWHM Full Width at Half Maximum
HPLC High-Performance Liquid Chromatography
MS Mass Spectrometry
MI Molecular Imprinting
NI Non-Imprinting
NMR Nuclear Magnetic Resonance
XPS X-ray Photoelectron Spectroscopy
Author contribution
The corannulene derivatives were provided by a group, led by Prof. Jay S. Siegel
from School of Pharmaceutical Science & Technology at Tianjin University,
China[13]. Fullerenes and hydrogenated fullerenes were supported by Alexandr
Talyzin from the Department of Physics at Umeå University, Sweden. The synthetic
procedure is a continuation of the master thesis of Piotr Jablonski[10].
The 13
C solid-state NMR was operated by Tobias Sparrman, who is a first research
engineer at the Department of Chemistry at Umeå University, Sweden. XPS
measurement was managed by Andrey Shchukarev, who is a research fellow at
Department of Chemistry at Umeå University, Sweden.
All of the experiments and some characterizations have been done by myself,
involving FT-IR and 1H NMR.
IV
V
Table of contents
Abstract .......................................................................................................................... I Author contribution ..................................................................................................... III 1. Introduction................................................................................................................ 1
2. Popular scientific summary including social and ethical aspects .............................. 2 2.1 Popular scientific summary ................................................................................. 2 2.2 Social and ethical aspects .................................................................................... 3
3. Experimental .............................................................................................................. 3 3.1 FTIR- Diffuse reflectance spectroscopy ............................................................. 3
3.2 1H NMR and solid state
13C NMR ....................................................................... 3
3.3 X-ray Photoelectron Spectroscopy ...................................................................... 3 3.4 Column preparation ............................................................................................. 4
3.5 HPLC-MS ............................................................................................................ 4 3.6 Activating silica ................................................................................................... 4 3.7 Substitution of (3-chloropropyl)-(trimethoxy)-silane with azide groups[11] ...... 4 3.8 Azide functionalization on activated silica[11,12] .............................................. 4
3.9 Click reaction on azide-functionalized silica ....................................................... 5 3.10 Solvent tests for C60-molecular imprinting ...................................................... 5
4. Results and discussion .......................................................................................... 6 4.1
1H NMR of substituted product ........................................................................... 6
4.2 FT-IR of azide silica and click reaction products ................................................ 7 4.3 Solid state
13C NMR of modified and imprinted silica ....................................... 8
4.4 XPS of azide silica and „click‟ products .............................................................. 8
4.5 HPLC-MS ............................................................................................................ 9
..................................................................................................................................... 12 ..................................................................................................................................... 12 5. Conclusions and Outlook ......................................................................................... 13
Acknowledgement ....................................................................................................... 13 References.................................................................................................................... 14
Appendix...................................................................................................................... 15
VI
1
1. Introduction
The existence of fullerenes, which has been known as the third form of carbon was
for the first time published in 1970 by Eiji Osawa[15]. Until 1985, the group of
scientists leading by Harry Kroto noticed that there were two stable ions present for
C60 (m/z=720) and C70 (m/z=840) on the mass spectrum[15]. These results were
observed by vaporizing a graphite disc, using laser source[15]. Fullerenes are hollow,
ellipsoid spheres consisting only of carbon atoms. For instance, there are twenty
hexagons and twelve pentagons in the structure of C60, forming a closed spherical
cage and resembling like a soccer ball. The most abundant are C60 and C70,
therefore, their appearance has become an interesting area in scientific community[1].
In the normal state, fullerenes are black solids. Although many aromatic rings are
involved in fullerenes structure their chemical properties are more likely super-alkene
compounds rather than benzene[15]. Another interest is the cage-liked structure of
fullerenes that could harbor metal atoms like potassium to become a
superconductor[14]. As electron acceptor molecules, fullerenes are highly soluble in
п-donor solvents such as toluene[2]. In fact, the highest solubility of fullerenes was
reported in piperidine as 53.29 g/L[3]. Piperidine is a cyclic aliphatic amine with the
free electron pair on nitrogen atoms, which are able to form charge transfer
complexes with electron deficient fullerenes.
Under high temperature, fullerenes can be hydrogenated by reacting under extremely
high pressure of hydrogen gas, resulting in products known as fulleranes[4].
Numerous synthetic approaches have been carried out to produce fulleranes.
However, the simplest way to synthesize hydrogenated fullerenes is the reduction
reaction, using zinc and hydrochloric acid in toluene[5,6]. The formation of
hydrogenated fullerene is the addition of H2 at a C=C double bond to gain two C-H.
Therefore, there is only an even number of hydrogen atoms in the structure of
fulleranes such as C60H18 or C60H36. It was reported that the high level of
hydrogenation was unstable, thereby, fully saturation of fullerenes are hardly able to
observe[6]. The partial unconjugated structure of fulleranes have changed the
physical and chemical properties, compared to its ordinary[1], which opens many
opportunities in the modification of fullerenes.
Figure 1. (A)3D Buckyball-C60; (B) 3D-buckybowl-corannulene
Fullerenes and fulleranes have many possible applications in batteries, gas storage,
medicine, or optical sensors. Depending on the level of hydrogenation, buckyballs
could obtain a wide range of electrochemical capacities and become novel material in
battery technology. Moreover, if the process of hydrogenation and de-hydrogenation
of C60 is reversible, these materials could be ideal for hydrogen storage for fuel cells.
This application is environmental friendly and could greatly contribute to the
development of electric vehicles[1]. In addition, fullerenes own a unique optical
property that only allow low intensity light to pass through them and turns opaque
with intense light. Its optical limiting phenomenon has been widely used as a sensor
protection for eyes[1]. The limited solubility of fullerenes in aqueous phase could be
(A) (B)
2
solved by chemical modification with hydrophilic side chains that, in addition,
broadens the potential in pharmaceutical applications. In fact, C60 is able to work as
an anticancer drug delivery system to treat targeted tumors, resulting in reducing
doses for patients and thereby minimize side effects[15].
Corannulene (C20H10) is one of the simplest fragments of a fullerene cage that
contains one pentane ring fused with five benzenes, forming a bowl shape. This
compound has a high host potential for fullerenes recognition. The combination of its
concave structure and the conjugated system maximizes the π-π interaction between
buckybowls and fullerenes[7]. Therefore, a modified silica based-stationary phase
with corannulene derivatives turns out to be promising for separation of fullerene
derivatives by liquid chromatography.
The click chemistry is one of the possible ways to immobilize corannulene
derivatives onto silica support. Although three main types of click reactions are
efficient, the Copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC) has been
reported as the most effective reaction in modifying materials[8]. In particular, the
CuAAC reaction occurs between a terminal alkyne and azide groups, forming stable
triazole rings. It was reported that the rate was improved significantly when using
Cu(I) as a catalyst compared to a catalyze free reaction[8]. In fact, a variety of copper
sources could be used with the appropriate oxidant/reductant species to obtain Cu(I).
In order to improve the retention and selectivity of fullerenes in chromatographic
separation, molecular imprinting technique is another possibility. In this approach,
C60 was used as a molecular template and added during the click reaction. The
formation of a template cavity was envisioned to be driven by π-π interactions
between C60 and corannulene-modified silica[9].
Firstly, a certain amount of corannulene-modified silica was synthesized as a
stationary phase. Secondly, chromatographic columns were packed with the prepared
materials. Finally, the separation ability was tested by liquid chromatography using a
MS detector. Numerous analytical techniques have been employed to characterize the
material throughout the synthetic processes, for instance, FT-IR spectroscopy, 1H and
13C solid state NMR spectroscopy and XPS.
Aim of the diploma work
The aim of the study was to extend the previous investigation[10] with other
derivatives (corannulene attached with oxygenated side chain linkers) to modify silica
surface by CuAAC-click chemistry. Buckybowl is a highly potential candidate to
separate fullerenes and their isomers by HPLC thanks to its strong π-π interaction
with guest molecules[7]. In addition, C60-molecular imprinting has been investigated
to increase the efficiency of the columns. Therefore, it is necessary to find a solvent
that works sufficiently for the CuAAC reaction as well as dissolving the template
molecules. The experiments were carried out with different organic solvents such as
toluene, pyridine, and piperidine. In this experiment, phenylacetylene was used as a
test substance for the click reaction.
2. Popular scientific summary including social and ethical
aspects
2.1 Popular scientific summary
Porous silica has been known as the most important stationary phase in liquid
chromatography. The chemical modification of silica surface makes it versatile,
especially in purification, qualitative or quantitative analysis. In this research, silica
was modified with corannulene derivatives that tend to separate fullerenes and
3
fulleranes. Fullerenes are new allotropes of carbon, which have many possible
applications in our life. One of the most interesting applications of fullerenes is the
capable of trapping atoms inside the cage. In cancer treatment, the radioactive gas
radon could be trapped in the C60 structure combined with antibodies attaching
outside fullerene that preferentially kills unhealthy tissue[15]. By controlling the
pathway of how radon gas reacts with tumor, it possibly prevents the tracer from
damaging healthy tissue rather than the targeted tumor cells. Therefore, the
purification after synthesis of fullerenes/fulleranes is indispensable for further
investigations. HPLC using corannulene-modified silica could be considered for this
purpose. HPLC is the abbreviation for High Performance Liquid Chromatography.
The sample has to be dissolved in a selected solvent before introducing into the
column. The mobile phase is flushed through the column filled with a properly
selected porous or gel bed. The mixtures/analytes are separated as the result of
intermolecular interactions with the stationary phase.
2.2 Social and ethical aspects
This thesis was reported with all the experimental data that was observed during lab
work. Additionally, there were no animal tests or other biological samples.
3. Experimental
3.1 FTIR- Diffuse reflectance spectroscopy
Bruker IFS 66v/S spectrometer (Bruker Optik GmbH, Ettlingen, Germany) was used.
KBr was purchased from Merck, Germany and used as a reference/matrix. The silica
was ground manually by mortar and pestle with KBr. FT-IR diffuse reflectance
technique was used for this analysis under high vacuum condition (~7 mbar) and the
spectrum was recorded within 256 scans in the region from 400-4000 cm-1
. The
baseline correction and normalization of spectra were controlled by OPUS software
(version 5.5).
3.2 1H NMR and solid state
13C NMR
Bruker Avance III 400 MHz spectrometer with a BBO-H/F SmartprodeTM
was used
to obtain proton NMR spectra. Deuterated chloroform (CDCl3) was used as solvent.
The sample was recorded within 16 scans and controlled by Topspin 3.6.2.
Bruker Avance III 500 MHz spectrometer with a 4 mm HX CP MAS probe that was
employed for 13C solid state NMR spectroscopy. The rotors were filled with azide-
silica and modified materials and spun at 10 kHz. The program used 90o excitation
pulse for 1H, followed by a 1.5 ms 13C spin lock at 60 kHz during which 1H was
ramped (45-90 kHz). The Spinal-64 1H decoupling condition was applied at 83 kHz.
The spectrum was recorded with a 2s relaxation delay and 6000 scans. The CH2
signal at 38.48 ppm of adamantine was used as an external reference.
3.3 X-ray Photoelectron Spectroscopy
Kratos Axis Ultra DLD electron spectrometer, equipped with monochromatic Al Kα
source, power supplied at 150 W was performed on the silica samples. The full wide
spectra were measured with analyzer pass energy at 160.0 eV, and 20.0 eV for each
photoelectron lines. The spectra were recorded by Kratos software. The binding
energy (BE) scale of all spectra was referenced to the Si-2p line set at 103.3 eV.
During measurement, the charge neutralizing is done by a built in system of the XPS
spectrometer.
4
3.4 Column preparation
A Knauer (Berlin, Germany) air-driven pneumatic pump was used at high pressure
350 bars to pack the column. Two metal HPLC columns (40 mm x 2.1 mm) from
Isolation Technologies (Middleboro, MA, USA) were packed with C4O2 and C6O3-
modified silica. These were dispersed in toluene (VWR, AnalR Normapur) and
flushed into the columns under high pressure.
3.5 HPLC-MS
The evaluation on LC-MS was performed by using a Shimadzu (Kyoto, Japan) LQ
LC-20AD and Agilent HP 1100 series G1310A pumps, they were connected by a
mixer before passing the column. As a detector LCQ Fleet (Thermo Scientific, USA)
ion trap mass spectrometer equipped with atmospheric photoionization accessory
(APPI), and a Rheodyne (Rohnert Park, CA, USA) 7010 injector with 20 µL metal
loop were used. The instrument was controlled by LCQ tune plus program. Toluene
(VWR, AnalR Normapur) and the mixture of toluene:methanol (Fisher Chemical,
HPLC grade) were used as mobile phases.
The MS detector was tuned by using low concentration of C60 in toluene (ca. 5.2
µg/ml) to optimize the operation condition and the positive mode was used. The
C60/C70 mixture, higher fullerenes as well as different levels of hydrogenated
fullerenes had concentrations of approximately 0.01-1.0 mg/ml in toluene.
3.6 Activating silica
2.0 gram of silica (Kromasil 200 Å, 5 µm) was added in a beaker, filled with 25 ml of
1M HCl. The reaction was allowed to run for one hour at 100 oC. The silica was then
filtered on a glass filter. After that the silica was washed with deionized water and
once with methanol to remove excess acid, following by drying in a vacuum oven
overnight at 40 oC. The silanol groups were formed in this step by protonating oxygen
atoms on silica surface.
3.7 Substitution of (3-chloropropyl)-(trimethoxy)-silane with azide groups[11]
10 ml of (3-chloropropyl)-(trimethoxy)-silane (≥ 97%; Sigma-Aldrich) was added to
250 ml in a dried round bottom flask, containing 125 ml of dry DMF. The water was
removed from DMF by a glass contour solvent system (SG, Water, USA) by flushing
the flask with argon gas for three times before collecting the solvent. 5.20 g of
sodium azide (> 99%; Merck) was added to the mixture, followed by KI (20 mg) as a
catalyst. The mixture was stirred under nitrogen gas, at 110 oC, overnight. The
reaction was quenched by diluting with diethyl ether and transferred into a separatory
funnel. The organic layer was washed thrice with 25 ml of deionized water and once
with 25 ml of brine to remove all the unreacted reagents. The combined aqueous
phases were extracted once with diethyl ether. The combined organic phase was dried
with anhydrous MgSO4 and filtered before evaporating in a rotary evaporator. The
product (3-azidopropyl)-(trimethoxy)-silane with high viscosity was collected and
confirmed by 1H NMR.
3.8 Azide functionalization on activated silica[11,12]
0.50 g of activated silica was added to a pressure resistant vial and 0.59 ml of (3-
azidopropyl)-(trimethoxy)-silane was pipetted in the presence of dry DMF (1ml) as a
solvent. The reaction was heated to 110 oC in a GC oven simultaneously with rotation
by a rotor, for 24 hours. The modified silica was filtrated and washed twice with
deionized water and once with methanol. The aizde-silica was then dried in a vacuum
5
oven overnight at 40 oC. The product was characterized by FT-IR spectroscopy and
13C solid state NMR.
3.9 Click reaction on azide-functionalized silica
0.50 g of azide silica was added to a vial containing 1.0 ml of dry DMF, followed by
80.0 mg of oxygenated corannulene with terminal alkyne (C4O2). CuSO4 and a
reductant (sodium L-ascorbate) were then added to this mixture. The solution was
allowed to react at 60 oC, for 24 hours. The reaction was quenched by adding 25.0 ml
of 0.1 M EDTA, following by filtering on a glass filter and washing with deionized
water and methanol[12]. The final product was dried in the vacuum oven at 40oC
overnight and characterized by FT-IR spectroscopy, solid state 13C-NMR.
Figure 2. The mechanism of ‘click’ reaction between azide group and terminal alkyne of corannulene
attached with C4O2 linker
3.10 Solvent tests for C60-molecular imprinting
0.25 g of azide silica was added to a vial containing 1.0 ml of dry DMF. 0.29 ml of
phenyl acetylene was added, followed by CuSO4 and a reductant (sodium L-
ascorbate). The solution was allowed to react at 60 oC, for 24 hours. The reaction was
quenched by adding 25 ml of 0.1 M EDTA, following by filtering on a glass filter and
washing with deionized water and methanol[12]. The final product was dried in the
vacuum oven at 40 oC overnight and characterized by FT-IR spectroscopy. The same
procedure was applied for piperidine, pyridine, and toluene.
0.50 g of azide silica was added to a vial containing 1.0 ml of a chosen solvent,
followed by 80 mg of corannulene attached with a six carbon side chain. 80 mg of
C60 (0.5 equiv.) was dissolved in 2 ml of the chosen solvent before adding to the vial,
followed by CuSO4 and a reductant (sodium L-ascorbate). The calculated amount of
C60 was based on the idea that one fullerene was surrounded by two corannulenes to
form a cavity (Figure 3). The solution was allowed to react at 60 oC, for 24 hours.
The reaction was quenched by adding 25 ml of 0.1 M EDTA, following by filtering
on a glass filter and washing with deionized water and methanol[12] . The material
was packed in a metal column and the template was removed by flushing with toluene
at 0.3 ml/min, overnight. The imprinted material was dried in the vacuum oven at 40 oC overnight and characterized by FT-IR and
13C solid state NMR.
6
Figure 3. The suggestion structure of C6-modified silica and the imprinted process during click
reaction, using C60 as template molecules
4. Results and discussion
4.1 1H NMR of substituted product
(3-chloropropyl)(trimethoxy)silane was substituted with sodium azide (NaN3) to
obtain the terminal azide functional groups that allow click reaction in the following
steps. In order to examine whether the reaction occurred, 1H NMR was performed on
the starting material and the substituted product.
Figure 4. 1H spectra and structures of (3-chloropropyl)(trimethoxy)silane(A); and azide silane(B)
It can be seen from Figure 4a below that there were totally three groups of proton.
Two of them were at 0.78 ppm and 1.87 ppm corresponding to protons on the first
(A)
(B)
7
and the second carbon, respectively. The other at 3.55 ppm is overlapping signal of
trimethoxy groups and protons on the third carbon, where the substitution occurred.
After the reaction (Figure 4b), the proton at substituted position has shifted from 3.52
to 3.20 ppm. As the results, the reaction was successful and the substitution yield was
about 91.9%.
4.2 FT-IR of azide silica and click reaction products
After completing the substitution, (3-azidopropyl)-(trimethoxy)-silane was attached to
activated silica. The attachment occurred between azide silane reagents and surface
silanol groups. After that the azide groups reacted with ω-terminal alkyne
corannulene derivatives via click chemistry to immobilize corannulene on silica
support.
Figure 5 shows that the silica was successfully modified with (3-azidopropyl)-
(trimethoxy)-silane due to strong band of azide stretching (N=N=N) at 2150 cm-1
.
The immobilization of corannulene derivatives on silica support was successful as
shown by the significant decrease of the azide band and there is a small signal in the
region over 3000 cm-1
, arising from aromatic C-H stretching of corannulene.
However, there were some residual azide groups on silica surface or not all of them
were accessible for coupling.
The characterization of „click‟ products in different solvents such as DMF, piperidine,
pyridine and toluene were shown in Figure 6. In the previous studies[14], DMF was
used successfully as a solvent, thereby, it was also used as a reference, marked as red
color in Figure 6b in this experiment. After reaction, the azide signal completely
disappeared when using piperidine (Figure 6c), indicating that the modification of
silica was successful. The other evidences were the aromatic C-H stretching/C=C
bending bands at 3060 cm-1
, 1480 cm-1
, respectively. In addition, the spectrum was
similar to the one using DMF as solvent. Piperidine was chosen for imprinting
reaction (appendix 1).
In contrast, there was an incomplete reaction in pyridine, which can be seen from
Figure 6d. Additionally, there was a band at 1650 cm-1
, relating to C=N that possibly
came from pyridine residual in the product. The reaction was not able to be
performed in toluene. Since there was a strong signal of azide in the spectrum (Figure
6e) and no aromatic appearance.
(a)
(b)
(c)
(d)
(e)
Figure 6. FT-IR spectra of azide silica (a) and
click reaction products using DMF (b); piperidine
(c); pyridine (d); toluene (e) as solvent
Figure 5. FT-IR spectra of azide silica (a) and
C4O2-modified silica (b); C6O3-modified silica
(c)
(a)
(b)
(c)
A.U
A.U
8
4.3 Solid state 13
C NMR of modified and imprinted silica
Further confirmation was obtained from 13
C solid state NMR spectroscopy (Figure
7). After the reaction, aromatic carbon signals appeared (from 110-150 ppm) in both
materials, which means that silica was successfully modified with corannulene
derivatives. In addition, in Figure 7(b,c), there was a new signal at 70 ppm was the
carbon that bound directly to oxygen in the linker. The region from 10-55 ppm
belongs to aliphatic carbons, which can be seen easily in Figure 7a.
The 13C NMR solid-state spectrum of molecularly imprinted product was compared
to non-imprinted material, which is shown in Figure 8. There was a new peak at 136
ppm (Figure 8b), which is characteristic for C60. In the solid state, pure C60 has a
high speed of rotation and gives rise to a sharp peak at 143 ppm (296 K)[16]. The
main reason to the change in the chemical shift as well as a broadening of the peak is
probably due to the π-π interaction between corannulene and C60. Despite the hard
attempt to remove template molecules by flushing with toluene, the result proved that
C60 interacts strongly with corannulene and is very difficult to remove from the
material.
4.4 XPS of azide silica and ‘click’ products
X-ray photoelectron spectroscopy is an efficient way to examine the atomic
compositions and thus was used to analyze functionalized silica and „click‟ products.
Table 1 showed that the substituted reaction was complete with the presence of azide
but no chlorine.
The signal of sp2 carbon (about 288.0 eV) from triazole ring is present in both
modified material, indicating the attachment of corannulene derivatives were
complete for both. However, in the C4O2-material, there was a trace signal of copper
catalyst in spectrum (appendix 2). XPS is known as a surface characterization
technique, therefore, in order to examine the coverage of corannulene in the bulk of
porous silica elemental analysis is necessary.
Figure 7. 13
C solid-state NMR spectra of
azide silica (a); C4O2-modified (b); C6O3-
modified silica (c)
Figure 8. 13
C solid-state NMR spectra of
corannulene attached with six carbons linker
modified silica NI (a); C60-MI (b)
9
Table 1. XPS characterization of azide silica, presenting atoms in the material via binding
energy (BE), atom concentration (AC%), and assignments.
Line Azide silica
Assignment BE (eV) AC (%)
C 1s 284.7 7.4 C-C
286.4 6.8 C-O; C-N
O 1s 532.6 52.6 SiO2
533.7 2.8 H2O
N 1s
397.4 0.2 -
398.8 1.7 -N=N=N-
400.5 1.9 -N=N=N-
404.3 0.7 NO2?
Si 2p 103.3 25.9 SiO2
Table 2. XPS characterization of C4O2, C6O3-modified silica, presenting atoms in the
material via binding energy (BE), atom concentration (AC%), and assignments.
Line C4O2-modified silica
Assignment BE (eV) AC (%)
C 1s
284.3 33.4 C-C
285.9 11.4 C-N, C-O
288.1 0.8 C=C-N
290.8 1.6 π-π* excitation
O 1s 532.5 33.0 SiO2, C-O-C
N 1s 399.8 2.9 -N=N=N-
401.3 1.1 -N=N=N-
Si 2p 103.3 15.8 SiO2
Line C6O3-modified silica
Assignment BE (eV) AC (%)
C 1s
284.4 22.0 C-C
286.0 11.2 C-N, C-O
288.0 0.5 C=C-N
O 1s 532.5 42.6 SiO2, C-O-C
N 1s
398.7 0.3 -
399.9 2.1 -N=N=N-
401.0 1.6 -N=N=N-
404.1 0.3 NO2?
Si 2p 103.3 19.4 SiO2
4.5 HPLC-MS
The modified materials were characterized many analytical techniques before packing
into the metal columns (40 x 2.1 mm). The HPLC evaluation was performed with the
mixture of C60/C70 and fulleranes. The composition of mobile phase was adjusted to
see whether the separation could be improved, especially for the mixture of
hydrogenated fullerenes.
10
Table 3. The chromatographic parameters of both columns were calculated from the
extracted chromatogram (figure 9)
Column Retention time
(min)
FWHM
(min)
Efficiency
(effective
theoretical plates)
Resolution
C4O2 5.66 2.50 28.40
1.90 21.33 7.22 48.35
C6O3 2.15 0.63 64.50
2.54 6.92 1.58 106.27
The chromatographic parameters of both columns are shown in Table 3. The results
showed that using corannulene as an active site in chromatographic phase for
fullerenes purification was efficient. The fullerenes are well-separated on both
columns, which was proved by a resolution larger than 1.50 (baseline resolution
requirement[17]). For instance, the C60 eluted (5.83 min) earlier than C70 (21.50
min) on the C4O2 column. The chromatographic condition was 100% toluene, 1.5
ml/min flow rate.
Although the C4O2 has higher carbon content than the C6O3 (Table 2), the efficiency
of C6O3 column was greater than C4O2, resulting in sharper peaks and better
resolution. It could be possibly explained C4O2 column was filled incompletely or
unevenly during the packing process, resulting in a dead volume. As consequences,
the peaks were broadening, leading to the longer retention time.
Figure 9. The chromatogram (smoothed, normalized 1,0) of C60/C70 on C4O2 column (left);
C6O3 (right). The LC operation condition was 100% toluene at 1.5 ml/min flow rate
A.U
A.U A.U
A.U
11
In general, the order of elution was from high to low hydrogenation due to the
reduction in number of sp2 carbons that able to interact with the corannulene
stationary phase. From the results we can observe a pattern that (Figure 10, 11), the
higher level of hydrogenation, the lower retention on both columns. In fact, the
compound C60H36 was not able to interact efficiently with the stationary phase.
However, C60H18 was completely separated from C60H36 on both columns. In
addition, the solubilized high hydrogenated fullerenes are tending to degrade and
precipitate after one hour, which caused an increase in the back pressure.
The sample contains fullerenes hydrogenated at low level was injected (Figure 11)
and confirmed by mass spectrum (appendix 4), which showed the m/z=729, relating
to C60H9+. Furthermore, the mixture of toluene:methanol (80:20, v:v%) was also
tested (Figure 12).
Figure 10. The extracted chromatogram (smoothed, normalized 1,0) of C60H18/C60H36 on
C4O2 column (left); C6O3 (right). The LC operation condition was 100% toluene at 0.5
ml/min flow rate
Figure 11. The extracted chromatogram (smoothed, normalized 1,0) of low hydrogenation on
C4O2 column (left); C6O3 (right). The LC operation condition was 100% toluene at 1 ml/min
flow rate
C4O2 C6O3 A.U A.U
A.U A.U
A.U
A.U
A.U
A.U
A.U
A.U
12
Figure 12 showed that by adding 20%MeOH, C60 was able to be separated
completely from C60H8 on C4O2. However, the separation efficiency has not been
improved significantly by adding 10%MeOH (appendix 5). The strength of the eluent
decreased when increasing MeOH, which caused an increase in retention time. The
mobile phase was thus adjusted to improve the resolution. However, more of the
polar solvent, caused more precipitation in sample.
Figure 13 showed that C70H38 was not able to be separated efficiently on either
column. In other words, there was no retention of high level of hydrogenation of C70
on these columns. The mass spectrum of C70H38 is presented in appendix 6.
Figure 12. The extracted chromatogram (smoothed, normalized 1,0) of low hydrogenation
on C4O2 column 100% toluene (left); toluene:MeOH (80:20) (right) at 1 ml/min flow rate
100%Toluene 80%Toluene:20%MeOH
Figure 13. The extracted chromatogram (smoothed, normalized 1,0) of C70H38 on both
columns, 100% toluene at 1 ml/min flow rate
A.U
A.U
A.U A.U
A.U
A.U
A.U
13
5. Conclusions and Outlook
Porous silica derivatized with corannulene having oxygenated linkers were successful
synthesized by CuAAC click chemistry. The materials were characterized by FT-IR, 13
C solid state NMR spectroscopies and XPS. The C6O3 column has better separation
ability maybe due to a high degree of flexibility in space, which gives more efficient
interactions with fullerenes than C4O2. The number of the effective theoretical plates
of C6O3 was almost double to C4O2. In addition, the high level hydrogenation of
C60H18/C60H36 were sufficiently separated on both column, whereas, C70H38 has no
retention. For further investigation, the same length of side chain with other
electronegative groups such as amine could be investigated.
In theoretical terms, corannulene has a bowl-shaped structure that fits the convex
surface of carbon cages. Therefore, it could maximize the π-π interaction between
stationary phase and fullerenes, compared to the planar aromatic molecules. The
elemental analysis is required to determine the coverage of the lab scale synthesized
material. It is then possibly compared to the large scale commercial one.
Additionally, in order to test reproducibility of the packing more columns should be
prepared.
Piperidine was chosen as solvent for both the click reaction and molecular imprinting
experiment. The molecularly imprinted silica was unsuccessfully synthesized, as the
π-π interaction between fullerene and corannulene was too strong for C60 to be
washed out. Other ways should be investigated to remove the template such as
sonication, flushing with higher conjugated π system organic solvent to reduce the
strength of this bonding.
Acknowledgement
I would like to say thank you to all of my supervisors, Dan Johnels, Knut Irgum and
graduate student Piotr Jablonski, who are well-support during my thesis working
time. I do appreciate many good advices from Chau Huynh, who is a graduate student
in the same research group.
14
References
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[2] S. Talukdar, P. Pradhan, and A. Banerji, Electron donor-acceptor interactions
of C60 with n-and π-donors : A rational approach towards its solubility,
Fuller. Sci. Technol., vol. 5, no. 3, pp. 547–557, 1997.
[3] D. Mahdaoui, M. Abderrabba, C. Hirata, T. Wakahara, and K. Miyazawa, The
Influence of Water and Temperature on the Solubility of C60 in Pyridine
Solution, J. Solution Chem, vol. 45, no. 8, pp. 1158–1170, 2016.
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M. Shulga, Dag Noréus, Toyoto Sato, Andrzej Dzwilewski, Bertil Sundqvist,
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temperature: Hydrogenation and cage fragmentation, J. Phys. Chem. A, vol.
110, no. 27, pp. 8528–8534, 2006.
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Carbon Nanostructures Reaction Kinetics of C 60 Fullerene Ozonation,
February 2014, pp. 37–41, 2007.
[6] S. M. Luzan, Y. O. Tsybin, and A. V. Talyzin, Reaction of C60 with hydrogen
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11484–11492, 2011.
[7] E. M. Pérez and N. Martín, π−π Interactions in Carbon Nanostructures, R.
Soc. Chem, 2013.
[8] A. Marechal, R. El-Debs, V. Dugas, and C. Demesmay, “Is click chemistry
attractive for separation sciences?,” J. Sep. Sci., vol. 36, no. 13, pp. 2049–
2062, 2013, doi: 10.1002/jssc.201300231.
[9] L. Ye and K. Mosbach, “The technique of molecular imprinting - Principle,
state of the art, and future aspects,” J. Incl. Phenom., vol. 41, no. 1–4, pp. 107–
113, 2001.
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to create an HPLC column capable of, 2016.
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stationary phases for high-performance liquid chromatography,” Anal. Bioanal.
Chem, vol. 399, no. 10, pp. 3307–3322, 2011.
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exchange packings for chromatographic separation of proteins using ‘click
chemistry, J. Sep. Sci., vol. 35, no. 8, pp. 907–914, 2012.
[13] A. M. Butterfield, B. Gilomen, and J. S. Siegel, Kilogram-Scale Production of
Corannulene, 2012.
[14] T. Orre, Synthesis of modified silica using copper catalyzed click chemistry,,
2014.
[15] Royal Society of Chemistry, The Age of the Molecule, 1st edition, 1
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15
Appendix
Appendix 1 : The FT-IR of imprinting material with corannulene attached with six
carbons side chain.
A.U
16
Appendix 2: The XPS spectrum of C4O2-modified silica, showing the trace amount
of Cu 2p, which came from the use of CuSO4 as catalyst.
17
Appendix 3 : The mass spectrum of fullerene C60/C70
Appendix 4: The mass spectrum of low hydrogenated fullerene C60H8
C60C70-P2-2704-1st_01 #478 RT: 2.36 AV: 1 NL: 5.60E2
T: ITMS + c APCI corona Full ms [200.00-1000.00]
690 700 710 720 730 740 750 760 770 780 790
m/z
0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
75
80
85
90
95
100
Re
lative
Ab
un
da
nce
720.17
721.12
722.10750.40
751.49
723.12 755.42742.53
693.51767.64737.62 743.45717.95 763.12695.01 791.60724.17 777.46710.93 730.45 796.56700.77 780.73
C60C70-P2-2704-1st_01 #1381 RT: 7.10 AV: 1 NL: 1.63E2
T: ITMS + c APCI corona Full ms [200.00-1000.00]
770 780 790 800 810 820 830 840 850 860 870 880 890
m/z
0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
75
80
85
90
95
100
105
110
115
120
125
130
135
140
145
150
155
160
Re
lative
Ab
un
da
nce
840.06
841.09
842.15
769.52
853.22857.29843.15784.70
892.77776.57 793.32 807.45 817.39 873.26829.69799.05 882.43845.98839.24820.59 859.41
878.53
C60
C70
p1_C60C70lowH 80T20M 1st_01 #813 RT: 5.76 AV: 1 NL: 1.30E2
T: ITMS + c APCI corona Full ms [150.00-2000.00]
690 700 710 720 730 740 750 760 770 780 790 800 810 820
m/z
0
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
170
180
190
200
210
220
230
Re
lative
Ab
un
da
nce
729.22
730.21
731.26
728.16
732.17
727.11 751.61733.40699.61 715.62792.45753.63 799.43710.88 743.31691.12 817.15781.49776.59769.74 801.73 807.82
C60H9+
18
Appendix 5: The extracted chromatogram (smoothed, normalized 1,0) of C60H8 on
C4O2 column, toluene:MeOH (90:10) at 1 ml/min flow rate.
Appendix 6: The mass spectrum of hydrogenated fullerene C70H38
p1 7038 1ml 2_01 #74 RT: 0.31 AV: 1 NL: 4.16E3
T: ITMS + c APCI corona Full ms [100.00-2000.00]
800 820 840 860 880 900 920 940 960 980 1000 1020 1040 1060 1080
m/z
0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
75
80
85
90
95
100
Re
lative
Ab
un
da
nce
878.70
852.48
840.33
866.55825.50907.40887.61
986.85806.57 1031.09953.37 1069.63934.16
C70H39+
A.U
A.U
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