Polymeric light harvesting antenna

Ronald Merckx Promotor: Prof. Dr. Richard Hoogenboom Guide: Dr. Eng. Valentin-Victor Jerca

Academic year 2016-2017

A dissertation submitted to Ghent University in partial fulfilment of the

requirements for the degree of Master of Science in Chemistry

Polymeric light harvesting antenna

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Acknowledgements In the long run from middle school towards a job, a study at the university is one of the most

important and certainly the largest contribution towards both knowledge and self-

development. A master thesis should be the top of the bill of a 5 year long journey at the

university. In this road towards a Master degree in science, I personally learned a lot of things

about both myself and my skills in chemistry. The evolution I passed through would not have

been possible without certain persons I met along the way, by this short fragment, I would like

to express my gratitude towards these people.

My first resignation goes to Professor Dr. Richard Hoogenboom, whom made it possible that

I could pick and study a subject of my choice. Whenever I faced obstacles in my project both

practical and theoretical, he was able to provide the information that was needed to continue

the research in a so efficient manner as possible. I would also like to thank him for the

opportunity to be part of a most welcoming and friendly Supramolecular Chemistry group.

In this Supramolecular Chemistry group I would like to express my gratitude to someone in

particular, namely my supervisor dr. Valentin- Victor Jerca. Who was always very patient when

something went wrong due to my mistakes, but who was also very helpful when I did not

know how to continue. He helped me to develop and improve my practical skills. But also he

was not only my supervisor, he was more of very smart friend who was always there for a

joke, which created a very nice work environment.

Besides Victor I would like also to thank the other Supramolecular Chemistry group members

(Bart, Joachim, Maarten, Maji, Zhanyao, Xiaowen, Martin, Victor dlr, Ali, Annelore, Brynn,

Glenn and Mathias) and certainly the other thesis students (Willem, Ann, Maria, Wouter, Jana

and Tim) for contributing greatly to the pleasant atmosphere both outside and in the lab.

Additionaly I would like to thank Vincent and Jos from the professor Madder group for their

willingness to help me with various things in the lab.

Finally, I would like to thank my girlfriend and certain friends (Babs, Jasper, Chiel, Luca, Elke

and Philippe) for supporting me throughout the education and my thesis year. And last but

certainly not least I would like to thank my parents for the support. Without all these people

it would have not been possible to achieve this.

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Table of Contents Abbreviations .......................................................................................................................................... 7

1. Introduction .................................................................................................................................... 9

1.1 Photophysical processes in nature ....................................................................................... 10

1.2 Chlorophyll, carotene and other pigments .......................................................................... 11

1.2.1 Absorption spectra ....................................................................................................... 11

1.2.2 Light harvesting processes and the reaction centre .................................................... 13

1.2.3 Energy transfer from the antenna to the reaction centre ........................................... 14

1.3 Synthetic light harvesting ..................................................................................................... 15

1.3.1 Light harvesting systems .................................................................................................. 15

1.3.1.1 Conjugated polymers .................................................................................................... 15

1.3.1.2 Dendrimers.................................................................................................................... 16

1.3.1.3 Side chain polymers ...................................................................................................... 17

1.4 Photophysical processes in light-harvesting polymers ....................................................... 18

1.4.1 Energy transfer mechanisms on the molecular scale ...................................................... 18

1.4.2 Energy transfer in light-harvesting polymers .................................................................. 20

1.4.2.1 Intermolecular energy transfer .................................................................................... 20

1.4.2.2 Intramolecular energy transfer .................................................................................... 21

1.4.2.3 Energy migration in light-harvesting polymers ........................................................... 22

1.4.2.4 The antenna effect in light-harvesting polymers......................................................... 23

1.5 Goal of the project ................................................................................................................ 26

2. Results and discussion .................................................................................................................. 28

2.1 Photophysical characterization of the starting fluorophores ............................................. 28

2.1.1 UV-Vis characterization ................................................................................................ 28

2.1.2 Steady state fluorescence spectroscopy ...................................................................... 29

2.2 Synthesis of dye–labeled poly-(2-ethyl-2-oxazoline)s ......................................................... 34

2.2.1 Synthesis of single-labeled poly-(2-ethyl-2-oxazoline)s .............................................. 34

2.2.2 Synthesis of α,-ω-dye-labeled poly-(2-ethyl-2-oxazoline)s ......................................... 38

2.3 Light harvesting antenna polymers based on (1-pyrenylpropyl)-2-oxazoline (PyOx) ........ 43

2.3.1 Synthesis route ............................................................................................................. 44

2.3.2 Energy transfer in pPyOx-PEtOx-Cou copolymers ....................................................... 47

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2.3.3 Energy transfer in pPyOx-PEtOx-Cou copolymers in film ............................................ 48

2.4 Light harvesting antennas based on poly-(2-isopropenyl-2-oxazoline) modified polymers

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2.4.1 Synthesis and characterization of poly-(2-isopropenyl-2-oxazoline) modified polymers

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2.4.2 Synthesis of the model compounds ............................................................................. 51

2.4.3 Energy transfer in PIPRO modified copolymers........................................................... 53

2.4.4 Energy transfer in modified PIPRO copolymers in film ............................................... 54

3. Conclusions & Outlook ................................................................................................................. 56

4. Appendix A .................................................................................................................................... 58

4.1 Materials ............................................................................................................................... 58

4.2 Equipment ............................................................................................................................. 58

4.3 Compound synthesis............................................................................................................. 61

4.3.1 Organic compounds ...................................................................................................... 61

4.3.2 Polymer synthesis ......................................................................................................... 65

6. References......................................................................................................................................... 72

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Abbreviations ACN Acetonitrile

ATP Adenosine Triphosphate

BODIPY boron-dipyrromethene

CROP Cationic Ring Opening Polymerization

DCM Dichloromethane

DMF N, N-dimethylformamide

DMSO Dimethylsulfoxide

DP Degree of polymerization

EtOAc Ethyl acetate

FRET Förster Resonance Energy Transfer

HOMO Highest Occupied Molecular Orbital

LHC Light Harvesting Complex

LUMO Lowest Unoccupied Molecular Orbital

MeOH Methanol

MeOts Methyl p-toluenesulfonate

NADPH Nicotinamide adenine Sinucleotide Phosphate

NMR Nuclear magnetic resonance spectroscopy

PAH Polycyclic aromatic hydrocarbon

PEtOx Poly-(2-ethyl-2-oxazoline)

PIPRO Poly-(2-isopropenyl-2-oxazoline)

PMMA Poly-(methyl methacrylate)

PSU Photosynthetic Unit

PyOx 2-(3-(3,8-dihydropyren-1-yl) propyl)-4,5-dihydrooxazole

RPM Rotations per Minute

SEC (HFIP) Hexafluoroisopropanol Size Exclusion Chromatography

SOCl2 Thionyl chloride

TEA Triethylamine

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1. Introduction

The world energy consumption is ca. 4,7 ∗ 1020 J (450quadrillion Btu) and is expected to grow

2% each year for the next 25 years.1 As the world population grows, the demand for natural

resources, in the form of energy increases, while the supply of coal, oil and gas drastically

decreases. Earth’s resources alone are not enough to cover this consumption, so the

anthropological impact should decrease or other sources need to be explored. Concerns about

global warming have led to high interest in the field of renewable energy.2,3,4 In this area there

were numeral attempts to create energy in a green , more sustainable way (e.g. solar cells,

water and wind turbines, etc...)

In order to improve the existing solar cells, different paths have been explored. Recently,

several major advances have been made in the design of dyes and electrolytes for dye-

sensitized solar cells.5 Further efforts, including metal-ligand complexes and other light

harvesting systems were explored to improve solar cell performances.6,7 This master

dissertation will focus on light harvesting systems, more specific on light harvesting antenna’s.

Light harvesting and energy transfer processes of photosynthesis are the fastest and most

efficient known to mankind. In order for researchers to improve our light and energy

harvesting systems, gaining information and understanding about these processes is

crucial.8,9,10 By doing so, one could find a clean and sustainable source of energy, which might

benefit society in an enormous way.

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1.1 Photophysical processes in nature

Photosynthesis, synthesis aided by light is one of the most important processes in plants, algae

and photosynthetic bacteria. In a process driven by light energy, glucose molecules (or other

sugars) are constructed from water and carbon dioxide, and oxygen is released as a byproduct.

The glucose molecules serve as fuel for cells: their chemical energy can be harvested through

processes like cellular respiration and fermentation which generates a compound named

Adenosine Triphosphate (ATP). 11 This chemical energy in the form of a molecule can be used

to energize other processes taking place in the organism.

The total photosynthetic reaction can be summarized as follows:

6𝐶𝑂2 + 6𝐻2𝑂 → 𝐶6𝐻12𝑂6 + 602

The photosynthetic process is split up in two steps: light and dark reactions. In green plants

the light reactions occur in the thylakoid membrane and convert the incoming sunlight into

molecules, and are thereby referred to as light dependent. In the stroma within the

chloroplast the dark reactions take place, where they convert carbon dioxide into 6 membered

sugar rings (Fig. 1). The adjective dark is added, because these reactions does not require light

to take place. ATP and NADPH (which are products originating the light reactions) are used to

drive the dark reaction. 11,12

Figure 1. Light and dark reactions contributing to photosynthesis. (Source: Khan Academy)

The Calvin cycle is a biochemical pathway embedded in the dark reactions which uses carbon

dioxide, ATP and NADPH to form sugars. The first product formed in this cycle is a three carbon

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compound named glycerol-3-phosphate. Two of these structures can be combined by a series

of metabolic steps to form a glucose ring. Which is the end product of the photosynthesis.

Since the subject is focused on light harvesting, the dark reactions will not be considered in

this work. Only the first stages are of importance to this study. Absorption, excitation and

transfer of energy will be taken into account, with thereafter the transfer of this energy of the

pigment molecules to the reaction centre. 8,12

1.2 Chlorophyll, carotene and other pigments

Pigments are molecules that can absorb light in the visible area (400-700 nm) of the light

spectrum. The colour of this pigment depends on the light that is not absorbed, but reflected

by the pigment. In green plants and other photosynthetic organisms all light but green is

absorbed by the pigment chlorophyll. A wide variety of chlorophyll compounds and derivates

are known. One of the most common is chlorophyll a, which is present in all photosynthetic

organisms. Other molecules absorb at different wavelengths. Xanthophyll’s, carotenoids,

chlorophyll b, c and e are other examples of pigments that can be found in algae and

protistants. 13

These pigments function as antenna complexes. In order to achieve an efficient and

maximized use of the light, photosynthetic organisms have developed these light harvesting

antennas.8 The complex mixture of several pigments co-operate to collect the sunlight and

transfer it to the reaction centre. In specific conditions, this antenna can be modified to fit the

purpose (absorb the requested wavelength).

1.2.1 Absorption spectra

In order to provide the energy for the photosynthesis, the light harvesting antenna must

absorb the right amount of sunlight and then transfer it to the reaction centre. However, the

pigments does not absorb light equally at every wavelength (Fig. 2). The absorption spectrum

of a pigment shows one or more peaks at a specific wavelengths.

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Figure 2. Energy spectrum of sunlight. (Source: http://www.viridiansolar.co.uk/)

Chlorophyll a absorbs energy in the visible region of the spectrum, between 400-450 nm and

600-700 nm (from the violet-blue region and the yellow-orange-red region). Chlorophyll b

absorbs around 400-500 nm and 630-680 nm (Fig. 3). (Blue-green region and orange-red

region). 14,13

Figure 3. UV-VIS Absorption spectra and molecular structures of chlorophyll a & b in diethyl ether.13

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Pigments have more than one option after absorbing sunlight.12 If a pigment molecule the

energy might:

Be released as heat: by atomic vibrations, the energy can be released from the pigment

molecule.

Be emitted as a longer wavelength: also called fluorescence. When the electron

returns to a lower energy state again, a fluorescence photon can be emitted, which

has a longer wavelength than the incident one.

Energy transformation: Having an electron at higher energy level through light

absorption, this has become a potential electron donor. Which may react with an

electron acceptor, leading to a transformation of light energy into chemical energy

Resonance energy transfer: the excitation energy may be transferred to a

neighbouring molecule. An electron of the receptor is pushed to a higher excited state

as the photo-excited electron returns back to the ground state.

In an ideal world for photosynthesis, only resonance energy transfer would occur.

Fluorescence is one of the most abundant processes that occur.12

1.2.2 Light harvesting processes and the reaction centre

Photosynthetic processes are driven by the energy of the sunlight. The photosynthetic

pigments and proteins are a good example of supramolecular chemistry, which means they

are non-covalently bound. These pigment-protein complexes contain two types: the light

harvesting complex I and II, see figure 4 (further referred as LHCI and LHCII).

Figure 4. Illustrating LHI and LHII (Ref: Humphrey et al., 1996).12

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These complexes are part of an even greater system called photosystem I and II (further

referred PS I and II), which is on its turn a part of a photosynthetic unit (PSU). This unit also

contains the reaction centre (other types of pigment-protein complexes).11,12

1.2.3 Energy transfer from the antenna to the reaction centre

Energy transfer can happen in a variety of mechanisms within the photosynthetic unit. The

Förster mechanism works over distances of 20-50 Angstroms.15 This mechanism is depended

upon relative orientation of the pigments and the spectral overlap between the emission

band of the donor and the absorption band of the acceptor.

In the Förster mechanism, the excitation energy is transferred by resonance energy transfer

to a neighbouring molecule. During the process, an electron in the receptor molecule is

excited to higher energy levels due to the excitation energy transfer.15 This happens when a

photo-excited electron returns back to the ground state in the absorbing molecule. This is

called Förster Resonance Energy Transfer (FRET), where light falls into an array of

photosensitive molecules and is subsequently transferred to a specific photochemical reactive

site.5,16

Light harvesting and the energy transfer process is started with the absorption of photon, this

is typically done in a time range of femtoseconds (10−15 seconds). Followed by a transition of

an electron from the ground state tot an excited state. The excited state is not stable and thus

performs relaxation back to the first excited singlet state. How this excited states behaves

depends on the chemical environment. Because of the similar energy states of the antenna

molecules, the excited state energy has a high probability to be transferred to a neighbour

antenna molecule. 17

Transfer between the antenna molecules happens by interaction of the transition dipole

moments of the molecules. The natural occurring photosynthetic organisms are highly

efficient. Under optimal conditions more than 90% of the absorbed light is transferred to the

acceptor. The whole process takes less than a few hundred picoseconds, to transfer the energy

from the antenna to the reaction centre.12

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1.3 Synthetic light harvesting

Nature has been the cradle for many molecular and supramolecular structures. By mimicking

nature’s most efficient processes researchers try to produce synthetic compounds which

possesses the desired functionalities.18,19 For example in nature, the transport of O2 and CO2

in the body is due to a metal ligand bond between the iron and the protein Hemoglobin. In

the supramolecular world crown-ethers and cryptands have been found to associate with

compounds in such a strong way they can be used in drug delivery.20 Similarly, the light

harvesting is also mimicked from the pigment aiding photosynthesis, the most essential

reaction in nature.

1.3.1 Light harvesting systems The light harvesting antenna is an array of molecules which can transfer energy from the

incident light to a reaction centre for further process. The goal of antennas is to broaden the

range of light that can be absorbed by the system. Each antenna possesses a specific

wavelength range, by modifying the structure of the antenna, the range can be manipulated.

Since the discovery of the first synthetic polymer by Leo Bakeland, the polymer field has

known an exponential growth. Bakelite, the oldest recorded synthetic plastic was often used

due its hardness, high heat resistivity and excellent insulator properties. In the world as we

know it today polymers are an indispensable source of materials. The field of polymers itself

has provided tons of new materials and publications for the last decades.

To create artificial light harvesting antennas, polymeric materials are often used. These can

be divided in three major groups: π- conjugated backbone polymers (referred to as conjugated

polymers), dendrimers and polymers with a non-conjugated backbone, bearing side chains

with π-conjugated chromophores. (Referred to as side chain conjugated polymers). The main

focus in this work will be on the latest category including the synthesis and their photophysical

properties.21

1.3.1.1 Conjugated polymers

The electronic structure of π-conjugated polymers generally originates from the sp2pz wave

functions of the carbon atoms in the repeated units. In total, the π-electron of one monomer

is conjugated with the other adjacent repeated units. Due to this conjugation, the energy to

promote an electron from the Highest unoccupied molecular orbital (HOMO) to the lowest

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unoccupied molecular orbital (LUMO) is lowered.2,21 In contrast to the side chain polymers,

here the backbone provides the light harvesting properties. Materials based on conjugated

polymers have very high extinction coefficients, good procesability and tunability of the gap

between the HOMO and LUMO energies.

The major drawbacks of the conjugated polymers are the difficult synthesis, limited tunability

and the chain length. The length is difficult to control in synthesis, which gives a broad weight

distribution. So the reproducibility between batches is very low (e. g. PT, PF, PP, PPV, PA).

1.3.1.2 Dendrimers

Vögtle et al. reported the synthesis of the first dendrimer in 1978.22 Dendrimer popularity

since then has greatly increased. In 2005 more than 5,000 scientific papers and patents were

published about dendrimer chemistry. Due to their highly branched synthetic macromolecular

structure they were a novelty in the polymer field. The structure consists of three

components: a central core, interior branches and surface functional groups. Dendrimers are

symmetric and monodisperse molecules, which are synthesized by a polymeric reaction.

Distinction can be made between different generations of dendrimers in terms of shape and

size, which are different than linear polymers.23

From the outer shell to the core, there is a decrease of number of chromophores in the

dendrimeric structure which makes it a very attractive candidate for light harvesting

systems.10 Due to the rigid structure and fixed position of the chromophores in the branches,

orientation of the energy transfer can be obtained. High energy transfer efficiencies were

noted for dendritic structures used in light-collecting systems.8 Dendritic branches can be π-

conjugated as well, here they act as both light-absorbing chromophores and energy/electron

transfer roads to the reaction centre.10

However, there are a few major disadvantages to dendrimers. This step-by-step synthesis

makes it very labour intensive. While the size is increasing generation by generation, the

reactivity of the reacting sites becomes lower and lower due to sterical hindrance.24 This leads

to a low yield and a difficult synthesis, which drives the price of the compounds to higher

values. Besides the hard synthesis, there is also a problem by purificating them with

conventional methods such as SEC, etc...25,23

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1.3.1.3 Side chain polymers

Looking towards the structure, the side-chain conjugated polymers have a backbone which is

not conjugated (e.g. PMMA and polystyrene). The side chains however do contain conjugated

chromophores, which can act as a donor in the process of energy transfer. With this kind of

polymer systems Fox, Webber, Guillet and coworkers tried to mimic photosynthesis.26,27 who

also came up with the concept of “light-harvesting polymers”.

The length of conjugated systems highly determines the optoelectronic properties of main-

chain conjugated polymers. Where in side chain polymer systems, the properties of the

chromophores are mostly unaffected.28 Thus, side-chain polymers combine best of two

worlds. The typical polymer properties are maintained (e.g. film-casting ability, mechanical

stability and processing advantages, ability to create polymeric structures), while the well-

defined electronic, photonic and morphological properties are gained from the side chains.21

By the controlled radical polymerization (CRP), random and block copolymers can be formed

with a precisely controlled polymer length. One is able to make batches of side-chain

conjugated polymers with very high reproducibility. Due this high degree of control, different

chromophores can be installed in the side chains. Which can be tuned and optimized for

different properties. For example, the energy donor and acceptor can be attached to the

polymer backbone to induce energy transfer.

When building in these chromophores into the copolymer, the concentrations can be tuned

of the different chromophores in such a way that the copolymer fits the purpose (e.g. polymer-

based white OLED).29,30,9

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1.4 Photophysical processes in light-harvesting polymers

A wide variety of polymeric structures has been assembled with a built-in chromophore.

Chromophores attached to the polymer backbone can either be the same as in the side chain

or might differ. In (co-) polymers with different kinds of chromophores, one often acts as an

energy donor (possesses a higher excited state energy) while the other acts as an energy

acceptor (lower excited state energy).

1.4.1 Energy transfer mechanisms on the molecular scale

The basic principle of energy transfer can be illustrated by the following equation:

D* + A → D + A*

Energy transfer is mostly described by one of the two following processes. The first one is

Dexter energy transfer, which is known as the orbital overlap mechanism. The second is the

dipole-dipole interaction (Coulombic resonance), better known as Förster energy transfer (Fig.

5).21,31 This thesis will focus on the last mechanism, where a donor molecule (further referred

to as D) will transfer energy of sunlight towards an acceptor molecule (further referred to as

A).

These two mechanisms are better known as coulomb and exchange mechanisms. Although

they describe both energy transfer they differ in operative range. While the coulomb

mechanism is effective over 20-50 Angstroms, the exchange mechanism is only active over

not more than a few Angstroms.

Figure 5. Comparison of the two different energy transfer mechanisms. (Copyright: Organic composite nanomaterial’s: energy transfers and tunable luminescent behaviors, New J. Chem., 2011, 35, 973-978)

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The main difference between the two mechanisms is that for Förster transfer the interaction

happens through space (between D* and A) due to the interaction of dipolar electrical fields

of the D* with those of the A. While in the Dexter mechanism, the interaction is purely due to

the overlap of the orbitals between the D* and A. No orbital overlap or van der Waals are

necessary for the dipole-dipole interaction. It has been observed that the LUMO electron of

the Donor is exchanged with the HOMO electron in the acceptor. 21

The rate of energy transfer can be expressed as:

𝑘𝐸𝑛𝑇(𝐷𝑒𝑥𝑡𝑒𝑟) = 𝐾𝐽 exp (−2𝑅𝐷𝐴

𝑅°𝐷𝐴)

In which K is related to specific orbital interactions, J is the normalized spectral overlap

integral, and 𝑅°𝐷𝐴 is the separation of D* and A when they are in van der Waals contact. Due

to the exponential function in the equation one can understand that the transfer rate is highly

dependent on the distance. This transfer is only efficient on very small distances, typically 5-

10 Angstrom. The mechanism requires the donor and acceptor to be very close to each other,

if not in direct contact (Fig. 6). 21

For the Förster mechanism the rate of energy transfer is proportional to the inverse sixth

power of the separation between D* and A:

𝑘𝐸𝑛𝑇(𝐹ö𝑟𝑠𝑡𝑒𝑟)𝛼µ𝐷

2 µ𝐴

𝑅𝐷𝐴6

In comparison to the Dexter mechanism, here the distance between the D* and the A can be

significantly large. 𝑅𝐷𝐴 can take values up to more than 30 Angstrom.21

Figure 6. Different types of energy transfer. (A) Between a polymer and a small molecule. (B) Intramolecular transfer. (C) Energy migration. Copyright 1985 John Wiley & Sons:21

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1.4.2 Energy transfer in light-harvesting polymers

The Dexter and Förster mechanism above describe energy transfer, the last one is the transfer

of choice for copolymer light-harvesting antennas. Nevertheless, it is more complicated than

energy transfer between two small molecules. One can distinguish 3 major types of energy

transfer in a polymeric systems: intramolecular energy transfer, intermolecular between a

small molecule and the polymer and energy migration.

1.4.2.1 Intermolecular energy transfer

In literature almost no reports are found for commercial polymers (PMMA, PS, ...) with

intermolecular energy transfer properties. Kang et al. reported two types of cationic

polyfluorene copolymers (FHQ, FPQ) with the same π-conjugated structure but different

counterions (bromide (BR), tetraphenylborate (PB)) (Fig. 7). The compounds have been

studied as a fluorescence resonance energy transfer (FRET) donor to fluorescein-labeled DNA

(ssDNA-Fl).

The counterions accompanying the polymer chain for charge compensation are expected to

disturb complexation with DNA and modify the molecular fine-structure of the donor-

acceptor complex. This was reported to create competition between the desired FRET and

charge transfer quenching. The FRET signal was enhanced 2∼8.6 times by changing bromide

to tetraphenylborate as a counterion due to weaker complexation between the D and A units,

the larger D-A separation and the reduced photoinduced charge transfer quenching.32

Figure 7. Molecular structures of cationic polyfluorene copolymers with different counterions and their complexation schematics with ssDNA-FI.32

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1.4.2.2 Intramolecular energy transfer

Intramolecular energy transfer processes are systems where both the donor (D) and the

acceptor chromophore (A) are present on the same polymer chain. The most common type

of transfer is the Förster resonance energy transfer (FRET) process. By the means of FRET,

different properties of the polymer can be studied: end-to end distance, chain dynamics and

conformation. 21

More practical applications can be found in biochemistry, FRET can be used to determine

peptide/DNA lengths and interactions. Aside of this distances, dynamical properties can also

be obtained from the FRET process. Hong and coworkers used FRET as a technique to monitor

molecular events such as molecular cleavage and conformational transitions of a polymer

chain(Fig. 8).33,34 When the polymer scaffold is sensitive towards changes in pH different

properties are obtained dependent on the acidity of the solution in which it is present. If the

pH is lower than the pKa the molecule will return to the coil structure, which brings the end-

groups (pyrene and coumarin 343) in proximity of each other and FRET can be observed

between the donor at one end of the chain towards the acceptor at the other end. This

property disappears when pH is increased above the pKa and only fluorescence of the pyrene

will be observed.35 It is noteworthy that this is the only literature report that described the

FRET of this pyrene and coumarin 343 pair, suggesting that FRET may also occur when the two

dyes will be present on the same polymer chain.

Figure 8. pH sensitive polymeric sensor using Fluorescence Resonance Energy Transfer (FRET). 35

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1.4.2.3 Energy migration in light-harvesting polymers

When polymers have more than just one built-in chromophore on the chain, the energy

transfer process will be more complicated then described above. Absorptivity is raised by

increasing the number of chromophores in the polymer backbone, usually these

chromophores are the donors in the transfer process. The distance between these

chromophores will depend on the geometry and the flexibility of the polymer chain. 36–38

Adjacent chromophores which are relatively close, can transfer the excitons on to each other.

This is described by the “random walk phenomenon”, this walk can happen in three different

ways. First, hopping of the exciton from one chromophore group to the other that is next to

it on the polymer backbone. Secondly, moving the exciton along the conjugated polymer

backbone. Thirdly, due to folding of the polymer chain, there can be a temporary collision of

the chromophore groups, where they can transfer the excitons.21 Collini and coworkers

reported that for a poly-(phenylene vinylene) chain intrachain energy transfer is preferred

over interchain transfer when the polymer isn’t present in a coil conformation. When the

conformation shifts towards a coil conformation due to solvent or external factors, the

interchain energy transfer becomes more dominant (Fig. 9).39

Figure 9. Intramolecular energy exchange in polymers. Copyright 1985 John Wileys & Sons.39

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However it is possible for the exciton to stay on a chromophore group for a finite period of

time, before moving to the next one during the random walk process. If a single step is meant,

this is called “energy transfer”, for multiple steps the term “energy migration” is used. 21,37

There are three kinds of process that can contribute to the energy migration. The first one is

called the “nearest neighbour” principle. ‘n’ is the number of monomer blocks between the

two units carrying the chromophores, for the nearest neighbour principle n = 0. The second

type is called the “non-nearest neighbour”. Transfer here happens with n having a value of n

= [1, 3], because the chromophoric part is hindered to transfer to adjacent chromophores

(steric or structural). The third kind of energy migration is called the “loop transfer”, where n

> 3 (Fig. 10). By folding in a particular way, chromophores can interact with each other. This

folding is due to a strong solvent effect on the polymer, or when the polymer is simply long

enough to fold. 21

Figure 10. Three types of energy migration. a) Nearest neighbour transfer (n=0). b) Non-nearest neighbour transfer (n=1, 2, 3). c) Loop transfer (n > 3).

1.4.2.4 The antenna effect in light-harvesting polymers

Fox and coworkers published that phosphorescence occurred in the styrene-vinylnaphthalene

copolymer. The copolymer emission of naphthalene was much higher than the emission of

the two polymers separate in solution.40 Schneider and Springer confirmed this effect later

with the styrene-vinylnaphthalene copolymer.41 The higher emission is explained by energy

migration along the polymer chain. The same phenomenon was used in photosensitized

dechlorination of 2, 2’, 3, 3’, 6, 6’-hexachlorobinphenyl compounds. Here aromatic

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chromophores such as anthracene, phenanthrene and naphthalene are incorporated in the

polymer and can absorb light from the near-UV-visible region. The excitation energy was used

here to induce photochemical reactions. This was referred as photozymes. This system might

provide an ideal way to increase the efficiency to remove PCBs in the environment.42

This has been mimicked from chloroplast compounds, which can be found in plants. Here the

chlorophyll pigments act as the antenna and transfers the light energy. This effect is referred

as the “ antenna effect”.23,44,43

A wide variety of copolymers containing naphthalene and phenanthrene was studied by

Guillet and coworkers, concerning singlet energy transfer and migration. The energy was

trapped in anthracene traps. The singlet energy transfer was thought to be done exclusively

by energy migration, Guillet found that it was the combination of energy migration and Förster

transfer to the acceptor. Due to the short singlet lifetime (typical shorter than 100ns),

collisional energy transfer is a relatively minor factor, since the conformation relaxation time

is much longer than the lifetime of the singlet. So the energy transfer and migration are

dominated by the Förster energy dipole-dipole transfer mechanism.27

Figure 11. a and b) molecular structure of the copolymer c)model for energy quenching. Ref 15-16 copyright 2001-2002 American Chemical Society

25

Meyer et al. investigated energy transfer on copolymers of polystyrene (PS) loaded with Ru-

(II) and Os-(II) complexes (Fig. 11). Every Transition metal has 4-5 neighbours with an average

distances of 2-3 angstroms between them. The rate constant for nearest neighbour is in the

order of 109 𝑠−1, while a migration constant of 108𝑠−1 was observed. Experiments pointed

out that the intrapolymer energy transfer quenching was due to a combination of the random

walk principle, energy migration and energy transfer events. It was shown that a distance of

average 2 to 3 Angstroms was sufficient to promote through space energy migration and

energy transfer.44,45

Several more light-harvesting systems with a polymer scaffold have been reported in

literature, Dichtel et. al published about dendrimer scaffolds containing one a set of

naphthopyranone dyes located at the interior and another set of coumarin chromophores

located in the adjacent outer layer surrounding a porphyrin acceptor.46 FRET-capable

supramolecular polymers based on a BODIPY-bridged pillar[5]arene dimer with BODIPY guests

were reported bu Meng and coworkers.47

26

1.5 Goal of the project

The inspiration behind this project are the light-harvesting systems in nature which absorb

light and transfer the energy to photosynthetic reaction centers through a precisely spaced

array of chromophores that can transport energy over long distances. This highly efficient

energy transfer is done typically through a series of fluorescence resonance energy transfer

(FRET) events. Aside from the cellular context, light-harvesting systems could be utilized to

sensitize solar cells, drive photo catalysts or even be used as optical sensors. The design and

development of artificial light-harvesting systems is a contemporary academic and industrial

challenge having important economic and ecological implications.

Pyrene and coumarin 343 have been chosen as the FRET pair (i.e. donor and acceptor) due to

several important advantages that those two fluorophores have to offer. Pyrene is an

alternant polycyclic aromatic hydrocarbon consisting of four fused benzene rings with a large,

flat aromatic system, showing high thermal stability, extensive electron delocalization and

electron accepting nature. It is the most studied as a fluorophore, and has several advantages

such as long singlet lifetime, sensitive to the changes of the polarity of its microenvironment,

and strong tendency to form excimers which possesses high fluorescence quantum yield.

Coumarin 343 emits in the blue green spectral region, has a very high molar extinction

coefficient, displays large Stokes shifts and has large fluorescence quantum yield, therefore

fulfilling all the required conditions to be a very good acceptor in FRET experiments. Pyrene

(donor molecule) has an absorption in the range of 300-350 nm and emits between 370 and

450 nm in the range of absorption for coumarin 343 (acceptor molecule). Thus, the coumarin

343 absorption is expected to overlap with the emission of the pyrene which is one of the

main requirements for the energy transfer. Although pyrene absorbs in the UV region of the

spectrum and solar light is observed to have a maximum intensity between 400-700 nm, this

is considered not to be a problem due to the high molar extinction coefficient and quantum

yield.

As polymeric scaffold to covalently attach the dyes, the poly-(2-oxazoline)s were chosen due

to several advantages offered by this class of polymers. They can be synthesized in a controlled

and living manner by CROP. Different functionalities can be easily introduced by using

initiators or terminating agents that have the desired chemical groups. Finally, the monomers

can be modified with different chemical substituents in the 2-position to match our purposes.

27

Poly-(2-alkyl-2-oxazoline)s have also good film forming properties, are soluble in a large

number of solvents and have relatively high glass transition temperatures.

In the first part of the project we will attempt to synthesize single or dual labeled polymers

with pyrene and/or coumarin groups in order to highlight the type of energy transfer present

in these polymers inter or intramolecular. For single labeled poly-(2-ethyl-2-oxazoline)

polymers we will make use of the “termination method”. This implies initiation with a suitable

initiator (i.e. methyl tosylate) and end-capping the living chains with 1-pyrenebutyric acid or

coumarin 343. For the α,-ω-labeled polymers initiation will be done with a compound that

bears pyrene unit (i.e. 1-bromomethyl pyrene) and then same “termination method” will be

used to introduce coumarin groups. The next step will be the physico-chemical

characterization of this polymers. Then the photophysical properties will be investigated.

In the second part of the project we will explore the possibility to obtain copolymer bearing

donor units in the side chain and acceptor units at the end chain similar to an antenna light

harvesting system. This will be achieved by cationic ring opening copolymerization of EtOx

with a specially designed monomer bearing pyrene units (i.e. 2-(1-Pyrenylpropyl)-2-oxazoline)

and subsequent end capping of the living chains with coumarin 343. First the pyrene monomer

will be synthesized, then copolymers with different amount of pyrene units will be obtained.

Also in this case a physico-chemical characterization of the polymer is mandatory followed by

the investigation of energy transfer properties.

In the last part, we will make used of the highly reactivity of the oxazoline cycle towards

carboxylic acids and obtained light harvesting polymers by chemically modifying a poly(2-

isopropenyl-2-oxazoline) scaffold with 1-pyrenebutyric acid and/or coumarin 343. Using this

method, we will be able to synthesize polymers with different ratios of donor and acceptor.

Therefore, enabling us to precisely tune up the energy transfer efficiency based on chemical

composition. As stated before physico-chemical characterization and photophysical

investigations will follow after synthesis.

The ultimate goal of this project will be to find the optimum between polymeric architecture,

location of fluorophores onto the polymer chain (side-chain, end-chain) and ratio between

donor and acceptor groups in order to obtain the polymer with the best light harvesting

properties in solution and/or in solid state.

28

2. Results and discussion

2.1 Photophysical characterization of the starting fluorophores

Figure 12. Molecular structure of 1-pyrenebutyric acid (1) and Coumarin 343 (2).

During our literature survey, we could not find any energy transfer investigations regarding

this exact pair of fluorophores while their photophysical properties are promising for FRET.

Hossain et al reported that intramolecular FRET might take place between this pair if the dyes

were covalently attached to peptides. Therefore, the need of a thoroughly investigation of the

photophysical properties of this organic fluorophores in solution is mandatory. The chemical

structure of the fluorophores is depicted in Figure 12.

2.1.1 UV-Vis characterization

The study of molecules with conjugated π systems is typically done with the use of UV-Vis

spectroscopy. The absorption spectra of the two compounds is given in Figure 13. The 1-

pyrenebutyric acid (Py-COOH) spectrum in CHCl3 shows the characteristic vibration pattern of

the pyrene group at 314, 328 and 345 nm, respectively. For coumarin 343 (Cou343) the

maximum of absorption corresponding to π-π* transition is located at 449 nm.

Figure 13. Normalized absorption spectra of the two fluorophores in CHCl3.

29

2.1.1.1 Molar extinction coefficient

The molar extinction coefficient (𝜀), also referred as molar attenuation coefficient is a

parameter which gives information about how strongly a chemical attenuates light at a

specific wavelength. This parameter depends on chemical structure and is typically expressed

in L ∙ mol−1 ∙ cm−1. The 𝜀 was obtained by plotting the absorbance of several known solutions

against concentration. A higher 𝜀 value was found for coumarin 343 compared to 1-

pyrenebutyric acid (Fig. 14).

2.1.2 Steady state fluorescence spectroscopy

The emission spectrum of 1-pyrenebutyric acid in chloroform (Fig. 15) reveals the known

vibronic structure of the monomer band between 370 and 450 nm and no measurable excimer

emission above 480 nm. The fluorescence maximum of coumarin 343 is centered at 472 nm.

Figure 14. Linear correlation of absorbance with concentration for a) 1-Pyreneburtyric acid and b) Coumarin 343 in CHCl3.

Figure 15. a) Emission sprectra of Py-COOH and Coumarin 343. b) Spectral overlap between the absorption spectrum of coumarin 343 and the emission spectrum of 1-pyrenebutyric acid in CHCl3.

𝜺𝑷𝒚−𝑪𝑶𝑶𝑯 = 𝟒𝟎𝟖𝟓𝟎 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

𝑹𝟐 = 𝟎. 𝟗𝟗𝟖

𝜺𝑪𝒐𝒖 𝟑𝟒𝟑 = 𝟓𝟔𝟓𝟎𝟎 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

𝑹𝟐 = 𝟎. 𝟗𝟗𝟕

30

The excitation maxima are well separated (pyrene 𝜆𝑚𝑎𝑥 = 377 nm, coumarin 343 𝜆𝑚𝑎𝑥 = 478

nm). The superposition of the excitation spectrum of coumarin 343 and the emission spectrum

of the pyrene shows a good spectral overlap (Fig. 15). Also, both the dyes are highly

fluorescent, therefore fulfilling the requirement for FRET to occur.

With the maximum values of the emission and the absorption spectrum one can determine

the Stokes shift. This is defined as the difference (in wavelength or frequency units) between

the positions of the band maxima of the absorption and emission spectra of the same

electronic transition (Table 1). For homo-FRET, a fluorophore with a small stokes shift and thus

a great excitation-emission overlap is necessary, but for hetero-FRET we need a large stokes

shift and good separation between emission and excitation maxima. Higher Stokes shift was

observed for Py-COOH as compared to Cou343 (Table 1).

Table 1. Photophysical characterization of 1-Pyreneburytic acid and Coumarin 343.

Compound Solvent 𝛌𝐚𝐛𝐬−𝐦𝐚𝐱 (nm) 𝛌𝐞𝐦𝐢−𝐦𝐚𝐱 (nm) Stokes Shift (nm)

1-pyrenebutyric acid CHCl3 345 378 33

Coumarin 343 CHCl3 449 472 23

2.1.2.1 Energy transfer study in chloroform

Looking at the UV-Vis spectra in Fig. 13 one can observe that the maximum wavelengths of

the two absorption bands are well separated (pyrene 𝜆𝑚𝑎𝑥 = 345 nm, coumarin 343 𝜆𝑚𝑎𝑥 =

449 nm). Furthermore coumarin 343 has very low absorption between 300 and 375 nm and

the absorption band of pyrene is located in this spectral region. To study the energy transfer

between Py-COOH and Cou343 the emission spectra of Py-COOH and Cou343 mixture (1:1

molar ratio) was measured with the excitation wavelength fixed at 345 nm (corresponding to

Py-COOH absorption maximum). Figure 16 shows the fluorescence spectra of Py-COOH,

Cou343 and their mixture in CHCl3. From figure 16 a), we can observe that the fluorescence

intensity of pure Py-COOH is much higher than the one for Cou343. For the 1:1 mixture we

can notice a very small decrease in the intensity of the Py-COOH with respect to pure Py-COOH

emission. Also, the Cou343 emission increases with respect to the pure Cou343 (Fig 16 a). This

can be ascribed to the transfer of energy of the excited state from Py-COOH molecules to

Cou343 molecules via FRET.

31

The Förster theory shows that FRET efficiency (EFRET) varies as the sixth power of the distance

between two molecules (equation 1). To calculate the FRET efficiency, one has to determine

the spectral overlap integral 𝐽 and the interchromophoric distance 𝑟.48

𝐸𝐹𝑅𝐸𝑇 =

1

1 + (𝑟

𝑅0)

6 (1)

Where 𝑅0 is the distance between the two molecules at 50% FRET efficiency and 𝑟 being the

interchromophoric distance for the experiment. 𝑅0 can be calculated using equation 2.16

𝑅0 = 0.02108 (𝜅2 𝜙𝐷 𝑛−4 𝐽)16 (2)

Where 𝜙𝐷is the fluorescence quantum yield of the donor in the absence of the acceptor, 𝜅2

is the orientation factor of transition dipole moment between donor and acceptor; 𝜅2 can

take values between 0 and 4.49 In the present case we have considered 𝜅² =2

3 (when both

dyes are freely rotating and can be considered isotropically oriented during the excited state

lifetime), 𝑛 is the refractive index of the medium, and 𝐽 is the spectral overlap integral

calculated as:

𝐽 = ∫ 𝜀𝐴𝑐𝑐𝑒𝑝𝑡𝑜𝑟(𝜆)𝜆4𝐼𝐷(𝜆) 𝑑𝜆 (3)

Figure 156. a) Emission spectra of pure Py-COOH (pink curve), Cou343 (grey curve) and Py-COOH + Cou343 ( 1:1 molar ratio) mixture in CHCl3. b) Emission spectra of Py-COOH in the presence of various amounts of Cou343 in CHCl3. Excitation

wavelength was 345 nm in all cases, Py-COOH concentration was always constant 10 µM.

32

Where 𝜀𝐴𝑐𝑐𝑒𝑝𝑡𝑜𝑟(𝜆) is the acceptor molar extinction coefficient at the wavelength 𝜆, 𝜆 is the

wavelength and 𝐼𝐷(𝜆) is the normalized emission spectrum of the donor. By integrating this

over the entire spectrum one can calculate the 𝐽 value, the unit of 𝐽(𝜆) is 𝑀−1 ∙ 𝑐𝑚−1 ∙ 𝑛𝑚4.

The refractive index of CHCl3, and the quantum yield of the 1-pyrenebutyric acid were used

from literature.49

The efficiency of FRET can be determined by steady-state measurements and is expressed as

equation (4).

𝐸𝐹𝑅𝐸𝑇 = 1 −𝐹𝐷𝐴

𝐹𝐷 (4)

Where 𝐹𝐷𝐴 and 𝐹𝐷 are the donor fluorescence intensities in the presence and in the absence

of the acceptor, respectively.

The actual distance 𝑟 between donor and acceptor is given by equation 5:

𝑟 =𝑅0

(1

𝐸𝐹𝑅𝐸𝑇− 1)

6

(5)

The FRET efficiency was determined by the donor quenching method.

It is well known that FRET efficiency largely depends on the molecular proximity of donor-

acceptor in the mixture.50 Therefore it is interesting to check the FRET efficiency by varying

the acceptor concentration in the mixture. Accordingly we measured the fluorescence spectra

of pyrenebutyric acid mixed with varying concentration of coumarin 343(further referred as

Cou343). It was observed that the FRET efficiency increases with the increase in acceptor

concentration in the mixture (Fig. 16 b). This may be due to closer proximity of the Py-COOH

and Cou343 with increase in Cou343 concentration. The values of spectral overlap 𝐽(𝜆),

energy transfer efficiency (EFRET), Förster radius (𝑅0) and the donor-acceptor distance (𝑟) have

been calculated and are listed in Table 2.

From the obtained FRET efficiencies, it can be concluded that energy transfer is low (see Table

2). When increasing the concentration of acceptor groups (Fig. 16b) we noticed a higher donor

quenching (i.e. lower emission intensity) but the increased emission signal corresponding to

the acceptor is due to the auto-fluorescence of the coumarin. Therefore, we cannot state that

at high acceptor ratio we have a higher FRET efficiency. The decrease of donor intensity is not

33

an evidence of FRET since donor deactivation can also be caused by other quenching

mechanisms. The only sure evidence of energy transfer is when the decrease of donor

intensity is accompanied by an increase in acceptor intensity. FRET efficiency for this pair of

fluorophore is very low, namely 1.8% at a 1 to 1 molar ratio, even though we have a good

overlap between emission of the donor and absorption of the acceptor. The inefficient energy

transfer could be arising from the unfavorable relative orientation of transition dipoles of the

donor and acceptor, i.e., a small k2 value correlated with the lower fluorescence yield of the

Py-COOH taken from literature data (φfl = 0.077).49

Table 2. Energy transfer parameters for Py-COOH-Cou343 pair in CHCl3. The donor concentration was fixed at 10µM.

To conclude this pair of fluorophores is unsuitable for FRET experiments at least in the

investigated conditions. However, in the literature we could find reports regarding FRET

between these dyes when linked at peptide chain. That means that the fluorophores need to

be brought into close proximity in order to have efficient FRET. In the next sections we will

expound this strategy using different poly-(2-oxazoline)s scaffolds.34

Acceptor (Cou343)

concentration (µM)

𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)

𝑬𝑭𝑹𝑬𝑻

(%)

5 12.21 4.82 12.73 0.3

10 12.96 4.87 9.51 1.8

50 12.01 4.81 7.92 4.8

100 12.50 4.84 6.05 20.8

34

2.2 Synthesis of dye–labeled poly-(2-ethyl-2-oxazoline)s

Well-defined polymers with precise functionality, narrow polydispersity and controlled

molecular weight are becoming prerequisite in light harvesting applications. The polymer

scaffold will provide film-forming ability, mechanical stability and processing advantages. FRET

is sensitive to intramolecular and intermolecular interactions and has also been used for the

quantitative characterization of the interchain distance.51 Therefore we first started our study

by the synthesis of single-labeled polymers, namely the donor and acceptor molecules are

placed on different polymeric chains. Then we proceeded with the synthesis of dual-labeled

poly-(2-ethyl-2-oxazolines)s polymers where the donor is located at one end and the acceptor

at the other end side. The polymer choice of 2-oxazolines was substantiated by literature.52

The poly-(-2-oxazoline)s form a dipole layer which is assumed to bring the dyes closer to each

other and so to improve the probability for energy transfer.53

2.2.1 Synthesis of single-labeled poly-(2-ethyl-2-oxazoline)s

Figure 16. CROP of poly-(2-ethyl-2-oxazoline)s

The living nature of the CROP of 2-substituted-2-oxazolines allows facile preparation of well-

defined polymers (see Fig. 17). Ring-opening polymerization (ROP) is a form of chain-growth

polymerization, in which the terminal end of a polymer chain acts as a reactive center where

further cyclic monomers can react by opening its ring system and form a longer polymer chain.

Depending on the monomers, the properties of the resulting poly-(2-oxazoline)s can be easily

varied. Methyl and ethyl side groups result in water-soluble polymers, whereas aromatic

groups result in hydrophobic polymers. The living chains were end-capped with the

35

corresponding dye in the presence of TEA, therefore providing easy access to the desired

labeled polymer (see Fig. 18).

Figure 17. Chemical structure of PEtOx modified polymers with the fluorescent dyes.

The polymers were characterized by 1H NMR spectroscopy and size exclusion chromatography

(SEC). The molecular weight of the polymers was varied from 3000 Da to 7000 Da (DP 30, 50,

70) in order to maintain enough chain flexibility and not being too high to have sufficient FRET

efficiency between the chain ends. The ω-end dye labeling efficiency was over 90% as

determined by 1H NMR analysis (see Table 3). The SEC–analysis proved that the obtained

polymers have narrow polydispersities (Đ < 1.1) and the acceptor-labeled polymers have the

same molar mass as the donor-labeled polymers (Fig. 19 a and b). Also using the SEC analysis

with UV and RI detection we were able to prove that the polymers are labeled with the

corresponding dyes (Fig. 19 c and d).

Table 3. Structural characterization of single dye-labeled PEtOx.

Polymer code Yield (%) DPa f (%)b Mn (Da)c Đc

PEtOx-Py 97.0 30 94 5900 1.09

PEtOx-Py 80.2 50 95 10900 1.09

PEtOx-Py 80.1 70 94 12600 1.09

PEtOx-Cou 81.0 30 92 5900 1.08

PEtOx-Cou 94.4 50 91 10200 1.08

PEtOx-Cou 96.4 70 95 13100 1.10

a Theoretical degree of polymerization. b Functionality calculated from the aromatic signals of the dye and the methyl signal of the polymer side chain from the 1H NMR spectra. c Determined from DMA SEC using PMMA calibration.

36

2.2.1.1 Energy transfer in single dye labeled poly-(2-ehtyl-2-oxazoline)s

The intermolecular energy transfer in solution was investigated by steady state fluorescence

spectroscopy. First stock solutions were prepared from which more diluted samples were

prepared on which the FRET was investigated. The solutions always had a content of 10 µM

of pyrene fluorophore. The solutions were irradiated with 345 nm light which corresponds

with the maximum of the PEtOx-chains modified with the pyrene moiety. The emission spectra

of the DP 30 endcapped with Py-COOH or Cou343 were similar to the corresponding organic

fluorophores (Fig 20 a). When mixing the polymers to have a 1:1 molar ratio between the

fluophores an energy transfer could be detected (Fig 20 a, c and d). From the obtained FRET

efficiencies, it can be concluded that energy transfer is low, since the highest transfer is 8%

for PEtOx chains with DP=30 (Table 4). When increasing the concentration of acceptor groups

(Fig. 20 b) we noticed a higher donor quenching (i.e. lower emission intensity) but the

Figure 18. SEC traces for the PEtOx dye labeled polymer a) Cou-labeled DP 30, 50, 70 b) Py labeled DP 30, 50, 70 c) UV and RI signal for PEtOx-70-Cou d) UV and RI signal for PEtOx-70-Py.

37

increased emission signal corresponding to the acceptor is due to the auto-fluorescence of

the coumarin, similar to the results obtained for the organic fluorophores. Therefore, we

cannot state that FRET efficiency is increasing.

The values of spectral overlap integral J(λ), and Förster radius (R0) are similar to the ones

calculated for Py-COOH and Coum343 (see Table 4). Still, covalently linking the dyes onto

polymeric scaffolds (PEtOx DP30 mix 1:1) leads to a decrease in the donor-acceptor distance

(r) of 2.17 nm in comparison to organic fluorophores (same concentration and mixed in a 1:1

ratio). Therefore, the FRET efficiency increases from 1.8% (organic dyes) to 8.9% for the PEtOx

labeled mixed polymers, due to the smaller distance between the two dyes thanks to the

intermolecular interactions of the polymer chains. The r value is increasing with the DP of the

polymer because the chromophores are no longer in the proximity of each other.

Consequently, EFRET is decreasing for higher DP’s.

Figure 19. a) Emission spectra of pure PEtOx-Py (pink curve), PEtOx-Cou (grey curve) and PEtOx-Py + PEtOx-Cou (1:1 molar ratio) mixture in CHCl3 with a DP=30. b) Emission spectra of PEtOx-Py DP=30 in the presence of various amounts of PEtOx-Cou DP=30 in CHCl3. PEtOx-Py concentration was 10 µM. c) idem as a), but with DP= 50. d) idem as a), but with DP= 70. In

all cases excitation wavelength was 345 nm and the pyrene concentration was 10µM.

38

Table 4. Energy transfer parameters for mixing single dye labeled PEtOx mixed in 1:1 ratio in CHCl3. The donor concentration was fixed at 10µM.

2.2.2 Synthesis of α,-ω-dye-labeled poly-(2-ethyl-2-oxazoline)s

The intermolecular mixing of PEtOx was considered not to give an efficient energy transfer at

equimolar ratio between the fluorophores, thus an intramolecular approach was persued. For

this the pyrene and coumarin units were placed on the same PEtOx chain, in order to increase

the probability of the two dyes to be in close proximity.24 To produce such polymers another

initiator must be used. In this case we choose 1-(bromomethyl) pyrene to initiate the CROP of

2-ethyl-oxazoline and introduce the pyrene group in the α-position, followed by end capping

with coumarin 343. Following this strategy α,-ω- dye labeled polymers can be easily obtained

(Fig. 21). Since no literature could be found regarding CROP of 2-oxazolines with this initiator

we first performed a kinetic investigation.

Figure 20. Chemical structure of py-CH2-PEtOx-Cou polymers.

The polymerization kinetics for the living CROP of EtOx initiated with 1-(bromomethyl) pyrene

was performed at 140 °C under microwave irradiation using ACN as solvent. The first-order

kinetic plot of monomer consumption with respect to reaction time revealed a linear

relationship thus demonstrating a constant amount of propagating species indicative of the

absence of terminator and as well as relatively fast initiation (Fig. 22a). The polymerization

rate constant was calculated form the linear fit up to 80 % conversion, assuming that the

concentration of propagating species is equal to the initial initiator concentration. The value

Polymer codes DP 𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

PEtOx-Mix 30 14.85 4.98 7.34 8.9

PEtOx-Mix 50 14.22 4.95 7.58 7.2

PEtOx-Mix 70 16.77 5.09 8.17 5.5

39

obtained is similar for the polymerization of EtOx initiated with benzyl bromide.51 Subsequent

SEC analysis revealed a linear increase in number average molecular weight (Mn) with

conversion and low dispersities (Đ) which can be seen in Figure 22b), demonstrating that the

polymerization proceeded in a controlled manner.

Figure 21. a) First-order kinetic plots of monomer consumption. b) Molecular weight(Mn) and dispersity (Ð) against conversion plot for the polymerization of EtOx initiated with BrMePy.

In order to verify and to investigate the chain length influence on FRET efficiency we

synthesize three polymers with different DP values, namely 30, 50 and 70. The polymers were

obtained in high yields with low polydispersities and high degree of functionalization (Table

5). The different chain lengths were confirmed by SEC analysis.

The small high molecular weight shoulder in Figure 23 can be explained by the occurrence of

chain coupling reactions as commenly observed for the CROP of 2-oxazolines at 140°C in ACN.

Figure 22. a) SEC traces for the py-CH2-PEtOx-Cou compounds DP= 30 ,50 ,70. b) UV & RI detector response for Py-CH2-PEtOx-Cou.

40

The succesful labeling of PEtOx with both fluorophores is sustained by SEC-UV detection (see

Fig. 23 b). Polymers without coumarin end groups were also obtained in order to be used in

the photophysical study.

Table 5. Structural characterization of the dual-labeled PEtOx.

2.2.2.1 Energy transfer in α,- ω-dye labeled poly(-2-ethyl-2-oxazoline)s

Once the polymers were synthesized, fluorescent experiments were conducted to check if

intramolecular energy transfer between the chain ends is taking place. Since PEtOx is water

soluble, the compounds were also measured in an aqueous media. When the chromophores

are both present on the same polymer chain a clear FRET is taking place. The presence of the

signal corresponding to the coumarin emission and the decrease in of the pyrene signal

undoubtedly proves this fact in the spectrum (Fig. 24).

Py-CH2-PEtOx-Cou Yield (%) DPa f (%)b Mn (Da)c Đc

DP 30 95 28 92 6000 1.09

DP 50 92 49 91 9500 1.09

DP 70 89 68 90 13600 1.12

a Real DP calculated from the aromatic signals of the pyrene and the methyl signal of the polymer side chain from the 1H NMR. b Functionality, calculated from the aromatic signals of the coumarin dye and the methyl signal. c Determined from DMA SEC using PMMA calibration.

Figure 23. Emission spectra for Py-CH2-PEtOx-Cou polymers in a) CHCl3 and b) H2O. In all cases excitation wavelength was 345 nm and the pyrene concentration was 10µM.

41

In order to better understand the process, the influence of chain length and the effect of

solvent on the FRET process was investigated (Table 6). Analysis of emission spectra reveal

that the spectral overlapping integral J(λ) between the fluorescence spectrum of pyrene and

absorption spectra of coumarin unit is similar to the organic fluorophores or single labeled

polymers (see Table 2, 4 and 6), although the fluorophores are linked on the same polymeric

chain. Also, the intermolecular distance (r) decreases significantly from 9.51 to 4.87 nm in

chloroform. Therefore, the polymeric chain plays a vital role in reducing the intermolecular

distance providing a favorable condition for efficient energy transfer. Regarding the effect of

chain length, one can conclude that we have a decrease in FRET efficiency with increasing DP

due to the higher distance between the chromophores located at the chain ends. The optimal

DP in terms of FRET efficiency is DP 30 (Table 6).

Table 6. Energy transfer parameters for dual labeled PEtOx in CHCl3.

Sample Solvent 𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

Py-CH2-PEtOx-Cou 30 CHCl3 12.05 4.81 4.87 48.2



Py-CH2-PEtOx-Cou 30 H2O 8.51 6.05 4.02 92.1

Py-CH2-PEtOx-Cou 50 H2O 8.98 6.10 4.57 85.1

Py-CH2-PEtOx-Cou 70 H2O 10.16 6.23 5.87 58.9

Another important parameter in FRET experiments is the solvent nature and pyrene is

reported to be a very sensitive solvatochromic photoprobe in different environments.51 On

going from CHCl3 to water we noticed a 6-fold increase of the emission intensity of the PEtOx

DP30 labeled only with pyrene (Fig. 24). This behavior can be explained in terms of solvent-

solute interactions and has been investigated previously by Winnick et al. for the pyrene

molecule.54 Normally going from an apolar to a polar solvent the molar extinction coefficient

of the O-O band is dramatically increasing leading to higher fluorescence (increased intensity).

Also, the FRET efficiency is greatly improved in water reaching 92%, even if the J(λ) value

(degree of overlapping) decreases as compared to chloroform. In this case the decrease in

42

distance between the fluophores can probably be explained by the different polymer

conformations in water as compared to CHCl3.

2.2.3. Energy transfer for α,-ω-dye labeled poly-(2-ethyl-2-oxazoline)s in film

Besides bringing the two dyes in closer proximity to one another, one of the main advantages

of polymers is the procesability. By dissolving certain amounts of polymer in chloroform and

spin-coated upon quartz glass polymer films could be obtained very easily. In order to check

the presence of FRET in films, we mixed the single labeled polymers in different ratios and

then we spin-coated them always keeping constant the total molar concentration in solution.

Although in solution the FRET efficiency was low, efficient FRET was found in the film (see

Table 7). When irradiated the polymers with 345 nm light (λmax of pyrene) we noticed a

decrease of the pyrene intensity and the appearance of a signal corresponding to the emission

of coumarin. In the emission spectrum of pyrene, we observed also the presence of a peak at

460 nm attributed to excimer formation. The amount of pyrene present will be the same both

in solution and in film, but in the spin-coating mixture not everything will be available for FRET

due to excimer formation.

Table 7. Energy transfer parameters for single and dual labeled PEtOx mixed in film.

Sample 𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

PEtOx30-Mix 1 to 1 22.39 5.16 2.40 99.5

PEtOx50-Mix 1 to 1 19.25 5.03 2.55 99.0

PEtOx70-Mix 1 to 1 16.09 4.88 2.70 96.7

Py-CH2-PEtOx-Cou 30 22.42 5.16 2.40 99.0

Py-CH2-PEtOx-Cou 50 23.51 5.20 2.63 98.4

Py-CH2-PEtOx-Cou 70 23.30 5.18 2.86 97.3

When we use a 1:1 ratio between donor and acceptor units no emission of the pyrene

monomer can be detected, thus FRET efficiency is close to 100%. However, the fluorescence

intensity of coumarin is low. This means that pyrene fluorescence is quenching also through

other mechanisms. If we increase the pyrene concentration in the spin coating mixture a

higher signal corresponding to coumarin emission could be detected accompanied by a very

steep decrease of the pyrene fluorescence intensity peak (see Figure 25a, b, c). This effect is

43

similar to an antenna, so we need more pyrene units to harvest light and then transfer it to

the coumarin acceptor. This strategy will be pursued in the next chapters. If we increase the

chain length (i.e. DP 50 or 70) a lower FRET efficiency is obtained for the same reasons

discussed in the section regarding energy transfer in solution.

For the dual-labeled polymers we can observe that the FRET efficiency is very high in the film.

However, the overlap of the excimer peak with the coumarin emission could lead to

misinterpreted results in both cases and time-resolved fluoresce spectroscopy should be used

in order to quantitatively asses the FRET efficiency, which could not be performed in this thesis

due to time constraints.

2.3 Light harvesting antenna polymers based on (1-pyrenylpropyl)-2-oxazoline (PyOx)

Since intramolecular transfer is preferred in solution, a more elegant way was persued to

produce polymers with an efficient FRET. Therefore, driven by the obtained results, we set up

Figure 24. Emission spectra of PEtOx-labeled polymers in film a) DP30 b) DP50 c)DP70 d) dual-labeled PEtOx. All spectra were measured at 550V and with the excitation wavelength 345nm.

44

a strategy to synthesize poly-2-oxazoline polymers that incorporate pyrene units in the chain

and endcap them with a coumarin unit at one end. A statistical copolymer was prepared by

combining PyOx with 2-ethyl-2-oxazoline, and finally end-capping the polymer with coumarin

343 (Fig. 27). By an excess of donor molecules present upon the polymer chain an efficient

energy transfer was expected.

2.3.1 Synthesis route

Figure 25. Synthesis route to form pyrene-oxazoline.

Preparation of the PyOx monomer 13 started from the 1-pyrenebutyric acid. First the 1-

pyrenebutyric acid 1 was converted into an acid chloride (Figure 26).

Figure 26. Chemical structure of pPyOx-PEtOx-Cou.

Then the acid chloride was reacted with 2-chloroethylamine hydrochloride under argon

atmosphere without any intermediate separation steps, as the synthesis yield was close to

100% for the reaction with thionyl chloride. The formation of the acid chloride was confirmed

by 1H NMR spectroscopy (Fig. 28). The shift of the signal corresponding to CH2 protons next to

the chloride from 2.5 to 3.0 ppm. Due to the presence of high amounts of by-product the

amide had to be purified before it can be used in the next step. The purification was done by

flash chromatography on silica gel using a solvent mixture of hexane/ EtOAc (80/20). The

intermediate amide structure is confirmed by the presence of the multiplet signal at 3.75 ppm

corresponding to the ethylene (CH2-CH2) protons from the chloroethyl group (Fig. 28). In order

45

to obtain the monomer 13, structure 12 was ring closed in presence of KOH and 18-crown-8-

ether.The final product displayed the characteristic triplet signals of the oxazoline present at

3.8 and 4.2 ppm corresponding to the two methylene protons next to the nitrogen and the

oxygen, respectively. The pure PyOx was dried by azeotropic distillation with cyclohexane.

To study the FRET efficiency a series of statistical copolymers of EtOx and PyOx, respectively

were synthesized by varying the monomer ratio in the feed. The synthesis of the polymers

was performed in a similar manner to the PEtOx labeled polymers. Polymerization mixture

with 1.5, 5.5, 8.5 and 16.5 mol% of PyOx were prepared and polymerized to full conversion

and endcapped with Coumarin 343. Finally the PEtOx-pPyOx-Cou copolymer was obtained

(Fig. 29). The polymers were characterized by 1H NMR and SEC analysis.

Figure 27. 1H NMR of 1-pyrenebutyric acid 1 (red), 4-(1,8-dihydropyren-1-yl)butanoyl chloride (green), N-(2-

chloroethyl)-4-(1,8-dihydropyren-1-yl)butanamide 12 (blue) and the 2-(1-pyrenebutyric)-2-oxazoline 13 (purple). The spectra were recorded in CDCl3.

46

Table 8. Structural characterization of the pPyOx-PEtOx-Cou copolymers.

pPyOx - PEtOx-Cou %mol PyOx feed

%mol PyOx copolymera

Py/Coub Yield (%)

Mn (Da)c

Đc

10 16.5 19.8 10:1 92 9700 1.11

5 8.5 9.5 5:1 82 10600 1.11

3 5.5 7.2 3:1 78 8500 1.10

1 1.5 2.0 1:1 76 10800 1.10

a %mol PyOx in the copolymer determined by 1H NMR. b Ratio of pyrene to coumarin in the copolymer determined by 1H NMR. c Determined from DMA SEC using PMMA calibration.

All copolymerizations were performed under microwave irradiation at 140°C. The final

polymers had a narrow polydispersity regardless of the PyOx molar ratio (Fig 29 and Table 8).

Although all the polymers have approximately the same degree of polymerization, according

to the SEC traces (Fig. 29 a) they have different molecular weights. This can be understood by

the different amounts of PyOx units present. So the molecular weight of the polymer chains

increase because the PyOx has a higher molecular weight than EtOx. The presence of both

PyOx and the coumarin units on the same polymeric chains is confirmed by analyzing the signal

of UV detectors at maximum wavelength of PyOx and Coumarin respectively (Fig. 29 b).

Figure 28. a) SEC traces for pPyOx-PEtOx-Coumarin copolymers b) SEC RI and UV detector for pPyOX-PEtOx.

47

2.3.2 Energy transfer in pPyOx-PEtOx-Cou copolymers

The statistical copolymers were very soluble in CHCl3 and with lowers amounts (1.5, 5.5 %) of

pyrene units were also soluble in H2O. Both in chloroform and water a significant FRET was

observed. The FRET was accompanied by a decrease of pyrene emission and an increase in

coumarin emission. One can notice that besides the signal 370-420 nm, the pyrene also emits

at approximately 450 nm and this can ascribed to excimer formation (Fig. 30). These are

unstable excited molecules which are formed by the combination of two smaller molecules

and rapidly dissociates with emission of radiation. The contribution of this peak to coumarin

emission is small and it can be subtracted because we have synthesized also the PyOx

copolymers without the coumarin end groups.

Table 9. Energy transfer parameters for pPyOx-PEtOx-Cou copolymers in CHCl3. .


( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

PEtOx-pPyOx-Cou 10 CHCl3 22.56 5.34 6.05 32.2




PEtOx-pPyOx-Cou 3 H2O 14.58 6.62 4.85 86.7


The PyOx amount in the copolymer was calculated in such a way so as the final ratios between

Pyrene and Coumarin to be 1:1, 3:1, 5:1 and 10:1 (see table 9). From Figure 30 b and Table 9

we can notice that the FRET efficiency is over 30% in all cases and comparable to the one

Figure 29. Emission spectra of pPyOx-PEtOx-Cou copolymers in a) CHCl3 and b) H2O.

48

formed in the case of dual-labeled PEtOx polymers with DP 30. The main advantage of this

strategy is that we manage to obtain a higher degree of polymerization (i.e. DP = 60). So, the

availability of the chromophore in the side chain structure is higher than for the α,-ω -dual

labeled polymers. The increasing number of donor units only brings a small contribution (60

intensity units pPyOx (1:1) as compared to pPyOx (10:1)) to the overall emission of the

acceptor. Another important aspect is that for the copolymers with Pyrene side chain units

we have excimer formative while for α,-ω -dye labeled ones no excimer signals could be

detected. For the water soluble polymers we observed an increased FRET efficiency of the

donor, but a lower fluorescence intensity of the acceptor. A more detailed study needs to be

performed in order to better understand this behavior. One can speculate that as the pyrene

fluorescence is increased in polar solvents due to solute-solvent interactions, the same thing

should be valid for the Coumarin but in the opposite direction, polar solvents lower the

fluorescence intensity.

2.3.3 Energy transfer in pPyOx-PEtOx-Cou copolymers in film

For the solar cell application the FRET of the copolymer was also tested in film (Fig. 31). In the

glassy matrix the formation of pyrene excimers is obvious and increases with the PyOx content

in the copolymer (Fig. 31). When coumarin is added at the chain end we noticed a shift of the

emission peak dye to FRET. However, due to the presence of such high amounts of pyrene

excimers the FRET efficient cannot be determined accurately from steady state

measurements. The only valid case is the pPyOx-PEtOx-Cou (1 to 1) where we have a very low

amount of excimers. In this case one can notice that FRET efficiency is close to 100%.

Figure 30. Emission spectra of spincoated pPyOx-PEtOx-Cou copolymers irradiated at 345nm.

49

2.4 Light harvesting antennas based on poly-(2-isopropenyl-2-oxazoline) modified

polymers

In the search for an efficient energy transfer between the two dyes, intramolecular energy

transfer was approached and appeared to be very efficient. This due to the higher probability

of interaction between the two dyes when situated on the same chain, in contrast to when

the dyes are on separate chains. To apply this knowledge on polymers, poly-(2-Isopropenyl-2-

oxazoline) 9 was used as scaffold for further modifications due to the accessible and

quantitative reaction of the oxazoline cycle with the carboxylic acids.55 The polymer was

synthesized by anionic polymerization by Dr. Adriana Jerca and the structure is depicted in

figure 32.

Figure 31. Chemical structure of poly-(2-isopropenyl-2-oxazoline).

Due to the oxazoline rings present on the polymer as side chain units, it was possible to modify

the polymer with the fluorophores that possess a carboxylic group so as to obtain similar

polymers with the pPyOx-PEtOx-Cou copolymers. The percentages used can be found in table

11. We synthesized copolymers modified only with one fluorophore and also copolymers

modified with both fluorophores to have both donor and acceptor units on the same

polymeric chain.

2.4.1 Synthesis and characterization of poly-(2-isopropenyl-2-oxazoline) modified polymers

Figure 32. Chemical structure of poly-(2-isopropenyl-2-oxazoline) modified with 1-pyrenebutyric acid and coumarin 343.

50

The different PIPRO 10 modified structures (Fig. 33) were synthesized by the following

method. The needed percentage of acid (1-pyrenebutyric acid or coumarin 343) was weighed

together with PIPRO polymer in a conical vial, DMF was added and the reaction mixture was

heated to 140°C for 5 hours. The copolymers were obtained with good purity and high yields.

The modified copolymers were characterized by 1H NMR, SEC and UV-Vis spectroscopy.

The successful modification of PIPRO with the fluorophores was confirmed by 1H NMR

spectroscopy (Fig. 34). After ring opening the distinctive signals of -CH2-O- from the oxazoline

ring at 4.12 ppm were shifted downfield at 4.37 ppm, while signals from CH2-NH-C=O appear

like a shoulder at 3.5 ppm. Furthermore aromatic protons from the pyrene and the coumarin

moiety show up at 7.5 – 8.3 ppm and 8.4 and 7.1 ppm, respectively. The signal present at 6.1

ppm can be attributed to the NH from the newly formed amide bond. The SEC data proved

that the Mn increases after the modification reaction but the dispersities remained constant,

proving that no side reactions were present (Table 10).

Table 10. Structural characterization of the modified PIPRO polymers.

Polymer code Yield (%) % mol Pya % mol Coua Mn (Da)b Đb

PIPRO-Py 10 95 8.9 0 20300 1.23

PIPRO-Py 5 92 3.8 0 19479 1.22

PIPRO-Cou 10 90 0 7.5 22000 1.24

PIPRO-Cou 3 91 0 1.2 19600 1.27

PIPRO-Cou 1 93 0 0.5 18900 1.21

PIPRO-Py-Cou 10:10 89 9.6 7.4 21900 1.36

PIPRO-Py-Cou 10:3 88 9.1 2.0 19900 1.25

PIPRO-Py-Cou 10:1 91 9.5 0.3 18900 1.28

PIPRO-Py-Cou 5:1 87 4.7 0.9 18800 1.23

a Calculated with the molar extinction coefficient of the model compounds using UV-vis spectroscopy. b Obtained by DMA-SEC analysis using PMMA calibration.

51

Figure 33. 1H NMR of PIPRO-Py-Cou 10:10 (blue), PIPRO-Cou 10 (green) and PIPRO-Py 10 (red).

2.4.2 Synthesis of the model compounds

Due to the fact that we synthesized polymers with very low content of fluorophores and due

to peak overlap, the degree of modification could not be determined directly by 1H NMR

spectroscopy (Fig. 34). Therefore model compounds with the same molecular structure as the

modified copolymers were made (Fig. 35), the modification degree could be calculated in a

more exact manner from the absorption using the Beer–Lambert law.

Figure 34. Molecular structure of model compounds (2-propionamidoethyl 4-(pyren-2-yl)butanoate (11) and 2-propionamidoethyl 11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1-ij]quinoline-10-carboxylate) (12).

52

The model compounds were obtained by reacting 2-ethyl-2-oxazoline with the 1-

pyrenebutyric acid or coumarin 343 using similar conditions as in the PIPRO modification. The

structures were confirmed by 1H NMR spectroscopy (See Appendix A). By making a dilution

series, a linear correlation was found between the absorbance of the model compounds and

the concentration. By applying the Lambert-Beer equation, we were able to determine the

molar extinction coefficient (Fig. 36).

𝐴 = 𝐶 ∗ 𝜀 ∗ 𝑙 (6)

Where A is the absorbance, 𝜀 the molar absorption coefficient, c the concentration of the dye

and l the thickness of the cuvet.

The UV-vis spectra of PIPRO modified copolymers had the absorption maxima corresponding

to the pyrene unit at 345 nm and an absorption maxima at 475 nm which can be attributed

to coumarin. The degree of modification in the copolymer was slightly lower than in the feed

especially when we used coumarin 343. Nevertheless accurate determination of the

fluorophore could be done using the model compounds (Fig. 37).

Figure 35. Linear correlation of absorbance with concentration for a) Pyrene-Etox and b) Coumarin-EtOx.

𝜺𝑪𝒐𝒖−𝑬𝒕𝑶𝒙 = 𝟒𝟏𝟕𝟎𝟓 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

R² = 0.997

𝜺𝑷𝒚−𝑬𝒕𝑶𝒙 = 𝟑𝟖𝟐𝟏𝟏 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

R² = 0.998

53

2.4.3 Energy transfer in PIPRO modified copolymers

To investigate the effect of solvent also H2O was tested, but all the PIPRO modified polymers

were insoluble in water except with a 5:1 ratio of pyrene to coumarin. This can be explained

by the hydrophobic behavior of pyrene. For the fluorescence investigation we persued the

two established routes, namely: i) first we made mixtures of single dye modified polymers and

checked for intermolecular FRET; then ii) we studied the intramolecular fluorescence

properties of the polymers modified with both dyes (Fig. 38).

When we mixed the two copolymers very little FRET could be detected, based on the change

in intensity of the pyrene (Fig. 38 a). The signal corresponding to Coumarin emission was

greatly affected by the excimer emission of pyrene. Therefore we can conclude that no

efficient intermolecular transfer can take place when we use this pair of polymeric

fluorophores. The situation changes once we have the dyes in close proximity on the same

Figure 36. Absorption spectra for the modified PIPRO copolymers.

Figure 37. a) Intermolecular energy transfer for dual-modified PIPRO polymers. b) Emission spectra of PIPRO dual-modified polymers in CHCl3 irradiated at 345 nm

54

polymeric chain. Although when we go to higher pyrene ratios we have also some emission,

corresponding to pyrene excimers, a clear peak in this case could be distinguished and

attributed to coumarin emission at 470 nm. The highest FRET signal is registered for the

polymer modified with 9.1% Pyrene and 2 % Coumarin. Further increasing the Coumarin

content doesn’t improve the fluorescence of the acceptor. FRET efficiency could not be

determined in the case of PIPRO modified polymers steady-state method due to the presence

of excimers and the fact that the ratio between monomer and excimer is affected by the

chemical composition of the copolymers. For the PIPRO polymer only modified with pyrene

(4.6%), the ratio of monomer/excimer peak intensity is 0.6. When 2% coumarin is added to

the polymer chain, it is impossible to predict that the ratio will remain the same. Due to this

the different data could not be compared due to different monomer/excimer ratios. Thus the

decrease in donor intensity could not be used to assess the FRET efficiency. We can conclude

that there is an optimal ratio between pyrene and coumarin in order to maximize the

fluorescence of the acceptor.

2.4.4 Energy transfer in modified PIPRO copolymers in film

In the film, the pyrene excimer formation is strongly present in the emission spectrum of the

modified PIPRO copolymers, similar to the pPyOx-PEtOx-Cou polymers (Fig. 39). Again when

coumarin is present on the same polymeric chain with the pyrene a decrease of the pyrene

emission signal is noticed accompanied by the appearance of a new peak at 480 nm

corresponding to coumarin emission. However, due to the presence of such high excimer

signals no accurate FRET efficiency cannot be determined.

Figure 38. Emission spectra of spincoated modified PIPRO copolymers irradiated at 347nm.

55

56

3. Conclusions & Outlook During this project, the synthesis and the photophysical properties of several modified

polymers, based on poly-(2-oxazoline) scaffolds, with pyrene and coumarin units were

investigated. The aim of this project was to obtain light harvesting polymers based on FRET.

Therefore, three synthetic approaches were developed.

The first one was the synthesis of poly-(2-ethyl-2-oxazoline) polymers with labeled with

fluorescent end-groups. For the single labeled polymers, we used the “terminator method”

and end capped the living chains with the 1-pyrenebutyric acid or coumarin 343. While for the

α,-ω-labeled polymers we initiated the chains with 1-bromomethyl pyrene and end capped

them with coumarin 343. In all cases the polymerization of EtOx was living and controlled

leading to the target molecular weights and polymers with Ð lower than 1.15. The end capping

reaction was very efficient allowing us to have a degree of functionalization of the end groups

higher than 90%. The photophysical investigations lead to the conclusion that both

fluorophores must be located on the same polymeric chain in order to have an efficient FRET

transfer between fluorophores in solution.

The second strategy was to synthesize a monomer containing pyrene units that could be

copolymerized with EtOx by CROP and then used the “terminator method” to introduce the

coumarin units. In this way, an antenna type polymer could be obtained. 2-(1-Pyrenylpropyl)-

2-oxazoline monomer was successfully synthesized although with a rather low overall yield of

10 % (due to purifications). CROP copolymerization with EtOx followed by end capping

reaction lead to the desired polymeric structures. Copolymers with different pyrene content

were obtained with low dispersities and high degree of functionalization of the end groups.

FRET efficiency in solution for these copolymers depended on the ratio between donor and

acceptor groups. Namely if the ratio was closer to 1 than the FRET efficiency was higher. A

very interesting effect was discovered when we investigated photophysical properties of

these polymers in water. The FRET efficiency increased dramatically from 47.7% to 80.3%.

Water is considered a less good solvent, in which the polymer coil will be present in a collapsed

state leading to closer proximity and thus higher FRET efficiencies.

The third strategy was based on a polymer analogous reaction. Namely poly-(2-isopropenyl-

2-oxazoline) synthesized by anionic polymerization was modified with the two fluorophores

57

making use of the reactivity of the oxazoline cycle towards carboxylic acids. Consequently,

copolymers with different donor/acceptor ratios could be easily obtained. Also in this case we

noticed that FRET is present only when both fluorophores are located on the same chain.

Quantitative determination of FRET was not possible for this polymers by steady state

fluorescent measurements due to the formation of pyrene excimers that overlapped with the

coumarin emission signal.

In order to use the polymers for light harvesting properties in future work, the properties were

also tested in film. Even though in solution the FRET efficiency of the single labeled polymers

was rather low, in film the situation changed. If we spin coat a mixture of the two polymers

(1:1 molar ratio) and obtain the film we notice that we have close to 100% FRET efficiency. For

α,-ω-labeled PEtOx polymers we also have a higher EFRET than in solution, due to the reduced

distance between donor and acceptor groups. For the PyOx and PIPRO copolymers we always

have generation of pyrene excimers that affects the FRET and prevails us from accurately

determining the FRET efficiency.

As general conclusion, we can say that each type of polymeric architecture obtained during

this project has his own advantages and disadvantages. If our applications are targeted for

solution than PyOx and PIPRO copolymers are the right choice, but if we go towards solid state

application than, α, ω-labeled PEtOx polymers should be used.

Future work will include the optimization of the composition of the copolymers and testing

the synthesized polymeric structures in an organic solar cell. By this, various ways could be

explored to optimize the output voltage and synthesize a more efficient solar cell.

58

4. Appendix A

4.1 Materials All the common used solvents were HPLC grade and include: dichloromethane (DCM, >99.8%,

Sigma Aldrich), diethyl ether (>99.9%, Sigma Aldrich), acetonitrile (ACN, >99.9%, Sigma

Aldrich), chloroform (CHCl3, >99%, Fisher Chemical) and N, N-dimethyl formamide (DMF,

>99%, Biosolve). Dry solvents, like methanol (99.9%, Extra Dry, AcroSeal®) and dimethyl

sulfoxide (99.7%) were purchased from Acros Organics. Dry DCM, TEA, THF and ACN were

obtained from a custom made J.W. Meyer solvent purification system and were dried over

aluminium oxide column.

The following chemicals were used as received: 1-pyrenebutyric acid (>98.0%, TCI), and the

coumarin 343 (>97.0%, TCI). 2-Ethyl-2-oxazoline (EtOx was kindly donated by Polymer

Chemistry and purified by distillation over BaO. Poly-(2-isopropenyl-2-oxazoline) polymers

were synthesized by Dr. Adriana Jerca by anionic polymerization of 2-isopropenyl-2-oxazoline

(Mn = 18500, Đ= 1.21). Br-Me-Pyrene (>92.0%, Sigma Aldrich) was recrystallized from CHCl3 (3

times).

4.2 Equipment NMR Proton and carbon-13 nuclear magnetic resonance spectra were recorded on a Bruker

Avance 300 MHz at room temperature. NMR spectra were measured in chloroform-d (CDCl3)

from Euriso-top. The chemical shifts are given in parts per million (δ), relative to CDCl3 at 77.36

ppm.

Gas chromatography (GC) was performed on a GC8000 from CE instruments with a DB-5MS

column (60 m x 0.249 mm x 0.25 µm) from J&W scientific. Injections are performed with a CTC

A200S auto sampler and detection is done with a flame ionization detector (FID) which burned

a H2 /air mixture. The carrier gas (H2) is pushed through the column with a pressure of 100

bar. Injector and detector are kept at a constant temperature of 300 °C.

Size exclusion Chromatography (SEC) Size-exclusion chromatography (SEC) was performed on

a Agilent 1260-series HPLC system equipped with a 1260 online degasser, a 1260 ISO-pump, a

1260 automatic liquid sampler (ALS), a thermostatted column compartment (TCC) at 50°C

59

equipped with two PLgel 5 μm mixed-D columns and a mixed-D guard column in series, a 1260

diode array detector (DAD) and a 1260 refractive index detector (RID). The used eluent was

DMA containing 50 mM of LiCl at a flow rate of 0.500 mL/min. The spectra were analyzed

using the Agilent Chemstation software with the GPC add on. Molar mass and dispersity values

were calculated against PMMA standards from PSS.

UV/VIS spectrometer UV/Vis spectra are recorded on a Varian Cary 100 Bio UVVIS

spectrophotometer equipped with a Cary temperature and stir control.

Fluorescence spectrometer The emission and excitation spectra are recorded on a Varian

Cary Eclipse fluoro-spectrophotometer also equipped with a Cary temperature and stir

control. The excitation and emission wavelength were varied according to the sample

absorption maximum, same was done for the detectors voltage, so the peaks did not go out

of scale. The slit width of the excitation and emission were kept at 5 nm during the

measurements. Samples were measured in quartz cuvettes with a fluorescence spectrometer

pathlength of 1.0 nm in the wavelength range of 200-700 nm.

Microwave All the microwave polymerizations were performed in a Biotage initiator

Microwave System with a temperature range of 40-250°C equipped with a variable magnetic

stirrer (300-900 RPM).

Chromatographic columns on aluminum oxide and silica were performed on Merck Alox 90

standard aluminum oxide and on Davisil chromatographic silica mecia LC60A 70-200 micron

respectively.

Silica TLC’s were taken on pre-coated Macherey-Nagel ALUGRAM SIL G/UV254 plates.

Aluminium oxide TLC’s were taken on Merck TLC Aluminum oxide 60 F254 neutral.

Flash chromatography was performed on a Grace Reveleris® flash chromatography system

using silica Reveleris flash cartridges.

60

Glove box The system used is a VIGOR Sci-Lab SG 1200/750 Glovebox System, the purity level

obtained is at most, < 1 ppm O2+ H2O

Deionized water was prepared with a resistivity less than 18.2 MΩ x cm using an Arium 611

from Sartorius with the Sartopore 2 150 (0.45 + 0.2 µm poresize) cartridge filter.

Spin Coater All films were made with 30 seconds of spinning at 500 RPM/sec up to 2000 RPM.

Before spin-coating the solutions were filtered and the quartz plates were extensively cleaned

with acetone and blown dry with nitrogen gas.

.

61

4.3 Compound synthesis

4.3.1 Organic compounds

5.3.1.1 Synthesis of (2-propionamidoethyl 4-(pyren-2-yl) butanoate (Pyrene-EtOx)

0.16 g (0.55 mmoles) of Pyrene-COOH was dissolved in 1 mL of DMF, then 0.05g (0.50 mmoles)

of 2-ethyl-2-oxazoline was added under continuous stirring. Then the vial was capped and the

mixture was heated to 100°C for 5 hours. The solution was precipitated in crushed ice, and

the white precipitate was filtered off and washed three times with water on the filter. The

precipitate was dried overnight in the vacuum oven at 50°C. Further purification was

performed by column chromatography (SiO2) eluent: hexane/EtOAc (30/70).

Yield: 80%

UV-vis: λmax CHCl3: 345 nm

1H NMR (300 MHz, CDCl3) δ 8.32 – 7.68 (m, 9H; Pyrene), 5.58 (s, 1H; NH), 4.14 – 3.97 (t, 2H,O-

CH2-CH2), 3.41 (dd, J = 10.8, 5.6 Hz, 2H, CH2- CH2-NH), 3.36 – 3.28 (t, 2H, Pyrene- CH2- CH2),

2.38 (t, J = 7.3 Hz, 2H,O=C- CH2-CH3), 2.21 – 1.96 (m, 4H, CH2- CH2- CH2-C=O), 1.02 (t, J = 7.6

Hz, 3H, CH2-CH3).

Figure 39. 1H NMR of model compound EtOx-Pyrene.

a

b

c

d

e

f

g

h

f

g d c e a b

h

62

5.3.1.2 Synthesis of (2-propionamidoethyl 4-(pyren-2-yl)butanoate and 2-propionamidoethyl

11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1-ij]quinoline-10-carboxylate)

(Cou-EtOx)

0.1 g (0.35 mmoles) of Pyrene-COOH was dissolved in 1 mL of DMF, than 1.96 (20 mmoles) of

2-ethyl-2-oxazoline was added under continuous stirring. Then the vial was capped and the

mixture was heated to 140°C for 15 minutes, at this point the reaction was completed. Then

the solution was precipitated in crushed ice, and the dark orange precipitate was filtered off

and washed three times with water on the filter. The precipitate was dried overnight in the

vacuum oven at 50°C. Further purification was performed by column chromatography (SiO2)

eluent: hexane/acetone (80/20).

Yield: 50%

UV-vis: λmax CHCl3: 440 nm

1H NMR (400 MHz, CDCl3) δ 8.25 (s, 1H, C-CH=C-C=O), 6.89 (s, 1H, C-CH-C), 6.54 (s, 1H, NH),

4.31 – 4.27 (t, 2H, O-CH2-CH2), 3.58 (q, J = 9.9, 5.0 Hz, 2H, CH2-N-CH2), 3.28 (dd, J = 11.4, 5.6

Hz, 4H, CH2-CH2-NH), 2.81 (t, J = 6.4 Hz, 2H, CH2-CH2-C), 2.73 – 2.66 (m, 2H, CH2- CH2-C), 2.20

(q, J = 7.6 Hz, 2H, CH2-CH2-CH2), 1.96 – 1.85 (m, 2H, O=C-CH2-CH3), 1.14 – 1.07 (m, 3H,CH2-

CH3).

Figure 40. 1H NMR of model compound EtOx-Pyrene.

a b

k

d e

g h

i

a

b c f

d

e

j

j

d a

g

k c h

b

i

f

63

5.3.1.3 Synthesis of 1-pyrenebutyric acid chloride

5g (0.017 moles) of 1-pyrenebutyric acid was weight in a round bottom flask. Then 25 mL of

dry DCM were added under argon the mixture was cooled in an ice bath. Then 3.09 (0.026

moles) thionyl chloride was added dropwise. The mixture was left to stirr at room temperature

overnight. The DCM was evaporated under reduced pressure and then was three times

redissolved in DCM and evaporated again in order to remove the excess of SOCl2, until a yellow

solid was obtained.

Yield: 99%.

1H NMR (300 MHz, CDCl3) δ 8.22 – 7.70 (m, 9H, Pyrene), 3.40 – 3.27 (t, 2H, Py-CH2-CH2), 2.93

(t, J = 7.1 Hz, 2H, CH2-CH2-CH2), 2.19 (dt, J = 14.6, 7.2 Hz, 2H, CH2-CH2-C=O).

5.3.1.4 Synthesis of N-(2-chloroethyl)-4-(pyren-1-yl)butanamide

4.95 g (0.016 moles) of 1-pyrenebutyric acid chloride and 1.57 g (0.020 moles) of 2-

chloroethylamine hydrochloride was suspended in dry DCM (50 mL) in an argon atmosphere

and the reaction mixture was cooled to 0°C. 4.898 g (0.048 moles) of TEA was added dropwise,

and the reaction mixture was stirred overnight. Water was added to the reaction and the

aqeous phase was extraced with DCM (2x 15 mL). The combined organic phases were washed

with two times 9% HCl solution, then with saturate sodium bicarbonate, water and brine.

Finally the organic phase was dried over MgSO4 and the solvent was removed under reduced

pressure. The compound was further purified by flash chromatography on silicagel eluting

with a hexane/ethyl acetate mixture (50/50).

Yield: 20%

1H NMR (300 MHz, CDCl3) δ 8.29 – 7.63 (m, 9H, pyrene), 5.71 (s, 1H, O=C-NH-CH2), 3.62 – 3.39

(m, 4H, Cl-CH2-CH2-NH), 3.27 (t, J = 15.3, 8.3 Hz, 2H, O=C-CH2-CH2), 2.28 – 2.01 (m, 4H, Pyrene-

CH2-CH2-CH2).

5.3.1.5 Synthesis of 2-(3-(pyren-1-yl)propyl)-4,5-dihydrooxazole

1.17 g ( 0.004 moles) N-(2-chloroethyl)-4-(3, 6-dihydropyren-1-yl) butanamide was dissolved

in 50 mL THF and 0.047 g (0.000178 moles) 18-crown-6-ether was added under argon

atmosphere. Then 0.57 g (0.010 moles) KOH was added under vigorous stirring. The reaction

was left to stir overnight. The THF was removed under reduced pressure followed by addition

of DCM (50 mL) and washing with water (3x15mL). The water phase was extracted with DCM

64

three times. The combined organic phases were washed with brine and dried over MgSO4. The

solvent was removed yielding a yellowish crude product. The compound was further purified

by flash chromatography on silica gel eluting with a hexane/EtOAc mixture (70/30).

Yield: 51%

1H NMR (400 MHz, CDCl3) δ 8.26 – 7.73 (m, 9H, Pyrene), 4.21 – 4.05 (t, 2H, O-CH2-CH2), 3.75

(t, J = 9.4 Hz, 2H, N-CH2-CH2), 3.42 – 3.26 (t, 2H, CNO-CH2-CH2), 2.43 – 2.28 (m, 2H,Pyrene-CH2-

CH2 ), 2.23 – 2.04 (t, 2H, CH2-CH2-CH2).

65

4.3.2 Polymer synthesis

5.3.2.1 Microwave –assisted synthesis of single-labeled poly-(2-ethyl-2-oxazoline) s

To study the effect of chain length, PEtOx with three different polymerization degree (DP= 30,

50 and 70) were synthesized. All polymers were synthesized under similar conditions. The

monomer concentration was 4 M for all polymerizations. The monomer/initiator [M]/ [I] ratio

was 30/1, 50/1 and 70/1, respectively. Reactions were carried out in capped microwave vials.

The vials were dried in the oven at 200°C and cooled under argon to room temperature before

use. The solutions were prepared in a glove box (Vigor Sci-Lab SG 1200/750) under argon. The

polymerizations mixtures were heated to 140°C under microwave irradiation for 216 s ([M]/

[I] = 30), 6 min ([M]/ [I] = 50) and 8 min ([M]/ [I] = 70) to reach almost full conversion.

For the end-capping reactions, the vial was cooled down to room temperature and a solution

of the corresponding acid in DMF 1.5-fold excess was added via a syringe through the septum

of the capped microwave vial containing the living oligomers (EtOx). Thereafter TEA was

added similarly in a two-fold excess. The given amounts were varied according to the used

monomer to initiator ratios. The reaction solution was heated to 100°C for 20 hours. After

cooling to room temperature the reaction mixture was diluted with chloroform and the

solution was washed three times with saturated aqueous sodium hydrogen carbonate. Then

the solution was washed with brine, dried over MgSO4 and filtered. The solvent was

evaporated under reduced pressure resulting a foamy polymer.

The polymers were dissolved again in 3mL chloroform and precipitated in ice cold diethyl

ether. The solutions were centrifuged for 10 minutes (7500 rpm) at 5°C. The solution was cast

off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under

reduced pressure resulting a white or yellow foam for the polymers modified with 1-

pyrenebutyric acid or coumarin 343, respectively. Finally the polymers were dried overnight

in the vacuum oven at 50°C. 1H NMR and SEC analysis confirmed the initial DP.

1H NMR (300 MHz, CDCl3) δ 8.30 – 7.68 ( 9H, Pyrene), 4.08 (s, 2H, CH2-c=O-O), 3.38 (4H, N-

CH2-CH2), 2.95 (2H, O-O=C-CH2), 2.89 (2H, CH3-N-CH2), 2.26 (2H, CH3-CH2-C=O and CH2-CH2-

Py), 1.05 (3H, CH3-CH2-C=O).

66

Figure 41. 1H NMR of PEtOx-Pyrene DP30.

Figure 42. 1H NMR of PEtOx-Coumarin DP30.

a

b

h

d

e f

g

b

c

i

a

e

h

b + c

d

i

e + f +g

a

a b d

f

c

g

f

e

e h

h g

d

b c a e h

i

i

j

j

67

5.3.1.2 Microwave assisted of dual-labeled poly-(2-ethyl-2-oxazoline)s

All polymers were synthesized under similar conditions. The monomer concentration was 4 M

for all polymerizations. The monomer/initiator [M]/ [I] ratio was 30/1, 50/1 and 70/1,

respectively. Reactions were carried out in capped microwave vials. The vials were dried in

the oven at 200°C and cooled under argon to room temperature before use. The solutions

were prepared in a glove box (Vigor Sci-Lab SG 1200/750) under argon. The polymerizations

mixtures were heated to 140°C under microwave irradiation for 216 s ([M]/ [I] = 30), 6 min

([M]/ [I] = 50) and 8 min ([M]/ [I] = 70) to reach almost full conversion. In this case the initiator

used was 1-(Bromomethyl)pyrene.









evaporated under reduced pressure resulting in foamy polymers.


ether. The solutions were centrifuged form 10 minutes (7500 rpm) at 5°C. The solution was

cast off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under

reduced pressure resulting in a yellow foamy polymer. Finally the polymers were dried

overnight in the vacuum oven at 50°C.

1H NMR (300 MHz, CDCl3) δ 8.36 – 7.87 (9H, Pyrene), 6.86 (1H, C-H-C), 5.27 (2H, pyrene-CH2),

4.33 (2H, CH2-CH2-O), 3.29 (4H, NH-CH2-CH2), 2.75 (4H, CH2-N-CH2), 2.26 (4H, C-O=C-CH2-CH3

and C-CH2-CH2), 1.15 – 0.88 (3H, CH2-CH3).

68

5.3.2.3 Synthesis of dye functionalized poly-(2-isopropenyl-2-oxazoline) copolymers

The functionalized copolymers were obtained by oxazoline ring-opening addition in the

presence of the fluorescent dyes. In a typical run, 1.8 mL of dried DMF were added to 0.025 g

of the corresponding dye and 0.1 g (0.0009 moles) PIPRO in a vial. The mixture was heated at

140°C with stirring for 5 hours. The reaction mixture was diluted with 1 ml of CHCl3 and

precipitated in cold diethyl ether. The solution was centrifuged for 10 min at 7500 rpm. The

product was purified twice by reprecipitation from chloroform in diethyl ether and dried at

50°C overnight under vacuum.

1H NMR (300 MHz, CDCl3) δ 8.02 (9H, Pyrene), 6.06 (1H, NH), 4.08 (4H, O-CH2 and O=O-CH2),

3.65 (4H, N-CH2 + NH-CH2), 3.32 (Pyrene-CH2), 2.42 (2H, CH2 -CH2-CH2), 1.75 (2H, O=C-CH2-

CH2), 1.25 – 0.84 (3H, CH2-CH3).

c

a

b c

g

f e h

i j

d

c d

j h

g a

g h i f e

c+ j

Figure 43. 1H NMR of Py-CH2-PEtOx-Cou with DP=30.

69

5.3.2.4 Microwave assisted copolymerization of 2-(1-pyrenylpropyl)-2-oxazoline and EtOx

The polymerization mixtures for the microwave assisted synthesis of a series of statistical

EtOx-PyOx copolymers were prepared in the glove box. In each microwave vial EtOx, PyOx and

ACN was added resulting in a 4 M total monomer concentration. The total monomer to

initiator ratio was 60. The amount of PyOx used was 1.5; 5.5; 8.5 and 16.5 %mol to obtain a

series with compositions similar to the PIPRO modified copolymers. The vials were capped and

were subsequently heated to 140°C for 12 min to reach full conversion.









evaporated under reduced pressure resulting in foamy polymers.


ether. The solutions were centrifuged form 10 minutes (7500 rpm) at 5°C. The solution was

cast off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under

reduced pressure resulting in white or yellow foam for the polymers modified with 1-

pyrenebutyric acid and coumarin 343, respectively. Finally the polymers were dried overnight

in the vacuum oven at 50°C.

1H NMR (300 MHz, CDCl3) δ 8.41 – 7.57 (9H, pyrene), 3.35 (6H, N-CH2-CH2 + Pyrene-CH2 ), 2.30

(s, 4H), 1.02 (3H, CH2-CH3).

70

Figure 44. 1H NMR of pPyOx-PEtOx-Cou 10:1

71

72

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Scientific article Polymeric light harvesting antenna

R. Merckxa, V. Jercaa, R. Hoogenbooma*

a Supramolecular Chemistry Group, Ghent University, Department of Organic and Macromolecular Chemistry, Ghent 9000, Belgium

*[email protected] Keywords: Light harvesting, stimuli-responsive materials, FRET, sustainable development, Dyes,

Fluorescence, oxazolines.

The synthesis of one pyrene containing monomer and the polymerization/ modification of three different polymers will be described. Successful FRET was described between the couple by Hong et al 1.One of the requirements for an efficient FRET is the proximity of the two dyes. Therefore,

the aim was to modify the dyes onto a polymer scaffold to improve the energy transfer and spincoat polymeric films with light harvesting properties.

Introduction

The world energy consumption is ca. 4.7 ∗ 1020J (450quadrillion Btu) and is expected to grow 2% each year for the next 25 years.2 As the world population grows, the demand for natural resources, in the form of energy increases, while the supply of coal, oil and gas drastically decreases. Earth’s resources alone are not enough to cover this consumption, so the anthropological impact should decrease or other sources need to be explored. Concerns about global warming have led to high interest in the field of renewable energy.3,4,5 In this area there were numeral attempts to create energy in a green , more sustainable way (e.g. Solar cells, water and wind turbines, etc...). The inspiration behind this project are the light-harvesting

systems in nature which absorb light and transfer the energy to photosynthetic reaction centers through a precisely spaced array of chromophores that can transport energy over long distances. This highly efficient energy transfer is done typically through a series of fluorescence resonance energy transfer (FRET) events (Fig. 1). Aside from the cellular context, light-harvesting systems could be utilized to sensitize solar cells, drive photo catalysts or even be used as optical sensors. The design and development of artificial light-harvesting systems is a contemporary academic and industrial challenge having important economic and ecological implications.

Pyrene and coumarin 343 have been chosen as the FRET pair (i.e. donor and acceptor) due to several important advantages that those two fluorophores have to offer. Pyrene is an

Figure 45. Schemeticaly illustrated Förster resonance energy transfer.

https://biblio.ugent.be/person/802002296007

76

alternant polycyclic aromatic hydrocarbon consisting of four fused benzene rings with a large, flat aromatic system, showing high thermal stability, extensive electron delocalization and electron accepting nature. It is the most studied as a fluorophore, and has several advantages such as long singlet lifetime, sensitive to the changes of the polarity of its microenvironment, and strong tendency to form excimers which possesses high fluorescence quantum yield. Coumarin 343 emits in the blue green spectral region, has a very high molar extinction coefficient, displays large Stokes shifts and has large fluorescence quantum yield, therefore fulfilling all the required conditions to be a very good acceptor in FRET experiments. Pyrene (donor molecule) has an absorption in the range of 300-350 nm and emits between 370 and 450 nm in the range of absorption for coumarin 343 (acceptor molecule). Thus, the coumarin 343 absorption is expected to overlap with the emission of the pyrene which is one of the main requirements for the energy transfer. Although pyrene absorbs in the UV region of the spectrum and solar light is observed to have a maximum intensity between 400-700 nm, this is considered not to be a problem due to the high molar extinction coefficient and quantum yield.

As polymeric scaffold to covalently attach the dyes, the poly-(2-oxazoline)s were chosen due to several advantages offered by this class of polymers. They can be synthesized in a controlled and living manner by CROP. Different functionalities can be easily introduced by using initiators or terminating agents that have the desired chemical groups. Finally, the monomers can be modified with different chemical substituents in the 2-position to match our purposes. Poly-(2-alkyl-2-oxazoline)s have also good film forming properties, are soluble in a large number of solvents and have relatively high glass transition temperatures.

In the first part of the project we will attempt to synthesize single or dual labeled polymers with pyrene and/or coumarin groups in order to highlight the type of energy transfer present in these polymers inter or intramolecular. For single labeled poly-(2-ethyl-2-oxazoline) polymers we will make use of the “termination method”. This implies initiation with a suitable initiator (i.e. methyl tosylate) and end-capping the living chains with 1-pyrenebutyric acid or coumarin 343. For the α,-ω-labeled polymers initiation will be done with a compound that bears pyrene unit (i.e. 1-bromomethyl pyrene) and then same “termination method” will be used to introduce coumarin groups. The next step will be the physico-chemical characterization of this polymers. Then the photophysical properties will be investigated.

In the second part of the project we will explore the possibility to obtain copolymer bearing donor units in the side chain and acceptor units at the end chain similar to an antenna light harvesting system. This will be achieved by cationic ring opening copolymerization of EtOx with a specially designed monomer bearing pyrene units (i.e. 2-(1-Pyrenylpropyl)-2-oxazoline) and subsequent end capping of the living chains with coumarin 343. First the pyrene monomer will be synthesized, then copolymers with different amount of pyrene units will be obtained. Also in this case a physico-chemical characterization of the polymer is mandatory followed by the investigation of energy transfer properties.

In the last part, we will make used of the highly reactivity of the oxazoline cycle towards carboxylic acids and obtained light harvesting polymers by chemically modifying a poly(2-isopropenyl-2-oxazoline) scaffold with 1-pyrenebutyric acid and/or coumarin 343. Using this method, we will be able to synthesize polymers with different ratios of donor and acceptor. Therefore, enabling us to precisely tune up the energy transfer efficiency based on chemical

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composition. As stated before physico-chemical characterization and photophysical investigations will follow after synthesis.

The ultimate goal of this project will be to find the optimum between polymeric architecture, location of fluorophores onto the polymer chain (side-chain, end-chain) and ratio between donor and acceptor groups in order to obtain the polymer with the best light harvesting properties in solution and/or in solid state.

Experimental

The used materials, synthesis and characterization of the target compounds is described in

the supporting information.

Results and Discussion

According to their UV-Vis and fluorescence spectra the pyrenebutyric acid and the coumarin 343 donor-acceptor pair is suited for FRET experiments. Looking at the UV-vis spectra (Fig. 2) one can observe that the maximum wavelengths of the two absorption bands are well separated (Pyrene 𝜆𝑚𝑎𝑥 = 345 nm, Coumarin 343 𝜆𝑚𝑎𝑥 = 450 nm).

Furthermore Coumarin 343 has very low absorption between 300 and 375 nm and the absorption band of pyrene is located in this spectral region. To study the energy transfer between Py-COOH and Cou343 the emission spectra of Py-COOH and Cou343 mixture (1:1 molar ratio) was measured with the excitation wavelength fixed at 345 nm ( corresponding to Py-COOH absorption maximum). Figure 3 shows the fluorescence spectra of Py-COOH, Cou343 and their mixture in CHCl3. From figure 3 a, we can observe that the fluorescence intensity of pure Py-COOH is much higher than the one for Cou343. For the 1:1 mixture we can notice a decrease in the intensity of the Py-COOH with respect to pure Py-COOH emission.

Figure 46. Normalized absorption spectra of the two fluorophores in CHCl3

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Also the Cou343 emission increases with respect to the pure Cou343 (Fig 3 a). This can be ascribed to the transfer of energy of the excited state from Py-COOH molecules to Cou343 molecules via FRET. It was observed that the FRET efficiency increases with the increase in acceptor concentration in the mixture. This may be due to closer proximity of the Py-COOH and Cou 343 with increase in Cou343 concentration. Calculation was done according to the formulas found in the experimental part.

It is well known that FRET efficiency largely depends on the molecular proximity of donor-acceptor in the mixture.7 Therefore it is interesting to check the FRET efficiency by varying the acceptor concentration in the mixture. Accordingly we measured the fluorescence spectra of pyrenebutyric acid mixed with varying concentration of coumarin 343. It was observed that the FRET efficiency increases with the increase in acceptor concentration in the mixture. This may be due to closer proximity of the Py-COOH and Cou 343 with increase in Cou343 concentration. The values of spectral overlap 𝐽(𝜆), energy transfer efficiency (EFRET), Förster radius (𝑅0) and the donor-acceptor distance (𝑟) have been calculated and listed in table 1. Highest energy transfer efficiency was found to be 20.7% for an acceptor concentration of 100 µM. In literature there are several reports where increase in FRET efficiency was been observed with increasing acceptor concentration.7

Table 11. Energy transfer parameters for Py-COOH-Cou343 pair in CHCl3. The donor concentration was fixed at 10µM.

Well-defined polymers with precise functionality, narrow polydispersity and controlled molecular weight are becoming prerequisite in light harvesting applications. The polymer

Acceptor (Cou343)

concentration (µM)

𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

5 12.21 4.82 12.73 0.3

10 12.96 4.87 9.51 1.8

50 12.01 4.81 7.92 4.8

100 12.50 4.84 6.05 20.8

Figure 47. a) Emission spectra of pure Py-COOH (pink curve), Cou343 (grey curve) and Py-COOH + Cou343 ( 1:1 molar ratio) mixture in CHCl3. b) Emission spectra of Py-COOH in the prescence of various amounts of Cou343 in CHCl3. Excitation

wavelength was 345 nm in all cases, Py-COOH concentration was always constant 10 µM.

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scaffold will provide film-forming ability, mechanical stability and processing advantages. FRET is sensitive to intramolecular and intermolecular interactions and has also been used for the quantitative characterization of the interchain distance.8 Therefore we first started our study by the synthesis of single-labeled polymers, namely the donor and acceptor molecules are placed on different polymeric chains. Then we proceeded with the synthesis of dual-labeled poly-(2-ethyl-2-oxazolines)s polymers where the donor is located at one end and the acceptor at the other end side. The polymer choice of 2-oxazolines was substantiated by literature.9 The poly 2-oxazolines form a dipole layer which is assumed to bring the dyes closer to each other and so to improve the probability for energy transfer.10 The living nature of the CROP of 2-substituted-2-oxazolines allows facile preparation of well-defined polymers. Depending on the monomers, the properties of the resulting poly-2-oxazolines can be easily varied. Methyl and ethyl side groups result in water-soluble polymers, whereas aromatic groups result in hydrophobic polymers. The living chains were end-capped with the corresponding dye in the presence of TEA, therefore providing easy access to the desired labeled polymer (see Fig. 4).

Figure 48: Molecular structure of PEtOx modified with pyrene 6 and coumarin 7.

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The intermolecular energy transfer in solution was investigated by steady state fluorescence

spectroscopy. First stock solutions were prepared, afterwards from this more diluted samples

were prepared on which the FRET was investigated. The solutions always had a content of 10

µM of pyrene fluorophore. The solutions were irradiated with 345 nm light which corresponds

with the maximum of the PEtOx-chains modified with the pyrene moiety. The emission spectra

of the DP 30 endcapped with Py-COOH or Cou343 were similar to the corresponding organic

fluorophores (Fig 5 a). When mixing the polymers as to have a 1:1 molar ratio between the

fluophores an energy transfer could be detected (Fig 5 a, c and d). From the obtained FRET

efficiencies, it can be concluded that energy transfer is very low, since the highest transfer is

8% and obtained for PEtOx chains with DP=30 (table 2). When increasing the concentration of

acceptor groups (Fig. 5 b) we noticed a higher donor quenching (i.e. lower emission intensity)

but the increased emission signal corresponding to the acceptor is due to the auto-

fluorescence of the coumarin. Therefore we cannot say that FRET efficiency is increasing.

Table 12. Energy transfer parameters for mixing single dye labeled PEtOx mixed in 1:1 raio in CHCl3. The donor concentration was fixed at 10µM.

Polymer codes DP 𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

PEtOx-Mix 30 14.85 4.98 7.34 8.9

Figure 49. a) Emission spectra of pure PEtOx-Py (pink curve), PEtOx-Cou (grey curve) and PEtOx-Py + PEtOx-Cou ( 1:1 molar ratio) mixture in CHCl3 with a DP=50. b) Emission spectra of PEtOx-Py in the prescence of various amounts of PEtOx-Cou in

CHCl3. PEtOx-Py concentration was 10 µM. c) idem as a), but with DP= 50. d) idem as a), but with DP= 70. In all cases excitation was 345 nm and the pyrene concentration was 10µM.

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The intermolecular mixing of PEtOx was considered not to give an efficient energy transfer at equimolar ratio between the fluophores, thus an intramolecular approach was persued. For this the pyrene and coumarin units were placed on the same PEtOx chain, in order to increase the probability of the two dyes to be in close proximity.11 To produce such polymers another initiator must be used. In this case we choose 1-(Bromomethyl) pyrene to initiate the CROP of 2-ethyl-oxazoline and introduce the pyrene group in the α-position, followed by end capping with coumarin 343. Following this strategy α,-ω –dye labeled polymers can be easily obtained (Fig. 6). Since no literature could be found regarding CROP of 2-oxazolines with this initiator we had to do a kinetic investigation.

Figure 50. Chemical structure of py-CH2-PEtOx-Cou polymers.

The polymerization kinetics for the living CROP of EtOx initiated with 1-(bromomethyl) pyrene was

performed at 140 °C under microwave irradiation using ACN as solvent. Subsequent SEC analysis

revealed a linear increase in number average molecular weight (Mn) with conversion and low

dispersities (Đ) (See experimental), demonstrating that the polymerization proceeded in a

controlled manner. The first-order kinetic plots of monomer consumption with respect to

reaction time revealed a linear relationship thus demonstrating a constant amount of

propagating species indicative of the abscence of terminator and as well as relatively fast

initiation (Fig. 7).The polymerization rate constant was calculated form the linear fit up to 80

% conversion, assuming that the concentration of propagating species is equal to the initial

initiator concentration. The value obtained is similar for the polymerization of EtOx initiated

with benzyl bromide.8 In order to verify and to investigate the chain length influence on FRET

efficiency we synthesize three polymers with different DP values, namely 30, 50 and 70. The

polymers were obtained in high yields with low polydispersities and high degree of

functionalization (See experimental). The different chain lengths were confirmed by SEC

analysis. The small high molecular weight shoulder can be explained by chain coupling

reactions. The succesful labeling of PEtOx with both fluorophores is sustained by SEC-UV

detection (See experimental). Polymers without coumarin end groups were also obtained in

order to be used in the photophysical study

PEtOx-Mix 50 14.22 4.95 7.58 7.2

PEtOx-Mix 70 16.77 5.09 8.17 5.5

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Figure 51. a) first-order kinetic plots of monomer consumption in function of reaction time. b) Molecular weigth and polydispersity increase in in respect to conversion for the living polymerization determined by SEC.

Once the polymers were synthesized, fluorescent experiments were conducted to check if intramolecular energy transfer between the chain ends is taking place. Since PEtOx is water soluble, the compounds were also measured in an aqueous media. When the chromophores are both present on the same polymer chain a clear FRET is taking place. The presence of the signal corresponding to the coumarin emission and the decrease in of the Pyrene signal undoubtedly proves this fact in the spectrum (Fig. 24).

In order to better understand the process, the influence of chain length and the effect solvent on the FRET process was investigated (table 3). If we consider the effect of chain length, once can conclude that we have a decrease in FRET efficiency due to the higher distance between the chromophores located at the chain ends. The optimal DP in terms of FRET efficiency is DP 30 (Table 3). Another important parameter in FRET experiments is the solvent nature and pyrene is reported to be a very sensitive photoprobe in different environments.8 On going from CHCl3 to water we noticed a 6-fold increase of the emission intensity of the PEtOx DP30 labeled only with pyrene (Fig. 8).

Figure 52. Emisssion spectra for Py-CH2-PEtOx-Cou polymers in a) CHCl3 and b) H2O. . In all cases excitation wavelength was 345 nm and the pyrene concentration was 10µM.

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Table 13. Energy transfer parameters for dual labeled PEtOx mixed in CHCl3.


( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)




Py-CH2-PEtOx-Cou 30 H2O 8.51 6.05 4.02 92.1

Py-CH2-PEtOx-Cou 50 H2O 8.98 6.10 4.57 85.1

Py-CH2-PEtOx-Cou 70 H2O 10.16 6.23 5.87 58.9

This behavior can be explained in terms of solvent-solute interactions and has been investigated previously by Winnick et al. for the pyrene molecule.12 Normally going from an apolar to a polar solvent the molar extinction coefficient of the O-O band is dramatically increasing leading to higher fluorescence (increased intensity). Also the FRET efficiency is greatly improved in water reaching 92%. In this case the decrease in distance between the fluophores can probably be explained by the different polymer conformations in water as compared to CHCl3. Besides bringing the two dyes in proximity of one another, one of the main advantages of polymers is the procesability. In this case by dissolving certain amounts of polymer in chloroform and spin-coated upon quartz glass polymer films could be obtained very easily. Similar to solution the effect of chain length can be noticed in film.

Table 14. Energy transfer parameters for single and dual labeled PEtOx mixed in film.

Sample 𝑱 ∙ 𝟏𝟎𝟏𝟓

( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)

PEtOx30-Mix 1 to 1 22.39 5.16 2.40 99.5

PEtOx50-Mix 1 to 1 19.25 5.03 2.55 99.0

PEtOx70-Mix 1 to 1 16.09 4.88 2.70 96.7

Py-CH2-PEtOx-Cou 30 22.42 5.16 2.40 99.0

Py-CH2-PEtOx-Cou 50 23.51 5.20 2.63 98.4

Py-CH2-PEtOx-Cou 70 23.30 5.18 2.86 97.3

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The pyrene emission decreases with increasing chain length for the same reasons discussed in the previous section. In order to check the presence of FRET in films, we mixed the single labeled polymers in different ratios and then we spin-coated them always keeping constant the total molar concentration in solution.

Although in solution the FRET efficiency was low in film the situation changed (Fig. 9). When irradiated the polymers with 345 nm light (λmax of pyrene) we noticed a decrease of the pyrene intensity and the appearance of a signal corresponding to the emission of coumarin. In order to obtain high FRET signals small polymeric chains should be used or more dyes should be present upon the polymer chain. Where the excess of pyrene donor is present upon the same chain as the coumarin acceptor (Table 4). This is illustrated in figure 25 below where the orange line represents a polymeric film, consisting of a 9:1 ratio pyrene and coumarin moieties. If compared with the same mixture in solution no energy transfer was found but the intramolecular energy transfer is preferred. This effect is similar to an antenna, so we need more pyrene units to harvest light and then transfer it to the coumarin acceptor. This strategy will be persued in the next chapters. For the dual-labeled polymers we can observe that the FRET efficiency is very high which could be explained by the formation of Pyrene excimers at 460 nm. The overlap of the excimer peak with the coumarin emission could lead to misinterpreted results.

Figure 53. Emission spectra of PEtOx-labeled polymers in film a) DP30 b) DP50 c)DP70 d) dual-labeled PEtOx in film. All spectra were measured at 550V and with the excitation wavelength 345nm.

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Therefore, driven by the results obtained in film, we set up our strategy to synthesize poly-2-

oxazoline polymers that incorporate pyrene units in the chain and endcap them with a

coumarin unit at one end. Since intramolecular transfer is preferred in solution, a more

elegant way was persued to produce polymers with an efficient FRET. Preparation of the PyOx

monomer 13 started from the 1-pyrenebutyric acid. First the 1-pyrenebutyric acid 1 was

converted into an acid chloride (Figure 10) and subquently coupled with 2-chloroethylamine

hydrochloride and ring closed with KOH and 16-crown-8 ether.

Figure 54. Synthesis route to form (1-Pyrenylpropyl)-2-oxazoline (PyOx).

A statistical copolymer was prepared by combining PyOx with 2-ethyl-2-oxazoline, and finally end-capping the polymer with coumarin 343 (Fig. 11). By an excess of donor molecules present upon the polymer chain an efficient energy transfer was expected.

Figure 55. Chemical structure of pPyOx-PEtOx-Cou.

Figure 56. Emission spectra of pPyOx-PEtOx-Cou copolymers in a) CHCl3 and b) H2O.

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These are unstable excited molecules which are formed by the combination of two smaller molecules and rapidly dissociates with emission of radiation. The contribution of this peak to coumarin emission is small and it can be subtracted because we have synthesized also the PyOx copolymers without the coumarin end groups. The PyOx amount in the copolymer was calculated in such a way so as the final ratios between Pyrene and Coumarin to be 1:1, 3:1, 5:1 and 10:1 (see table 5). From Figure 30 b and Table 10 we can notice that the FRET efficiency is over 30% in all cases and comparable to the one formed in the case of dual-labeled PEtOx polymers with DP 30. The main advantage of this strategy is that we manage to obtain a higher degree of polymerization (i.e. DP = 60). So the availability of the chromophore in the side chain structure is higher than for the α,-ω- dual labeled polymers. The increasing number of donor units only brings a small contribution (60 intensity units pPyOx (1:1) as compared to pPyOx (10:1)) to the overall emission of the acceptor. Another important aspect is that for the copolymers with Pyrene side chain units we have excimer formative while for α,-ω- dye labeled ones no excimer signals could be detected. For the water soluble polymers we observed an increased FRET efficiency of the donor, but a lower fluorescence intensity of the acceptor (Fig. 12).

Table 15. Energy transfer parameters for pPyOx-PEtOx-Cou copolymers in CHCl3.


( 𝐌−𝟏 ∙ 𝐜𝐦−𝟏 ∙ 𝐧𝐦𝟒)

𝑹𝟎

(nm)

𝒓

(nm)


(%)







A more detailed study needs to be performed in order to better understand this behavior. One can speculate that as the pyrene fluorescence is increased in polar solvents due to solute-solvent interactions, the same thing should be valid for the Coumarin but in the opposite direction, polar solvents lower the fluorescence intensity.

In the glassy matrix the formation of pyrene excimers is obvious and increases with the PyOx content in the copolymer (Fig. 13). When coumarin is added at the chain end we noticed a shift of the emission peak dye to FRET. Still due to the presence of such high amounts of pyrene excimers the FRET efficient cannot be determined accurately from steady state measurements. The only valid case is the pPyOx-PEtOx-Cou (1 to 1) where we have a very low amount of excimers. In this case one can notice that FRET efficiency is close to 100%.

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In the search for an efficient energy transfer between the two dyes, intramolecular energy transfer was approached and appeared to be very efficient. This due to the higher probability of interaction between the two dyes when situated on the same chain, in contrast to when the dyes are on separate chains. To apply this knowledge on polymers, poly-(2-isopropenyl-2-oxazoline) 9 was used as scaffold for further modifications due to the accessible and quantitative reaction of the oxazoline cycle with the carboxylic acids (Fig. 14).13

Figure 58. Chemical structure of poly-(2-isopropenyl-2-oxazoline) modified with 1-pyrenebutyric acid and coumarin 343.

The SEC data proved that the Mn increases after the modification reaction but the dispersities remained constant, proving that no side reactions were present. The copolymers were obtained with good purity and high yields. The modified copolymers were characterized by 1H NMR, SEC and UV-Vis spectroscopy (See experimental). Due to the fact that we synthesized polymers with very low content of fluorophores and due to peak overlap, the degree of modification could not be determined directly by 1H NMR (see experimental). Therefore model compounds with the same molecular structure as the modified copolymers were made (Fig. 15), the modification degree could be calculated in a more exact manner from the absorption using the Beer–Lambert law.

Figure 57. Emission spectra of spincoated pPyOx-PEtOx-Cou copolymers irradiated at 345nm.

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Figure 59. Molecular structure of model compounds (2-propionamidoethyl 4-(pyren-2-yl)butanoate (11) and 2-propionamidoethyl 11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1-ij]quinoline-10-carboxylate) (12).

To investigate the effect of solvent also H2O was tested, but all the PIPRO modified polymers were insoluble in water except 5:1 ratio. This can be explained by the hydrophobic behavior of pyrene. For the fluorescence investigation we persued the two established routes, namely: i) first we made mixtures of single dye modified polymers and checked for intermolecular FRET; then ii) we studied the intramolecular fluorescence properties of the polymers modified with both dyes (Fig. 16). When we mixed the two copolymers very little FRET could be detected, based on the change in intensity of the pyrene (Fig. 16 a). The signal corresponding to Coumarin emission was greatly affected by the excimer emission of pyrene. Therefore we can conclude that no efficient intermolecular transfer can take place when we use this pair of fluorophores. The situation changes once we have the dyes in close proximity on the same polymeric chain. Although when we go to higher pyrene ratios we have also some emission, corresponding to pyrene excimers a clear peak in this case could be distinguished and attributed to coumarin emission at 470 nm. The highest FRET signal is registered for the polymer modified with 9.1% Pyrene and 2 % Coumarin. Further increasing the Coumarin content doesn’t improve the fluorescence of the acceptor. FRET efficiency could not be determined in the case of PIPRO modified polymers steady-state method due to the presence of excimers and the fact that the ratio between monomer and excimer is affected by the chemical composition of the copolymers. If for the single pyrene modified PIPRO (4.6%) polymer we have a 0.6 ratio of monomer/excimer. When we use 2% coumarin we can’t predict that the ratio will remain the same so we cannot use the decrease in the donor to assess the

Figure 60. a) intermolecular energy transfer for dual-modified PIPRO polymers. b) Emission spectra of PIPRO dual-modified polymers in CHCl3 irradiated at 345 nm.

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FRET efficiency. We can conclude that there is an optimal ratio between pyrene and coumarin in order to maximize the fluorescence of the acceptor.

In the film, the pyrene excimer formation is again present in the emission spectrum of the modified PIPRO copolymers, similar to the pPyOx-PEtOx-Cou polymers (Fig. 17). Again when coumarin is present on the same polymeric chain with the pyrene a decrease of the pyrene emission signal is noticed accompanied by the appearance of a new peak at 480 nm corresponding to coumarin emission. Still due to the presence of such high excimer signals no accurate FRET efficiency cannot be determined.

Conclusion

During this project, the synthesis and the photophysical properties of several modified polymers, based on poly-(2-oxazoline) scaffolds, with pyrene and coumarin units were investigated. The aim of this project was to obtain light harvesting polymers based on FRET. Therefore, three synthetic approaches were developed. The first one was the synthesis of poly-(2-ethyl-2-oxazoline) polymers with labeled with fluorescent end-groups. For the single labeled polymers, we used the “terminator method” and end capped the living chains with the 1-pyrenebutyric acid or coumarin 343. While for the α,-ω-labeled polymers we initiated the chains with 1-bromomethyl pyrene and end capped them with coumarin 343. In all cases the polymerization of EtOx was living and controlled leading to the target molecular weights and polymers with Ð lower than 1.15. The end capping reaction was very efficient allowing us to have a degree of functionalization of the end groups higher than 90%. The photophysical investigations lead to the conclusion that both fluorophores must be located on the same polymeric chain in order to have an efficient FRET transfer between fluorophores in solution. The second strategy was to synthesize a monomer containing pyrene units that could be copolymerized with EtOx by CROP and then used the “terminator method” to introduce the coumarin units. In this way, an antenna type polymer could be obtained. 2-(1-Pyrenylpropyl)-2-oxazoline monomer was successfully synthesized although with a rather low overall yield of 10 % (due to purifications). CROP copolymerization with EtOx followed by end capping reaction lead to the desired polymeric structures. Copolymers with different pyrene content were obtained with low dispersities and high degree of functionalization of the end groups. FRET efficiency in solution for these copolymers depended on the ratio between donor and acceptor groups. Namely if the ratio was closer to 1 than the FRET efficiency was higher. A

Figure 61. Emission spectra of spincoated modified PIPRO copolymers irradiated at 347nm.

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very interesting effect was discovered when we investigated photophysical properties of these polymers in water. The FRET efficiency increased dramatically from 47.7% to 80.3%. Water is considered a less good solvent, in which the polymer coil will be present in a collapsed state leading to closer proximity and thus higher FRET efficiencies. The third strategy was based on a polymer analogous reaction. Namely poly-(2-isopropenyl-2-oxazoline) synthesized by anionic polymerization was modified with the two fluorophores making use of the reactivity of the oxazoline cycle towards carboxylic acids. Consequently, copolymers with different donor/acceptor ratios could be easily obtained. Also in this case we noticed that FRET is present only when both fluorophores are located on the same chain. Quantitative determination of FRET was not possible for this polymers by steady state fluorescent measurements due to the formation of pyrene excimers that overlapped with the coumarin emission signal. In order to use the polymers for light harvesting properties in future work, the properties were also tested in film. Even though in solution the FRET efficiency of the single labeled polymers was rather low, in film the situation changed. If we spin coat a mixture of the two polymers (1:1 molar ratio) and obtain the film we notice that we have close to 100% FRET efficiency. For α,-ω-labeled PEtOx polymers we also have a higher EFRET than in solution, due to the reduced distance between donor and acceptor groups. For the PyOx and PIPRO copolymers we always have generation of pyrene excimers that affects the FRET and prevails us from accurately determining the FRET efficiency. As general conclusion, we can say that each type of polymeric architecture obtained during this project has his own advantages and disadvantages. If our applications are targeted for solution than PyOx and PIPRO copolymers are the right choice, but if we go towards solid state application than, α,-ω-labeled PEtOx polymers should be used. Future work will include the optimization of the composition of the copolymers and testing the synthesized polymeric structures in an organic solar cell. By this, various ways could be explored to optimize the output voltage and synthesize a more efficient solar cell.

Acknowledgments

I would like to thank my promotor Prof. dr. ir. Richard Hoogenboom for the privilege of working in his lab and my supervisor dr. Valentin- Victor Jerca for his advice concerning the processing of the dyes onto polymers. I would also like to thank Vincent of the UBCR group for the advice and the help concerning the fluorescence measurements.

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Supporting information

Materials

All the common used solvents were HPLC grade and include: dichloromethane (DCM, >99.8%, Sigma Aldrich), diethyl ether (>99.9%, Sigma Aldrich), acetonitrile (ACN, >99.9%, Sigma Aldrich), chloroform (CHCl3, >99%, Fisher Chemical) and N, N-dimethyl formamide (DMF, >99%, Biosolve). Dry solvents, like methanol (99.9%, Extra Dry, AcroSeal®) and dimethyl sulfoxide (99.7%) were purchased from Acros Organics. Dry DCM, TEA, THF and ACN were obtained from a custom made J.W. Meyer solvent purification system and were dried over aluminium oxide column.The following chemicals were used as received: 1-pyrenebutyric acid (>98.0%, TCI), and the coumarin 343 (>97.0%, TCI). 2-Ethyl-2-oxazoline (EtOx was kindly donated by Polymer Chemistry and purified by distillation over BaO. Poly-(2-isopropenyl-2-oxazoline) polymers were synthesized by Dr. Adriana Jerca by anionic polymerization of 2-isopropenyl-2-oxazoline (Mn = 18500, Đ= 1.21). Br-Me-Pyrene (>92.0%, Sigma Aldrich) was recrystallized from CHCl3 (3 times). Equipment NMR Proton and carbon-13 nuclear magnetic resonance spectra were recorded on a Bruker Avance 300 MHz at room temperature. NMR spectra were measured in chloroform-d (CDCl3) from Euriso-top. The chemical shifts are given in parts per million (δ), relative to CDCl3 at 77.36 ppm. Gas chromatography (GC) was performed on a GC8000 from CE instruments with a DB-5MS column (60 m x 0.249 mm x 0.25 µm) from J&W scientific. Injections are performed with a CTC A200S auto sampler and detection is done with a flame ionization detector (FID) which burned a H2 /air mixture. The carrier gas (H2) is pushed through the column with a pressure of 100 bar. Injector and detector are kept at a constant temperature of 300 °C. Size exclusion Chromatography (SEC) Size-exclusion chromatography (SEC) was performed on a Agilent 1260-series HPLC system equipped with a 1260 online degasser, a 1260 ISO-pump, a 1260 automatic liquid sampler (ALS), a thermostatted column compartment (TCC) at 50°C equipped with two PLgel 5 μm mixed-D columns and a mixed-D guard column in series, a 1260 diode array detector (DAD) and a 1260 refractive index detector (RID). The used eluent was DMA containing 50 mM of LiCl at a flow rate of 0.500 mL/min. The spectra were analyzed using the Agilent Chemstation software with the GPC add on. Molar mass and dispersity values were calculated against PMMA standards from PSS. UV/VIS spectrometer UV/Vis spectra are recorded on a Varian Cary 100 Bio UVVIS spectrophotometer equipped with a Cary temperature and stir control. Fluorescence spectrometer The emission and excitation spectra are recorded on a Varian Cary Eclipse fluoro-spectrophotometer also equipped with a Cary temperature and stir control. The excitation and emission wavelength were varied according to the sample absorption maximum, same was done for the detectors voltage, so the peaks did not go out of scale. The slit width of the excitation and emission were kept at 5 nm during the measurements. Samples were measured in quartz cuvettes with a fluorescence spectrometer pathlength of 1.0 nm in the wavelength range of 200-700 nm. Microwave, all the microwave polymerizations were performed in a Biotage initiator Microwave System with a temperature range of 40-250°C equipped with a variable magnetic stirrer (300-900 RPM). Chromatographic columns on aluminum oxide and silica were performed on Merck Alox 90 standard aluminum oxide and on Davisil chromatographic silica mecia LC60A 70-200 micron respectively. Silica TLC’s were taken on pre-coated Macherey-Nagel ALUGRAM SIL G/UV254 plates. Aluminium oxide TLC’s were taken on Merck TLC Aluminum oxide 60 F254 neutral.

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Flash chromatography was performed on a Grace Reveleris® flash chromatography system using silica Reveleris flash cartridges. Glove box The system used is a VIGOR Sci-Lab SG 1200/750 Glovebox System, the purity level obtained is at most, < 1 ppm O2+ H2O. Deionized water was prepared with a resistivity less than 18.2 MΩ x cm using an Arium 611 from Sartorius with the Sartopore 2 150 (0.45 + 0.2 µm poresize) cartridge filter. Spin Coater All films were made with 30 seconds of spinning at 500 RPM/sec up to 2000 RPM. Before spin-coating the solutions were filtered and the quartz plates were extensively cleaned with acetone and blown dry with nitrogen gas. Synthesis

Organic compounds

Synthesis of (2-propionamidoethyl 4-(pyren-2-yl) butanoate (Pyrene-EtOx)

0.16 g (0.55 mmoles) of Pyrene-COOH was dissolved in 1 mL of DMF, then 0.05g (0.50 mmoles) of 2-ethyl-2-oxazoline was added under continuous stirring. Then the vial was capped and the mixture was heated to 100°C for 5 hours. The solution was precipitated in crushed ice, and the white precipitate was filtered off and washed three times with water on the filter. The precipitate was dried overnight in the vacuum oven at 50°C. Further purification was performed by column chromatography (SiO2) eluent: hexane/EtOAc (30/70). Yield: 80%; UV-vis: λmax CHCl3: 345 nm. 1H NMR (300 MHz, CDCl3) δ 8.32 – 7.68 (m, 9H; Pyrene), 5.58 (s, 1H; NH), 4.14 – 3.97 (t, 2H,O-CH2-CH2), 3.41 (dd, J = 10.8, 5.6 Hz, 2H, CH2- CH2-NH), 3.36 – 3.28 (t, 2H, Pyrene- CH2- CH2), 2.38 (t, J = 7.3 Hz, 2H,O=C- CH2-CH3), 2.21 – 1.96 (m, 4H, CH2- CH2- CH2-C=O), 1.02 (t, J = 7.6 Hz, 3H, CH2-CH3).

2 Synthesis of (2-propionamidoethyl 4-(pyren-2-yl)butanoate and 2-propionamidoethyl 11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1-ij]quinoline-10-carboxylate) (Cou-EtOx)

0.1g (0.35 mmoles) of Pyrene-COOH was dissolved in 1 mL of DMF, than 1.96 (20 mmoles) of 2-ethyl-2-oxazoline was added under continuous stirring. Then the vial was capped and the mixture was heated to 140°C for 15 minutes, at this point the reaction was completed. Then the solution was precipitated in crushed ice, and the dark orange precipitate was filtered off and washed three times with water on the filter. The precipitate was dried overnight in the vacuum oven at 50°C. Further purification was performed by column chromatography (SiO2) eluent: hexane/acetone (80/20). Yield: 50%; UV-vis: λmax CHCl3: 440 nm.

1H NMR (400 MHz, CDCl3) δ 8.25 (s, 1H, C-CH=C-C=O), 6.89 (s, 1H, C-CH-C), 6.54 (s, 1H, NH), 4.31 – 4.27 (t, 2H, O-CH2-CH2), 3.58 (q, J = 9.9, 5.0 Hz, 2H, CH2-N-CH2), 3.28 (dd, J = 11.4, 5.6 Hz, 4H, CH2-CH2-NH), 2.81 (t, J = 6.4 Hz, 2H, CH2-CH2-C), 2.73 – 2.66 (m, 2H, CH2- CH2-C), 2.20 (q, J = 7.6 Hz, 2H, CH2-CH2-CH2), 1.96 – 1.85 (m, 2H, O=C-CH2-CH3), 1.14 – 1.07 (m, 3H,CH2- CH3)

Synthesis of 1-pyrenebutyric acid chloride

5g (0.017 moles) of 1-pyrenebutyric acid was weight in a round bottom flask. Then 25 mL of dry DCM were added under argon the mixture was cooled in an ice bath. Then 3.09 (0.026 moles) thionyl chloride was added dropwise. The mixture was left to stirr at room temperature

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overnight. The DCM was evaporated under reduced pressure and then was three times redissolved in DCM and evaporated again in order to remove the excess of SOCl2, until a yellow solid was obtained. Yield: 99%.

1H NMR (300 MHz, CDCl3) δ 8.22 – 7.70 (m, 9H, Pyrene), 3.40 – 3.27 (t, 2H, Py-CH2-CH2), 2.93 (t, J = 7.1 Hz, 2H, CH2-CH2-CH2), 2.19 (dt, J = 14.6, 7.2 Hz, 2H, CH2-CH2-C=O).

Synthesis of N-(2-chloroethyl)-4-(pyren-1-yl)butanamide

4.95 g (0.016 moles) of 1-pyrenebutyric acid chloride and 1.57 g (0.020 moles) of 2-chloroethylamine hydrochloride was suspended in dry DCM (50 mL) in an argon atmosphere and the reaction mixture was cooled to 0°C. 4.898 g (0.048 moles) of TEA was added dropwise, and the reaction mixture was stirred overnight. Water was added to the reaction and the aqeous phase was extraced with DCM (2x 15 mL). The combined organic phases were washed with two times 9% HCl solution, then with saturate sodium bicarbonate, water and brine. Finally the organic phase was dried over MgSO4 and the solvent was removed under reduced pressure. The compound was further purified by flash chromatography on silicagel eluting with a hexane/ethyl acetate mixture (50/50). Yield: 20%

1H NMR (300 MHz, CDCl3) δ 8.29 – 7.63 (m, 9H, pyrene), 5.71 (s, 1H, O=C-NH-CH2), 3.62 – 3.39 (m, 4H, Cl-CH2-CH2-NH), 3.27 (t, J = 15.3, 8.3 Hz, 2H, O=C-CH2-CH2), 2.28 – 2.01 (m, 4H, Pyrene-CH2-CH2-CH2).

Synthesis of 2-(3-(pyren-1-yl)propyl)-4,5-dihydrooxazole

1.17 g ( 0.004 moles) N-(2-chloroethyl)-4-(3, 6-dihydropyren-1-yl) butanamide was dissolved in 50 mL THF and 0.047 g (0.000178 moles) 18-crown-6-ether was added under argon atmosphere. Then 0.57 g (0.010 moles) KOH was added under vigorous stirring. The reaction was left to stir overnight. The THF was removed under reduced pressure followed by addition of DCM (50 mL) and washing with water (3x15mL). The water phase was extracted with DCM three times. The combined organic phases were washed with brine and dried over MgSO4. The solvent was removed yielding a yellowish crude product. The compound was further purified by flash chromatography on silica gel eluting with a hexane/EtOAc mixture (70/30). Yield: 51%

1H NMR (400 MHz, CDCl3) δ 8.26 – 7.73 (m, 9H, Pyrene), 4.21 – 4.05 (t, 2H, O-CH2-CH2), 3.75 (t, J = 9.4 Hz, 2H, N-CH2-CH2), 3.42 – 3.26 (t, 2H, CNO-CH2-CH2), 2.43 – 2.28 (m, 2H,Pyrene-CH2-CH2 ), 2.23 – 2.04 (t, 2H, CH2-CH2-CH2).

Polymer synthesis

Microwave –assisted synthesis of single-labeled poly-(2-ethyl-2-oxazoline)s

To study the effect of chain length, PEtOx with three different polymerization degree (DP= 30, 50 and 70) were synthesized. All polymers were synthesized under similar conditions. The monomer concentration was 4 M for all polymerizations. The monomer/initiator [M]/ [I] ratio was 30/1, 50/1 and 70/1, respectively. Reactions were carried out in capped microwave vials. The vials were dried in the oven at 200°C and cooled under argon to room temperature before use. The solutions were prepared in a glove box (Vigor Sci-Lab SG 1200/750) under argon. The polymerizations mixtures were heated to 140°C under microwave irradiation for 216 s ([M]/ [I] = 30), 6 min ([M]/ [I] = 50) and 8 min ([M]/ [I] = 70) to reach almost full conversion. For the end-capping reactions, the vial was cooled down to room temperature and a solution of the

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corresponding acid in DMF 1.5-fold excess was added via a syringe through the septum of the capped microwave vial containing the living oligomers (EtOx). Thereafter TEA was added similarly in a two-fold excess. The given amounts were varied according to the used monomer to initiator ratios. The reaction solution was heated to 100°C for 20 hours. After cooling to room temperature the reaction mixture was diluted with chloroform and the solution was washed three times with saturated aqueous sodium hydrogen carbonate. Then the solution was washed with brine, dried over MgSO4 and filtered. The solvent was evaporated under reduced pressure resulting a foamy polymer. The polymers were dissolved again in 3mL chloroform and precipitated in ice cold diethyl ether. The solutions were centrifuged for 10 minutes (7500 rpm) at 5°C. The solution was cast off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under reduced pressure resulting a white or yellow foam for the polymers modified with 1-pyrenebutyric acid or coumarin 343, respectively. Finally the polymers were dried overnight in the vacuum oven at 50°C. 1H NMR and SEC analysis confirmed the initial DP.

1H NMR (300 MHz, CDCl3) δ 8.30 – 7.68 ( 9H, Pyrene), 4.08 (s, 2H, CH2-c=O-O), 3.38 (4H, N-CH2-CH2), 2.95 (2H, O-O=C-CH2), 2.89 (2H, CH3-N-CH2), 2.26 (2H, CH3-CH2-C=O and CH2-CH2-Py), 1.05 (3H, CH3-CH2-C=O).

Microwave assisted of dual-labeled poly-(2-ethyl-2-oxazoline)s

All polymers were synthesized under similar conditions. The monomer concentration was 4 M for all polymerizations. The monomer/initiator [M]/ [I] ratio was 30/1, 50/1 and 70/1, respectively. Reactions were carried out in capped microwave vials. The vials were dried in the oven at 200°C and cooled under argon to room temperature before use. The solutions were prepared in a glove box (Vigor Sci-Lab SG 1200/750) under argon. The polymerizations mixtures were heated to 140°C under microwave irradiation for 216 s ([M]/ [I] = 30), 6 min ([M]/ [I] = 50) and 8 min ([M]/ [I] = 70) to reach almost full conversion. In this case the initiator used was 1-(Bromomethyl)pyrene.

For the end-capping reactions, the vial was cooled down to room temperature and a solution of the corresponding acid in DMF 1.5-fold excess was added via a syringe through the septum of the capped microwave vial containing the living oligomers (EtOx). Thereafter TEA was added similarly in a two-fold excess. The given amounts were varied according to the used monomer to initiator ratios. The reaction solution was heated to 100°C for 20 hours. After cooling to room temperature the reaction mixture was diluted with chloroform and the solution was washed three times with saturated aqueous sodium hydrogen carbonate. Then the solution was washed with brine, dried over MgSO4 and filtered. The solvent was evaporated under reduced pressure resulting in foamy polymers.

The polymers were dissolved again in 3mL chloroform and precipitated in ice cold diethyl ether. The solutions were centrifuged form 10 minutes (7500 rpm) at 5°C. The solution was cast off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under reduced pressure resulting in a yellow foamy polymer. Finally the polymers were dried overnight in the vacuum oven at 50°C.

1H NMR (300 MHz, CDCl3) δ 8.36 – 7.87 (9H, Pyrene), 6.86 (1H, C-H-C), 5.27 (2H, pyrene-CH2), 4.33 (2H, CH2-CH2-O), 3.29 (4H, NH-CH2-CH2), 2.75 (4H, CH2-N-CH2), 2.26 (4H, C-O=C-CH2-CH3 and C-CH2-CH2), 1.15 – 0.88 (3H, CH2-CH3).

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Synthesis of dye functionalized poly-(2-isopropenyl-2-oxazoline) copolymers

The functionalized copolymers were obtained by oxazoline ring-opening addition in the presence of the fluorescent dyes. In a typical run, 1.8 mL of dried DMF were added to 0.025 g of the corresponding dye and 0.1 g (0.0009 moles) PIPRO in a vial. The mixture was heated at 140°C with stirring for 5 hours. The reaction mixture was diluted with 1 ml of CHCl3 and precipitated in cold diethyl ether. The solution was centrifuged for 10 min at 7500 rpm. The product was purified twice by reprecipitation from chloroform in diethyl ether and dried at 50°C overnight under vacuum.

1H NMR (300 MHz, CDCl3) δ 8.02 (9H, Pyrene), 6.06 (1H, NH), 4.08 (4H, O-CH2 and O=O-CH2), 3.65 (4H, N-CH2 + NH-CH2), 3.32 (Pyrene-CH2), 2.42 (2H, CH2 -CH2-CH2), 1.75 (2H, O=C-CH2-CH2), 1.25 – 0.84 (3H, CH2-CH3).

Microwave assisted copolymerization of 2-(1-pyrenylpropyl)-2-oxazoline and EtOx

The polymerization mixtures for the microwave assisted synthesis of a series of statistical EtOx-PyOx copolymers were prepared in the glove box. In each microwave vial EtOx, PyOx and ACN was added resulting in a 4 M total monomer concentration. The total monomer to initiator ratio was 60. The amount of PyOx used was 1.5; 5.5; 8.5 and 16.5 %mol to obtain a series with compositions similar to the PIPRO modified copolymers. The vials were capped and were subsequently heated to 140°C for 12 min to reach full conversion.

For the end-capping reactions, the vial was cooled down to room temperature and a solution of the corresponding acid in DMF 1.5-fold excess was added via a syringe through the septum of the capped microwave vial containing the living oligomers (EtOx). Thereafter TEA was added similarly in a two-fold excess. The given amounts were varied according to the used monomer to initiator ratios. The reaction solution was heated to 100°C for 20 hours. After cooling to room temperature the reaction mixture was diluted with chloroform and the solution was washed three times with saturated aqueous sodium hydrogen carbonate. Then the solution was washed with brine, dried over MgSO4 and filtered. The solvent was evaporated under reduced pressure resulting in foamy polymers.

The polymers were dissolved again in 3mL chloroform and precipitated in ice cold diethyl ether. The solutions were centrifuged form 10 minutes (7500 rpm) at 5°C. The solution was cast off and the resulting precipitate was dissolved in 5 mL of DCM and was evaporated under reduced pressure resulting in white or yellow foam for the polymers modified with 1-pyrenebutyric acid and coumarin 343, respectively. Finally the polymers were dried overnight in the vacuum oven at 50°C.

1H NMR (300 MHz, CDCl3) δ 8.41 – 7.57 (9H, pyrene), 3.35 (6H, N-CH2-CH2 + Pyrene-CH2 ), 2.30 (s, 4H), 1.02 (3H, CH2-CH3).

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The Förster theory shows that FRET efficiency (EFRET) varies as the sixth power of the distance between two molecules (equation 1). To calculate the FRET efficiency, one has to determine the spectral overlap integral 𝐽 and the interchromophoric distance 𝑟.

𝐸𝐹𝑅𝐸𝑇 =

1

1 + (𝑟

𝑅0)

6 (7)

Where 𝑅0 is the distance between the two molecules at 50% FRET efficiency and 𝑟 being the interchromophoric distance for the experiment. 𝑅0 can be calculated using equation 2.

𝑅0 = 0.02108 (𝜅2 𝜙𝐷 𝑛−4 𝐽)16 (8)

Where 𝜙𝐷is the fluorescence quantum yield of the donor in the absence of the acceptor, 𝜅2 is the orientation factor of transition dipole moment between donor and acceptor;𝜅2 can take

values between 0 and 4. In the present case we have considered 𝜅² =2

3 (when both dyes are

freely rotating and can be considered to isotropically oriented during the excited state lifetime), 𝑛 is the refractive index of the medium, and 𝐽 is the spectral overlap integral calculated as:

𝐽 = ∫ 𝜀𝐴𝑐𝑐𝑒𝑝𝑡𝑜𝑟(𝜆)𝜆4𝐼𝐷(𝜆) 𝑑𝜆 (9)

Where 𝜀𝐴𝑐𝑐𝑒𝑝𝑡𝑜𝑟(𝜆) is the acceptor molar extinction coefficient at the wavelength 𝜆, 𝜆 is the

wavelength and 𝐼𝐷(𝜆) is the normalized emission spectrum of the donor. By integrating this over the entire spectrum one can calculate the 𝐽 value, the unit of 𝐽(𝜆) is 𝑀−1 ∙ 𝑐𝑚−1 ∙ 𝑛𝑚4. The refractive index of CHCl3, geometry factor and the quantum yield of the 1-pyrenebutyric acid were used from literature.6 The efficiency of FRET can be determined by steady-state measurements and is expressed as equation (4). From this 𝑟 can be calculated as follows:

𝐸𝐹𝑅𝐸𝑇 = 1 −𝐹𝐷𝐴

𝐹𝐷 (10)

Where 𝐹𝐷𝐴 and 𝐹𝐷 are the donor fluorescence intensities with and without an acceptor respectively. The FRET efficiency was determined by donor quenching method.

The actual distance 𝑟 between donor and acceptor is given by:

𝑟 =

𝑅0

(1

𝐸𝐹𝑅𝐸𝑇− 1)

6

(11)

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SEC Analysis

Table 16. Structural characterization of single dye-labeled PEtOx.

Table 17. Structural characterization of the dual-labeled PEtOx.

Table 18. Structural characterization of the pPyOx-PEtOx-Cou copolymers.

pPyOx - PEtOx-Cou %mol PyOx feed

%mol PyOx copolymera

Py/Coub Yield (%)

Mn (Da)c

Đc

10 16.5 19.8 10:1 92 9700 1.11

5 8.5 9.5 5:1 82 10600 1.11

3 5.5 7.2 3:1 78 8500 1.10

1 1.5 2.0 1:1 76 10800 1.10

a %mol PyOx in the copolymer determined by 1H NMR. b Ratio of pyrene to coumarin in the copolymer determined by 1H NMR. c Determined from DMA SEC using PMMA calibration.

Polymer code Yield (%) DP a f (%) b Mn (Da)c Đc

PEtOx-Py 97.0 30 84 5900 1.09

PEtOx-Py 80.2 50 85 10900 1.09

PEtOx-Py 80.1 70 94 12600 1.09

PEtOx-Cou 81.0 30 98 5900 1.08

PEtOx-Cou 94.4 50 91 10200 1.08

PEtOx-Cou 96.4 70 99 13100 1.10

a Theoretical degree of polymerization. b Functionality calculated from the aromatic signals of the dye and the methyl signal of the polymer side chain from the 1H NMR. c Determined from DMA SEC using PMMA calibration.

Py-CH2-PEtOx-

Cou

Yield (%) DPa f (%)b Mn (Da)c Đc

DP 30 95 28 92 6000 1.09

DP 50 92 49 91 9500 1.09

DP 70 89 68 90 13600 1.12

a Real DP calculated from the aromatic signals of the pyrene and the methyl signal of the polymer side chain from the 1H NMR. b Functionality, calculated from the aromatic signals of the coumarin dye and the methyl signal. c Determined from DMA SEC using PMMA calibration.

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Table 19. Structural characterization of the modified PIPRO polymers

Polymer code Yield (%) % mol Pya % mol Coua Mn (Da)b Đb

PIPRO-Py 10 95 8.9 0 20300 1.23

PIPRO-Py 5 92 3.8 0 19479 1.22

PIPRO-Cou 10 90 0 7.5 22000 1.24

PIPRO-Cou 3 91 0 1.2 19600 1.27

PIPRO-Cou 1 93 0 0.5 18900 1.21

PIPRO-Py-Cou 10:10 89 9.6 7.4 21900 1.36

PIPRO-Py-Cou 10:3 88 9.1 2.0 19900 1.25

PIPRO-Py-Cou 10:1 91 9.5 0.3 18900 1.28

PIPRO-Py-Cou 5:1 87 4.7 0.9 18800 1.23

a Calculated with the molar extinction coefficient of the model compounds using UV-vis spectroscopy. b Obtained by DMA-SEC analysis using PMMA calibration.

Molar absorption coefficients

Figure 62. Linear correlation of absorbance with concentration for a) Pyrene-Etox and b) Coumarin-EtOx.

𝜺𝑪𝒐𝒖−𝑬𝒕𝑶𝒙 = 𝟒𝟏𝟕𝟎𝟓 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

R² = 0.997

𝜺𝑷𝒚−𝑬𝒕𝑶𝒙 = 𝟑𝟖𝟐𝟏𝟏 𝑳 ∙ 𝒎𝒐𝒍−𝟏 ∙ 𝒄𝒎−𝟏

R² = 0.998

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