Master’s Thesis
To confer the academic degree of
Diplom-Ingenieur
In the Master’s Program
Management in Chemical Technologies
Author
Patrick Breiteneder, BSc Submission
Institut für Chemie der Polymere Thesis Supervisor
Assoz. Univ.-Prof. Dr. Ian Teasdale Assistant Thesis Supervisor
Mgr. Paul Strasser, MSc May 2021
Bifunctional BODIPYs for green-light
photolabile
macromolecular applications
JOHANNES KEPLER UNIVERSITÄT LINZ Altenberger Straße 69 4040 Linz, Austria jku.at
Statutory Declaration
I hereby declare that the thesis submitted is my own unaided work, that I have not used other than the sources indicated, and that all direct and indirect sources are acknowledged as references.
This printed thesis is identical with the electronic version submitted.
Place, Date
Signature
04.05.2021
Acknowledgement
I want to address my gratitude to the Institut für Chemie der Polymere. The positive working atmosphere allowed me to not only work efficiently but also to enjoy my time
I want to address my special thanks to Paul Strasser who always supported me technically and also on a personal level. Additionally, I want to thank Ian Teasdale for his competent advice in order to solve the numerous problems which came up during my work.
Abstract
The interest in stimuli responsive polymers has increased significantly over the last few years. The search for non-invasive techniques of disease treatment and structural manipulation of macromolecular systems is going on until this day.
Especially light sensitive polymeric materials show promising results in the area of precise drug release in high local concentration. These materials have implemented photolabile groups referred to as photocages. Most available photocages nowadays are restricted to absorption in the UV range, which limits their penetration depth in materials and hinders application in biomedical settings.
Unlike other commonly available photocages, boron dipyrromethenes (BODIPYs) not only offer excellent extinction coefficients but also an absorption range in the visible light spectrum. However, their use in macromolecular applications in sense of a photo-labile group remains scarce. The work presented in this master thesis focuses on the synthesis of a bi-functionalised BODIPY compound and its polymeric implementation in order to produce green-light photo-degradable oligomers and hydrogels. The synthesised BODIPY compounds were fully characterized via nuclear magnetic resonance spectroscopy (NMR), mass spectroscopy (MS) and UV-Vis measurements and their incorporation and degradation in oligomers verified via gel permeation chromatography (GPC) and UV-Vis. Finally, the synthesis of BODIPY containing hydrogels could be proven via absorption and emission measurements and their wavelength dependent degradation shown at both 365 nm and 510 nm irradiation.
Zusammenfassung
Das Interesse an stimulus-sensitiven Polymeren hat sich während den letzten Jahren drastisch erhöht. Die Suche nach nichtinvasiven Methoden für Krankheitsbehandlungen und die strukturelle Abänderung von makromolekularen Systemen ist bis heute fortwährend. Lichtsensitive Polymere im speziellen zeigen vielversprechende Resultate im Bereich der gezielten Wirkstofffreisetzung. Solche Materialien haben lichtsensitive Moleküle inkorporiert, welche unter Lichteinstrahlung abgebaut werden können. Diese Materialien werden als
„Photocages“ bezeichnet. Die meisten der derzeit zugänglichen „Photocages“
absorbieren Licht im UV-Bereich was deren Anwendungsbereiche limitiert, da UV- Licht nur eine geringe Eindringtiefe hat und zu möglichen Schäden bei medizinischen Anwendungen führen kann. Anders als andere lichtsensible Substanzen haben BODIPYs nicht nur exzellente Extinktionskoeffizienten, sondern haben auch Absorptionsbereiche im sichtbaren Lichtspektrum. Trotz all dessen bleibt deren Verwendung für lichtsensible Makromoleküle bisher rar. Diese Arbeit beschäftigt sich mit der Bi-Funktionalisierung von BODIPYs und deren Implementierung in Polymerketten, um in grünem Licht abbaubare Oligomere und Hydrogele zu produzieren. Die hergestellten BODIPYs wurden mittels Kernspinresonanzspektroskopie, Massenspektrometrie sowie durch Absorptions- und Emmissionsspektroskopie charakterisiert. Die Inkorporation und der Oligomerabbau konnte mittels Gelpermeationschromatographie sowie mittels UV- Vis Messungen verifiziert werden. Abschließend konnte die Synthese der BODIPY beinhaltenden Hydrogele sowie deren wellenlängenabhängiges Abbauverhalten via Absorption und Emissionsmessungen gezeigt werden.
Index
Abstract: ... VI
1. Introduction: ... 1
1.1 Stimuli responsive polymers ... 1
1.2 Photocages ... 3
1.3 BODIPY ... 6
2. Materials and methods ... 13
3. Experimental procedure ... 14
3.1 Compound 1 (BODIPY) synthesis ... 14
3.2 Hydrolysis of compound 1 (BODIPY) ... 15
3.3 Hydroxylation of compound 1 (BODIPY) ... 16
3.4 Compound 4 (Diol-BODIPY) synthesis ... 17
3.4.1 Hydroxylation of compound 2 (Hydrolysed-BODIPY) ... 17
3.4.2 Hydrolysis of compound 3 (Hydroxy-BODIPY) ... 18
3.5 Compound 4 (Diol-BODIPY) one-pot synthesis ... 19
3.6 Compound 5 (Dibenzylisocyanate-Diol) synthesis ... 20
3.7 Oligomer synthesis ... 21
3.7.1 Diol-ED-2003-Oligomer 1 (1:1) ... 21
3.7.2 Diol-ED-2003-Oligomer 2.1 (1:3) ... 21
3.7.3 Diol-ED-2003-Oligomer 2.2 (1:3) ... 22
3.7.4 Diol-ED-2003-Oligomer 3.1 (1:15) ... 22
3.7.5 Diol-ED-2003-Oligomer 3.2 (1:15) ... 23
3.8 Hydrogel synthesis ... 24
3.8.1 Gel experiment 1 (Diol:Jeffamine = 1:2) ... 24
3.8.2 Gel experiment 2 (Diol:Jeffamine = 1:28) ... 25
3.8.3 Gel experiment 3 (Diol:Jeffamine = 1:140) ... 25
3.8.4 Gel experiment 4 (no BODIPY) ... 26
4. Results and discussion ... 27
4.1 Optical product characteristics ... 27
4.2 Compound 1 (BODIPY) ... 28
4.3 Compound 2 (Hydrolysed-BODIPY) ... 32
4.4 Compound 3 (Hydroxy-BODIPY) ... 35
4.5 Compound 4 (Diol-BODIPY) ... 38
4.6 Compound 5 (Dibenzylisocyanate-BODIPY) ... 41
4.7 Absorption measurement comparison ... 45
4.8 Emission spectra comparison ... 47
4.9 Oligomer characterisation and degradation studies ... 48
4.9.1 UV-Vis ... 48
4.9.2 GPC-measurements ... 55
4.10 Gel synthesis and degradation ... 60
4.10.1 Degradation studies under 510 nm irradiation ... 60
4.10.2 Degradation studies under 365 nm irradiation ... 63
4.10.3 Degradation studies of a control gel ... 65
4.10.4 Kinetic studies of the hydrogel degradation ... 66
5. Outlook and conclusion ... 70
6. References ... 73
7. Supporting information ... 83
1
1. Introduction:
1.1 Stimuli responsive polymers
The development of smart materials and polymers has grown special interest in current biomedical research. Smart polymeric materials are referred to as compounds that are stable at ambient conditions but are responsive to stimuli such as pH or light 1. Especially light sensitive polymers have been developed in order to rely on light as the stimuli, for example, for an effective drug release. These delivery systems allow a selective transport and release at the target sites.
Compared to other stimuli, it enables a spatiotemporal release of the active compounds in a high local concentration. These polymeric materials have implemented photosensitive groups which can result in different structural changes upon irradiation as depicted in Figure 1 1. Depending on the composition the irradiation either leads to (a) complete polymer degradation, (b) degradation of a linker releasing smaller molecules or to (c) changes in polymer polarity 1, of which (a) was further investigated in this work in form of oligomers and hydrogels.
2 Figure 1: Examples of the possible polymer degradation caused by light-stimulus 1.
Current polymeric hydrogel structures including photocleavable linkers often rely on structures only susceptible to UV-irradiation 2. To overcome these problems, green-light sensitive compounds were developed in this thesis.
3
1.2 Photocages
Figure 2: Schematic drawing of the principle of a photocleavage reaction.
Photocages are light-sensitive structures which are susceptible to irradiation with a specific wavelength. A schematic depiction of a photodegradation is visible in Figure 2. Photocages are readily used in a biological context allowing a direct release of bound compounds. The drug can therefore be administered in its inactive form and can be precisely activated upon a light stimulus at a specific place. This mechanism is frequently used to selectively release drugs or proteins 3,4. Another possibility is the loading of the active compound into a hydrogel and its release upon degradation. The photocleavage process in general opens new possibilities in the bio medicinal area, as it is a non-invasive and precise method of drug administration 5.
One of the most readily used photocages are ortho-nitrobenzyl or coumarin compounds which require UV-light for the cleavage process 6. Ortho-nitrobenzyl photocages are used because of their simple structure and easy availability as well as their fully investigated and understood photocleavage reactions 6,7. Coumarin photocages have established themselves as photoactivatable phosphate releasing groups. A special characteristic of coumarin derivatives is that it can undergo a reversible dimerization depending on the irradiated wavelength 1. Some applications may require additional chemicals like sensitizers to activate the photocages in order to make use of their photo cleavable properties. These sensitizers initially need to activate protecting groups through electron transfer processes, resulting in a deprotection of the photocage 8,9. In Figure 3 the
BODIPY
R1 R2 R1 R2
4 structures of some o-nitrobenzyl and coumarin derivatives can be seen alongside their respective photocleavage reaction.
Figure 3: Photocleavage of a o-nitrobenzyl compound (top), and a photocleavage of a coumarin compound (bottom) 7.
Because these classes of photocages require UV- light as stimulus, their use in a biological context is limited.
To allow for an efficient application of photocages it is therefore thought after to achieve absorption in the so-called biological window around 650-850 nm to increase efficiency. In Figure 4 the absorption spectra of the different components of some biological tissues and liquids can be seen 10.
O O
NO2
CHO NO2
+ HO hv O
O O O P O OH
OH hv
Nu O O
O O
Nu
+ HO P OH O OH
5 Figure 4: Absorbance spectra of different biological substances and the depiction of the biological
window 10.
In addition to the low tissue penetration, UV-irradiation shows possible harmful side-reactions 11. It can induce oxidative stress and inflammatory skin reactions and depending on the irradiation time and intensity this can result in DNA-damage and apoptosis 12.
Initiation of the cleavage process by visible light is therefore especially of interest because of its possible uses in biomedicine 13. Different structures like GABA-Ruthenium complexes have been developed enabling the use of visible light degradation 14. However, because these ruthenium complexes have a very wide absorption spectrum it can hinder fluorescence observations 8. The perhaps biggest potential is predicted to lie in 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) compounds. Since their development they have greatly increased in their use 15,16, with applications in the field of photochemical degradation and dyes for biological uses 17.
6
1.3 BODIPY
The unique properties of BODIPYs arise in their photo-stability and high molar absorption coefficients. Unlike most other commonly available photocages, BODIPYs have their absorption range in the visible light spectrum. The exact wavelength can be fine-tuned with the derivatisation of the chromophore which can be of special interest for uses in chemical sensors or fluorescent organic devices 18. Meso-methyl BODIPYs in particular are used in particular as photocages as they are able to release active organic compounds upon irradiation with light of a suitable wavelength 13. Especially the high emission quantum yields of meso- methyl derived BODIPY structures and the possible usage for photo-release reactions are of interest. The high extinction coefficients and narrow absorption bands raise additional interest 19,20.
In Figure 5 the photocleavage process in the meso-position of such a BODIPY compound and the release of the active substrate can be seen.
Figure 5: Schematic drawing of the photo- release process of BODIPY and an associated substrate.
Further advantages of BODIPY compounds compared to many other photocages are their biological compatibility as well as their sterically compact structure 21. Because BODIPYs show a low sensitivity towards pH changes and are stable under physiological conditions 22, applications in the field of tissue regeneration and drug delivery would be possible 23,24.
7 Some basic BODIPY structures can be seen in Figure 6. The simplest structure on the top left is not reported in literature and predicted to be unstable, as the system is not sterically protected. In order to reduce the risk for any side reactions of BODIPY-compounds it is important that the framework is substituted 25. The simplest synthesised structure on the top right is often viewed as reference to which other BODIPY-structures can be compared to.
Like stated above, the absorbance of the chromophore can be altered via chemical derivatisations. However, the differences in the fluorescence properties of these lower alkylated BODIPY compounds are rather small. Higher alkylated BODIPY- compounds like it can be seen in the lower right structure tend to have a bit higher absorption maxima (528 nm) compared to lower substituted analogous like in the upper right structure (507 nm) 26.
Figure 6: Basic BODIPY-structures with different substituents, adapted from literature 26.
8 Next to parameters like light intensity, wavelength and duration of irradiation, also structural aspects play a role, affecting the speed and efficiency of the photocleavage 27. Depending on the exact structure of the BODIPY compound the absorption wavelengths can be modified and tailored towards specific needs, as already mentioned 28,13. Not only is the type of substituents important but also its position in the system. Substituents in position 8 have a low impact on the photodynamic properties, whereas large electron-donating groups in position 2 and 6 can lead to a substantial red shift of the absorbance 29. Especially the introduction of heavy atoms like halogens can not only increase the absorption wavelength but can also increase the photocleavage efficiency. This is due to an increased intersystem crossing (ISC) efficiency if the leaving group is liberated from the triplet excited state. The fluorescence quantum yields decrease with an increased molecular weight of the substituents in position 2 and 6 (I > Br > Cl > H), most likely due to increased ISC. It was found that the photocleavage efficiency can be increased proportionally to ISC quantum yields. In general, substituents which increase ISC and therefore lower the barrier for a release on the triplet state can improve the photocleavage efficiency drastically 13.
Furthermore, an increase of the conjugated system as through styryl- functionalised BODIPY-compounds can lead to absorption windows even over 700 nm which allows a deep tissue penetration 3. In Figure 7 an example for a 3,5-distyryl functionalised BODIPY structure can be seen.
Figure 7: Chemical structure of 3,5-distyryl functionalized BODIPY with absorption wavelengths around 700 nm 3.
9 Also, aryl-modified BODIPYs show absorption windows at the near IR spectrum.
The high rigidity of their π-system leads to high fluorescence quantum yields 30. It has been reported that BODIPY-compounds with an alkylated boron centre have higher fluorescence quantum yields than a fluorinated boron centre. However, it is also dependant which kind of alkyl group is attached to the boron atom. It has been shown that via irradiation existing alkyl groups can be cleaved and substituted with fitting solvent molecules 31. In Figure 8 the cleavage of alkyl-groups and the introduction of a methoxy-functionality is depicted. It is described that the compound on the right has a higher fluorescence quantum yield than the compound on the left by the factor 6.
Figure 8: Light induced substitution reaction on the boron-centre of a BODIPY compound for modification of photochemical properties of the fluorophore with their corresponding fluorescence
quantum yields 31.
The peak absorption can not only be altered through the introduction of different ligands, the BODIPY backbone itself can be modified. By the introduction of a nitrogen in position 8, aza-BODIPYs can be produced which can have absorption areas which extend into the near- infrared 32. However, because the meso-position can no longer be alkylated the uses as photocages are limited. In Figure 9 a comparison between the BODIPY and the aza-BODIPY structure can be seen.
10 Figure 9: Comparison of the aza-BODIPY (left) and BODIPY structure (right).
Whereas BODIPYs are typically very well soluble in most organic solvents 33, a challenge arises in their poor water solubility 34. This can be generally improved through the introduction of ionisable groups on the fluorophore or by the incorporation of the BODIPY into the backbone of a hydrophilic polymer 35. While many of these alterations can improve the solubility, the photodynamic properties of the compounds are changed as well which can make them unsuitable for their intended usage 34. An advantage of the introduction of ionisable groups lies within their easy implementation and small size. It is however possible that these ionic functionalities undergo non-specific interactions with chemicals or other biomolecules. Non-ionic water soluble BODIPY compounds are therefore superior in avoiding such side reactions, however they often depend on long sidechains like poly(ethyleneglycol) to achieve a sufficient water solubility 36.
Another method that is used in order to improve the water-solubility of BODIPY-compounds involves the introduction of sulfonate groups. However, with a conventional 2,6-sulfonation, the BODIPY structures can become unsuitable for photocleavage due to the strong electron-withdrawing effect of the substituent destabilizing the cleavage intermediate. To overcome this problem the sulfonate groups were positioned via an alkyl-residue at the 3 and 5 positions. The resulting compound, depicted in Figure 10, showed a better water-solubility and adjustable cell permeability depending on the degree of sulfonation. Possible uses may therefore arise as modulation agents for cell surface receptors 37.
11 Figure 10: Chemical structure of a water-soluble BODIPY-compound 37.
A factor that can hinder an effective photocleavage of BODIPYs is aggregation caused quenching (ACQ) 38. It has been a long reported problem also with other light emitting compounds, especially in the near-infrared light spectrum 39. This can lead to their dark and non-emitting appearance in high concentration or as a solid.
The small stokes-shift and the therefore high self-absorption of the emitted light can be seen as the main reason for this quenching effect 40. In highly concentrated solutions or in solid state usage the emission could therefore be massively reduced 41. This has to be taken into account when incorporated into polymeric structures to avoid undesired quenching when integrated in close proximity.
BODIPY- compounds opened new ways in various application as, for example, in photodynamic therapy, which is seen as one of the most promising methods in the treatment of cancer 42. Photodynamic therapy involves a photosensitive substance like BODIPY, which is injected into specific part of the body. Upon irradiation singlet oxygen can be produced in a sequence of steps which can consequently damage the target tissue 43. BODIPYs have also found its use in other cancer treatments.
It has been covalently bound to capsaicin. Capsaicin has not only found its treatment in anti-inflammatory treatments but also shows promising results in anti- tumor activity. With the usage of this covalently bound CAP-BODIPY complex the efficiency in prostate cancer treatments could be increased 44. The various fields of application show the importance of BODIPY and its derivatives 45.
12
13
2. Materials and methods
All used chemicals were purchased at Sigma Aldrich or VWR. Dry solvents have been stored over 4 Å molecular sieves. Before each use N2 was bubbled through the solvent. Air and moisture sensitive chemicals were stored in an argon filled glovebox. All other chemicals have been used without further purification. Solvents were reduced with the use of rotary evaporators.
Dry column vacuum chromatography was performed using 0.015 –0.040 mm silica gel. For the thin film chromatography ALUGRAM SIL G/UV254 plates were used.
The spots were observed under λ= 365 nm irradiation.
The 1H-NMR as well as the 13C-NMR measurements were carried out on a Bruker Advance III 300 MHz spectrometer. All spectra have been recorded in CDCl3 at a temperature of 25 °C. The chemical shifts are shown in parts per million (d). All spectra have been referenced against residual protons of the solvent peak of d = 7.26 ppm. The coupling constants (J) are given in hertz (Hz).
The UV-Vis measurements of the BODIPY-compounds as well as the oligomers were carried out in MeOH on a SpectraMax M2e from Molecular Devices. The peak intensities were normalized for a better comparability.
The kinetic studies were carried out on a Jobin Yvon Fluorolog 3 spectrometer allowing irradiation at the different wavelengths with roughly the same intensity.
GPC measurements were performed with a Viscotek GPCmax equipped with a Viscotek TDA 305 Triple Detector. Dimethyl formamide (DMF) with 10 mM lithium bromide (LiBr) was used as eluent with a flow rate of 0.75 ml min-1. The system was calibrated with polystyrene standards.
Mass spectra were recorded using an Agilent 6520 ESI-QTOF in positive mode.
14
3. Experimental procedure
3.1 Compound 1 (BODIPY) synthesis
The whole reaction was carried out under argon and in the dark. Kryptopyrrole (1.92 ml, 14.23 mmol, 2 eq,) was dissolved in 80 ml dry dichloromethane (DCM).
Acetoxyacetyl chloride (0.92 ml, 8.54 mmol, 1.2 eq) was added under stirring and the reaction was allowed to proceed over night at room temperature.
Triethylamine (TEA) (5.95 ml, 42.69 mmol, 6.0 eq) was added to the reaction mixture and after stirring for 15 min borontrifluoride diethyletherate (BF3.OEt2) (8.04 ml, 64.03 mmol, 9.0 eq) was added slowly. The reaction was stirred for 60 min after which a second batch of TEA (5.95 ml, 42.69 mmol, 6.0 eq) got added and stirred for 15 min. Finally, a second batch of BF3.OEt2 (8.04 ml, 9.0 eq, 64.03 mmol) was added as well and stirred for 60 min.
Upon completion the DCM was evaporated until 5 ml remained. The product mixture was diluted with ethyl acetate (EtOAc) and washed with brine. The water fractions were subsequently extracted with additional EtOAc and the combined organic phase was dried with MgSO4, filtered and the solvent evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) to yield compound 1 (BODIPY) (628 mg, 1.67 mmol, yield 23.5 %) as a shiny red solid.
15
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.05 (t, J= 7.5, 6H), 2.15 (s, 3H), 2.26 (s, 6H), 2.39 (q, J = 7.6, 4H), 2.50 (s, 6H), 5.30 (s, 2H)
ESI-MS (positive mode) calculated (C20H27BF2N2O2) 376.21, found m/z [M+H]+ 377.22.
λmax (abs.) = 544 nm; λmax (em.) = 564 nm
3.2 Hydrolysis of compound 1 (BODIPY)
Compound 1 (100.0 mg, 0.266 mmol, 1 eq) was dissolved in 6 ml tetrahydrofuran (THF). Lithiumhydroxide monohydrate (LiOH.H2O) (33.0 mg, 0.797 mmol, 3 eq) was dissolved in 6 ml deionized water. The aqueous LiOH.H2O solution was added under stirring and the reaction was allowed to proceed for 4 h in the dark.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) to yield compound 2 (Hydrolysed-BODIPY) (31.2 mg, 0.093 mmol, yield 35.1 %) as a shiny red solid.
16
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.04 (t, J= 7.5, 6H), 2.38 (q, J = 7.4, 4H), 2.40 (s, 6H), 2.50 (s, 6H), 5.30 (s, 2H)
ESI-MS (positive mode) calculated (C18H25BF2N2O) 334.20, found m/z [M+H]+
335.21.
λmax (abs.) = 538 nm; λmax (em.) = 557 nm
3.3 Hydroxylation of compound 1 (BODIPY)
The whole reaction was carried out under argon and in the dark. Compound 1 (BODIPY) (70.0 mg, 0.186 mmol, 1 eq) was dissolved in 1.5 ml dry DCM.
N-Bromosuccinimide (NBS) (35.7 mg, 0.186 mmol, 1 eq) was dissolved in 1.5 ml dry DCM and added to the solution. The reaction was carried out for 45 min.
A mixture of 1.5 ml deionized water and 2 ml of DMF was added and the reaction was allowed to proceed for 4 h.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) to yield compound 3 (Hydroxy-BODIPY) (17.80 mg, 0.045 mmol, yield 24.4 %) as a shiny red solid.
DCM, 45 min, RT, dark
17
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.06 (t, J= 7.6, 3H), 1.10 (t, J= 7.5, 3H), 2.15 (s, 3H), 2.28 (s, 6H), 2.40 (q, J= 7.5, 4H), 2.53 (s, 3H), 4.73 (s, 2H), 5.32 (s, 2H) ESI-MS (positive mode) calculated (C20H27BF2N2O3) 392.21, found m/z [M+Na] 415.20.
λmax (abs.) = 544 nm; λmax (em.) = 565 nm
3.4 Compound 4 (Diol-BODIPY) synthesis
3.4.1 Hydroxylation of compound 2 (Hydrolysed-BODIPY)
The whole reaction was carried out under argon and in the dark. Compound 2 (70.0 mg, 0.209 mmol, 1 eq) was dissolved in 1.5 ml dry DCM. NBS (37.3 mg, 0.209 mmol, 1 eq) was dissolved in 1.5 ml dry DCM and added to the solution. The reaction got carried out for 45 min. A mixture of 1.5 ml deionized water and 2 ml DMF was added and the reaction was allowed to proceed for 4 h.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) to yield compound 4 (Diol-BODIPY) (12.60 mg, 0.036 mmol, yield 17.2 %) as a shiny red solid.
DCM
18
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.07 (t, J= 7.5, 3H), 1.11 (t, J= 7.5, 3H), 2.42 (q, J=7.5, 2H), 2.46 (s, 6H), 2.50 (q, J= 7.5, 2H), 2.53 (s, 3H), 4.73 (s, 2H), 4.96 (s, 2H)
13C-NMR (CDCl3), 300 MHz): d (ppm) = 12.6, 13.0, 14.5, 15.6, 17.1, 54.9, 56.1 ESI-MS (positive mode) calculated (C18H25BF2N2O2) 350.20, found m/z [M+Na]
373.20.
λmax (abs.) = 538 nm; λmax (em.) = 557 nm
3.4.2 Hydrolysis of compound 3 (Hydroxy-BODIPY)
Compound 3 (40.0 mg, 0.102 mmol, 1 eq) was dissolved in 1.5 ml DCM. An aqueous mixture of 0.41 ml 0.1 M NaOH (0.4 eq, 0.041 mmol) and 2.5 ml MeOH was prepared separately and stirred for 10 min. The solutions then got combined and the reaction was allowed to proceed for 4 h at room temperature in the dark.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) to yield compound 4 (5.2 mg, 0.015 mmol, yield 14.6 %) as a shiny red solid.
19
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.07 (t, J= 7.5, 3H), 1.11 (t, J= 7.5, 3H), 2.42 (q, J=7.5, 2H), 2.46 (s, 6H), 2.50 (q, J= 7.5, 2H), 2.53 (s, 3H), 4.73 (s, 2H), 4.96 (s, 2H)
13C-NMR (CDCl3, 300 MHz): d (ppm) = 12.6, 13.0, 14.5, 15.6, 17.1, 54.9, 56.1 ESI-MS (positive mode) calculated (C18H25BF2N2O2) 350.20, found m/z [M+Na]
373.20.
λmax (abs.) = 538 nm; λmax (em.) = 557 nm
3.5 Compound 4 (Diol-BODIPY) one-pot synthesis
The whole reaction was carried out under argon and in the dark. Compound 1 (120.0 mg, 0.319 mmol, 1 eq) was dissolved in 2.5 ml dry DCM. NBS (60 mg, 0.337 mmol, 1.05 eq) was dissolved in 2.5 ml dry DCM and added to the solution.
The reaction got carried out for 45 min.
An aqueous mixture of 1.6 ml 0.2 M NaOH (1.0 eq, 0.320 mmol) in 10 ml MeOH was prepared separately and stirred for 10 min. The solutions got combined and the reaction was allowed to proceed for 4 h at room temperature in the dark.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane). The product was a mixture of unidentified side-products and no compound 4 (Diol-BODIPY) could be isolated.
20
3.6 Compound 5 (Dibenzylisocyanate-Diol) synthesis
The whole reaction was carried out under argon, at room temperature and in the dark. Compound 4 (50 mg, 142.77 mmol, 1 eq) was dissolved in 3 ml dry toluene.
Benzylisocyanate (36 mg, 270.37 mmol, 1.95 eq) got added under stirring along with ~0.5 mg DABCO. The whole reaction was allowed to proceed for 24 h.
The product mixture was diluted with EtOAc and washed with brine. The water fractions were extracted with EtOAc. The combined organic phase was dried with MgSO4, filtered and the solvent got evaporated under reduced pressure. The crude product was purified by dry column vacuum chromatography (0 %- 100 % EtOAc in heptane) twice to yield compound 5 (Dibenzylisocyanate-Diol) (18.0 mg, 0.029 mmol, yield 20.3 %) as a shiny red solid.
1H-NMR (CDCl3, 300 MHz): d (ppm) = 1.04 (t, J= 7.2, 6H), 2.31 (s, 6H), 2.43 (q, J = 7.5, 4H), 2.64 (s, 3H), 4.39 (s, 4H), 5.36 (s, 4H), 7.28 (m, 10H)
13C-NMR (CDCl3, 300 MHz): d (ppm) = 12.1, 13.4, 14.8, 17.1, 45.3, 57.9, 127.7 λmax (abs.) = 542 nm; λmax (em.) = 565 nm
21
3.7 Oligomer synthesis
3.7.1 Diol-ED-2003-Oligomer 1 (1:1)
The whole reaction was carried out under argon and in the dark. Jeffamine ED- 2003 (10.41 mg, 5.44 μmol, 0.95 eq) was dissolved in 0.5 ml dry toluene.
Compound 4 (2 mg, 5.71 μmol, 1 eq) was added to the solution along with ~0.5 mg DABCO. 4,4’-Methylen-bis-(cyclohexylisocyanate) H12-MDI (2.85 mg, 10.86 μmol, 1.90 eq) was added slowly to the solution and the reaction was allowed to proceed for 24 h at room temperature. The product was a very dark purple, almost black viscous substance.
3.7.2 Diol-ED-2003-Oligomer 2.1 (1:3)
The whole reaction was carried out under argon, at room temperature and in the dark. Compound 4 (2 mg, 5.71 μmol, 1 eq) was dissolved in ~0.5 ml dry toluene along with ~0.5 mg DABCO. H12-MDI (5.99 mg, 22.84 μmol, 4 eq) was added to the solution and the reaction was allowed to proceed for 24 h. Jeffamine ED-2003 (32.81 mg, 17.13 μmol, 3 eq) was added and the mixture was stirred for 1 h.
The product was further purified using 6-8 kDa dialysis tubes (one time H2O, 15 min; two times EtOH, 1 h; one time H2O, 1 h; one time EtOH, 1 h). Upon completion the solvent was removed under reduced pressure until dryness. The product was a dark purple viscous substance.
22 3.7.3 Diol-ED-2003-Oligomer 2.2 (1:3)
The whole reaction was carried out under argon, at room temperature and in the dark. Jeffamine ED-2003 (32.81 mg, 17.13 μmol, 3 eq) was dissolved in ~0.5 ml dry toluene. H12-MDI (5.99 mg, 22.84 μmol, 4 eq) got added and the reaction was allowed to proceed for 1 h. Compound 4 (2 mg, 5.71 μmol, 1 eq) was added along with ~0.5 mg DABCO. The reaction was stirred for 24 h.
The product got further purified using 6-8 kDa dialysis tubes (one time H2O, 15 min;
two times EtOH, 1 h; one time H2O, 1 h; one time EtOH, 1 h). Upon completion the solvent was removed under reduced pressure until dryness. The product was a dark purple viscous substance.
GPC (in DMF+ 10 mmol LiBr, 0.75 ml min-1) retention time: 14.5 min, Mn= 41000- 45000 g mol-1
3.7.4 Diol-ED-2003-Oligomer 3.1 (1:15)
The whole reaction was carried out under argon, at room temperature and in the dark. Compound 4 (2 mg, 5.71 μmol, 1 eq) was dissolved in ~0.5 ml dry toluene along with ~0.5 mg DABCO. H12-MDI (23.97 mg, 91.37 μmol, 16 eq) got added and the reaction was allowed to proceed for 24 h. Jeffamine ED-2003 (164.03 mg, 85.66 μmol, 15 eq) was added and the mixture was stirred for 1 h.
The product got further purified using 6-8 kDa dialysis tubes ((one time H2O, 15 min; two times EtOH, 1 h; one time H2O, 1 h; one time EtOH, 1 h). Upon completion the solvent was removed under pressure until dryness. The product was a pinkish-red soft solid.
GPC (in DMF+ 10 mmol LiBr, 0.75 ml min-1) retention time: 14.6 min, Mn= 57000- 64000 g mol-1
23 3.7.5 Diol-ED-2003-Oligomer 3.2 (1:15)
Jeffamine ED-2003 (164.03 mg, 85.66 μmol, 15 eq) was dissolved in ~0.5 ml dry toluene. The whole reaction was carried out under argon, at room temperature and in the dark. H12-MDI (23.97 mg, 91.37 μmol, 16 eq) got added and the solution was stirred for 1 h. Compound 4 (Diol-BODIPY) (2 mg, 5.71 μmol, 1 eq) was added along with ~0.5 mg DABCO. The whole reaction got stirred for 24 h.
The product got further purified using 6-8 kDa dialysis tubes ((one time H2O, 15 min; two times EtOH, 1 h; one time H2O, 1 h; one time EtOH, 1 h). Upon completion the solvent was removed under reduced pressure until dryness. The product was a pinkish-red soft solid depicted in Figure 32.
24
3.8 Hydrogel synthesis
3.8.1 Gel experiment 1 (Diol:Jeffamine = 1:2)
The whole reaction was carried out under argon and in the dark. Compound 4 (10.0 mg, 28.56 μmol, 1.0 eq) was dissolved in ~0.5 ml dry toluene. H12-MDI (13.19 mg, 50.26 μmol, 1.76 eq) and 1 mg DABCO were added to the toluene solution and the reaction was allowed to proceed over night at room temperature.
Jeffamine ED-2003 (102.82 mg, 53.69 μmol, 1.88 eq) was added in addition with poly(hexamethylene diisocyanate) (10.62 mg, 22.19 μmol, 1.16 eq). The reaction was stirred vigorously in order to achieve an even distribution of all reactants before gelation of the solution. The product got washed several times with acetone and deionized water in order to wash out any unreacted residues. The product was a dark pink, almost black coloured, soft and flexible gel. It showed a characteristic orange glow under λ= 365 nm.
25 3.8.2 Gel experiment 2 (Diol:Jeffamine = 1:28)
The whole reaction was carried out under argon and in the dark. Compound 4 (1.0 mg, 2.86 μmol, 1.0 eq) was dissolved in ~0.5 ml dry toluene. H12-MDI (13.19 mg, 50.26 μmol, 17.60 eq) and 1 mg DABCO were added to the toluene solution and the reaction was allowed to proceed over night at room temperature.
Jeffamine ED-2003 (152.0 mg, 79.38 μmol, 27.80 eq) was added in addition with poly(hexamethylene diisocyanate) (10.62 mg, 22.19 μmol, 11.66 eq). The reaction was stirred vigorously in order to achieve an even distribution of all reactants before gelation of the solution. The product got washed several times with acetone and deionized water in order to wash out any unreacted residues. The product was a pink coloured, soft and flexible gel. It showed the characteristic orange glow under λ= 365 nm.
3.8.3 Gel experiment 3 (Diol:Jeffamine = 1:140)
The whole reaction was carried out under argon and in the dark. Compound 4 (0.2 mg, 0.57 μmol, 1.0 eq) was dissolved in ~0.5 ml dry toluene. H12-MDI (13.19 mg, 50.26 μmol 88.0 eq) and 1 mg DABCO were added to the solution and the reaction was allowed to proceed over night at room temperature. Jeffamine ED-2003 (156.40 mg, 81.68 μmol, 143.0 eq) was added in addition with poly(hexamethylene diisocyanate) (10.62 mg, 22.19 μmol, 58.29 eq). The reaction was stirred vigorously in order to achieve an even distribution of all reactants before gelation of the solution. The product got washed several times with acetone and deionized water in order to wash out any unreacted residues. The product was a slightly pink coloured soft and flexible gel. It showed a slight orange glow under λ= 365 nm.
26 3.8.4 Gel experiment 4 (no BODIPY)
The whole reaction was carried out under argon and in the dark. Jeffamine ED-2003 (157.54 mg, 82.25 μmol, 3.0 eq) was dissolved in 1 ml dry toluene.
H12-MDI (13.20 mg, 50.26 μmol, 1.82 eq) was added to the solution and the reaction was allowed to proceed over night at room temperature.
Poly(hexamethylene diisocyanate) (10.63 mg, 22.19 μmol, 1.2 eq) was added and the reaction got stirred vigorously in order to achieve an even distribution of all reactants before gelation of the solution. The product got washed several times with acetone and deionized water in order to wash out any unreacted residues.
The product was a transparent elastic gel.
27
4. Results and discussion
4.1 Optical product characteristics
The produced BODIPY compounds all looked similar in their optical appearance.
In Figure 11 the characteristic red shimmering colour can be seen. The precise looks are strongly dependent on the crystal size.
Figure 11: Compound 1 (BODIPY) powder in a round bottom flask.
A very fine powder has an orange/ brown look whereas bigger crystals tend to have a red/golden colour. By a slow evaporation of the remaining solvents, bigger crystals can be produced which show a green shimmer. In Figure 12 a grown crystal of compound 1 (BODIPY) can be seen.
Figure 12: Compound 1 (BODIPY) crystal on a watch glass.
28 Compound 1 (BODIPY) itself in its unsolved state shows no fluorescence due to aggregation caused quenching. Upon dissolution a characteristic orange glow can be observed when irradiated with λ= 365 nm which can be seen in Figure 13. This characteristic applies to all synthesised BODIPYs.
Figure 13: Compound 1 (BODIPY) fluorescence under λ= 365 nm irradiation.
4.2 Compound 1 (BODIPY)
According to Figure 14, the successful synthesis of compound 1 (BODIPY) can be indicated by 1H-NMR-spectroscopy. Especially the presence of the triplet at 1.05 ppm, designated as signal a, and the quadruplet at 2.40 ppm, signal e, along with signal b at 5.30 ppm are strong indicators of a successful synthesis.
29 Figure 14: 1H-NMR of compound 1 (BODIPY) in CDCl3.
b
d e
f c a
30 In Addition, mass spectrometric measurements were performed, depicted in Figure 15. Despite showing various fragments a clear product peak can be assigned at 377.22 [M+H]+ and 399.20 [M+Na]+, further proving the successful synthesis of the BODIPY framework. The isotopic pattern of ~1:5:1 fits with the structure as well.
Figure 15: MS (ESI_QTOF)-spectrum of compound 1 (BODIPY).
31 Finally, the UV-Vis measurements, depicted in Figure 16 show a pure fluorophore with overlapping excitation and absorption spectra. The absorbance measurement shows a maximum at λ= 544 nm, the emission spectrum at λ= 564 nm and the excitation measurements at λ= 541 nm. The emission shows a stokes shift of the maxima of 20 nm and is nicely mirrored to the absorbance.
Figure 16: Absorbance (green dash/dotted line), emission (red dashed line) and excitation (blue bold line) spectra of compound 1 (BODIPY) in MeOH.
350 400 450 500 550 600 650 700 750 0.0
0.2 0.4 0.6 0.8 1.0
Normali zed intensi ty
Wavelength / nm
Normalized Absorbance Normalized Emission Normalized Excitation
32
4.3 Compound 2 (Hydrolysed-BODIPY)
To control the complete hydrolysis of compound 1 (BODIPY) a 1H-NMR was recorded. As can be seen from Figure 17 the signals a and e, the triplet and quadruplet respectively, remain the same as compared to compound 1, however, the singlet of signal d is shifted into the quadruplet. Signal c, present at 2.15 ppm in Figure 14 disappears whereas signal b is shifted upfield.
Figure 17: 1H-NMR of compound 2 (Hydrolysed-BODIPY) in CDCl3.
a d
e f b
33 As for compound 1 the MS spectra shows various fragments, still the product peak can be nicely assigned at 335.21 [M+H]+ which can be seen in Figure 18. The isotopic pattern of ~1:5:1 fits with the structure as well. In combination both spectra prove of a successful synthesis.
Figure 18: MS (ESI_QTOF)-spectrum of compound 2 (Hydrolysed-BODIPY).
34 Again, the purity of the fluorophore was examined by UV-Vis spectroscopy which is visible in Figure 19. As for compound 1 the absorbance and excitation spectra are overlapping indicating a pure product. The stokes shift of the emission remained around the same with 19 nm, nevertheless the absorbance maximum shifted to 538 nm compared to 544 nm for compound 1 possibly due to different inductive effects of the ester compared to the alcohol. The same effect can be seen in the emission spectrum with a maximum at λ= 557 nm and in the excitation measurements with maximum at λ= 539 nm.
Figure 19: Absorbance (green dash/dotted line), emission (red dashed line) and excitation (blue bold line) spectra of compound 2 (Hydrolysed-BODIPY) in MeOH.
350 400 450 500 550 600 650 700 750 0.0
0.2 0.4 0.6 0.8 1.0
Normalized intensity
Wavelength / nm
Normalized Absorbance Normalized Emission Normalized Excitation
35
4.4 Compound 3 (Hydroxy-BODIPY)
The structural differences in comparison to compound 1 (BODIPY) are due to the introduction of an alcohol group in position 3 of the BODIPY framework. To control the complete hydroxylation of compound 1 a 1H-NMR was recorded. As can be seen from Figure 20 the 2 protons at signal g next to the alcohol functionalization with a shift of d = 4.73 ppm can be used in order to identify a successful hydroxylation reaction.
The 6 protons of signal a can be seen as 2 triplets shifting into each other because of the loss of symmetry through the introduction of the alcohol group. Same is true for the 4 protons at e.
Figure 20: 1H-NMR of compound 3 (Hydroxy-BODIPY) in CDCl3. d a f e g
b
c
36 The MS spectra shows only little fragmentation, depicted in Figure 21. The product peak can be nicely assigned at 415.20 [M+Na]+. The isotopic pattern of ~1:5:1 fits with the structure as well. A fragment peak is measured at 375.21 which corresponds to compound 3 (Hydroxy-BODIPY) with an abstracted hydroxide functionalization. This is unexpected as the acetate is considered as the labile functionality. The substitution in the meso position therefore seems to have a considerable influence on the stability of different substituents. No hydrolysis of the ester functionality can be observed. No further relevant peaks can be observed. In combination both spectra prove of a successful synthesis.
Figure 21: MS (ESI_QTOF)-spectrum of compound 3 (Hydroxy-BODIPY).
37 Again, the purity of the fluorophore was examined by UV-Vis spectroscopy which is visible in Figure 22. The absorbance and excitation spectra are overlapping indicating a pure product. The stokes shift of 21 nm as well as the absorption maximum at 544 nm remained around the same compared to compound 1. A similar trend can also be seen in the emission spectrum with a maximum at λ= 565 nm and in the excitation measurements with maximum at λ= 544 nm.
Figure 22: Absorbance (green dash/ dotted line), emission (red dashed line) and excitation (blue bold line) spectra of compound 3 (Hydroxy-BODIPY) in MeOH.
350 400 450 500 550 600 650 700 750 0.0
0.2 0.4 0.6 0.8 1.0
Normali zed intensi ty
Wavelength / nm
Normalized Absorbance Normalized Emission Normalized Excitation
38
4.5 Compound 4 (Diol-BODIPY)
To control the complete synthesis of compound 4 a 1H-NMR was recorded. As can be seen from Figure 23 the 2 protons at signal g with a shift of d = 4.7 ppm in addition with the 2 protons at signal b with a shift of d = 4.9 ppm are used in order to verify a successful synthesis of compound 4. The 6 protons of signal d are split up as in compound 3 (Hydroxy-BODIPY) and shifted a bit higher as for compound 2 (Hydrolysed-BODIPY). Similar to the triplets the 4 protons at e also split into a doublet of a quadruplet, although the splitting is not well observed due to the overlap of signals. A HSQC spectrum was recorded additionally in order to assign all protons to the corresponding carbon atoms which can be seen in Figure S1.
Figure 23: 1H-NMR of compound 4 (Diol-BODIPY) in CDCl3.
a d
f e
g b
39 As for compound 3 the MS spectra shows only little fragmentation, depicted in Figure 24. The product peak can be nicely assigned at 373.20 [M+Na]+. The isotopic pattern of ~1:5:1 fits with the structure as well. A fragment peak is measured at 333.22, which corresponds to compound 4 (Diol-BODIPY) with an abstracted hydroxide functionalization. Which of the two hydroxide groups was cleaved could not be further determined as both result in a fragment peak with the same m/z. Considering the spectrum from compound 3 (Hydroxy-BODIPY) it suggests that the lower hydroxide in position 3 has a higher instability and tends to cleave more readily, however the hydrolysis of the acetate could have an influence here too.
The substitution in the meso position therefore seems to have a considerable influence on the stability of different substituents in compound 4 (Diol-BODIPY) as well. No other relevant peaks can be observed. In combination both spectra prove of a successful synthesis.
Figure 24: MS (ESI_QTOF)-spectrum of compound 4 (Diol-BODIPY).
40 Again, the purity of the fluorophore was examined by UV-Vis spectroscopy which is visible in Figure 25. As for the other compounds the absorbance and excitation spectra are overlapping indicating a pure product. The stokes shift of the emission remained around the same with 19 nm, nevertheless the absorbance maximum shifted to 538 nm compared to 544 nm for compound 1 possibly due to different inductive effects of the ester compared to the alcohol. The same effect can be seen in the emission spectrum with a maximum at λ= 557 nm and in the excitation measurements with maximum at λ= 538 nm.
Figure 25: Absorbance (green dash/ dotted line), emission (red dashed line) and excitation (blue bold line) spectra of compound 4 (Diol-BODIPY) in MeOH.
350 400 450 500 550 600 650 700 750 0.0
0.2 0.4 0.6 0.8 1.0
Normalized intensity
Wavelength / nm
Normalized Absorbtion Normalized Emission Normalized Excitation
41
4.6 Compound 5 (Dibenzylisocyanate-BODIPY)
Compound 5 was synthesized as a small molecular model compound for the subsequent macromolecular applications of compound 4 (Diol-BODIPY) and analysed via 1H-NMR spectroscopy, visible in Figure 26. The shifts between d = 0 - 2.6 ppm are nearly identical to compound 4 (Diol-BODIPY). This is because the chemical environment of these protons is not drastically altered by the addition of the isocyanate. The aromatic protons at signal h verify the successful introduction of the aromatic structures onto compound 4. A HSQC spectrum was recorded additionally in order to assign all protons to the corresponding carbon atoms which can be seen in Figure S2 (supporting information).
Figure 26: 1H-NMR of compound 5 (Dibenzylisocyanate-BODIPY) in CDCl3. a
d f e
g b
h
42 To allow for a proper photochemical analysis the fluorophore had to be especially pure and was therefore purified twice by DCVC and 1H-NMR spectra were recorded in-between. The aim of the purification process was to reduce the peak at d = 5.1 ppm, as this signal could not be assigned to the product. However, as it can be seen in the NMR comparison in Figure 27 below, almost no difference can be observed.
Figure 27: 1H-NMR comparison of the impurified and purified compound 5 (Dibenzylisocyanate- BODIPY) in CDCl3.
The purification was carried out with the use of dry column vacuum chromatography and the fractions were controlled with thin film chromatography plates in order to combine the different fractions. As it can be seen from Figure 28 below, the degree of separation was poor. The upper big spot is the product peak, and the smaller peak underneath are impurities which may explain the peak at d = 5.1 ppm. However, from the NMR-measurement as well as the absorption and emission measurements no other impurities could be detected.
--- no purification --- 1 time purification --- 2 times purification
43 Figure 28: Thin film chromatography spots of the different fractions collected during the
chromatographic purification of compound 5.
Again, the purity of the fluorophore was examined by UV-Vis spectroscopy which is visible in Figure 29. As for the other compounds the absorbance and excitation spectra are overlapping indicating a pure product. Unlike in the stokes shift of the other compounds of around 20 nm, the stokes shift of the emission increased to 24 nm. The absorbance maximum shifted to 542 nm compared to 538 nm for compound 4 possibly due to different inductive effects of the urethane groups. The same effect can be seen in the emission spectrum with a maximum at λ= 565 nm and in the excitation measurements with maximum at λ= 542 nm.
44 Figure 29: Absorbance (green dash/ dotted line), emission (red dashed line) and excitation (blue bold line) spectra of compound 5 (Dibenzylisocyanate BODIPY) in MeOH.
350 400 450 500 550 600 650 700 750 0.0
0.2 0.4 0.6 0.8 1.0
Normalized intensity
Wavelength / nm
Normalized Absorbtion Normalized Emission Normalized Excitation
45
4.7 Absorption measurement comparison
To analyse the influence of the various modifications of the BODIPY structure on its absorbance, the spectra were compared as plotted in Figure 30.
Figure 30: Comparison of the different absorbance spectra of the BODIPY compounds 1-5 in methanol with normalized absorption. Top (full spectrum), bottom (zoomed).
The acetate in the meso position of compound 1 (BODIPY) and compound 3 (Hydroxy-BODIPY) is electron donating (+I-effect) whereas the alcohol functionality in compound 2 (hydrolysed-BODIPY) and compound 4 (Diol-BODIPY) is electron withdrawing (-I-effect). The urethane groups in compound 5 (Dibenzylisocyanate-BODIPY) have a similar electron donating effect as the
350 400 450 500 550 600 650 700 750
0.0 0.2 0.4 0.6 0.8 1.0
500 520 540 560 580
0.6 0.8 1.0
Normalized Absorption
Wavelength / nm
compound 1 (BODIPY)
compound 2 (hydrolyzed-BODIPY) compound 3 (hydrox-BODIPY) compound 4 (diol-BODIPY)
compound 5 (dibenzylisocyanate-Diol)
Normalized Absorption
Wavelength / nm
compound 1 (BODIPY)
compound 2 (hydrolyzed-BODIPY) compound 3 (hydrox-BODIPY) compound 4 (diol-BODIPY)
compound 5 (dibenzylisocyanate-Diol)
46 acetate. However, no correlation between the inductive effect and the absorption wavelength could be found. Compound 3 (Hydroxy-BODIPY) with an absorption at the longest wavelengths has one donating and one withdrawing group.
Compound 2 (Hydrolysed-BODIPY) with just one electron withdrawing group shows an absorption at the shortest wavelengths. If the inductive effect would be responsible for the shifts in absorption, the extrema would be expected in compound 5 (Dibenzylisocyanate-BODIPY) with the highest electron donating effects and in compound 4 (Diol-BODIPY) with the highest electron withdrawing effects.
In general, the hydrolysis of the BODIPY tends to shift the peak absorption towards smaller wavelengths. The introduction of an alcohol in position 3 has no significant influence on the absorption. Derivatisations in the meso-position therefore may lead to bigger shifts in absorption. From the spectra it could be seen that the absorption maxima of the different compounds were similar, as no changes to the conjugated system itself were done. An extension of itself may possibly lead to absorptions at longer wavelengths like it can be seen in the 3,5 -distyryl functionalized BODIPY in Figure 7.
47
4.8 Emission spectra comparison
A similar trend can be observed in the emission spectra, depicted in Figure 31. The hydrolysis of the BODIPY tends to shift the emission peaks towards lower wavelengths whereas the introduction of hydroxyl functionality shows hardly any influence on the peak emission. The impact of the changes in the meso-position on the peak emission outweighs, like for the absorption, the derivatisations in position 3. Whereas the emission spectrum of compound 2 and compound 5 overlap, the absorption spectra vary. It therefore indicates a higher stokes shift of compound 5.
Figure 31: Comparison of the different emission spectra of the BODIPY- compounds 1-5 in methanol with normalized intensities. Top (full spectrum), bottom (zoomed).
500 550 600 650 700 750
0.0 0.2 0.4 0.6 0.8 1.0
520 540 560 580 600 620 640
0.4 0.6 0.8 1.0
Normalized Emission
Wavelength / nm
compound 1 (BODIPY)
compound 2 (hydrolized-BODIPY) compound 3 (hydrox-BODIPY) compound 4 (diol-BODIPY)
compound 5 (dibenzylisocyanate-Diol)
Normalized Emission
Wavelength / nm
compound 1 (BODIPY)
compound 2 (hydrolized-BODIPY) compound 3 (hydrox-BODIPY) compound 4 (diol-BODIPY)
compound 5 (dibenzylisocyanate-Diol)
48
4.9 Oligomer characterisation and degradation studies
The characterization of the oligomers was more difficult compared to the small molecular BODIPY compounds since NMR spectroscopy was hindered due to the large signals stemming from the Jeffamine. Therefore, they were characterized via UV-Vis spectroscopy and GPC. The same methodology was used for the investigation of the degradation behavior of the oligomers.
In order to receive preliminary information about the degradation process an oligomer was irradiated with 365 nm and 510 nm light in different solvents.
Diol-ED-2003-Oligomer 1 was chosen for the irradiation experiments as it was accessible at first. Absorbance spectra were recorded to follow the process and to estimate the degradation rate. GPC measurements were conducted for the oligomers to receive data regarding the molecular weight before and after the irradiation processes, in order to verify the degradation of the oligomeric chains.
4.9.1 UV-Vis
The produced oligomers were soft flexible solids. Their colours ranged from dark purple to slightly pink depending on their BODIPY concentration. In Figure 32 the optical appearance of a Diol-ED-2003-Oligomer 3.1 film can be seen.
Figure 32: Photo of the Diol-ED-2003-Oligomer 3.1
49 The oligomers were irradiated with λ= 365 nm and λ= 510 nm light in order to study the degradation behaviour. The experiments have been carried out in two different solvents as a degradation of the oligomers was observed in DMF without any irradiation. Therefore, the measurements have also been performed in MeOH to rule out any polymer instability.
4.9.1.1 Diol-ED-2003-Oligomer 1 in DMF
To investigate the degradation of the oligomer at λ= 365 nm the sample has been dissolved in DMF and irradiated for 5 min before each measurement. The measurements were carried out without any interruption or overnight storage. Only every second measurement is plotted to simplify the spectrum and ease the interpretation. As it can be seen in Figure 33 the absorption decreases linearly with each irradiation step. Additionally, the broad signal from λ= 600 - 780 nm shows a significant reduction in absorption. Since for the absorption measurements of the monomeric BODIPYs no absorption could be determined at these wavelengths, it was assumed that these signals are not originating from any BODIPY compounds.
The exact structures underlying this signal have not been further characterized.
After 60 min the absorption maximum was reduced to 66 % of the starting value.
