Doctoral Proposal .pdf

Enhanced Two-Photon Absorption Cross-Sections in
Conjugated Polyelectrolyte Aggregates for Use as Drug
Carriers

C. Jason Zeman
7 September 2016

Oral Qualifying Exam
University of Florida
Department of Chemistry
Division of Physical Chemistry

Dissertation Committee
Prof. Kirk Schanze
Prof. Alex Angerhofer
Prof. Valeria Kleiman
Prof. Philip Brucat
Prof. Jennifer Andrew

ABSTRACT

Since the discovery of water-soluble, conjugated polyelectrolytes, researchers
have been actively investigating them for their peculiar properties. The property of
particular interest to this proposal is the aggregation behavior of these polymers.
Aggregation may be induced by oppositely charged ions, which is easily observed by
quenching of fluorescence. Notably, conjugated polyelectrolytes in aggregate states are
known to experience stabilization of the conjugated backbone by interchain π-π
interactions. This property may help facilitate multiphoton absorption when aggregation
is induced for a properly designed polymer. Should this hypothesis hold true, conjugated
polyelectrolytes may be potential candidates for a drug delivery system for platinumbased antineoplastic drugs: a class of anticancer agents that are infamous for their
indifference in affecting cancerous and noncancerous cells alike. A conjugated
polyelectrolyte may be able to render these drugs inert by aggregating around them in a
chelation-like fashion, and thereupon becoming sensitized to multiphoton absorption
when acting as a drug carrier. It is hoped that upon multiphoton-induced excitation, a
charge-transfer event may facilitate the release of the drug. This proposal outlines a
systematic study on the effect that aggregation has on the two-photon absorption of a
particular polyelectrolyte. Both two-photon excited fluorescence and non-linear
transmission techniques will be used for complete characterization. Photolysis techniques
will be employed for determining the efficiency with which platinum based antineoplastic
drugs are released from aggregates after two-photon excitation.

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SPECIFIC AIMS
The controlled release of anticancer agents such as platinum based antineoplastic
drugs is an important contemporary research goal, as this class of drugs has been shown
to cause a number of negative side effects. One potential solution to this problem is
multiphoton-induced electron transfer to stimulate the release of the drug. This proposal
aims to study a specific class of drug carriers that may be idealized for this process.
The compounds to be studied are conjugated polyelectrolytes: a type of polymer
that is known to aggregate in the presence of oppositely charged, multivalent ions. It is
believed that these polymers may be designed in such a way that they may release a
platinum based anticancer agent after electronic excitation by IR light. More importantly,
it is believed that these polymers will show enhanced multi-photon absorption when
aggregated. The current state of this field of study has shown that most multi-photon
absorbing chromophores experience diminished multi-photon absorbing behavior in
aqueous solution, making their viability as drug carriers reduced dramatically. Conjugated
polyelectrolytes may not only be an exception to this apparent rule, but may experience
enhanced multi-photon absorbing character when carrying anticancer agents. This
proposal aims to study their efficacy in this regard, as there are currently no known reports
in the literature on the dependence of aggregation of conjugated polyelectrolytes on their
two-photon absorption cross-sections. Should this prove fruitful, research may progress
towards efficiency of drug release following photoexcitation.

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BACKGROUND AND SIGNIFICANCE
Platinum based antineoplastic drugs have been a golden standard in
chemotherapy for decades ever since the first of their variety, cisplatin, was shown to
increase survival rates for some types of cancers to upwards of 90% compared to just
5% without chemotherapy (1, 2, 3). Their method of action has been, and still is, an area

(Figure 1. All platinum based antineoplastic drugs that have been implemented for cancer
treatment in at least one country.)4

of great research interest. In recent years, cisplatin has evolved to several other types of
ligated Pt(II) complexes of varying complexity. Of the dozens of platinum based
antineoplastic drugs actively being studied today for their unique medical benefit, six have
gained marketing approval for human use, and several more have reached clinical trials
(4). The structures of those that are currently in use are shown in Figure 1.
These Pt(II) complexes all function medicinally via the same general pathway: loss
of anionic ligands followed by binding of Pt to the N7 position of Guanine in two separate

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strands of DNA, rendering DNA replication impossible and inevitably ending in apoptosis
of the cancerous cell (5). Unfortunately, these drugs act indiscriminately towards all
rapidly dividing cells and therein lies the risk (4). With side effects such as reduced kidney
function, nervous system damage, hearing loss, reduced bone marrow activity, and hair
loss when combined with other drugs, the need to be able to bias these drugs towards
cancerous cells is an obvious goal in the pursuit for the solution to this problem. One such
solution, is to render the drug temporarily inert until it is needed. This proposal outlines a
potential method for controlling the activity of these drugs through the use of conjugated
polyelectrolytes.

Conjugated Polyelectrolytes
Conjugated polyelectrolytes (CPEs) have been studied rigorously since their
discovery. Characterized as having an extensively π-conjugated backbone with ionically
charged side chains, CPEs are of great interest because of their high molar absorptivity,
high quantum yields, and aqueous processability (6). Because of these traits, they have
shown themselves to have great potential for their applications in solar cells, optics,
organic electronics, biomedical sensors and other electrochromic devices (7). Due to the
extensive number of applications and interesting properties of CPEs, it would be difficult
to sufficiently summarize in any detail all of the interesting characteristics that CPEs have
been shown to have.
One peculiar characteristic of CPEs is that they exhibit amplified quenching by
oppositely charged ionic quenchers (8). Where the Stern-Volmer quenching constant,

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KSV, as determined from Equation 1 (where I0 and I are the fluorescence intensity without
and with the quencher present respectively, and [Q] is the quencher concentration),
!"
!

= 1 + & '() [+]

(Equation 1)

is normally on the order of 101 – 103 M-1, KSV values on the order of 106 – 109 M-1 have
been reported for CPEs (9, 10). An indicative characteristic of amplified quenching is that
at low quencher concentrations, I0/I increases linearly with increasing quencher
concentration, but develops an upwards curvature (dubbed superlinearity) at higher
quencher concentrations, as seen in Figure 2 (8). Extensive research has been done to
explain how the emission is so efficiently quenched. While several explanations have

(Figure 2. A Stern-Vomer plot showing superlinearity for a CPE that is being quenched
by methyl viologen at various different Ca2+ concentrations. Superlinearity is experienced
more readily for higher Ca2+ concentrations. Reprinted with permission from Jiang et al.)6

been proposed, including charge-transfer to the quencher, Förster resonance energy
transfer, efficient interchain exciton mobility, or ion-pair complex formation between CPE

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and quencher, these effects are all likely contributors to amplified quenching (9, 10, 11).
Importantly, these effects are resultant of polymer aggregation in solution (8, 12, 13, 14).
The polymer aggregation behavior that gives rise to amplified quenching can
exhibit distinctly different identifiable characteristics in comparison to a non-aggregated
CPE. For reference, sample UV-Vis absorption and emission spectra are shown in Figure
3 (15). When a CPE is dissolved in a good solvent, the conjugated backbone exists in a
highly disorganized state with frequent breakage of conjugation. This state of high
dissolution has been referred to as the --phase (16, 17). Likewise, when solvated by a
poor solvent, CPEs tend to aggregate in such a way that minimizes contact with the
solvent. Via aggregation, conjugation of a single chain of a CPE is stabilized by other
CPEs and can be observed as a red shift to the --phase absorption. These tightly packed
chains have been aptly named compact aggregates (14).

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(Figure 3. Absorption (a) and Emission (b) spectra of a sulfate functionalized
polyphenylene ethynylene CPE in methanol (—), 1:1 methanol:water (- - -), and water (·· -) where emission has been normalized to reflect relative quantum yield. Reprinted with
permission from Tan et al.)15

Alternatively, CPEs may form loose aggregates in the presence oppositely
charged ions. It has been shown experimentally that this effect is only possible for
multivalent ions, allowing for the interchain crosslinking that gives rise to aggregation.
These loose aggregates that are tethered together via their ionic pendants can easily be
swelled with solvent and allow for intercalation of other compounds, size permitting.
Similar to compact type aggregates, loose aggregates also experience stabilization of the
conjugated backbones of the CPEs in the aggregate, making loose aggregates typically
indiscernible from compact aggregates in a UV-Vis absorption spectrum. (8, 14)
The most efficient way of recognizing aggregation is by fluorescence. While
emission for CPEs in the --phase is usually of high intensity, the spectral shape shows
clear vibronic structure with narrow bandwidth for the 0-0 transition. Contrastingly,
emission from an aggregate state shows no vibronic structure and a very broad and
bathochromically shifted band of lower intensity. Of course, if aggregation has been
induced by a quencher, it is possible for the CPEs to be completely non-emissive; a
characteristic of amplified quenching. (18)

Multi-photon Absorption
A concept first proposed in 1931 and finally confirmed in 1961 to occur (19, 20),
multi-photon absorption (MPA) has repeatedly shown great promise in the field of

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medicine for its use in fluorescence microscopy of biological samples, less photo-toxicity,
low tissue auto-fluorescence, and especially deep tissue penetration (21). These effects
are all resultant of the inherent lower photon energy associated with multi-photon
absorption in comparison to one photon absorption (1PA), and is summarized in Figure
4. Whereas in 1PA a single photon is absorbed by a molecule that is then promoted to an
energetically excited state, degenerate two photon absorption (2PA) of halved

(Figure 4. A simplified charaicature of one, two, and three photon absorption processes)

frequencies will achieve the same effect. Likewise, the same can be said for higher orders
of multi-photon absorption. Although this quantum electrodynamical model for MPA
typically explains the process as a cascade through intermediate or virtual states, we
know MPA is a single step that requires simultaneous absorption of two photons in
extreme proximity (22, 23, 24). The ramification of this requirement is that MPA is greatly
dependent on light intensity.

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