Water-soluble [2.2]paracyclophane chromophores with large two-photon action cross sections

Article abstract of DOI:10.1021/ja0440811

A series of α,ω-donor-substituted distyrylbenzene dimers held together by the [2.2]paracyclophane core were designed, synthesized, and characterized. Different substituents were chosen to modulate the strength of the donor nitrogen groups and to allow the

Full text of DOI:10.1021/ja0440811

Published on Web 12/30/2004
Water-Soluble [2.2]Paracyclophane Chromophores with Large Two-Photon
Action Cross Sections
Han Young Woo, Janice W. Hong, Bin Liu, Alexander Mikhailovsky, Dmitry Korystov, and
Guillermo C. Bazan*
Mitsubishi Chemical Center for AdVanced Materials, Department of Materials,
Institute for Polymers and Organic Solids, UniVersity of California, Santa Barbara, California 93106
Received September 28, 2004; E-mail: bazan@chem.ucsb.edu
Two-photon fluorescence microscopy (TPM),1 and higher mul-
Scheme 1
tiphoton variations, provide the best noninvasive means of fluo-
rescence microscopy for biological imaging. The advantages of
TPM have been described in detail elsewhere2 and include reduced
photodamage, improved depth penetration, the ability to image
turbid samples, and reduced background cellular autofluorescence.
The application of multiphoton microscopy in elucidating biological
processes is increasing exponentially.3
TPM-specific fluorophores for biomolecular tags are an important
consideration, since these provide a means to further reduce
photodamage and enable the use of more economical laser sources.2
Two parameters need to be optimized for best performance. One
Scheme 2 a
is the two-photon absorption cross section (δ, expressed in GM )
1 × 10-50 cm4‚s‚photon-1‚molecule-1), which provides the prob-
ability of two-photon absorption (TPA) at a particular frequency.
A large fluorescence quantum yield (η) is also desired for better
signal-to-noise. The product ηδ defines the two-photon action cross
section of a molecule.
Commonly used fluorescent reporters have δ values in the 1-100
GM range, which give ηδ products of 1-50 GM.2,4,5 Recent
molecular guidelines have shown how to obtain δ > 1000 GM.6
a Reagents and conditions: (i) t-BuOK, THF, 0 °C, 6 h; (ii) NaI, acetone/
One approach is a quasi linear D-π-D structure, where D is a
CH2Cl2, reflux, 2 days; (iii) NMe3, THF/H2O, room temperature, 24 h.
donor group and π is a π-conjugated bridge. The magnitude of δ
Table 1. Spectroscopy Summary
depends on the degree of intramolecular charge transfer (ICT) upon
excitation.7 However, in polar aqueous media, necessary for
biological samples, the ICT leads to reduced η values and lower
overall ηδ.8,9
In this contribution, we report the synthesis of water-soluble TPA
chromophores based on the [2.2]paracyclophane (pCp) core which
display exceptionally large ηδ values. The specific molecular
structures are shown in Scheme 1. Each molecule can be considered
to contain two distyrylbenzene fragments held together by their
internal rings via the pCp core and was expected to have a δ value
approximately twice that of the constituent distyrylbenzene chro-
mophore.10
The terminal groups determine the donor ability of the nitrogen
atom6b and whether the molecule is neutral (N series) and soluble
in nonpolar organic solvents, or cationic (C series) and soluble in
water. The overall set of compounds allows the examination of
the solvent influence on δ and the effect of donor strength on η.
The synthetic routes are given in Scheme 2. Four-fold Horner-
Emmons Wittig coupling reactions between 4,7,12,15-tetra(dieth-
ylphosphonatemethyl)-[2.2]paracyclophane11 and excess 4, 5, or 6
using potassium tert-butoxide in THF at 0 °C give 1N, 2N, or 3N,
respectively, in yields close to 60%. Chloride/iodide exchange of
1N, 2N, and 3N, via the Finkelstein reaction, followed by
quaternization with a large excess trimethylamine, provides 1C,
2C, and 3C, respectively, in 85-90% yields.
solvent
λabs
λem
ηa
λTPA
δ
(GM)b
1N
1C
2N
2C
3N
3C
toluene
water
toluene
water
toluene
water
434
435
420
410
441
431
486
553
468
505
492
537
0.92
0.04
0.95
0.42
0.92
0.52
725
725
700
700
770
750
1290
370
1690
700
2080
690
a Measured at 10-6 M relative to fluorescein. b Peak TPA cross section
at λTPA
.
Table 1 summarizes the absorption and photoluminescence (PL)
spectra. Figure 1 shows the spectra for 3N and 3C, which are typical
for the entire series of compounds. Neutral compounds were
measured in toluene and the charged counterparts in water.
The maxima in absorption (λabs) and emission (λem) for 1N are
similar to those reported for 4,7,12,15-tetra(4′-dihexylaminostyryl)-
[2.2]paracyclophane,11 which indicates negligible perturbation by
the chloride functionalities. There is a slight blue shift in the λabs
and λem of 2N, relative to those of 1N. The absorption and emission
of 3N are red-shifted relative to those of 1N. We note that the energy
differences between the electronic states of bichromophoric pCp
molecules are determined by through-space delocalization (electron
exchange) across the pCp core.12 The main absorption in the UV-
vis range corresponds to two closely separated states (IIa and IIb
in ref 12a), and the energy difference between these states
9
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J. AM. CHEM. SOC. 2005, 127, 820-821
10.1021/ja0440811 CCC: $30.25 © 2005 American Chemical Society

C O M M U N I C A T I O N S
causes a substantial decrease.7,16b The larger polarity of water could
provide for a similar charge transfer stabilization in the ground and
excited states. However, there is no theoretical examination of how
hydrogen bonding from the solvent influences the strength of the
donor groups at the termini of the π system and how these changes
influence δ. A more detailed structure/optical properties relationship
analysis therefore awaits theoretical examination. From a practical
perspective, the larger ηδ values of 2C and 3C, relative to that of
1C, indicate that maximizing the donor strength of the terminal
nitrogen atoms is not necessarily a useful design parameter for TPM
applications. We propose that weaker donors, such as triarylamine,
are a better choice, because the weaker ICT character of the excited
state leads to higher η. This search for an optimal balance is likely
to be applicable to other molecular systems. Despite the gap in
relating molecular structure to optical properties, 2C and 3C show
exceptionally large action cross sections in water.
Figure 1. Normalized absorption and PL spectra of 3N in toluene (a, a′)
and 3C in water (b, b′). PL spectra were collected by exciting at the λabs of
each sample. TPA spectra of 3N in toluene (c) and 3C in water (d) (GM )
10-50 cm4‚s‚photon-1).
determines the overall absorption band shape and ultimately λabs
.
A discussion of the relatively small perturbation of solvents on λabs
and λem is therefore not appropriate at this stage. In toluene, 1N,
2N, and 3N have high η values, in the range of 0.9.
Acknowledgment. The authors are grateful to Mitsubishi
Chemical Center for Advanced Materials (MC-CAM), the NIH
(GM62958-01), and the NSF (DMR-0097611) for financial support.
A comparison of the linear spectra of the C series in water with
those of the N series in toluene reveals the following trends (Table
1). The λabs values are nearly the same (1C/1N) or slightly blue-
shifted (2C/2N, 3C/3N). The emissions are red-shifted and broader,
with no vibronic structure (see Figure 1). Most importantly for the
design of efficient TPM fluorophores, the η values in water are
inversely proportional to the donor strength of the terminal groups
(1N ∼ 1C > 2N ∼ 2C > 3N ∼ 3C).13 A substantial drop in η is
observed for 1C (0.04), while the η values for 2C (0.42) and 3C
(0.52) remain high. We also note that, in a solvent of intermediate
polarity (DMSO), there are no measurable differences in the linear
spectroscopy of the N and C series (Supporting Information). The
charged groups therefore are not interacting with the chromo-
phores.14
Two-photon excitation spectra were measured using the two-
photon induced fluorescence technique4,14 using a femtosecond
pulsed laser source (see Supporting Information). The TPA
maximum (λTPA) and δ for 1N in toluene (725 nm and 1290 GM,
respectively) are similar to those previously reported for 4,7,12,-
15-tetra(4′-dihexylaminostyryl)-[2.2]paracyclophane (720 nm and
1410 GM).10 The λTPA values for 1N, 2N, and 3N follow the trend
of λabs. Compounds 2N and 3N have higher δ than 1N. Literature
precedent shows that arylamine donor groups in D-π-D structures
can provide for larger15 or similar6b cross sections relative to their
alkylamine counterparts. Higher δ values may be expected on the
basis of additional delocalization within the extended π electron
system.
Within the range of frequencies accessible with our instrumenta-
tion (620-900 nm), the TPA measurements in water show a
substantial decrease in δ, relative to those in toluene (Table 1),
with little change in the λTPA or the general band shape (Supporting
Information). Combining these results with the determination of η
obtained by linear spectroscopy methods provides for δη values
of (in GM) 294 and 359 for 2C and 3C, respectively, which are
the highest reported action cross sections for chromophores in water.
In summary, we provide a synthetic entry to water-soluble
paracyclophane-based TPA chromophores. These molecules have
allowed us to measure a substantial decrease in δ when the solvent
is changed from toluene to water. It has been predicted theoreti-
cally,16 by looking at the effect of external fields, that δ increases
to a maximum point, after which increasing the strength of the field
Supporting Information Available: Synthetic details and char-
acterization for all compounds, TPA measurements, and spectra. This
material is available free of charge via the Internet at http://pubs.acs.org.
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