Efficient synthesis of fluorescent rosamines: Multifunctional platforms for cellular imaging

Article abstract of DOI:10.1016/j.tetlet.2014.01.067

Substituted rosamines are efficiently prepared through a new organometallic addition to an imine-substituted xanthone as a novel primary amine equivalent. The synthesis reduces the number of synthetic steps to the targeted rosamines, for convenient and facile access to potential libraries of rosamine dyes. The prepared rosamine derivatives represent unique multifunctional platforms that possess radiolabeling capability and fluorescence. Rosamines have (i) useful non-specific binding properties in mammalian cells and plant root hair, and (ii) positive uptake or binding properties in microbial systems.

Full text of DOI:10.1016/j.tetlet.2014.01.067

Tetrahedron Letters 55 (2014) 1549–1551
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Efficient synthesis of fluorescent rosamines: multifunctional
platforms for cellular imaging
George A. Kraus a,b, , Tezcan Guney a,b, Aaron Kempema a,b, Joel M. Hyman c, Bahram Parvin c,
⇑
⇑
a Ames Laboratory, US DOE, Iowa State University, Ames, IA 50011, USA
b Department of Chemistry, Iowa State University, Ames, IA 50011, USA
c Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Substituted rosamines are efficiently prepared through a new organometallic addition to an imine-
substituted xanthone as a novel primary amine equivalent. The synthesis reduces the number of syn-
thetic steps to the targeted rosamines, for convenient and facile access to potential libraries of rosamine
dyes. The prepared rosamine derivatives represent unique multifunctional platforms that possess radio-
labeling capability and fluorescence. Rosamines have (i) useful non-specific binding properties in
mammalian cells and plant root hair, and (ii) positive uptake or binding properties in microbial systems.
Ó 2014 Elsevier Ltd. All rights reserved.
Received 6 December 2013
Revised 13 January 2014
Accepted 15 January 2014
Available online 24 January 2014
Keywords:
Rosamine
Fluorescent
Benzophenone imine
Organolithium
Xanthone
Acidic hydrolysis
In recent years, the use of live cell imaging for the understanding
of cellular processes has made large contributions to our under-
standing of living cells.1–3 Compounds with either ‘non-sticky’
properties (e.g., compounds that do not bind specifically and have
balanced intracellular in- and out-flux properties) or those with im-
proved uptake properties are a key part of live cell imaging strate-
gies. Recently, a combinatorial fluorescent library was screened to
identify compounds that can be employed as tags for tracking intra-
cellular localization.4 A number of hits were identified for a variety
of mammalian cells as well as a plant root hair system. In a similar
screen, a small subset of the same chemical library identified ros-
amines represented by 1 (Fig. 1a) that have the dual advantages
of non-stickyness in plant and mammalian species and intracellular
accumulation in microbial isolates. With the potential for imaging
applications,5,6 we selected rosamine derivatives of general struc-
ture 1 for synthesis and radiolabeling. As an extension of our previ-
ous research,7 we developed a concise synthesis of rosamine
analogs represented by 1, which has the potential for radioactive
fluorine incorporation to construct radiolabeled dye 2 (Fig. 1a).
We recognized that capitalizing on a protecting group-free
method to introduce a primary amine would play a key role to
improve efficiency. Figure 1b shows the juxtaposition of the classic
routes8–10 toward rosamines contrasted with our approach to
decrease the total number of steps. Traditional routes toward
rosamines generally utilize a nitro group as in 3 as the primary
amine equivalent which require additional steps to first reduce
a. General structure of targeted rosamines
18F
O
R
H
H
R1
R2
N
H
O
N
N
H
O
N
2
1
b. Synthetic approaches to primary amine functionalized rosamine analogs
1. Reduction
2. Protection
3. Organometallic addition
4. Deprotection
O
R1
R2
O2N
O
N
(3 or 4 steps)
Known
Ar
3
H
R1
N
H
O
N
Organometallic addition/
Acidic hydrolysis
O
R2
5
R1
R2
N
O
N
(1 step)
This work
Ph Ph
4
⇑
Corresponding authors. Tel.: +1 515 294 7794; fax: +1 515 294 0105.
E-mail addresses: gakraus@iastate.edu (G.A. Kraus), b_parvin@lbl.gov (B. Parvin).
Figure 1. Background information about rosamines.
0040-4039/$ - see front matter Ó 2014 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2014.01.067

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G. A. Kraus et al. / Tetrahedron Letters 55 (2014) 1549–1551
Tf2O, Pyr., CH2Cl2
O
O
0 oC to rt, 24 h, 89%
HO
O
OH
TfO
O
OTf
6
7
Piperidine,
DMSO,
rt, 1.5 h, 87%
NH
Ph Ph
Pd(OAc)2 (3 mol %),
rac-BINAP (4.5 mol %),
O
O
Cs2CO3, THF, 65 oC, 16 h, 84%
N
O
N
TfO
O
N
Ph Ph
9
8
Scheme 1. Synthetic steps toward rosamines.
the nitro group to an amine, followed by protection and deprotec-
tion during the organometallic addition. In contrast, by utilizing 4,
our approach effectively decreases the total number of synthetic
steps. Designing a succinct synthesis to rosamine 2 and similar
analogs represented by 5 creates facile access to a class of appeal-
ing multifunctional platforms for cellular imaging.
One way to make these compounds is by the addition of orga-
nometallic species to xanthones.11–13 We initially looked at the po-
tential of organometallic addition to xanthones bearing a NH2
(from hydrolysis of 9) using at least 2 equiv of the aryllithium spe-
cies. This approach afforded 5–10% yields of rosamines, which pre-
sumably arises from the deprotonation of the primary amine. The
deprotonation generates the enolate on the xanthone through
delocalization of the negative charge and has the overall effect of
deactivating the ketone for organometallic addition. As a result,
we decided to mask the amine functionality in our synthesis.
Our synthesis commences with the conversion of 3,6-dihy-
droxyxanthone 612 to bis-triflate 7.14 This is followed by the addi-
tion of one equivalent of piperidine to afford xanthone 8,13 as
depicted in Scheme 1. Amination of triflate 8 was achieved via pal-
ladium catalysis with commercially available benzophenone
imine, adapted from the conditions developed by Buchwald15 to
provide 9 in 84% yield. To the best of our knowledge, the transfor-
mation represents the first example that attaches an imine onto a
xanthone moiety. Overall, the imine adduct 9 functions as a
primary amine equivalent which introduces another method for
rapidly synthesizing substituted rosamines of high interest.16
Continuing the route toward assembling rosamines, unsymmet-
rical xanthone 9 is ideally poised for the critical organometallic
addition. The addition utilizes excess 40-bromo-(1,10-biphenyl)-4-
ol from which the dianion was generated with two equivalents
of n-BuLi. Reaction with 9, followed by acidification with HCl
smoothly provided rosamine 10 in 95% yield, which can potentially
be converted to fluorinated dye 2 by employing [18F]2-fluoroethyl
tosylate. By applying the same strategy of lithium–halogen ex-
change of the bromobiphenyl counterparts to generate aryllithiat-
ed species, analogs 11, 12, and 13 were successfully obtained in
78%, 54%, and 82% yields, respectively. Analogs 11 and 12
(Scheme 2) are potentially convenient substrates for the ensuing
fluorination with an 18F nucleophile.
1. ArLi
THF, -78 oC to rt, 16 h
O
O
Ar
2. 2M HCl
H
N
N
N
H
O
N
Ph Ph
Cl
9
10-13
In addition, the preparation of rosamine 12 revealed that lith-
ium–halogen exchange reactions can be accomplished even in
the presence of a primary tosylate elsewhere on the molecule. Ros-
amine 12 was isolated as an inseparable mixture with a free tosyl-
ate impurity (see 1H NMR spectrum provided in the Supporting
information). The yield of 12 was calculated as 54% by comparing
the integrated 1H NMR peaks ratios of the tosyl CH3 at 2.45 ppm
on rosamine 12 and the corresponding CH3 of the free tosylate at
2.27 ppm (12:ꢀOTs, 2:1). Overall, the tosylated rosamine 12 advan-
tageously includes a better leaving group than the chloride in 11
for further manipulation.
OH
Cl
O
H
N
O
N
H
N
H
O
N
H
Cl
10
Cl
11
95%
78%
O
In conclusion, our newly developed approach to rosamine ana-
logs generates the necessary amine intermediate utilizing a modi-
OTs
F
fied imine functionalization protocol onto
a xanthone. The
O
expeditious strategy allows facile access to the primary amine moi-
ety compared to previous routes. Combining the key organometal-
lic addition to the masked amine in one pot resulted in a flexible
and general synthesis for multifunctional rosamine analog
platforms for cellular imaging.
H
H
N
H
O
Cl
N
N
H
O
Cl
N
Acknowledgments
12
54%
13
82%
This work was funded by the Director, Office of Science, Office
of Biological and Environmental Research, Radiochemistry and
Imaging Instrumentation, of the U.S. Department of Energy under
Scheme 2. Key transformation to obtain rosamines 10–13.

G. A. Kraus et al. / Tetrahedron Letters 55 (2014) 1549–1551
1551
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Contract No. DE-AC02-05CH11231 to the University of California.
The funders had no role in study design, data collection and anal-
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