SCOTfluors: Small, Conjugatable, Orthogonal, and Tunable Fluorophores for In Vivo Imaging of Cell Metabolism

Article abstract of DOI:10.1002/anie.201900465

The transport and trafficking of metabolites are critical for the correct functioning of live cells. However, in situ metabolic imaging studies are hampered by the lack of fluorescent chemical structures that allow direct monitoring of small metabolites u

Full text of DOI:10.1002/anie.201900465

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Accepted Article
Title: SCOTfluors: Small, Conjugatable, Orthogonal and Tunable
Fluorophores for in vivo Imaging of Cell Metabolism
Authors: Sam Benson, Antonio Fernandez, Nicole Barth, Fabio de
Moliner, Mathew Horrocks, C Simon Herrington, Jose Luis
Abad, Antonio Delgado, Lisa Kelly, Ziyuan Chang, Yi Feng,
Miyako Nishiura, Yuichiro Hori, Kazuya Kikuchi, and Marc
Vendrell
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To be cited as: Angew. Chem. Int. Ed. 10.1002/anie.201900465
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SCOTfluors: Small, Conjugatable, Orthogonal and Tunable
Fluorophores for in vivo Imaging of Cell Metabolism
Sam Benson,a,ǂ Antonio Fernandez,a,ǂ Nicole D Barth, Fabio de Moliner, Mathew H Horrocks, C
a
a
b
c
d
d
a
a
a
Simon Herrington, Jose Luis Abad, Antonio Delgado, Lisa Kelly, Ziyuan Chang, Yi Feng, Miyako
e
e
e
a,*
Nishiura, Yuichiro Hori, Kazuya Kikuchi, Marc Vendrell
Abstract: The transport and trafficking of metabolites are critical for
the correct functioning of live cells. However, in situ metabolic
imaging studies are hampered by the lack of fluorescent chemical
structures that allow direct monitoring of small metabolites under
physiological conditions with high spatial and temporal resolution.
Herein we describe SCOTfluors as novel small-sized multi-colored
fluorophores for real-time tracking of essential metabolites in live
cells and in vivo and for the acquisition of metabolic profiles from
human cancer cells of variable origin.
Metabolites are essential biochemical components, with
their transport and localization regulating most biological
functions. Despite advances in fluorescence imaging to label
1
biomolecules, there are few approaches to image in situ small
metabolites in live cells and intact organisms. Most metabolites
do not contain groups that allow direct visualization and need to
be modified with exogenous chromophores. However,
fluorescent labels, in particular red and near-infrared (NIR)
fluorophores, are bulky structures that can impair how
metabolites traffic within cells. Our group has recently developed
fluorogenic amino acids to label peptides without affecting their
2
properties. Herein we describe a new strategy for direct imaging
of essential metabolites in live cells and in vivo using small-sized
multi-color fluorophores.
Figure 1. General synthetic procedure, structures and spectral properties of
SCOTfluors. a Values determined in EtOH. b QY in dioxane using acridine
orange, fluorescein, rhodamine 101 and Cy5 as standards.
Since
the
report
by
Ghosh
and
Whitehouse,3
Several strategies have been described to optimize the optical
properties of fluorophores for live-cell imaging. For instance, the
nitrobenzodioxazole (NBD) has been widely used because of its
small size and neutral character. These properties have
facilitated labeling biomolecules with retention of their native
5
replacement of oxygen atoms with geminal dimethyl groups in
rhodamine and fluorescein produced red-shifted fluorophores
with enhanced properties for bioimaging.6 We envisioned that
the synthesis of nitrobenzodiazoles with different groups
bridging the nitroaminoaniline core would render multi-color
fluorophores with tunable emission and enhanced capabilities
for metabolite imaging in live cells. The preparation of
SCOTfluors was achieved in two synthetic steps from the
common intermediate 1 (Figure 1). First, the aminoaniline core
was cyclized with different bridging groups. All these reactions
proceeded similarly for fluoride and chloride derivatives [full list
of analogues in Supporting Information (SI)]. Second,
halogenated compounds (2, Figure 1) underwent substitution
with primary and secondary amines to render the final
fluorophores (3-7, Figure 1). Triazole derivatives (4) were
synthesized by reaction with sodium nitrite in acidic media at r.t.,
thioderivatives (5) were obtained by condensation with N-
thionylaniline under heating, and selenium analogues (6) were
4
properties. However, NBD (emission ~ 540 nm) is incompatible
with other green fluorescent reporters (e.g., GFP) and has
limited application for in vivo use. To address these
shortcomings, herein we report a collection of fluorophores -
named SCOTfluors- with tunable emission covering the entire
visible spectrum. SCOTfluors include the smallest fluorophores
emitting in the NIR window (650-900 nm) reported to date
(
Figure 1).
[
a]
b]
S. Benson, Dr. A. Fernandez, N. D. Barth, Dr. F. de Moliner, Dr. L.
Kelly, Dr. Z. Chang, Dr. Y. Feng and Dr. M. Vendrell
Centre for Inflammation Research, The University of Edinburgh,
EH16 4TJ Edinburgh, UK. E-mail: marc.vendrell@ed.ac.uk.
Dr. M. H. Horrocks
[
UK Dementia Research Institute and EaStCHEM School of
Chemistry, The University of Edinburgh, EH9 3FJ Edinburgh, UK.
Prof. C. S. Herrington
Edinburgh Cancer Research Centre, EH4 2XR Edinburgh, UK
Prof. J. L. Abad and Prof. A. Delgado
[
c]
d]
[
2
prepared by reaction with SeO under reflux in EtOH. Finally,
Research Unit on Bioactive Molecules, Institute for Advanced
Chemistry of Catalonia, 08034 Barcelona, Spain and University of
Barcelona, Faculty of Pharmacy, Barcelona, Spain.
M. Nishiura, Dr. Y. Hori and Prof. K. Kikuchi
carbon derivatives (3 and 7) were synthesized by Cu-catalyzed
coupling using linear and cyclic ketones, respectively.
[e]
We examined the optical properties of SCOTfluors and
compared them to the original NBD (Figure 1 and S1). With the
exception of triazoles (4), all compounds showed longer
emission wavelengths than NBD, long Stokes shifts (around 80-
Graduate School of Engineering, Osaka University, Suita, Japan.
ǂ
These authors contributed equally to the work.
Supporting information for this article is given via a link at the end of
the document.
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1
00 nm), solvatochromic properties (Figure S2) and good
localization with ER Tracker Green (R=91%) but not
LysoTracker Blue (R=26%). Time-lapse imaging demonstrated
that the ceramide 8 translocated to the recycling lysosomes after
3 h (Figure 2C) as highlighted by the increased co-staining with
LysoTracker Blue (R=75%). Notably, these observations agree
photostability (Figure S3). Among SCOTfluors, Se- and C-
bridged derivatives display red and NIR emission respectively,
likely due to reduced HOMO-LUMO gaps that result in
bathochromic shifts in fluorescence emission, as with
7
9
heteroatom-bridged rhodamine and rhodol fluorophores. To the
with prior reports of lipid mobilization, and could not be obtained
best of our knowledge, this is the first example of C-bridged
nitrobenzodiazoles as fluorophores with NIR emission. Further-
more, C-bridged derivatives are readily accessible via one-step
coupling of aminoaniline 1 with different ketones, representing a
new platform for the direct synthesis of small NIR fluorophores.
SCOTfluors proved compatible for experiments in live cells,
showing no significant cytotoxicity in HeLa cells (Figure S4).
with a fluorescein ceramide analog (Figure S8). These results
confirm the suitability of our approach to prepare neutral NIR-
fluorescent probes to image biolipid function in cells.
We also examined whether SCOTfluors could be used to image
in vivo tissues with high metabolic activity. Fluorescent
deoxyglucose tracers can monitor glucose uptake in metabo-
lically-active cells and tissues,10 although few have been
reported for in vivo use. Herein we synthesized compound 9
(Figure 3I) as an in vivo-compatible glucose analog by
Then, we examined the properties of SCOTfluors for imaging the
trafficking of essential metabolites under physiological
conditions. Sphingolipids are critical components of membranes
in the regulation of cellular metabolism. The dysregulation of
conjugation of the nitrobenzoselenadiazole
6
with 2-
deoxyglucosamine. Of note, we performed the reaction with
chloride and fluoride derivatives of 6 and observed increased
reactivity and recovery for the latter (Figure S9). Compound 9
showed emission around 605 nm with a remarkable Stokes shift
of 115 nm (Figure S10), enabling multiplexed imaging with blue
and green fluorescent proteins (i.e. BFP and GFP, Figure 3A-F).
We examined the transport of 9 in HeLa cells transfected with
EGFP-tagged GLUT4, the main glucose transporter in
mammalian cells. Fluorescence microscopy showed the uptake
sphingolipid metabolism is associated with several diseases
8
(
e.g. Gaucher, Niemann-Pick ) and its intracellular localization is
crucial to understand metabolic disruption. We used the C-
bridged nitrobenzodiazole core to generate the NIR ceramide 8
(
Figure 2A) and monitor its intracellular localization over time by
co-staining with endoplasmic reticulum (ER) and lysosome
markers. Spectral analysis confirmed that the optical properties
of 8 were independent of the sphingoid base, and therefore
could be applicable to several types of biolipids (Figure 2C and
S5). Compound 8 showed insignificant aggregation in water
of
9 in GLUT4-EGFP cells and co-localization with the
transporters (Figure 3A-C). Notably, the uptake of 9 was blocked
by competition with excess glucose (Figure 3D-F) and was
increased by pre-treating HeLa cells with insulin11 (Figure 3H
and S11). These results confirm that compound 9 is a functional
substrate of GLUT4 and that enables dual tracking of glucose
uptake and its transporters under physiological conditions.
Finally, we tested compound 9 in vivo in zebrafish embryos to
visualize regions of high glucose uptake. In vivo administration
and imaging of compound 9 in wildtype zebrafish embryos
indicated bright red fluorescence staining in regions of the
developing brain (e.g. midbrain, hindbrain, Figure 3J), which
express GLUT2 transporters to supply glucose from circulation.12
We confirmed that the staining was dependent on the active
transport of 9 through GLUT2 by examining glut2 morpholino-
injected zebrafish, which have reduced levels of GLUT2. In vivo
images of 9-treated glut2 morpholino-injected zebrafish showed
much weaker fluorescence in the same regions (Figure 3J). We
(
Figure S6) and the incubation with liposomes highlighted its
fluorogenic behavior, with around 15-fold increase in emission
Figure 2B and S7). We exploited this property to visualize the
(
recycling of ceramide 8 in real time in human A549 cells using
fluorescence confocal microscopy.
2
also tested compound 6-NEt as a control and observed no
tissue-specific staining in wildtype or in glut2 morpholino-injected
zebrafish (Figure 3J), highlighting the role of deoxyglucose to
recognize GLUT2 transporters. Altogether, compound 9 can be
used to image glucose uptake in vivo and to perform non-
invasive studies of glucose transport in whole organisms.
Next, we used SCOTfluors to prepare the first red-fluorescent
analogue of lactic acid, an essential metabolite in muscle, blood
and cancer cells. Lactic acid is known as a carbon source in
cancer cells and its uptake in tumours has been recently linked
to aggressive oncological behaviour,13 yet little is known about
its traffic and diffusion inside cancer cells. We conjugated the
Figure 2. A) NIR-fluorescent ceramide 8. B) Emission of compound 8 in PBS
(black) and in phosphatidylcholine:cholesterol (7:1) liposomes (red). C) Time-
nitrobenzoselenadiazole
6
with L-isoserine to produce
lapse confocal microscopy images of A549 cells treated with 8 (50 µM, red),
LysoTracker Blue (magenta) and ER Tracker Green (green) after 15 min (co-
localization: white arrows) and 3 h (co-localization: yellow arrows). Total co-
localization coefficients (R) were determined using ImageJ. Scale bar: 15 µm.
compound 10 (Figure 4A, λem. ~ 605 nm) as a probe to study the
mobilization of lactic acid in live cells. First, we confirmed
increased uptake in hypoxic (1% O ) vs normoxic (20% O ) cells,
2 2
At short times (i.e. 15 min), the ceramide 8 was mainly found at
the Golgi apparatus around the ER, as shown by high co-
since lactic acid can accumulate in environments with low
concentrations of oxygen (Figure 4B).14
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Figure 3. Fluorescence images of GLUT4-EGFP HeLa cells treated with compound 9. A-F) Green (GLUT4-EGFP), red (9, 100 µM) and merged (Hoechst 33342)
images of HeLa cells without additional glucose (A-C) and in media containing 5 mM D-glucose (D-F). White arrows identify co-localization of GLUT4-EGFP and
9. Scale bar: 10 µm. G) Fluorescence emission spectra of BFP (blue), GFP (green) and 9 (red). H) Insulin-dependent (100 nM, 1 h) uptake of 9 (red, 100 µM) in
GLUT4-EGFP HeLa cells. I) Chemical structures of compounds 6-NEt and 9. J) In vivo images of the head in zebrafish embryos (28 hpf) after injection of 6-NEt
2
2
or 9 (both 50 pmol) to the yolk sac (blue arrowheads). Fluorescence images were taken of wildtype zebrafish or zebrafish that had been injected at one cell stage
with 4.2 ng anti-sense glut2 morpholino. Yellow arrows point at midbrain and hindbrain regions within the zebrafish embryo heads. Scale bar: 100 µm.
We performed flow cytometry analysis to observe that hypoxic
cells were significantly brighter than normoxic cells after
incubation with the same concentration of compound 10. We
also performed competition assays between 10 and excess of
lactic acid in normoxic cells, which markedly reduced the
fluorescence staining, suggesting a common transporter for
compound 10 and lactic acid in live cells (Figure 4B).
Encouraged by these results, we used total internal reflection
fluorescence (TIRF) microscopy to image the real-time diffusion
of lactic acid in normoxic and hypoxic cancer cells with super-
resolution. For these studies, we used HeLa cells that had been
treated or not with dimethyloxalylglycine (DMOG), a permeable
prolyl 4-hydroxylase inhibitor that upregulates hypoxia-inducible
factors.
We tracked the paths of over 1,000 individual particles in both
untreated (i.e. normoxic) and DMOG-treated (i.e. hypoxic) cells
after incubation with compound 10 and measured their
respective intracellular diffusion coefficients (Figures 4D-E and
S12, Movies 1-4). Remarkably, particles in hypoxic cells showed
higher mean diffusion coefficients than in normoxic cells, as well
as a reduction of the slow diffusion species. Altogether, these
results suggest that hypoxic tumors might display faster
recycling rates for intracellular lactic acid than normoxic tumors
and prove the utility of compound 10 as a new probe for imaging
lactic acid metabolism in live cells with high spatiotemporal
resolution.
Finally, given the multi-color capabilities and high stability of
SCOTfluors under physiological and oxidative environments
(
Figure S13), we employed them to analyze the metabolic
profiles of human cells from different origin. The groups of
Chang and Rotello previously reported the discrimination of
cancer cells using fluorescent dyes15 or host-guest arrays.16 In
this study, we incubated human cancer cell lines with
compounds 8, 10 and 11 (Figure 5) as respective analogues of
ceramide, lactic acid and glucose in order to obtain metabolic
uptake signatures. First, we plated the cells at similar densities
and incubated them with the probes under the same conditions.
Next, we measured their fluorescence emission in the NIR, red
and green regions to determine their intracellular levels of
ceramide, lactic acid and glucose, respectively. Notably,
different cancer cells presented variability in their metabolic
uptake, as represented by their intracellular glucose-lactate and
ceramide-lactate ratios (Figure 5). Whereas the biological
implications of these results remain to be defined, these results
demonstrate that SCOTfluors can generate multiplexed
metabolic readouts from live cells, which is not possible in other
imaging modalities.
Figure 4. A) Structure of compound 10. B) Fluorescence emission of live cells
after incubation with compound 10 (100 µM) at different oxygen tensions
(normoxia: blue; hypoxia: red) and in normoxic cells after competition with
lactic acid (no lactate: red; 5 mM lactate: blue). Values as means and s.e.m as
error bars. C) Time-lapse TIRF tracking of fluorescent particles in untreated
In conclusion, we developed SCOTfluors as small-sized
fluorophores covering the entire visible spectrum. SCOTfluors
are readily obtained by bridging aminoanilines with different
groups and include the smallest NIR-emitting fluorophores to
date. We validated SCOTfluors for real-time and in situ imaging
of different small metabolites (e.g. lipids, sugars) in live cells and
in vivo, as well as their combination to generate multi-color
(top) and DMOG-treated (10 µM, bottom) HeLa cells after incubation with
compound 10 (100 µM) [Movies 1-2 in SI]. Scale bars: 1 µm. D) Histograms of
the diffusion coefficients of fluorescent particles in normoxic (blue) and hypoxic
(red) cells. Dotted lines delineate fast and slow diffusion species. Mean
diffusion coefficients (D) were determined after averaging the tracks of multiple
particles for each condition (n=1,009 for untreated; n=2,906 for DMOG-
treated).
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fingerprints in cells. The tunability and versatility of SCOTfluors
will enable non-invasive bioimaging studies of essential
metabolites that cannot be performed with conventional
fluorophores.
[4]
a) Jenkinson, D. R.; Cadby, A. J.; Jones, S. Chem. Eur. J. 2017,
23,12585-12592; b) Yamada, K.; Nakata, M.; Horimoto, N.; Saito, M.;
Matsuoka, H.; Inagaki, N. J. Biol. Chem. 2000, 275, 22278-22283.
[5]
[6]
[7]
a) Alamudi, S.; Satapathy, R.; Kim, J.; Su, D.; Ren, H.; Das, R.; Hu, L.;
Alvarado-Martinez, E.; Lee, J.; Hoppmann, C.; Pena-Cabrera, E.; Ha,
H. H.; Park, H. S.; Wang, L.; Chang, Y. T. Nat. Commun. 2016, 7,
1
1964-11973; b) de Moliner, F.; Kielland, N.; Lavilla, R.; Vendrell, M.
Angew. Chem. Int. Ed. 2017, 56, 3758-3769; Angew. Chem. 2017, 129,
812-3823.
3
a) Kolmakov, K.; Belov, V. N.; Wurm, C. A.; Harke, B.; Leutenegger, M.;
Eggeling, C.; Hell, S. W. Eur. J. Org. Chem. 2010, 3593-3610; b)
Grimm, J. B.; Sung, A. J.; Legant, W. R.; Hulamm, P.; Matlosz, S. M.;
Betzig, E.; Lavis, L. D. ACS Chem. Biol. 2013, 8, 1303-1310; c) Jia, S.;
Ramos-Torres, K.M.; Koleman, S.; Ackerman, C. M.; Chang, C. J. ACS
Chem. Biol. 2018, 13, 1844-1852.
a) Calitree, B.; Donnelly, D. J.; Holt, J. J.; Gannon, M. K.; Nygren, C. L.;
Sukumaran, D. K.; Autschbach, J.; Detty, M. R. Organometallics, 2007,
26, 6248-6257; b) Koide, Y.; Urano, Y.; Hanaoka, K.; Terai, T.; Nagano,
T. J. Am. Chem. Soc. 2011, 133, 5680-5682; c) Grimm, J. B.; Sung, A.
J.; Legant, W. R.; Hulamm, P.; Matlosz, S. M.; Betzig, E.; Lavis, L. D.
ACS Chem. Biol. 2013, 8, 1303-1310.
Figure 5. Multi-color metabolic profiles of human cancer cells. Relative
fluorescence intensities of compounds 8, 10 and 11 were measured in
different human cell lines. Emission wavelengths: compound 8 (650 nm),
compound 10 (610 nm), compound 11 (550 nm). Values are presented as
[8]
[9]
Kitatani, K.; Idkowiak-Baldys, J.; Hannun, Y. A. T. Cell Signal. 2008, 20,
1010-1018.
Futerman, A.H. Biochim. Biophys. Acta 2006, 1758, 1885-1892.
6
fluorescence emission per 10 cells after normalization against the
[
10] a) Tian, Y. S.; Lee, H. Y.; Lim, C. S.; Park, J.; Kim, H. M.; Shin, Y. N.,
fluorescence intensity of the respective solutions in aqueous buffer.
Fluorescence fold ratios glucose vs. lactate (left) and ceramide vs. lactate
Kim, E. S.; Jeon, H. J., Park, S. B., Cho, B. R. Angew. Chem. Int. Ed.
2009, 48, 8027-8031; Angew. Chem. 2009, 121, 8171-8175; b) Lee, H.
(right) are calculated for cell type. Data presented as means and s.e.m as
Y.; Lee, J. J.; Park, J.; Park, S. B. Chem. Eur. J. 2011, 17, 143-150.
11] Hirayama, S.; Hori, Y.; Benedek, Z.; Suzuki, T.; Kikuchi, K. Nat. Chem.
Biol. 2016, 12, 853-859.
error bars.
[
[
[
Acknowledgements
The authors acknowledge funding from Medical Research Scotland (S.B.: 879-
12] Marin-Juez, R.; Rovira, M.; Crespo, D.; van der Vaart, M.; Spaink, H.
P.; Planas, J. J. Cereb. Blood Flow Metab. 2015, 35, 74-85.
2
015), MSCA Individual Fellowship (A.F.: 704912), OPTIMA (N.D.B.:
EP/L016559/1), Wellcome Trust Sir Henry Dale Fellowship (Y.F.:
00104/Z/12/Z) and the Spanish Ministry of Science, Innovation and
13] Faubert, B.; Li, K. Y.; Cai, L.; Hensley, C. T.; Kim, J.; Zacharias, L. G.;
Yang, C.; Do, Q. N.; Doucette, S.; Burguete, D.; Li, H.; Huet, G.; Yuan,
Q.; Wigal, T.; Butt, Y.; Ni, M.; Torrealba, J.; Oliver, D.; Lenkinski, R. E.;
Malloy, C. R.; Wachsmann, J. W.; Young, J. D.; Kernstine, K.;
DeBerardinis, R. J. Cell, 2017, 171, 358-371.
1
Universities (J.L.A, A.D.: CTQ2017-85378-R). M.V. acknowledges funds from
ERC Consolidator Grant (771443), Biotechnology and Biological Sciences
Research Council (BB/M025160/1) and the Royal Society (IEC\R3\170132).
The authors thank the technical support from the Flow Cytometry and the
Confocal Advanced Light Microscopy units at the University of Edinburgh.
[
14] Lee, D. C.; Sohn, H. A.; Park, Z. Y.; Oh, S.; Kang, Y. K.; Lee, K. M.;
Kang, M.; Jang, Y. J.; Yang, S. J.; Hong, Y. K.; Noh, H.; Kim, J. A.; Kim,
D. J.; Bae, K. H.; Kim, D. M.; Chung, S. J.; Yoo, H. S.; Yu, D. Y.; Park,
K. C.; Yeom, Y. I. Cell, 2015, 161, 595-609.
Keywords: fluorescence • metabolites • cancer • imaging agents • super
resolution • fingerprints
[
15] Lee, J.-S.; Kim, Y. K.; Kim, H. J.; Hajar, S.; Tan, Y. L.; Kang, N.-Y.; Ng,
S. H.; Yoon, C. N.; Chang, Y. T. Plos One 2012, 7, e32096.
[1]
a) Karton-Lifshin, N.; Segal, E.; Omer, L.; Portnoy, M.; Satchi-Fainaro,
R.; Shabat, D. J. Am. Chem. Soc. 2011, 133, 10960-10965; b) Chan,
J.; Dodani, S. C.; Chang, C. J. Nat. Chem. 2012, 4, 973-984; c)
Kielland, N.; Vendrell, M.; Lavilla, R.; Chang, Y. T. Chem. Commun.
[16] Le, N. D. B.; Yesilbag Tonga, G.; Mout, R.; Kim, S.-T.; Wille, M. E.;
Rana, S.; Dunphy, K. A.; Jerry, D. J.; Yazdani, M.; Ramanathan, R.;
Rotello, C. M.; Rotello, V. M. J. Am. Chem. Soc. 2017, 139, 8008-8012.
2
012, 48, 7401-7403; d) Hong, S. C.; Murale, D. P.; Lee, M.; Lee, S.
M.; Park, J. S.; Lee, J. S. Angew. Chem. Int. Ed. 2017, 56, 14642-
4647; Angew. Chem. 2017, 129, 14834-14839; e) Schischko, A.; Ren,
H.; Kaplaneris, N.; Ackermann, L. Angew. Chem. Int. Ed. 2017, 56,
576-1580; Angew. Chem. 2017, 129, 1598-1602; f) Fernandez, A.;
1
1
Vermeren, M.; Humphries, D.; Subiros-Funosas, R.; Barth, N. D.;
Campana, L.; MacKinnon, A.; Feng, Y.; Vendrell, M. ACS Cent. Sci.
2017, 3, 995-1005; g) Schulte, L. N.; Heinrich, B.; Janga, H.; Schmeck,
B. T.; Vazquez, O. Angew. Chem. Int. Ed. 2018, 57, 11564-11568;
Angew. Chem. 2018, 130, 11738-11742; h) Wang, W.; Lorion, M. M.;
Martinazzoli, O.; Ackermann L. Angew. Chem. Int. Ed. 2018, 57,
10554-10558; Angew. Chem. 2018, 130, 10714-10718.
[2]
a) Mendive-Tapia, L.; Zhao, C.; Akram, A.R.; Preciado, S.; Albericio, F.;
Lee, M.; Serrels, A.; Kielland, N.; Read, N.D.; Lavilla, R.; Vendrell, M.
Nat. Commun. 2016, 7, 10940-10948; b) Mendive-Tapia, L.; Subiros-
Funosas, R.; Zhao, C.; Albericio, F.; Read, N. D.; Lavilla, R.; Vendrell,
M. Nat. Prot. 2017, 12, 1588-1619; c) Subiros-Funosas, R.; Mendive-
Tapia, L.; Sot, J.; Pound, J.; Barth, N. D.; Varela, Y.; Goñi, F.; Paterson,
M.; Gregory, C.; Albericio, F.; Dransfield, I.; Lavilla, R.; Vendrell, M.
Chem. Commun. 2017, 53, 945-948.
[3]
Ghosh, P.B.; Whitehouse, M.W. Biochem. J. 1968, 108, 155-156.
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COMMUNICATION
Small fluorophores
Sam Benson, Antonio
Fernandez, Nicole Barth,
Fabio de Moliner, Mathew
H Horrocks, Simon
Herrington, Jose Luis Abad,
Antonio Delgado, Lisa
Kelly, Ziyuan Chang, Yi
Feng, Miyako Nishiura,
Yuichiro Hori, Kazuya
Kikuchi and Marc Vendrell
Herein we describe a
family of small multi-
color benzodiazole
fluorophores for non-
invasive imaging of
essential metabolites in
live cells and in vivo.
Page No. – Page No.
SCOTfluors: Small,
Conjugatable, Orthogonal
and Tunable
Fluorophores for in vivo
Imaging of Cell
Metabolism
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