Chromo-fluorogenic detection of aldehydes with a rhodamine based sensor featuring an intramolecular deoxylactam

Article abstract of DOI:10.1039/c1ob06448g

A chromogenic and fluorogenic detection of aldehydes was achieved via analyte triggered opening of the deoxylactam of N-(rhodamine B)-deoxylactam-ethylenediamine (dRB-EDA). The utility of the sensor was demonstrated by fluorescent labeling of aldehyde-dis

Full text of DOI:10.1039/c1ob06448g

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry
This journal is © The Royal Society of Chemistry 2011
Chromo-fluorogenic Detection of Aldehydes with a Rhodamine
Based Sensor Featuring an Intramolecular Deoxylactam
Zhu Li,1,2,3 Zhongwei Xue,1,3 Zhisheng Wu,1 Jiahuai Han2 and Shoufa Han1,*
1Department of Chemical Biology, College of Chemistry and Chemical Engineering, and the Key Laboratory for
Chemical Biology of Fujian Province; and 2State Key Laboratory of Cellular Stress Biology and School of Life
Sciences, Xiamen University, Xiamen, Fujian 361005, China; Tel: 86-0592-2181728;
E-mail: shoufa@xmu.edu.cn
Experimental Procedures
Bafilomycin A was purchased from Sigma. All other chemical reagents were obtained from Alfa Aesar
and used without further purification. N-(rhodamine B)-lactam-ethylenediamine was prepared according to
a published procedure.1 N-(rhodamine B)-deoxylactam-amine was prepared according to a literature.2 The
synthesis of N-(rhodamine B)-deoxylactam-2-aminoethanol will be reported elsewhere. Column
chromatography was performed on silica gel (300-400 mesh). NMR spectra (1H-400 MHz and 13C-100
MHz) were recorded on a Bruker instrument using tetramethyl silane as the internal reference. The
fluorescence spectra and Uv-vis absorption spectra were performed on a spectrofluorimeter (Spectamax M5,
Molecular Device) using the excitation wavelength (λex) of 560 nm. Melting points were determined on a
Yanaco MP-500 melting point apparatus，IR spectra were recorded on a Nicolet Avatar 330 FT-IR
spectrophotometer. The mass analysis was performed on Bruker En Apex ultra 7.0T FT-MS.
L929 cells, obtained from American Type Culture Collection, were grown at 37 °C under 5% CO2 in
Dulbecco’s Modified Eagle Medium (DMEM, Gibco; Invitrogen) supplemented with 10% fetal bovine
serium. Confocal microscopic images were obtained on LeicaSP2 using the following filters: λex@560 nm
and λem@580-650 nm. Fluorescence images were merged using Photoshop CS 3.0. Graph by GraphPad
Prism5 software.
Synthesis of dRB-EDA
N
O
N
N
O
N
N
NH2
N
NH2
O
dRB-EDA
RB-EDA
Scheme S1. synthesis of dRB-EDA
N-(rhodamine B)-lactam-ethylenediamine (2 g) was dissolved in anhydrous THF (30 ml). To the solution
was added lithium aluminium hydride (1 g). The mixture was stirred at room temperature under argon
overnight.
1-Butanol (200 ml) was added gradually to the solution to quench residual lithium aluminium
hydride, and then the mixture was washed with water (200 ml). The organic layer was collected, dried over
sodium sulfate, and then concentrated to remove the solvent. The residue was purified by silica gel coloum
chromatography using dichloromethane/hexanes/triethylamine (10:10:1 v/v/v) as the eluent to give 0.8 g of
pale yellow solid as the desired product (40% yield, melting points: 60-61oC). 1H-NMR (400 MHz,
CDCl3), δ:7.35 (d, 1H, J = 7.00 Hz), 7.30~7.25 (m, 1H), 7.20 (t, 1H, J = 7.30 Hz), 6.92 (d, 1H, J = 7.52 Hz),
6.68 (d, 2H, J = 8.72 Hz), 6.37~6.31 (m, 4H), 4.19 (s, 2H), 3.35 (q, 8H, J = 6.98 Hz), 2.58 (t, 2H, J = 5.30
Hz), 2.34 (t, 2H, J = 5.64 Hz), 1.93 (s, 2H), 1.18 (t, 12H, J = 7.02 Hz); 13C-NMR (100 MHz, CDCl3):152.88,
149.46, 147.91, 139.08, 130.37, 127.55, 126.77, 124.59, 121.78, 111.65, 107.52, 97.51, 67.98, 56.08, 51.55,
44.30, 40.12, 12.71 ppm; MS (C30H38N4O): calculated (MH+): 471.3118, found: 471.3122.
Comparison of the fluorescent spectral properties of the colored species in the assay solution with
rhodamine B
Formaldehyde was added into 20 mL of DMF solutions containing dRB-EDA (1 mg ml-1) to a final
concentration of 17.5 mM. The solution was incubated at room temperature for 2 hours and then an aliquot
was taken for analysis by mass spectrometry and fluorescence emission as compared to rhodamine B in DMF
solution.

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry
This journal is © The Royal Society of Chemistry 2011
3000
2000
1000
0
540
570
600
630
Wavelength (nm)
Fig. S1 Fluorescence emission spectra of dRB-EDA assay solution (in dark) as compared to that of
rhodamine B in DMF (in red)
Reaction rates of dRB-EDA in selected solvents
dRB-EDA, was respectively added into various solvents containing formaldehyde (17.5 mM) to a final
concentration of 1 mg ml-1. The rates of color development of the correponding solutions were directly
monitored as function of time by UV-vis absorbance at 560 nm.
4
CH2Cl2
3
CH3CN
DMF+2%H2O
DMF
2
1
0
0
1
2
3
4
Time (hours)
Fig. S2 Effects of solvents on the chromogenic reaction between dRB-EDA and formaldehyde
0.4
A
B
1.5
1.0
0.5
0.0
0.3
0.2
0.1
0
40
80
120
0
120
240
360
Time (minutes)
Time (minutes)
Fig. S3 Time dependant color formation of dRB-EDA (1 mg ml-1) in DMF containing formaldehyde (17.5
mM, A; or 3.5 mM, B).
Fig. S4 Time dependant color formation of dRB-EDA (1 mg ml-1) in DMF containing formaldehyde (0, 3.5, 7,
10.5, 14, 17.5). Incubation time: 0 (A), 0.5 (B), 1 (C), or 1.5 hour (D).

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry
This journal is © The Royal Society of Chemistry 2011
N
O
N
R
N
O
N
N
O
N
R-CHO
N
N
O
NH
NH2
O
O
N
NH
R
RB-EDA
3
Scheme S2. Possible mechanism for nonchromogenic response of RB-EDA with aldehydes.
Differential chromogenic resoponses of dRB-EDA and its structural analogs with formaldehyde in
DMF
dRB-EDA, dRB-EA, dRM-amine and RB-EDA were respectively added into DMF containing formaldehyde
(17.5 mM) to a final concentration of 1 mg ml-1. Using dRB-EDA solution as the control, the rates of color
development of the correponding solutions were directly monitored as function of time by UV-vis
absorbance at 560 nm.
Assay sensitivity of dRB-EDA for formadehyde in DMF:
Formaldehyde was added into a serial of DMF solutions containing dRB-EDA (1 mg ml-1) to a final
concentration of 0, 3.5, 7, 10.5, 14, 17.5, 35 mM. Using dRB-EDA solution as the control, the reaction
solutions incubated at room temperature for about 2 hours and then directly detected by UV-vis absorbance.
Assay sensitivity for aldehydes in DMF
Aliquots of the stock slution of hexanaldehyde, 4-hydroxylbenzaldehyde, D-(+)-glucose, or acetone were
respectively added into a serial of DMF solutions containing dRB-EDA (1 mg ml-1) to various
concentrations. Using dRB-EDA solution as the control, the reaction solutions incubated at room
temperature for about 2 hours and then directly detected by UV-vis absorbance at 560 nm.
0.2
a
b
c
d
0.1
0.0
0
10
20
30
40
Concentration (mM)
Fig. S5 Chromogenic detection of hexanaldehyde (a), 4-hydroxylbenzaldehyde (b), glucose (c) and acetone
(d) with dRB-EDA in DMF. The absorbance of dRB-EDA in the presence of analytes at 560 nm were
respectively recorded. The concentration of the analytes uased was 0, 10, 20, 30, 40 mM as indicated in the
figure.
Labeling of cell-surface sialoproteins with dRB-EDA in live cells
L929 cells, incubated with DMEM containing bafilomycinA1(50nM) for 6 hours, were treated with or
without sodium periodate(1mM, pH 6.8) for 5-20 minutes on ice. Cells were then washed with PBS for 2
times and incubated in dRB-EDA (0.1mg/ml) for 1 minute. Cells were then washed with cold PBS for 3
times and analyzed with Leica SP2 laser confocol scaning microscope.

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry
This journal is © The Royal Society of Chemistry 2011
Et2
N
O
NEt2
NH2
N
7.5
7.0
6.5
6.0
5.5
5.0
4.5
4.0
3.5
3.0
2.5
2.0
1.5
ppm
Fig. S6 1H-NMR spectrum of dRB-EDA
Et2
N
O
NEt2
NH2
N
150
140
130
120
110
100
90
80
70
60
50
40
30
20
10
ppm
Fig. S7 13C-NMR spectrum of dRB-EDA
Fig. S8 Mass spectrometry spectrum of dRB-EDA

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry
This journal is © The Royal Society of Chemistry 2011
Fig. S9 IR spectrum of dRB-EDA
Fig. S10 IR spectrum of RB-EDA
References
1
2
J. S. Wu, I. C. Hwang, K. S. Kim, J. S. Kim, Org. Lett., 2007, 9, 907.
Q. A. Best, R. Xu, M. E. McCarroll, L. Wang, D. J. Dyer, Org. Lett., 2010, 12, 3219.