Accepted Manuscript New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate Fanyong Yan, Tancheng Zheng, Shanshan Guo, Dechao Shi, Ziyi Han, Siyushan Zhou, Li Chen PII: DOI: Reference:
S1386-1425(15)30082-2 http://dx.doi.org/10.1016/j.saa.2015.07.033 SAA 13932
To appear in:
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy
Received Date: Revised Date: Accepted Date:
13 February 2015 4 June 2015 7 July 2015
Please cite this article as: F. Yan, T. Zheng, S. Guo, D. Shi, Z. Han, S. Zhou, L. Chen, New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy (2015), doi: http://dx.doi.org/10.1016/j.saa.2015.07.033
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate *
Fanyong Yan , Tancheng Zheng, Shanshan Guo, Dechao Shi, Ziyi Han, Siyushan Zhou, Li Chen State Key Laboratory of Hollow Fiber Membrane Materials and Processes; Key Lab of Fiber Modification & Functional Fiber of Tianjin; Tianjin Engineering Center for Safety Evaluation of Water Quality & Safeguards Technology; Tianjin Polytechnic University, Tianjin 300387, China *Corresponding author. Tel.: +86 22 83955766; fax: +86 22 83955766. E-mail addresses: [email protected]. Abstract: A bis-rhodamine based fluorescent probe R1 for naked-eye detection of Fe3+ with enhanced sensitivity compared to a mono-rhodamine derivative that shows selectivity for Hg2+, has been synthesized. The 1:1 stoichiometric structure of R1 and Fe3+ is confirmed using a Job’s plot estimation and density functional theory calculations. The reversibility of R1 is verified through its spectral response toward Fe3+ and S2- titration experiments. Using Fe3+ and S2- as chemical inputs and the fluorescence intensity signal as outputs, R1 can be utilized as an INHIBIT logic gate at molecular level. Fluorescent imaging for Fe3+ in living HL-7702 cells have also been successfully performed. Keyword: Fluorescent probe; Rhodamine; Fe3+; Logic gate Introduction The bioavailability of iron is of great interest because all known forms of life need iron and scientists have confirmed that ferric ion performs a key role in many biochemical processes at the cellular level [1-4]. In addition, the ferric ion is well-known as a fluorescence quencher due to its paramagnetic nature, to develop new fluorescent Fe3+ indicators, especially those that present selective Fe3+-amplified emission, is still a challenge. Rhodamine has a time-honored role in sensing [5-14], and its unique properties decide that rhodamine is a promising scaffold for the design of reversible and selective probes. However, most of the rhodamine probes were based on mono-rhodamine. We assumed that a multichannel molecular system that integrated two rhodamine fluorophores into a linker molecule to form a dual-rhodamine, instead of the common mono-rhodamine testing system, would achieve satisfactory ion detection and 1 / 16
imaging assays. To
launch
the
research,
we
employed
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene as a flexible linker to connect two rhodamine units to frame a reversible fluorescent probe for Fe3+ over other metal ions. The reversibility of R1 was realizable by addition of S2-. And using Fe3+ and S2- as chemical inputs and the fluorescence intensity signal as outputs, R1 could be utilized as an INHIBIT logic gate at molecular level. The structure of the complex formed between R1 and Fe3+ ion was enlightened by performing density functional theory (DFT) computations. Experiment Materials and instruments All the materials for synthesis were purchased from commercial suppliers and used without further purification. All reactions were monitored by TLC (thin-layer chromatography) with detection by UV. Deionized water was used throughout the experiment. The absorption spectra were recorded with a Purkinje General TU-1901 UV-vis spectrometer. Hitachi F-4500 fluorescence spectrophotometer was used for fluorescence measurement. 1H-NMR and
13
C-NMR were measured on a Bruker
Avance 600 MHz spectrometer with chemical shifts reported in ppm (in CDCl3; TMS as internal standard). Electrospray ionization (ESI) mass spectra were conducted by ABI-057-TY4675 instrument. All pH were carried out on a PHS-3W pH meter. Fluorescence images of HL-7702 cells were carried out with an Olympus IX71 inverted fluorescence microscope. All measurements were carried out at room temperature. General procedures of metal ion sensing Two millimolar of each inorganic salt (NaNO3, KNO3, Mg(NO3)2·6H2O, Ba(NO3)2, Al(NO3)3·9H2O, Pb(NO3)2, Cu(NO3)2·3H2O, Zn(NO3)2·6H2O, Cd(NO3)2·2H2O, Hg(NO3)2·0.5H2O,
Cr(NO3)3·9H2O,
Fe(NO3)2·6H2O,
Fe(NO3)3·6H2O,
Co(NO3)2·6H2O and Ni(NO3)2·6H2O) was dissolved in deionized water to afford 2 × 10-3 M solution. A 1.0 × 10 -3 M stock solution of R1 was prepared by dissolving R1 in absolute ethanol. HEPES buffer solutions (10 mM, pH 7.0) were prepared by dissolving HEPES in deionized water. The UV-vis and fluorescence spectra were measured in ethanol-HEPES buffer solutions (EtOH/HEPES = 1:9, v/v, pH 7.0). Fluorescence measurements were carried out with excitation and emission slit width of 10 and 5 nm at λex = 520 nm. 2 / 16
Synthesis of probe R1 The synthesis of probe R1 is according to procedure [15] that is depicted in Scheme 1. Rhodamine ethylenediamine (1) was facilely synthesized in high yield according to the
procedures
reported
in
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene
literature (2)
was
[16]. synthesized
according to the reported procedures [17]. A solution containing 5 g of (1) dissolved in 30 mL of ethanol was slowly added into a solution containing 0.6 g of (2) in 20 mL of ethanol and several drops of AcOH. The mixture was heated under reflux for 6 h. After removal of the solvent, the resulting crude product was purified by column chromatography with EtOH/CH2Cl2 (1:10~1:4, v/v). The yield was (1.71 g) 18.5%. 1
H NMR (600 MHz, CDCl3): δ 7.94 (s, 1H), 7.85-7.75 (m, 2H), 7.43-7.29 (m, 4H),
7.02 (m, 2H), 6.33 (m, 13H), 3.35-2.98 (m, 24H), 2.88-2.81 (m, 2H), 2.34 (t, 2H), 1.73-1.53 (m, 2H), 1.09 (t, 24H).
13
C NMR (75 MHz, CDCl3): δ 171.51, 168.13,
165.69, 163.66, 153.38, 148.91, 138.31, 132.67, 130.60, 129.46, 126.31, 125.15, 121.34, 120.11, 108.27, 97.66, 65.35, 44.35, 42.18, 38.04, 29.68, 12.54. ESI-MS [M + H]+: m/z = 1105.5602 (C68H77N8O4Cl). Binding constant The data obtained from fluorescence titration profile were fitted to be a 1:1 binding model according to the following equation.
ଵൗ ଶ
Y୪୧୫ − Y c 1 c 1 ଶ c ቐ1 + Y = Y + + − ቈ൬1 + + ൰ −4 2 c K ୟ c c K ୟ c c
ቑ
The association constant (Ka) is an important parameter, indicating the inclusion capacity of the host-guest complex. The association constant (Ka) can thus be obtained by a nonlinear least’s squares analysis of ∆Y versus [Fe3+], fitting to the experimental data obtained from the fluorescence titrations. Y was the recorded fluorescent intensity, Y0 was the start value without the addition of target molecule, Ylim was the limiting value (left as a floating parameter), CM was the target molecule concentration, and CL was the sensor concentration. Determination of the detection limit [18,19] The detection limit DL of R1 for Fe3+ was determined from the following equation: DL = K*Sb1/S. Where K = 3; Sb1 is the standard deviation of the blank solution; S is the slope of the calibration curve. Quantum chemical computations 3 / 16
All calculations were carried out using the molecular modeling DMol3 and simulated in EtOH environment. The DMol3 method from Material Studio is developed by Accelrys Inc., in which the wave functions are expanded in terms of an accurate numerical basis set. We used BLYP functional and DNP basis sets. The size of the DNP basis set is comparable to Gaussian 6-31G**, but DNP is more accurate than a Gaussian basis set of the same size [20-23]. Cells culture and fluorescence imaging Hepatocyte cell lines (HL-7702 cells) were cultured in Dulbecco’s modified eagle medium (DMEM) with 10% (v/v) fetal bovine serum (FBS) in an atmosphere of 5% CO2 at 37 °C. One day before imaging experiment, the cells were seeded in 24-well plates. The next day, the HL-7702 cells were incubated with 10.0 µM R1 for 30 min and then washed with PBS buffer three times before incubated with 15.0 µM Fe(NO3)3 for 30 min. After washing three times with PBS, the cells were imaged under an inverted fluorescence microscopy (Olympus, Japan). For all images, the microscope settings were held constant. Results and discussion The probe R1 was readily synthesized from the reaction of rhodamine B ethylenediamine with 2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene and characterized by 1 H NMR, mono-rhodamine
probe
13
C NMR and ESI-MS. As a contrast, we synthesized a
R2
with
rhodamine
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene
B
ethylenediamine that
presented
and high
selectivity to Hg2+. The synthesis of R2 and the fluorescent spectra of R2 with Hg2+ were shown in the supporting information. The binding properties of R1 toward different cations (Na+, K+, Mg2+, Ba2+, Al3+, Pb 2+, Cu2+, Zn2+, Cd 2+, Hg2+, Cr3+, Fe3+, Fe2+, Co2+ and Ni2+) were investigated by UV-visible and fluorescence spectroscopy. Fig. 1a displayed the UV-visible spectra changes of probe R1 in the presence and absence of various metal ions. Among the various metal ions examined (10 eq.), R1 showed highly selective “off-on” absorption enhancement only with Fe3+ at 560 nm in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0) solution, and the color of the solution changed from colorless to pink (Fig. 1a insert). It is reliable that R1 serves as a colorimetric probe for the detection of Fe3+ based on the color change. In fluorescent experiments, highest fluorescence intensities at 581 nm of R1-Fe3+ were obtained by excitation at 520 nm (Fig. 1b). In the presence of other metal ions, there was no evident fluorescence intensity enhancement. The 4 / 16
resulted in a prominent change, which can be ascribed to the spirolactam bond cleavage followed by the formation of a delocalized xanthene moiety of the rhodamine group. Achieving high selectivity toward the probe over the other competitive species coexisting in the sample is a very important feature to evaluate the performance of a fluorescence probe. Therefore, competition experiments with high concentrations of the Na+, K+, Mg2+, Ba2+, Al3+, Pb2+, Cu2+, Zn2+, Cd2+, Hg2+, Cr3+, Fe2+, Co2+ and Ni2+ (100 µM) were carried out in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0). No significant variations were observed for the absorption (at λmax 560 nm, Fig. 2a) and the emission (at λmax 581 nm, Fig. 2b) of R1 (10 µM) in the presence of Fe3+ and other metal ions. The addition of 20 µM Fe3+ could significantly enhance the R1 fluorescence, whereas 100 µM solutions of other metal ions did not increase the fluorescence. The competition experiments revealed that the R1 had remarkably high selectivity toward Fe3+ ions over other competitive cations in the water medium. The fluorescent titration studies of R1 with various concentrations of Fe3+ were carried out to explore the profiles of the spectroscopic changes. The solution of R1 turned from colorless to pink upon addition of Fe3+ ions (1-16 µM), and the fluorescence intensity were significantly enhanced with a new peak appearing at 581 nm and reached saturation when addition of 12 µM Fe3+, which were illustrated in Fig. 3. The fluorescence spectra were analyzed using the Job’s plot to determine the binding stoichiometry of R1-Fe3+ complex, by maintaining the total concentration of R1 and Fe3+ constant (10 µM) and changing the mole fraction of Fe3+ from 0 to 1. The fluorescence intensity at 581 nm of the R1-Fe3+ complex was achieved at a mole fraction of approximately 50% of Fe3+ ions, suggesting that a 1:1 stoichiometry was the likely binding mode of complex R1 with Fe3+ ions (Fig. 4). From fluorescence titration results, the binding constant (Ka) of R1 with Fe3+ was estimated to be 1.15 × 107 M-1 based on nonlinear curve fitting of the titration assuming 1:1 stoichiometry (Fig. 5). The detection limit of Fe3+ was 7.3 × 10 -8 M, which makes probe R1 a sensitive tool for the detection of Fe3+ in water. To apply the probe in complex environments, the pH influence on the sensitivity of R1-Fe3+ was tested under various pH conditions. As shown in Fig. 6, R1 was found to be non-fluorescent in EtOH-H2O and showed a positive response (strong fluorescence) between pH 4.0 and 9.0 upon the addition of Fe3+ at 520 nm excitation wavelength. 5 / 16
The results suggested that R1 was insensitive to pH near 7.0, so in this work, a pH value of 7.0 was chosen and maintained with EtOH/HEPES (1:9, v/v, 10 mM, pH 7.0) solution. The effect of reaction time on the present system was investigated and the results were shown in Fig. 7. It can be seen that 20 min later a plateau of fluorescence enhancement was achieved, and 20 min reaction time was selected in subsequent experiments. For better understanding the interaction between R1 and Fe3+, a reversible experiment was performed by alternating addition of Fe3+ and Na2S to the solution of probe R1. When Na2S (3 eq. of Fe3+) was added to the R1-Fe3+ solution, the fluorescent intensity at 581 nm of R1-Fe3+ restored to the original state of R1 (little fluorescence) (Fig. 8a). The stronger affinity between S2- and Fe3+ resulted in the decomplexation of the R1-Fe3+ complex. The fluorescent intensity was revived on further addition of Fe3+ indicating the reversible behavior of R1 for Fe3+. Due to the sequential and reversible recognition behavior of R1 to Fe3+ and S2-, an integrated molecular logic gates can be constructed [24,25]. The corresponding truth values were collected in Fig. 8b. The two input signals were input 1 (Fe3+) and input 2 (S2-). Input 1 elicited strong fluorescence enhancement at 581 nm, equivalent to a YES operation, represent as output ‘1’. On the contrary, input 2 caused fluorescence quenching, implementing the necessary NOT gate, represent as output ‘0’. The receptor acted in parallel on the spectrum output signals, which implements the required AND function. Hence, monitoring the fluorescence enhancement at 581 nm, upon addition of Fe3+ and S2-, and their reacting dose mixture resulted in an INHIBIT logic gate (integrated by combining NOT, YES, and AND gates). For theoretical calculation studies, two plausible structures of the Fe3+ complexes were generated from the available experimental data (Fig. 9). In Fig. 9a, R1 behaved as tridentate ligand using two oxygen atoms from amide carbonyl oxygen group and one nitrogen atom from Schiff base group to bind one Fe3+ ion. While, in Fig. 9b, R1 behaved as quadridentate ligand using two oxygen atoms and two nitrogen atom. The complexes were fully optimized using BLYP function and DNP basis sets as implemented in the program DMol3. In Fig. 9a, the Fe-O bonds length were 1.957 Å, 2.037 Å, which were much shorter than the sum of the Van der Waals radii of Fe and O (3.34 Å), and the Fe-N bond length (1.923 Å) was much shorter than the sum of the Van der Waals radii (3.35 Å). However, when R1 behaved as quadridentate ligand the 6 / 16
SCF was not converging. The results suggest that R1 can provide suitable space to accommodate the Fe3+ ion when it behaves as a tridentate ligand. The highest occupied and the lowest unoccupied molecular orbitals (HOMO and LUMO) of the complex were shown in Fig. 10. It was clearly shown the HOMO distribution
of
the
complex
was
located
essentially
over
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene moiety, while the LUMO was mainly distributed over xanthene ring structure. The energy gap between HOMO and LUMO was computed to be 0.457 eV. With the above data in hand, we next were interested in studying on the Fe3+ bioimaging of R1 in living cells system. HL-7702 cells were used as the living system. After HL-7702 cells were incubated with R1 (10 µM) in culture medium for 30 min at 37 °C, there showed no intracellular fluorescence (Fig. 11b). After further incubation with Fe(NO3)3 for another 30 min at 37 °C, a bright fluorescence appeared from within the cell (Fig. 11d). The bright-field transmission image of cells treated with R1 and Fe3+ (Fig. 11a and 11c) confirmed that the cells were viable throughout the imaging experiments. The results indicated that the R1 was cell permeable and could be used for imaging of Fe3+ in living cells. We also employed probe R1 to determine Fe3+ concentrations in water samples to study the practical applicability in environmental science. First of all, we quantified the fluorescence of R1 (10 µM) in the presence of various concentrations of Fe3+ (1-10 µM) and the corresponding calibration plot was prepared as the standard curve (Fig. S4). The water samples were collected from the tap water and lake water obtained from the university. In table 1, it can be seen that R1 could measure the centration of Fe3+ in water samples with good recovery and R.S.D. results, suggesting the potential application of real sample analysis by R1. Conclusion In summary, we synthesized a new bis-rhodamine-based fluorescent probe R1 for Fe3+. R1 showed excellent selectivity for Fe3+ in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0) over other common cations, such as Na+, K+, Mg2+, Ba2+, Al3+, Pb2+, Cu 2+, Zn2+, Cd2+, Hg2+, Cr3+, Fe2+, Co2+ and Ni2+. The detection limit of R1 for Fe3+ was 7.3 × 10-8 M, which presented a pronounced sensitivity toward Fe3+, and the association constant (Ka) of the complex was 1.15 × 107 M-1. The fluorescent changes of R1 upon the addition of Fe3+ and S2- can be utilized as an INHIBIT logic gate. Optimized geometry and HOMO-LUMO levels of Fe3+ complex of the R1 were computed with 7 / 16
BLYP/DNP. In addition, R1 was further demonstrated to be a potential probe for detecting Fe3+ in living cells. Acknowledgement We are grateful for the financial support from the National Natural Science Foundation of China (Nos. 21174103, 21374078 and 51303132). Electronic Supporting Information The synthesis of R2, 1H NMR,
13
C NMR and MS of R1 and R2, the fluorescent
spectra of R2 with Hg2+ are available. References [1] R.S. Eisenstein, Annu. Rev. Nutr 20 (2000) 627-662. [2] P. Aisen, M.W. Resnick, E.A. Leibold, Curr. Opin. Chem. Biol 3 (1999) 200-206. [3] B. D Autreaux, N.P. Tucker, R. Dixon, S. Spiro, Nature 437 (2005) 769-772. [4] J. W. Lee, J.D. Helmann, Nature 440 (2006) 363-367. [5] D.T. Quang, J.S. Kim, Chem. Rev 110 (2010) 6280-6301. [6] H.N. Kim, M.H. Lee, H.J. Kim, J.S. Kim, J. Yoon, Chem. Soc. Rev 37 (2008) 1465-1472. [7] X. Chen, X. Tian, I. Shin, J. Yoon, Chem. Soc. Rev 40 (2011) 4783-4804. [8] F. Yan, D. Cao, M. Wang, N. Yang, Q. Yu, L. Dai, L. Chen, J. Fluoresc 22 (2012) 1249-1256. [9] F. Yan, D. Cao, N. Yang, Q. Yu, M. Wang, L. Chen, Sensor. Actuat. B-Chem 162 (2012) 313-320. [10] M. Wang, F. Yan, Y. Zou, N. Yang, L. Chen, L. Chen, Spectrochim. Acta A 123 (2014) 216-223. [11] F. Yan, M. Wang, D. Cao, N. Yang, Y. Fu, L. Chen, L. Chen, Dyes Pigm 98 (2013) 42-50. [12] M. Wang, F. Yan, Y. Zou, L. Chen, N. Yang, X. Zhou, Sensor. Actuat. B-Chem 192 (2014) 512-521. [13] F. Yan, D. Cao, N. Yang, M. Wang, L. Dai, C. Li, L. Chen, Spectrochim. Acta A 106 (2013) 19-24. [14] F. Yan, M. Wang, D. Cao, N. Yang, B. Ma, L. Chen, J. Spectrosc 1 (2013) 1-7. [15] T. Tao, E. Kottmair, S.A. Beckley, US20050113546 A1. [16] X. Zhang, Y. Shiraishi, T. Hirai, Org. Lett 9 (2007) 5039-5042. [17] X. Zhao, R.S. Wei, L.G. Chen, D. Jin, X.L. Yan, New. J. Chem 38 (2014) 4791-4798. 8 / 16
[18] M. Zhu, M.J. Yuan, X.F. Liu, J.L. Xu, J. Lv, C. Huang, H. Liu, Y. Li, S. Wang, D. Zhu, Org. Lett 10 (2008) 1481-1484. [19] H.T. Niu, D.D. Su, X.L. Jiang, W.Z. Yang, Z.M. Yin, J.Q. He, J.P. Cheng, Org. Biomol. Chem 6 (2008) 3038-3040. [20] N.A. Benedek, I.K. Snook, K. Latham, I. Yarovsky, J Chem. Phys 122 (2005) 144102. [21] H. Kusama, H. Orita, H. Sugihara, Langmuir 24 (2008) 4411-4419. [22] Y. Inada, H. Orita, J. Comput. Chem 29 (2008) 225-232. [23] C. C. Tsai, I.T. Ho, J.H. Chu, L.C. Shen, S.L. Huang, W.S. Chung, J. Org. Chem 77 (2012) 2254-2262. [24] L.J. Tang, X. Dai, X. Wen, D. Wu, Q. Zhang, Spectrochim. Acta A 139 (2015) 329-334. [25] X. Zhou, X. Wu, J.Y. Yoon, Chem. Comm 51 (2015) 111-113.
9 / 16
Scheme 1. Synthesis of R1.
Fig. 1. (a) UV-vis absorption and (b) Fluorescent spectra of R1 (10 µM) and R1 with cations in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0).
Fig. 2. (a) UV-vis absorption spectra of R1 (10 µM) in the presence of Fe3+ (2.0 eq.) and other metal ions (10.0 eq.) at 560 nm. (b) Fluorescent spectra of R1 (10 µM) in the presence of Fe3+ (2.0 eq.) and other metal ions (10.0 eq.) at 581 nm.
10 / 16
Fig. 3. Emission titration spectra of R1 upon the addition of increasing amounts of Fe3+ (1-16 µM) in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0).
Fig. 4. Job’s plot of R1 with Fe3+. R1+Fe3+ was kept constant at 10 µM.
Fig. 5. Fluorescence titration profile of R1 (10 µM) with Fe3+ ion (0-1.6 eq.), from which the association constant was determined, Ka = 1.15 × 107 M-1 (R = 0.997).
11 / 16
Fig. 6. Fluorescence intensity of probe R1 (10 µM) in the absence and presence of 2 eq. Fe3+ at different pH.
Fig. 7. Effect of reaction time on the fluorescence intensity of R1 (10 µM) in the presence of Fe3+.
Fig. 8. (a) Fluorescence spectrum of R1 (10 µM) in the presence of Fe3+ (20 µM) and S2- (60 µM) in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0). (b) truth table, and logic scheme.
12 / 16
Fig. 9. Optimized geometries of two plausible structures of Fe3+ complexes.
Fig. 10. Calculated HOMO and LUMO of Fe3+ comlplex.
13 / 16
Fig. 11. Fluorescence images of Fe3+ in HL-7702 cells. Bright-field transmission image (a and c) and fluorescence image (b and d) of HL-7702 cells incubated with 0 µM, 15 µM of Fe3+, respectively. Table 1. Determination of Fe3+ in water samples. Sample Tap water
Lake water a
Fe3+ added
Fe3+ found
Recovery (%)
R.S.D.a (%)
(µM)
(µM)
0.0
0.0
-
-
2.0
2.15
107
2.6
4.0
4.10
102
2.1
0.0
7.10
1.0
8.03
93
2.4
n=3.
14 / 16
15 / 16
New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate Fanyong Yan*, Tancheng Zheng, Shanshan Guo, Dechao Shi, Ziyi Han, Siyushan Zhou, Li Chen
Highlights
1. A novel bis-rhodamine probe (R1) is successfully designed. 2. It exhibits an excellent selectivity and a high sensitivity toward Fe3+. 3. Using Fe3+ and S2− as inputs, R1 could be used as an INHIBIT logic gate. 4. The probe has been used for imaging of Fe3+ in living cells.
16 / 16
S1386-1425(15)30082-2 http://dx.doi.org/10.1016/j.saa.2015.07.033 SAA 13932
To appear in:
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy
Received Date: Revised Date: Accepted Date:
13 February 2015 4 June 2015 7 July 2015
Please cite this article as: F. Yan, T. Zheng, S. Guo, D. Shi, Z. Han, S. Zhou, L. Chen, New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy (2015), doi: http://dx.doi.org/10.1016/j.saa.2015.07.033
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate *
Fanyong Yan , Tancheng Zheng, Shanshan Guo, Dechao Shi, Ziyi Han, Siyushan Zhou, Li Chen State Key Laboratory of Hollow Fiber Membrane Materials and Processes; Key Lab of Fiber Modification & Functional Fiber of Tianjin; Tianjin Engineering Center for Safety Evaluation of Water Quality & Safeguards Technology; Tianjin Polytechnic University, Tianjin 300387, China *Corresponding author. Tel.: +86 22 83955766; fax: +86 22 83955766. E-mail addresses: [email protected]. Abstract: A bis-rhodamine based fluorescent probe R1 for naked-eye detection of Fe3+ with enhanced sensitivity compared to a mono-rhodamine derivative that shows selectivity for Hg2+, has been synthesized. The 1:1 stoichiometric structure of R1 and Fe3+ is confirmed using a Job’s plot estimation and density functional theory calculations. The reversibility of R1 is verified through its spectral response toward Fe3+ and S2- titration experiments. Using Fe3+ and S2- as chemical inputs and the fluorescence intensity signal as outputs, R1 can be utilized as an INHIBIT logic gate at molecular level. Fluorescent imaging for Fe3+ in living HL-7702 cells have also been successfully performed. Keyword: Fluorescent probe; Rhodamine; Fe3+; Logic gate Introduction The bioavailability of iron is of great interest because all known forms of life need iron and scientists have confirmed that ferric ion performs a key role in many biochemical processes at the cellular level [1-4]. In addition, the ferric ion is well-known as a fluorescence quencher due to its paramagnetic nature, to develop new fluorescent Fe3+ indicators, especially those that present selective Fe3+-amplified emission, is still a challenge. Rhodamine has a time-honored role in sensing [5-14], and its unique properties decide that rhodamine is a promising scaffold for the design of reversible and selective probes. However, most of the rhodamine probes were based on mono-rhodamine. We assumed that a multichannel molecular system that integrated two rhodamine fluorophores into a linker molecule to form a dual-rhodamine, instead of the common mono-rhodamine testing system, would achieve satisfactory ion detection and 1 / 16
imaging assays. To
launch
the
research,
we
employed
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene as a flexible linker to connect two rhodamine units to frame a reversible fluorescent probe for Fe3+ over other metal ions. The reversibility of R1 was realizable by addition of S2-. And using Fe3+ and S2- as chemical inputs and the fluorescence intensity signal as outputs, R1 could be utilized as an INHIBIT logic gate at molecular level. The structure of the complex formed between R1 and Fe3+ ion was enlightened by performing density functional theory (DFT) computations. Experiment Materials and instruments All the materials for synthesis were purchased from commercial suppliers and used without further purification. All reactions were monitored by TLC (thin-layer chromatography) with detection by UV. Deionized water was used throughout the experiment. The absorption spectra were recorded with a Purkinje General TU-1901 UV-vis spectrometer. Hitachi F-4500 fluorescence spectrophotometer was used for fluorescence measurement. 1H-NMR and
13
C-NMR were measured on a Bruker
Avance 600 MHz spectrometer with chemical shifts reported in ppm (in CDCl3; TMS as internal standard). Electrospray ionization (ESI) mass spectra were conducted by ABI-057-TY4675 instrument. All pH were carried out on a PHS-3W pH meter. Fluorescence images of HL-7702 cells were carried out with an Olympus IX71 inverted fluorescence microscope. All measurements were carried out at room temperature. General procedures of metal ion sensing Two millimolar of each inorganic salt (NaNO3, KNO3, Mg(NO3)2·6H2O, Ba(NO3)2, Al(NO3)3·9H2O, Pb(NO3)2, Cu(NO3)2·3H2O, Zn(NO3)2·6H2O, Cd(NO3)2·2H2O, Hg(NO3)2·0.5H2O,
Cr(NO3)3·9H2O,
Fe(NO3)2·6H2O,
Fe(NO3)3·6H2O,
Co(NO3)2·6H2O and Ni(NO3)2·6H2O) was dissolved in deionized water to afford 2 × 10-3 M solution. A 1.0 × 10 -3 M stock solution of R1 was prepared by dissolving R1 in absolute ethanol. HEPES buffer solutions (10 mM, pH 7.0) were prepared by dissolving HEPES in deionized water. The UV-vis and fluorescence spectra were measured in ethanol-HEPES buffer solutions (EtOH/HEPES = 1:9, v/v, pH 7.0). Fluorescence measurements were carried out with excitation and emission slit width of 10 and 5 nm at λex = 520 nm. 2 / 16
Synthesis of probe R1 The synthesis of probe R1 is according to procedure [15] that is depicted in Scheme 1. Rhodamine ethylenediamine (1) was facilely synthesized in high yield according to the
procedures
reported
in
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene
literature (2)
was
[16]. synthesized
according to the reported procedures [17]. A solution containing 5 g of (1) dissolved in 30 mL of ethanol was slowly added into a solution containing 0.6 g of (2) in 20 mL of ethanol and several drops of AcOH. The mixture was heated under reflux for 6 h. After removal of the solvent, the resulting crude product was purified by column chromatography with EtOH/CH2Cl2 (1:10~1:4, v/v). The yield was (1.71 g) 18.5%. 1
H NMR (600 MHz, CDCl3): δ 7.94 (s, 1H), 7.85-7.75 (m, 2H), 7.43-7.29 (m, 4H),
7.02 (m, 2H), 6.33 (m, 13H), 3.35-2.98 (m, 24H), 2.88-2.81 (m, 2H), 2.34 (t, 2H), 1.73-1.53 (m, 2H), 1.09 (t, 24H).
13
C NMR (75 MHz, CDCl3): δ 171.51, 168.13,
165.69, 163.66, 153.38, 148.91, 138.31, 132.67, 130.60, 129.46, 126.31, 125.15, 121.34, 120.11, 108.27, 97.66, 65.35, 44.35, 42.18, 38.04, 29.68, 12.54. ESI-MS [M + H]+: m/z = 1105.5602 (C68H77N8O4Cl). Binding constant The data obtained from fluorescence titration profile were fitted to be a 1:1 binding model according to the following equation.
ଵൗ ଶ
Y୪୧୫ − Y c 1 c 1 ଶ c ቐ1 + Y = Y + + − ቈ൬1 + + ൰ −4 2 c K ୟ c c K ୟ c c
ቑ
The association constant (Ka) is an important parameter, indicating the inclusion capacity of the host-guest complex. The association constant (Ka) can thus be obtained by a nonlinear least’s squares analysis of ∆Y versus [Fe3+], fitting to the experimental data obtained from the fluorescence titrations. Y was the recorded fluorescent intensity, Y0 was the start value without the addition of target molecule, Ylim was the limiting value (left as a floating parameter), CM was the target molecule concentration, and CL was the sensor concentration. Determination of the detection limit [18,19] The detection limit DL of R1 for Fe3+ was determined from the following equation: DL = K*Sb1/S. Where K = 3; Sb1 is the standard deviation of the blank solution; S is the slope of the calibration curve. Quantum chemical computations 3 / 16
All calculations were carried out using the molecular modeling DMol3 and simulated in EtOH environment. The DMol3 method from Material Studio is developed by Accelrys Inc., in which the wave functions are expanded in terms of an accurate numerical basis set. We used BLYP functional and DNP basis sets. The size of the DNP basis set is comparable to Gaussian 6-31G**, but DNP is more accurate than a Gaussian basis set of the same size [20-23]. Cells culture and fluorescence imaging Hepatocyte cell lines (HL-7702 cells) were cultured in Dulbecco’s modified eagle medium (DMEM) with 10% (v/v) fetal bovine serum (FBS) in an atmosphere of 5% CO2 at 37 °C. One day before imaging experiment, the cells were seeded in 24-well plates. The next day, the HL-7702 cells were incubated with 10.0 µM R1 for 30 min and then washed with PBS buffer three times before incubated with 15.0 µM Fe(NO3)3 for 30 min. After washing three times with PBS, the cells were imaged under an inverted fluorescence microscopy (Olympus, Japan). For all images, the microscope settings were held constant. Results and discussion The probe R1 was readily synthesized from the reaction of rhodamine B ethylenediamine with 2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene and characterized by 1 H NMR, mono-rhodamine
probe
13
C NMR and ESI-MS. As a contrast, we synthesized a
R2
with
rhodamine
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene
B
ethylenediamine that
presented
and high
selectivity to Hg2+. The synthesis of R2 and the fluorescent spectra of R2 with Hg2+ were shown in the supporting information. The binding properties of R1 toward different cations (Na+, K+, Mg2+, Ba2+, Al3+, Pb 2+, Cu2+, Zn2+, Cd 2+, Hg2+, Cr3+, Fe3+, Fe2+, Co2+ and Ni2+) were investigated by UV-visible and fluorescence spectroscopy. Fig. 1a displayed the UV-visible spectra changes of probe R1 in the presence and absence of various metal ions. Among the various metal ions examined (10 eq.), R1 showed highly selective “off-on” absorption enhancement only with Fe3+ at 560 nm in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0) solution, and the color of the solution changed from colorless to pink (Fig. 1a insert). It is reliable that R1 serves as a colorimetric probe for the detection of Fe3+ based on the color change. In fluorescent experiments, highest fluorescence intensities at 581 nm of R1-Fe3+ were obtained by excitation at 520 nm (Fig. 1b). In the presence of other metal ions, there was no evident fluorescence intensity enhancement. The 4 / 16
resulted in a prominent change, which can be ascribed to the spirolactam bond cleavage followed by the formation of a delocalized xanthene moiety of the rhodamine group. Achieving high selectivity toward the probe over the other competitive species coexisting in the sample is a very important feature to evaluate the performance of a fluorescence probe. Therefore, competition experiments with high concentrations of the Na+, K+, Mg2+, Ba2+, Al3+, Pb2+, Cu2+, Zn2+, Cd2+, Hg2+, Cr3+, Fe2+, Co2+ and Ni2+ (100 µM) were carried out in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0). No significant variations were observed for the absorption (at λmax 560 nm, Fig. 2a) and the emission (at λmax 581 nm, Fig. 2b) of R1 (10 µM) in the presence of Fe3+ and other metal ions. The addition of 20 µM Fe3+ could significantly enhance the R1 fluorescence, whereas 100 µM solutions of other metal ions did not increase the fluorescence. The competition experiments revealed that the R1 had remarkably high selectivity toward Fe3+ ions over other competitive cations in the water medium. The fluorescent titration studies of R1 with various concentrations of Fe3+ were carried out to explore the profiles of the spectroscopic changes. The solution of R1 turned from colorless to pink upon addition of Fe3+ ions (1-16 µM), and the fluorescence intensity were significantly enhanced with a new peak appearing at 581 nm and reached saturation when addition of 12 µM Fe3+, which were illustrated in Fig. 3. The fluorescence spectra were analyzed using the Job’s plot to determine the binding stoichiometry of R1-Fe3+ complex, by maintaining the total concentration of R1 and Fe3+ constant (10 µM) and changing the mole fraction of Fe3+ from 0 to 1. The fluorescence intensity at 581 nm of the R1-Fe3+ complex was achieved at a mole fraction of approximately 50% of Fe3+ ions, suggesting that a 1:1 stoichiometry was the likely binding mode of complex R1 with Fe3+ ions (Fig. 4). From fluorescence titration results, the binding constant (Ka) of R1 with Fe3+ was estimated to be 1.15 × 107 M-1 based on nonlinear curve fitting of the titration assuming 1:1 stoichiometry (Fig. 5). The detection limit of Fe3+ was 7.3 × 10 -8 M, which makes probe R1 a sensitive tool for the detection of Fe3+ in water. To apply the probe in complex environments, the pH influence on the sensitivity of R1-Fe3+ was tested under various pH conditions. As shown in Fig. 6, R1 was found to be non-fluorescent in EtOH-H2O and showed a positive response (strong fluorescence) between pH 4.0 and 9.0 upon the addition of Fe3+ at 520 nm excitation wavelength. 5 / 16
The results suggested that R1 was insensitive to pH near 7.0, so in this work, a pH value of 7.0 was chosen and maintained with EtOH/HEPES (1:9, v/v, 10 mM, pH 7.0) solution. The effect of reaction time on the present system was investigated and the results were shown in Fig. 7. It can be seen that 20 min later a plateau of fluorescence enhancement was achieved, and 20 min reaction time was selected in subsequent experiments. For better understanding the interaction between R1 and Fe3+, a reversible experiment was performed by alternating addition of Fe3+ and Na2S to the solution of probe R1. When Na2S (3 eq. of Fe3+) was added to the R1-Fe3+ solution, the fluorescent intensity at 581 nm of R1-Fe3+ restored to the original state of R1 (little fluorescence) (Fig. 8a). The stronger affinity between S2- and Fe3+ resulted in the decomplexation of the R1-Fe3+ complex. The fluorescent intensity was revived on further addition of Fe3+ indicating the reversible behavior of R1 for Fe3+. Due to the sequential and reversible recognition behavior of R1 to Fe3+ and S2-, an integrated molecular logic gates can be constructed [24,25]. The corresponding truth values were collected in Fig. 8b. The two input signals were input 1 (Fe3+) and input 2 (S2-). Input 1 elicited strong fluorescence enhancement at 581 nm, equivalent to a YES operation, represent as output ‘1’. On the contrary, input 2 caused fluorescence quenching, implementing the necessary NOT gate, represent as output ‘0’. The receptor acted in parallel on the spectrum output signals, which implements the required AND function. Hence, monitoring the fluorescence enhancement at 581 nm, upon addition of Fe3+ and S2-, and their reacting dose mixture resulted in an INHIBIT logic gate (integrated by combining NOT, YES, and AND gates). For theoretical calculation studies, two plausible structures of the Fe3+ complexes were generated from the available experimental data (Fig. 9). In Fig. 9a, R1 behaved as tridentate ligand using two oxygen atoms from amide carbonyl oxygen group and one nitrogen atom from Schiff base group to bind one Fe3+ ion. While, in Fig. 9b, R1 behaved as quadridentate ligand using two oxygen atoms and two nitrogen atom. The complexes were fully optimized using BLYP function and DNP basis sets as implemented in the program DMol3. In Fig. 9a, the Fe-O bonds length were 1.957 Å, 2.037 Å, which were much shorter than the sum of the Van der Waals radii of Fe and O (3.34 Å), and the Fe-N bond length (1.923 Å) was much shorter than the sum of the Van der Waals radii (3.35 Å). However, when R1 behaved as quadridentate ligand the 6 / 16
SCF was not converging. The results suggest that R1 can provide suitable space to accommodate the Fe3+ ion when it behaves as a tridentate ligand. The highest occupied and the lowest unoccupied molecular orbitals (HOMO and LUMO) of the complex were shown in Fig. 10. It was clearly shown the HOMO distribution
of
the
complex
was
located
essentially
over
the
2-cholro-1-formyl-3-(hydroxymethylene)-1-cyclohexene moiety, while the LUMO was mainly distributed over xanthene ring structure. The energy gap between HOMO and LUMO was computed to be 0.457 eV. With the above data in hand, we next were interested in studying on the Fe3+ bioimaging of R1 in living cells system. HL-7702 cells were used as the living system. After HL-7702 cells were incubated with R1 (10 µM) in culture medium for 30 min at 37 °C, there showed no intracellular fluorescence (Fig. 11b). After further incubation with Fe(NO3)3 for another 30 min at 37 °C, a bright fluorescence appeared from within the cell (Fig. 11d). The bright-field transmission image of cells treated with R1 and Fe3+ (Fig. 11a and 11c) confirmed that the cells were viable throughout the imaging experiments. The results indicated that the R1 was cell permeable and could be used for imaging of Fe3+ in living cells. We also employed probe R1 to determine Fe3+ concentrations in water samples to study the practical applicability in environmental science. First of all, we quantified the fluorescence of R1 (10 µM) in the presence of various concentrations of Fe3+ (1-10 µM) and the corresponding calibration plot was prepared as the standard curve (Fig. S4). The water samples were collected from the tap water and lake water obtained from the university. In table 1, it can be seen that R1 could measure the centration of Fe3+ in water samples with good recovery and R.S.D. results, suggesting the potential application of real sample analysis by R1. Conclusion In summary, we synthesized a new bis-rhodamine-based fluorescent probe R1 for Fe3+. R1 showed excellent selectivity for Fe3+ in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0) over other common cations, such as Na+, K+, Mg2+, Ba2+, Al3+, Pb2+, Cu 2+, Zn2+, Cd2+, Hg2+, Cr3+, Fe2+, Co2+ and Ni2+. The detection limit of R1 for Fe3+ was 7.3 × 10-8 M, which presented a pronounced sensitivity toward Fe3+, and the association constant (Ka) of the complex was 1.15 × 107 M-1. The fluorescent changes of R1 upon the addition of Fe3+ and S2- can be utilized as an INHIBIT logic gate. Optimized geometry and HOMO-LUMO levels of Fe3+ complex of the R1 were computed with 7 / 16
BLYP/DNP. In addition, R1 was further demonstrated to be a potential probe for detecting Fe3+ in living cells. Acknowledgement We are grateful for the financial support from the National Natural Science Foundation of China (Nos. 21174103, 21374078 and 51303132). Electronic Supporting Information The synthesis of R2, 1H NMR,
13
C NMR and MS of R1 and R2, the fluorescent
spectra of R2 with Hg2+ are available. References [1] R.S. Eisenstein, Annu. Rev. Nutr 20 (2000) 627-662. [2] P. Aisen, M.W. Resnick, E.A. Leibold, Curr. Opin. Chem. Biol 3 (1999) 200-206. [3] B. D Autreaux, N.P. Tucker, R. Dixon, S. Spiro, Nature 437 (2005) 769-772. [4] J. W. Lee, J.D. Helmann, Nature 440 (2006) 363-367. [5] D.T. Quang, J.S. Kim, Chem. Rev 110 (2010) 6280-6301. [6] H.N. Kim, M.H. Lee, H.J. Kim, J.S. Kim, J. Yoon, Chem. Soc. Rev 37 (2008) 1465-1472. [7] X. Chen, X. Tian, I. Shin, J. Yoon, Chem. Soc. Rev 40 (2011) 4783-4804. [8] F. Yan, D. Cao, M. Wang, N. Yang, Q. Yu, L. Dai, L. Chen, J. Fluoresc 22 (2012) 1249-1256. [9] F. Yan, D. Cao, N. Yang, Q. Yu, M. Wang, L. Chen, Sensor. Actuat. B-Chem 162 (2012) 313-320. [10] M. Wang, F. Yan, Y. Zou, N. Yang, L. Chen, L. Chen, Spectrochim. Acta A 123 (2014) 216-223. [11] F. Yan, M. Wang, D. Cao, N. Yang, Y. Fu, L. Chen, L. Chen, Dyes Pigm 98 (2013) 42-50. [12] M. Wang, F. Yan, Y. Zou, L. Chen, N. Yang, X. Zhou, Sensor. Actuat. B-Chem 192 (2014) 512-521. [13] F. Yan, D. Cao, N. Yang, M. Wang, L. Dai, C. Li, L. Chen, Spectrochim. Acta A 106 (2013) 19-24. [14] F. Yan, M. Wang, D. Cao, N. Yang, B. Ma, L. Chen, J. Spectrosc 1 (2013) 1-7. [15] T. Tao, E. Kottmair, S.A. Beckley, US20050113546 A1. [16] X. Zhang, Y. Shiraishi, T. Hirai, Org. Lett 9 (2007) 5039-5042. [17] X. Zhao, R.S. Wei, L.G. Chen, D. Jin, X.L. Yan, New. J. Chem 38 (2014) 4791-4798. 8 / 16
[18] M. Zhu, M.J. Yuan, X.F. Liu, J.L. Xu, J. Lv, C. Huang, H. Liu, Y. Li, S. Wang, D. Zhu, Org. Lett 10 (2008) 1481-1484. [19] H.T. Niu, D.D. Su, X.L. Jiang, W.Z. Yang, Z.M. Yin, J.Q. He, J.P. Cheng, Org. Biomol. Chem 6 (2008) 3038-3040. [20] N.A. Benedek, I.K. Snook, K. Latham, I. Yarovsky, J Chem. Phys 122 (2005) 144102. [21] H. Kusama, H. Orita, H. Sugihara, Langmuir 24 (2008) 4411-4419. [22] Y. Inada, H. Orita, J. Comput. Chem 29 (2008) 225-232. [23] C. C. Tsai, I.T. Ho, J.H. Chu, L.C. Shen, S.L. Huang, W.S. Chung, J. Org. Chem 77 (2012) 2254-2262. [24] L.J. Tang, X. Dai, X. Wen, D. Wu, Q. Zhang, Spectrochim. Acta A 139 (2015) 329-334. [25] X. Zhou, X. Wu, J.Y. Yoon, Chem. Comm 51 (2015) 111-113.
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Scheme 1. Synthesis of R1.
Fig. 1. (a) UV-vis absorption and (b) Fluorescent spectra of R1 (10 µM) and R1 with cations in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0).
Fig. 2. (a) UV-vis absorption spectra of R1 (10 µM) in the presence of Fe3+ (2.0 eq.) and other metal ions (10.0 eq.) at 560 nm. (b) Fluorescent spectra of R1 (10 µM) in the presence of Fe3+ (2.0 eq.) and other metal ions (10.0 eq.) at 581 nm.
10 / 16
Fig. 3. Emission titration spectra of R1 upon the addition of increasing amounts of Fe3+ (1-16 µM) in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0).
Fig. 4. Job’s plot of R1 with Fe3+. R1+Fe3+ was kept constant at 10 µM.
Fig. 5. Fluorescence titration profile of R1 (10 µM) with Fe3+ ion (0-1.6 eq.), from which the association constant was determined, Ka = 1.15 × 107 M-1 (R = 0.997).
11 / 16
Fig. 6. Fluorescence intensity of probe R1 (10 µM) in the absence and presence of 2 eq. Fe3+ at different pH.
Fig. 7. Effect of reaction time on the fluorescence intensity of R1 (10 µM) in the presence of Fe3+.
Fig. 8. (a) Fluorescence spectrum of R1 (10 µM) in the presence of Fe3+ (20 µM) and S2- (60 µM) in 1:9 EtOH/HEPES buffer (v/v, 10 mM, pH 7.0). (b) truth table, and logic scheme.
12 / 16
Fig. 9. Optimized geometries of two plausible structures of Fe3+ complexes.
Fig. 10. Calculated HOMO and LUMO of Fe3+ comlplex.
13 / 16
Fig. 11. Fluorescence images of Fe3+ in HL-7702 cells. Bright-field transmission image (a and c) and fluorescence image (b and d) of HL-7702 cells incubated with 0 µM, 15 µM of Fe3+, respectively. Table 1. Determination of Fe3+ in water samples. Sample Tap water
Lake water a
Fe3+ added
Fe3+ found
Recovery (%)
R.S.D.a (%)
(µM)
(µM)
0.0
0.0
-
-
2.0
2.15
107
2.6
4.0
4.10
102
2.1
0.0
7.10
1.0
8.03
93
2.4
n=3.
14 / 16
15 / 16
New fluorescence probe for Fe3+ with bis-rhodamine and its application as a molecular logic gate Fanyong Yan*, Tancheng Zheng, Shanshan Guo, Dechao Shi, Ziyi Han, Siyushan Zhou, Li Chen
Highlights
1. A novel bis-rhodamine probe (R1) is successfully designed. 2. It exhibits an excellent selectivity and a high sensitivity toward Fe3+. 3. Using Fe3+ and S2− as inputs, R1 could be used as an INHIBIT logic gate. 4. The probe has been used for imaging of Fe3+ in living cells.
16 / 16
