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Surfaces, Interfaces, and Catalysis; Physical Properties of Nanomaterials and Materials
Carbon Dot with pH Independent Near Unity PLQY in Aqueous Medium: Electrostatics Induced FRET at Sub-Micromolar Concentration Ananya Das, Debjit Roy, Mrinal Mandal, Chitra Jaiswal, Malancha Ta, and Prasun K. Mandal J. Phys. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acs.jpclett.8b02193 • Publication Date (Web): 19 Aug 2018 Downloaded from http://pubs.acs.org on August 19, 2018
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Carbon Dot with pH Independent Near Unity PLQY in Aqueous Medium: Electrostatics Induced FRET at Sub-Micromolar Concentration Ananya Das,a Debjit Roy,a Mrinal Mandal,a Chitra Jaiswal,b Malancha Ta,b Prasun K. Mandal a,c,* a
Department of Chemical Sciences, bDepartment of Biological Sciences, cCentre for Advanced Functional Materials, Indian Institute of Science Education and Research (IISER) - Kolkata, Mohanpur, West-Bengal, 741246, India. Supporting Information
ABSTRACT: Hereby, we report the synthesis, and dynamical behaviour of carbon dot (CD) with near 100% PLQY in water for a very large range of pH (1-12). This CD exhibits rotational correlational time of only ~ 130 ps, signifying the whole CD is not exhibiting PL. Unlike most of the carbon based nano particles (acting as quencher of fluorescence) this CD could act as a donor and Förster model could account for the experimental observables for the resonance energy transfer (RET) experiment quite well. Based on two dynamical measurements it could be shown that the fluorescing moiety is located inside the core of the CD. Importantly, for this CD, RET experiment could be performed with very low concentration (500 nM) of the acceptor. This kind of electrostatics driven RET at very low concentration is quite important in bio-imaging. This ultrabright CD is nontoxic and useful for bioimaging in mesenchymal stem cells.
The quest for making a fluorophore or photoluminescence nanomaterial with near unity photoluminescence quantum yield (PLQY) specially in water is extremely important, however highly challenging and hence extremely demanding among researchers. Most of the very bright semi-conductor quantum dots (QDs) have toxic metals like Cd or Pb and exhibit feeble photoluminescence in water.1 Carbon dots (CDs) are gaining increasing importance as photo-luminescent material over many other nanomaterials towards bio-imaging.2-10 Most of the CDs have been synthesized from citric acid 11-14 and either toxic ethylenediamine or ethanolamine. Photoluminescence quantum yield (PLQY) of CDs is an important optical parameter for the usage of CD as LEDs and in bio-imaging. Most of the CDs reported so far possess low PLQY,15-22 specially in aqueous medium,17 hence, their usage in bio-imaging is quite restricted. Most of the CDs exhibit excitation wavelength dependent emission behavior.17-30 Most of the CDs exhibit multi-exponential PL decay.6,7,31,32 All the above mentioned properties restrict the usage of CD quite significantly. QDs have been considered as bright photo-luminescent material and used in the resonance energy transfer (RET) process towards probing different biomolecular processes.33,34 QDs with the surrounding ligands are electrically neutral and requires very high concentration (mM) of acceptor molecules for the observation of RET. Such high concentration could be quite detrimental in the biological medium. Hence, it is quite necessary to find highly photoluminescent (in aqueous medium), highly photostable, non-toxic nanomaterial of size less than 10 nm which can take part in RET process with acceptor concentration in the
range of nM to a few µM. It is well known that electrostatics plays an important role towards controlling the interaction between nanomaterials and biomolecules.35 If CDs can be made highly photoluminescent and highly photostable and with surface charge (positive or negative) then, electrostatic effect can be utilized to adhere oppositely charged acceptor dyes with necessary RET requirements. Then it should be possible to observe RET process with acceptor concentration in the range of nM to a few µM. It would also be interesting to investigate if Förster method can account for the experimental observables of the RET process. In order to achieve these very difficult targets mentioned in first paragraph one has to consider several modifications. For example it is quite important to synthesize highly luminescent CDs in aqueous medium starting from non-toxic precursors. It is necessary that the CDs exhibit excitation wavelength independent emission as well as single exponential PL decay with long excited state lifetime. As CDs have been extensively used in bio-imaging, hence, it is also important to know whether CDs can be employed in the resonance energy transfer (RET) experiments and whether Förster model can explain the experimental observables pertinent to RET investigations. In the current effort we could achieve all the above-mentioned novel properties for the CD.
Scheme 1: Synthesis scheme of CD from non-toxic precursors
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Figure 1. FTIR (a), Raman spectroscopy (b), PXRD (c), TEM image, HRTEM (inset), size distribution (inset) (d), Hydrodynamic diameter in aqueous medium from DLS measurement (e), Zeta potential (f) of CD. Characterization. CD has been synthesized starting from nontoxic precursors like citric acid and TRIS (Scheme 1). The most bright spot has been separated by column chromatography (see methods section for details). FTIR spectrum of purified CD is depicted in Figure 1a confirms the presence of functional groups such as –O-H (stretching, peak at around 3417 cm-1), sp3-C-H (asymmetric and symmetric stretching at 2977 and 2888 cm-1 respectively). The strong signal peaks at 1671, 1647 cm-1 could be attributed to >C=O stretching of -CONH. The signal peaks at 1578, 1460,1419 cm-1 could be assigned to stretching band of sp2 carbon atoms (-C=C-); the peaks at 1384, 1260, 1126, 1042 and 1014 cm-1 could be assigned to different modes of –C-N- and –CO- groups present in CDs. The Raman spectrum (Figure 1b) depicts two broad peaks at around 1340 cm-1 and 1616 cm-1 which can be attributed to D band (sp3) and G band (sp2) respectively. Elemental analysis, 1HNMR and 13CNMR spectra for the CD have been measured (Supporting information Figure S1 and S2). Regarding the functional groups present Raman, FTIR and NMR spectra are consonant with each other (Supporting information Table S1). The X-ray powder diffraction (PXRD) spectrum (Figure 1c) exhibited three broad peaks centered at 2θ values of 14, 17.4 and 28°, corresponding to graphitic interplanar spacing of 6.4 Å, 4.9 Å and 3.2 Å, which can be attributed to highly disordered carbon species.36,37 The broad peak centered at 44° corresponds to a d-spacing of ~0.21 nm. TEM image of the CD has been depicted in Figure 1d (Supporting information Figure S3 ). The size distribution of CD has been shown in the inset of Figure 1d. As can be noticed from this Figure 1d, the size of CD is ~ 6 nm, similar to what has been reported in literature.17-19,20,38 From the HRTEM image the interplanar distance has been calculated to be 0.21 nm (inset of Figure 1d). Thus, the magnitude of inter-planar spacing obtained from PXRD and HRTEM are consonant with each other. From DLS measurement the average hydrodynamic diameter of CD in aqueous medium has been obtained to be 12.2 nm (Figure 1e) (DLS measurement in DMEM medium Supporting infor
mation Figure S4). As can be seen from the zeta potential distribution (Figure 1f) the surface charge on the CD is negative (-4 mV with an SD of 3.45 mV). This CD has been characterized using spectroscopic techniques as well (Figure 2). The CD exhibits absorption maximum at around 330 nm (Figure 2a) and the emission maximum at around 413 nm (Figure 2b). Quite importantly this CD exhibits very high PLQY(~100%) in aqueous medium (Figure 2c) and this near unity PLQY is maintained for a very large pH range of ~1 to 12 (Figure 2d). This property is quite useful and important as to the best of our knowledge no QD or CD of near unity PLQY (in water) can retain its high photoluminescence for such large pH range of 1 to 12. The photoluminescence of this CD is also quite stable for at least six months. PLQY has been observed to be non-variant for six months (Supporting inforomation Figure S5). Even the PLQY has been observed to be non-variant over entire range of pH for six months (Supporting information Figure S5.). This CD has been shown to be quite photostable. Photostability of this CD is much better than CdSe semiconductorr QDs and comparable to CdSe/CdS core/shell QDs (Supporting information Figure S6). Thus, small sized, highly photostable, CDs with near unity PLQY (in aqueous medium and in pH range of ~1 to 12) could be prepared from non toxic precursors. We would like to go through the spectroscopic characterization in a deeper way. Unlike most of the literature reports in our case the emission maximum is independent of excitation wavelength (see inset of Figure 2b). Very few literature reports show excitation wavelength independent PL emission maximum in CD. 14,39,40 Most of the CDs reported in the literature possess low PLQY.15-22,30 Quite interestingly we have observed that the PLQY is dependent on the excitation wavelength (Figure 2c). The magnitude of PLQY remain ~100% for excitation wavelength (λex) above 300 nm, however, as the λex goes below 300 nm the PLQY falls very rapidly and comes down to 30% for λex = 270 nm or lower.
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Figure 2. Absorption and excitation spectra of CD (a), emission spectra of CD (b), excitation wavelength dependent PLQY(c), pH dependent PLQY (d), PL decay at different monitoring wavelengths (e) (excitation wavelength = 340 nm, for other excitation wavelengths see supporting information. Figure S7, Table S2). Careful observation points the fact that excitation spectrum of CD in water although resembles absorption spectrum at the higher wavelength range, but the former deviates significantly from the latter from 300 nm and below (Figure 2a). The abovementioned two observations are quite correlated and indicate that for excitation energy much above the absorption maximum (300 nm), non-radiative decay pathway gets dominated. This could perhaps be because of the interaction with the trap states for excitation energies much higher than 300 nm. This observation is quite similar to what has been reported for semiconductor QDs.41,42 Unlike most of the literature reports this CD exhibits single exponential PL decay with a quite long excited state lifetime of 15 ns in water (Figure 2e). We would like to point out here that out of hundreds of CDs reported in literature, there are very few CDs with single exponential PL decay, and there are even fewer CDs with single exponential PL decay with an excited state life time of >10 ns (Figure 2e). Longer lifetime is beneficial not only because it allows to follow excited state dynamics for a longer period of time, it also allows RET to be observed for a longer donor acceptor distance (see next sections). Quite importantly, PL decay has been noted to be independent of monitoring wavelength (Figure 2e). This observation implies that there exist no low-lying trap states.
Rotational anisotropy decay. In order to decipher nanoscopic structural information of the CD we have performed rotational anisotropy decay in aqueous medium. If the whole spherical CD exhibits photoluminescence then it is not expected to show any
rotational anisotropy decay.43 For example, spherical QDs do not exhibit any rotational anisotropy decay. Observation of rotational anisotropy decay (Figure 3) signifies that fluorescing unit is anisotropic/anisometric in nature. Rotational anisotropy decay has been measured with 377 nm with an IRF of less than 100 ps. Rotational correlational time obtained for different solvents are much higher than IRF of the LASER. The single exponential fit yielded highly reasonable rotational correlation time with minimal (
