Effect of Lateral Size of Graphene Quantum Dots on Their Properties

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Effect of Lateral Size of Graphene Quantum Dots on Their Properties and Application Fangwei Zhang, Fei Liu, Chong Wang, Xiaozhen Xin, Jingyuan Liu, Shouwu Guo, and Jingyan Zhang ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.5b10602 • Publication Date (Web): 03 Jan 2016 Downloaded from http://pubs.acs.org on January 7, 2016

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ACS Applied Materials & Interfaces

Effect of Lateral Size of Graphene Quantum Dots on Their Properties and Application Fangwei Zhang,a Fei Liu,b Chong Wang,b Xiaozhen Xin,b Jingyuan Liu,b Shouwu Guoa* and Jingyan Zhangb* a

Department of Electronic Engineering, School of Electronic Information and Electrical

Engineering, Shanghai Jiao Tong University, Shanghai 200240, P. R. China. b

State Key Laboratory of Bioreactor Engineering, Shanghai Key Laboratory of New Drug

Design, School of Pharmacy, East China University of Science and Technology, Shanghai, 200237. P. R. China. Corresponding Authors * Shouwu Guo, Tel/Fax: 0086-21-34206915, E-mail: [email protected], or Jingyan Zhang, Tel/Fax: 0086-21-64253846, [email protected]

1 Environment ACS Paragon Plus


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ABSTRACT Well-defined graphene quantum dots (GQDs) are crucial for their biological applications and the construction of nano scaled optoelectronic and electronic devices. However, as-synthesized GQDs reported in many work assume a very wide lateral size distribution, thus their apparent properties cannot truthfully reflect intrinsic properties of the well-defined GQDs, and more importantly the applications of GQDs will be affected and limited as well. In this work, we demonstrated that different sized GQDs with a narrow size distribution could be obtained via gel electrophoresis of the crude GQDs prepared through photo-Fenton reaction of graphene oxide (GO). It is illustrated that the photoluminesce (PL) emissions of the well-defined GQDs are originated mainly from the periphery carboxylic groups and conjugated carbon backbone planes through fluorescence and UV-vis spectroscopies. More importantly, our findings challenge the notion that the excitation wavelength dependent PL property of the as-synthesized GQDs is the intrinsic property of the size-defined GQDs. Preliminary data at cellular level indicated that the small sized GQDs exhibit weaker quenching DNA dye ability, but higher toxicity to the cells comparing to the as-synthesized GQDs. The discovery is essential to explore applications of the GQDs in pharmaceutics, and to understand the origin of the optoelectronic properties

of

GQDs.

KEYWORDS: Graphene quantum dot; Gel electrophoresis; Lateral size; Photoluminescence


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1. INTRODUCTION Graphene quantum dots (GQDs) have attracted great research interests, and have shown many potential applications in diverse areas.1-5 With a strong quantum confined electronic state and unique edge structure effect, GQDs exhibit an open band gap structure, affording them distinctive optoelectronic properties,6 thus have been explored in the constructions of nano-scaled optoelectronic and electronic devices.7-10 In addition, the intrinsic single atomic layer thickness, nanometer lateral size, and peripheral functional groups provide GQDs with unique application in biomedical research.6 The well-defined GQDs including the size and surface status are critical for their applications. For example, large sized GQDs could alleviate immune-mediated liver damage, but relatively small sized GQDs could not.11 However, the as-synthesized GQDs that prepared with various strategies, in most cases assume a very wide lateral size distribution.12-16 This will severely affect the understanding of their intrinsic properties and their applications. For instance, the photoluminances (PL) emission wavelength of many GQDs reported in the literature, is dependent on the excitation wavelength.17 However, that is different from a quantum dot, whose PL emission peak is independent on the excitation wavelength, but the intensity of the PL emission is dependent on the excitation wavelength, when their size, morphology and surface functional groups are defined. Several mechanisms were proposed to illustrate the origin of the excitation wavelength dependence of the PL property of GQDs.18-28 Generally accepted explanation is that quantum size, zig-zag edge sites, recombination of localized electron-hole pairs, and defect effects were involved in the PL emission.4, 23, 28-32 The first three of them were regarded as intrinsic state emission and the last one defined as the defect state emission.



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However, the dominant factor remains unclear. Zhu et al prepared three different GQDs with different methods, in which they wished to differentiate the contribution of these factors, but the amination of the GQDs in fact altered the physical property of the starting GQDs, thus the comparison of the GQDs with the aminated GQDs seems questionable.25 Meanwhile, to obtain well-defined GQDs, especially within a certain size region, several methods have been employed to the separation of as-synthesized GQDs.28, 33-35 Fuyuno et al obtained different sized GQDs by size-exclusion HPLC,34 however, the PL spectra of the different sized GQDs suggested that they still have a wide size distribution. The GQDs were also separated by ammonium sulphate through salting out strategy.35 The different fractions obtained by this method had fairly different sizes, but exhibited quite similar PL spectra with multiple peaks at one excitation wavelength. Jiang et al. acquired different sized amino-functionalized GQDs with molecular sieving column,33 and the two fractions obtained from the column assumed different thickness, but unexpectedly, their UV-Visible and PL spectra were basically identical. Hence, to obtain GQDs with welldefined physical property, in this work, the as-synthesized GQDs were graded with gel electrophoresis, different sized GQDs with a narrow size distribution were obtained and characterized, and their PL property was investigated. The results clearly showed that different sized GQDs exhibit different PL emission and UV-visible spectra. And more importantly, the PL emission wavelength of the well-defined GQDs is excitation wavelength independent in general. However, this is not the case when GQDs are in smaller sizes (less than ~6 nm in this work), because their surface status becomes a dominant factor to their PL property. In addition, it is found that different sized GQDs exhibit very different behaviours when they interact with cells.


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2. EXPERIMENTAL SECTION 2.1 Materials and Instruments: GQDs were prepared from the GO by photon-Fenton reaction as described in our previous work.36 Atomic Force Microscopy (AFM) images were acquired in tapping mode using a Multimode Nanoscope V scanning probe microscopy system (Bruker, USA), and AN-NSC 10 AFM cantilever tips (SHNIT Co., Russia) with force constant of ~37 Nm-1 and resonance vibration frequency of ~300 kHz were used. The specimens for AFM measurement were prepared by solution casting the aqueous suspensions of GQDs on a freshly cleaved mica surface and drying in air. Transmission Electron Microscope (TEM) images were obtained using a JEM-2100F transmission electron microscope (JEOL, Japan) operated at 200 kV. The samples were prepared by solution casting the aqueous suspensions of GQDs on the copper grids with ultrathin carbon films and drying under ambient condition in a drier. The UVvis measurements were performed on a Cary 50 spectrometer (Varian, USA). The fluorescence spectra were acquired by a Cary Eclipse spectrofluorometer (Varian, USA). Fluorescence images of cells were obtained by a fluorescence microscopy (Nikon ECLIPSE, Ti-S). The optical photographic were captured by Panasonic a DMX-LX7 camera with a Leica lens (Panasonic, Japan). Agarose gel electrophoresis was carried out with a DYY-6C electrophoresis apparatus (Liuyi Instrumental Co., China). The agarose gels were visualized and digitized with the FR200A gel image analysis system and analyzed by Smart View software. 2.2 Gel electrophoresis of GQDs and recovery of the GQDs from gel: Agarose gel experiments were carried out using 1.5% agarose gel. Typically, to run the gel electrophoresis, the aqueous suspension of 400 µL (~2 mg mL-1) of the as-synthesized GQD sample mixed with 40 µL of glycerin, and then was injected into the sample well. A voltage of 50 V was first applied for 6 min, then the voltage was set at 120 V for around 25 min until the brown band



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reached middle of the gel. The different color bands were carefully cut out under UV light after the electrophoresis process. The incised gel bands were then soaked in purified water for 24 h to recover the corresponding GQDs. The resulting GQDs solutions were then centrifuged (4000 rpm, 15 min) and dialyzed (dialysis tube with a cutoff of 1000 Da for GQDs-1, and 8000-14,000 Da for the others) to remove the electrolytes. 2.3 Cellular experiments. Human breast cancer cells MCF-7 cells (purchased from Shanghai Cell Bank, Chinese Academy of Sciences) were cultured in RPMI 1640 medium with 10% fetal bovine serum and 1% antibiotics (penicillin and streptomycin) at 37 ℃ under 5% CO2. For the cell morphology observation, MCF-7 cells were detached by trypsin and plated in 24-well plates at a density of 5×104 cells per well to culture sequentially. After 12 h of attaching, medium containing 50 µg mL-1 of the GQDs was added into the well followed by removing the medium with serum and washing cells with PBS buffer. After 3 h of incubation, the cell morphology was observed and recorded by invert microscope. For the fluorescence quenching experiment, after 12 h of attaching, the cell medium was replaced with the medium without serum but containing 0.5 µg mL-1 of Hoechst 33258, followed by the washing with PBS buffer two times. After 30 min for incubation, the medium was removed and the cells were washed with PBS buffer. Then medium containing 20 µg mL-1 of GQDs were added and the cells were incubated for 4 h continuously, while the control cells were incubated in the medium without GQDs. The cells were washed two times with PBS buffer and then fixed with 4% paraformaldehyde (pH = 7.4). After 15 min of fixing, the coverslips were laid onto slides after washing with PBS buffer. The prepared slides were imaged by fluorescence microscope. 3. RESULTS AND DISCUSSION


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3.1 Different sized GQDs were obtained by gel electrophoresis of the as-synthesized GQDs. Gel electrophoresis is widely employed in biology to separate biomacromolecules and their fragments based on size and charge statues. By simply varying the ratio of monomer and cross-linker that composed gels, gels with different pore size networks could be prepared, and are suitable for separating proteins, nucleic acids or their fragments.37 As we reported previously, the GQDs synthesized from photo-Fenton reaction of GO sheets assume defect free carbon basal plane with a large number of peripheral carboxylic groups, and a relatively wide lateral size distribution (Figure S1). In principle, each GQD can be considered as a large molecule, and its charge statues is determined by its peripheral carboxylic groups. Thereby, in theory, the as-synthesized GQDs could be separated into different sized GQDs through gel electrophoresis. There are possibly some ultra-small sized GQDs generated during the photon-Fenton reaction of GO,36 electrophoresis was thus first tried with polyacrylamide gel (PAGE). Unfortunately, most of the GQDs were trapped in the gel, possibly due to the small pore size of the 10% PAGE. An agarose gel with larger matrix pores, often used in nucleic acid molecules separation,37 was then employed. As shown in Figure 1, under a constant voltage, three bands with different colours, blue, green, and orange on the gel were observed under UV light (302 nm), and were labelled as GQDs-1, GQDs-2, and GQDs-3, respectively. In contrast, under white light, only band GQDs-3 is barely visible. On the basis of general principle of gel electrophoresis, the separation of GQDs in agarose gel is determined both by the lateral dimension of the GQDs and the number of their peripheral carboxylic groups. The number of carboxylic groups of each GQD is proportional to its lateral size, the larger the size is, the more carboxylic groups it contains. However, the



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surface area of the GQDs grows square proportionally with its lateral dimension. Hence, the density of carboxylic

Figure 1. Agarose gel (0.75%) of the as-prepared GQDs aqueous solution. The images were taken under white and UV light (302 nm). The running buffer was Tris-HAc (40 mM, pH = 8.35). The voltages were set at 50 V for 6 min and then 120 V for 23 min.

groups in larger lateral sized GQDs is lower than that in smaller sized GQDs. Accordingly, the running speed of the smaller GQDs is faster than that of the larger ones in the gel. As a result, the as-synthesized GQDs were graded according to their lateral sizes in agarose gel, and their sizes should be in the following order: the blue luminescent GQDs-1 < green GQDs-2 < orange GQDs-3. The result agrees well with the literature that GQDs with different sizes emitted different PL.28, 38-39 To identify accurately the size of GQDs in each gel band, the bands were recovered from the agarose gel by cutting out the gel stripes according to their colour under UV light (302 nm) and incubating them in Milli-Q H2O for 24 h, respectively. The resulted aqueous solutions were centrifuged, and the supernatants were dialyzed. As expected, the extracted solutions exhibit the same colour as the corresponding gel bands under the UV-light, revealing that the GQDs were


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recovered successfully from the agarose gel. The recovery efficiency was further confirmed by comparing the intensity of the GQDs gel bands under UV light before and after the water soaking (Figure S2). Gel electrophoresis of GQDs not only can provide a partition method of obtaining the GQDs with different sizes, it can also be employed in monitoring the preparation of the GQDs via monitoring the intensity of the colour bands under UV-light of the samples (Figure S3). This method makes it possible to track the reaction process within an hour. However, gel electrophoresis is not a ubiquitous method to obtain different sized GQDs with all as-synthesized GQDs. Gel electrophoresis experiment with the GQDs made from glucose (GQDs-glucose)21 showed that the GQDs-glucose were trapped inside the gel, suggesting that either their net charge is zero, or they contain extremely high concentration of zigzag sites that are easily trapped in the gel matrix. Similarly, experiment with GO/DMF (GQDs-DMF)10 displayed that some of them moved forward in the gel, but some of them moved backwards (Figure S4), indicating that some GQDsDMF are negatively charged, and some are positively charged. Apparently, these two kinds of GQDs can’t be efficiently separated by gel electrophoresis. 3.2 Characterization of the isolated GQDs. TEM images of the GQDs-1, GQDs-2, and GQDs-3 were acquired, and the results were shown in Figure 2a-c. Clearly, the average sizes of the GQDs is in an order of GQDs-1 < GQDs-2 < GQDs-3, which is highly in consistent with the aforementioned gel electrophoresis results. Figure S5 shows the high resolution TEM (HRTEM) image of the GQDs-2. The basal plane of GQDs-2 maintains the pristine graphene structure with a lattice parameter of 0.23 nm (for (100) facet of graphite) and 0.34 nm (for (002) facet of graphite),36, 40 indicating that no damage to the



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GQDs is occurred during the electrophoresis. The isolated GQDs were further

Figure 2. TEM images of the GQDs-1 (a), GQDs-2 (b), and GQDs-3 (c). Scale bar equals 20 nm. Inset in (a) is the blow up of the GQDs-1, and the scale bar equals 10 nm. (d-f) are taping mode AFM phase images of the GQDs-1, GQDs-2, and GQDs-3, respectively. Scale bar equals 100 nm.

i characterized by taping mode atomic AFM (Figure 2d-f). The average sizes of the GQDs estimated from their AFM images are 5.5, 12.5, and 16 nm in diameter for GQDs-1, GQDs-2, and GQDs-3, respectively (Figure S6). Notably, the diameter data were measured directly from AFM images of the GQDs, and calibrated with the AFM tip deconvolution parameters obtained using a simplified model shown in Figure S7.41 Obviously, the estimated lateral sizes of GQDs based on their AFM images are in along


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with the electrophoresis and TEM results. The thickness of the most GQDs detected in taping mode AFM images is around 0.8 nm, showing a single layer motif (Figures S6 and S8).36, 42 These results together demonstrate that the GQDs isolated by gel electrophoresis show different sizes with a narrow size distribution. 3.3. PL property of the GQDs-1, GQDs-2, and GQDs-3. Although GQDs exhibit unique optoelectronic property owing to the quantum confinement effect, the apparent properties of the GQDs are usually collected on the mixture of GQDs with different sizes,

Figure 3. PL properties of the GQDs separated by gel electrophoresis. (a) The comparison of the PL spectra of as-synthesized GQDs with GQDs-1, GQDs-2, and GQDs-3 that excited at 340 nm (concentrations were not calibrated). (b), (c), and (d) are the PL spectra of the GQDs-1, GQDs-2, and GQDs-3 that excited from 320 to 400 nm,



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respectively. The insets in each figure are the pictures of each sample in cuvettes taken under UV-Vis light (302nm).

and it thus cannot reflect the intrinsic property of the certain sized GQDs. The PL properties of the as-synthesized GQDs, GQDs-1, GQDs-2, and GQDs-3 thus were investigated, and their spectra were displayed in Figure 3a. Overall, under the same excitation wavelength, the PL spectrum of the as-synthesized GQDs (Figure 3a, black line) is broad suggesting that it is a combination of the PL spectra of different sized GQDs. While the PL spectra of GQDs-1 and GQDs-2, and GQDs-3 are much narrow, especially for GQDs-1, the FWHM is about 90 nm, which is much narrower than many reported work.39 It has been demonstrated that the π*→n transition of carbonyl or carboxylic groups connected to carbon nanotubes and other fullerene carbon nanostructured materials produce PL emission around 430 nm with excitation wavelength at around 320 nm.43 Therefore, the emission at 450 nm of the GQDs-1 is partially contributed by the π*→n transition of carbonyl or carboxylic of GQDs. The other contribution to PL of the GQDs-1 might be the conjugated basal carbon skeleton. Notably, the main PL emission peak of the GQDs-1 shifted gradually to the longer wavelength with the increase of the excitation wavelength (Figure 3b). However, the size distribution of the GQDs-1 is narrow based on AFM statistic result (Figure S6), implying that the emission wavelength shifts with the excitation wavelength of GQDs-1 is possibly caused by other reasons (see discussion later). Figure 3c showed the PL spectra of the GQDs-2. Overall it can be de-convoluted into two peaks centered at ~425 and 510 nm, respectively. Similar to the GQDs-1, the emission at 425 nm is originated from the π*→n transition of the carboxylic


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groups, and the emission at 510 nm is from the conjugated carbon backbone of the GQDs.17 In contrast to the GQDs-1, the emission wavelength of the GQDs-2 is independent on the excitation wavelength. Similarly, as illustrated in Figure 3d, the PL emission peak of GQDs-3 centered at 513 nm is mainly from the conjugated carbon backbone. The slight red shift of the emission wavelength of the GQDs-3 with the increase of the excitation wavelength is likely due to the wider size distribution compared to GQDs-1 and GQDs-2, as evidenced by their broad PL peak. Notably, PL intensity of the larger sized GQDs-3 is much weaker because the contribution of the quantum effect apparently is less prominent. To further understand the phenomenon of the emission wavelength dependent on the excitation wavelength we observed in the GQDs-1, but not in GQDs-2, two adjacent blue gel bands were cut out from the gel and recovered, the bottom band was named as GQDs-1a, the top band was named as GQDs-1b as illustrated in Figure 4. GQDs-1a are marginally smaller than



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Figure 4. Recovery of GQDs-1a and GQDs-1b from the gel bands. Their pictures were taken under UV-vis light in cuvettes (302 nm). The PL spectra of two samples were acquired at excitation wavelengths of 340 and 380 nm. GQDs-1b, because GQDs-1a run slightly faster than GQDs-1b in the gel. The PL spectra of these two samples were compared. When they were excited at 340 nm, their emission spectra are almost identical, but when they were excited at 380 nm, the emission wavelengths of both samples shifted to longer wavelengths, and it was 23 nm shift for the GQDs-1b, which was 6 nm more than that for the GQDs-1a. Since the GQDs-1a and GQDs-1b are very similar in terms of size distribution and surface status, the difference in emission wavelengths shift apparently is caused by their small difference in size. As mentioned earlier, PL spectra of the GQDs were contributed by the conjugated carbon skeleton and surface status. These two contributions are both excitation wavelength independent in principle, however, when the excitation wavelength shifts to a longer wavelength, the relative ratio of these two contributions will change, the overall apparent emission spectrum thus will be excitation wavelength dependent. In the case of GQDs1, the contribution of surface status is dominant because of their smaller sizes, thus main peak of PL centers at ~450 nm, the contribution from the carbon skeleton shows up only when the GQDs-1 are excited at a longer wavelength. While for the GQDs-2, the larger carbon skeleton is a major contributor to the PL, and peripheral carboxylic groups have a minor effect, thus the change of the excitation wavelength would not cause a larger shift to the emission wavelength. Similar is true for the GQDs-3. These results together showed that the excitation wavelength dependence of the emission wavelength of the GQDs is not their intrinsic property when they are larger than 5 nm, it is caused by the wide size distribution of the GQDs. Our result is different from the literature report that the GQDs with sizes from 1.5 to 5.5 nm, prepared by the hydrothermal method, exhibited emission wavelength independent PL property.44 The authors


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proposed that the surface status of the GQDs prepared by hydrothermal method dominated the PL property, and thus small size variation would not cause the shift of emission wavelength.44 This possibly is true, the GQDs generated from mild photon-Fenton reaction of GO unlikely have many zig-zag sites (this is in along with their low quantum yield discussed later), and thus only when they are in small sizes, the contribution of surface state is dominant. Nevertheless, through gel electrophoresis method, the information of the size and PL property of the GQDs could be obtained concomitantly. The PL quantum yields of GQDs-1, GQDs-2, and GQDs-3 are higher than the assynthesized GQDs (0.3%, 0.07 %, and 0.02 % vs 0.006% using quinine sulphate as a reference), but are far comparable to organic PL dyes. The low quantum yield of the GQDs possibly is caused by the way how they were prepared, in which the precursor GO was not treated as harshly as in hydrothermal method that generated high concentration of zig-zag site.12 3.4. UV-Vis absorption of the GQDs-1, GQDs-2, and GQDs-3. Different sized GQDs also exhibit dissimilar UV-Visible spectra. The as-synthesized GQDs display a wide absorption at UV and visible regions (black line), but GQDs-1 adsorb primarily UV light (blue line), and GQDs-2 and GQDs-3 are in between (Figure S9). All the GQDs samples show absorption at 225 nm, which could be assigned to π→π* transition of aromatic sp2 domains;45 but the intensity of the peak at 225 nm of GQDs-1 is much higher than that of GQDs-2 and GQDs-3, indicating that smaller sized GQDs-1 assume better defect free aromatic sp2 domains. The shoulder at 300 nm appeared in the spectra of GQDs-1, GQDs-2, and GQDs-3 (enlarged spectra in inset of Figure S9), is assigned to the n→π* transition of C=O bonds.46 Apparently, the density of carboxylic groups in the GQDs-1



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and GQDs-2 is higher as shown in gel electrophoresis. In contrast, the peak at 300 nm of the GQDs-3 sample is almost invisible, revealing that the surface density of carboxylic groups is relatively low in the GQDs-3, and their size distribution is wider. The result is in along with the observation that GQDs-1 and GQDs-2 move faster than GQDs-3 in the agarose gel (Figure 1). The red shift of the onsets of the absorption around 300 nm in the UV-Vis spectra of the GQDs-1, GQDs-2, and GQDs-3 also indicates the lateral dimension increases from GQDs-1 to GQDs-3. 3.5 GQDs obtained by gel electrophoresis exhibit unique effect to the cells. Interestedly, with these isolated GQDs, it is possible to understand some of their biological effect, because they have well defined properties. Figure 5 compared the

Figure 5. a) Bright field images of MCF-7 cells incubated with 50 µg mL-1 of the assynthesized GQDs and GQDs-1 for 3 h. b) MCF-7 cells were pre-incubated with 0.5 µg mL-1 of Hoechst for 30 min, and then incubated with 20 µg mL-1 of the GQDs for 4 h.


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Excitation wavelength, 340 nm. Scale bar: 50 µm. c) The PL of Hoechst was quenched by the various concentrations of the as-synthesized GQDs and GQDs-1 in vitro, respectively.

effects of the as-synthesized GQDs and the GQDs-1 to the MCF-7 cells. The bright field images of the MCF-7 cells that incubated with the as-synthesized GQDs and GQDs-1 for 3 h clearly show the difference. With the GQDs-1, the cells were round and unhealthy, but with the as-synthesized GQDs, cells are in normal healthy state. Furthermore, when MCF-7 cells were pre-stained with nuclei dye Hoechst, and then incubated with the assynthesized GQDs and GQDs-1, the fluorescence of the nuclei in the former was quenched rapidly, while it was not in the latter, which agrees well with the in vitro molecular level experiment as shown in Figure 5c. The fluorescence of the Hoechst was quenched upon the addition of as-synthesized GQDs, but it cannot be quenched by the GQDs-1 under the same condition. The slight increase of the fluorescence intensity of the Hoechst with GQDs-1 was caused by the contribution of the fluorescence of the GQDs-1 themselves (Figure 3a). The result suggests that smaller sized GQDs-1 have a weaker fluorescence quench capability because of their poor electron transport ability comparing to the larger sized GQDs. Though more work in depth is necessary to understand these preliminary findings, it indicates unambiguously that the well-defined GQDs exhibit different property comparing to the as-synthesized GQDs that with a wide size distribution. More importantly, it’s necessary to have well-defined GQDs for the research of GQDs in biomedical applications, such as in drug delivery. 4. CONCLUSION



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In summary, we employed convenient gel electrophoresis strategy to obtain GQDs in average sizes of 5.5, 12.5, and 16 nm with a narrow size distribution. With these well-defined GQDs, we demonstrated that the PL emission of the well-defined GQDs are originated mainly from the periphery carboxylic groups and conjugated carbon skeleton. More importantly, it is revealed that the contribution of surface status to PL property of the GQDs is dominant when GQDs are in small sizes (less than 6 nm in this work), the contribution of the carbon skeleton only shows up when they are excited at a longer wavelength. While for the large sized GQDs (~12 and 16 nm), the larger carbon skeleton is a major contributor to their PL, and peripheral carboxylic groups have a minor effect, their PL emission spectra is independent of the excitation wavelength. We thus can infer that the dependence of emission wavelength on the excitation wavelength of the GQDs that reported in the literature most were caused by their wide size distributions. Preliminary experimental data also showed that the well-defined small sized GQDs (~ 5.5 nm) obtained from gel electrophoresis exhibit unique behaviour to cells and to small molecules. The results should be beneficial to deep understanding the origin of the optoelectronic property of GQDs and their biological applications as drug, gene delivery systems. Supporting Information. UV-Vis spectra of the different sized GQDs, AFM images of the assynthesized GQDs, High resolution TEM of the different sized GQDs and other supporting information are included. Acknowledgment. This research was carried out with financial support from the national “973 Program” of China (Nos. 2014CB260411, 2015CB931801), National Science foundation of China (No. 11374205), the state key laboratory of bioreactor engineering (No. 2060204), 111 Project (No. B07023), the Shanghai Committee of Science and Technology (No. 11DZ2260600), and national “863” Program of China (No. 2012AA022603) for financial support of this work.


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Table of Contents Different sized graphene quantum dots (GQDs) with a narrow size distribution could be obtained via gel electrophoresis of the crude GQDs prepared through photo-Fenton reaction of graphene oxide. The isolated GQDs, due to different sizes, exhibit unique photoluminesce properties, and

interact with the cells differently.


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Effect of Lateral Size of Graphene Quantum Dots on Their Properties

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