**3. Carbon-based QDs: synthesis methods, modification, cytotoxicity, and application in cancer cell imaging**

 Carbon-based quantum dots include a wide group, which is discussed more in this chapter on carbon quantum dots (CQDs) and graphene quantum dots (GQDs) *Quantum Dots in Cancer Cell Imaging DOI: http://dx.doi.org/10.5772/intechopen.107671*

 **Figure 3.**  *Schematic diagram of the structure of carbon-based QDs.* 

( **Figure 3** ). CQDs, also called carbon dots, are a new category of carbon nanomaterials with a size below 10 nm that were first discovered in 2004 [ 32 ]. The shape of CQDs, is spherical and has a crystal lattice with surface chemical groups, which possess quantum confinement effect (QCE) and intrinsic state luminescence [ 33 ]. A major feature of quantum dots is the QCE, which occurs when quantum dots are smaller than their exciton Bohr radius. According to the results of previous studies, it seems that the small size (below 10 nm) and the increase in the thickness of the shell in quantum dots create a strong confinement effect, which ultimately increases the amount of luminescence [ 34 – 36 ]. In addition to having excellent optical properties, CQDs are less cytotoxic, environmental, and biohazardous than traditional semiconductor quantum dots. Moreover, in addition, CQDs have good water solubility, chemical stability, and photobleaching resistance, ease of surface functionalization, and large-scale preparation [ 37 ]. This group of quantum dots is widely used in the field of biosensing and bioimaging [ 38 , 39 ]. GQDs is one of the most attractive and newest members of the graphene family, which have an exposed graphene network

#### **Figure 4.**

 *The typical approaches for the synthesis of CQDs. (A) hydrothermal method, (B) chemical oxidation, (C) emulsion-templated carbonization, (D) chemical vapor deposition, (E) hydrothermal method, (F) microwave-assisted pyrolysis synthesis method. Reprinted with permission from [ 36 ].* 

 **Figure 5.**

 *The typical approaches for the synthesis of GQDs [ 5 ].* 

and are composed of single or multiple sheets of graphene fragments [ 40 ]. This type of QDs has many applications in the field of biomedical, including cell imaging [ 41 ].

 The synthesis methods of both quantum dots are almost similar and include: acidic exfoliation method, laser ablation, hydrothermal method, solvothermal method, electrochemical method, precursor pyrolysis, microwave-assisted synthesis chemical vapor deposition (CVD), etc. [ 5 , 42 ]. The most common and important synthesis methods are shown in **Figures 4** and **5** . In CQDs, due to having oxygen-containing groups, the possibility of covalent bonding with other functional groups is very high. Covalent bonding by chemical agents such as amine groups is a current approach for surface modification of CQDs [ 43 ]. In the case of GQDs, surface functionalization with folic acid, arginine-glycine-aspartic acid (RGD) and polyethyleneimine (PEI) were performed in recent years [ 5 ].

 Previous studies in the in vitro and in vivo showed that both QDs at low concentrations have little cytotoxicity even if are synthesized from toxic ingredients [ 44 ]. The size and concentration of quantum dots are two important factors that affect cytotoxicity [ 45 ]. For example, it has been reported that GQDs with a concentration of less than 50 μg/ml have less than 10% cytotoxicity, while at a concentration of 200 μg/ml, they have more than 20% cytotoxicity [ 46 ].

 Carbon-based quantum dots are used in various fields such as environment, energy, sensing, and imaging. Wang et al. synthesized polymer-coated nitrogen-doped carbon nanodots by direct solvothermal reaction. The CQDs were stable and water soluble with a particle size in the range of 5–15 nm. The prepared quantum dots did not show obvious cytotoxicity. In the in vivo study that was performed on the glioma-bearing nude mice,

*Quantum Dots in Cancer Cell Imaging DOI: http://dx.doi.org/10.5772/intechopen.107671*

#### **Figure 6.**

 *Application of CQDs in cancer cell imaging. (A) In- and ex-vivo imaging of glioma-bearing mice intravenously administered with the pN-CNDs. reprinted with permission from [ 41 ]. (B) NIR fluorescence images of a representative mouse bearing a HeLa tumor that received intravenous injection of LAAM TC-CQDs at the indicated time points. (C) In- and ex-vivo NIR fluorescence images of a representative U87-tumor-bearing mouse (a) and indicated organs and tumor (b) after intravenous injection of LAAM TC-CQDs at the indicated time points. Reprinted with permission from [ 42 ].* 

#### **Figure 7.**

 *Application of GQDs in cancer cell imaging. (A) In vivo imaging of HeLa tumor-bearing nude mice after injection of (GQD/DBM)3EuPhen/GQD (5 mg/kg) (a). Ex vivo images of isolated organs of mice at 10 h after injection (b) and PL intensities of (GQD/DBM)3EuPhen/GQD from isolated organs (c). Reprinted with permission from [ 43 ]. (B) Confocal laser scanning microscopy of Hela, A549, and HEK293A cells incubated with GQD–FA. Reprinted with permission from [ 44 ].* 

the high fluorescence emission was observed at 30-min post injection of the pN-CNDs ( **Figure 6A** ). They stated that surface coating of the QDs with a hydrophilic polymer, in addition to increasing the accumulation of the particles in the glioma, extended the circulation time in the bloodstream, which increases the chance of binding to the target tumor site [ 47 ]. In another study Li and co-worker showed that CQDs functionalized with multiple paired α-carboxyl and amino groups that bind to the large neutral amino acid transporter 1 (which is expressed in most tumors), selectively accumulate in human tumor xenografts in mice and in an orthotopic mouse model of human glioma. They reported that 8 h after intravenous injection into mice bearing HeLa tumors, maximum fluorescence emission was observed in the tumor area, while no fluorescence was observed in other areas ( **Figure 6B** ). In order to investigate the capability of LAAM TC-CQDs for brain cancer imaging and treatment, the prepared CQDs injected intravenously into mice bearing U87 gliomas. Their results showed the maximum accumulation of carbon quantum dots in the brain occurs 8–12 h after injection. In addition, by euthanizing the mice after 12 h, ex vivo results showed that LAAM TC-CQDs were significantly accumulated in the brain tumor compared to other organs ( **Figure 6C** ) [ 48 ]. In one study, the GQD–europium complex composites was used as a probe for in vivo fluorescence imaging of HeLa tumor-bearing nude mice. They stated that the maximum amount of fluorescence emission occurs at the tumor site 2 h after injection. While fluorescence is not observed in other organs ( **Figure 7A** ) [ 49 ]. Wang et al. reported that Folic acid (FA)-conjugated GQDs were capable to selective imaging of Hela cancer cell in comparison to A549 and HEK293A cell line ( **Figure 7B** ). Their result showed that GQD– FA enter to Hela cells with FR-induced endocytosis, which is consistent with the fact that HeLa cells overexpress FR while A549 and HEK293A express FR at a low level [ 50 ].
