New insight into the engineering of green carbon dots: Possible applications in emerging cancer theranostics

Journal Pre-proof New insight into the engineering of green carbon dots: Possible applications in emerging cancer theranostics Fatemeh Radnia, Nasrin Mohajeri, Nosratollah Zarghami PII:

S0039-9140(19)31180-4

DOI:

https://doi.org/10.1016/j.talanta.2019.120547

Reference:

TAL 120547

To appear in:

Talanta

Received Date: 6 July 2019 Revised Date:

5 November 2019

Accepted Date: 7 November 2019

Please cite this article as: F. Radnia, N. Mohajeri, N. Zarghami, New insight into the engineering of green carbon dots: Possible applications in emerging cancer theranostics, Talanta (2019), doi: https:// doi.org/10.1016/j.talanta.2019.120547. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. 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. © 2019 Published by Elsevier B.V.

New Insight into the Engineering of Green Carbon Dots: Possible Applications in Emerging Cancer Theranostics Fatemeh Radnia1,2, Nasrin Mohajeri1, and Nosratollah Zarghami 1,3* 1.

Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran 2.

Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran

3.

Department of Biochemistry and Clinical Laboratories, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran

Corresponding author: Nosratollah Zarghami, MDLab, Ph.D. Department of Clinical Biochemistry and Laboratory Medicine Department of Medical Biotechnology Faculty of Advanced Medical Sciences Tabriz University of Medical Sciences, Tabriz, Iran. Postal code: 13191-45156. E-mail: [email protected] Tel: +984133355788, Fax: +984133355789

1

Abstract Fluorescence imaging via carbon dots (CDs) has found multifarious applications in the biomedical sciences including biosensing, cancer cell bioimaging, drug delivery and tracking therapeutic response. Presently, the latest generation of fluorescence CDs known as green-CDs has attracted ever-increasing attention due to the use of natural sources, low-cost synthesis, nanoscale size, promising biocompatibility, superior photoluminescence, and ease of functionalization for versatile applications, which in turn could have higher priority over the traditional toxic fluorescent agents. In this review, we aim to have a new insight into the engineering green-CDs and their physicochemical properties. Moreover, we discuss the possible applications of green-CDs in self and active targeting, therapeutics delivery, and finally their promising future in cancer theranostics.

Keywords: Drug delivery; Fluorescence materials; Green-carbon dots; Targeted imaging; Theranostics

2

Table of contents 1. Introduction 2. Overview of natural green-CDs 2.1. Synthesis methods 2.2. Optical properties 2.3. Surface chemistry and chemical structure 3. Targeted cancer bioimaging with green-CDs (Diagnostic) 3.1. Self-targeted imaging 3.2. Actively targeted imaging 4. Cancer image-guided drug delivery with green-CDs (Therapeutic) 4.1. Drug loading, uptake, and release 4.2. Drug and green-CD interactions 5. Conclusion and future prospective (Theranostic)

1. Introduction Nanoparticles (NPs) based on fluorescence agents are key components for diagnostic imaging studies and advanced therapy. There is an increasing trend on developing novel imaging nanomaterials to address the major challenges facing current imaging agents, which is most importantly their toxicity [1]. Carbon dots (CDs), the latest generation of carbon nanomaterials with considerable surface area, various functional groups, and enhanced drug loading capacity, have gained a growing interest compared with conventional quantum dots (QDs) [2,3]. Also, their extraordinary potentials such as high quantum yield, multicolor wavelength emission, and water stability are of their remarkable interests [3–5]. Historically, CDs were primarily discovered by Xu et al. as side-products during the study on single-walled carbon nanotubes (CNTs) in 2004 [6], while during the next two years, Sun research group synthesized and introduced the pioneer generation dots as “CDs” [7]. Over the 3

past few years, some chemical precursors including citric acid, ethylene glycol, benzene, and poly ethylene imine were the main initial materials for CDs synthesis [8–10]. The only limitation was attributed to their synthesis sources and approaches which had weaknesses including the use of toxic chemical materials, requiring high temperature, time-consuming

process, and the

exorbitant cost of synthesis. The most recent and innovative approach to improve CDs and to overcome these restrictions is based on the use of green precursors which caused growing attention to benign preparation of CDs, entitled “green-CDs” (Table 1).While using fluorescent CDs for in vivo assays and ultimately clinical translations, their safety and biological properties seem to be absolutely challenging . The bio-related applications of fluorescence imaging techniques have been optimistically followed since the advent of green-CDs [11–14]. Their successful performance is based on low to no toxicity while maintaining all the desirable features. To date, green-CDs have been derived from various natural precursors such as fruits (pomelo peels [15], orange juice [16], and coffee grounds [17]), beverages (cow milk [18], beer [19], and black tea [20]), animal derivatives (chicken egg [21], honey [22], crab shell [23], and chitosan [24]), human derivatives (human hair [25], and urine [26]), waste materials (food [27– 29], and agriculture waste [30]), plant leaves and vegetables [32–34]. Also small biomolecules such as proteins and amino acids [35–37], nucleic acids [38], vitamins [39], and carbohydrates [40,41] have been employed for the synthesis of these green-CDs. Therefore, the greatest feature of green-CDs is the use of eco-friendly green sources of raw materials, which has brought them into many different experimental studies on cancer bioimaging [42] and diagnostics or therapeutic [19, 43–45]. Interestingly, imaging agents have played an essential role as a platform for real-time multimodal imaging-associated cancer therapy [14,46,47]. Theranostic, as the integration of diagnostic and therapeutic approaches in one entity, is receiving considerable interest and might be a promising alternative to invasive and non-specific cancer prevention and treatments [48,49]. Up to date, several methods have been reported for cancer diagnosis or therapy but merging these two goals in one photoluminescent (PL) biocompatible platform like green-CD is highly desirable [50–52]. There are many comprehensive review articles for synthesis methods, physicochemical properties, surface engineering, and fluorescence-based bioimaging of green-CDs. However, their significant potential for drug delivery and targeting with different ligands for image-guided therapy as well as their recent developments are still lacking. Perhaps the most important reason 4

for the success of green-CDs in cancer theranostic could be the natural origin of carbon dot with the lowest toxicity. In a recent review paper, Wang et al. have focused on synthesis and application of multicolor CDs

from various origins in gene and drug delivery for cancer

theranostics, while they have not considered the harmless of green CDs for cancer cell theranostics [53]. In the present review paper, we discuss the recent advancements of green-CDs in image-guided cellular self and active-targeting. Subsequently, we focused on drug delivery application of green-CDs which has recently received growing attention; the interactions of green-CDs with therapeutic agents, loading efficiency, cellular uptake, and drug release have been fully elucidated. 2. Overview of natural green-CDs CDs contain C, H, N, and O related functional groups, which are arranged by biological molecules and could be efficient for the theranostic application. Based on the source of CDs synthesis, the benefit of natural resources rather than expensive chemical entities is not only their easier availability, affordability, and intrinsic non-toxicity, but also due to the existence of different carbohydrates, lipids, proteins, and other natural ingredients (e.g. biomass and plants) excessive surface passivation or doping is not required. The green-CDs are almost selfpassivated under nucleation process [8,16,54,55] (Table 2). 2.1. Synthesis methods Although it has been only about two decades of CDs arrival, several procedures have been suggested for their preparation, which can be mainly categorized into “Top-down” and “Bottomup” processes [56,57]. Top-down system for the creation of CDs from bulk carbon resources such as graphite powder or CNTs is generally performed under severe physical or chemical circumstances [7,58,59]. In contrast, bottom-up approaches such as ultra-sonication, microwave pyrolysis, and hydrothermal heating are used to prepare CDs from small organic molecules including glucose, fructose, and other natural resources [60–62]. Due to the existence of complete and comprehensive reviews on these synthesis approaches, herein the advantages and disadvantages of each one are more dedicated. The high-intensity wavelength of ultra-sonic approach is responsible for surface chemical modifications of CDs. Particularly for more efficient modification, ultra-sound from electrical power can be used, however, some chemical events such as cavitation preserve a low-yield [63]. 5

Pyrolysis is an irretrievable thermal breakdown reaction causing physical and chemical changes in inorganic ingredients. This method is not a common approach of CDs synthesis, due to the necessity for high temperature and energy-consumption which is not desirable. Microwave-assisted synthesis is based on electromagnetic radiation treatment of a mixture of precursor molecules. Microwave-assisted synthesis is known to be a rapid and cost-effective approach which is appropriate for scaled-up synthesis, while there is less control over the size distribution of CDs [64]. Fluorescent garlic CDs were synthesized using microwave-assisted heating, with bioimaging application and anti-oxidative effects which could be effective as benign theranostic nanoparticle [65]. It has been reported that N, S, P doped CDs exhibiting stable fluorescence were fabricated from onion peel powder (OPP) via short microwave treatment. It showed great cellular and blood compatibility despite degraded polyphenols in the extraction [66]. Additionally, other researchers have also reported the microwave-assisted synthesis of green-CDs [67–69]. Nowadays, hydrothermal treatment as a promising liquid phase preparation method has developed rapidly due to the merits of simplicity, energy efficiency, and generating nontoxic and efficient low-cost CDs. This method contains four consecutive stages: dehydration, polymerization, carbonization, and passivation that occur in one step remarkably [70–73]. According to the articles, most of the fruits, fruit juices, peels, vegetables, etc. [22,31,74–85] have gone through hydrothermal treatment for green-CDs formation. Green-CDs derived from milk using a hydrothermal method has been also reported elsewhere thatserved as highresolution fluorescent probes for glioma cancer cell line imaging [18]. Also, in another study amino-functionalized CDs were created by hydrothermal treatment of chitosan with excellent optical properties, without any post surface passivation [86]. Carbonizing of bovine serum albumin (BSA) through a hydrothermal approach has been suggested as a green strategy for highly fluorescent CDs synthesis, with no post modification [35]. The most recent approaches to prepare naturally occurring CDs, are thermal (roasting) processing and extraction which are mainly concerned with food, animal derivatives, and beverages. As mentioned above, food and animal derivatives are among the most reported natural sources for green-CDs synthesis. Recently, the emergence of fluorescent CDs in roasted chicken and its physicochemical properties were investigated. It was confirmed that the size, fluorescence 6

properties and cytotoxicity of CDs are affected by roasting temperature [87]. In another study, it has been shown that pizza contains fluorescence NPs with a mean size of 3.33 nm, which could be originated from proteins, lipids, and carbohydrates of pizza ingredients during the baking process [27]. Another food-borne fluorescent carbon nanostructure was produced during the roasting of pike peel. The formation process of the CDs during the roasting includes different steps of polymerization, dehydration, nucleation, aggregation, emergence, and blossom [88]. Some other reported food-borneNPs, are the CDs discovered in some popular beverages such as Coca-Cola and commercial beer which have been easily extracted and studied [89,90]. There are many other studies on food-borne green-CDs to evaluate their toxicity and efficacy [28,29,88– 93], but it still has potential uncertainty to human health and needs more future examinations. Overall, both hydrothermal and microwave-assisted have been greatly used for the preparation of CDs, but according to the recent publications, it seems that hydrothermal method is more economical and practical so broadly accepted for green-CDs synthesis. 2.2. Optical properties 2.2.1 Absorbance Green-CDs are demonstrated to have an absorption peak in UV expanse and a tail extending in the visible range. The π-π* transition of C=C and n-π* transition of C=O are respectively associated with absorption in 230-270 nm and 300-330 nm. Also, the Optical absorption of CDs in the UV region is achieved based on surface functional groups that absorb strongly in the specific wavelengths [33,54]. 2.2.2. Fluorescence The most brilliant property of CDs is their fluorescence emission depending on numerous phenomena such as size, quantum confinement, surface defects, and aromatic structures [111, 112]. Fluorescence emission from intrinsic and defect states of CDs are the two dominant PL mechanisms that have been proposed yet. It is more facile to manage PL properties of CD through governing their defect states and functional groups instead of altering their core structure [99,100]. CDs are functionalized by polymers/organic molecules and doped with non-carbon elements (e.g., N, S, P) due to the improved PL intensity [115, 116]. But it has been shown that the use of natural resources and green synthesis methods developed to address this goal in one 7

easy and cost-effective way [103]. For instance, in a recent study on CDs derived from Pyrus Pyrifolia fruit, due to the various phytochemicals of primary source, the superficial layer of CDs could be constructed with many electron-donating moieties which effectively caused sensitive and specific probing of Al3+ ion via chelation enhanced fluorescence mechanism without the consumption of any expensive or toxic agents [104]. 2.3. Surface chemistry and chemical structure Most of the green-CDs are formless sphere-shape NPs with the typical mean diameter size < 10 nm [8]. The core of CD noticeably contains sp2 hybridized carbon while the surface structure contrasts vividly based on the synthesis procedure and precursors utilized for preparation. Furthermore, the significant structural feature of CDs is likely due to different surficial functional groups such as amine, hydroxyl and carboxyl [105] which are associated with high solubility and a notable possibility for conjugation of different polymeric [20,106], organic, therapeutic [107,108] and targeting agents [44,109]. 3. Targeted cancer bioimaging with green-CDs Cancer cell detection in early stages remains a major challenge. Fluorescence imaging due to significant sensitivity and spatiotemporal resolution could hopefully shed light on this purpose [110,111]. CDs with their excellent fluorescence properties can get functionalized via targeting agents such as folic acid, antibody, aptamer through functional groups which facilitate “active targeting” of the cancer cells. Surprisingly, some of them also have “self-targeting” ability without the need for targeting agent [112]. Since green-CDs have been broadly studied with this purpose, Table 3 summarizes their imaging targets, imaging models, and targeting ligands. In the next sections, we followed the green-CDs as platforms to cancer cell targeting and tracking through fluorescent irradiation. 3.1. Self-targeted imaging Both targeting and fluorescence labeling of cancer cells could occur by fluorescent green-CDs. Bhunia SK et al. described that folic acid-derived CDs could selectively detect folate receptor (FR) in positive cancer cell without additional targeting probe. In vitro study of folate receptor in several cell lines, which were categorized into three groups: FR++ (SKOV3 and HeLa), FR+ (HepG2, MCF7) and FR̄ (A549, CHO), resulted in a direct association between cellular uptake of 8

photoluminescent CD probe and the folate receptor expression level. Spectroscopic data proved the presence of folate residue on CD surface which causes the cancer-targeting ability [113]. Similar results obtained by Zhao et al. in which folic acid-derived CDs synthesized through green synthesis approach had ultrahigh QY (94.5%) and target ability to tumor cells by folate residue on its surface as shown in Fig. 1. Owing to the great biocompatibility of folic acid, it showed the least amount of toxicity to HeLa and A549 cell lines. Fluorescence microscopy image analysis showed that HeLa cells could readily uptake the CDs while A549 was hardly fluorescence labeled. Therefore, this would be a favorable biocompatible NP for fluorescence targeted imaging [114]. Kumavat et al. reported grape seed extract-derived green graphene QDs with the ability of nucleus-targeting without peripheral targeting agents. Nucleus imaging and staining was claimed to reach by this biocompatible graphene QD and can form a novel and economical organelle imaging agent [115]. In another report by Qiao L et al., CDs derived from D-glucose and L-aspartic acid possessed the ability for targeted Imaging of C6 glioma cells. With respect to flow cytometry and confocal laser scanning microscopy results, the CDs with the molar ratio of Glu/Asp (7:3) termed CD73 could target C6 glioma cells with the highest selectivity in comparison to L929 cells. This work indicates a self-targeting function of fluorescent CDs which makes them superb candidates for the future theranostic applications [13]. 3.2. Actively targeted imaging Green-CDs were conjugated with different targeting agents to cancer cell tracking. As demonstrated by Zhang J et al., green-CDs derived from active dry yeast were functionalized by folic acid covalently to create fluorescent detection probes for FR-over expressed cancer cells. These folate-CDs could distinguish HepG2 cells (FR+) from the other cells by showing great targeting function. the internalization of folate-CD into tumor cell is via receptor-mediated endocytosis. Fig. 2 schematically shows that in the lack of free folate, a larger amount of folateCDs can get into the cells due to binding to FR and hence generating bright fluorescence. In the presence of free folate, the competitive folate sticks to target FR and prevents Folate-CDs from attaching, resulting in less fluorescence signals. Noteworthy it showed that covalent modification of CDs did not reveal any detectable changes in fluorescence spectra of CD against folate-CD complex [116]. In another study, green fluorescent CD derived from dandelion plant and 9

ethanediamine was conjugated to folic acid designing folate-CDs for targeted imaging and detection of FR+ tumor cells. In vitro study revealed that the folate-CDs could precisely distinguish FR+ cells in diverse cancer cell collections. The complex showed not only cancer cell targeting ability but also fascinating photostability through labeling nuclei of MCF-7 and cytoplasm of HepG2 [44]. Tryptophan and phenylalanine CDs conjugated to DNA-aptamer as targeting agent was synthesized by Wang Z et al. The DNA MUC1-aptamer-CD served as a probe for cancer cell recognition maintaining both the bright fluorescence and targeting ability. According to in vitro study, MUC-1 overexpressed MCF-7 cell detection could be highlighted through fluorescent signals [117]. Also, in another study, the glucose derived CD was covalently conjugated with anti-MUC1 antibody or MUC1 aptamer. While both of the functionalized CDs exposed to MUC1 protein in solution, the fluorescence of CDs decreased due to the generation of sandwich structure and fallowing CD aggregation [118]. Recently, a CD core- molecularly imprinted polymers (MIP) shell structure was discovered by Demir B et al. for Image-guided targeting of cancer biomarkers. Source of the CD core was starch and L-tryptophan respectively. MIPs shell here were synthesized for accurate detection of glucuronic acid (GlcA) which is a determinant of hyaluronan, a biological marker for a variety of cancers. According to confocal laser microscopy images, a greater concentration of core-shell NPs existed on HeLa cells than on non-cancerous cells, representing that these MIPs were extremely applicable for distinguishing among tumor and intact cells [119]. In a study by Tan et al., N-doped CDs originated from BSA and formic acid got properly functionalized with TATpeptide for the first time, which could be applied in both one- and two-photon fluorescence cellular imaging, as well as cell nuclear-targeted bioimaging. It has been reported that TATpeptide functionalized CDs were detected in the cell nucleus, whereas there was no cellular nuclear fluorescence signals for the free-CDs [120]. In another recent study, TAT peptide-conjugated CD was designed with a promising application for nuclear-targeting of mouse melanoma cancer cells. The CDs were fabricated using tryptophan and formic acid which was selected from seventeen amino acids as the precursor and instigated the highest quantum yield and two-photon cellular imaging. CDs were effectively conjugated with TAT peptide through the reaction between the amino group of the peptide and carboxylic group of the CDs. According to in vitro bioimaging study, similar to the previous 10

study, free dots have only cytoplasmic distribution, whereas fluorescence signals are clearly detectable from the nucleus of TAT-CD incubated cells [121]. As it is known, besides CDs retained fluorescence property after conjugation, it also sometimes resulted in enhanced fluorescence intensity of CDs remarkably which could be so desirable in biomedical purposes. 4. Cancer image-guided drug delivery with green-CDs The nanotechnology advent can potentially relate diagnosis and therapy. Therapeutics, various imaging nanomaterials and targeting ligands coordination in a particular nanocarrier allows simultaneous selective treatment and monitoring the response to treatment which is known as “nanotheranostics” [108,122,123]. Molecular imaging facilitates non-invasive monitoring of drug delivery and nanocarrier distribution [124,125]. The diverse imaging modalities like optical imaging (bioluminescence and fluorescence), CT (computed tomography), PET (positron emission tomography), MRI (magnetic resonance imaging), ultrasound have been developed for precise tumor localization and diagnosis to reduce invasive use of tissue biopsy [126–129]. Among the as-mentioned, optical fluorescent imaging has gained growing attention due to the merits of low cost, high signal sensitivity, high contrast, efficient detection of the disease in early stages [130]. The preliminary research study of fluorescent nanomaterials as a drug delivery platform was based on QDs. Bagalkot V et al. designed a multifunctional nanoparticle composed of QDs conjugated to Doxorubicin (DOX) and RNA-aptamer capable of sensing single cancer cells while intracellularly releasing an effective dose of the drug in a traceable manner [131]. However, clinical translation of semiconductor QDs has been limited due to high toxicity. As a desirable alternative, in vitro and in vivo drug delivery with CDs has been shown by Qinghui Zeng group that conjugated DOX to CDs electrostatically serving as a targeted and traceable delivery complex for specified cancer therapy in a diseased mouse model. Its targeting ability attributed to pH sensitivity of non-covalent interaction and pH variance between tumor and normal cells which caused in vivo drug release in tumor area with high efficiency and stability [132].

11

In this regard, scientists assume that green-CDs due to their inexpensive and straightforward synthetic routes, excellent fluorescence property in addition to high biocompatibility, great surface area, enhanced cellular uptake and easy conjugation with therapeutics [38,106,133,134] might be extravagant image guided-drug delivery platform. Most of the reports on green-CDs have been in the field of cellular and molecular imaging and ion-sensing applications with favorable results [8,135–138]. Recently, according to the tremendous bio-related application of green-CDs and widespread attention to cancer theranostics concurrently, they are capable of serving as trace-based drug delivery systems. Emphasis is placed on the fact that drug loading efficiency, cellular transmission, and release in the target site are relevant to drug-CD conjugation chemistry. In this part, we focus on green-CDs loading efficiency, drug release, and in vitro/in vivo applications. Also, possible drug-CD interactions are mentioned and compared according to the performance of the final nanocomplex, Table 4. 4.1. Drug loading, uptake, and release CDs prepared through green hydrothermolysis of milk could serve as an environment-friendly drug delivery system capable of nucleus-targeted treatment and fluorescent tracking simultaneously. The data revealed that the comparative cellular uptake of CD-DOX complexes was lower than the free-DOX based on the detected fluorescent signals, which presumably owning the slow release profile of DOX from the CD-DOX complex since unreleased DOX and CDs might not be distinguished. The maximum amount of DOX released at pH 5.0, which showed that CD-DOX could facilitate selective release in acidic environments. Increased therapeutic efficacy of DOX was based on specified drug release in cancer cell nuclei which was confirmed by confocal scanning microscopy (Fig. 3). Making the investigation of free DOX and CD-DOX apoptosis in cancer cells, the complex showed a higher level of apoptosis and a much lower degree of necrosis than free DOX [50]. Eventually, DOX as an effective chemotherapy drug has incredible role in different cancer cell theranostics. Towards addressing free DOX challenges such as systemic toxicity and severe complications, DOX was conjugated to CDs in order to transport energy to DOX by nanoparticle surface energy transfer similar to fluorescence resonance energy transfer. This method has ON/OFF switch to track the DOX loading, uptake, and release [139].

12

Further consideration of milk-derived graphene quantum dot was done by Thakur M et al. for berberine hydrochloride (BHC) anticancer drug delivery through graphene QDs@Cys-BHC complex. BHC drug loading efficiency on GQD was evaluated to be 88%. The complex showed pH-dependent sustained drug release behavior pursuant to Higuchi release model. Apoptosis analysis performed to find out the efficiency of green synthesized drug delivery agent, while cells treated with GQDs@Cys-BHC complex indicated almost all cells passed in late apoptosis with almost a few cells in necrosis phase [140]. D’Souza et al. developed dried shrimps-derived CDs overload by boldine anti-cancer therapeutic could serve as shining carriers for drug delivery application. The MCF-7 cells incubated with boldine-CDs at 48 h displayed proper fluorescence signals, suggested the effective uptake of boldine-CDs by MCF-7 cells through receptormediated endocytosis. In contrast to the free boldine which crosses the cell membrane by passive diffusion, the complex retained the drug activity with the minimal side effects for healthy cells through PH sensitivity of the complex against acidic pH of the tumor site. It might be due to the increase of hydrophilicity and solubility of boldine at acidic PH [133]. Bayda et al. showed that a simple and green synthesized carbon nanoparticle (CNP) from black tea, could serve as a tunable and safe drug delivery nanocarrier with excellent biocompatible properties. Tea-CNP escaped lysosomal entrapment very well, which is another vital feature for drug delivery applications. DOX was loaded on tea-CNPs, and its function was evaluated by in vitro and in vivo studies. The results of this investigation showed that the complex was more effective and toxic to cancer cells than the free DOX. The tumor size of the DOX-CNP treated mice was diminished compared to the tumors of mice treated with free DOX which is due to an improved pharmacokinetics profile [141]. Moreover, as pointed out by Zhang M et al., the fluorescent CDs derived from chitosan, containing amino groups, could successfully be attached to more than one drug without additional modifications which are greatly desired in drug delivery. They designed a PH sensitive drug delivery complex for DOX anticancer agent and anticoagulant heparin. Loading efficiency of DOX was reported 32.2 and for heparin was about 28.45%. CDs–Hep/DOX retained the therapeutic activity of DOX and showed long-term anti-tumor efficacy. Also, the hemolysis degree of CDs–Hep/DOX was greatly lower than that of free-DOX which is attributed to the synergistic effect of heparin. To test the toxicity of CD-Hep for in vivo application, the spider plant model was chosen which illustrated that CD-drug complex was non-toxic and did not hinder plant growth [142]. In another study of drug delivery vehicles, chitosan-CDs were 13

coated on calcium alginate (CA) beads to prepare stimuli-responsive and a sustained drug release system while it releases the drug according to the methicillin-resistance Staphylococcus aureus (MRSA) pathogen amount. The Drug loading efficiency of CD-CA and CA were 78% and 19% respectively which indicated that CD had promoted drug and the beads interaction leading to high drug encapsulation efficiency [143]. In the first study of lignin as the carbon source of fluorescent CDs by Rai S et al., it was synthesized for traceable drug delivery application. Here the green-CD drug loading efficiency was evaluated to be 67.4%. The drug loading mostly causes fluorescence quenching in CDs. Accordingly, the fluorescence emission strength of the lignin CD was increasingly growing during drug release. Maximum curcumin release was about 86.37% [144]. Given that CDs derived from beer same as other CDs were able to merge optical and therapeutic properties to create functional image-guided drug delivery system. After beer CD and DOX conjugation, the fluorescence signals emitted from CDs and DOX provides the ability to trace the entry and release of drug to MCF7 cell line through confocal microscopy. As the concentration raised, the beer CD-DOX presented lower cytotoxicity against free-DOX. The notable reason for this occurrence would be the slow and controlled release of DOX from beer CD-DOX construct which limits DOX side effects. A stronger electrostatic force between the drug and the nuclei can be a substantial factor in the facile and continuous release of the drug from CD after entering the cell cytoplasm [19]. Escherichia coli genomic DNA was used as the source of fluorescent CDs due to its high content of nitrogen and phosphorus elements without the additional doping step. As DNA derived CDs performed properly for cell imaging, hence evaluating it as drug delivery agent with tracking ability was considered by Ding H et al. As a confirmation, propidium iodide(PI) was used as a trial cargo to interact onto DNA-CDs. PI cannot be up taken by live eukaryotic cells, and the wide entrance showed the special ability of DNA-CDs for delivering drugs into cells [38]. Another example of fluorescence nanomaterial is vancomycin that has gone through an environment-friendly single-step method to prepare carbon dot. Flutamide (FLU)-anticancer drug was significantly attached to the CDs via weak van der Waals interactions. The medium pH about 5.2 triggered the drug release from CD, which was attributed to pH-responsive release profile. The fluorescence emission strength of CD remarkably increased during drug detachment 14

from CDs. Also, it was observed that FLU and CD-FLU at the same concentrations, showed similar cytotoxicity which means that CDs had no interfering activity with FLU pharmacodynamics and released FLU effectively inhibited MCF-7 growth [145]. CD-DNA hydrogel was generated by Singh S et al. for DOX delivery with the possibility to screen drug loading and release. CDs were prepared by green synthesis method from cysteamine. CD not only served as a linker for DNA hydrogel network formation, moreover participated in capturing the drug via electrostatic interaction. The drug loading efficiency of 94% was reported for as prepared CD-DNA hydrogel, in which both the negatively charged components (CD and DNA) effected on positively charged DOX loading. The fluorescence strength of CDs was reduced by ~51 % after the Dox addition to CD-DNA hydrogel solution which might be attributed to the quenching effect of DOX encapsulation. Deep-UV fluorescence of the CDs was reserved in the hydrogel which allowed for real-time tracking and demonstrated the level of drug loading efficiency. Therefore, by CDs and DNA conjugation and eventually forming a hydrogel, a novel eco-friendly and biocompatible method for drug delivery has been created with superior performance [146]. Berberine, an alkaloid from the berberis plant with anticancer effect, went through a green hydrothermal approach to generate fluorescent CDs. It was assumed to have promising cancer theranostic application. Through direct injection of Ber-CDs into the tumor site of HepG 2 tumor-bearing mice, the fluorescence signals could be detected at the tumor site. Ex vivo fluorescence imaging also confirmed the finding via stronger fluorescence intensity observed in tumor, kidney, and liver in comparison to other organs indicating Ber-CDs selective tumor accumulation. Also, cancer cell lines were more sensitive to Ber-CDs rather than common cell lines which exhibited Ber-CD antitumor activity. Ber-CD had lower anti-tumor efficiency towards free berberine, but due to its bright optical properties and tumor cell selectivity would have innovative therapeutical application in vivo. Besides, the size and weight of the tumor treated with Ber-CDs are less than free berberine treated ones, which confirmed the better antitumor efficacy Fig. 4 [147]. In another study, Ghosh et al. have recently worked on gene delivery using green CDs and obtained promising theranostic application for Triple-negative breast cancer (TNBC) gene therapy. CDs were synthesized from sweet lemon peel, conjugated with polyamidoamine 15

(PAMAM) dendrimer to prepare CD-PAMAM conjugates (CDPs) and the complex was furtherly connected to RGDS peptide for targeting v 3 integrin which is over expressed in TNBC. The gene of interest was complexed with nanoparticles with high efficiency. Moreover, selective fluorescence quenching of CD in the presence of Cu(II) ion occurred. Since Cu(II) ion concentration remains regulated in TNBC, CDP3 could be an efficient fluorescent gene delivery tool for TNBC treatment [148]. 4.2. Drug and green-CD interactions Generally, there are two main strategies to design drug-CDs conjugates. One is based on noncovalent interaction such as electrostatic attractions, hydrogen bond, hydrophobic interaction, and the other method is covalent bonding mostly via an amide bond between the carboxylic group from CD and the amine group of the drug. Also, there are some molecular linkers for CD and drug interaction [149]. Here we describe the green-CD conjugation principles to another molecule. Different therapeutics can be attached to green-CDs as a drug delivery platform that mentioned above. Some drugs i.e., DOX are weak amphipathic base, so could be positively charged in the physiological pH which is due to amine moieties protonation. They can be simply loaded on negatively charged CD via electrostatic interactions [19,38,143,144]. For instance, Yuan et al. utilized cow milk-derived CD as the green PL carrier for DOX to have higher efficiency. The drug loading occurred through two non-covalent mechanisms:1) electrostatic interactions due to CD negative zeta potential and DOX positivity. 2) H-bonds according to CD and DOX hydrophilic functional groups [50]. In another hand, the covalent bonding between the greenCDs and drugs can be formed either directly or linker-mediated. Vancomycin derived CDs, with functional groups (carboxyl, carbonyl, hydroxyl, and amino) were coated by flutamide anticancer drug via covalent amide bonds [145]. In a report on cow milk-derived graphene quantum dot, cysteamine hydrochloride (Cys-HCl), a biocompatible molecule, was used as the linker to create CD@cys-BHC complex for BHC delivery via Covalent interaction which showed green-CD efficiency while covalently interacted to the therapeutic [140]. Drugs which have synergic effects could be loaded on green-CDs simultaneously due to their suitable and active surface chemistry without any interference effect. Chitosan derived N-doped CD has been synthesized for DOX delivery application. At first, heparin was conjugated to CD through the formation of 16

an amide bond, which maintains the stability and biological activity of heparin. DOX was also added to the nanostructure (CD-Hep) by electrostatic interaction (Zeta potential DOX+ and CD-). The final nanostructure could safely perform in vitro and in vivo examinations as traceable dual drug delivery system for cancer therapy [142]. Accordingly, green-CDs can be conjugated to drugs through covalent interactions, non-covalent bonding, or both concurrently. So, it can create different types of connections with high performance while maintaining the properties of each component. Perhaps more importantly, electrostatic and covalent interactions were almost the same in drug loading yield. Whereas electrostatic binds through the drug release may be a greater tendency for the drug to conjugate the desired target rather than CDs. 5. Conclusions and future perspectives The evolutionary advantages indicated by theranostic agents inspired a considerable amount of efforts for green-CDs synthesis and applications. We have concentrated on the latest trends and advances of fluorescence synthesized CDs for target-specific bioimaging and also drug delivery with the potency for real-time optical tracking of the therapeutic and diagnostic process. A primary conclusion according to as-mentioned summarized research progresses, green-CDs can be developed as amazing drug delivery systems because of their particular features, including superior cargo-loading efficiency, photostability, appropriate quantum yield, and longtime fluorescence. Another area of interest for further development is the targeting of tumor cells and cellular organelles through green-CDs. Although, the intracellular and nuclear localization of majority of the CDs has been likely related with their surface functionalization, we can see that “naked” green-CDs could successfully locate in targeted areas including cancer cells, cytoplasm, and organelles. Green-CDs not only have shown great potential as self-targeted imaging but also, the formation of covalent or electrostatic bonding of targeting ligands with green-CDs is possible based on their dynamic/self-passivated surface chemistry. The bio-inspired CDs are supposed to have unique optical properties due to hetero-atom doping, stems from proteins and carbohydrates of natural precursors. They have notably retained fluorescence properties in conjugation with targeting or therapeutic agents which makes them appropriate for the biomedical application. It is worthy to note that in vivo fluorescence imaging suffers from weak tissue penetration and autofluorescence background. These are the main challenges of fluorescent NPs clinical 17

translation. Near-infrared spectroscopy (NIR), corresponding to the wavelength range of 700– 900 nm, with the advantages of decreased autofluorescence and higher depth of penetration, is suitable for in vivo imaging. If we focus on how to shift green-CDs emission into the NIR area, with all of their remarkable properties, there would be no limits for their clinical use. Today, since green-CDs have worked well in the fields of drug delivery or targeted-imaging, it would be an initiation of an amazing revolution in cancer theranostic. To fully understand the concept of green-CD promises, extra research efforts must be directed concerning different types of therapeutics (drugs, nucleic acids, antibodies,) that can be loaded on CDs and evaluating them in vivo performance more widely. Obviously, with the progress in the development of green-CDs and their various practical applications, they are undoubtedly the next generation material among carbon structures.

Conflicts The authors declare that they have no competing interests. Acknowledgement: This work was supported by Tabriz University of Medical Science through the Academic Research Fund under Projects NO.59018. Also, we would like to acknowledge the contribution and assistance of Dr. Ebrahim Mostafavi in this maneuscript.

18

References [1]

J. Li, X. Chang, X. Chen, Z. Gu, F. Zhao, Z. Chai, Y. Zhao, Toxicity of inorganic nanomaterials in biomedical imaging, Biotechnol. Adv. 32 (2014) 727–743. doi:https://doi.org/10.1016/j.biotechadv.2013.12.009.

[2]

S.Y. Lim, W. Shen, Z. Gao, Carbon quantum dots and their applications, Chem. Soc. Rev. 44 (2015) 362–381. doi:10.1039/C4CS00269E.

[3]

A. Valizadeh, H. Mikaeili, M. Samiei, S.M. Farkhani, N. Zarghami, A. Akbarzadeh, S. Davaran, Quantum dots: synthesis, bioapplications, and toxicity, Nanoscale Res. Lett. 7 (2012) 480. doi:https://doi.org/10.1186/1556-276X-7-480.

[4]

Z. Peng, X. Han, S. Li, A.O. Al-Youbi, A.S. Bashammakh, M.S. El-Shahawi, R.M. Leblanc, Carbon dots: biomacromolecule interaction, bioimaging and nanomedicine, Coord. Chem. Rev. 343 (2017) 256–277. doi:https://doi.org/10.1016/j.ccr.2017.06.001.

[5]

H. Ding, S.-B. Yu, J.-S. Wei, H.-M. Xiong, Full-color light-emitting carbon dots with a surface-state-controlled luminescence mechanism, ACS Nano. 10 (2015) 484–491. doi:https://doi.org/10.1021/acsnano.5b05406.

[6]

X. Xu, R. Ray, Y. Gu, H.J. Ploehn, L. Gearheart, K. Raker, W.A. Scrivens, Electrophoretic analysis and purification of fluorescent single-walled carbon nanotube fragments, J. Am. Chem. Soc. 126 (2004) 12736–12737. doi:https://doi.org/10.1021/ja040082h.

[7]

Y.-P. Sun, B. Zhou, Y. Lin, W. Wang, K.A.S. Fernando, P. Pathak, M.J. Meziani, B.A. Harruff, X. Wang, H. Wang, Quantum-sized carbon dots for bright and colorful photoluminescence, J. Am. Chem. Soc. 128 (2006) 7756–7757. doi:https://doi.org/10.1021/ja062677d.

[8]

V. Sharma, P. Tiwari, S.M. Mobin, Sustainable carbon-dots: recent advances in green carbon dots for sensing and bioimaging, J. Mater. Chem. B. 5 (2017) 8904–8924. doi:10.1039/C7TB02484C.

[9]

M. Zhou, Z. Zhou, A. Gong, Y. Zhang, Q. Li, Synthesis of highly photoluminescent carbon dots via citric acid and Tris for iron (III) ions sensors and bioimaging, Talanta. 143 19

(2015) 107–113. doi:https://doi.org/10.1016/j.talanta.2015.04.015. [10]

C. Wang, Z. Xu, C. Zhang, Polyethyleneimine‐functionalized fluorescent carbon dots: water stability, pH sensing, and cellular imaging, ChemNanoMat. 1 (2015) 122–127. doi:https://doi.org/10.1002/cnma.201500009.

[11]

K. Yang, M. Liu, Y. Wang, S. Wang, H. Miao, L. Yang, X. Yang, Carbon dots derived from fungus for sensing hyaluronic acid and hyaluronidase, Sensors Actuators B Chem. 251 (2017) 503–508. doi:https://doi.org/10.1016/j.snb.2017.05.086.

[12]

J. Zhang, Y. Yuan, G. Liang, S. Yu, Scale‐Up Synthesis of Fragrant Nitrogen‐Doped Carbon Dots from Bee Pollens for Bioimaging and Catalysis, Adv. Sci. 2 (2015) 1500002. doi:https://doi.org/10.1002/advs.201500002.

[13]

L. Qiao, T. Sun, X. Zheng, M. Zheng, Z. Xie, Exploring the optimal ratio of d-glucose/laspartic acid for targeting carbon dots toward brain tumor cells, Mater. Sci. Eng. C. 85 (2018) 1–6. doi:https://doi.org/10.1016/j.msec.2017.12.011.

[14]

R. Atchudan, T.N.J.I. Edison, S. Perumal, N.C.S. Selvam, Y.R. Lee, Green synthesized multiple fluorescent nitrogen-doped carbon quantum dots as an efficient label-free optical nanoprobe for in vivo live-cell imaging, J. Photochem. Photobiol. A Chem. 372 (2019) 99–107. doi:https://doi.org/10.1016/j.jphotochem.2018.12.011.

[15]

W. Lu, X. Qin, S. Liu, G. Chang, Y. Zhang, Y. Luo, A.M. Asiri, A.O. Al-Youbi, X. Sun, Economical, green synthesis of fluorescent carbon nanoparticles and their use as probes for sensitive and selective detection of mercury (II) ions, Anal. Chem. 84 (2012) 5351– 5357. doi:https://doi.org/10.1021/ac3007939.

[16]

S. Sahu, B. Behera, T.K. Maiti, S. Mohapatra, Simple one-step synthesis of highly luminescent carbon dots from orange juice: application as excellent bio-imaging agents, Chem. Commun. 48 (2012) 8835–8837. doi:10.1039/C2CC33796G.

[17]

P.-C. Hsu, Z.-Y. Shih, C.-H. Lee, H.-T. Chang, Synthesis and analytical applications of photoluminescent carbon nanodots, Green Chem. 14 (2012) 917–920. doi:10.1039/C2GC16451E.

[18]

L. Wang, H.S. Zhou, Green synthesis of luminescent nitrogen-doped carbon dots from 20

milk and its imaging application, Anal. Chem. 86 (2014) 8902–8905. doi:https://doi.org/10.1021/ac502646x. [19]

Z. Wang, H. Liao, H. Wu, B. Wang, H. Zhao, M. Tan, Fluorescent carbon dots from beer for breast cancer cell imaging and drug delivery, Anal. Methods. 7 (2015) 8911–8917. doi:10.1039/C5AY01978H.

[20]

A. Konwar, N. Gogoi, G. Majumdar, D. Chowdhury, Green chitosan–carbon dots nanocomposite hydrogel film with superior properties, Carbohydr. Polym. 115 (2015) 238–245. doi:https://doi.org/10.1016/j.carbpol.2014.08.021.

[21]

J. Wang, C. Wang, S. Chen, Amphiphilic egg‐derived carbon dots: Rapid plasma fabrication, pyrolysis process, and multicolor printing patterns, Angew. Chemie Int. Ed. 51 (2012) 9297–9301. doi:https://doi.org/10.1002/anie.201204381.

[22]

X. Yang, Y. Zhuo, S. Zhu, Y. Luo, Y. Feng, Y. Dou, Novel and green synthesis of highfluorescent carbon dots originated from honey for sensing and imaging, Biosens. Bioelectron. 60 (2014) 292–298. doi:https://doi.org/10.1016/j.bios.2014.04.046.

[23]

Y.-Y. Yao, G. Gedda, W.M. Girma, C.-L. Yen, Y.-C. Ling, J.-Y. Chang, Magnetofluorescent carbon dots derived from crab shell for targeted dual-modality bioimaging and drug delivery, ACS Appl. Mater. Interfaces. 9 (2017) 13887–13899. doi:https://doi.org/10.1021/acsami.7b01599.

[24]

X. Liu, J. Pang, F. Xu, X. Zhang, Simple approach to synthesize amino-functionalized carbon dots by carbonization of chitosan, Sci. Rep. 6 (2016) 31100. doi:https://doi.org/10.1038/srep31100.

[25]

Y. Guo, L. Zhang, F. Cao, Y. Leng, Thermal treatment of hair for the synthesis of sustainable carbon quantum dots and the applications for sensing Hg 2+, Sci. Rep. 6 (2016) 35795. doi:https://doi.org/10.1038/srep35795.

[26]

J.B. Essner, C.H. Laber, S. Ravula, L. Polo-Parada, G.A. Baker, Pee-dots: biocompatible fluorescent carbon dots derived from the upcycling of urine, Green Chem. 18 (2016) 243– 250. doi:10.1039/C5GC02032H.

[27]

S. Cong, N. Wang, K. Wang, Y. Wu, D. Li, Y. Song, S. Prakash, M. Tan, Fluorescent 21

nanoparticles in the popular pizza: properties, biodistribution and cytotoxicity, Food Funct. 10 (2019) 2408–2416. doi:10.1039/C8FO01944D. [28]

Y. Song, Y. Wu, H. Wang, S. Liu, L. Song, S. Li, M. Tan, Carbon quantum dots from roasted Atlantic salmon (Salmo salar L.): Formation, biodistribution and cytotoxicity, Food Chem. 293 (2019) 387–395. doi:https://doi.org/10.1016/j.foodchem.2019.05.017.

[29] X. Song, L. Cao, S. Cong, Y. Song, M. Tan, Characterization of Endogenous Nanoparticles from Roasted Chicken Breasts, J. Agric. Food Chem. 66 (2018) 7522–7530. doi:https://doi.org/10.1021/acs.jafc.8b01988. [30] X.W. Tan, A.N.B. Romainor, S.F. Chin, S.M. Ng, Carbon dots production via pyrolysis of sago waste as potential probe for metal ions sensing, J. Anal. Appl. Pyrolysis. 105 (2014) 157–165. doi:https://doi.org/10.1016/j.jaap.2013.11.001. [31]

S. Thambiraj, R. Shankaran, Green synthesis of highly fluorescent carbon quantum dots from sugarcane bagasse pulp, Appl. Surf. Sci. 390 (2016) 435–443. doi:https://doi.org/10.1016/j.apsusc.2016.08.106.

[32]

L. Zhu, Y. Yin, C.-F. Wang, S. Chen, Plant leaf-derived fluorescent carbon dots for sensing, patterning and coding, J. Mater. Chem. C. 1 (2013) 4925–4932. doi:10.1039/C3TC30701H.

[33]

V. Ramanan, S.K. Thiyagarajan, K. Raji, R. Suresh, R. Sekar, P. Ramamurthy, Outright green synthesis of fluorescent carbon dots from eutrophic algal blooms for in vitro imaging, ACS Sustain. Chem. Eng. 4 (2016) 4724–4731. doi:https://doi.org/10.1021/acssuschemeng.6b00935.

[34]

J. Wang, Y.H. Ng, Y.-F. Lim, G.W. Ho, Vegetable-extracted carbon dots and their nanocomposites for enhanced photocatalytic H 2 production, Rsc Adv. 4 (2014) 44117– 44123. doi:10.1039/C4RA07290A.

[35]

Q. Yang, L. Wei, X. Zheng, L. Xiao, Single particle dynamic imaging and Fe 3+ sensing with bright carbon dots derived from bovine serum albumin proteins, Sci. Rep. 5 (2015) 17727. doi:https://doi.org/10.1038/srep17727.

[36]

Z. Wang, C. Xu, Y. Lu, X. Chen, H. Yuan, G. Wei, G. Ye, J. Chen, Fluorescence sensor 22

array based on amino acid derived carbon dots for pattern-based detection of toxic metal ions, Sensors Actuators B Chem. 241 (2017) 1324–1330. doi:https://doi.org/10.1016/j.snb.2016.09.186. [37]

H. Qi, M. Teng, M. Liu, S. Liu, J. Li, H. Yu, C. Teng, Z. Huang, H. Liu, Q. Shao, Biomass-derived nitrogen-doped carbon quantum dots: highly selective fluorescent probe for detecting Fe3+ ions and tetracyclines, J. Colloid Interface Sci. 539 (2019) 332–341. doi:https://doi.org/10.1016/j.jcis.2018.12.047.

[38]

H. Ding, F. Du, P. Liu, Z. Chen, J. Shen, DNA–carbon dots function as fluorescent vehicles for drug delivery, ACS Appl. Mater. Interfaces. 7 (2015) 6889–6897. doi:https://doi.org/10.1021/acsami.5b00628.

[39]

S.K. Bhunia, N. Pradhan, N.R. Jana, Vitamin B1 derived blue and green fluorescent carbon nanoparticles for cell-imaging application, ACS Appl. Mater. Interfaces. 6 (2014) 7672–7679. doi:https://doi.org/10.1021/am500964d.

[40]

H. Peng, J. Travas-Sejdic, Simple aqueous solution route to luminescent carbogenic dots from carbohydrates, Chem. Mater. 21 (2009) 5563–5565. doi:https://doi.org/10.1021/cm901593y.

[41]

Q. Tang, W. Zhu, B. He, P. Yang, Rapid conversion from carbohydrates to large-scale carbon quantum dots for all-weather solar cells, ACS Nano. 11 (2017) 1540–1547. doi:https://doi.org/10.1021/acsnano.6b06867.

[42]

C.J. Jeong, A.K. Roy, S.H. Kim, J.-E. Lee, J.H. Jeong, I. In, S.Y. Park, Fluorescent carbon nanoparticles derived from natural materials of mango fruit for bio-imaging probes, Nanoscale. 6 (2014) 15196–15202. doi:10.1039/C4NR04805A.

[43]

C.-Y. Liang, W. Xia, C.-Z. Yang, Y.-C. Liu, A.-M. Bai, Y.-J. Hu, Exploring the binding of carbon dots to calf thymus DNA: From green synthesis to fluorescent molecular probe, Carbon N. Y. (2018). doi:https://doi.org/10.1016/j.carbon.2018.01.009.

[44]

X. Zhao, J. Zhang, L. Shi, M. Xian, C. Dong, S. Shuang, Folic acid-conjugated carbon dots as green fluorescent probes based on cellular targeting imaging for recognizing cancer cells, RSC Adv. 7 (2017) 42159–42167. doi:10.1039/C7RA07002K. 23

[45]

M. Dadashpour, A. Firouzi-Amandi, M. Pourhassan-Moghaddam, M.J. Maleki, N. Soozangar, F. Jeddi, M. Nouri, N. Zarghami, Y. Pilehvar-Soltanahmadi, Biomimetic synthesis of silver nanoparticles using Matricaria chamomilla extract and their potential anticancer activity against human lung cancer cells, Mater. Sci. Eng. C. 92 (2018) 902– 912. doi:https://doi.org/10.1016/j.msec.2018.07.053.

[46]

C.T. Matea, T. Mocan, F. Tabaran, T. Pop, O. Mosteanu, C. Puia, C. Iancu, L. Mocan, Quantum dots in imaging, drug delivery and sensor applications, Int. J. Nanomedicine. 12 (2017) 5421. doi:10.2147/IJN.S138624.

[47]

H. Wang, X. Li, B.W.-C. Tse, H. Yang, C.A. Thorling, Y. Liu, M. Touraud, J.B. Chouane, X. Liu, M.S. Roberts, Indocyanine green-incorporating nanoparticles for cancer theranostics, Theranostics. 8 (2018) 1227. doi:10.7150/thno.22872.

[48]

H. Huang, J.F. Lovell, Advanced functional nanomaterials for theranostics, Adv. Funct. Mater. 27 (2017) 1603524. doi:https://doi.org/10.1002/adfm.201603524.

[49]

M.K. Kumawat, M. Thakur, R. Bahadur, T. Kaku, R.S. Prabhuraj, A. Ninawe, R. Srivastava, Preparation of graphene oxide-graphene quantum dots hybrid and its application in cancer theranostics, Mater. Sci. Eng. C. (2019) 109774. doi:10.1039/C4NR05712K.

[50]

Y. Yuan, B. Guo, L. Hao, N. Liu, Y. Lin, W. Guo, X. Li, B. Gu, Doxorubicin-loaded environmentally friendly carbon dots as a novel drug delivery system for nucleus targeted cancer therapy, Colloids Surfaces B Biointerfaces. 159 (2017) 349–359. doi:https://doi.org/10.1016/j.colsurfb.2017.07.030.

[51]

X. Wei, L. Li, J. Liu, L. Yu, H. Li, F. Cheng, X. Yi, J. He, B. Li, Green Synthesis of Fluorescent Carbon Dots from Gynostemma for Bioimaging and Antioxidant in Zebrafish, ACS Appl. Mater. Interfaces. (2019). doi:https://doi.org/10.1021/acsami.9b00074.

[52]

P. Gong, L. Sun, F. Wang, X. Liu, Z. Yan, M. Wang, L. Zhang, Z. Tian, Z. Liu, J. You, Highly fluorescent N-doped carbon dots with two-photon emission for ultrasensitive detection of tumor marker and visual monitor anticancer drug loading and delivery, Chem. Eng. J. 356 (2019) 994–1002. doi:https://doi.org/10.1016/j.cej.2018.09.100.

24

[53]

H. Wang, J. Bi, B.-W. Zhu, M. Tan, Multicolorful Carbon Dots for Tumor Theranostics, Curr. Med. Chem. 25 (2017) 2894–2909. doi:10.2174/0929867324666170316110810.

[54] W.-J. Wang, J.-M. Xia, J. Feng, M.-Q. He, M.-L. Chen, J.-H. Wang, Green preparation of carbon dots for intracellular pH sensing and multicolor live cell imaging, J. Mater. Chem. B. 4 (2016) 7130–7137. doi:10.1039/C6TB02071B. [55]

H. Miao, L. Wang, Y. Zhuo, Z. Zhou, X. Yang, Label-free fluorimetric detection of CEA using carbon dots derived from tomato juice, Biosens. Bioelectron. 86 (2016) 83–89. doi:Label-free fluorimetric detection of CEA using carbon dots derived from tomato juice.

[56]

Z. Yang, Z. Li, M. Xu, Y. Ma, J. Zhang, Y. Su, F. Gao, H. Wei, L. Zhang, Controllable synthesis of fluorescent carbon dots and their detection application as nanoprobes, NanoMicro Lett. 5 (2013) 247–259. doi:https://doi.org/10.1007/BF03353756.

[57]

P. Miao, K. Han, Y. Tang, B. Wang, T. Lin, W. Cheng, Recent advances in carbon nanodots: synthesis, properties and biomedical applications, Nanoscale. 7 (2015) 1586– 1595. doi:10.1039/C4NR05712K.

[58]

J. Zhou, C. Booker, R. Li, X. Zhou, T.-K. Sham, X. Sun, Z. Ding, An electrochemical avenue to blue luminescent nanocrystals from multiwalled carbon nanotubes (MWCNTs), J. Am. Chem. Soc. 129 (2007) 744–745. doi:https://doi.org/10.1021/ja0669070.

[59]

X.T. Zheng, A. Ananthanarayanan, K.Q. Luo, P. Chen, Glowing graphene quantum dots and carbon dots: properties, syntheses, and biological applications, Small. 11 (2015) 1620–1636. doi:https://doi.org/10.1002/smll.201402648.

[60]

H. Li, X. He, Y. Liu, H. Huang, S. Lian, S.-T. Lee, Z. Kang, One-step ultrasonic synthesis of water-soluble carbon nanoparticles with excellent photoluminescent properties, Carbon N. Y. 49 (2011) 605–609. doi:https://doi.org/10.1016/j.carbon.2010.10.004.

[61]

C. Wang, Z. Xu, H. Cheng, H. Lin, M.G. Humphrey, C. Zhang, A hydrothermal route to water-stable luminescent carbon dots as nanosensors for pH and temperature, Carbon N. Y. 82 (2015) 87–95. doi:https://doi.org/10.1016/j.carbon.2014.10.035.

[62]

Y. Choi, N. Thongsai, A. Chae, S. Jo, E.B. Kang, P. Paoprasert, S.Y. Park, I. In, Microwave-assisted synthesis of luminescent and biocompatible lysine-based carbon 25

quantum dots, J. Ind. Eng. Chem. 47 (2017) 329–335. doi:https://doi.org/10.1016/j.jiec.2016.12.002. [63]

J.H. Bang, K.S. Suslick, Applications of ultrasound to the synthesis of nanostructured materials, Adv. Mater. 22 (2010) 1039–1059. doi:https://doi.org/10.1002/adma.200904093.

[64]

P. Karfa, S. De, K.C. Majhi, R. Madhuri, P.K. Sharma, Functionalization of Carbon Nanostructures, (2018).

[65]

C. Yang, R. Ogaki, L. Hansen, J. Kjems, B.M. Teo, Theranostic carbon dots derived from garlic with efficient anti-oxidative effects towards macrophages, RSC Adv. 5 (2015) 97836–97840. doi:10.1039/C5RA16874K.

[66]

K. Bankoti, A.P. Rameshbabu, S. Datta, B. Das, A. Mitra, S. Dhara, Onion derived carbon nanodots for live cell imaging and accelerated skin wound healing, J. Mater. Chem. B. 5 (2017) 6579–6592. doi:10.1039/C7TB00869D.

[67]

Q. Wang, X. Liu, L. Zhang, Y. Lv, Microwave-assisted synthesis of carbon nanodots through an eggshell membrane and their fluorescent application, Analyst. 137 (2012) 5392–5397. doi:10.1039/C2AN36059D.

[68]

C. Yu, T. Xuan, Y. Chen, Z. Zhao, Z. Sun, H. Li, A facile, green synthesis of highly fluorescent carbon nanoparticles from oatmeal for cell imaging, J. Mater. Chem. C. 3 (2015) 9514–9518. doi:10.1039/C5TC02057C.

[69]

S. Chandra, S.H. Pathan, S. Mitra, B.H. Modha, A. Goswami, P. Pramanik, Tuning of photoluminescence on different surface functionalized carbon quantum dots, Rsc Adv. 2 (2012) 3602–3606. doi:10.1039/C2RA00030J.

[70]

M. Yuan, R. Zhong, H. Gao, W. Li, X. Yun, J. Liu, X. Zhao, G. Zhao, F. Zhang, One-step, green, and economic synthesis of water-soluble photoluminescent carbon dots by hydrothermal treatment of wheat straw, and their bio-applications in labeling, imaging, and sensing, Appl. Surf. Sci. 355 (2015) 1136–1144. doi:https://doi.org/10.1016/j.apsusc.2015.07.095.

[71]

Y. Liu, Y. Zhao, Y. Zhang, One-step green synthesized fluorescent carbon nanodots from 26

bamboo leaves for copper (II) ion detection, Sensors Actuators B Chem. 196 (2014) 647– 652. doi:https://doi.org/10.1016/j.snb.2014.02.053. [72]

P. Roy, P.-C. Chen, A.P. Periasamy, Y.-N. Chen, H.-T. Chang, Photoluminescent carbon nanodots: synthesis, physicochemical properties and analytical applications, Mater. Today. 18 (2015) 447–458. doi:https://doi.org/10.1016/j.mattod.2015.04.005.

[73]

B. De, N. Karak, Recent progress in carbon dot–metal based nanohybrids for photochemical and electrochemical applications, J. Mater. Chem. A. 5 (2017) 1826–1859.

[74]

J. Yu, N. Song, Y.-K. Zhang, S.-X. Zhong, A.-J. Wang, J. Chen, Green preparation of carbon dots by Jinhua bergamot for sensitive and selective fluorescent detection of Hg2+ and Fe3+, Sensors Actuators B Chem. 214 (2015) 29–35. doi:https://doi.org/10.1016/j.snb.2015.03.006.

[75]

H. Ding, Y. Ji, J.-S. Wei, Q.-Y. Gao, Z.-Y. Zhou, H.-M. Xiong, Facile synthesis of redemitting carbon dots from pulp-free lemon juice for bioimaging, J. Mater. Chem. B. 5 (2017) 5272–5277. doi:10.1039/C7TB01130J.

[76]

A. Prasannan, T. Imae, One-pot synthesis of fluorescent carbon dots from orange waste peels, Ind. Eng. Chem. Res. 52 (2013) 15673–15678. doi:https://doi.org/10.1021/ie402421s.

[77]

J. Shen, S. Shang, X. Chen, D. Wang, Y. Cai, Facile synthesis of fluorescence carbon dots from sweet potato for Fe3+ sensing and cell imaging, Mater. Sci. Eng. C. 76 (2017) 856– 864. doi:https://doi.org/10.1016/j.msec.2017.03.178.

[78]

V.N. Mehta, S. Jha, H. Basu, R.K. Singhal, S.K. Kailasa, One-step hydrothermal approach to fabricate carbon dots from apple juice for imaging of mycobacterium and fungal cells, Sensors Actuators B Chem. 213 (2015) 434–443. doi:https://doi.org/10.1016/j.snb.2015.02.104.

[79]

A.-M. Alam, B.-Y. Park, Z.K. Ghouri, M. Park, H.-Y. Kim, Synthesis of carbon quantum dots from cabbage with down-and up-conversion photoluminescence properties: excellent imaging agent for biomedical applications, Green Chem. 17 (2015) 3791–3797. doi:10.1039/C5GC00686D. 27

[80]

B.S.B. Kasibabu, S.L. D’souza, S. Jha, S.K. Kailasa, Imaging of bacterial and fungal cells using fluorescent carbon dots prepared from carica papaya juice, J. Fluoresc. 25 (2015) 803–810. doi:https://doi.org/10.1007/s10895-015-1595-0.

[81]

S. Zhao, M. Lan, X. Zhu, H. Xue, T.-W. Ng, X. Meng, C.-S. Lee, P. Wang, W. Zhang, Green synthesis of bifunctional fluorescent carbon dots from garlic for cellular imaging and free radical scavenging, ACS Appl. Mater. Interfaces. 7 (2015) 17054–17060. doi:https://doi.org/10.1021/acsami.5b03228.

[82]

C.-L. Li, C.-M. Ou, C.-C. Huang, W.-C. Wu, Y.-P. Chen, T.-E. Lin, L.-C. Ho, C.-W. Wang, C.-C. Shih, H.-C. Zhou, Carbon dots prepared from ginger exhibiting efficient inhibition of human hepatocellular carcinoma cells, J. Mater. Chem. B. 2 (2014) 4564– 4571. doi:10.1039/C4TB00216D.

[83]

J. Xu, T. Lai, Z. Feng, X. Weng, C. Huang, Formation of fluorescent carbon nanodots from kitchen wastes and their application for detection of Fe3+, Luminescence. 30 (2015) 420–424. doi:https://doi.org/10.1002/bio.2754.

[84]

H. Huang, Y. Xu, C.-J. Tang, J.-R. Chen, A.-J. Wang, J.-J. Feng, Facile and green synthesis of photoluminescent carbon nanoparticles for cellular imaging, New J. Chem. 38 (2014) 784–789. doi:10.1039/C3NJ01185B.

[85]

S. Liu, J. Tian, L. Wang, Y. Zhang, X. Qin, Y. Luo, A.M. Asiri, A.O. Al‐Youbi, X. Sun, Hydrothermal treatment of grass: a low‐cost, green route to nitrogen‐doped, carbon‐rich, photoluminescent polymer nanodots as an effective fluorescent sensing platform for label‐free detection of Cu (II) ions, Adv. Mater. 24 (2012) 2037–2041.

[86]

Y. Yang, J. Cui, M. Zheng, C. Hu, S. Tan, Y. Xiao, Q. Yang, Y. Liu, One-step synthesis of amino-functionalized fluorescent carbon nanoparticles by hydrothermal carbonization of chitosan, Chem. Commun. 48 (2012) 380–382. doi:10.1039/C1CC15678K.

[87]

X. Song, H. Wang, R. Zhang, C. Yu, M. Tan, Bio-distribution and interaction with dopamine of fluorescent nanodots from roasted chicken, Food Funct. 9 (2018) 6227–6235. doi:10.1039/C8FO01159A.

[88]

J. Bi, Y. Li, H. Wang, Y. Song, S. Cong, C. Yu, B.-W. Zhu, M. Tan, Presence and 28

formation mechanism of foodborne carbonaceous nanostructures from roasted pike eel (Muraenesox cinereus), J. Agric. Food Chem. 66 (2017) 2862–2869. doi:https://doi.org/10.1021/acs.jafc.7b02303. [89] S. Li, C. Jiang, H. Wang, S. Cong, M. Tan, Fluorescent nanoparticles present in CocaCola and Pepsi-Cola: physiochemical properties, cytotoxicity, biodistribution and digestion studies, Nanotoxicology. 12 (2018) 49–62. doi:doi.org/10.1080/17435390.2017.1418443. [90]

H. Wang, S. Liu, Y. Song, B.-W. Zhu, M. Tan, Universal existence of fluorescent carbon dots in beer and assessment of their potential toxicity, Nanotoxicology. 13 (2019) 160– 173. doi:doi.org/10.1080/17435390.2018.1530394.

[91] S. Cong, J. Bi, X. Song, C. Yu, M. Tan, Ultrasmall fluorescent nanoparticles derived from roast duck: their physicochemical characteristics and interaction with human serum albumin, Food Funct. 9 (2018) 2490–2495. doi:10.1039/C8FO00178B. [92] Y. Li, J. Bi, S. Liu, H. Wang, C. Yu, D. Li, B.-W. Zhu, M. Tan, Presence and formation of fluorescence carbon dots in a grilled hamburger, Food Funct. 8 (2017) 2558–2565. doi:10.1039/C7FO00675F. [93] H. Wang, Y. Xie, S. Liu, S. Cong, Y. Song, X. Xu, M. Tan, Presence of fluorescent carbon nanoparticles in baked lamb: their properties and potential application for sensors, J. Agric. Food Chem. 65 (2017) 7553–7559. doi:10.1021/acs.jafc.7b02913. [94]

L. Cao, X. Song, Y. Song, J. Bi, S. Cong, C. Yu, M. Tan, Fluorescent nanoparticles from mature vinegar: their properties and interaction with dopamine, Food Funct. 8 (2017) 4744–4751. doi:10.1039/C7FO01475A.

[95] C. Jiang, H. Wu, X. Song, X. Ma, J. Wang, M. Tan, Presence of photoluminescent carbon dots in Nescafe® original instant coffee: applications to bioimaging, Talanta. 127 (2014) 68–74. [96]

H. Liao, C. Jiang, W. Liu, J.M. Vera, O.D. Seni, K. Demera, C. Yu, M. Tan, Fluorescent nanoparticles from several commercial beverages: their properties and potential application for bioimaging, J. Agric. Food Chem. 63 (2015) 8527–8533. 29

doi:https://doi.org/10.1021/acs.jafc.5b04216. [97] J. Wang, J. Qiu, A review of carbon dots in biological applications, J. Mater. Sci. 51 (2016) 4728–4738. doi:https://doi.org/10.1007/s10853-016-9797-7. [98]

N. Murugan, M. Prakash, M. Jayakumar, A. Sundaramurthy, A.K. Sundramoorthy, Green synthesis of fluorescent carbon quantum dots from Eleusine coracana and their application as a fluorescence ‘turn-off’sensor probe for selective detection of Cu2+, Appl. Surf. Sci. 476 (2019) 468–480. doi:doi.org/10.1016/j.apsusc.2019.01.090.

[99]

K.P. Loh, Q. Bao, G. Eda, M. Chhowalla, Graphene oxide as a chemically tunable platform for optical applications, Nat. Chem. 2 (2010) 1015. doi:https://doi.org/10.1038/nchem.907.

[100] J. Manioudakis, F. Victoria, C.A. Thompson, L. Brown, M. Movsum, R. Lucifero, R. Naccache, Effects of nitrogen-doping on the photophysical properties of carbon dots, J. Mater. Chem. C. 7 (2019) 853–862. doi:10.1039/C8TC04821E. [101] Q. Xu, T. Kuang, Y. Liu, L. Cai, X. Peng, T.S. Sreeprasad, P. Zhao, Z. Yu, N. Li, Heteroatom-doped carbon dots: synthesis, characterization, properties, photoluminescence mechanism and biological applications, J. Mater. Chem. B. 4 (2016) 7204–7219. doi:10.1039/C6TB02131J. [102] Y. Park, J. Yoo, B. Lim, W. Kwon, S.-W. Rhee, Improving the functionality of carbon nanodots: doping and surface functionalization, J. Mater. Chem. A. 4 (2016) 11582– 11603. doi:10.1039/C6TA04813G. [103] J. Liao, Z. Cheng, L. Zhou, Nitrogen-doping enhanced fluorescent carbon dots: green synthesis and their applications for bioimaging and label-free detection of Au3+ ions, ACS Sustain. Chem. Eng. 4 (2016) 3053–3061. doi:https://doi.org/10.1021/acssuschemeng.6b00018. [104] J.R. Bhamore, S. Jha, R.K. Singhal, T.J. Park, S.K. Kailasa, Facile green synthesis of carbon dots from Pyrus pyrifolia fruit for assaying of Al3+ ion via chelation enhanced fluorescence mechanism, J. Mol. Liq. 264 (2018) 9–16. doi:https://doi.org/10.1016/j.molliq.2018.05.041. 30

[105] L. Bao, Z. Zhang, Z. Tian, L. Zhang, C. Liu, Y. Lin, B. Qi, D. Pang, Electrochemical tuning of luminescent carbon nanodots: from preparation to luminescence mechanism, Adv. Mater. 23 (2011) 5801–5806. doi:https://doi.org/10.1002/adma.201102866. [106] A.R. Chowdhuri, S. Tripathy, C. Haldar, S. Roy, S.K. Sahu, Single step synthesis of carbon dot embedded chitosan nanoparticles for cell imaging and hydrophobic drug delivery, J. Mater. Chem. B. 3 (2015) 9122–9131. doi:10.1039/C5TB01831E. [107] T. Feng, X. Ai, G. An, P. Yang, Y. Zhao, Charge-convertible carbon dots for imagingguided drug delivery with enhanced in vivo cancer therapeutic efficiency, ACS Nano. 10 (2016) 4410–4420. doi:https://doi.org/10.1021/acsnano.6b00043. [108] N. Gao, W. Yang, H. Nie, Y. Gong, J. Jing, L. Gao, X. Zhang, Turn-on theranostic fluorescent nanoprobe by electrostatic self-assembly of carbon dots with doxorubicin for targeted cancer cell imaging, in vivo hyaluronidase analysis, and targeted drug delivery, Biosens. Bioelectron. 96 (2017) 300–307. doi:https://doi.org/10.1016/j.bios.2017.05.019. [109] J. Pardo, Z. Peng, R.M. Leblanc, Cancer targeting and drug delivery using carbon-based quantum dots and nanotubes, Molecules. 23 (2018) 378. doi:https://doi.org/10.3390/molecules23020378. [110] D. Feng, Y. Song, W. Shi, X. Li, H. Ma, Distinguishing folate-receptor-positive cells from folate-receptor-negative cells using a fluorescence off–on nanoprobe, Anal. Chem. 85 (2013) 6530–6535. doi:https://doi.org/10.1021/ac401377n. [111] Q. Duan, M. Che, S. Hu, H. Zhao, Y. Li, X. Ma, W. Zhang, Y. Zhang, S. Sang, Rapid cancer diagnosis by highly fluorescent carbon nanodots-based imaging, Anal. Bioanal. Chem. 411 (2019) 967–972. doi:https://doi.org/10.1007/s00216-018-1500-1. [112] M. Zheng, S. Ruan, S. Liu, T. Sun, D. Qu, H. Zhao, Z. Xie, H. Gao, X. Jing, Z. Sun, Selftargeting fluorescent carbon dots for diagnosis of brain cancer cells, ACS Nano. 9 (2015) 11455–11461. doi:https://doi.org/10.1021/acsnano.5b05575. [113] S.K. Bhunia, A.R. Maity, S. Nandi, D. Stepensky, R. Jelinek, Imaging cancer cells expressing the folate receptor with carbon dots produced from folic acid, ChemBioChem. 17 (2016) 614–619. doi:https://doi.org/10.1002/cbic.201500694. 31

[114] H. Liu, Z. Li, Y. Sun, X. Geng, Y. Hu, H. Meng, J. Ge, L. Qu, Synthesis of Luminescent Carbon Dots with Ultrahigh Quantum Yield and Inherent Folate Receptor-Positive Cancer Cell Targetability, Sci. Rep. 8 (2018) 1086. doi:https://doi.org/10.1038/s41598-01819373-3. [115] M.K. Kumawat, M. Thakur, R.B. Gurung, R. Srivastava, Graphene quantum dots for cell proliferation, nucleus imaging, and photoluminescent sensing applications, Sci. Rep. 7 (2017) 15858. doi:https://doi.org/10.1038/s41598-017-16025-w. [116] J. Zhang, X. Zhao, M. Xian, C. Dong, S. Shuang, Folic acid-conjugated green luminescent carbon dots as a nanoprobe for identifying folate receptor-positive cancer cells, Talanta. 183 (2018) 39–47. doi:https://doi.org/10.1016/j.talanta.2018.02.009. [117] Z. Wang, B. Fu, S. Zou, B. Duan, C. Chang, B. Yang, X. Zhou, L. Zhang, Facile construction of carbon dots via acid catalytic hydrothermal method and their application for target imaging of cancer cells, Nano Res. 9 (2016) 214–223. doi:https://doi.org/10.1007/s12274-016-0992-2. [118] N. Ma, W. Jiang, T. Li, Z. Zhang, H. Qi, M. Yang, Fluorescence aggregation assay for the protein biomarker mucin 1 using carbon dot-labeled antibodies and aptamers, Microchim. Acta. 182 (2015) 443–447. doi:https://doi.org/10.1007/s00604-014-1386-3. [119] B. Demir, M. Lemberger, M. Panagiotopoulou, P. Medina-Rangel, S. Timur, T. Hirsch, B. Tse Sum Bui, J. Wegener, K. Haupt, Tracking Hyaluronan: Molecularly Imprinted Polymer Coated Carbon Dots for Cancer Cell Targeting and Imaging, ACS Appl. Mater. Interfaces. (2018). doi:https://doi.org/10.1021/acsami.7b16225. [120] M. Tan, X. Li, H. Wu, B. Wang, J. Wu, N-doped carbon dots derived from bovine serum albumin and formic acid with one-and two-photon fluorescence for live cell nuclear imaging, Colloids Surfaces B Biointerfaces. 136 (2015) 141–149. doi:doi.org/10.1016/j.colsurfb.2015.09.008. [121] Y. Song, X. Li, S. Cong, H. Zhao, M. Tan, Nuclear-targeted of TAT peptide-conjugated carbon dots for both one-and two-photon fluorescence imaging, Colloids Surfaces B Biointerfaces. 180 (2019) 449–456. doi:doi.org/10.1016/j.colsurfb.2019.05.015.

32

[122] M.S. Muthu, D.T. Leong, L. Mei, S.-S. Feng, Nanotheranostics˗ application and further development of nanomedicine strategies for advanced theranostics, Theranostics. 4 (2014) 660. doi:10.7150/thno.8698. [123] J. Ge, Q. Jia, W. Liu, M. Lan, B. Zhou, L. Guo, H. Zhou, H. Zhang, Y. Wang, Y. Gu, Carbon Dots with Intrinsic Theranostic Properties for Bioimaging, Red‐Light‐Triggered Photodynamic/Photothermal Simultaneous Therapy In Vitro and In Vivo, Adv. Healthc. Mater. 5 (2016) 665–675. doi:https://doi.org/10.1002/adhm.201500720. [124] X. Bai, S. Wang, S. Xu, L. Wang, Luminescent nanocarriers for simultaneous drug or gene delivery and imaging tracking, TrAC Trends Anal. Chem. 73 (2015) 54–63. doi:https://doi.org/10.1016/j.trac.2015.04.027. [125] D. Li, Y. Fan, M. Shen, I. Bányai, X. Shi, Design of dual drug-loaded dendrimer/carbon dot nanohybrids for fluorescence imaging and enhanced chemotherapy of cancer cells, J. Mater. Chem. B. 7 (2019) 277–285. doi:10.1039/C8TB02723D. [126] S.M. Janib, A.S. Moses, J.A. MacKay, Imaging and drug delivery using theranostic nanoparticles, Adv. Drug Deliv. Rev. 62 (2010) 1052–1063. doi:https://doi.org/10.1016/j.addr.2010.08.004. [127] D.M.J. Lambregts, M. Maas, M.P.M. Stokkel, R.G.H. Beets-Tan, Magnetic resonance imaging and other imaging modalities in diagnostic and tumor response evaluation, in: Semin. Radiat. Oncol., Elsevier, 2016: pp. 193–198. doi:https://doi.org/10.1016/j.semradonc.2016.02.001. [128] S. Kunjachan, J. Ehling, G. Storm, F. Kiessling, T. Lammers, Noninvasive imaging of nanomedicines and nanotheranostics: principles, progress, and prospects, Chem. Rev. 115 (2015) 10907–10937. doi:https://doi.org/10.1021/cr500314d. [129] A.A. Jamali, M. Pourhassan-Moghaddam, J.E.N. Dolatabadi, Y. Omidi, Nanomaterials on the road to microRNA detection with optical and electrochemical nanobiosensors, TrAC Trends Anal. Chem. 55 (2014) 24–42. doi:https://doi.org/10.1016/j.trac.2013.10.008. [130] H. Kobayashi, M. Ogawa, R. Alford, P.L. Choyke, Y. Urano, New strategies for fluorescent probe design in medical diagnostic imaging, Chem. Rev. 110 (2009) 2620– 33

2640. doi:https://doi.org/10.1021/cr900263j. [131] V. Bagalkot, L. Zhang, E. Levy-Nissenbaum, S. Jon, P.W. Kantoff, R. Langer, O.C. Farokhzad, Quantum dot− aptamer conjugates for synchronous cancer imaging, therapy, and sensing of drug delivery based on bi-fluorescence resonance energy transfer, Nano Lett. 7 (2007) 3065–3070. doi:https://doi.org/10.1021/nl071546n. [132] Q. Zeng, D. Shao, X. He, Z. Ren, W. Ji, C. Shan, S. Qu, J. Li, L. Chen, Q. Li, Carbon dots as a trackable drug delivery carrier for localized cancer therapy in vivo, J. Mater. Chem. B. 4 (2016) 5119–5126. doi:10.1039/C6TB01259K. [133] S.L. D’souza, B. Deshmukh, J.R. Bhamore, K.A. Rawat, N. Lenka, S.K. Kailasa, Synthesis of fluorescent nitrogen-doped carbon dots from dried shrimps for cell imaging and boldine drug delivery system, Rsc Adv. 6 (2016) 12169–12179. doi:10.1039/C5RA24621K. [134] V.N. Mehta, S.S. Chettiar, J.R. Bhamore, S.K. Kailasa, R.M. Patel, Green synthetic approach for synthesis of fluorescent carbon dots for lisinopril drug delivery system and their confirmations in the cells, J. Fluoresc. 27 (2017) 111–124. doi:https://doi.org/10.1007/s10895-016-1939-4. [135] L. Xiao, H. Sun, Novel properties and applications of carbon nanodots, Nanoscale Horizons. 3 (2018) 565–597. doi:10.1039/C8NH00106E. [136] N. Kaur, V. Sharma, P. Tiwari, A.K. Saini, S.M. Mobin, “Vigna radiata” based green Cdots: Photo-triggered theranostics, fluorescent sensor for extracellular and intracellular iron (III) and multicolor live cell imaging probe, Sensors Actuators B Chem. 291 (2019) 275–286. doi:https://doi.org/10.1016/j.snb.2019.04.039. [137] L. Zhang, Y. Wang, W. Liu, Y. Ni, Q. Hou, Corncob residues as carbon quantum dots sources and their application in detection of metal ions, Ind. Crops Prod. 133 (2019) 18– 25. doi:https://doi.org/10.1016/j.indcrop.2019.03.019. [138] V. Roshni, S. Misra, M.K. Santra, D. Ottoor, One pot green synthesis of C-dots from groundnuts and its application as Cr (VI) sensor and in vitro bioimaging agent, J. Photochem. Photobiol. A Chem. 373 (2019) 28–36. 34

doi:https://doi.org/10.1016/j.jphotochem.2018.12.028. [139] B. Wang, S. Wang, Y. Wang, Y. Lv, H. Wu, X. Ma, M. Tan, Highly fluorescent carbon dots for visible sensing of doxorubicin release based on efficient nanosurface energy transfer, Biotechnol. Lett. 38 (2016) 191–201. doi:10.1007/s10529-015-1965-3. [140] M. Thakur, A. Mewada, S. Pandey, M. Bhori, K. Singh, M. Sharon, M. Sharon, Milkderived multi-fluorescent graphene quantum dot-based cancer theranostic system, Mater. Sci. Eng. C. 67 (2016) 468–477. doi:https://doi.org/10.1016/j.msec.2016.05.007. [141] S. Bayda, M. Hadla, S. Palazzolo, V. Kumar, I. Caligiuri, E. Ambrosi, E. Pontoglio, M. Agostini, T. Tuccinardi, A. Benedetti, Bottom-up synthesis of carbon nanoparticles with higher doxorubicin efficacy, J. Control. Release. 248 (2017) 144–152. doi:https://doi.org/10.1016/j.jconrel.2017.01.022. [142] M. Zhang, P. Yuan, N. Zhou, Y. Su, M. Shao, C. Chi, pH-Sensitive N-doped carbon dots– heparin and doxorubicin drug delivery system: preparation and anticancer research, Rsc Adv. 7 (2017) 9347–9356. doi:10.1039/C6RA28345D. [143] S. Majumdar, G. Krishnatreya, N. Gogoi, D. Thakur, D. Chowdhury, Carbon-Dot-Coated Alginate Beads as a Smart Stimuli-Responsive Drug Delivery System, ACS Appl. Mater. Interfaces. 8 (2016) 34179–34184. doi:https://doi.org/10.1021/acsami.6b10914. [144] S. Rai, B.K. Singh, P. Bhartiya, A. Singh, H. Kumar, P.K. Dutta, G.K. Mehrotra, Lignin derived reduced fluorescence carbon dots with theranostic approaches: Nano-drug-carrier and bioimaging, J. Lumin. 190 (2017) 492–503. doi:https://doi.org/10.1016/j.jlumin.2017.06.008. [145] S.L. D’souza, B. Deshmukh, K.A. Rawat, J.R. Bhamore, N. Lenka, S.K. Kailasa, Fluorescent carbon dots derived from vancomycin for flutamide drug delivery and cell imaging, New J. Chem. 40 (2016) 7075–7083. doi:10.1039/C6NJ00358C. [146] S. Singh, A. Mishra, R. Kumari, K.K. Sinha, M.K. Singh, P. Das, Carbon dots assisted formation of DNA hydrogel for sustained release of drug, Carbon N. Y. 114 (2017) 169– 176. doi:https://doi.org/10.1016/j.carbon.2016.12.020. [147] F. Zhang, M. Zhang, X. Zheng, S. Tao, Z. Zhang, M. Sun, Y. Song, J. Zhang, D. Shao, K. 35

He, Berberine-based carbon dots for selective and safe cancer theranostics, Rsc Adv. 8 (2018) 1168–1173. doi:10.1039/C7RA12069A. [148] S. Ghosh, K. Ghosal, S.A. Mohammad, K. Sarkar, Dendrimer functionalized carbon quantum dot for selective detection of breast cancer and gene therapy, Chem. Eng. J. 373 (2019) 468–484. doi:https://doi.org/10.1016/j.cej.2019.05.023. [149] X. Ke, V.W.L. Ng, R.J. Ono, J.M.W. Chan, S. Krishnamurthy, Y. Wang, J.L. Hedrick, Y.Y. Yang, Role of non-covalent and covalent interactions in cargo loading capacity and stability of polymeric micelles, J. Control. Release. 193 (2014) 9–26. doi:https://doi.org/10.1016/j.jconrel.2014.06.061. [150] Z.L. Wu, P. Zhang, M.X. Gao, C.F. Liu, W. Wang, F. Leng, C.Z. Huang, One-pot hydrothermal synthesis of highly luminescent nitrogen-doped amphoteric carbon dots for bioimaging from Bombyx mori silk–natural proteins, J. Mater. Chem. B. 1 (2013) 2868– 2873. doi:10.1039/C3TB20418A. [151] A. Tyagi, K.M. Tripathi, N. Singh, S. Choudhary, R.K. Gupta, Green synthesis of carbon quantum dots from lemon peel waste: applications in sensing and photocatalysis, RSC Adv. 6 (2016) 72423–72432. doi:10.1039/C6RA10488F. [152] R. Bandi, B.R. Gangapuram, R. Dadigala, R. Eslavath, S.S. Singh, V. Guttena, Facile and green synthesis of fluorescent carbon dots from onion waste and their potential applications as sensor and multicolour imaging agents, RSC Adv. 6 (2016) 28633–28639. doi:10.1039/C6RA01669C. [153] C. Zhu, J. Zhai, S. Dong, Bifunctional fluorescent carbon nanodots: green synthesis via soy milk and application as metal-free electrocatalysts for oxygen reduction, Chem. Commun. 48 (2012) 9367–9369. doi:10.1039/C2CC33844K. [154] B. Kumari, R. Kumari, P. Das, Visual detection of G-quadruplex with mushroom derived highly fluorescent carbon quantum dots, J Pharm Biomed Anal. 157 (2018) 137–144. doi:10.1016/j.jpba.2018.05.013. [155] Q. Zhuang, P. Guo, S. Zheng, Q. Lin, Y. Lin, Y. Wang, Y. Ni, Green synthesis of luminescent graphitic carbon nitride quantum dots from human urine and its bioimaging 36

application, Talanta. 188 (2018) 35–40. doi:10.1016/j.talanta.2018.05.060. [156] M. Shahshahanipour, B. Rezaei, A.A. Ensafi, Z. Etemadifar, An ancient plant for the synthesis of a novel carbon dot and its applications as an antibacterial agent and probe for sensing of an anti-cancer drug, Mater Sci Eng C Mater Biol Appl. 98 (2019) 826–833. doi:10.1016/j.msec.2019.01.041. [157] N. Pourreza, M. Ghomi, Green synthesized carbon quantum dots from Prosopis juliflora leaves as a dual off-on fluorescence probe for sensing mercury (II) and chemet drug, Mater Sci Eng C Mater Biol Appl. 98 (2019) 887–896. doi:10.1016/j.msec.2018.12.141. [158] R. Shariati, B. Rezaei, H.R. Jamei, A.A. Ensafi, Application of coated green source carbon dots with silica molecularly imprinted polymers as a fluorescence probe for selective and sensitive determination of phenobarbital, Talanta. 194 (2019) 143–149. doi:10.1016/j.talanta.2018.09.069. [159] F. Ansari, D. Kahrizi, Hydrothermal synthesis of highly fluorescent and non-toxic carbon dots using Stevia rebaudiana Bertoni, Cell Mol Biol. 64 (2018) 32–36. [160] B. Rooj, A. Dutta, S. Islam, U. Mandal, Green Synthesized Carbon Quantum Dots from Polianthes tuberose L. Petals for Copper (II) and Iron (II) Detection, J Fluoresc. 28 (2018) 1261–1267. doi:10.1007/s10895-018-2292-6. [161] T. Pal, S. Mohiyuddin, G. Packirisamy, Facile and Green Synthesis of Multicolor Fluorescence Carbon Dots from Curcumin: In Vitro and in Vivo Bioimaging and Other Applications, ACS Omega. 3 (2018) 831–843. doi:https://doi.org/10.1021/acsomega.7b01323. [162] S. Ahmadian-Fard-Fini, M. Salavati-Niasari, D. Ghanbari, Hydrothermal green synthesis of magnetic Fe3O4-carbon dots by lemon and grape fruit extracts and as a photoluminescence sensor for detecting of E. coli bacteria, Spectrochim Acta A Mol Biomol Spectrosc. 203 (2018) 481–493. doi:10.1016/j.saa.2018.06.021. [163] N. Vasimalai, V. Vilas-Boas, J. Gallo, M. de Fátima Cerqueira, M. Menéndez-Miranda, J.M. Costa-Fernández, L. Diéguez, B. Espiña, M.T. Fernández-Argüelles, Green synthesis of fluorescent carbon dots from spices for in vitro imaging and tumour cell growth 37

inhibition, Beilstein J. Nanotechnol. 9 (2018) 530. doi:10.3762/bjnano.9.51. [164] X. Zhao, S. Liao, L. Wang, Q. Liu, X. Chen, Facile green and one-pot synthesis of purple perilla derived carbon quantum dot as a fluorescent sensor for silver ion, Talanta. 201 (2019) 1–8. doi:10.1016/j.talanta.2019.03.095. [165] Y. Zu, J. Bi, H. Yan, H. Wang, Y. Song, B.W. Zhu, M. Tan, Nanostructures derived from starch and chitosan for fluorescence bio-imaging, Nanomaterials. 6 (2016) 1–13. doi:10.3390/nano6070130. [166] X. Teng, C. Ma, C. Ge, M. Yan, J. Yang, Y. Zhang, P.C. Morais, H. Bi, Green synthesis of nitrogen-doped carbon dots from konjac flour with “off–on” fluorescence by Fe 3+ and L-lysine for bioimaging, J. Mater. Chem. B. 2 (2014) 4631–4639. doi:10.1039/C4TB00368C. [167] H. Wang, Y. Xie, X. Na, J. Bi, S. Liu, L. Zhang, M. Tan, Fluorescent carbon dots in baked lamb: Formation, cytotoxicity and scavenging capability to free radicals, Food Chem. 286 (2019) 405–412. doi:10.1016/j.foodchem.2019.02.034.

38

Figure captions Figure 1. Folic acid-derived CDs for concurrent targeting and imaging of FR positive cancer cells (Reprinted with permission from ref [114]. Figure 2. Folate-conjugated CDs serving as effective probe for FR-positive cancer cell detection. Schematic illustration of preparation of dry yeast-derived CDs and folate-CDs (a), HepG2 cells after incubation in the presence of free-folate (b), Folate-CDs (c); Reproduced with permission from ref [116]. Figure 3. Milk-derived CDs for nucleus-targeted DOX delivery. (a) Optical images of the CD solution under natural and ultraviolet light (365 nm) are shown. (b) Confocal fluorescence images of ACC-2 cells that had been treated with milk-derived CDs, free DOX, and CD-DOX. CDs were predominantly located in the cytoplasm with a smaller quantity visible in the nucleus. Free DOX was located both in the nucleus and cytoplasm but was mainly concentrated at the nuclear membranes. Notably, the CD-DOX complexes showed distinct nuclear localization, which was in contrast to the CDs and free DOX. Adapted with permission from ref [50]. Figure 4. Fluorescent imaging and anti-tumor effect of Ber-CDs in HepG2 tumor-bearing mice. (a) Ex vivo fluorescent imaging of Ber-CDs in different organs of HepG2 tumor-bearing mice. (b) Biodistributions of Ber–CDs based on the fluorescence intensity of organ and serum. (c) tumor picture and (d) Tumor weight from mice after administration of Ber or Ber–CDs. (e) IC50 values of Ber or Ber-CDs on cancer cell lines (HepG2, MCF-7, A549, SMC-7721, and H22) and normal cell lines (HL-7702, HUVEC, and C2C12) to evaluate anti-tumor effect of Ber–CDs. Reproduced with permission from ref [147].

39

Table 1. A comparison between green and chemical carbon dots: the synthesis condition and toxicity.

Green synthesis

Chemical synthesis

Use of green (naturally occurring), eco-friendly, safe precursors as a source of carbon for CD synthesis via biological techniques

Mainly use of inorganic and metallic precursors, stabilizing agents and reducing agents for CD synthesis

Environmental friendly, use of non-toxic and safe reagents

Highly use of strong acids (HNO3/H2SO4) and chemical stabilizing agents

Single-step method, cost-effective

Mostly complex synthesis steps while the necessity for special equipment, costly chemicals

Consume less energy, easy to scale-up

Demanding high energy

Facilitating waste management

Generating risky products/by-products for the environment and human health

Green-CDs

Chemical-CDs

Non-toxic Self-passivated (reducing chemical exposure)

Unwanted reactivity and toxicity, uncertainty of composition Extra passivation step with chemicals

Availability and affordability/renewable

Non-renewable source of synthesis

40

Table 2. Green CDs sources and their synthesis methods: advantages and disadvantages. Green source

Synthesis methods

Advantages

Disadvantages

Apple juice [78], Bagasse [31], bee pollens [12], Bombyx mori silk [150], Cabbage [79], Carica papaya juice [80], Dried shrimp[133], Garlic [81], Ginger [82], Grape peel[83], Grape juice, Grass [85], Honey [22], Jinua bergamot [74], Lemon juice[75], Lemon peel [151], Milk [18], Orange juice [16], Orange waste peel [76], Onion waste [152], Pomelo peels [15], Soy milk [153], Strawberry juice [70], Sugar cane juice [31], Sweet potato [77], wheat straw/ bamboo [71], mushroom [154], human urine [155], (Henna)plant [156], Prosopis juliflora leaves [157], cedrus [158], Steviarebaudiana Bertoni [159], tuberose petals [160], Curcumin [161], (grape fruit- lemon-turmeric) extracts [162], spices [163], bovine serum albumin/formic acid [120], tryptophan (Trp)/formic acid [121], chitosan [86,106], Purple perilla [164]

Hydrothermal treatment

• • • •

Cost-effective eco-friendly non-toxic moderate-pressure method of synthesis convenient operation, simple synthesis process large-scale synthesis no need for special equipment energy efficient

slow kinetics of crystallization

Egg shell membrane [67], Flour[68], Garlic [65], Onion peel [66], Chitosan/starch [165]

Microwave heating

• • • •

Rapid and scalable cost-effective eco-friendly directly transfer heat to the reactants (in contrast to the conventional methods)

Poor control over size

Konjac flour [166], Plant leaf [32]

Pyrolysis

producing smaller size of CDs

necessity for high temperature and energy

Chicken egg [21]

Plasma irradiation

• Rapid • creating amphiphilicity CDs

CD pattern expanding need to other QDs

Muraenesox cinereus (fish) [88] roasted Atlantic salmon [28], pizza [27] chicken [87], Chicken breast [29], Lamb [93], Duck [91], Grilled burger [92] Baked lamb [93,167]

Thermal processing (roasting)

Ease of synthesis during food/beverages preparation

Uncertainty of CD composition

Commercial beverages [96] Commercial beer [90] Chinese mature vinegar [94] Coca-cola [89]

Extraction

Ease of synthesis during food/beverages preparation

Uncertainty of CD composition

• • • •

41

Table 3. Self and actively targeted imaging via green-CDs.

Precursor (green/bio)

QY(%)

Imaging target

Imaging model

Targeting ligand

Application

Ref.

Folic acid

94.50

Cancer cell membrane

A549 (C-) HeLa (C+)

Folate residue

FR-mediated cancer cell targeting

[114]

Folic acid

9

Cancer cell membrane

HeLa, SKOV3, HepG2, MCF7, CHO and A549 cell

Folate residue

Imaging cancer cell expressing the folate receptor

[113]

Grape seed extract

31.79

Nucleus

L929,HT-1080 , MIA PaCa-2 , HeLa, MG-63 cells

_

Selective nucleus labelling and targeting in theranostics

[115]

Dandelion

13.90

Cell membrane/ cytoplasm

MCF7 , HepG2(FR+) PC12 cell (FR- )

Folic acid

For recognizing folate receptor positive cancer cells

[44]

Active dry yeast

11.60

Cell membrane/ cytoplasm

HepG2 and PC12 cell line

Folic acid

Folate receptor positive cancer cells targeting and imaging

[116]

D-glucose L-aspartic acid

_

Intracellular

C6 glioma cell

_

Self- targeted imaging cancer cells

[13]

Tryptophan and Phenylalanine/ Glucose

21

Cell membrane/ cytoplasm

MUC-1 overexpressed MCF-7

MUC1aptamer/ MUC1 antibody

For cancer cell targeting and imaging

Hyaluronan (a biomarker for certain cancers)

HeLa cell (+) HaCaT cell (-)

MIP shell for GlcA

For cancer cell targeting and imaging

[119]

Starch

25.1 ± 2

[117,118]

BSA + formic acid

17.1

Nucleus

tongue squamous cancer cells

TATpeptide

for live cell nuclear-targeted imaging

[120]

Trp + formic acid

58.4

Nucleus

Mouse melanoma B16-F10 cancer cells

TATpeptide

Nuclear-targeting cellular imaging

[121]

42

Table 4. Representative drug/gene delivery systems based on green-CDs. Precursor

Drug delivery

Loading efficiency

CDs-conjugation

Cow milk

Doxorubicin

87%

Electrostatic Hydrogen bond

Cow milk

BHC (berberine hydrochloride)

88%

Covalent

Dried shrimps

Boldine (herbal alkaloid)

_

Black tea

Doxorubicin

Chitosan

Drug release factor PH

In vitro

In vivo

Application

Ref.

ACC-2, L929

_

Apoptosis induced by free DOX and CDDOX complex

[50]

PH

L929, HeLa, MDA-MB231

_

Traceable targeted drug delivery to cancer cells

[140]

Non-covalent

PH

MCF7

_

Evaluating MCF7 death by free boldine in comparison to CDboldine

[133]

60%

Electrostatic

PH

MDA-MB231, DLD1

Higher DOX efficacy in vitro and in vivo

[141]

Heparin Doxorubicin

28.45% 32.2%

Chemical bonds Electrostatic

PH

HeLa, MCF-7 and A549

Nude mice model of breast cancer Spider plant

To deliver Dox with higher efficiency

[142]

Chitosan

Allicin

77.98%

Electrostatic

amount of MRSA pathogen

Lignin

Curcumin

67.40%

Hydrophobic interaction

Beer

Doxorubicin

_

E.coli genomic DNA

Rhodamine 6G/ doxorubicin

Vancomycin

MRSA strains

_

Increased drug encapsulation

[143]

PH

A549 and SW480

_

As nano-drug carrier and bioimaging agent

[144]

Electrostatic

PH

MCF7

_

[19]

_

Electrostatic interactions

PH

_

Flutamide

_

Covalent H-bond

Simultaneous imaging and drug delivery

[145]

Berberis plant

_

_

_

pH / Hixson– Crowell model _

S. cerevisiae, E. coli, HEK 293 cells MCF-7, SHSY5Y cells

For cell imaging and anticancer drug delivery to MCF-7 cells DNA-CD as bioimaging and drug delivery platform

Self-theranostic application of green CDs

[147]

sweet lemon peel

p-DNA

_

Electrostatic/steric interaction

promising theranostic tool for TNBC in future

[148]

_

43

MDA-MB231 cell

_

tumor site of HepG 2 tumorbearing mice _

[38]

Highlights

•

The role of fluorescent green-CDs to guide therapy and monitoring the therapeutic response (Therapeutic).

•

Image-guided active and self-targeting of cancer cells via green-CDs (Diagnostic).

•

Future prospects of the green-CD application as a platform to integrate diagnostics and therapeutics (Theranostic).

Declaration of interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.