Magnetofluorescent Carbon Dots Derived from Crab Shell for Targeted Dual-Modality Bioimaging and Drug Delivery.

We propose a one-pot microwave-assisted pyrolysis method for fabrication of magnetofluorescent carbon quantum dots (MFCQDs), using a combination of waste crab shell and three different transition-metal ions, Gd3+, Mn2+, and Eu3+, referred to as Gd@CQDs, Mn@CQDs, and Eu@CQDs, respectively. Chitin from waste crab shell acted not only as a carbon source but also as a chelating ligand to form complexes with transition-metal ions. Gd@CQDs exhibited a high r1 relaxivity of 4.78 mM-1·s-1 and a low r2/r1 ratio of 1.33, suggesting that they show excellent potential as a T1 contrast agent. Mn@CQDs and Eu@CQDs showed high r2 relaxivity values of 140.7 and 28.32 mM-1·s-1, respectively, suggesting their potential for use as T2 contrast agents. Further conjugation of Gd@CQDs with folic acid (FA) enabled specific targeting to folate receptor-positive HeLa cells, as confirmed via in vitro magnetic resonance and fluorescence imaging. Doxorubicin (DOX) was selected as a model drug for conjugation with FA-Gd@CQDs. The as-prepared nanocomposites showed significantly higher cytotoxicity toward HeLa cells than free DOX. No apparent cytotoxicity was observed in vivo (zebrafish embryos) or in vitro (cell viability), suggesting that MFCQDs show potential for development as diagnostic probes or theranostic agents.

Facile synthesis of gold/gadolinium-doped carbon quantum dot nanocomposites for magnetic resonance imaging and photothermal ablation therapy.

Composites of gold nanomaterials and imaging agents show promise in cancer therapy. Here we have demonstrated a rapid, facile, environmentally friendly, and organic solvent-free method for the synthesis of a gold/gadolinium-doped carbon quantum dot (Au/GdC) nanocomposite for magnetic resonance imaging (MRI) and photothermal ablation (PTA) therapy. The gadolinium-doped carbon quantum dots (Gd@CQDs) were synthesized using a one-pot, microwave-assisted method, and used as reducing and stabilizing agents to both form the Au/GdC nanocomposite and prevent its agglomeration. Formation of the Au/GdC nanocomposite is achieved by simple mixing of Gd@CQDs and a gold precursor, without the addition of any other reducing agents, surface passivating agents, surfactants, or organic solvents. The Au/GdC nanocomposite shows paramagnetism, surface plasma resonance in the near infrared region (NIR), and excellent photostability. Furthermore, it provides high longitudinal relaxivity (r1 = 13.95 mM-1 s-1), indicating its potential for use as a T1 contrast agent in MRI. Furthermore, in vitro and in vivo studies using HeLa cells and zebrafish embryos as cancer and animal cell models, respectively, confirmed the low toxicity and excellent biocompatibility of the Au/GdC nanocomposite. Notably, our results demonstrate the ability of the Au/GdC nanocomposite to efficiently destroy cancer cells using PTA. Therefore, this work reveals a simple and powerful strategy to fabricate an Au/GdC nanocomposite for MRI and photothermal ablation of cancer cells.

Structural simulation and protein engineering to convert an endo-chitosanase to an exo-chitosanase.

To obtain an enzyme for the production of chito-disaccharides (GlcN(2)) by converting endo-chitosanase to exo-chitosanase, we chose an endo-chitosanase from Bacillus circulans MH-K1 (Csn) as the candidate for protein engineering. Using molecular modeling, two peptides with five amino acids (PCLGG) and six amino acids (SRTCKP) were designed and inserted after the positions of D(115) and T(222) of Csn, respectively. The inserted fragments are expected to form loops that might protrude from opposite walls of the substrate-binding cleft, thus forming a 'roof' over the catalytic site that might alter the product specificity. The chimeric chitosanase (Chim-Csn) and wild-type chitosanase (WT-Csn) were both over-expressed in Escherichia coli and purified nearly to homogeneity. The products formed from chitosan were analyzed by ESI-MS (electrospray ionization-mass spectrometry). A mixture of GlcN(2), GlcN(3) and GlcN(4) was obtained with WT-Csn, whereas Chim-Csn formed, with a smaller catalytic rate (3% of WT-Csn activity), GlcN(2) as the dominant product. Measurements of viscosity showed that, with similar amounts of enzyme activity, Chim-Csn catalyzed the hydrolysis of chitosan with a smaller rate of viscosity decrease than WT-Csn. The results indicate that, on inserting two surface loops, the endo-type chitosanase was converted into an exo-type chitosanase, which to our knowledge is the first chitosanase that releases GlcN(2) from chitosan as the dominant product.