NANOCOMPOSITES OF NITROGEN-DOPED GRAPHENE OXIDE AND MANGANESE OXIDE FOR PHOTODYNAMIC THERAPY AND MAGNETIC RESONANCE IMAGING

Abstract

The present invention relates to a NDG-Mn.sub.3O.sub.4 nanocomposite comprising a nitrogen doped graphene (NDG) and Mn.sub.3O.sub.4 nanoparticles. The NDG-Mn.sub.3O.sub.4 nanocomposite is useful in bimodal performance including photodynamic therapy (PDT) and magnetic resonance imaging (MRI). The NDG-Mn.sub.3O.sub.4 nanocomposites of the present invention caused significant cell death under laser irradiation, while control and Mn.sub.3O.sub.4 nanoparticles showed negligible cell death.

Claims

1. A nanocomposite comprising a nitrogen doped graphene oxide conjugated with Mn.sub.3O.sub.4 nanoparticle.

2. The nanocomposite as claimed in claim 1, wherein particle size of the nanocomposite is in a range of 5 nm to 15 nm.

3. The nanocomposite as claimed in claim 1, wherein particle size of the nanocomposite is 10±1.7 nm.

4. The nanocomposite as claimed in claim 1, wherein the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticle is present in the nanocomposite in a ratio of 1:1.

5. The nanocomposite as claimed in claim 1, wherein the nanocomposite is obtained using milling process.

6. A process of preparation of a nanocomposite as claimed in claim 1, wherein the process comprises the steps of: (a) reacting manganese (II) acetylacetonate with oleylamine at a temperature in a range of 150° C. to 170° C. for a period of 8 to 12 hours to obtain Mn.sub.3O.sub.4 nanoparticles; (b) reacting a suspension of graphene oxide (GO) with hydrazine hydrate in presence of a base at a temperature range of 85° C. to 95° C. for a period of 1 to 4 hours to obtain nitrogen doped graphene oxide; (c) milling the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticles for a time period in a range of 14 hours to 17 hours to obtain a nanocomposite comprising the nitrogen doped graphene oxide conjugated with the Mn.sub.3O.sub.4 nanoparticle.

7. The process as claimed in claim 6, wherein manganese (II) acetylacetonate and oleylamine is present in a molar ratio of 1:25.

8. The process as claimed in claim 6, wherein the base is ammonium hydroxide or potassium hydroxide.

9. The process as claimed in claim 6, wherein the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticles is in 1:1 ratio.

10. The process as claimed in claim 6, wherein the temperature in step (a) is 160° C. and in step (b) is 90° C.

11. A method of treating a cancer or imaging targeted tissue in a subject, comprising: (a) administering to the subject in need thereof an effective amount of a nanocomposite comprising nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticles; and (b) exposing cancer cell or tissue of the subject to laser irradiation in presence of fluorescein diacetate (FDA) and propidium iodide (PI) for a sufficient amount of time to obtain a desired response.

12. The method as claimed in claim 11, wherein the cancer is breast cancer, prostate cancer, brain cancer, colorectal cancer, pancreatic cancer, ovarian cancer, lung cancer, cervical cancer, liver cancer, head/neck/throat cancer, skin cancer, bladder cancer and a hematologic cancer.

13. The method as claimed in claim 11, wherein the cancer is lung cancer.

14. The method cancer as claimed in claim 11, wherein the imaging is Magnetic Resonance Imaging (MRI).

Description

BRIEF DESCRIPTION OF THE FIGURES

[0030] FIG. 1: High-resolution transmission electron microscopy (HRTEM) images of the NDG-Mn.sub.3O.sub.4 nanocomposite (a) low magnification image, (b) magnified image, (c) energy-dispersive X-ray spectroscopy of NDG-Mn.sub.3O.sub.4 nanocomposite and (d) particle size distribution of NDG-Mn3O4 nanocomposite.

[0031] FIG. 2: XRD pattern of (a) Mn.sub.3O.sub.4 NPs, (b) NDG and (c) NDG-Mn.sub.3O.sub.4 nanocomposite.

[0032] FIG. 3: FT-IR spectra of (a) Mn.sub.3O.sub.4 NPs, (b) NDG and (c) NDG-Mn.sub.3O.sub.4 nanocomposite.

[0033] FIG. 4: Cytotoxicity analysis showing cell viability of A549 cells treated with different concentrations of Mn.sub.3O.sub.4 and NDG—Mn.sub.3O.sub.4 nanocomposites. Values are means of three replicates ± standard error.

[0034] FIG. 5: Cell viability of A549 cells incubated with PBS (control), Triton X-100 (negative control) and different concentrations of, Mn.sub.3O.sub.4 and NDG-Mn.sub.3O.sub.4 nanocomposites in presence of 670 nm laser irradiation (0.1 W/cm.sup.2) for 5 min. Data is represented as mean values of three replicates (±) standard deviations. *p< 0.05, **p< 0.01 and ***p< 0.001 versus respective control groups.

[0035] FIG. 6: Fluorescence microscopy of A549 cells stained with fluorescein diacetate (green emission for live cells) and propidium iodide (red emission for dead cells) with PBS (control), Mn.sub.3O.sub.4 and NDG—Mn.sub.3O.sub.4 nanocomposites with/without laser irradiation (670 nm, 0.1 W/cm.sup.2) for 5 min.

[0036] FIG. 7: Effect of SNWE treatment on apoptotic markers in the MCF-7 and MDA-MB-231 cells. The protein expressions was analysed by immunofluorescence method, the A) BAX, B) Caspase3, and C) p53, expression were increased in the SNWE treatment which shows the induction of caspase dependent apoptosis in breast cancer cells. The cells were immunostained with anti p53, BAX, Caspase3 antibodies and FITC labelled secondary antibodies. DAPI was used as counter stain for nucleus and the images were acquired with fluorescence microscope.

[0037] FIG. 8: T.sub.1-weighted MR imaging of NDG-Mn.sub.3O.sub.4 nanoparticles in aqueous suspension and the T1 relaxivity plot of aqueous suspension of NDG-Mn.sub.3O.sub.4 nanoparticles. The concentration range of 0.06-1.0 mM of Mn is equivalent to approximately 18-152 .Math.g/mL of NDG-Mn.sub.3O.sub.4 nanoparticles.

DETAILED DESCRIPTION OF THE INVENTION

[0038] The embodiments herein and the various features and advantageous details thereof are explained more comprehensively with reference to the non-limiting embodiments that are detailed in the following description. Descriptions of well-known components and processing techniques are omitted so as to not unnecessarily obscure the embodiments herein. The examples used herein are intended merely to facilitate an understanding of the ways in which the embodiments herein may be practiced and to further enable those of skill in the art to practice the embodiments herein. Accordingly, the examples should not be construed as limiting the scope of the embodiments herein.

[0039] Unless otherwise specified, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions may be included to better appreciate the teaching of the present invention.

[0040] As used in the description herein, the meaning of “a,” “an,” and “the” includes plural reference unless the context clearly dictates otherwise. Also, as used in the description herein, the meaning of “in” includes “in” and “on” unless the context clearly dictates otherwise.

[0041] As used herein, the terms “comprise”, “comprises”, “comprising”, “include”, “includes”, and “including” are meant to be non- limiting, i.e., other steps and other ingredients which do not affect the end of result can be added. The above terms encompass the terms “consisting of” and “consisting essentially of”.

[0042] The present disclosure relates to a NDG-Mn.sub.3O.sub.4 nanocomposite comprising a nitrogen doped graphene (NDG) and Mn.sub.3O.sub.4 nanoparticles. The NDG-Mn.sub.3O.sub.4 nanocomposite is useful in bimodal performance including photodynamic therapy (PDT) and magnetic resonance imaging (MRI). The NDG-Mn.sub.3O.sub.4 nanocomposites of the present invention caused significant cell death under laser irradiation.

[0043] In an embodiment, the present invention relates to a nanocomposite comprising a nitrogen doped graphene oxide conjugated with Mn.sub.3O.sub.4 nanoparticle.

[0044] In another embodiment of the present invention, particle size of the nanocomposite is in a range of 5 nm to 15 nm. Preferably, the particle size of the nanocomposite is 10±1.7 nm.

[0045] In another embodiment of the present invention, the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticle is present in the nanocomposite in a ratio of 1:1.

[0046] In another embodiment of the present invention, the nanocomposite is obtained using milling process.

[0047] In yet another embodiment, the present invention relates to a process of preparation of a nanocomposite as claimed in claim 1, wherein the process comprises the steps of: [0048] (a) reacting manganese (II) acetylacetonate with oleylamine at a temperature in a range of 150° C. to 170° C. for a period of 8 to 12 hours to obtain Mn.sub.3O.sub.4 nanoparticles; [0049] (b) reacting a suspension of graphene oxide (GO) with hydrazine hydrate in presence of a base at a temperature range of 85° C. to 95° C. for a period of 1 to 4 hours to obtain nitrogen doped graphene oxide; [0050] (c) milling the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticles for a time period in a range of 14 hours to 17 hours to obtain a nanocomposite comprising the nitrogen doped graphene oxide conjugated with the Mn.sub.3O.sub.4 nanoparticle.

[0051] In another embodiment of the present invention, the manganese (II) acetylacetonate and oleylamine used in the process of preparing nanocomposite is present in a molar ratio of 1:25.

[0052] In another embodiment of the present invention, the base used in the process of preparing nanocomposite is ammonium hydroxide or potassium hydroxide.

[0053] In another embodiment of the present invention, the nitrogen doped graphene oxide and the Mn.sub.3O.sub.4 nanoparticles used in the process of preparation of nanocomposite is present in 1:1 ratio.

[0054] In another aspect of the present invention, the temperature in step (a) is 160° C. and in step (b) is 90° C.

[0055] In yet another embodiment, the present invention relates to a method of treating a cancer or imaging targeted tissue in a subject, comprising: [0056] (a) administering to the subject in need thereof an effective amount of a nanocomposite comprising nitrogen doped graphene oxide and the Mn3O4 nanoparticles; and [0057] (b) exposing cancer cell or tissue of the subject to laser irradiation in presence of fluorescein diacetate (FDA) and propidium iodide (PI) for a sufficient amount of time to obtain a desired response.

[0058] In another embodiment of the present invention, the cancer is lung cancer, breast cancer, prostate cancer, brain cancer, colorectal cancer, pancreatic cancer, ovarian cancer, cervical cancer, liver cancer, head/neck/throat cancer, skin cancer, bladder cancer and a hematologic cancer.

[0059] In another aspect of the present invention, the imaging is Magnetic Resonance Imaging (MRI).

[0060] According to the present invention, NDG—Mn.sub.3O.sub.4 nanocomposites did not cause any cytotoxicity unless activated by laser irradiation that resulted in concentration dependent cytotoxicity in lung cancer cells (FIG. 5). These biochemical findings were supported by fluorescence microscopy observations, suggesting that NDG-Mn.sub.3O.sub.4 nanocomposites initiate cytotoxic properties only under laser irradiation (FIG. 6).

[0061] In an embodiment of the present invention, the 670 nm laser was used in order to keep the wavelength within the red optical window (620-750 nm).

[0062] In another embodiment of the present invention, the NDG-Mn.sub.3O.sub.4 nanocomposites killed 68% of cancer cells which is more effective than GQD-PDA-Mn.sub.3O.sub.4 nanoparticles (51% cell death). The mechanism of laser-induced toxicity of NDG—Mn.sub.3O.sub.4 nanocomposites can be multifactorial. The laser irradiation during PDT triggered the disruption of cellular membranes resulting in a higher cellular uptake of the GQD-PDA-Mn.sub.3O.sub.4 nanoparticles compared to graphene quantum dots. This selective transport across the cell membrane might have been influenced by the size, shape and surface chemistry of nanoparticles.

[0063] In another embodiment of the present invention, the NDG-Mn.sub.3O.sub.4 nanocomposites caused .sup.1O.sub.2 generation under laser irradiation in a time-dependent manner and longer exposure to laser irradiation produced excessive ROS generation (FIG. 7).

[0064] In another embodiment of the present invention, the results of MRI demonstrated a concentration dependent enhancement of signal intensity with increasing concentration of NDG-Mn.sub.3O.sub.4 nanocomposites (FIG. 8).

[0065] The present disclosure may be more fully understood by reference to the following examples:

EXAMPLES

Preparation of Mn.SUB.3.O.SUB.4 Nanoparticles

[0066] Manganese (II) acetylacetonate was dissolved in oleylamine (molar ratio of manganese (II) acetylacetonate: oleylamine = 1:25) and the mixture was heated at 160 C for 10 h under a nitrogen cover. The resulting product was cooled to room temperature to form a brownish suspension, which was centrifuged at 9000 rpm for 15 min and the supernatant was removed to obtain a brown residue. The precipitate was washed multiple times with ethanol to acquire pure Mn.sub.3O.sub.4 nanoparticles, which were dried under vacuum before use.

Preparation of Nitrogen-Doped Graphene Oxide (NDG)

[0067] Initially, graphite oxide (GO) was synthesized from graphite powder using a modified Hummers method. Briefly, graphite powder (0.5 g) and NaNO.sub.3 (0.5 g) were added to 23 mL of H.sub.2SO.sub.4 and the mixture was stirred for 10 min in an ice bath. Subsequently, KMnO.sub.4 (3 g) was slowly added and after proper mixing, the ice bath was replaced with water bath (35) for 1 h, resulting in the formation of a thick paste. Thereafter, 40 mL of deionized water was added, and the mixture was stirred for 30 min at 90 C. Finally, 100 mL of deionized water was added, followed by the slow addition of 3 mL of H.sub.2O.sub.2. The mixture was allowed to cool, filtered and washed with deionized water. The resulting thick brown paste was dispersed in water and centrifuged at 1000 rpm for 2 min. This step was repeated 4-5 times, until all unsettled particles were removed. The resultant paste was dispersed in water with mild sonication to obtain a suspension of graphene oxide (GO). For nitrogen doping, the resulting suspension was taken in a round bottom flask, to which 4 mL of NH.sub.4OH and 4 mL hydrazine hydrate were added simultaneously. The mixture was stirred for a few minutes, and the flask (equipped with cooling condenser) was put in a water bath controlled at 90° C. for 3 h. The product was collected after been filtered through micropore filters (Whatman filter paper, pore size-20 .Math.m, W&R Balston Limited, Maidstone, Kent, UK), washed by deionized water and freeze-dried.

Preparation of Nanocomposites of NDG and Mn3O4(NDG—Mn3O4)

[0068] Equal amounts of Mn.sub.3O.sub.4 nanoparticles and NDG were milled using a Fritsch Pulverisette P7 planetary ball mill (Idar-Oberstein, Germany). The nanomaterials powder and stainless steel balls (5 mm diameter) with the ball to powder weight ratio of 1:1 were introduced into the stainless steel container. The milling of the powder was performed for 16 h, with intermittent pausing of milling process at regular intervals.

Characterization of Nanoparticles

[0069] The synthesized nanoparticles were characterized for size and physicochemical properties using high resolution transmission electronmicroscopy (JSM-7610F, JEOL, Tokyo, Japan), X-ray diffraction analysis (D2 Phaser X-ray diffractometer, Bruker, Ettlingen, Germany) and FT-IR spectroscopy (Perkin Elmer 1000 FT-IR spectrometer,Waltham, MA, USA). A microplate reader determined the absorbance at 570 nm (Molecular Devices, USA). Percentage cell viability was calculated, and cell-survival curves were constructed.

[0070] The results of high resolution transmission electron microscopy (HRTEM) displayed the existence of spherical shaped Mn.sub.3O.sub.4 nanoparticles on the surface of NDG within the range of 5-15 nm (FIG. 1). The Mn.sub.3O.sub.4 NPs are well distributed on the surface of NDG as the magnified image indicates the shape and crystallinity of these NPs. The Mn.sub.3O.sub.4 NPs are not bonded covalently but are held by physisorption on the NDG surface by Vander Waals interactions. The elemental composition of NDG-Mn.sub.3O.sub.4 nanocomposite, analyzed by energy-dispersive X-ray spectroscopy, showed intense signals at 0.65, 5.88, and 6.65 keV strongly suggesting that ‘Mn’ was the major element, which has an optical absorption in this range owing to the surface plasmon resonance (SPR). Other signals that were found in the range of 0.0-0.5 keV signified the absorption of carbon, nitrogen and oxygen, confirming the formation of NDG-Mn.sub.3O.sub.4 nanocomposite. The average particle size of the NDG-Mn.sub.3O.sub.4 nanocomposite was found to be 10 ± 1.7 nm (FIG. 1).

[0071] The XRD pattern of Mn.sub.3O.sub.4NPs shown in FIG. 2a exhibits characteristics peaks at 18.2 (101), 29.1 (112), 31.2 (200), 32.5 (103), 36.3 (211), 38.2 (004), 44.6 (220), 50.8 (105), 53.8 (312), 58.7 (321), 60.0 (224), and 64.8 (314), which points to the formation of manganese oxide NPs (FIG. 2a) These peaks reveal that the as-obtained Mn.sub.3O.sub.4NPs exist in single phase hexagonal wurtzite structure, besides, the data clearly matched with the standard Mn.sub.3O.sub.4 phase reported in the literature (JCPDS Card No. 24-0734). Notably, the sharp diffraction peaks point toward the highly crystalline and well-disperse nature of nanoparticles which clearly matched with the Hausmannite crystal phase. On the other hand, the XRD pattern of NDG-Mn.sub.3O.sub.4 nanocomposite showed the appearance of a broad peak at ~22.4 (002) (FIG. 2b) that confirmed the reduction of graphene oxide and formation of NDG. Furthermore, there is no broadening or shift of the (002) peak, proving that there is no change in the interlayer spacing of graphene after nitrogen-doping. No significant change in the full width at half-maximum (FWHM) of the (002) diffraction peak indicates the similar crystallite size before and after nitrogen doping. In case of the composite, the XRD pattern of which is shown in FIG. 2c, characteristic diffraction peaks of both Mn.sub.3O.sub.4 and N-doped graphene are present, which clearly indicate the formation of hybrid material.

[0072] FT-IR spectra of Mn.sub.3O.sub.4 NPs displayed the characteristic peak of Mn-O, stretching mode in the range of 624 cm.sup.-1 while the vibrational frequency associated to the Mn-O distortion vibration poisoned at 525 cm.sup.-1 (FIG. 3a). The characteristic narrow and broad bands located at 3420 and 1600 cm.sup.-1 were related to the hydroxyl (—OH) groups absorbed by the samples or potassium bromide. FT-IR spectra of NDG are shown in FIG. 3b. FT-IR spectra of NDG-Mn.sub.3O.sub.4 displayed the graphene oxide intense bands for C=C stretching (~1630 cm.sup.-1), C—O—C stretching (~1209 cm.sup.-1), C—O stretching (~1050 cm.sup.-1). The nitrogen doping in the sample was confirmed by the presence of two characteristic peaks at ~1325 and ~1570 cm.sup.-1, which were attributed to the stretching of the C—N bond from the secondary aromatic amine, which pointed toward bonding between carbon and nitrogen including the existence of other absorption bands of ‘Mn’ at 624 and 525 cm.sup.-1 clearly indicating the formation of HRG—Mn.sub.3O.sub.4 nanocomposite (FIG. 3c).

Cell Viability Analysis

[0073] The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method was used for testing the cytotoxicity of Mn.sub.3O.sub.4 and NDG—Mn.sub.3O.sub.4 nanoparticles. A549 lung cancer cells were seeded into 96-well plate (4 × 10.sup.4 cells per well) in RPMI and incubated at 37° C. for 4 h in a 5% CO.sub.2 incubator. Different concentrations (6.25-100 .Math.g/mL) of Mn.sub.3O.sub.4 and NDG—Mn.sub.3O.sub.4 nanoparticles were added to the 96-well plate. Phosphate buffer saline (PBS) was used as a control whereas triton-X100 was used as negative control. The cells were treated with a 670 nm laser irradiation at 0.1 W/cm.sup.2 for 5 min and further incubated for 24 h. Aqueous solution of MTT (50 .Math.L) was added to each well in the 96-well plate 4 h before the termination of 24 h incubation. The upper layer of the solution was discarded. The MTT solubilization solution, DMSO (100 .Math.L) was added to each well to dissolve the formazan crystals by pipette stirring and then observed the absorbance at 590 nm, which was converted to cell viability using the following equation. Cell viability (%) = (absorbance of sample cells/absorbance of control cells) × 100

[0074] The results of cell viability analysis using MTT assay showed that exposure of Mn.sub.3O.sub.4 and NDG-Mn.sub.3O.sub.4 in the concentration range of 6.25-100 .Math.g/mL did not cause any cytotoxicity (FIG. 4). However, NDG-Mn.sub.3O.sub.4 nanocomposites displayed significant cells death under laser irradiation for 5 min, while PBS (control) and Mn3O4 NPs showed negligible cell death (FIG. 5). Almost 100% cells were viable when treated with PBS whereas 41% for cancer cells survived after the treatment of 100 .Math.g/mL concentration of NDG-Mn.sub.3O.sub.4 nanocomposites along with 5 min of laser irradiation. The effect of NDG-Mn.sub.3O.sub.4 nanocomposites on the cytotoxicity of A549 cells was concentration-dependent and only the concentrations of 25 .Math.g/mL and above were found to be effective in killing the cells under laser irradiation (FIG. 5).

In-Vitro Photodynamic Therapy

[0075] Fluorescence microscopy was used for morphological analysis of cancer cells following treatment with nanoparticles and laser irradiation. Fluorescein diacetate (FDA) and propidium iodide (PI) were used to visualize the live and dead cells, respectively. A549 cells (2 × 10.sup.4 cells per well) were seeded in a 24 well plate and incubated at 37° C. for 24 h in an atmosphere of 5% CO.sub.2. Mn.sub.3O.sub.4 and NDG-Mn.sub.3O.sub.4 nanoparticles (50 .Math.g/mL) were added to the wells and the plate was incubated for 4 h. After incubation, the cells were irradiated for 5 min with a 670 nm laser, followed by another incubation for 24 h. Both the dyes were added to wells and the plate was incubated for 5 min. Then, the cells were washed three times with PBS to remove excess dyes, and the fluorescence images were acquired by fluorescence microscope with 490 nm excitation and 525 nm emission wavelengths.

[0076] The results of in-vitro photodynamic therapy are shown in FIG. 6. Without laser irradiation, none of the treatments including PBS, Mn.sub.3O.sub.4, or NDG-Mn.sub.3O.sub.4 caused any cellular damage as almost all the cells appeared green. After 5 min laser irradiation, NDG-Mn.sub.3O.sub.4 nanocomposites killed 68% of the cancer cells (shown as red dots) whereas the treatments of PBS and Mn.sub.3O.sub.4 did not cause any significant cellular damage under laser irradiation (FIG. 6).

Analysis of Singlet Oxygen Generation

[0077] 1,3-Diphenylisobenzofuran (DBPF) was used to detect singlet oxygen (.sup.1O.sub.2) generation by NDG-Mn.sub.3O.sub.4 nanocomposites under 670 nm laser irradiation (0.1 W/cm.sup.2). Fifty microliters of ethanolic solution of DPBF (1 mg/mL) were added to the nanocomposites solution under stirring and irradiated with laser for different time points. The absorbance of solution was measured by UV-Visible spectrophotometer. The decrease in absorbance at 426 nm indicated the degradation of DPBF in presence of .sup.1O.sub.2 which was generated by laser-induced activation of NDG-Mn.sub.3O.sub.4 nanocomposites.

[0078] To evaluate the .sup.1O.sub.2 generation from NDG-Mn.sub.3O.sub.4 nanocomposites under laser irradiation, we measured the absorbance of 1,3-diphenylisobenzofuran (DPBF) after laser irradiation (670 nm, 0.1 W/cm.sup.2) at different time points (FIG. 7). The DPBF absorbance decreased with increasing the laser irradiation time, indicating the generation of singlet oxygen from NDG-Mn.sub.3O.sub.4 nanocomposites is directly proportional to the duration of laser irradiation (FIG. 7).

MRI Relaxivity Analysis

[0079] A series of aqueous suspensions of NDG-Mn.sub.3O.sub.4 nanoparticles (with Mn concentration from 0 to 1 mM) were prepared and imaged in 0.2 mL Eppendorf tubes using a 3T clinical MRI instrument (GE Signa Excite Twin-Speed, GE Healthcare, Milwaukee, WI, USA). The specific relaxivity (r.sub.1) was calculated from linear curve generated from concentration of NDG-Mn.sub.3O.sub.4 nanocomposites versus 1/T.sub.1 (s.sup.-1).

[0080] For testing the effectiveness of NDG-Mn.sub.3O.sub.4 nanocomposites toward diagnostic standpoint, we investigated whether these nanoparticles have MRI contrast properties or not. Various concentrations of nanoparticles were subjected to imaging by 3T MRI scanner. The result demonstrated a concentration dependent enhancement of signal intensity with increasing concentration of NDG-Mn.sub.3O.sub.4 nanocomposites. The r1 value was found to be 0.09 mM.sup.-1s.sup.-1 (FIG. 8).

[0081] The foregoing description of the specific embodiments will so fully reveal the general nature of the embodiments herein that others can, by applying current knowledge, readily modify and/or adapt for various applications such specific embodiments without departing from the generic concept, and, therefore, such adaptations and modifications should and are intended to be comprehended within the meaning and range of equivalents of the disclosed embodiments. It is to be understood that the phraseology or terminology employed herein is for the purpose of description and not of limitation. Therefore, while the embodiments herein have been described in terms of preferred embodiments, those skilled in the art will recognize that the embodiments herein can be practiced with modification within the spirit and scope of the embodiments as described herein.

ADVANTAGES OF THE INVENTION

[0082] The present invention provides a nanocomposite comprising nitrogen doped graphene oxide conjugated with Mn.sub.3O.sub.4 nanoparticle for the effective treatment of cancer using photodynamic therapy.

[0083] The present disclosure provides a nanocomposite comprising nitrogen doped graphene oxide conjugated with Mn.sub.3O.sub.4 nanoparticle for the magnetic resonance imaging (MRI).