Porphyran, a poly-anionic sulfated polyose isolated from marine ruddy algae Porphyra vietnamensis has been investigated as a reduction agent for size controlled synthesis of gold nanoparticles. The prepared AuNps showed SPR centered at 520 nanometers with mean atom size of 13±3 nanometers. FTIR spectra suggested that the sulphate mediety is chiefly responsible for decrease of aurochloric acid. The capping of the AuNps with porphyran was apparent from the negative zeta potency value which besides rendered electrostatic stableness. Therefore, porphyran act as both cut downing every bit good as cresting agent. These porphyran capped AuNps are extremely stable in a broad scope of pH and electrolytic concentration. Porphyran capped AuNps showed enhanced cytotoxicity on human glioma cell lines ( LN-229 ) as compared to native porphyran. Consequently, porphyran capped AuNps have been used for drug bringing application by lading the antineoplastic drug doxorubicin hydrochloride ( DOX ) . Spectroscopic scrutinies revealed that DOX conjugated onto porphyran capped AuNps via H bond between protonated aminoalkane group of DOX with porphyran capped AuNps. In acetate buffer ( pH 4.5 ) , release rate of DOX was found to be well higher as compared to physiological buffer ( pH 7.4 ) from DOX loaded porphyran capped AuNps. Further, these drug loaded porphyran capped AuNps demonstrated higher cytotoxicity on LN-229 cell line as compared with an equal dosage of native DOX solution. This established the potency of porphyran capped AuNps as a bearer for antineoplastic drug bringing.
Keywords: gold nanoparticles, porphyran, cytotoxicity, doxorubicin hydrochloride, and drug bringing
Novel colloidal bearers such as liposomes, polymeric nanoparticles and solid lipid nanoparticles are widely used for bringing of assorted biomolecules and pharmaceutical actives because they offer batch of advantages like improved efficaciousness, targeted bringing and decreased toxicity as compared to conventional drug bringing system.1-3 In the present province of personal businesss, metal nanoparticles are besides widely investigated, peculiarly gold nanoparticles ( AuNps ) for their alone belongingss such as comparable size with biomolecules, adhering ability to assorted molecules and optical belongingss in the seeable and NIR parts make them possible campaigners for chemical every bit good as biological applications and these can be synthesized easily.4 Recent explosion of research includes biological, diagnostic and curative applications of AuNps.5-11 Several research workers improved the belongingss of biomolecules and drugs such as amino acids,12 protein,13 DNA,14 insulin,15 ciprofloxacin16 and plasmid17 after junction with AuNps without changing their biochemical belongingss. But, in bulk instances chemically synthesis of AuNps18-19 were utilized for junction of drug molecules. In this procedure, chemical decrease of gold salt to AuNps was carried out by utilizing rough cut downing agent such as tri-sodium citrate and Na borohydrate and organic dissolvers doing them unsuitable for biological applications. To get the better of these challenges, the involvement in this field has been increased from past few old ages for use of such cut downing agent will synthesise size controlled AuNps and give sufficient stableness to AuNps under strong electrolytic and pH conditions and doing them suited for curative and drug bringing applications. Recently, Dhar et al. , synthesized extremely stable AuNps by utilizing gellan gum as a reduction agent for bringing of anticancer drug.20 Besides, in another survey biodegradable chitosan mediated synthesis of AuNps was improved the transmucosal bringing of insulin.21 In the present survey, we have synthesized one pot size controlled AuNps by utilizing of course occurred cut downing agent followed by junction of drug molecule on the surface of AuNps.
Porphyran, a poly-anionic sulfated polyose is isolated from marine ruddy algae ( Porphyra vietnamensis ) found preponderantly in rainy season on the west seashore of India, particularly in the states of Maharashtra, Goa and Karnataka.22 Porphyran consisting the hot-water soluble part of cell wall is the chief constituent of the ruddy algae. Porphyran comprises of disaccharide units dwelling of 3-linked-D-galactosyl residues jumping with 4-linked 3, 6- anhydro-l-galactose and the 6-sulfate residues.23 Porphyran is a dietetic fibre that contains about 40-50 % of seaweed constituents. Pharmacological maps of stray porphyran from several porphyra species were demonstrated in different structural and functional surveies. Previously few surveies have been reported on the antioxidant and anticancer activity of porphyran.23-24
Here, we have explored the usage of stray porphyran for synthesis of stable AuNps, such that it acts as a reduction every bit good as cresting agent. Prepared AuNps were evaluated through suited techniques to analyze morphology, surface charge and analyze functional group responsible for decrease of gold ions and adhering on AuNps. Effect of pH status and presence of electrolyte on synthesized AuNps was carried out. Six month stableness survey of these nanoparticles was performed at ambient temperature. We hypothesize that the porphyran capped AuNps would ensue in enhanced cytotoxicity as compared to native porphyran through MTT assay method on human glioma cell line ( LN-229 ) .26-28 Further to show pertinence for drug bringing we have investigated the burden of antineoplastic drug doxorubicin hydrochloride ( DOX ) on porphyran capped AuNps. DOX is a powerful anthracycline antibiotic used for assorted malignant neoplastic disease therapies such as malignances, carcinoma and sarcomas, Lack of tumour aiming capacity of DOX consequences in narrow biodistribution and unwanted side effects which remain as major jobs to be solved.25 This led the company ALZA to develop a merchandise DOXIL® that improved the public presentation by encapsulating the DOX in a liposome, thereby heightening the antineoplastic action of DOX. Here, we have presented the junction of DOX via H bond onto extremely stable porphyran capped AuNps may take to improved cytotoxic consequence on human malignant neoplastic disease cell line. Besides, in-vitro release surveies of DOX loaded porphyran capped AuNps was carried out in different pH to show the consequence of pH on the release of DOX.
2. Experimental Methods
2.1. Materials and reagents. Doxorubicin hydrochloride was gifted from RPG Life Sciences Limited, Mumbai ( India ) . Hydrochloroauric acid ( HAuCl4 ) was obtained from Sisco India Ltd. Mumbai. The human glioma cell line ( LN-229 ) was porches from American type civilization aggregation ( ATCC, USA ) . The xanthous tetrazolium MTT ( 3- ( 4, 5-dimethylthiazolyl-2 ) -2, 5-diphenyltetrazolium bromide ) was obtained from Sigma-Aldrich ( USA ) . Analytic class reagents are from Merck India Ltd. , India. All the samples were prepared in deionized H2O.
2.2. Isolation and word picture of porphyran. Porphyran was isolated from marine ruddy algae ( Porphyra vietnamensis ) .29,30 In brief, dried ruddy algae were soaked in 7.5 % formol for 12 H at ambient temperature. An equal volume of H2O was added and solution refluxed on boiling H2O for 8 h. The mixture was centrifuged at 10,000 revolutions per minute for 20 min and supernatant was filtered through diatomaceous earth. The ensuing filtrate was adjusted to pH 7 with Na hydrated oxide and 75 % volume was evaporated at 65 0C on Rota evaporator. Four fold volume of methyl alcohol was added to residuary solution to precipitate porphyran content. The mixture was centrifuged at 10,000 revolutions per minute for 20 min and supernatant was discarded. The precipitated porphyran was washed with 80 % aqueous methyl alcohol. The residue was freeze dried to give pale xanthous colour powdered porphyran. Entire sugar and sulphate content in porphyran was determined by the phenol sulphuric acid method31 and Kawai method.32 This freezing dried porphyran was used for farther surveies.
2.3. Synthesis of gold nanoparticles utilizing porphyran. In a typical experiment, 100 milliliter an aqueous solution of HAuCl4 ( 1X10-4 M ) incorporating porphyran ( 0.01 % w/v ) was reduced to AuNps by seting the pH of solution with Na hydrated oxide to 11 followed by heat up to boiling for 10 min yielded dark ruby ruddy colored AuNps. The AuNps scattering was exhaustively dialyzed for 24 H to take ionic drosss used during the decrease. After dialysis, the pH of the AuNps scattering was measured to be 7.
2.4. UV/Visible spectroscopy measuring. The UV/Visible spectra of native porphyran solution ( 0.1 % W/V ) , porphyran capped AuNps was monitored by UV/Visible spectrometry ( Jasco double beam UV/Visible spectrophotometer, Model V-570, Japan ) . Change in surface plasmon resonance ( SPR ) of porphyran capped AuNps was monitored.
2.5. Loading of DOX on porphyran capped AuNps. A deliberate sum of DOX was added to porphyran capped AuNps scattering ( pH 7 ) , obtained as described above, ensuing in a concluding DOX concentration of 10-4 M in solution. The mixture of DOX and AuNps scattering was incubated for 24 H at room temperature and so centrifuged at 10,000 revolutions per minute for 10 min. The obtained pellet after centrifugation was separated from the supernatant solution and redispersed in deionized H2O prior to farther word picture.
2.6. Determination of percent drug lading on porphyran capped AuNps. Drug burden was calculated from the difference between the initial DOX concentration and the otiose DOX determined in the supernatant liquids. The drug concentration in supernatant was determined by measurings of its UV optical density at 480 nanometers utilizing UV/Visible spectrometry and the per centum burden of DOX on porphyran capped AuNps was estimated by expression ( 1 )
2.7. Spectroscopy, zeta possible measuring, microscopic survey and X-ray diffraction. Fourier transform infra ruddy ( FTIR ) spectra of native porphyran, porphyran capped AuNps, native DOX and DOX loaded porphyran capped AuNps was recorded in KBr pellets utilizing FTIR spectrophotometer ( Jasco, Japan ) . The scan was performed in the scope 400 to 4000 cm-1. The zeta potency ( ZP ) was determined by utilizing the Zetasizer 300 HAS ( Malvern, UK ) . The zeta potency of porphyran capped AuNps and DOX loaded porphyran capped AuNps was determined as such without dilution. Which measured the surface charge on polyose capped AuNps and DOX loaded porphyran capped AuNps. The morphology and size distribution of the porphyran capped AuNps and DOX loaded porphyran capped AuNps scattering was carried out by high declaration transmittal negatron microscopy ( HRTEM ) measuring casting of nanoparticle scattering on carbon-coated Cu grids and allowed to dry at room temperature. Measurements were done on TECHNAI G2 F30 S-TWIN instrument operated at an accelerated electromotive force of 300 KV with a lattice declaration of 0.14 nanometers and point image declaration of 0.20 nanometers. The atom size analysis was carried out utilizing Gattan package ( Pleasanton, CA, USA ) . X-ray diffraction ( XRD ) measuring of porphyran capped AuNps was carried out by fixing movies of nanoparticle scattering on glass substrates by simple solvent vaporization method at room temperature. The diffraction measurings were carried out on X’pert Pro X ray diffractometer ( Netherlands ) instrument runing at 40 kilovolts and a current of 30 ma at a scan rate of 0.388/min.
2.8. pH and electrolytic stableness survey. In the pH stableness survey, the pH of porphyran capped AuNps scattering was adjusted utilizing 0.1N hydrochloric acid ( pH 2, 3, 4, 5 and 6 ) and 0.1M Na hydrated oxide ( pH 7, 8, 9, 10, 11 and 12 ) utilizing calibrated pH metre ( Delux pH metre 101, India ) . Change in SPR of porphyran capped AuNps scattering was recorded after 24 H utilizing a UV/Visible spectrophotometer. Besides, the consequence of electrolyte was studied by adding changing molar concentration of Na chloride ( 101 to 10-6 M ) to porphyran capped AuNps scattering. The alteration in SPR was recorded after 24 H utilizing UV/Visible spectrophotometer.
2.9. Stability survey. The stableness survey of porphyran capped AuNps composite was carried out at ambient temperature. The alteration in SPR of the nanoparticle scattering was recorded up to six month utilizing UV/Visible spectrometry.
2.10. In-vitro drug release survey. The dialysis tubing incorporating DOX loaded porphyran capped AuNps was transferred to a beaker incorporating 100 milliliter of phosphate buffer ( pH 7.4 ) by maintain temperature at 37 0C with uninterrupted stirring at 100 revolutions per minute. Sink status was maintained by sporadically taking 2 mL sample and replacing equal volume of buffer. The sum of released DOX was analyzed with a spectrophotometer at 485 nanometer. A similar release survey was carried out in ethanoate buffer ( pH 4.5 ) . The experiments were performed in triplicate for each of the samples.
2.11. In-vitro MTT check. Human glioma cell lines ( LN-229 ) was cultured in Dulbecco ‘s modified bird of Jove ‘s medium ( DMEM ) supplemented with 1.5 gm/mL sodium hydrogen carbonate, 4 millimeter glutamine and 5 – 10 % foetal bovine serum ( Gibco, USA ) . The civilizations were maintained in a humidified ambiance of 5 % CO2 at 37 0C in an brooder. For cytotoxicity testing, the cells were utilized when they reached 70 – 80 % meeting. The cells were diluted as needed and seeded as 3 X 103 for LN-229 in 200 µL of media per good, consecutive plated in level underside 96 good home bases ( Becton Dickinson Labwane, USA ) . This figure of cells was selected to avoid possible over meeting of the cells at the terminal of the four twenty-four hours experiment while still supplying adequate cells for equal formazan production. After plating, the 96 good home bases were so incubated for 24 H to let attachment of the cells prior to the disposal of assorted samples for proving. After complete attachment of cells the civilization medium was replaced with 200 µL of solution incorporating mixture of fresh medium and porphyran solution ( 0.01 % w/v ) , porphyran capped AuNps scattering, native DOX solution and DOX loaded porphyran capped AuNps in separate Wellss. Control wells incorporating cells received merely 200 µL of medium. After add-on of all the trial samples, the home bases were incubated up to 72 H for native porphyran and porphyran capped AuNps and 48 H for native DOX and DOX loaded porphyran capped AuNps in CO2 brooder. The cells could be maintained in Wellss for this period without the demand for re-feeding. All experiments were performed in triplicate. MTT check was based on the measuring of the mitochondrial activity of feasible cells by the decrease of the tetrazolium salt MTT ( 3- ( 4,5-dimethyathiazol-2-yl ) -2, 5-diphenyl tetrazolium bromide ) to organize a bluish non-water-soluble merchandise, formazan. MTT ( 5 mg/mL, 20 µL ) was added to respective set of cells and the home bases were incubated for an extra 4 h. After 4 H of incubation, the medium was removed and DMSO ( 200 µL, Sigma-Aldrich, USA ) was added to fade out the formazan crystals ensuing from the decrease of the tetrazolium salt merely by metabolically active cells. The optical density of dissolved formazan was measured at 570 nanometers utilizing a Bio-Rad microplate reader ( Model 680, Heraeus, USA ) . Since the optical density straight correlated with the figure of feasible cells, the per centum viability was calculated from the optical density.
3. Consequences and treatment
In this work, we have used porphyran isolated from marine ruddy algae ( Porphyra vietnamensis ) for decrease of HAuCl4, where it acts as a reduction and stabilising agent ( scheme 1 ) . The synthesized AuNps were farther utilized as a nanocarrier for the bringing of antineoplastic drug such as DOX.
3.1. Chemical analysis of porphyran. This pale yellow colored stray porphyran was dissolved in H2O on warming. It contained 78.92 % of entire sugar and 9.62 % of sulphate. Further, UV/Visible spectra of stray porphyran ( 0.1 % w/v ) in deionized H2O showed no extremum from 260 to 280 nanometers bespeaking absence of the protein and nucleic acid like constituents in the stray porphyran ( Figure 1A ) . The rating of functional group nowadays in the porphyran was carried out by FTIR analysis. The FTIR spectra of the porphyran ( Figure 1B ) depicted typical sets at 1645, 1417, 1230, 1158, 1019, 933 and 819 cm-1. The signal at 1230 cm?1 was assigned to the asymmetric stretching quiver of sulphate group, and the signal at 819 cm?1 was declarative of a sulphate group attached to a primary hydroxyl group. Another weak set at 933 cm?1 was due to the 3, 6-anhydro-D-galactose unit in the polyose. From these consequences and similar determination reported earlier we confirmed that the stray porphyran contained 3, 6-anhydrogalactose unit and sulphate group.33
3.2. Synthesis and rating of AuNps. This stray porphyran was used as a reduction agent and stabilising for synthesis of AuNps. Preliminary survey revealed that the decrease of gold ions in presence of porphyran did non happen up to pH 7, but as pH increased up to 11 decrease occured as indicated by the colour alteration. At pH 11 the colour of solution changed from colorless to dark ruby ruddy on heating for 10 min. Figure 2A shows the UV/Visible spectra recorded from the scattering obtained by the decrease of HAuCl4 utilizing 0.01 % w/v of porphyran at pH 11, the set matching to the SPR occurred at 520 nanometer. SPR of AuNps appears in the seeable part and can be used to supervise form, size and collection of the nanoparticles. It was observed that HAuCl4 decrease occurs quickly and the strength remained unchanged, without any displacement in the extremum wavelength even after 24 H of decrease clip. Inset exposure showed the colour of AuNps reduced at 0.01 % w/v porphyran ( Figure 2A ) . Figure 2B depicted the FTIR spectra of porphyran reduced AuNps, where signal of asymmetric stretching quiver of sulphate group shifted to 1283 cm?1 and the signal at 819 cm?1 ( sulfate group attached to a primary hydroxyl group ) was diminished after synthesis of AuNps bespeaking the engagement of the sulfur group of porphyran in synthesis of AuNps.
Zeta potency of porphyran reduced AuNps was found to be – 31.05 millivolt. The negative charge indicated that the AuNps were decently wrapped with poly-anionic porphyran. In general, particle collection is less likely to happen for charged atoms with optimal zeta potency ( ~ ±30 millivolt ) due to electrostatic repulsions.20 HRTEM images ( Figure 3A ) revealed that the porphyran capped AuNps appeared to be spherical in form with narrow size distribution and inset Figure 3A showed mean atom size about 13 ± 3 nanometer. The selected country of negatron diffraction form of the porphyran capped AuNps demoing the rings designated 1, 2, 3 and 4 arise due to the contemplations from ( 111 ) , ( 200 ) , ( 220 ) and ( 311 ) ( Figure 3B ) . This was further confirmed by the pulverization X-ray diffractogram recorded from the sample ( Inset Figure 3B ) , which may be indexed as the set for face centered three-dimensional ( Federal Communications Commission ) constructions of gold. The XRD form therefore clearly illustrated that the widening of Bragg ‘s extremums indicated the formation of nanoparticles34 and these are in crystalline signifier.
3.3. Stability survey at different pH status and electrolytic concentration. For varied drug bringing applications,35 we studied the stableness of porphyran capped AuNps by supervising the SPR over sensible period of clip at different pH and electrolytic conditions. It should be noted that a ruddy displacement in UV/Visible spectra is associated with either an addition in the average size of the atoms or collection of nanoparticles or a combination of both.36 In instance of pH survey, the pH of porphyran capped AuNps scattering was adjusted from pH 2-12. The sample was incubated nightlong and analyzed for any alteration in the SPR. Figure 4A depicted that alteration in peak strength and SPR displacement was non observed in pH scope of 3 to 12. Inset exposure suggested that the colour of porphyran capped AuNps turned to blue bespeaking the collection of nanoparticles at pH 2. Besides, add-on of electrolyte ( NaCl ) up to 1 X 10-2 M caused no major collection ( Figure 4B ) . Borohydrate or citrate reduced AuNps sum at little alteration in their pH and electrolytic condition.35 This rating was easy confirmed by the SPR place of porphyran capped AuNps and its sums, the undistinguished alteration in its place under alteration in the pH and electrolytic conditions bespeaking the inordinate stableness of porphyran capped AuNps. Besides, in long term stableness survey, AuNps did non demo displacement in SPR ( Figure 4C ) and inset HRTEM image of porphyran capped AuNps revealed that no alteration was observed in atom size and form over six month stableness period.
3.4. In-vitro cytotoxicity of porphyran capped AuNps. Several research workers have made efforts to look into the macrophage-stimulating activity of the porphyran by agencies of in-vitro and in-vivo survey. Yamamoto et Al. reported that the unwritten disposal of several seaweeds could do a important lessening in the incidence of carcinogenesis.37 In recent old ages, algal polyoses have been reported to hold free-radical scavenging activity and act as an antioxidant for the bar of oxidative harm in life organisms.38, 39 Previously, Kwon et Al. reported the anti-proliferative consequence of porphyran on human stomachic carcinoma cell line, where 0.25 % and 0.5 % of porphyran showed important suppression of cell growing as compared to control.24
In order to show the consequence of the nanocarrier such as AuNps on the cytotoxic activity of porphyran, we have carried out cytotoxicity survey of equal concentration of native porphyran and porphyran capped AuNps on LN-229 cell line utilizing in-vitro MTT check method. Both native porphyran and porphyran capped AuNps revealed cytotoxic consequence on LN-229 cell line. It was interesting to observe a quadruple addition in cytotoxicity of porphyran capped AuNps as compared to native porphyran ( Figure 5 ) . This clearly demonstrated the function of AuNps for the sweetening of cytotoxic consequence of porphyran on human glioma cell lines. Previous study justified that porphyran exhibited cytotoxicity on human stomachic carcinoma cell line via initiation of programmed cell death related signaling through activation of proapoptotic molecules ( Bax and caspase-3 ) , suppression of anti-apoptotic molecule ( Bcl-2 ) , suppression of IGF-I receptor and lessening in the degree of Akt activation.24 In our instance, betterment in cytotoxicity of porphyran capped AuNps can be attributed to the greater uptake potency by endocytosis of the AuNps as compared to native porphyran.21,40
3.5. Evaluation of DOX loaded porphyran capped AuNps. After the successful synthesis of stable porphyran capped AuNps, we have envisaged this system for drug bringing application through subsequent burden of a bioactive molecule. Therefore, we have selected a low anthracycline pealing incorporating antineoplastic drug DOX ( pKa=8.2 ) for lading on porphyran capped AuNps. Loading efficiency of DOX on porphyran capped AuNps was found to be 60 % after 24 h incubation at room temperature. Figure 6A represented HRTEM image of DOX loaded porphyran capped AuNps revealed undistinguished alteration in atom size ( 14±3 nanometer ) and inset image represented uniformly redispersed DOX loaded porphyran capped AuNps scattering. The lessening in the zeta potency ( from -31.05 millivolt to -19 millivolt ) of DOX loaded porphyran capped AuNps was ascribed to the presence of positively charged DOX on the surface of porphyran capped AuNps. Thus even after DOX burden, porphyran capped AuNps remained as a stable scattering owing to the electrostatic repulsive force through the negative surface charge. It was thought that along with the electrostatic interaction other attractive forces including H bond could be playing a major function easing the drug lading procedure. The H adhering hypothesis between protonated aminoalkane groups of the DOX molecule with porphyran capped AuNps is besides supported by FTIR, where NH stretching set of native DOX at 3314 cm-1 shifted to 3413 cm-1 in instance of DOX loaded porphyran capped AuNps suggested the formation of H bond between protonated aminoalkane group of DOX with porphyran capped AuNps ( Figure 6B ) .
3.6. In-vitro drug release survey. Figure 7A depicted the release of DOX from DOX loaded porphyran capped AuNps in ethanoate buffer ( pH 4.5 ) and phosphate buffer ( pH 7.4 ) . At the terminal of 7, 98 % and 15 % of DOX was released in ethanoate and phosphate buffer severally. This consequence revealed that release of DOX was found to be higher in acidic pH as compared to basic pH. This pH dependent release may assist to better efficaciousness of DOX because consumption of drug loaded nanoparticle through endocytosis procedure leads to exposure to an acidic environment,41 which may originate rapid release of DOX from DOX loaded porphyran capped AuNps. Such efficient release would finally ensue in improved cytotoxic efficaciousness against tumour cells. Besides, really low release of DOX in basic pH will assist to cut down toxicity of DOX to the normal tissue because physiological pH of organic structure is maintained at pH 7.4, such pH status may suppress the release of DOX from DOX loaded porphyran capped AuNps.42
3.6. In-vitro cytotoxicity of DOX loaded porphyran capped AuNps. To set up the capablenesss of the porphyran capped AuNps drug transporting engineering, we determined the cytotoxicity of native DOX solution and DOX loaded porphyran capped AuNps on LN-229 cell line utilizing in-vitro MTT check method. Figure 7B illustrated dose dependent cytotoxic consequence of DOX in the signifier of either DOX loaded porphyran capped AuNps or native DOX on the LN-229 cells after 48 H exposure. DOX loaded porphyran capped AuNps exerts a higher cytotoxic consequence than native DOX on LN-229 cells at the same dosage. At the terminal of 48 H, the lessening in cell viability with native DOX and DOX loaded porphyran capped AuNps in the concentration scope studied ( 1.0-20 µg/mL ) was found to be between 60-35 % and 48-20 % , severally.
Previously, Serpe et Al. reported higher cytotoxicity of doxorubicin when incorporated in solid lipid nanoparticles due to the fast internalisation of doxorubicin loaded solid lipid nanoparticles followed by the drug ‘s release inside the cells.43 In our survey, we observed a important addition in the cytotoxicity of DOX on LN-229 when loaded on AuNps compared to native DOX. The addition in cytotoxicity of DOX loaded porphyran capped AuNps may be due to the enrichment in internalisation of DOX loaded porphyran capped AuNps by an endocytosis mechanism as compared to the inactive diffusion mechanism of native DOX into cells.44 In an earlier cytotoxicity survey of antineoplastic drug loaded AuNps, Chen et Al. have reported the intracellular accretion of amethopterin conjugated AuNps owing to AuNps mediated endocytosis.40
In decision, we have reported size controlled synthesis of AuNps by utilizing stray Marine porphyran from ruddy algae. These nanoparticles exhibited stableness in a broad scope of pH and electrolyte concentration. Further, pertinence of these nanoparticles as bearers for the bringing of the cationic anticancer drug was demonstrated by successful burden of DOX onto porphyran capped AuNps. In-vitro cell line survey revealed higher cytotoxicity of porphyran capped AuNps and DOX loaded porphyran capped AuNps in human glioma cell lines as compared to native porphyran and native DOX solution. Further, in-vivo toxicity survey of porphyran capped AuNps and in-vivo anti-tumor activity of DOX loaded porphyran capped AuNps are under probe in our research lab.