Preparation And Performance Of Melanin Nanoparticles From The Apricot Kernel

Melanin is widely present in nature and has many distinct biological functions.However,most kinds of melanin are insoluble in water and common organic solvents,which greatly limits the applications in the fields of biology and medicine.Nanotechnology can increase the solubility and bioavailability of natrual products.At the same time,new characteristics such as surface effects and size effects can be generated.Therefore,nanotechnology can be used to expand applications of melanin.So for,there has been limited studies about increasing solubility of natural melanin by nanotechnology,and thus to inprove solubility of melanin through the nanotechnology is worth investigating.In this paper,natural melanin was extracted from waste core-shell of wild apricot by alkali-solution and acid-isolation method.Firstly,the optimization of the extraction process was carried out using the single-variable method and response surface analysis（RSA）.Secondly,the melanin nanoparticles were prepared and its metal ion chelation,photothermal conversion performance,drug controlled release and in vitro safety were investigated.This paper aims at providing scientific basis for preparation and application of melanin nanoparticles.Also,it provides a new direction for the high-value utilization of the core-shell form wild apricot.The main findings are as follows:（1）Melanin from core-shell was extracted by alkali-solution and acid-isolation assisted by ultrasound.The optimum conditions for extraction were:sample-solvent ratio 1:10（g/mL）,sodium hydroxide concention 1.5 mol/L,precipitation pH 1 and the ultrasonic time 35 min.Under these conditions,the maximal yield of melanin was up to 4.68%.UV-vis of melanin from apricot kernel showed that the slope of the linear curve was-0.00186,which was consistent with the typical melanin.FT-IR of the melanin from the apricot kernel showed that the product have functional groups such as hydroxyl group,carboxyl group,amino group,alkyl group and aromatic ring.The results of SEM showed that the melanin from the apricot kernel was cuboid.The particle size of the melanin was about 1⁵μm.The results of solubility test showed that the natural melanin from the apricot kernel was insoluble in water and most organic solvents,but only soluble in alkaline solution or DMSO.Melanin from apricot kernel has the ability to scavenge DPPH radicals,OH radicals and ABTS radicals,and the radical scavenging ability was lower than Vc.The half inhibitory concentrations(IC₅₀)of melaninwere46.21μg/mL,143.49μg/mL,4.48μg/mL,respectively.Under the condition of pH=7.4,the maximum chelation rates between product and Fe（III）,Cu（II）,Zn（II）were 29.21%,6.49%and 2.26%.The maximum chelating amount were 1.80mmol/g,0.48 mmol/g and 0.17 mmol/g,respectively.（2）Natural melanin nanoparticles（NMNP）were prepared by ultrasonic method according to the different solubility of melanin in various solvents.In order to further improve the dispersion stability,NMNP were successfully modified by mPEG₂₀₀₀-NH₂.The particle size of the NMNP-PEG was about 15²0 nm.The results of UV-vis of NMNP showed that the slope of the linear curve was-0.002,which was consistent with the melanin from apricot kernel.The results of FT-IR showed that the NMNP prepared by this method were still retained the key functional groups of the melanin from apricot kernel.（3）The metalion chelation,photothermal conversion and stimuli-responsive drug release of NMNP-PEG were tested.The results showed that the NMNP-PEG could chelate Fe（III）,Cu（II）and Zn（II）through phenolic hydroxyl group,catechol group and amine group under the condition of pH=7.4.NMNP-PEG were found to possess higher chelation rate than natural melanin.The maximum chelation rates of Fe（III）,Cu（II）and Zn（II）were 48.51%,10.10%and 4.54%.NMNP-PEG have excellent photothermal conversion performance and stability,and near-infrared photothermal conversion efficiency was about 28.57%.The maximum drug-loading rate of NMNP-PEG was found to be 61.54%（w/w）.The drug release was only 11.90%without stimulation.When the pH was reduced to 5.0 or reactive oxygen species（ROS）was present,the cumulative release rate of DOX could reached 40.83%and21.92%after 48 h,which proved NMNP-PEG have a strong pH/ROS dependency.（4）In vitro safety of NMNP-PEG were evaluated.NMNP-PEG lost their absorbance coupled with color fading in the presence of hydrogen peroxide solution,which proved that NMNP-PEG were biodegradable.The hemolysis test showed that NMNP-PEG did not cause hemolytic reaction in the concentration range of 5⁴00μg/mL.NMNP-PEG hemolysis rate was less than 5%,which meets the requirements for medical purpose.This also indicated that the NMNP-PEG have good biocompatibility.Using MCF-7 breast cancer cell line,the cytotoxicity of the NMNP-PEG was investigated through CCK-8.The results showed that NMNP-PEG have no significant cytotoxicity in the concentration range of 0.78⁴00μg/mL.