WO2016094991A1 - Method for producing pegylated silica nanoparticles as carriers of hydrophobic pharmaceutical drugs, thus obtained nanoparticles and uses thereof - Google Patents

Abstract

The present invention relates to a method for producing PEGylated silica nanoparticles as carriers of hydrophobic pharmaceutical drugs, to the silica nanoparticles obtained by this or other methods, and to the use thereof for the treatment of cancer. The nanoparticles effectively retain the pharmaceutical drug inside them, preventing the premature release of the drug in the external medium during intraveinous transport, reducing the collateral effects associated with chemotherapy and the dose concentration required for an antitumoral treatment.

Description

 PROCEDURE FOR OBTAINING PEGUILATED SILICAN NANOPARTICULES CARRYING HYDROPHOPIC PHARMACEUTICALS, SO NANOPARTICULES

 OBTAINED AND ITS USES

FIELD OF INVENTION

 [01] The present invention falls within the field of nanotechnology comprising the scientific platforms of material chemistry and solid state chemistry with emphasis on the interaction of nanostructures with biosystems, also known as nanobiotechnology.

 [02] More specifically, the present invention relates to a process for obtaining pegylated silica nanoparticles carrying hydrophobic drugs, the silica nanoparticles obtained or not by said process and their use in anti-tumor and anti-inflammatory treatments.

BACKGROUND OF THE INVENTION

 [03] Cancer is a disease caused by exacerbated cell growth, invading tissues and organs. This disease is derived from external or environmental factors and in approximately 90% of cases external factors are the main causes of the disease. If discovered early, the disease can be remedied by several treatments, among them are chemotherapy, radiation, hormones, among others. In fact, chemotherapy is the most common type of treatment and involves the use of antitumor drugs.

[04] Cancer drugs have several side effects when applied to the body, as a result of the low selectivity of the drug that reaches healthy body cells and, consequently, causes high toxicity. In addition, there is a short circulation time in the blood and high 15 000202

 2

doses a. applied, further causing such negative effects.

 [05] In the literature, there are several types of anticancer drugs, among them Camptothecin, Cisplatin and 5-fluorouracil as agents used to fight lung cancer. Also worth mentioning are organic substances. such as Anamycin, Doxorubicin and Lurtotecan, applied to the treatment of kidney (carcinoma) and stomach tumor cells. Docetaxel has the function of fighting metastatic breast cancer, esophageal adenocarcinomas, among others, along with Doxorubicin and Paclitaxel.

 [06] One drug that has been showing good effects is violacein, a violet-colored pigment produced by the bacterium Chromobacterium violaceum found in the Amazon. Studies conducted by researcher Nelson Duran and collaborators, from the State University of Campinas, showed the characteristics of this drug and its high anti-tumor, antiulcerogenic, antiviral, antibacterial and anti-leishmania ability. It is an extremely important drug in many medical applications, which underscores the need to produce molded nanoparticles for drugs with such characteristics.

[07] Due to the hydrophobic characteristics of certain antitumor drugs such as Camptothecin and Violacein, their solubilization in (hydrophilic) blood plasma becomes impossible, impairing their biological activity. Given these factors, it is worth stressing the importance of producing nanoparticles with hydrophobic inner surface, ideal for drug internalization, as well as their subsequent transport to target cells due to the presence of hydrophilic organic functions on the outer surface of nanoparticles. , providing solubility and locomotion. The drug will be compatible with the internal medium of the nanoparticle which, in turn, will hold stability in the blood. Therefore, the nanoparticle becomes suitable for the process of transport and release of the drug in biological targets.

 [08] In the prior art, several nanoparticles are reported, and there has been a large increase in the number of technologies involving nanoparticle production applied in the context of controlled drug release.

[09] In 2001, Nakanishi and colleagues developed a polymeric micellar nanocarrier called NK911, where the doxorubicin antitumor drug was inserted. The outer surface of this carrier was subjected to polyethylene glycol functionalization to avoid its uptake by the immune system and favors colloidal stability in hydrophilic solvents. Said technology described in the journal resembles the present invention in that it comprises nanoparticles capable of carrying drugs within them and comprises polyethylene glycol (PEG) on the outer surface. However, the present invention differs from the technology described in the article in that it comprises nanostructure functionalized silica, simultaneously with PEG on the outer surface and with hydrophobic organic functions on the inner surface, providing effective drug transport while maintaining it in a stable manner. effective within the nanoparticle due to the strong interactions between hydrophobic groups and the drug itself. The described technology does not hold said inner groups, providing drug exit during transport. In addition, micellar carriers do not have a rigid surface which gives the silica nanoparticle greater physical integrity. Advantageously, the silica nanoparticles proposed in the present invention have a rigid surface that allows the surface to functionalize internally with drug-interacting hydrophobic organic groups. Additionally, they are distinct nanoparticles in relation to the nanostructure and chemical composition.

[010] In 2005, Latallo and colleagues developed nanoparticles consisting of acetylated dendritic polymers whose outer surfaces were functionalized with folic acid binders aimed at obtaining nanoparticles capable of specifically targeting tumor cells. Said cell types show strong expression of membrane receptors for folic acid. Thus, the insertion of this chemical species on the nanoparticle surface guarantees its specificity to the target cells. The technology described in the article resembles the present invention in that it comprises antitumor drug carrier nanoparticles, however, they do not have PEG on the outer surface, which is important for the biocompatibility of the nanoparticle in the organism, and it does not understand the inner surface. hydrophobic to keep the drug entrapped during intravenous transport. Thus, although it has specificity in the attack on cancer cells, the technology described in the article presents problems regarding the possible premature release of the drug in the organism, previously reaching the target cell so that the treatment efficiency is impaired. Advantageously, the present invention comprises PEG-functionalized silica nanoparticles on the outer surface, providing biocompatibility in the body, and keeping the drug trapped within the hydrophobic interior until the target cell is reached. Additionally, they treat different nanoparticles in relation to the nanostructure and chemical composition.

 WO2009038659 relates to organically modified silica nanoparticles with covalently linked photosensitizers applied for photodynamic drug release. The technology described in WO2009038659 resembles the present invention in that it comprises aromatic groups on the inner surface of the nanoparticle. However, said organic groups are inserted in the middle of a complex molecular structure characterized by photosensitizers, that is, they are part of an organic entity whose function is solely and exclusively linked to the imaging properties of the nanoparticle in tumors. In addition, the document does not mention the drug-encapsulating property of the nanoparticle - it has no antitumor potential - and it does not have polyethylene glycol (PEG) groups on its outer surface - biocompatibility problems and short blood circulation time. Unlike the nanoparticles claimed in WO2009038659, the present invention comprises hydrophobic groups on the inner surface capable of retaining the drug in the pores during transport in the blood fluid to the reach of the cancer cell when it is released. In addition, the nanoparticle comprises PEG groups on the outer surface, which are important for nanoparticle biocompatibility, high colloidal stability essential for locomotion of the nanostructure in the blood (hydrophilic fluid) and long time of blood circulation.

US20100255103 relates to silica nanoparticles for biomedical applications. Technology resembles the present invention in that it comprises a silica nanoparticles whose outer surface is coated with PEG and by encapsulating hydrophobic drugs comprising camptothecin. The nanoparticles claimed in the patent have the following technical problems (solved by the present invention): the interior of the patent nanoparticles comprises magnetic cores such as iron oxide and gold; These are inorganic entities occupying a significant internal volume, impairing the ability to incorporate drug; furthermore, the patent does not claim pore volume, of fundamental importance for said drug encapsulation capacity. In the present invention there are results of physicochemical characterization proving the pore volume of the pegylated nanoparticles, ideal for the incorporation of active biological molecules, characterizing one of the protection strategies in the present patent application; The interior of the nanoparticles claimed in the US patent is free of hydrophobic groups capable of retaining the drug during intravenous locomotion. Applying said nanostructures in the blood there is a likelihood that the drug will exit the interior of the pores into the fluid, causing the drug to disperse in the blood, which is undesirable. In this context, the nanoparticles proposed in the present invention comprise porous interior hydrophobic groups for retention of the biological active during the intravenous pathway to the target cell range.

US20130274226 relates to silica nanoparticles with conjugated agents. The technology comprises silica nanoparticles whose outer surface is coated with PEG functions and by encapsulating hydrophobic drugs in their _. inside. The present invention differs from the technology disclosed in US20130274226 in that it comprises nanoparticles whose inner surface comprises hydrophobic groups capable of retaining the drug during intravenous locomotion, an important feature in order to avoid possible losses of the biological active to the blood fluid during the carriage to the target cell. The technology also has the following problems: The technology uses drug-conjugated silane reagents to form covalent bonding of the drug within the nanoparticle. In this context, there are difficulties in drug release during intravenous transport, since drug release depends on the degradation of the covalent chemical bond of the silane-drug radical. In contrast, the present invention maintains the drug retained within the hydrophobic interior during carriage as a function of interactions with hydrophobic groups. In addition, covalent incorporation of the drug into the nanoparticle prevents the incorporation of other hydrophobic drugs into a single batch of nanoparticles, requiring another nanoparticle production process containing another specific type of hydrophobic biological asset bound as another silane drug - note that for each Batch only one type of drug can be incorporated, as it is in the form of silane (silane-drug), requiring its addition in the process together with the other precursors of nanoparticle to ensure condensation inside, making it impossible to be added later in a situation in which a batch of still drug-free silica nanoparticles is incorporated. Advantageously, the present invention has the versatility of encapsulating a range of drugs with hydrophobic chemical characteristics, since only one batch is required to produce silica nanoparticles. Once nanoparticles are obtained, any type of hydrophobic antitumor drug can be incorporated in the shipyard without the need to produce other batches, saving raw material, energy and production time.

 Thus, the proposed nanoparticles simultaneously comprise hydrophilic outer surface due to the presence of polyethylene glycol and hydrophobic inner surface related to the presence of hydrophobic functional groups. Additionally, nanoparticles have as a differential the versatility to incorporate hydrophobic drugs.

 [015] Moreover, the proposed process has the differential that it does not use toxic solvents in its stages, encompassing two strategies of functionalization of the outer surface of nanoparticles, as well as understanding mild and specific conditions to produce nanoparticles with the mentioned characteristics.

BRIEF DESCRIPTION OF THE INVENTION

 [016] The present invention relates to a process for obtaining pegylated silica nanoparticles carrying hydrophobic drugs, the silica nanoparticles obtained or not by said process and their use in anti-tumor and anti-inflammatory treatments.

The process of obtaining drug-bearing pegylated silica nanoparticles comprises the following steps: dissolution of surfactant in ammonia catalyst and alcohol reaction solvent, stirring of the solution, addition of inorganic silicon precursor in hydrophobic silane, addition of solution obtained in the previous step in the solution obtained in the first step, agitation of the obtained solution, addition of inorganic silicon precursor, centrifugation, supernatant disposal and redispersion in ethanol, functionalization of the external surface of the silica nanoparticles, preparation of an ethanolic solution of HCl, addition of nanoparticles to the solution prepared in the previous step, application ultrasound, centrifugation, supernatant disposal and redispersion in ethanol, suspension of nanoparticles obtained in aqueous solution, addition of hydrophobic drug, mixing of the components comprised in the previous step, separation of nanoparticles, obtaining pegylated nanoparticles carrying hydrophobic drugs.

 Nanoparticles obtained or not by said process comprise polyethylene glycol (PEG) -functioning outer surface, hydrophobic-functionalized porous inner surface, and encapsulated hydrophobic drug.

[019] Finally, the invention also describes the use of the carrier of hydrophobic drugs pegylated nanoparticles in ^■ antitumor and anti-inflammatory treatments.

BRIEF DESCRIPTION OF THE FIGURES

 [020] Figure 1: Molecular structure of PTES (a), TEOS (b) and CTAB (c).

 [021] Figure 2: Arrangement of phenyl groups in the spaces between CTAB molecules.

 [022] Figure 3: Representative image of the starting nanoparticle.

 [023] Figure 4: Representative image of externally functionalized nanoparticle.

 [024] Figure 5: Molecular structure of the silanes used.

 [025] Figure 6: Representation of nanoparticle production and its functionalization.

 [026] Figure 7: Summary of the production of the starting nanoparticles.

[027] Figure 8: General representation of the functionalization reaction of the starting nanoparticle with PEGio silane. [028] Figure 9: General representation of the functionalization reaction of GPS silane starting nanoparticle and coupling of PEG40 to terminal epoxy function.

 [029] Figure 10: Summary process of surfactant extraction.

 [030] Figure 11: Functionalizations performed and their location in the molecular vehicle.

 [031] Figure 12: Infrared Vibrational Spectra (FTIR) for starting nanoparticles before and after CTAB removal.

 [032] Figure 13: Thermogravimetric (TG) curves and their respective differential thermal analysis (DTA) curves of the starting nanoparticles.

 [033] Figure 14: NPH30-SiOH N2 adsorption-desorption isotherms and NPH30-CTAB-SiOH.

 [034] Figure 15: Micrograph (TEM) of internally functionalized silica nanoparticles with phenyl groups after CTAB extraction.

 [035] Figure 16: Infrared Vibrational Spectra (FTIR) for NPH30-SiO-1 OPEG10.

 Figure 17: NPH30-SiO-10PEG10 TG and DTA curves.

 [037] Figure 18: NPH30-SiO-10PEG10 N2 adsorption-desorption isotherms.

 Figure 19: NPH30-SiO-10PEG10 carbon NMR spectrum.

 Figure 20: NPH30-SiO-10PEG10 silicon NMR spectrum.

 [040] Figure 21: Infrared Vibrational Spectrum (FTIR) for NPH30-SiO-25PEG10.

 Figure 22: TG and DTA curves of NPH30-SiO-25PEG10. Figure 23: NPH30-SiO-25PEG10 N2 adsorption-desorption isotherms.

 Figure 24: NPH30-SiO-25PEG10 carbon NMR spectrum.

Figure 25: NPH30-SiO-25PEG10 silicon NMR spectrum. 2

 11

[044] Figure 26: Infrared Vibrational Spectrum (FTIR) for NPH30-SiO-50PEG10.

 Figure 27: TG and DTA curves of NPH30-SiO-50PEG10.

 [046] Figure 28: N2 adsorption-desorption isotherms in NPH30-SiO-50PEG10.

 [047] Figure 29: NPH30-SiO-50PEG10 carbon NMR spectrum.

 [048] Figure 30: NPH30-SiO-50PEG10 silicon NMR spectrum.

 [049] Figure 31: Micrograph (TEM) of silica nanoparticles functionalized internally with phenyl groups and externally with polyethylene glycol.

 [050] Figure 32: Infrared Vibrational Spectra (FTIR) for NPH30-SIO-10GPS-PEG40.Error! Reference source not found. Figure 33: TG and DTA curves of NPH30-SiO-10GPS-PEG40.

 [051] Figure 34: N2 adsorption-desorption isotherms in NPH30-SiO-10GPS-PEG40.

 Figure 35: 13 C NMR spectra of NPH30-SiO-10GPS-PEG40.

 Figure 36: NPH30-SiO-10GPS silicon NMR spectrum.

 [054] Figure 37: Colloidal stability graph of nanoparticles at 250 g / mL in PBS.

 [055] Figure 38: Dose response curves of nanoparticles incubated in red blood cells.

 [056] Figure 39: Study of nanoparticle aggregation in PBS for one hour.

 [057] Figure 40: Displays the curcumin calibration curve in ethanol as well as the information for each spectrum acquired at each of the five known concentrations.

[058] Figure 41: Graphs the concentration of curcumin in the pegylated silica nanoparticles after each separation by centrifugation and decantation. [059] Figure 42: shows the image of suspensions of silica nanoparticles NPH30-SiOH, NPH30-1 OPEGsoo, NPH30-50PEG ₅ and oo-NPH30 lÕPEG5ooo in the presence of curcumin and said drug added to water in the absence of molecular vehicles ( H2O) after 24 hours of decantation and prior to removal of the supernatant.

DETAILED DESCRIPTION OF THE INVENTION

 [060] The present invention comprises the process of obtaining drug-bearing pegylated silica nanoparticles, the steps of which comprise:

a) surfactant dissolution between 0.65 and 0.75 g, preferably 0.75 g, in ammonia catalyst in a concentration between 0.03 and 0.07 mol / L, preferably 0.05 mol / L, and solvent of alcoholic reaction in volumes between 2.5 and 3.8 mL, preferably 3.2 mL;

b) stirring the solution obtained in the previous step at a temperature between 40 and 80 ° C, preferably 60 ° C;

c) adding inorganic silicon precursor between 1 and 3 mL, preferably 1.5 mL, in hydrophobic silane;

d) adding dropwise the solution obtained in step (c) to the solution obtained in step (a);

e) stirring the solution obtained in the previous step for 120 minutes at temperatures between 40 and 80 ° C, preferably 60 ° C;

el) addition of inorganic silicon precursor after 60 minutes of reaction at volumetric quantities between 100 and 150 i, preferably 124 μL;

e2) adding inorganic silicon precursor, after 90 minutes of reaction, in volumetric amounts between 100 and 150 µl, preferably 124 L; f) Centrifugation for 40 to 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000; discarding the supernatant and redispersing in ethanol;

g) Functionalization of the external surface of the starting silica nanoparticles in two different ways (1 or 2):

gl.l) dispersion of starting nanoparticles in masses between 50 and

500 mg, preferably 300 mg, in ethanol solvent in volumes between 20 and 60 mL, preferably 40 mL; stirring the solution between 40 and 90 minutes, preferably 75 minutes at a temperature between 40 and 80 ° C, preferably 60 ° C;

gl.2) Addition of silane-PEG to the solution of step gl.l;

gl.3) Stirring the solution from step gl.2 between 10 and 20 hours, preferably 13 hours;

gl.4) Centrifugation of the solution comprised in step gl.3 between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000; discarding the supernatant and redispersing in ethanol; repeated at least twice; or

g2.1) dispersion of starting nanoparticles in masses between 50 and 500 mg, preferably 300 mg, in ethanol solvent in volumes between 40 and 80 mL, preferably 62 mL and stirring of the solution between 20 and 60 minutes, preferably 30 minutes at room temperature. between 60 and 90 ° C, preferably 85 ° C;

g2.2) Addition of an intermediate silane with reactive functional groups to the obtained solution;

g2.3) Stirring the solution between 10 and 20 hours, preferably 12 hours in an inert gas atmosphere;

g2. ) Centrifuging the solution between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000 and discarding the supernatant and redispersing in ethanol; repeated at least twice; g2.5) Dispersion of intermediate nanoparticles obtained in step g2. in masses between 50 and 500 mg, preferably 150 mg, in ethanol solvent in volumes between 10 and 30 mL, preferably 20 mL and stirring of the solution between 15 and 45 minutes, preferably 30 minutes at a temperature of 95 ° C;

g2.6) - Addition of a strong base to the obtained solution;

g2.7) Dissolution of PEG-COOH (PEG-terminal carboxylic acid) comprising mass between 5 and 70 mg, preferably 15.1 mg in ethanol solvent in volumes between 500 and 1500 μΙ, preferably 750 pL;

g2.8) Addition of the PEG-COOH solution from step g2.7 to the solution obtained in step g2.6 for coupling reaction between reactive functional groups of intermediate nanoparticle with reactive functional groups. and PEG-COOH carboxylic acid;

g2.9) Stirring the solution obtained in step g2.8 between 10 and 20 hours, preferably 12 hours at a preferred temperature of 95 ° C; g2.10) Centrifuging the solution obtained in step g2.9 between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000, discarding the supernatant and redispersing in ethanol; repeated at least twice.

h) Preparation of an ethanololic solution of HCl in a volume ratio of preferably 1: 9 HCl: ethanol;

(i) Addition of nanoparticles comprised in steps gl. 5 or g2.12 in the solution prepared in step h; ultrasound application at times between 10 and 30 minutes, preferably 15 minutes;

j) Centrifuging the solution obtained in step (i) between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000; discarding the supernatant and redispersing in ethanol; repeated at least twice; and storage until encapsulation of the hydrophobic drug; (k) Suspension of nanoparticles obtained in step (j) in aqueous solution (component 1) is in the range of 5 to 15 mg, preferably 10 mg, and in concentrations up to 1 mg / mL; and adding 0.5 to 2 mg, preferably 1 mg, of hydrophobic drug (component 2); 1) Mixing the components comprised in step (k) by means of ultrasonic homogenization at times between 30 and 60 minutes;

m) separating the silica nanoparticles carrying hydrophobic drugs comprising said encapsulated drug (product) from unencapsulated hydrophobic drug (component 1) by separation means; ,

n) Obtaining silica nanoparticles carrying hydrophobic drugs comprising said encapsulated drug.

 [061] The process described herein comprises the following characteristics:

 the surfactant preferably comprises hexadecyltrimethylammonium bromide;

 - the inorganic silicon precursor comprises silicon alkoxides and organosilanes, preferably tetraethylorthosilicate;

- the hydrophobic silane is used in amounts between 10 and 40 Si-mol%, preferably 30% ^'Si-mol;

 - the hydrophobic group comprised in hydrophobic silane comprises hydrophobic groups selected from aromatic groups and derivatives; and hydrocarbon groups and derivatives;

 - aromatic groups and derivatives are selected from the group of phenyl, acetophenyl, and pyrene functions;

 - hydrocarbon and derivative groups are selected from the group of octyl and octadecyl functions;

 - PEG silane is used in amounts between 2 and 50 mol% Si;

- PEG comprised in silane-PEG comprises at least 350 Da; - intermediate silane with reactive functional groups is used in amounts between 2 and 50 mol% Si;

 reactive functional groups comprised in intermediate silane comprise epoxy functions;

 - the strong base used preferably comprises potassium hydroxide;

 - the PEG-COOH used comprises at least 350 Da;

 - the hydrophobic drug is used in amounts from 1 to 20% by mass;

- hydrophobic drug used is selected from the group of drugs comprising curcumin _; camptothecin, doxorubicin, violacein, tamoxifen, beta-lapacone and the like, preferably camptothecin.

 The separation means described in the process comprise centrifugation or decantation.

 Centrifugation is performed at rotational forces between 1500 and 2000 rpm, preferably for 2 minutes.

 [064] The decantation is carried out in time of at least 24 hours.

 Alternatively, the sequence of steps k, l, m and n may be performed by the following sequence of steps:

 k ') Suspension of nanoparticles obtained in aqueous solution, addition of hydrophobic drug and keep for 4 hours;

 1 ') Centrifugation of nanoparticles from step (k) under the same conditions established in step (j) and obtaining pegylated nanoparticles carrying hydrophobic drugs.

The pegylated hydrophobic drug carrier nanoparticles of the present invention comprise the following characteristics: outer surface functionalized with polyethylene glycol (PEG); porous inner surface functionalized with hydrophobic groups;

 - encapsulated hydrophobic drug;

 - encapsulated curcumin.

 More specifically, pegylated nanoparticles carrying hydrophobic drugs comprise the following characteristics:

 - PEG of at least 350 Da;

 - PEG functionalized in amounts between 2 and 50 mol-Si;

 - functionalized hydrophobic groups in amounts between 10 and 40 mol% Si, preferably 30 mol% Si;

 - hydrophobic groups selected from aromatic groups and derivatives; and hydrocarbon groups and derivatives;

 - aromatic groups and derivatives selected from the groups of phenyl, acetophenyl, and pyrene functions;

 - hydrocarbon-based groups and derivatives selected from the group of octyl and octadecyl functions;

 - hydrophobic drug in amounts of 1 to 20% by mass;

 hydrophobic drug selected from the group of drugs comprising curcumin, camptothecin, doxorubicin, violacein, tamoxifen, beta-lapacone and the like, preferably curcumin and camptothecin;

 - particle diameter between 40 and 100 nm;

 - zeta potential between -5.3 and -35 mV;

- surface area between 760 and 943 m ² / g;

pore volume between 1.0 and 1.8 cm ³ / g;

-cause maximum hemolytic effects between 2 and 14% and generate 50% hemolysis at concentrations between 35 and 75 micrograms / mL. [068] The present invention further comprises the use of pegylated hydrophobic drug carrier nanoparticles in anti-tumor and anti-inflammatory treatments.

 [069] The process of producing the pegylated hydrophobic drug carrier nanoparticles is initiated with the surfactant, the silane to be internally disposed in the nanoparticle, the inorganic silicon precursor that will form the inorganic wall, and the catalyst. After this process, an additional amount of inorganic silicon precursors are implemented by controlling time so that excessive (excess reaction time) or precarious (reaction time) polymerization is avoided as they promote the formation of silicon walls. silica very thick and significantly thin respectively. Therefore, adequate nucleation time control is required. After the internal compartment is properly separated from the external environment by the already nucleated silica wall according to the reaction parameters (reaction time and temperature), the external surface can be functionalized with the silane of interest, followed by the removal of the surfactant. Methods of hierarchical functionalization are described in the literature according to the descriptions of this paragraph.

[070] In a detailed and sequential manner, the production process of the hydrophobic drug-bearing pegylated nanoparticles is initiated by the co-condensation process involving the inorganic silicon precursor (TEOS), comprising the inorganic source of silica, hydrophobic silane comprising hydrophobic groups selected from the aromatic and derivative groups (phenyl, acetophenyl and pyrene) and hydrocarbon and derivative groups (octyl and octadecyl) which are located on the inner surface of the silica, the surfactant preferably comprising hexadecyltrimethylammonium bromide (CTAB), the function of which is to form spherical micelles, whereby aqueous ammonium hydroxide as catalyst and ethanol as reaction solvent as a template. Figure 1 shows the molecular structure of the mentioned components.

[071] In the process, TEOS and hydrophobic silane mutually hydrolyze and initiate their polymerization around the micelles, so that they are muted. that ^"silica. negatively charged electrostatically interacts with. . the positively charged micellar surface and hydrophobic groups arrange themselves within the small spaces between the hydrophobic tails of the surfactants. This situation can best be seen in Figure 2.

 After this step, a certain amount of TEOS is added again to increase the thickness of the inorganic silica layer, effectively separating the hydrophobic interior from the hydrophilic external medium. Thus, the starting nanoparticle is obtained, as indicated in Figure 3.

: [073] After the formation of nanoparticle containing phenyl group inside the silica sphere, the outer surface is also subjected to a novel process of functionalization (after functionalization), whose goal is to insert bodily functions which confer new ^'properties nanoparticles , as observed in Figure A .. In the image, the group ^' R indicated in. nanoparticle represents any organic function to be grafted to the outer surface through a previously condensed silane.

As shown in the nanoparticle structure in Figure 14 the main strategy is to produce antagonistic nanoparticles, i.e. covering hydrophobic interior and hydrophilic outer surface due to the applications in which they are expected. [075] In the present invention, the major silanes involved in external surface functionalizations are compounds whose R groups encompass strategic organic functions, such as glycidoxypropyltrimethoxy silane (GPS silane), as well as fundamental functions for the acquisition of important new chemical properties, including 2. -

[methoxy (polyethylenoxy) propyl] trimethoxysilane (PEG10 silane), but is not limited to said silanes. The chemical structures of both chemical species can be seen in Fig. 5. GPS silane is used with intermediate silane.

 [076] Following external functionalization of the nanoparticles, the surfactant is removed by extraction with ethanolic HCl solution and sonication as discussed above.

 [077] In general and simplified, the process of producing the pegylated hydrophobic drug carrier nanoparticles can be seen in Figure 6. Note that initially there is the co-condensation of TEOS and hydrophobic silane around the organic mold. nucleation process with extra additions of TEOS and external functionalization with a silane of interest.

[078] Developed nanoparticles have potential application as carriers of antitumor hydrophobic drugs and, as such, must have the ability to trap a drug within them for later delivery to the cancer cell and colloidal stability in the blood to ensure biological target range. In this context, the importance of the presence of internal hydrophobic groups is noteworthy due to their hydrophobic character similar to the antitumor drugs that also have this characteristic, resulting in a strong interaction between Van der Waals and both, favoring their effective insertion. and storage within the nanostructure. PEG (hydrophilic) externally will contribute to colloidal stability for intravenous transport as blood has hydrophilic characteristics. In addition, polyethylene glycol also contributes to "softening" the protein / nanoparticle interaction, especially those involved in the recognition and withdrawal of supposedly foreign agents in the blood, as occurs in the immune system.

CONCRETIZATION EXAMPLES

 The lowest purity of the reagents cited within the scope of the invention was the analytical grade (P.A). In the case of organosilanes, phenyltrimethoxysilane - (PTES) Sigma Aldrich, 2-

[methoxy (polyethylenoxy) propyl] trimethoxysilane (short chain PEG silane) Gelest and 3-glycidoxypropyltrimethoxysilane (GPTMS) Sigma Aldrich, purity is equal to or greater than 98%. For PEG-COOH, 98% purity is also verified.

[080] Fourier transform infrared (FTIR) spectra were obtained on an ABB Bomem B-series model FTLA2000-102 spectrometer using the transmittance method in the range 4000-200 cm ^-1 with an accuracy of 4 cm ^-1 and 32 accumulations. Samples were diluted and compacted into potassium bromide pellets (KBr, analytical purity). The ²⁹ Si nuclear magnetic resonance (RN) spectra were made on a Bruker Avance 300 Hz HPDEC ( ²⁹ Si - → ¹ H) method with 1024 scans, acquisition time 0 , 0299 s, pulse interval of 60.0 s and frequencies of 59, 624 MHz on channel 1 ( ²⁹ Si) and 300, 130 MHz on channel 2 (¾). The analysis was performed with a 90 ° single pulse excitation of 5.0 με in channel 1 ( ²⁹ Si) followed by signal acquisition under a continuous uncoupling pulse in channel 2 ( ¹ H), initiated simultaneously with channel 1 pulse. , until the signal relaxation in that channel . ¹³ C NMR spectra were obtained with the cross-polarization and magic angle spinning (CPMAS) method of the neighboring IH nuclei (20480 scans), acquisition time 0.0499 s, pulse interval 3.0 s and frequencies 75.475 MHz on channel 1 ( ¹³ C) and 300, 131 MHz on channel 2 (¾) The analysis is performed with a 90 ° pulse excitation of 2.7 με on channel 2 ( ¹ H), followed by a pulse 4000 μ≤ on channel 1 ( ¹³ C), followed by signal acquisition Along with channel 1 ( ¹³ C) pulse, another pulse for channel 2 ( ^X H) decoupling is produced, which extends to relaxation signal on channel 1. the sample holder rotation speed was 10 kHz for both ^■ analysis (SiC), made under the magic angle (54.74 °). the thermogravimetric analyzes (TGA) were performed on a TA Instruments 500 thermal analyzer, TGA module 2050. Analyzes used approximately 5.0 mg of dry samples placed in a platinum crucible. The heating rate was maintained at 10.0 ° C / min and the air flow at 100 mL / min. The morphology of the nanostructures was analyzed by transmission electron microscopy in bright field mode (TEM, Zeiss Libra 120, operating at 80KV). Dynamic light scattering (DLS) to evaluate particle sizes and zeta potential (PZ) were obtained with the Malvern ZetaSizer-Nano instrument. DLS results were obtained using deionized water as dispersant (1.0 mg / mL). Nitrogen adsorption measurements were made at liquid nitrogen temperature on an ASAP 2020 Micromeritics analyzer. Samples were treated at 120 ° C for 12 hours prior to analysis. Surface area was calculated from the isotherm adsorption branch using the Brunauer-Emmett-Teller (BET) method. Pore diameter and volume were calculated from the isotherm adsorption branch using the Barrett-Joyner-Halenda (BJH) method and from the single-point adsorption amount (P / Po = -0.94), respectively. The stability evaluations of nanoparticle suspensions are made by light absorption analysis in the ultraviolet-visible range with a Shimadzu model UV-1650 PC spectrophotometer.

 Production of starting nanoparticles (step A)

[081] The production of nanoparticles with internal phenyl groups was performed in a sol gel type reaction. In this process, 0.75 g of CTAB was dissolved in 20 mL of 0.05 mol / L ammonia solution, followed by the addition of 3.2 mL of ethanol in this reaction medium. Said solution was stirred at 60 degrees Celsius. At the same time, a mixture of 1.49 mL of TEOS and 130 L of PTES was prepared and then poured into the system containing the organic mold. The ethanol co-solvent which favors the solubilization of the reagents is the catalyst (NH ₃ ). The solution was stirred for 120 min (from time t = 0) with two additions of 124 µl TEOS at t = 60 min and t = 90 min. After this step, CTAB-containing nanoparticles and phenyl groups were centrifuged and redispersed in ethanol. Approximately 1 mL of the suspension was placed in eppendorf which was evaporated in an oven at 60 ° C for 24 hours. The isolated material was characterized by TGA-DTA and IV.

[082] In order to evaluate the amount of CTAB in nanoparticles produced in step A and to obtain parameters for future calculations, the organic mold was removed, obtaining only the nanoparticle with phenyl groups inside, whose characterization was performed. using the same characterization techniques already described. The nanoparticles without the mold were subjected to Zeta potential analysis (PZ), whose objective was to determine the surface charge. Zeta potential measurements are important for confirming functionalization with PEG, as this process decreases the negative charges on the surface of the silica nanoparticle. Therefore, this characterization was performed before and after functionalization. Figure 7 summarizes the synthetic process of the starting nanoparticle, consisting of a silica nanocarrier containing internal phenyl groups, CTAB (represented by yellow circles) and free outer surface for surface modification.

 Functionalization of nanoparticles produced in step A with PEGio silane (short chain)

 [083] The CTAB-containing nanoparticles and phenyl groups produced in step A were subjected to the functionalization process with short-chain PEG silane, constituting step B described in this topic. In this process, 300 mg of the starting nanoparticle is dispersed in 40 mL of absolute ethanol for 60 minutes and then homogenized for a further 15 minutes at 60 ° C. Then PEGio silane is added based on the number of moles of silicon, ie, considering 300 mg of initial NP, the amount of functionalized organosilane corresponds to 10%, 25% or 50% mol-Si relative to number of moles of total silicon present in said nanoparticle mass. Each reaction was performed under constant stirring at 60 ° C for one hour, and an additional 12 hours of functionalization at room temperature. The products were centrifuged and dispersed in ethanol. Removal of the organic mold was performed according to the procedure described below.

[084] Samples were subjected to FTIR, TG-DTA, surface area, NMR, PZ and MET analyzes. It is noteworthy that the determination of the amount of silicon of the nanochargers obtained in step A was performed by thermogravimetry. Therefore, from the resulting mass, the relative amount of matter for silica and hence silicon is calculated. Figure 8 summarizes the process of external surface modification of the starting nanoparticles with PEGio silane. The figure facilitates understanding of the functionalized Si mol number concept. It is notorious that the values of 10, 25 and 50 mol% Si represent a percentage relative to. total number of silicon atoms present in the silica "shell" present prior to surface modification.

Functionalization of nanoparticles produced in step A with long chain PEG

 [085] GPS silane is the intermediate chemical species for obtaining long-chain PEG-containing nanoparticles. It is important to mention that in this reaction a long-chain PEG silane was not used due to the absence of this product for purchase on the market until mid-May 2013. Production of the surface glycoxy-oxide nanoparticles was performed using 300 mg nanoparticles. synthesized in step A dispersed in 62 mL of absolute ethanol for 30 minutes in the sonicator. Next, GPS silane (10 mol% silicon) is added to the flask containing the nanoparticles already suspended in the solvent, followed by stirring of the system for 12 hours at 85 ° C under nitrogen atmosphere. The products were centrifuged at 15000 rpm for one hour and dispersed in ethanol.

Coupling of PEG-COOH in the glycidoxy group-containing nanoparticle (NP-GPS) was performed followed by the molar ratio of PEG: glycidoxy = 1: 3. For this a 150 mg mass of NP-GPS was previously dispersed in 20 mL of ethanol through the sonicator for 30 minutes. In parallel, 15.1 mg of PEG-COOH was dissolved in 875 μ. of ethanol. In a round bottom flask containing the surface modified nanoparticles with glycidoxy groups, potassium hydroxide was added at the molar ratio KOH: glycidoxy = 1.6: 1. Said polymer previously Solubilized in absolute ethanol was added to the system which at 95 ° C was subjected to the coupling reaction for 12 hours under constant stirring. The products were centrifuged at 15000 rpm and dispersed in ethanol. Removal of the organic mold was performed according to the procedure described below.

 [087] Samples were subjected to FTIR, TG-DTA, surface area, NMR and PZ analysis.

 Figure 9 indicates the production of the silica nanoparticle holding PEG10 in a summarized manner. Note that the synthetic procedure of said molecular vehicle comprises two main steps characterized, first, by the functionalization of GPS silane, followed by the coupling reaction of PEG40 in the epoxy function (significantly reactive) of the mentioned organosilane.

Soft Template Extraction (CTAB)

 [089] As noted in previous topics, after complete obtaining of the nanoparticles the organic mold is still present, requiring a standard processol for all situations that favor the removal of the surfactant that once directed the shape acquired by the nanocarriers. In this context, the isolated products containing the organic mold were transferred to a flask containing an ethanolic HCl solution at a volumetric ratio of HCl: Ethanol = 1: 9 and ultrasounded for 10 minutes to remove the mold. The standard stoichiometric ratio was 5 mg of products: 1 mL of HCl / Ethanol solution.

[090] Figure 10 generally represents the removal process of the starting nanoparticle soft template equally applied to the other nanoparticles obtained in the present invention. Evaluation of colloidal stability of nanoparticles

 [091] The nanoparticles were subjected to the colloidal stability test in order to evaluate the influence of polyethylene glycol on these dispersion aspects. In this context, the colloids were dispersed in PBS (1x) at a concentration of 250 pg / mL and centrifuged at rotations of 0, 94, 2348, 9391 and 18407 rcf - the preparation of PBS is described below. The 1 mL volume of each supernatant was subjected to analysis by UV-Vis absorption electron spectroscopy and the absorbance at 263 nm wavelength measured.

Heraolitic Assays

 The nanoparticles produced were subjected to hemolytic assays.

 [093] Hemolytic tests were performed using Type B blood from the Hemocenter of the State University of Campinas to the Solid State Chemistry Laboratory of the Institute of Chemistry. Concomitantly two PBS solutions were prepared according to the following procedure:

 a) PBS (2x): Two tablets of saline phosphate buffer were diluted in 200 mL of deionized water (2 mM phosphate, 0.54 mM KCl and 27.4 mM NaCl);

 b) PBS (1x): 100 mL of the above solution was diluted in 100 mL of deionized water (1 mM phosphate, 0.27 mM KCl and 13.7 mM NaCl).

The volume of 10 mL of blood was centrifuged at 14,000 rpm at 4 ° C for 10 minutes. The supernatant was discarded and 20 mL of PBS (2x) was added to the pellet which was subsequently resuspended. After this process, two new washes were performed following the current procedure, obtaining 20 mL of a red blood cell solution in PBS (2x). Of this total, 2.5 mL were diluted in 12.5 mL of PBS (2x) to obtain the red cell stock solution.

 Nanoparticle pretreatment (initially in ethanol) for hemolytic assays comprises a procedure in which all are centrifuged at 15,000 rpm at 4 ° C for 30 min and dispersed in deionized water. This process is repeated twice more to obtain final colloidal dispersions in deionized water at concentrations of 1 mg / mL.

 The influence of nanoparticle aggregation state as a function of time in hemolysis

 [096] Performing hemolytic assays requires a prior assessment of the aggregation behavior of each nanoparticle in PBS for one hour. Thus, 25 pL of each molecular carrier was added, respectively, to a mixture of 25 pL PBS (2x) and 950 pL PBS (1x). The dynamic light scattering technique intermittently evaluated the particle size variation and polydispersity index within one hour, similar to the standard red cell incubation time. Colloidal behavior was evaluated at 0, 5, 15, 30, 45 and 60 minutes.

 Evaluated the above physicochemical aspects, 25 µl of each nanoparticle was added in eppendorf containing a mixture of 25 µl PBS (2x) and 850 µl PBS (1x). The nanoparticles were kept in the respective vials for one hour. After this step, 100 pL of red blood cell stock solution was added while maintaining one hour of incubation.

[098] The above experiment was again performed maintaining all experimental conditions except for the interaction time between the nanoparticles and the PBS mixture (prior to the addition of the red cell stock solution), which in this case was one minute. After the interaction period between nanoparticles and red blood cells the systems are centrifuged at 14,000 rpm at 4 ° C for 10 minutes and 100 æl of the supernatant is subjected to UV-Vis absorption wavelength electron spectroscopy analysis. 540 nm. The absorbance values are reference for the percentage calculations of the hemolytic rate.

 [0100] As for the statistical aspect in the reliable and accurate interpretation of the results, all experiments were performed in triplicate.

Results and discussion

 In the following topics certain designations have been established for each type of nanoparticle as explained below:

a) N: nanoparticle;

b) PH30: 30% of theoretically functionalized phenyl groups;

c) SiOH: free surface silanol groups for functionalization;

d) xPEG10: x% of theoretically functionalized short chain PEG groups (n = 10);

e) xGPS: x% of theoretically functionalized glycidoxy groups; f) XGPS-PEG40: x% of theoretically functionalized large chain PEG-coupled glycidoxy groups (n = 40);

g) NPH30-CTAB-SiOH: starting nanoparticle containing 30% theoretically functionalized phenyl groups on the inner surface, further containing the surfactant;

h) NPH30-SiOH: starting nanoparticle containing 30% of theoretically functionalized phenyl groups on the inner surface with surfactant already removed; (i) NPH30-C AB-SiO-xPEG10: nanoparticles containing 30% of theoretically functionalized phenyl groups in the presence of CTAB and x% of theoretically functionalized PEGio groups;

j) NPH30-SiO-xPEGio: nanoparticles containing 30% of theoretically functionalized phenyl groups in the absence of CTAB and x% of theoretically functionalized PEGio groups;

k) NPH30-CTAB-SiO-xGPS: nanoparticle containing 30% of theoretically functionalized phenyl groups in the presence of CTAB and x% of theoretically functionalized glycidoxy groups;

 1) NPH30-SiO-xGPS: nanoparticle containing 30% of theoretically functionalized phenyl groups in the absence of CTAB and x% of theoretically functionalized glycidoxy groups;

m) NPH30-CTAB-SiO-xGPS-PEG / io: nanoparticle containing 30% of theoretically functionalized phenyl groups in the presence of CTAB and x% of theoretically functionalized PEG40-coupled glycidoxy groups; n) NPH30-SiO-xGPS-PEG40: nanoparticle containing 30% of theoretically functionalized phenyl groups in the absence of CTAB and x% of theoretically functionalized PEG40-coupled glycidoxy groups.

 All designations prior to the term SiO (representing the "shell" of the nanoparticle), such as PH30-CTAB-, PH30-, refer to the chemical entities present within the nanoparticle whereas names written after the SiO nomenclature, such as xPEGio, xGPS and xGPS-PEG-jo, come from the chemical species present on the outer surface.

The following is a picture of all the embodiments performed in this invention. Figure 11 shows the "shell" of internally functionalized silica with phenyl groups and externally with glycidoxy (GPS), open-ring glycidoxy (Open GPS), PEG40-coupled glycidoxy (GPS-PEG-jo) and PEGio functions. The numbers and terms written in blue and red, respectively, will be the basis for the interpretation of the carbon and silicon NMR spectra presented in the invention. Note that the functionalizations shown in the figure below are not necessarily present in the same nanoparticle, showing only the types of surface modifications possible on the inner surfaces and those occurring only on the outer surface. As an example, note that, although present on the same surface, GPS silane and PEGio silane are not concomitantly present on the same nanoparticle because they encompass different molecular vehicle development contexts.

Starting nanoparticles

[0104] This topic addresses the results related to nanoparticles containing functionalized phenyl groups on the inner surface and in the absence of any outer surface modification. Such materials are the main starting products for any step and deserve a unique approach. Figure 12 shows the infrared spectra of the starting nanoparticles before and after removal of the organic mold. The main silica bands are observed at 1072 cm ^-1 (asymmetric Si-O-Si stretch), 800 cm ^-1 (symmetrical Si-O-Si stretch) and 962 cm ^-1 (Si-OH stretch), 464 cm ^{- 1} (Si-O-Si angular deformation). Confirmation of mold extraction can be obtained by observing the absence of typical CTAB bands in the NPH30-SiOH spectrum. The soft template bands can be observed at: 2926 cm ^-1 (asymmetric CH stretch in CTAB carbon chain CH2 groups), 2854 cm ^-1 (symmetrical CH stretch) and 1232 cm ^-1 (CN stretch). that the visualization of the latter band is made difficult by the overlap with the bands from the asymmetrical stretches of the SiC (Si-O-Si) tetrahedral units by approximately 1200 cm ^-1 and 1130 cm " ¹ .- Analyzing both spectra, it is noticeable the absence of these bands after the removal of the soft template, characterizing a significant parameter for the follow-up of the organic mold extraction reactions presented in the next topics. Bands at 1596 cm ^-1 , 1431 cm ^-1 (both derived from the aromatic ring stretching), 739 cm ^-1 and 698 cm ^-1 from the angular deformation of the aromatic ring functionalized internally in the nanoparticle. The above discussions will be important for the characterization analysis of surface modified nanoparticles, as the presence of silica bands and aromatic rings should be noted in all situations. The absence and presence of CTAB bands will be essential for monitoring their extraction in the later stages of the process.

[0105] Figure 13 shows the thermogravimetric (TG) curves and their respective differential thermal analysis (DTA) curves for the starting nanoparticles before and after removal of the organic mold. The NPH30-SiOH DTA curve exhibits a major event at 577 ° C, suggesting the thermal decomposition of the phenyl groups and a minor one occurring at 308 ° C from residual CTAB release. Disregarding the discrete event, which in this case is not significant in relation to the total percentage of the sample, there is only the decomposition of functionalized aromatic species beginning and ending at 324 ° C and 716 ° C, respectively. Considering that the final sample consists only of silica (SiO2) and, in the graph, the final mass is 73%, it is known that the percentage of phenyl is slightly less than 27% by mass. In the case of CTAB-SiOH-NPH30 in which CTAB is present, the two DTA recorded significant events relating to the decomposition of the mold ^"organic occurring at 308 ° C and weight loss of the phenyl groups at 577 ° C. In the TG curve of the latter molecular vehicle CTAB decomposition begins and ends at 135 ° C and 381 ° C, respectively. It should be noted that the onset of phenyl group decomposition (approximately 324 ° C) occurs parallel to the end of surfactant release, making it impossible to calculate precisely the percentage of both components in the sample. However, it is possible to confirm the extraction of the commented macromolecule by comparing these data with those of the vibrational spectrum.

[0106] Figure 14 presents the results of NPH30-SiOH nitrogen adsorption-desorption analysis as well as NPH3G-CTAB-SiOH. For the isotherm of this last nanoparticle, its profile did not show a significantly defined pattern, resembling only the type IV isotherm according to the IUPAC classification for porous materials. Type IV isotherms are related to mesoporous materials. A more detailed analysis concludes that the desorption curve does not coincide with the adsorption curve as P / Po tends to 0, that is, the observed hysteresis cycle is not closed within the analyzed range. This phenomenon is possibly linked to the presence of CTAB which, together with the range of possibly interconnected pores, makes gas release difficult, making adsorption a slightly irreversible process, which is expected considering the complexity of said hybrid system. The pore distribution of the starting molecular carrier holding the amphiphilic macromolecule exhibits a wide variety of pores, comprising a material of complex porous nature due mainly to the high content of internally functionalized phenyl groups in the nanoparticle. Such an effect generates increased internal volume due to the arrangement of the aromatic rings between the surfactant tails. Consequently, in the nucleation stage during the nanoparticle production, the inorganic lattice deposited around the CTAB engenders pores of varying sizes, characterizing the desired uneven surface. According to the pore distribution graph, whose calculations are based on the Barret, Joyner and Halenda model (BJH method), the predominance of pores between 4 nm and 10 nm is observed. Mesoporous materials have pore sizes ranging from 2 to 50 nm), which classifies said material as mesoporous. Analyzing the nanoparticle nitrogen isotherm profile after CTAB extraction, it is clear that the profile variation is directly related to the type IV isotherm. In this context, there is a slight hysteresis similar to that classified as type H3 according to IUPAC. This result indicates a complex mesoporous structure according to the isotherm profile itself due to the presence of the phenyl groups, which help in the formation of the complex porous structure. The pore distribution graph according to the BJH method indicates greater pore predominance in a size range between 2 nm and 10 nm, reflecting CTAB extraction. The removal of said organic mold can be confirmed by analyzing the surface area variation calculated by the method proposed by Brunauer, Emmet-Teller (BET method) before and after this process. For NPH30-CTAB-SiOH and NPH30-SiOH the surface area ranged from 548 m ² / g to 898 m2 / g, respectively. Complementing this information, the total pore volume of both nanoparticles was 0.96 mL / g and 2.2 mL / g, respectively.

[0107] This result set confirms CTAB extraction since there was an increase in total pore volume and surface area after surfactant extraction. The results also corroborate to obtain a nanoparticle with mesostructure with porous complexity reflected in the size diversification and, possibly, the possibility of wide interconnection between the cavities.

 Figure 15 shows the micrograph of the NPH30-SiOH in which disordered porous arrangements can be observed featuring an irregular topography. The presence of .30% mol-Si of phenyl groups within the nanoparticles provides an increase in the internal cavity, generating this characteristic on the outer surface not yet coated with PEG. When drying the sample for electron microscopy, it is impossible to avoid colloid aggregation, as shown in Figure 10. In solution, however, the nanoparticles show apparent colloidal stability, a feature that will be discussed quantitatively below with the starting and functionalized nanoparticles. polyethylene glycol. According to Figure 10 it is possible to estimate the average colloidal diameter around 65 nm.

 NPH30-SiO-10PEGio Surface-Modified Short-Chain Polyethylene Glycol Nanoparticles

[0109]. The infrared vibrational spectra of the NPH30-SiOPLOG shown in Figure 16 show the characteristic bands of silica and aromatic rings in all situations. Also shown in the Figure is the spectrum of PEGio silane used in functionalization. The main polymer bands can be observed at 1080 cm ^-1 (asymmetric COC stretch, similar to Si-O-Si in silica), 1040 cm ^-1 (symmetrical COC stretch, observed as a shoulder), 2867 cm ^-1 (stretch CH in ethylene glycol groups C¾) and 1460 cm ^-1 (angular deformation CH in ethylene glycol CH2 groups). After functionalization NPH30-CTAB-SiOH (on the spectrum denoted as P ^ C-SiOH) with said organosilane product ^'is generated NPH30-CTAB-SiO-10EEGio. (PC-lOPEGio). In this last spectrum the visualization of the polymer bands is difficult due to the overlap with the silica bands (at 1100 cm " ¹ and 962 cm ^-1 ) and the CTAB (at 2854 cm ^-1 ). Even after the removal of the mold, whose absence of its typical bands is noticeable in the spectrum of the final nanoparticle NPH30-SiO-10PEGio (P-lOPEGio), the visualization of the PEG bands is impaired as they overlap with the silica bands, requiring the use of other characterization techniques to confirm the functionalization.

 [0110] Through thermal analysis (Figure 17) it was possible to confirm the presence of the polymer in the nanoparticle, although the confirmation of the condensation of PEG silane on the colloid surface cannot yet be confirmed using this technique alone. DTA curves exhibit a more significant event occurring at 292 ° C for the NPH30-CTAB-SiO-10PEGio (P-C-SiO-10PEGio) sample relative to the starting nanoparticle, showing the presence of the outer macromolecule. At this temperature an additional mass loss step is observed on the P-C-SiO-10PEG10 TG curves. After removal of the mold, the decomposition of polyethylene glycol is observed at 216 ° C, ending at 375 ° C on the NPH30-SiO-10PEG10 TG curves. As noted, the release of aromatic groups at 324 ° C occurs in parallel with the elimination of the polymer; NPH30-SiO-10PEGio DTA curve shows a less evident temperature difference near 300 ° C compared to NPH30-CTAB-SiO-lOPEGio, since the final nanoparticle does not contain CTAB whose degradation already occurs at that temperature, contributing to the decrease in peak intensity observed.

The nitrogen adsorption-desorption analyzes shown in Figure 18 indicated a nanoparticle profile similar to the starting nanoparticles, allowing to state that after functionalization the structures did not change pore profile as desired. Specifically, the isotherm of Nitrogen resembles a type IV isotherm according to the IUPAC classification, ie nanocarriers are mesoporous in nature.

 According to the pore size distribution in the system in question, there is a predominance of pores having diameters between 2 and 10 nm, similar to the result for the starting nanoparticles.

According to the BET method, the surface area of the starting nanoparticle is 898 m ² / g. After functionalization, said value undergoes slight increase to 943 m ² / g, indicating little ^{'differences.} For the total pore volume determined by the BJH method, the starting nanocarriers have 2.2 cmVg as the NPH30-SiO-10PEG ₁₀ holds 1.48 cm ³ / g. This difference is possibly linked to the presence of PEGio silane, possibly characterizing steric effects against nitrogen gas adsorption in the nanoparticle.

In Figure 24 the 13 C and ²⁹ Si NMR spectra of NPH30-SiO-10PEG10 are shown. Using Figure 11 as a reference for the location of each nucleus one should disregard the GPS, open GPS and GPS-PEG40 groups because they are not present in the current nanoparticle.

Initially analyzing the NMR spectrum shown in Figure 14, one has the molecular carrier before and after surface modification. Prior to grafting, the only carbon nuclei present come from the phenyl group represented by nuclei 1 (130.5 ppm), 2 (129.7 ppm), 3 (128.4 ppm) and 4 (133.8 ppm) observed. in both situations. Peaks at approximately 30 ppm and 17 ppm can be attributed to the residual CTAB carbon nuclei and therefore can be neglected. The presence of polyethylene glycol in 10 monomer units is confirmed in the NPH30-SiO-10PEG10 spectrum whose nuclei 7, 8 and 9 appear at 72 ppm, 60.8 ppm and 60.8 ppm, respectively, after operation. However, the peak area of the main carbon atoms of PEG is low, indicating that the functionalization of 10% mol Si is not yet satisfactory, and it is necessary to study the surface modification with 25% mol Si and 50% molar. mol Si.

 Although the presence of said polymer is evident even in small quantities, confirmation of silane condensation can be completed by analyzing the silicon NMR spectrum.

 [0117] In the study of silicon NMR, it is necessary to understand the terms used for each type of nucleus according to its coupling with neighboring nuclei - Figure 11 helps in this understanding. The designations and their meanings are as follows:

Q ⁴ : Silicon core bonded with 4 substituents OSi, Si (O-Si) 4, located mainly within the "shell" of the silica;

Q ³ : refers to the silicon core bonded to 3 O-Si substituents and an OH, HO-Si (O-Si) 3 group present on the inner and outer surfaces of the molecular carrier, as well as within the silica layer - used for confirmation of condensation of silanes; Q ² : although less, it is the silicon atom bonded to 2 O-Si groups and 2 OH, (HO) 2Si (O-Si) 2 groups, arranged spatially in the same way as Q ³ nuclei;

T ³ : These are silicon cores bonded to 3 O-Si substituents and a carbon substituent contained in the group R, R-Si (O-Si) 3, present on the inner surface, inside or on the shell. molecular carrier; T ² : silicon nuclei attached to 2 O-Si substituents, one OH group and one carbon substituent contained in one group, R-Si (0-Si) 20H, present on the inner surface, inside or on the surface. "shell" molecular vehicle.

 It is important to note that the designation R refers to the phenyl, glycidoxy or PEG10 groups present in the nanoparticles.

[0119] Figure 25 shows NMR spectra of the NPH30-SiO-1OPEGio sample, before and after functionalization, containing the major silicon sites and their corresponding area in percent. The chemical displacements of the silicon peaks have specific values and do not change after functionalization, since the chemical entity that represents them does not undergo structural changes. In fact, what is observed are variations in peak area before and after grafting. Silicon NMR peaks at -110 ppm, -100 ppm and -90 ppm are clear for Q ⁴ , Q ³ and Q ^{2 sites} , respectively, -80 ppm and -70 ppm from T ³ and T ^{2 sites} at this time. order.

In surface modification, the OH groups from the Q ³ and Q ^{2 sites} undergo nucleophilic substitution with ossilanes, generating new T ³ and T ^{2 sites} . In this context, the area ratio (Q ⁴ + Q + Q ² ) / (T ³ + T ² ) should decrease after functionalization, as the sum of T ³ + T ² increases. Analyzing the graph gives (Q "+ Q + Q ² ) / (T ³ + T ² ) SiOH = 3.61 and (Q ⁴ + Q + Q ² ) / (T ³ + T ² ) PEGIO = 3.39, indicating the expected result, so the condensation of the PEGio functional groups can thus be confirmed.

However, as noted above, it is necessary to increase the amount of PEGio silane in surface modification in order to improve its degree of condensation on the external surface which has so far been found unsatisfactory in due to the small increase in the main carbon peaks of polyethylene glycol in carbon NMR.

 NPH30 ~ SiO-25PEGio

The nanoparticle containing internal phenyl (30% mol-Si) groups as well as externally functionalized 25% mol-Si PEG10 groups was called NPH30-SiO-25PEG10. Their respective structural characterizations showed satisfactory results according to the pre-established expectations for the construction of the said molecular vehicle. PC-SiOH subtitles, PC-P and 25PEGio 25PEGio refer respectively to NPH30-SiOH, NHPH30 CTAB--SiO-SiO-25PEGio and NPH30 _25PEGi-0.

[0123] Figure 26 shows the infrared spectrum of NPH30-SiO-25PEGio- In all situations shown in the figure the major silica bands are noticeable. This result evidences the presence of this main chemical entity that constitutes the fundamental structural unit of the nanoparticle. In the case of phenyl groups, their presence is also easily confirmed before and after functionalization and in the presence and absence of organic mold. Keeping the focus on polyethylene glycol, it is worth noting again the difficulty associated with observing the polymer bands overlapped by the silica bands. In the spectrum of NPH30-SiO-25PEGio, a small band widening close to 1040 cm ^-1 is observed in relation to the spectrum of NPH30-SiOH, supposedly evidencing the presence of PEG in the nanoparticle. As previously mentioned, in this region there are bands from the symmetrical COC stretches of the polymeric entity. overlapped by the silica bands from the asymmetrical Si-O-Si stretch. Knowing these difficulties, it is necessary to analyze the results obtained concurrently with other characterization techniques. [0124] The thermal analysis data shown in Figure 27, are important for elucidation of ^'distinct chemical species present in the nanostructured system. According to the DTA curves, note ^are a typical event occurring at 577 ° C also present in all steps linked to the thermal decomposition of the phenyl groups. Regarding the 25% mol-Si functionalized silica nanoparticle PEG silane in the presence of the organic mold, one is noted. event is 274 ° C referring to the decomposition of CTAB together with the polymeric entity cited. After removal of the soft template, obtaining the final nanoparticle, the event mentioned shifts to 224 ° C, due to the thermal decomposition only of polyethylene glycol, which occurs in a smaller temperature range compared to the event involving the simultaneous presence. PEG and CTAB. This set of results is important to indicate the presence of the three organic species in NPH30-SiO-25PEGio, mainly by observing the displacement of decomposition events in the presence or absence of certain compounds.

The thermogravimetric curves also presented in Figure 17 complement the above information regarding the chemical entities present in the molecular vehicle. Comparing the curve of NPH30-CTAB-SiOH with NPH30-CTAB-SiO-25PEG10, a new decomposition plateau is observed due to the PEG present. Comparing the curves of NPH30-SiOH and NPH30-SiO-25PEGio, both exempt from CTAB, it is evident that the thermal decomposition of polyethylene glycol between 217 ° C and 377 ° C is the only possible organic entity in this range. temperature, as CTAB is absent and, as already mentioned, the phenyl group decomposes from 324 ° C. In fact, it becomes complicated to estimate quantitatively the individual percentage of each chemical species commented, once in the temperature range between 324 ° C and 381 ° C CTAB, PEG and phenyl mass loss events occur simultaneously.

Following are shown in Figure 28 the nitrogen adsorption isotherms and pore distribution of NPH30-SiO-25PEG10. The graph of pore volume as a function of relative pressure shows the maintenance of the functionalized nanoparticle profile in relation to the starting one, since it has the same type IV isotherm profile with slight type H3 hysteresis. This result reflects the mesoporous and complex nature of randomly arranged pores, that is, without definite ordering. Moreover, it can be stated that the functionalization did not modify the nanocarriers in these aspects, which is significantly important from a structural point of view. Analyzing the surface areas of the nanoparticles before and after functionalization estimated by the BET method, the increase from 898 m ² / g to 988 m ² / g before and after surface modification, respectively, is observed.

In terms of pore distribution, functionalized nanoparticles have a similar profile to the starting nanoparticles similar to what occurs in nitrogen isotherms. Figure 18 shows the predominance of pores between 2 and 10 nm in both nanocarriers, classifying NPH30-SiO-25PEGio as a mesoporous material. Pore volumes indicated phenomenologically interesting values, that is, after functionalization the total pore volume ranges from 2.2 cm ³ / g to 1.74 cmVg. This result infers the possible spherical impedance of the pores by the superficially functionalized silanes, preventing the entry of gases inside the nanoparticle.

The following are the carbon and silicon NMR spectra of NPH30-SiO-25PEG10. As already mentioned previously, based on Figure 11 for understanding, GPS and GPS-PECio species should be disregarded, since they are not involved in the context of this nanoparticle.

Figure 29 shows the ¹³ C NMR spectrum of the nanoparticle in which nuclei from the phenyl groups at 130.4 ppm (nucleus 1), 129 ppm (nucleus 2), 126.8 ppm (nucleus 3) are readily identified. ) and 134.1 ppm (core 4). It should be noted that said nuclear species are present before and after functionalization as expected. Regarding the identification of polyethylene glycol in the molecular carrier, one should identify the nucleus 7 peak of the polymer at 70.5 ppm in the NPH30-SiO-25PEGio spectrum. Its absence in the starting nanoparticle spectrum is noticeable; It is inferred that after surface modification PEG is present in the system due to the appearance of the peak at 70.5 ppm in the modified molecular vehicle. However, the information contained in ¹³ C NMR does not confirm the functionalization of PEG10 silane, but only indicates its presence. To complement this information it is necessary to analyze the 29Si NMR spectrum of the NPH30-SiO-25PEGio shown in Figure 20.

According to the silicon NMR spectrum of NPH30-SiO-25PEG10, the decrease of the sum of Q areas and increase of T areas after surface modification is clearly observed. This result indicates that the number of Q ³ and Q ^{2 sites} linked to the (S1O2) 3-Si-OH and (SiO2) 2-Si- (OH) 2 structures, respectively, decreases, which is expected since in Functionalization the silanols groups react covalently with the PEG silane providing the increase of species (Si02) 4-Si in the structure. Similarly, the spectrum indicates that the number of T ³ and T ^{2 sites} linked to the (Si02) 3 -Si R and (S1O2) 2-Si- (OH) (R) structures increases, since in the organosilane condensation has if the formation of three or two siloxane bridges Si-O-Si and the presence of covalent bond between a silicon and carbon core intrinsically linked to silane. Complementarily, there was a decrease from 3.61 to 3.21 in the (Q + Q ³ + Q ² ) / (T ³ + T ² ) ratio, confirming the above observation quantitatively. Accordingly, the functionalization of PEG10 silane in said molecular carrier can be confirmed. NPH30-SiO-50Pegio

[0131] The nanoparticle containing internal phenyl (30% mol-Si) groups as well as externally functionalized 50% mol-Si PEGió groups was named NPH30-SiO-50PEGio ■ Their respective structural characterizations showed satisfactory results according to pre-established expectations for the construction of said molecular vehicle. Subtitles PC-SiOH, PC-50PEG10 and P-50PEG10 refer to NPH30-SiOH, NHPH30-CTAB-SiO-50PEG10 and NPH30-SiO-50PEG10. Initiating such studies by infrared vibrational spectroscopy whose spectrum is observed in Figure 21, the main silica bands were observed in all the synthetic steps presented. In this context, there are bands at 1082 cm ^-1 (asymmetric Si-O-Si stretch), 800 cm ^-1 (symmetrical Si-O-Si stretch), 962 cm ^-1 (Si-OH stretch) and 462 cm ^{- 1} (Si-O-Si angular deformation) confirming the presence of the inorganic matrix CTAB extraction control can be monitored by noting their respective bands at 2924 cm ^-1 (asymmetric CH stretch) and 2852 cm " ¹ (CH stretch from the carbon chain of said surfactant, as well as those present at 1483 cm ^-1 (asymmetric CH stretch of the H3C-N + group CH3) and 1232 cm ^-1 (CN stretch) Error! Indicator not defined in all situations shown in Figure 21, except for P ~ 5OPEG10 where they are absent and therefore indicating removal of organic mold. In the case of phenyl groups, their respective Bands are confirmed at 1596 cm ^-1 (C = C stretch), 740 cm ^-1 (CH angular deformation) and 698 cm ^-1 (C = C angular deformation).

[0132] Confirmation of polyethylene glycol by infrared spectroscopy on the nanoparticle is difficult mainly due to the overlap of its main bands with those from silica. According to Figure 21, after functionalization there is a slight widening of the band present at 1100 cm ^-1 , partially indicating the presence of one of the main PEG bands at 1080 cm ^-1 associated with the asymmetric COC stretch. This polymeric entity was analyzed by thermal analysis and NMR aiming at its confirmation in the nanoparticle.

Figure 22 shows the TG and DTA curves of NPH30-SiO-50PEG10. Differential thermal analysis records an exothermic event at 267 ° C for NPH30-CTAB-SiO-50PEGio, indicating the thermal decomposition of surfactant and polyethylene glycol. After removal of the organic mold, the event is shifted to 221.7 ° C, associated only with the presence of the polymeric species in the absence of the soft template in this situation. From these considerations, observing the thermogravimetric curve of the NPH30-CTAB-SiOH allows visualization of CTAB mass loss starting at 135 ° C and ending at 381 ° C (22.2% mass). During this process the onset of PEG decomposition occurring within the temperature range of 216 ° C to 378 ° C (observed on the curve of NPH30-SiO-50PEG10) is noticeable. Estimation of the functionalization content of PEG silane in the nanoparticle can be obtained by comparing the curves of NPH30-SiOH and NPH30-SiO-50PEGio at a temperature of 900 ° C, in which all organic components are absent. In this situation the final mass for each of them is 73% and 67%, respectively, ie the 6% difference between them is linked to the polyethylene glycol mass. Through stoichiometric calculations it was can confirm the functionalization at the content of 1.17. 10 ^-7 mol Si-PEG / mg give nanoparticle or 1.06 mol% Si. In the region between 324 ° C and 716 ° C occurs the release of the phenyl group whose mass amount is 22.7% or 2.95. 10 ^-6 mol Si-phenyl / mg nanoparticle, corresponding to 28.7 mol% Yes. Note that the theoretical amount of functionalization of said aromatic groups is 30 mol-Si, corroborating the experimental results.

 The functionalization rates of the phenyl and PEG groups are consistent with the expected values. The 28.7% mol Siphenyl graft is considerably close to the theoretical result, which is not the case with the polymeric entity whose 1.06% functionalization value, which is different from the theoretical value (50%), linked to possible effects. between the carbonic chains of the polymer which, in this system, interfere with the low reaction yield. On the other hand, said functionalization content was sufficient to generate new chemical properties to the nanoparticle, such as improved colloidal stability and influence on hemolysis, topics discussed below.

The NPH30-SiO-50PEGio was subjected to nitrogen adsorption-desorption analysis whose isotherm is shown in Figure 23. By analyzing the pattern, its nitrogen isotherm profile resembles that of the so-called nitrogen isotherm profile. type IV with slight hysteresis of type H3, according to the IUPAC classification in the context of porous materials. The observed pattern reflects a mesoporous material of complex porosity indicating a marked level of surface irregularities as the production of nahoparticles holding irregular surfaces. The formation of a slight hysteresis at P / Po = 0.40, typical of type IV isotherms for mesoporous materials, is notorious. Comparing the starting carrier with the final nanoparticle functionalized externally, a decrease in surface area (BET method) is observed from 898 m ² / g to 771 m ² / g, respectively. Such result suggests the possible steric hindrance. pores by externally condensed silanes and thus slightly preventing the adsorption of nitrogen gases in their entirety. This phenomenon is most evident when comparing the total pore volume of both nanoparticles whose calculated values were 2.2 mL / g and 1.3 mL / g for NPH30-SiOH and NPH30-SiO-50PEGio, respectively. Since thermal analysis and infrared spectra confirmed CTAB extraction, the expected result would be relatively similar surface area and pore volume between both nanoparticles, which is not the case. Fortunately, such a result complements the confirmation of external functionalization. According to the pore distribution graph, also in Figure 23, there is a predominance of pores between 2 nm and 10 nm for NPH30-SiO-50PEGio characterizing it as a mesoporous material.

Nuclear Magnetic Resonance techniques are essential tools in the structural elucidation of organic and inorganic compounds. In this sense, it becomes necessary to complement the information obtained through infrared and thermal analysis techniques, as analyzed above. From the ¹³ C NMR of NPH30-SiO-50PEG10 Figure 24 shows the main peaks of the aromatic ring nuclei denominated 1 to 4 in chemical shifts of 130 ppm, 128.9 ppm, 127.4 ppm and 134.0 ppmError! Indicator not defined., Respectively, present at all stages as expected. The presence of PEG after functionalization can be confirmed by observing its main peaks at 28.0 ppm (carbon 6), 29.8 ppm (carbon 5), 59.8 ppm (carbons 8 and 9) and 70.4 ppm. (carbon 7). The latter is related to the main carbon nucleus by virtue of its more evident characteristic and its chemical displacement value significantly different from any other nucleus in the spectrum, facilitating its visualization and interpretation.

The ¹³ C NMR spectrum allows the conclusion of the polyethylene glycol in the molecular carrier. However, confirmation of the condensation of phenyl group and PEG containing silanes requires the ²⁹ Si NMR technique. Figure 30 shows the results of said technique in the context of NPH30-SiOH and NPH30-SiO-50PEG10 (before and after external surface modification). In both cases the major silicon sites are observed at -109 ppm (Q4), -100 ppm (Q3), -92 ppm (Q2), -77 ppm (T3) and -68 ppm (T2) Error! Indicator not set. . In the case of NPH30-SiOH, the condensation of phenyl group-containing silanes on the inner surface of the nanoparticle can be confirmed by visualizing peaks T3 and T2, related to the silicon species (Si-O) 3-Si-R and (Si -O) 2-Si (OH) -R, respectively, attributed exclusively to the only functional group in that situation. The sum of areas T3 and T2, 21.7%, correlates with the percentage of mol of silane containing phenyl group, differing by 7% from that value found by TG. However, the accuracy of the NMR technique makes its result very close to reality.

After functionalization, the decrease of the summed Q3 and Q2 areas as well as the slight increase of the T3 and T2 areas, confirming the condensation of PEG silane on the outer surface, becomes evident. Analyzing the behavior of the sites before and after the functionalization, we have that the ratio (Q ⁴ + Q ³ + Q ² ) / (T ³ + T ² ) _Si OH = 3.61 decreases slightly given by ( Q ⁴ + Q ³ + Q ² ) / (T ³ + T ² ) SOPEGIO = 3.39. Estimation of the mol number of condensed PEG silane silicon is not efficient via NMR since the functionalization content calculated by thermogravimetry (1.06%) fits the experimental error of the resonance technique (1%).

Transmission electron microscopy of the NPH30-SiO-50PEG10 whose image can be seen in Figure 31 shows the morphology obtained after functionalization with polyethylene glycol. As expected, uniformly spherical and porous colloids with an average diameter of approximately 63 nm are observed, similar to the value found for nanoparticles prior to surface modification. This result indicates that the presence of the polymer grafted on the surface does not interfere in their spatial dimensions ^■ possibly due to the shorter length of the polymer chain functionalization and low rate (1%), thus maintaining the unique structural aspects of starting nanoparticle. This result is interesting from the point of view of the interaction of nanostructures with biosystems together with the colloidal stability interface, that is, after the surface modification process it is important to maintain the structural integrity of the nanotechnological object.

 Surface-Modified Long Chain Polyethylene Glycol Nanoparticles

[0140] In this topic we will discuss the results obtained in obtaining the surface-modified nanoparticle with long chain polyethylene glycol. Initiating the structural characterizations, we initially have the infrared vibrational spectrum of the NPH30 ~ SiO-10GPS-PEG40 shown in Figure 32. It is observed that in all the steps the CTAB was extracted, a characteristic evident when observing the absence of your typical bands as commented in the previous topics. The spectra show that bands from silica and phenyl groups are present in all situations as expected. At the nanocarrier containing long chain polyethylene glycol should be focused initially on the glycidoxy group. Their spectrum shown at the top of the figure records the following bands that, at first, should be noted after their external functionalization: 2943 cm ^-1 (asymmetric CH CH2 stretch), 2839 cm- ¹ (CH2 CH symmetrical stretch), 1076 cm ^-1 (asymmetric COC stretch), 815 cm-1 (symmetrical COC stretch), 1253 cm-1 (asymmetrical stretches on the epoxy ring) and 910 cm-1 (symmetrical stretches on the epoxy ring).

[0141] The nanoparticle functionalized with the GPS group showed barely noticeable changes in the profile of its spectrum. An enlargement of the bands close to 1250 cm ^-1 and 1070 cm ^-1 , typical of the. asymmetric stretches in the epoxy ring and asymmetric COC stretches, respectively. The changes are clearer compared to the spectrum of the NPH30-SiO-10PEGio, in which the bandwidth underwent slightly pronounced variations. After PEG coupling in the three-membered ring, an increase in bandwidth is observed in the region of 1100-1070 cm ^-1, where we observed the asymmetric COC stretching of polyethylene glycol. Although there is evidence of the functionalization of the glycidoxy group and the coupling of PEG in the molecular vehicle, it is necessary to use techniques such as thermal analysis and nuclear magnetic resonance to reach more robust conclusions.

Figure 33 shows the TG and DTA curves of NPH30-SiO-10GPS-PEG40 showing the complexity of events due to the presence of an intermediate component, the glycidoxy group, whose function is to condense on the external surface of the silica and act as a support for inserting polyethylene glycol in this region. The starting nanoparticle TG and DTA curves present the release events of surfactants and phenyl groups in this order of occurrence, as shown in discussed in item 3.1. The NPH30-CTAB-SIO-1 OGPS DTA peaks show a significant event at 300 ° C related to simultaneous release of CTAB and GPS. In the respective TG curves the beginning and end of the glycidoxy function decomposition are observed at 281 ° C and 329 ° C, respectively. After coupling PEG into the system, a change in the molecular vehicle TG curve profile is observed, indicating the beginning of polymer decomposition at 216 ° C and termination at 460 ° C. Removing the organic mold shows the NPH30-SiO-10GPS-PEG40 TG curve whose profile is almost similar to the NPH30-CTAB-Siio-IOGPS-PEG40 and the mass difference between both after 900 ° C is only 1%, referring only to surfactant. It can then be said that in the process of glycidoxy group functionalization, PEG coupling and removal of the organic mold - comprising 3 solution steps - CTAB molecules randomly escaped from the nanoparticle pores so that in the extraction step the amount In order to be eliminated by treatment with ethanolic HCl solution, it was already small, generating a similar profile between both commented curves. In the DTA, this phenomenon is evident, since the height ratio between the peaks present at 300 ° C and 577 ° C, and 256 ° C and 577 ° C from the nano-carriers before and after elimination, respectively, is the same. .

Taking into account the changes in the behavioral patterns of the curves as production is followed at each step, it is possible to confirm the presence of long chain PEG in the molecular vehicle, a fact that was not yet precisely evident. using the infrared spectroscopy technique. However, it is not yet possible to conclude the occurrence of covalent bonding between the glycidoxy group and PEG, and the evaluation by NMR is necessary. [0144] The NPH30 10GPS-PEG-SiO ₄ was subjected to the adsorption-desorption of nitrogen and its analysis results are shown in Figure 34. It is evident a significant similarity between the profile of the isotherm of said nanoparticle with NPH30- SiOH, both referring to type IV with mild hysteresis of type H3 according to the IUPAC classification. This result proposes a complex mesoporous structure maintained before and after functionalization.

The surface areas of NPH30-SiOH and NPH30-SiO-10GPS-PEG ₄ o calculated by the BET method are 898 m ² / g and 760 m ² / g, respectively. The total pore volume values calculated by the BJH method are respectively 2,2 cm ³ / g and 1,5 cm ³ / g. These latest results suggest the possible pore occlusion due to GPS silane functionalization followed by PEG coupling, contributing to the reduction of the total pore volume - it is worth noting that occluded pores physically prevent nitrogen adsorption in the innermost cavities of the nanoparticle contributing to the sorption of smaller quantities of gas and, consequently, lower bound volume. The decrease in surface area as a function of post-rhodification reflects the same conclusion regarding the total pore volume result.

 By analyzing the pore distribution graph of the nanoparticle, the spatial predominance of cavities between 2 nm and 10 nm can be corroborated to the conclusion of the molecular vehicle complexity in the context of surface chemistry.

Figure 35 shows the carbon NMR of NPH30-SiO-10GPS-PEG40, as well as the intermediate and starting nanoparticles. In the NPH30-SiOH spectrum the 4 peaks already mentioned are from the aromatic rings as the only organic entity present before functionalization. By condensing the glycidoxy group, changes in the spectrum become evident. Analyzing the NPH30- SiO-lOGPS, we have the 4 peaks of the phenyl group already studied; three carbon-related nuclei 11 (8.2 ppm), 12 (22.7 ppm) and 13 (71.9 ppm) from the propyl group of the glycidoxy function; two nuclei referring to carbon 16 of the epoxy group (44.7 ppm) and carbon 14 adjacent to said three-membered ring (70.8 ppm). Theoretically, the carbon 15 peak present in the epoxy group should appear at 51.4 ppm, which is not easily observed. Such information generates discussions regarding the possibility of opening the ring prior to the PEG coupling, which is confirmed by observing in the same spectrum the 16 'carbon peak. At the opening of the ring, a diol is formed (see Open GPS group in Figure 11) which alters the chemical offsets of carbons 15 and 16, now called 15 'and 16' to differentiate and identify them in the spectrum. Knowing that the peak of carbon 16 at 44.7 ppm is not very evident, the one regarding nucleus 15 hardly appears, at 63.7 ppm and 70.7 ppm, there are peaks referring to atoms 16 'and 15' respectively, and the last two cited should not be present in such a spectrum. It can therefore be concluded that the NPH30-SIO-10GPS has in. its structure few closed glycidoxy groups and reasonable amounts of this function presenting the open 3-membered ring, making it unable to couple to PEG40. In the spectrum of NPH30-SiO-10GPS-PEG40 there is a reduction in carbon peak 16, which in this spectrum is renumbered 17 due to its chemical shift to 71.2 ppm. In this region, its identification is difficult due to the overlap with the peak of atom 14. However, it is evident the increase of the peak intensity present in 60.1 ppm, coming from the carbon nuclei of the polyethylene glycol chain, confirming its presence. Coupling of PEG engendered after surface chemical reaction between epoxy ring and polymer carboxylic acid can be confirmed, except for the best of judgment, by the presence of a peak 226. ppm (core 18) refers to the carbon ester generated after said chemical transformation.

In Figure 36 are shown the silicon NMR spectra on NPH30-SiOH and NPH30-SiO-10GPS. It should be noted that the silica structure before and after PEG40 coupling does not cover variations of areas between existing silicon sites, since the chemical reaction occurs between the polymer carboxylic acid and the epoxy group of the GPS function, ie is: silanol groups are not involved, eliminating the need for characterization of NPH30-SiO-IOGPS-PEG40 by ²⁹ Si NMR, which would have the same profile as NPH30-SiO-10GPS. Analyzing the figure, silicon sites are typical in their respective chemical displacements, as commented earlier. Analyzing the relationship ΣQ ⁿ / ⁿ ΣT, it has been (Q2 + Q3 + Q4) / (T2 + T3) and SiOH = 3.61 (Q2 + Q3 + Q4) / (T2 + T3) = PEG40 3,26, indicating a decrease in the ratio and, therefore, an increase in the areas referring to the condensed silane sites.

Table 1 shows the particle sizes and zeta potential of each molecular vehicle encompassed by the present invention. Acceptable polydispersity indices (IPD) for a satisfactorily monodisperse system should be less than 0.4, which occurred in all situations. presented. It is observed that surface modification does not significantly alter the average size of colloids (as previously mentioned). Surface loads showed interesting differences from the point of view of surface modification. Knowing that the surface of NPH30-SiOH, as well as any silica nanostructures in basic medium has negative charges, it is expected that after PEG (neutral) functionalization there will be a tendency to surface neutrality even though such parameters are unreliable. to predict this surface phenomenon, since in this process there is a Gaussian distribution of loads and sizes. After the functionalization of NPH30-SiOH with PEG10 in amounts equivalent to 10%, 25% and 50% mol-Si its surface load decreased from -20.5 ± 5.80 mV to -13.5 ± 5.6 mV, - 5.4 + 5.6 mV and -8.81 ± 5.41 mV, respectively, a qualitative indicator, but essential for the complementarity of structural characterizations. For surface modification with PEG40, the initial load decreased to -5.98 ± 4.66 mV.

Table 1: Particle Size and Zeta Potential of Nanoparticles

 Surface Load

 Non-particle Size (nm) (mV)

 Potential Size Deviation

 Medium IPD Zeta Standard

 NPH30-SiOH 104.6 0.116 -20.5 5.5

NPH30-SiO-10PEG10 116, 1 0.240 -13.5 5, 6

NPH30-SiO-25PEG10 109.3 0.104 -5.4 7.3

NPH30-SiO-50PEG10 126.6 0.240 -8.8 5.4

 NPH30-SiO-1OGPS-

PEG40 146.5 0.263 -6.0 4.7

Colloidal Stability Studies

[0150] The set of structural characterizations confirmed the achievement of all nanoparticles as well as the functionalizations on the internal (phenyl) and external (PEG) surfaces. In this context, it was also possible to prove the production of nanoparticles having a porous complexity in which the significant variety size was obtained, reflecting the desired uneven texture. The materials showed large surface areas, important in the study of interaction of nanostructures with biosystems.

 Although elucidated, nanoparticles need to be subjected to the study of colloidal stability in order to evaluate the influence of PEG on the acquisition of this new biologically interesting physicochemical property. The graph shown in Figure 37 shows the appearance of colloidal stability (in PBS at 250 pg / ml concentration) of each nanoparticle synthesized in the present invention. In the absence of rotation, the presence of 100% of each nanoparticle in the supernatant was taken as a parameter.

[0152] According to the graph, the instability of NPH30-SiOH is observed throughout the centrifuge's rotational range, since the percentage of this nanoparticle in the supernatant over the entire studied range was close to 0. As the Short-chair PEG functionalization content ranges from 10% mol-Si to 50% mol-Si. An increased colloidal stability at 94 rcf is observed, ie, the NPH30-SiO-50PEGio has a higher percentage of nanoparticles suspended in relation to the others, confirming the direct relationship between the functionalization rate of PEGio silane and the stable colloidal characteristic. In this situation NPH30-SiO-IOGPS-PEG40 becomes more stable compared to NPH30-SiO-10PEGio and more unstable compared to the others. This result indicates that in the 10% functionalization content the nanoparticle holding longer chain PEG exhibits greater colloidal stability. From the rotation of 2348 rcf all nanoparticles exhibited unstable behavior as expected, since the centrifugal force acting in this situation is sufficient to generate coalescence. The influence of nanoparticle aggregation state as a function of time in hemolysis

 [0153] The universe of nanoparticles demands the evaluation of its colloidal behavior in different dispersion media from the one in which the colloid is synthesized (ethanol, deionized water, among others) to the biological fluid where it will be applied, such as PBS. In this context, the behavior of nanoparticles in this saline solution was evaluated in order to correlate the possibility of aggregation phenomena supposedly associated with different equally possible hemolytic effects.

 Prior to the application of nanoparticles in the process of red blood cell interaction, a study of the effect of aggregation on PBS was carried out during the same incubation period with red blood cells. The resulting graph is shown in Figure 38.

 The average size of nanoparticles in deionized water is approximately 100 nm according to the data commented on in the characterization topics. However, when added in PBS buffer the nanoparticles aggregate so that the average particle size reaches the order of 1000 nm (see graph below). This behavior is related to the strong electrostatic interactions between the negatively charged nanoparticles and the electrolytes present in the PBS buffer. In the time interval covered in this study, there was an increase in the average size of colloids as a function of time in all situations, reinforcing the gradual interaction between the colloidal system and the buffer solution in which it is dispersed. Note that the polydispersity index remained close to 1 in virtually all steps, reflecting significantly polydispersed systems.

Confirming the gradual trend in the aggregation of nanoparticles in PBS for one hour, it is necessary to study the influence of this phenomenon on hemolysis and therefore the results were grouped at two distinct times for comparative effects as explained in the experimental procedure (nanoparticles dispersed in PBS for 1 minute and 60 minutes prior to addition gives red cell stock solution).

 Evaluating the results shown in Figure 39, it is found that nanoparticles in contact with PBS for one minute generate higher hemolysis contents than those maintained for one hour. Knowing the aggregation characteristic of nanoparticles as a function of time, it is trivial to understand that increasing the size of nanoparticles minimizes the lysis of red blood cells with respect to the colloidal systems studied. In the aggregation phenomenon the total intrinsic surface area of each carrier decreases, that is, the number of silanol groups directly accessible to the red blood cells decays and, consequently, the less pronounced nanoparticle-cell specific interactions reflect less hemolytic effect by the substantially aggregated nanoparticles.

Analyzing individually the group of nanoparticles holding polyethylene glycol of short chair, it is observed direct influence of the increased functionalization rate of PEGio silane with the minimization of hemolysis, as observed in all previous situations and according to the literature data mentioning the suppression of hemolysis as a function of the presence of said polymer. Unlike experiments involving the influence of a common surface area for all nanoparticles on hemolytic aspects, the only molecular vehicle to have hemolysis higher than the starting nanoparticle (free of external functionalization) was NPH30-SYLOOPEGIO. The other nanoparticles have hemolytic effects. lower rates than NPH30-SiOH as expected. The 10% mol-Si-containing PEGio silane nanoparticle has a larger surface area compared to the starting colloid and, according to ¹³ C NMR, the results also show that the amount of functionalized polyethylene glycol is significantly lower compared to those with 25% and 50% mol-Si, that is, the NPH30-SiOH and NPH30-SiO-10PEGio have significant structural similarities, differing only by the tiny presence of PEGio for the latter. Certainly the phenomenon present here is similar to that discussed in the previous topic, based on the higher adsorption of blood proteins by NPH30-SiOH compared to NPH30-Si010PEGio, giving the latter a greater hemolytic effect due to the greater presence of unprotected SiO- Compared to other nanoparticles belonging to the short-chain PEG group, the decrease in hemolytic effect may be associated with the decrease in negative charges due to the functionalized polymer as well as its polymeric chain possibly shielding the remaining silanol groups. Special attention to NPH30-SiO-50PEGio should be highlighted: this molecular carrier has the second smallest surface area in relation to the nanoparticles of the summed to the high PEG content grafted to its surface. As noted for the presence of the polymer, the smaller surface area as well as the aggregating effect are factors that directly contribute to the minimization of hemolysis in the context of mesoporous silica nanoparticles. The graph reflects such assumptions by observing the optimal performance of the NPH30-SiO-50PEGio with only 1.6% and 1.18% hemolysis at times 1 minute and 60 minutes, respectively.

Finally, at NPH30-SiO-10GPS-PEG ₀ , its aggregation effect remained slightly constant as a function of time, as hemolysis rates at 1 minute and 1 were relatively similar and equal to 4.9% and 4.6% respectively. Although significant discrepancies between the results are not noticeable, the functionalization of this last nanoparticle showed efficient results to prove direct influence of long chain polyethylene glycol on hemolysis. At this stage of the study the molecular vehicle mentioned showed efficiency comparable to NPH30-SiO-25PEGio, except that for the latter the aggregation as a function of time reflects significant difference in hemolytic effect.

Rescuing the characterization results of NPH30-SIO-IOGPS-PEG40 via ¹³ C-NMR, there was a question to be clarified regarding the visualization of a supposedly carbon-18 peak (carbon ester between the glycidoxy group bonding). and the carboxylic function of HOOC-PEG40) Assuming no such binding, the interaction of the polymer with the nanoparticle via electrostatic interactions would be unfeasible, as it does not have charged sites to make them viable. Thus, after centrifugation of the nanoparticle, the significantly hydrophilic PEG40 present in the supernatant would be automatically discarded and the final product would consist of the nanoparticle functionalized only with glycidoxy groups. As discussed earlier, the epoxy ring of said functions can be opened to form a diol group which, like silanol functions, would interact with red blood cells in a similar manner causing toxicity and thus acquiring substantially the same behavior as NPH30-SiOH. In all situations observed so far, this molecular vehicle exhibited behavior that directs the results to the reduction of hemolysis, indirectly indicating the presence of PEG40 in its structure. If this polymer is present then it should necessarily be covalently attached to the nanoparticle and therefore the peak in its ¹³ C NMR spectrum is from carbon 18.

 [0161] The following examples refer to the Certificate of Addition, filed under the number BR 13 2015 030941 3, on December 10, 2015, which reflects the improvement of this process by improving the drug encapsulation steps. hydrophobic to pegylated silica nanoparticles carrying hydrophobic drugs comprising said encapsulated drug, and their use in anti-tumor and anti-inflammatory treatments.

[0162] In each of the suspensions of silica nanoparticles NPH30-SiOH, NPH30-10PEG500 (or NPH30-SiO-10PEGio), oo NPH30-50PEG ₅ (or NPH30-SiO-50PEGio) and ₂ ooo NPH30-10PEG (or NPH30 -SiO-10PEG4 ₀₎ 1 mg / ml (10 mg) was added 1 mg of curcumin. The nanoparticle + curcumin mixtures were subjected to ultrasonic homogenization for 30 minutes. After this step the systems were separated in two ways:

 1) Centrifugation of nanoparticles at 0, 500, 1000 and 1500 rpm and collection of the respective supernatants for analysis of curcumin concentration by UV-Vis spectroscopy;

 2) Decantation of nanoparticles for 24 hours and collection of respective supernatants for analysis of curcumin concentration by UV-Vis spectroscopy.

To calculate the drug concentration in each of the situations, the ethanol curcumin calibration curve was developed at five known concentrations (1, 5, 10, 30 and 50 μg / mL). For each concentration, the area under the curcumin absorption curve in the region between 200 and 550 nm was estimated for graph acquisition relating the area as a function of concentration. Finally, to determine the concentration of encapsulated curcumin in each of the pegylated silica nanoparticles in the above mentioned situations, 1 ml of the respective supernatant was subjected to UV-Vis spectroscopy so that area value under the curve similarly estimated As described in the paragraph above, it was compared to the calibration curve and concentration-related, thus acquiring the drug mass value per unit volume of suspension.

Figure 40 shows the curcumin calibration curve in ethanol as well as the information for each spectrum acquired at each of the five known concentrations. According to the results, the line drawn at the five points is adequate to estimate the concentration calculation of curcumin in the concentration range between 1 and 50 μg / mL, since the value of R ² = 0.99. The equation of the line is described as:

 Curcumin concentration (x) = [area under the curve (y) - 22.9] / 14.3

Figure 41 shows the graph of curcumin concentration in the pegylated silica nanoparticles after each separation by centrifugation and decantation. In all situations, the influence of PEG on curcumin encapsulation efficiency is noticeable, since all pegylated nanoparticles comprise a higher concentration of encapsulated drug over non-functionalized nanoparticle (NPH30-SiOH). In all situations it is evident that increasing the amount of PEG on the outer surface increases the amount of encapsulated curcumin due to the hydrophilic character of the polymeric chain, coupled with the increased solubility of the pegylated silica nanoparticles in water. Additionally, it is observed that at speeds above 1000 rpm and under the condition of decantation, the increase of the polymer chain (comparison between NPH30-10PEG ₅ oo and NPH30-10PEG ₂₀ oo) causes slight decrease in the amount of encapsulated curcumin possibly due to predominant steric effects on NPH30-10 PEG2000 over NPH30-10PEGsoo · However, functionalization with PEG2000 showed higher encapsulation efficiency of the drug compared to non-functionalized nanoparticle (NPH30-SiOH) confirming the importance of polyethylene glycol for greater drug efficiency within the nanoparticle.

 For comparison of the best method for separating pegylated silica nanoparticles comprising curcumin and unencapsulated curcumin, the concentration of the drug present in water (in the absence of silica nanoparticles) was estimated at all centrifugation conditions and at decantation situation. Looking at Figure 41, the presence of curcumin in water at all the rotational forces studied (0 to 1500 rpm) is clear. However, the concentration of free drug in water decreases with increasing rotational force, tending to zero in the 1500 rpm situation where the concentration of curcumin is only 1.6 μg / mL. Such result indicates that in the supernatant of nanoparticles centrifuged at 1500 rpm there are silica nanoparticles comprising encapsulated curcumin and said drug possibly free in small amount in the aqueous environment.

To optimize the separation method a decantation process of the nanoparticles was carried out over a period of 24 hours. In this situation the amount of curcumin in water (in the absence of nanoparticles) is zero, as suggested in the graph in Figure 2. Therefore, in the supernatant of silica nanoparticles decanted for 24 hours there is possibly only said nanocarriers covering the encapsulated drug. characterizing decantation as the best separation method studied.

 An important point to note is that, when added to the described silica nanoparticle suspensions, curcumin is not only encapsulated in nanocarriers, but also possibly interacting with the outer surface of molecular vehicles. that is, the drug referred to as encapsulated in any context described above may also be interacting superficially through specific interactions to be confirmed in further work and in greater detail.

[0170] Figure 42 shows the image of the suspensions NPH30 SiOH-silica nanoparticles NPH30-10PEG ₅ oo o NPH30-10PEG NPH30-50PEG ₅₀ ^'in 5ooo. presence of curcumin and said drug added in water in the absence of molecular vehicles (H2O) after 24 hours of decantation and prior to removal of the supernatant. The hydrophobic characteristic of the drug is evident in the clear supernatant in the H2O bottle characterizing the problem of the insolubility of hydrophobic drugs in the blood (hydrophilic) whereas in the presence of silica nanoparticles their yellow colored supernatants show the stability of the suspended drug upon application of the drugs. molecular vehicles.

 Finally, the presence of PEG on the outer surface of the colloids was of fundamental importance for the best encapsulation efficiency and / or interaction with curcumin and characterizes such systems as potential candidates for antitumor application, since (hydrophilic) polyethylene glycol confers longer circulation times of nanoparticles in the bloodstream, a crucial property for reaching target cells effectively.

Claims

 Process for obtaining pegylated silica nanoparticles carrying hydrophobic drugs characterized by the following steps:

(a) dissolving surfactant between 0,65 and 0,75 g in ammonia catalyst at a concentration between 0,03 and 0,07 mol / L and an alcoholic reaction solvent in volumes between 2,5 and 3,8 mL; b) stirring the solution obtained in step (a) at a temperature between 40 and 80 ° C;

c) adding inorganic silicon precursor between 1 and 3 mL in hydrophobic silane;

d) adding dropwise the solution obtained in step (c) to the solution obtained in step (a);

e) stirring the solution obtained in step (d) for 120 minutes at temperatures between 40 and 80 ° C;

el) addition of inorganic silicon precursor after 60 minutes of reaction at volumetric quantities between 100 and 150 xL; e2) addition of inorganic silicon precursor after 90 minutes of reaction, in volumetric quantities between 100 and 150 i; f) Centrifugation for 40 to 80 minutes, between 10,000 and 20,000 rpm; discarding the supernatant and redispersing in ethanol; g) Functionalization of the external surface of silica nanoparticles;

h) Preparation of an ethanolic HCl solution in a preferred volume ratio of 1: 9 HCl: ethanol; (i) Addition of nanoparticles comprised in steps gl. 5 or g2.12 in the solution prepared in step h; ultrasound application at times between 10 and 30 minutes, preferably 15 minutes; j) Centrifuging the solution obtained in step (i) between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000; discarding the supernatant and redispersing in ethanol; repeated at least twice; and storage until encapsulation of the hydrophobic drug.

k) Suspension of nanoparticles obtained in step (j) in aqueous solution (component 1) in masses between 5 and 15 mg, preferably 10 mg, and in concentrations of up to 1 mg / mL; and adding 0.5 to 2 mg, preferably 1 mg, of hydrophobic drug (component 2);

 1) Mixing the components comprised in step <k) by means of ultrasonic homogenization at times between 30 and 60 minutes;

m) separating the silica nanoparticles carrying hydrophobic drugs comprising said encapsulated drug (product) from unencapsulated hydrophobic drug (component 1) by separation means;

n) Obtaining silica nanoparticles carrying hydrophobic drugs comprising said encapsulated drug.

Process according to Claim 1, characterized in that the surfactant dissolution is preferably 0.75 g, the ammonia catalyst concentration is preferably 0.05 mol / l, and the volume of alcoholic reaction solvent is preferably 3. 2 mL;

Process according to Claim 1, characterized in that the stirring of the solution in step (b) is at a preferential temperature of 60 ° C.

 Process according to Claim 1, characterized in that the addition of inorganic silicon precursor is preferably 1.5 ml in hydrophobic silane.

 Process according to Claim 1, characterized in that the stirring in step (e) is preferably 60 ° C.

 Process according to Claim 1, characterized in that the addition of inorganic precursor, after 60 minutes and 90 minutes of reaction, is in a preferred volume of 124 µl.

 Process according to Claim 1, characterized in that the centrifugation of step (f) is preferably for 60 minutes at 15,000 rpm.

 Process according to Claim 1, characterized in that step (g) may comprise the following steps:

gl.l) dispersion of starting nanoparticles in masses between

50 and 500 mg, preferably 300 mg, in ethanol solvent in volumes between 20 and 60 mL, preferably 40 mL; stirring the solution between 40 and 90 minutes, preferably 75 minutes under temperature between 40 and 80 ° C preferably 60 ° C _r;

gl.2) Addition of silane-PEG to the solution of step gl.l;

gl.3) Stirring the solution from step gl.2 between 10 and 20 hours, preferably 13 hours;

gl.4) Centrifugation of the solution comprised in step gl.3 between 40 and 80 minutes, preferably 60 minutes, between 10,000 and

20,000 rpm, preferably 15,000; discarding supernatant and re-dispersing in ethanol; repeated at least twice.

Process according to Claims 1 and 8, characterized in that the step (g) may alternatively comprise the following steps:

g2.1) dispersion of ■ starting nanoparticles in masses between 50 and 500 mg, preferably 300 mg, in ethanol solvent in volumes between 40 and 80 mL, preferably 62 mL and stirring of the solution between 20 and 60 minutes, preferably 30 minutes in temperature between 60 ° C is 90 ° C, preferably 85 ° C;

g2.2) Addition of an intermediate silane with reactive functional groups to the obtained solution;

g2.3) Stirring the solution between 10 and 20 hours, preferably 12 hours in an inert gas atmosphere;

g2.4) Centrifuging the solution between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000 and discarding the supernatant and re-dispersing in ethanol; repeated at least twice;

g2.5) Dispersion of intermediate nanoparticles obtained in step g2. in masses between 50 and 500 mg, preferably 150 mg, in ethanol solvent in volumes between 10 and 30 mL, preferably 20 mL and stirring of the solution between 15 and 45 minutes, preferably 30 minutes at a temperature of 95 ° C, preferably;

g2.6) Addition of a strong base to the obtained solution;

g2.7) Dissolution of PEG-COOH (PEG with terminal carboxylic acid) comprising mass between 5 and 70 mg, preferably 15.1 mg in ethanol solvent in volumes between 500 and 1500 μΐι, preferably 750 æL;

g2.8) Addition of the PEG-COOH solution from step g2.7 to the solution obtained in step g2.6 for coupling reaction between reactive functional groups of the intermediate nanoparticle with reactive functional groups and the carboxylic acid of PEG-COOH;

g2.9) Stirring the solution obtained in step g2.8 between 10 and 20 hours, preferably 12 hours at a preferred temperature of 95 ° C;

g2.10) Centrifuging the solution obtained in step g2.9 between 40 and 80 minutes, preferably 60 minutes, between 10,000 and 20,000 rpm, preferably 15,000, discarding the supernatant e.g. re-dispersion in ethanol; repeated at least twice.

Process according to Claim 1, characterized in that the separation means of step (m) comprise centrifugation or decantation.

 Process according to Claim 10, characterized in that the centrifugation is carried out at rotational forces between 1500 and 2000 rpm, preferably for 2 minutes.

Process according to Claim 10, characterized in that the decantation takes place at least 24 hours.

Process according to Claim 1, characterized in that the surfactant preferably comprises hexadecyltrimethylammonium bromide.

Process according to Claim 1, characterized in that the inorganic silicon precursor comprises silicon alkoxides and organosilanes, preferably tetraethylorthosilicate.

Process according to Claim 1, characterized in that the hydrophobic silanp hair is used in amounts of 10 to 40 mol% Si, preferably 30 mol% Si.

16. Process according to claim 1, wherein the hydrophobic group comprised in the hydrophobic silane comprises hydrophobic ^groups' selected from aromatic groups and derivatives; and hydrocarbon groups and derivatives.

Process according to Claim 1, characterized in that the aromatic groups and derivatives are selected from the group of phenyl _/ acetophenyl and pyrene functions.

 Process according to Claim 1, characterized in that the hydrocarbon-based groups and derivatives are selected from the group of octyl and octadecyl functions.

^19: Process according to claim 1, characterized in PEG-silane is used in amounts from 2 to 50 mol-% Si.

Process according to claim 1, characterized in that the PEG is comprised. in silane-PEG comprise at least 350 Da. ^■ ■

 Process according to Claim 1, characterized in that the intermediate silane with reactive functional groups is used in amounts between 2 and 50 mol% Si.

Process according to Claim 1, characterized in that the reactive functional groups comprised in the intermediate silane comprise epoxy functions.

Process according to Claim 1, characterized in that the strong base used preferably comprises potassium hydroxide.

 Process according to Claim 1, characterized in that the PEG-COOH used comprises at least 350 Da.

 Process according to Claim 1, characterized in that the hydrophobic drug is used in amounts of 1 to 20% by mass.

 Process according to Claim 1, characterized in that the hydrophobic drug used is selected from the group of drugs comprising curcumin, camptothecin, doxorubicin, violacein, tamoxifen, beta-lapacone and the like, preferably curcumin or camptothecin.

 Pegylated hydrophobic drug carrier nanoparticles characterized by being obtained by the process described in claims 1 to 26 and comprising polyethylene glycol (PEG) -functioning outer surface, hydrophobic-grouped porous inner surface, and encapsulated hydrophobic drug.

 28. Pegylated nanoparticles carrying hydrophobic drugs comprising polyethylene glycol (PEG) -functional outer surface, hydrophobic-functionalized porous inner surface, and encapsulated hydrophobic drug.

Pegylated hydrophobic drug carrier nanoparticles according to claim 28, characterized in that the hydrophobic drug used is selected from the group of drugs comprising curcumin, camptothecin, doxorubicin, violacein, tamoxifen, beta-lapacone and the like, preferably curcumin or camptothecin.

 30. Pegylated nanoparticles carrying hydrophobic drugs comprising polyethylene glycol (PEG) functionalized outer surface, porous inner surface functionalized with hydrophobic groups, and encapsulated curcumin.

 Nanoparticles according to any one of claims 28 to 30, characterized in that the PEG comprises at least 350 Da and is functionalized in amounts between 2 and 50 mol-Si.

 Nanoparticles according to any one of claims 28 to 30, characterized in that the hydrophobic groups are functionalized in amounts between 10 and 40 mol% Si, preferably 30 mol% Si.

 Nanoparticles according to Claim 32, characterized in that the hydrophobic groups are selected from aromatic groups and derivatives; and hydrocarbon groups and derivatives.

 Nanoparticles according to claim 33, characterized in that the aromatic groups and derivatives are selected from the groups of the phenyl, acetophenyl, and pyrene functions.

Nanoparticles according to claim 33, characterized in that the hydrocarbon-based groups and derivatives are selected from the group of octyl and octadecyl functions.

36. Nanoparticles, ^"according to. one of the claims. 28 to 30, characterized in that they comprise hydrophobic drug in amounts from 1 to 20% by mass.

Nanoparticles according to any one of claims 28 to 30, characterized in that they comprise a diameter between 40 and 100 nm.

 Nanoparticles according to any one of claims 28 to 30, characterized in that it comprises a zeta potential of between -5.3 and -35 mV.

Nanoparticles according to any one of claims 28 to 30, characterized in that it comprises a surface area of between 760 and 943 m ² / g.

Nanoparticles according to any one of claims 28 to 30, characterized in that it comprises pore volume between 1.0 and 1.8 cm ³ / g.

 Nanoparticles according to any one of claims 28 to 30, characterized in that they cause maximum hemolytic effects of between 2 and 14%.

 Use of pegylated hydrophobic drug carrier nanoparticles, as described in claim 27, characterized in that they are applied in anti-tumor and anti-inflammatory treatments.

 Use of pegylated hydrophobic drug carrier nanoparticles as described in claims 28 to 41, characterized in that they are applied in anti-tumor and anti-inflammatory treatments.