Method for synthesizing zeolite nanoparticles in phosphate-buffered saline - related nanoparticles, suspensions, and pharmaceutical compositions

Abstract

The present invention relates to a method for synthesizing nanoparticles comprising or containing at least one zeolite nanocrystal, comprising: - preparing a first composition / solution 1 comprising an aluminum source and a source of alkali metal M, particularly K ions; - preparing a second composition / solution 2 comprising a silicon source and a source of alkali metal M, particularly K ions, wherein compositions / solutions 1 and 2 contain no organic structuring agents; - mixing the two compositions / solutions 1 and 2 together; - crystallizing the mixture; and - separating the nanoparticles thus formed. According to the present invention, both the first composition / solution 1 and the second composition / solution 2 consist of the sources and phosphate-buffered saline. The present invention also relates to a colloidal suspension of the obtained nanoparticles and a pharmaceutical composition containing the nanoparticles.

Description

[Technical Field]

[0001] This invention relates to a method for synthesizing zeolite nanoparticles, suspensions of these nanoparticles, pharmaceutical compositions containing these nanoparticles, and nanoparticles obtained according to the method of this invention. These suspensions and nanoparticles can be used, in particular, in the medical field, and especially for the treatment of hypoxic solid tumors. [Background technology]

[0002] Zeolites are a type of crystalline microporous material. They are formed by a three-dimensional arrangement of TO4 (T = Si, Al, or other heteroatoms) that are bonded to each other by sharing an oxygen atom. In aluminosilicate zeolites, AlO4 5- The presence of ions creates a negative charge in the skeleton, and in order to maintain electrical neutrality, so-called compensating cations (alkali metals or alkaline earth metals), such as Na, are produced. + , K + Ca 2+ Sr 2+ This must be compensated for. The voids are defined by the aluminosilicate framework formed by these three-dimensional chains. The voids in the tetrahedral crystal network are interconnected and occupied by alkali ions and water molecules. Dehydration of the zeolite removes the present water molecules. Zeolites also possess a micropore network formed from cavities, channels connecting the cavities, and pores connecting the channels or cavities to the outside. Zeolites can be natural or synthetic. There are currently more than 250 types of zeolites.

[0003] Patent Document 1 discloses FAU and EMT-type zeolite nanocrystals in which sodium ions are replaced by Fe or Gd ions and which can be filled with gaseous dioxygen. These nanocrystals may be used for the diagnosis and treatment of brain tumors, including glioblastoma. The zeolite nanocrystals in these two documents are synthesized in water, which limits their medical use. In fact, over long periods and at high concentrations, these nanocrystals appear to cause allergic problems.

[0004] Non-patent document 1 discloses a method for synthesizing LTL-type nanoparticles. The composition of the mixture used for crystallization is as follows: 5K2O:10SiO2:0.5Al2O3:200H2O. This solution is an aqueous solution and does not contain PBS.

[0005] Non-patent document 2 does not disclose the synthesis of LTL-type nanozeolite. This document shows that ions present in nanozeolite can be moved to the outside and replaced. [Prior art documents] [Patent Documents]

[0006] [Patent Document 1] International Publication No. 2019 / 030200 [Non-patent literature]

[0007] [Non-Patent Document 1] Kharchenko et al., “formation of copper nanoparticles in LTL nanosized zeolite: kinetics study”, The journal of physical chemistry C, 5 November 2016, volume 120, No. 46 [Non-Patent Document 2] F. Mayer et al., “nanozeolite-LTL with GdIII deposited in the large and EUIII in the small cavities as a magnetic resonance optical imaging probe”, Chemistry-A European Journal, John Wiley & sons, Inc, 12 February 2014, DE volume 20, No 12

Summary of the Invention

Problems to be Solved by the Invention

[0008] One object of the present invention is, therefore, to propose a novel pharmaceutical composition containing zeolite nanoparticles that can be used for the diagnosis and / or treatment of certain medical conditions, particularly hypoxic solid tumors and, in particular, brain tumors.

[0009] Another object of the present invention is to propose a novel method for synthesizing such nanoparticles.

[0010] Another object of the present invention is to provide a colloidal suspension of these nanoparticles.

[0011] Another object of the present invention is to provide zeolite nanoparticles with reduced toxicity and / or improved oxygen adsorption and / or oxygen release ability.

Means for Solving the Problems

[0012] The first aspect of the present invention comprises the following steps, namely: - preparing a first composition / solution 1 containing an aluminum source and a source of ions of an alkali metal M, particularly K (potassium); - A second composition / solution 2 is prepared, comprising a silicon source and a source of alkali metal M, particularly K (potassium) ions, wherein compositions / suspensions 1 and 2 contain no organic structuring agents; - The compositions / solutions 1 and 2 are mixed together, and the mixture is left to stand under stirring; - The mixture is crystallized at a temperature of 50°C or higher; and - The nanoparticles formed in this manner may be separated. The present invention relates to a method for synthesizing nanoparticles consisting of at least one zeolite nanocrystal, or containing at least one zeolite nanocrystal.

[0013] The features of the present invention are that the first composition / solution 1 and the second composition / solution 2 each consist of the supply source and an aqueous buffer (PBS) comprising water and 137 mM / L NaCl, 2.7 mM / L KCl, 10 mM / L Na2HPO4, and 1.76 mM / L KH2PO4, with a pH of 7.2, and the mixture of compositions / solutions 1 and 2 has the following molar composition (I), namely, [ka] It is having.

[0014] The method of the present invention is easy to implement and makes it possible to obtain a stable colloidal suspension of LTL-type nanoparticles in PBS.

[0015] Preferably, the composition is a suspension.

[0016] The above compositions / solutions 1 and 2 consist of PBS, a silicon or aluminum source, and an alkali metal source.

[0017] Depending on the embodiment, it may sometimes be necessary to heat the mixture of PBS buffer and other reagents in order to obtain a clear suspension.

[0018] Regarding temperature, the first composition / solution 1 can be prepared at room temperature. The second composition / solution 2 can be prepared, for example, at a temperature below 30°C, particularly 25°C. The mixing of the two compositions / solutions can be carried out at a temperature of 0°C or higher and below 30°C, particularly 25°C. The mixture must then be left to stand under stirring for at least 4 hours, particularly 16 hours, at a temperature of 0°C or higher and 30°C or lower, particularly 25°C. The mixture can be crystallized at a temperature of 90°C or higher and 200°C or lower, particularly 170°C. [Brief explanation of the drawing]

[0019] The features and advantages of the present invention will become clear from the accompanying drawings and the following description.

[0020] [Figure 1] Figure 1 shows the dimensional distribution of the K-LTL-PBS nanoparticles of the present invention. The nanoparticles are dispersed in a PBS solution. [Figure 2] Figure 2A shows scanning electron microscope images of K-LTL-PBS and Gd-LTL-PBS type nanoparticles (nanocrystals) with scales of 1 μm and 500 nm. Figure 2B shows transmission electron microscope images of K-LTL-PBS type nanoparticles (nanocrystals) with scales of 100 nm and 40 nm. [Figure 3] Figure 3 shows the amount of adsorbed dinitrogen as a function of the obtained relative pressure for K-LTL-PBS type nanoparticles under the operating conditions described below. [Figure 4] Figure 4 shows the viability of U87-MG cells exposed to colloidal suspensions of two nanoparticles containing 10 μg / ml and 100 μg / ml K-LTL-water or Gd-LTL-water nanoparticles, respectively, compared to the viability obtained for K-LTL-PBS and Gd-LTL-PBS nanoparticles of the same concentration. [Figure 5] Figure 5 shows the percentage of hemolysis as a function of the dose (in μg / ml) of K-LTL and Gd-LTL type nanoparticles synthesized in water or PBS on the left side. [Figure 6]Figure 6A shows the change in oxygen concentration in water obtained by contacting K-LTL-water and Gd-LTL-water particles that have been pre-filled with O2. Figure 6B shows the change in oxygen concentration in PBS obtained by contacting K-LTL-PBS and Gd-LTL-PBS particles of the present invention that have been pre-filled with O2. [Modes for carrying out the invention]

[0021] The aluminum source can be selected from any aluminum source capable of providing monomeric oxyhydroxide species. For the purposes of the present invention, "any aluminum source capable of providing monomeric oxyhydroxide species" means that the aluminum source does not provide polymeric oxyhydroxide species of aluminum and is capable of dissolving all of the aluminum in a clear aqueous solution. In particular, the aluminum source can be selected from alumina, hydrated alumina, aluminum powder, Al, Al2(SO4)3, aluminum hydroxide Al(OH)3, sodium aluminate (NaAlO2), potassium aluminate (KAlO2), and kaolin clay.

[0022] The silicon source can be selected from all silicon sources that can provide the monomeric oxyhydroxide species of Si2-Si6. Therefore, a larger unit [SiOOH] n The amount of is limited in the composition / solution 2.

[0023] In particular, the silicon source can be selected from silica hydrogel, silicic acid, colloidal silica, calcined silica, tetraalkyl orthosilicate, silica hydroxide, precipitated silica, silica fume, and clay. Colloidal silica is preferred in all embodiments of the method of the present invention.

[0024] The alkali metal source M can be selected from a source of Na (sodium) and a source of alkali metal K (potassium). Hydroxides of these two alkali metals are preferred as alkali metal sources.

[0025] According to one particular embodiment, M is K, the aluminum source is aluminum hydroxide, and the mixture of the compositions / solutions 1 and 2 is crystallized at a temperature of 90 °C or higher and 200 °C or lower, particularly 170 °C, for a period of 4 hours or more and 16 hours or less.

[0026] According to a particular variation, 2.8 ≤ x1 ≤ 8, 0.3 ≤ y ≤ 1.5 and 100 ≤ z ≤ 300, and according to a further particular variation, x1 = 5, y = 0.5 and z = 200.

[0027] Advantageously, the purification / separation of the nanoparticles is carried out using a membrane, particularly a dialysis membrane. The operation of this method is simple and inexpensive and is usually achieved using commercially available membranes (dialysis membranes), and it can also save water and is also capable of retaining PBS ions within the pore network of the nanosieve.

[0028] According to a second aspect of the present invention, the present invention relates to a method for producing a colloidal suspension of nanoparticles, in which method nanoparticles are synthesized according to the method of the present invention, and the synthesized nanoparticles are dispersed in substantially sterilized water or in a substantially sterilized PBS buffer composed of water and 137 mM / L of NaCl, 2.7 mM / L of KCl, 10 mM / L of Na2HPO4 and 1.76 mM / L of KH2PO4 and having a pH of 7.2. Thus, a suspension of zeolite nanoparticles is obtained in water or PBS.

[0029] It should also be noted that the synthesis method of the present invention can also directly obtain a suspension of LTL-type nanoparticles in PBS.

[0030] According to another aspect, the present invention relates to PBS ions, particularly Na + , K + ions, HPO4 2- , H3O + , H2PO4 - , Cl - , particularly Na+ and K + The present invention relates to nanoparticles comprising or containing at least one LTL-type zeolite nanocrystal containing ions. The nanoparticles may advantageously contain at least two of the above ions. Depending on pH, it may contain HPO4 2- H3O + H2PO4 - It may contain various amounts of ions. These ions, whatever they may be, are located on the surface of the crystalline network of the nanozeolite, which can be confused with the pore network and / or (as compensating ions) the gaps in the crystalline network and / or the surface of the nanoparticles. Preferably, Na + and K + Cations are found as compensatory ions in the gaps of the crystal network.

[0031] Advantageously, the nanoparticles further contain, in their crystalline network, cations of a metal selected from Fe, Gd, Cu, and Ce, in an amount of 0.2% by weight or more and 5% by weight or less, and / or gases selected from oxygen, dinitrogen, carbon dioxide, and mixtures thereof, contained in their pore volume. When the nanoparticles contain the above-mentioned ions, they can be useful as a comparison product in the diagnosis of solid tumors, particularly brain tumors. When the nanoparticles contain the above-mentioned gases, they can have a therapeutic effect by releasing oxygen, and this therapeutic effect is particularly pronounced in the presence of an anticancer agent. This anticancer agent can be selected from the anticancer agents described later with respect to the combination products of the present invention.

[0032] According to a particular embodiment that can be combined with each of the above embodiments, the zeolite has the following chemical formula (II), namely, [ka] The equation has such that x is greater than 9 and / or y is less than 27 and / or a is greater than 6. Advantageously, a is greater than 6 and y is less than 27. Advantageously, a is 10 or less and y is 26 or less. The equation between the vertical lines above represents the compensating ions (K) present in the nanozeolite. + and Na + ) and water molecules are shown.

[0033] According to a particular embodiment of the present invention, which can be combined with the embodiments described above, the zeolite has the following chemical formula (III), namely, [ka] has

[0034] Advantageously, each of the nanoparticles is 0.4 cm 3 / g or more and 1.5cm 3 Total pore volume of less than / g, and / or 200m 2 / g or more and 1000m 2 Specific surface area S less than or equal to / g BET It has the following characteristics. In particular, each of the nanoparticles is 0.5 cm 3 / g or more and 1.4cm 3 Less than / g, especially 0.73cm 3 / g or 0.85cm 3 Total pore volume per g, and / or 300 m 2 / g or more and 900m 2 / g or less, especially 538m 2 / g or 490m 2 Specific surface area S / g BET It may have.

[0035] In another aspect, the present invention relates to a colloidal suspension of nanoparticles comprising, or containing, at least one LTL-type zeolite nanocrystal as defined above, in a dispersant selected from alcohol, water, phosphate-buffered saline, and mixtures thereof. Preferably, PBS is selected.

[0036] In another aspect, the present invention relates to a pharmaceutical composition comprising a plurality of nanoparticles or a suspension according to the present invention, which are composed of or contain at least one LTL-type zeolite nanocrystal according to the present invention, as an active agent or contrast agent as a pharmaceutical.

[0037] In another embodiment, the present invention relates to a combination product comprising or consisting of an anticancer agent and a pharmaceutical composition according to the present invention, for administration thereof simultaneously or at time intervals. The anticancer agent is advantageously administered first. The anticancer agent can be selected from temozolomide® (4-methyl-5-oxo-2,3,4,6,8-pentazabicyclo[4.3.0]nona-2,7,9-triene-9-carboxamide metazolastone), doxorubicin (8S,10S)-10-(4-amino-5-hydroxy-6-methyl-tetrahydro-2H-pyran-2-yloxy)-6,8,11-trihydroxy-8-(2-hydroxyacetyl)-1-methoxy-7,8,9,10-tetrahydrotetracene-5,12-dione), and cisplatin (cis-diaminedichloroplatinum(II)).

[0038] According to another embodiment of the present invention, the present invention has applications as a pharmaceutical for the treatment of solid tumors, particularly brain tumors, particularly glioblastoma, or as a contrast agent, particularly as a contrast agent in magnetic resonance imaging (MRI). The present invention relates to nanozeolite or suspension according to the present invention.

[0039] The inventors have demonstrated that LTL-type nanoparticles according to the present invention, and therefore those synthesized in PBS salt buffer, exhibit lower cytotoxicity and are therefore more likely to be accepted by the human body.

[0040] PBS has the same salt concentration as the human body and also contains dihydrogen phosphate / hydrogen phosphate ion pairs, which help maintain the pH of blood in the human body. Although the inventors are not bound by this description, it appears that some of the PBS ions contained in the pore network of nanozeolite may stabilize the nanoparticles and reduce their toxic potential by preventing their degradation in the biological environment. In addition, PBS ions can also modulate the charge on the surface of the nanoparticles, which may affect the behavior of the nanoparticles and their interaction with living cells and tissues. Therefore, the nanoparticles of the present invention avoid or mitigate the aforementioned allergic phenomena.

[0041] Regardless of the embodiments and forms of the present invention, LTL zeolite nanocrystals may have a sequin (flattened parallelepiped) shape. These nanocrystals may have an average length of 5 nm or more and 20 nm or less, and an average width of 2 nm or more and 15 nm or less. In particular, the average length may be 30 nm and the average width may be 10 nm. Throughout this application, the nanoparticles are preferably made from LTL-type zeolite single crystals.

[0042] The pore network of zeolites can extend in only one direction.

[0043] The pharmaceutical compositions of the present invention may be formulated for oral, mucosal (intranasal), or parenteral administration.

[0044] definition For the purposes of this invention, the predicate "average dimension" means the average dimension (number or intensity) measured by dynamic light scattering (DLS). The average dimension is the diameter of spheres that scatter at the same rate as the particles being measured.

[0045] The basic dimensional distribution obtained by DLS is the intensity distribution. The operating conditions are as defined in the remainder of this application.

[0046] For the purposes of this invention, the term "nanoparticles" means solid particles with a maximum dimension of less than 100 nm. These may be single crystals or aggregates of crystals.

[0047] The term "composition" means a homogeneous or heterogeneous mixture of at least two compounds that may be in different physical states. Therefore, this term includes suspensions, solutions, and dispersions. This term does not limit the physical state of the substance.

[0048] The term "PBS" refers to phosphate-buffered saline, which is an aqueous solution of NaCl (0.137 M), KCl (0.0027 M), disodium hydrogen phosphate (0.01 M), and potassium dihydrogen phosphate (0.0018 M). Its pH is 7.2.

[0049] The term "LTL-type zeolite nanoparticles" refers to nanoparticles containing or composed of LTL-type zeolite single crystals. LTL-type zeolites crystallize in a hexagonal crystal system. The chemical formula for LTL-type zeolite nanocrystals is as follows: [ka] Here, x + y ≤ 36, and Na + and K + Ions exist as compensating ions, ensuring the electrical neutrality of the nanocrystals. Water molecules may be absent from the nanocrystals (c=0).

[0050] The term "solution" refers to an aqueous solution having substantially the same refractive index as water, that is, an aqueous solution with a refractive index of approximately 1.333 ± 0.030 as measured under standard conditions.

[0051] The technical terms "pore volume" or "pore network" refer to the collection of cavities, channels connecting these cavities to one another, and pores connecting the channels and cavities to the outside of the nanoparticle.

[0052] The term "pharmaceutically active agent" means an agent that imparts therapeutic properties, such as therapeutic or preventive effects, to a composition containing it, and even if the agent has harmful side effects, these side effects do not outweigh the therapeutic and / or preventive effects.

[0053] "Pharmaceutically acceptable agent" means any agent that does not cause adverse effects or adverse effects of a magnitude that would impair the health of the patient, and that can be taken orally, injected, or applied in contact with the skin and mucous membranes. [Examples]

[0054] Synthesis and characterization of nanoparticles The raw materials used in the following examples are listed below. That is, - Sodium hydroxide (Sigma Aldrich, 98%) - Potassium hydroxide (Sigma Aldrich, 90%) - Aluminum hydroxide (Type A2100, Alfa Aesar, 76.5% by mass Al(OH)3, 23.5% by mass H2O) - Aluminum powder (325 mesh, Alfa Aesar 99.5%) - Colloidal silica (SiO2 (Ludox HS-30, 30 wt% SiO2, pH=9.5-10.2 Sigma Aldrich)) - Colloidal silica (SiO2 (Ludox SM-30, 30 wt% SiO2, pH=9.7-10.3 Sigma Aldrich)) - Gadolinium nitrate Gd(NO3)3,6H2O(Aldrich, 99.9%) - Phosphate-buffered saline (PBS) (Sigma Aldrich, pH=7.15-7.25) These starting reagents were used as received from the manufacturer and were not further purified.

[0055] Analytical methods and operating conditions used The various zeolite materials obtained in the examples were characterized using various dimensional scales.

[0056] Imaging using scanning electron microscopes (SEM) and transmission electron microscopes (TEM): The morphology of the nanoparticles was investigated using a scanning electron microscope (SEM) at a voltage of 20 kV with a Tescan Mira I LMH system.

[0057] Transmission electron microscopy (TEM): The colloidal suspension was sonicated for 15 minutes, and then 2-3 drops of the particulate suspension were dried on a 300-mesh copper electron microscope grid covered with a carbon film. Images were obtained using a Tecnai G2 30 FEI microscope (Vacc=300kV, LaB6).

[0058] Dynamic light scattering (DLS) analysis: The hydrodynamic diameters of zeolite materials in various suspensions were measured using a Malvern Zetasizer Nano instrument. The surface charge of the samples was determined by measuring the zeta potential of the suspensions. This analysis was performed on purified samples with a solid concentration of 1 wt% dispersed in PBS solution at pH 7.2. The backscattering configuration (HeNe laser with a scattering angle of 173° and a wavelength of 632.8 nm with an output power of 3 mW) allows for measurements at high sample concentrations because complete transmission of incident light through the sample is not required.

[0059] Chemical analysis by ICP-MS (Inductively Coupled Plasma Mass Spectrometry) The chemical composition of the zeolite samples was characterized by inductively coupled plasma mass spectrometry using an Agilent Technologies 7900 instrument. The samples were prepared according to the following procedure: (i) 50 mg of the zeolite sample was dissolved in 3 ml of hydrofluoric acid (HF) (Sigma-Aldrich 40-45%), (ii) 0.5 ml of solution (HNO3 / HCl = 1:3 volume / vol) was added, and the mixture was then heated at 110°C for 1 hour in a polytetrafluoroethylene (PTFE) bottle (100 ml), and then (iii) 96.5 ml of distilled water and 2 g of boric acid (H3BO3) were added. The resulting solution was stirred overnight to promote the dissolution of boric acid. Finally, 10 ml of the solution was diluted 10-fold with distilled water before analysis.

[0060] Example 1: Synthesis of K-LTL type zeolite nanocrystals in PBS solution (K-LTL-PBS) Alumina suspension C is prepared by dissolving 2 g of KOH in 8 g of PBS, followed by the slow addition of 0.496 g of aluminum hydroxide.

[0061] Silicate suspension D is prepared by mixing 10 g of colloidal silica (LUDOX® SM-30) with 1.1 g of KOH and 2.88 g of PBS.

[0062] The two suspensions C and D are mixed together and then left to stand at room temperature under stirring for 24 hours.

[0063] The initial molar composition of the mixture is as follows: [ka]

[0064] The resulting mixture is crystallized in an oven at 170°C for 16 hours.

[0065] The obtained nanoparticle suspension was purified using a dialysis membrane (Visking Dialysis Tubing, code number DTV.12000.08, size No. 8 (25.4 mm, 30 m), molecular weight 12-14000 Daltons) until the pH reached 7.2. 80 to 100 ml (2.0 to 2.5 g) of colloidal suspension containing 2.5% by mass of K-LTL type zeolite nanocrystals was obtained. The crystal yield ranged from 66 to 76% by mass. The final suspension was sonicated for 2 hours.

[0066] The formula for the K-LTL-PBS zeolite nanoparticles obtained by ICP is as follows. [ka]

[0067] The formulas in parentheses correspond to the crystal network, and the values ​​between the vertical lines represent the number of moles of compensating ions and the number of moles of water.

[0068] Example 2: Synthesis of a colloidal suspension of nanoparticles according to the present invention A certain amount of the nanoparticles from Example 1 were dispersed in 10 ml of PBS under sonication for 2 hours. Colloidal suspensions containing 1% by mass, 2% by mass, 3% by mass, and 4% by mass of the K-LTL-PBS type nanoparticles dispersed in PBS were obtained.

[0069] These nanoparticles are stable in colloidal suspension at room temperature for at least one week.

[0070] Example 3: Synthesis of zeolite nanoparticles charged with metal ions Five ml each of the two colloidal suspensions obtained in Example 2, each containing 2.5% by mass of nanoparticles, was separately added to 25 ml of a 0.03 mM gadolinium(III) nitrate hexahydrate (Gd(NO3)3·6H2O) solution. The suspensions were then left to stand at room temperature for 1 hour with stirring. This ion exchange process was repeated twice. The final suspensions were then washed three times with PBS to finally obtain a Gd-LTL-PBS zeolite suspension in PBS at pH=7.2.

[0071] Example 4: Dioxygen and / or CO into a nanoparticle pore network 2 embedded Each colloidal aqueous suspension obtained in Example 2 or Example 3 was then splashed with O2, CO2, or carbogen at a flow rate of 800 ml / min for 30 minutes. In this way, nanoparticles were obtained in which the pore network was at least partially filled with O2 and / or CO2.

[0072] Example 5: Morphological properties evaluation of nanoparticles obtained in Examples 1, 3, and 4 The morphology and dimensions of synthesized K-LTL-PBS and Gd-LTL-PBS nanoparticles were measured using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Figure 1 shows a curve illustrating the dimensional distribution of K-LTL-PBS nanoparticles dispersed in PBS.

[0073] Scanning electron microscope images of K-LTL and Gd-LTL zeolite nanoparticles (see Figure 2A) show small clusters of nanoparticles. However, images obtained by transmission electron microscopy (see Figure 2B) show that these nanoparticles are small individual nanocrystals with a rectangular shape measuring 10 nm in width and 30 nm in length.

[0074] In addition, high-resolution transmission electron microscope images show that the nanoparticles are highly crystalline. These are nanoparticles formed from individual nanocrystals.

[0075] Example 6: Method and results of chemical analysis of K-LTL nanoparticles The chemical formulas of the zeolite materials for K-LTL and Gd-LTL nanoparticles are as follows (determined by ICP method): [ka]

[0076] The formulas in parentheses correspond to a crystalline tetrahedron network, and the values ​​between the vertical lines represent the number of moles of compensating ions and the number of moles of water.

[0077] Example 7: Measurement of porosity by dinitrogen adsorption / desorption The porosity of the K-LTL-PBS and Gd-LTL-PBS-PBS zeolites synthesized in the above examples was characterized by adsorption / desorption measurements of gaseous nitrogen.

[0078] The texture properties of the nanoparticles obtained in the above examples were measured by a dinitrogen adsorption / desorption isotherm measured at a temperature of -196°C using a "Micromeritics 3Flex" surface characterization system. All samples were degassed overnight under vacuum at 250°C before analysis. The volume of the outer surface and micropores of the zeolite nanoparticles were evaluated by the t-plot method.

[0079] Nitrogen adsorption / desorption measurements of two types of zeolites yielded hybrid isotherms of types I and IV according to the IUPAC classification (see Figure 3). Specific surface area S of zeolite nanoparticles. BET The measurement results for total pore volume and micropore volume are shown in Table 1 below. Micropores are defined as pores with dimensions of less than 0.08 mm.

[0080] [Table 1]

[0081] Table 2 below shows the chemical compositions of the K-LTL-PBS and Gd-LTL-PBS nanoparticles of the present invention.

[0082] [Table 2]

[0083] The pharmacological properties of the nanoparticles of the present invention were studied. The results are shown in the following examples.

[0084] Example 8: Human culture cytotoxicity test The cells used were derived from the human glioblastoma cell line U87-MG, purchased from the American Type Culture Collection (ATCC). These cells were cultured in 1 g / L DMEM (Sigma-Aldrich) supplemented with 10% fetal bovine serum (Eurobio), 2 mM glutamine (Sigma-Aldrich), and penicillin (100 U / ml) / streptomycin (100 μg / ml) (Sigma-Aldrich). The cells were cultured at 37°C, 5% CO2, and 95% humidity.

[0085] Cell viability was assessed 72 hours after exposure to zeolite nanoparticles using the WST-1(Rock) test, according to the manufacturer's instructions. The results can be seen in Figure 4. This figure also shows the percentage of cell viability after exposure to nanoparticles synthesized in water instead of PBS. With respect to the K-LTL-PBS and Gd-LTL-PBS nanoparticles of the present invention, a higher percentage of viable cells was observed compared to nanoparticles synthesized from the same mixture but using water instead of PBS (Na-LTL-water and Gd-LTL-water).

[0086] Example 9: Red blood cell survival rate test To confirm the lack of toxicity of the nanoparticles of the present invention, red blood cells were exposed to increasing concentrations of nanoparticles according to the present invention (from the above examples).

[0087] The results are shown in Figure 5. Therefore, with respect to nanoparticles synthesized in water, the maximum percentage of hemolysis was observed to be 1.12%, 0.649%, 0.00%, and 0.00% for K-LTL-water, Gd-LTL-water, K-LTL-PBS, and Gd-LTL-PBS nanoparticles, respectively.

[0088] Furthermore, in the presence of a large amount of nanoparticles (500 μg / ml), the maximum percentage of hemolysis obtained for these nanoparticles is 0.0% for Gd-LTL-PBS particles.

[0089] These results confirm that using PBS during nanoparticle synthesis reduces its toxicity.

[0090] Example 10: Study on oxygen release by zeolite nanoparticles in aqueous solution and low-oxygen conditions By using a low-oxygen chamber (IN VIVO2 500 (trademark), 3M) and adjusting the amount of N2, a stable and precisely controlled atmospheric gas composition with an accuracy of 0.1% O2 was achieved.

[0091] The PBS solution (Sigma-Aldrich) was equilibrated with the gas mixture contained in the hypoxic chamber over a period of one hour prior to the experiment.

[0092] A sealed tube containing 12 ml of PBS equilibrated at 37°C and a dissolved oxygen sensor (SevenGo (Duo) pro® / OptiOx®, Mettler Toledo) were used inside a hypoxic chamber. Prior to the experiment, a baseline was established by measuring the oxygen content of the PBS for 30 minutes.

[0093] The nanoparticles of the above example, pre-filled with oxygen, were then added to the system, and the oxygen dissolved in the PBS solution was measured continuously for 1 hour. The results are shown in Figure 6. K-LTL-water and / or Gd-LTL-water nanoparticles have been observed to have superior release capacity (4.07 times and 5.12 times higher, respectively, than baseline values) due to their chemical composition. In the case of K-LTL-PBS and / or Gd-LTL-PBS nanoparticles, the O2 filling within the nanoparticles was controlled. When the network extends in one direction, using PBS rather than water during particle synthesis can lead to differences in the physicochemical properties of the zeolite, such as the surface, pore volume, and charge distribution of the zeolite, which can consequently affect the interaction between the zeolite and oxygen, and thus affect oxygen release. Release kinetics over longer periods of time are of interest for more continuous reoxygenation of target tissue throughout the entire treatment protocol. A slower release rate allows cells to continuously adapt to reoxygenation and thus maintain the molecular-level processes associated with reoxygenation. This also allows for longer intervals between injections and radiation therapy sessions. Longer reoxygenation also reduces the number of injections that need to be administered.

Claims

1. The following steps, namely, - A first composition / solution 1 is prepared, comprising an aluminum source and a source of alkali metal M, particularly K, ions; - A second composition / solution 2 is prepared, comprising a silicon source and a source of alkali metal M, particularly K ions, wherein compositions / solutions 1 and 2 contain no organic structuring agents; - The compositions / solutions 1 and 2 are mixed together, and the mixture is left to stand under stirring; - The mixture is crystallized at a temperature of 50°C or higher; and - The nanoparticles formed in this manner may be separated. A method for synthesizing nanoparticles consisting of at least one zeolite nanocrystal, or containing at least one zeolite nanocrystal, The first composition / solution 1 and the second composition / solution 2 are each supplied with the aforementioned source, water, and 137 mM / L NaCl, 2.7 mM / L KCl, and 10 mM / L Na ２ HPO ４ , 1.76 mM / L KH ２ PO ４ A buffer (PBS) consisting of and having a pH of 7.2, and a mixture of the above composition / suspension 1 and 2 having the following molar composition (I), i.e. 【Chemistry 1】 A method characterized by having the following:

2. The method according to claim 1, characterized in that the nanoparticles are separated using a membrane.

3. The method according to claim 1 or 2, characterized in that M is K, the aluminum source is aluminum hydroxide, and the mixture is crystallized at a temperature of 90°C to 200°C, particularly 170°C, for a period of 4 hours or more and 16 hours or less.

4. Na ＋ , K ＋ , HPO ４ ２－ , H ２ PO ４ － , H ３ O ＋ and Cl － A nanozeolite comprising, or consisting of, LTL-type zeolite nanocrystals, characterized by further containing ions of Na, K, HPO, H, PO, H, O and Cl.

5. The nanozeolite according to claim 4, characterized in that the crystalline network further contains cations of a metal selected from Fe, Gd, Cu, and Ce, in particular in an amount of 0.2% by weight or more and 5% by weight or less, and / or a gas selected from oxygen, dinitrogen, carbon dioxide, and mixtures thereof, contained in the pore volume.

6. The aforementioned LTL-type zeolite has the following chemical formula (II), that is, 【Chemistry 2】 The nanozeolite according to claim 4 or 5, wherein x is greater than 9 and / or y is less than 27 and / or a is greater than 6.

7. The zeolite mentioned above has the following chemical formula (III), i.e. 【Transformation 3】 It has, or the following chemical formula, 【Chemistry 4】 The nanozeolite according to claim 6, characterized by having the following features.

8. 0.5 cm ３ / g or more and 1.4cm ３ Less than or equal to 0.73 cm / g, especially 0.73 cm ３ / g or 0.85cm ３ Total pore volume per g, and / or 300 m ２ / g or more and 900m ２ / g or less, especially 538m ２ / g or 490m ２ Specific surface area S of / g ＢＥＴ A nanozeolite according to any one of claims 4 to 7, characterized by having the following features.

9. The nanozeolite according to any one of claims 4 to 8, characterized by having a sequin, that is, a flattened parallelepiped shape.

10. A nanozeolite according to any one of claims 4 to 9, characterized by having an average length of 5 nm or more and 20 nm or less, and an average width of 2 nm or more and 15 nm or less.

11. A nanozeolite according to any one of claims 4 to 10, characterized in that the pore network extends in only one direction.

12. A colloidal suspension of nanoparticles, characterized by containing the nanoparticles described in any one of claims 4 to 11 in a dispersant selected from alcohol, water, phosphate-buffered saline (PBS), and mixtures thereof.

13. A pharmaceutical composition characterized by comprising, as an active product or contrast agent as a pharmaceutical, a plurality of nanoparticles comprising or containing at least one nanozeolite as described in any one of claims 4 to 11, or a suspension as described in claim 12.

14. A combination product comprising or consisting of an anticancer agent and the pharmaceutical composition described in claim 13, for administration simultaneously or at time intervals.

15. Nanoparticles according to any one of claims 4 to 11, for use as pharmaceuticals, particularly for the treatment of solid tumors, especially brain tumors, especially glioblastoma, or for use as contrast agents, especially contrast agents for magnetic resonance imaging.

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