Method for obtaining functionalized polymer surface using photosensitizer, functionalized polymer material and use thereof

Abstract

The present invention describes a method for obtaining a functionalized polymer surface (MPn-PSm) from a polymer or copolymer of an appropriate functionalization (X); the polymer or copolymer can be a halogen or leaving group, such as polyvinyl chloride (MP1) and (chloromethyl) polystyrene-Merrifield (MP2), as well as a curcumin photosensitizer (PS1), meso-tetra(aryl)porphyrin, a chlorin or bacterial chlorin (PS2) halogenated and functionalized by a nucleophilic group, particularly including P2 of the meso-imidazolyl-porphyrin, chlorin or bacterial chlorin (PS3) type. The structures of all photosensitizers in this application contain functional groups (Y) hydroxyl (OH), sulfhydryl (SH) or NH2 nucleophiles. The application also describes a process in which photosensitizer PS1-PS3 is covalently bonded to a functional polymer material MP1-MP2 by a nucleophilic substitution reaction to prepare an MPn-PSm product. The product formed prevents the proliferation of microorganisms, thereby preventing serious infections - one of the main causes of death in patients using these devices.

Description

[0001] Field of the Invention

[0002] The present invention, applicable to the field of polymer functionalization, essentially involves a method for bonding photosensitive molecules to polymer surfaces, each of which contains suitable chemical functional groups for establishing thermodynamically stable covalent chemical bonds. The resultant material consists of a pair of photosensitive polymer materials (MPn-PSm). In the dark, or preferably when illuminated by light of an appropriate wavelength, these functionalized materials promote photodynamic action, preventing the formation of microbial colonies on the surface while simultaneously eliminating microorganisms present in the medium in contact with the surface.

[0003] More specifically, the present invention is applicable to polymeric biomedical devices used in life support, such as endotracheal tubes, catheters, stylets, reservoirs, gloves, tracheostomy tubes, IV needles, nasal oxygen cannulas, endotracheal suction tubes (with and without valves), packaging, and instrument storage bags.

[0004] Invention basis

[0005] Immunosuppressed patients using these devices in hospital settings typically undergo standard procedures involving the use of antibiotics to control infection; in most cases, this leads to the emergence of bacteria that are multidrug-resistant to commercially available antibiotics.

[0006] Endotracheal tubes are an example of a common device used to assist mechanical ventilation in patients with respiratory problems, postoperative or traumatic injuries; the surfaces of such devices are prone to microbial colonization, which in most cases leads to mechanical ventilation-associated pneumonia. Data reported by Zeitoun, SS et al. (2001) showed that intubated patients faced a 21-fold higher risk of death from microbial infection than non-intubated patients, and microbial infection was one of the main causes of death in these intubated patients.

[0007] Blood bags are another example of a device used to transport and store a donor's blood until it is needed for a transfusion. Transfusion infections are very serious because they can lead to systemic infection in the patient.

[0008] Organs intended for transplantation should be transported in packaging that does not pose a risk of microbial contamination or compromise the integrity of the organ; therefore, ensuring the sterility of the medical devices used to transport and store the organs is crucial.

[0009] The present invention describes a method for combining photosensitizers (PS) of curcumin groups and derivatives (PS1), porphyrins and derivatives (chlorins and bacteriochlorins) (PS2 and PS3) with polymer surfaces (MP1 and MP2) containing appropriate functional groups (X) to generate (MPn-PSm) type products; each component of the photosensitizer (appropriate nucleophilic group) establishes a stable covalent chemical bond (ether, amine, thioether) with the polymer surface (MP1 and MP2) containing the appropriate functional group (X).

[0010] The product developed by the present invention (MPn-PSm) is stable in aqueous solutions of different pH values ​​after irradiation with light of appropriate wavelength at physiological temperature.

[0011] Products composed of polymer surfaces (MPn-PSm) functionalized with covalently bonded photosensitizers (PS) can prevent microbial infections in the absence of light, thereby reducing the risk of mortality. In the presence of light of the appropriate wavelength, this product inhibits microbial and biofilm formation through photodynamic action, thereby reducing mortality caused by infections caused by microorganisms that are multidrug-resistant to commercially available antibiotics.

[0012] Level of technological development

[0013] Photodynamic therapy (PDT) requires three elements to inactivate microorganisms: a photosensitizer (PS), a light source of appropriate wavelength, and oxygen.

[0014] Photosensitizers (PS) are chemical entities that absorb light of a specific wavelength, changing its chemical and / or physical properties. Photosensitizers (PS) absorb the energy of light by transitioning to an excited state and thereby transferring energy. There are different mechanisms by which photosensitizers (PS) act: type I and type II. In the type I mechanism, the photosensitizer (PS) reacts directly with molecules, generating reactive free radicals (ROS) and radical ions. In the type II mechanism, energy transfer occurs to molecular oxygen, generating singlet oxygen that can inactivate microorganisms (Pucelik et al. (2018) (Plos One, 13(1):e0191777).

[0015] However, no studies have shown that the effects of reactive oxygen species (ROS) generated by photodynamic therapy (PDT) can lead to microbial resistance.

[0016] Bezman et al. (1978) (Photochemistry and Photobiology, 28, 325-329, 1978) showed that the photosensitizer (PS) Rose Bengal can be covalently bonded to polystyrene and chloromethylstyrene copolymer beads through ester-type bonds to inactivate Escherichia coli by photodynamics. This ester-like covalent bond is easily hydrolyzed in biological media and under the action of light. On the other hand, the preparation method and MPn-PSm type products described in the present invention involve different photosensitizers (PS); the photosensitizers contain selected functional groups (NH2, OH, SH) to form different stable covalent bonds (amine, ether and thioether) with different polymers (including polyvinyl chloride (PVC) and Merrifield) that are stable in biological media and under light.

[0017] Patent WO 1993 / 000815 discloses a photo-bactericidal composition comprising a textile fiber-type polymer (cellulose) and a photosensitizer (porphyrin or phthalocyanine) linked solely by electrostatic interactions, for surface disinfection. The photosensitizer used is a non-functionalized meso-tetrapyridylporphyrin or phthalocyanine type, adsorbed to the polymer solely by electrostatic bonds.

[0018] Document EP1203052 / US6420455B1 discloses a polymeric composition (which exhibits surface antimicrobial activity) and articles using such a composition. However, there is no mention of a covalent bond with a photosensitizer, at least one of which is a xanthene (also known as "xanthene"). The interaction between the photosensitizer and the polymer material is purely physical (physical mixing), with some electrostatic interactions, and no interactions that are essentially chemical in nature. Although the document mentions the effect of the photosensitizer in the absence of light stimulation, the difference in the level of interaction with and without light stimulation is more obvious, because the surface functionalization proposed in the present invention involves promoting biological surface properties while maintaining the mechanical requirements and other functions of the base polymer material.

[0019] Document RU 2663061 proposes a polymer-based antimicrobial agent endowed with bactericidal properties. The molecule contains at least one nitrogen atom with a pair of free electrons, resulting in the biocide being coordinated to a metal complex via a labile electrostatic bond. The metal complex may contain a magnesium porphyrin as a central atom, in addition to other chemical groups (such as phthalocyanine). The only similarity lies in the use of porphyrin-based photosensitizers, but their structure and photodynamic properties differ from those described in the present invention. Furthermore, the invention, with respect to the polymers used, is not intended to functionalize the surfaces of biomedical devices using covalent bonding.

[0020] The novelty of the present invention is based on the functionalization of the polymer surface of polyvinyl chloride, halomethyl polystyrene, or copolymers thereof; the polyvinyl chloride, halomethyl polystyrene, or copolymers thereof containing leaving groups (bromine (Br), chlorine (Cl), iodine (I), or fluorine (F)), particularly polyvinyl chloride (MP1) or Merrifield (MP2) containing a chlorine (Cl) leaving group, bonded to a photosensitizer. These compounds contain appropriate functional groups (hydroxyl (OH), nitrogen (N-), or sulfhydryl (SH)); upon forming stable covalent bonds in biological media and under light irradiation, MPn-PSm-type products are formed; upon irradiation with light of an appropriate wavelength, the products can be used as biomedical devices with antimicrobial effects.

[0021] It is also important to emphasize that the materials obtained by the method of the present invention can significantly reduce the risk of infection and pneumonia in hospitalized and intubated patients. Therefore, polymer materials covalently functionalized with photosensitizers (PS) can promote improvements in hospital environments and bring huge benefits to public health.

[0022] Summary of the Invention

[0023] This invention describes a method for preparing a polymer product composed of a polymer or copolymer containing appropriate functional groups. The polymer product (MPn-PSm) developed according to this invention inhibits microbial growth in a dark environment and exhibits antimicrobial activity upon exposure to light of an appropriate wavelength. The polymer material (MPn-PSm) formed by irreversibly covalently bonding a photosensitizer (PSm) is particularly suitable for microbial inactivation in medical devices, endotracheal tubes, organ storage and transport packaging, blood storage and transport bags, and food storage and transport packaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to fully present the subject matter of the present invention, the accompanying drawings are as follows:

[0026] Figure 1 shows a functionalization scheme for a polymeric material (MPn) containing a leaving group (X); the leaving group can be a halogen atom, which reacts with a photosensitizer (PSm) to produce a functionalized polymeric material called (MPn-PSm); the photosensitizer can be of the curcumin type and derivatives or tetrapyrrole macrocycles, more specifically porphyrins and derivatives (dihydrochlorins or bacteriochlorins) containing a nucleophilic group (Y); the nucleophilic group can be of -OH, -N- or -SH type.

[0027] FIG2 shows data from an example of UV-Vis spectroscopy characterization, comparing a polymer material (MP1) (black curve) and a curcumin photosensitizer (PS1) (blue curve) used as an endotracheal tube functionalized with curcumin photosensitizer (MP1-PS1) (green curve).

[0028] Figure 3 shows examples of infrared spectroscopy (FT-IR) characterization of a non-functionalized endotracheal tube (ET) (black curve) with curcumin (PS1) (red curve) and a functionalized endotracheal tube (polyvinyl chloride (PVC)-curcumin, MP1-PS1) (blue curve). FT-IR characterization analysis was performed using a spectrophotometer equipped with a Smart Orbit accessory in the range of 500-4000 cm -1 Measured within the range.

[0029] Figure 4 shows examples of characterization by scanning electron microscopy (SEM): (a) an endotracheal tube composed of a solid polymeric material (MP1); (b) an endotracheal tube composed of a functionalized polymeric material (MP1) covalently bonded to curcumin (PS1) (MP1-PS1). SEM micrographs were obtained using a ZEISS LEO 440 (Cambridge, England) instrument equipped with an OXFORD detector (model 7060) operating at a 15 kV electron beam, 2.82 A current, and a 200 pA I probe. The samples were coated with 6 nm gold using a BAL-TEC MED 020 coating system metallizer (BAL-TEC, Liechtenstein) and stored in a desiccator until analysis. Metallization conditions: chamber pressure = 2.00 x 10 -2 mbar; current = 60 mA; deposition rate = 0.60 nm / s.

[0030] FIG5 shows an example of fluorescence spectroscopy characterization of an endotracheal tube (MP1-PS1) composed of covalently bonded functionalized materials in the excitation wavelength range of 350-600 nm; a) Fluorescence image obtained from the outer surface of the functionalized endotracheal tube (MP1-PS1);

[0031] FIG6 shows the results of microbiological analysis of the reduction of Staphylococcus aureus biofilm on endotracheal tubes functionalized with curcumin by covalently bonding MP1-PS1; orange ( / / / ) represents the percentage of bacterial growth reduction in biofilms on the surfaces of endotracheal tubes (control) and endotracheal tubes functionalized with curcumin by covalently bonding MP1-PS1 (orange) in a dark environment; red ( / / / ) represents the percentage of bacterial growth reduction after irradiation with an LED light source from the 450 nm region for 12 minutes (a total of 50 J / cm 2 ) after treatment with curcumin.

[0032] FIG7 is a UV-visible spectrum of solutions formed by immersing the polymer material MP1-PS1 under different pH conditions (pH values ​​of 2, 7, and 10).

[0033] Detailed description of the invention

[0034] The present invention relates to functionalizing polymeric or copolymeric materials by covalently bonding them with photosensitizers (PSs); the polymeric or copolymeric materials exhibit antimicrobial activity in the dark or, preferably, under illumination (PDT), contributing to the reduction of microbial adhesion and inactivation in biomedical devices made with these materials. These polymers are preferably from the class of vinyl polyhalides; wherein the halide or halogen is a diatomic molecule of an element from Group 17 of the Periodic Table containing a leaving group of fluorine (F), chlorine (Cl), bromine (Br), or iodine (I), preferably chlorine (Cl).

[0035] The present invention describes a method for obtaining functionalized polymer surfaces (MPn-PSm) from polymers or copolymers of suitable functionalization (X); the polymers or copolymers may be halogens or leaving groups, in particular polyvinyl chloride (MP1) and (chloromethyl) polystyrene-Merrifield (MP2), as well as photosensitizers (PSs) and derivatives (PS1) of the curcumin type, meso-tetraarylporphyrins and derivatives, chlorins and bacteriochlorins halogenated and functionalized with nucleophilic groups (PS2), in particular those of the meso-imidazolyl-porphyrin type and derivatives (chlorins and bacteriochlorins) (PS3). All photosensitizers (PSs) of the present invention contain a functional group (Y) in their structure - a nucleophile of the hydroxyl (OH), thiol (SH) or NH2 type.

[0036] Specifically, the present invention describes a method for preparing MPn-PSm products by covalently bonding (D) photosensitizers (PSs) (PS1, PS2, and PS3) to functionalized polymers MP1 and MP2 via a nucleophilic substitution reaction in the form of ether, amine, or thioether bonds. Furthermore, the present invention relates to the use of the MPn-PSm polymer products for reducing microbial adhesion and inactivating microorganisms via photodynamic therapy (PDT). Specific applications include probes, catheters, reservoirs, tracheostomy tubes, intravenous needles, nasal oxygen cannulas, hemodialysis catheters, rectal probes, packaging for organ transport and storage, urethral probes, and tracheal suction probes for use by humans and animals in hospital settings.

[0037] Products formed from the MPn-PSm polymer surfaces developed according to the present invention can prevent microbial proliferation, which is the cause of many serious infections and currently one of the leading causes of death in hospitalized patients using these devices. Furthermore, these MPn-PSm polymer materials have important applications in the preparation of food storage packaging.

[0038] The method proposed in the present invention can simply and economically obtain MPn-PSm type polymer materials functionalized with photosensitizers; in the MPn-PSm, n is 1-2 and m is 1-3, namely MP1-PS1, MP1-PS2, MP1-PS3, MP2-PS1, MP2-PS2 and MP2-PS3 (according to Table 1); the polymer materials are obtained by direct nucleophilic substitution reaction between commercially available MP1 or MP2 type polymer materials and each of PS1, PS2 or PS3 type photosensitizers; the MP1 or MP2 type polymer materials contain a leaving group in their structure, in particular a halogen (fluorine, chlorine, bromine or iodine); the PS1, PS2 or PS3 type photosensitizers contain a nucleophilic group OH, SH or N-.

[0039] Due to the structure of the polymer material MP1 (polyvinyl chloride (PVC) with the general formula [C2H3Cl]n) or MP2 [(chloromethyl)polystyrene with the general formula [C9H9Cl]n], the presence of the halogen leaving group is easily identifiable.

[0040]

[0041] Source of photosensitizer

[0042] Photosensitizer PS1 with a hydroxyl nucleophilic group [having the functionality of a nucleophilic reagent, commonly known as "curcumin", and the International Union of Pure and Applied Chemistry (IUPAC) name is (1E,6E)-1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione] and its derivatives:

[0043]

[0044] Photosensitizers of the PS2 and PS3 families include porphyrins functionalized with hydroxyl, amine or thiol nucleophilic groups (Formula II to Formula IX; two β-pyrrole positions (---) in the form of double bonds) and their reduced derivatives, chlorins (one position (---) in the form of a single bond and the other position (---) in the form of a double bond) and bacterial chlorins (two positions (---) in the form of single bonds).

[0045] Symmetrical porphyrins (Formulas II to IV) with double bonds at two positions (---) are synthesized according to the nitrobenzene or nitrobenzene-NaY method; the method comprises mixing pyrrole with four equivalents of a halogenated aldehyde having a selected structure and containing a nucleophile Y (OH, N-, or SH) at one of the other positions at a temperature between 100° C. and 140° C. under aerobic conditions, using a mixture of acetic acid or propionic acid and nitrobenzene as a solvent, and using a reusable NaY zeolite-type catalyst with or without the use of a catalyst.

[0046] After hot NaY filtration, the porphyrin is precipitated directly from the reaction medium after cooling, or precipitated after adding methanol, or purified by flash chromatography to obtain a symmetrical photosensitizer of the PS2 family (Formula II to Formula IV). Between 100°C and 140°C, under aerobic conditions, using a mixture of acetic acid or propionic acid and nitrobenzene as a solvent, without or with a reusable catalyst of the NaY zeolite type, according to the nitrobenzene or nitrobenzene-NaY method, one of the other positions contains a nucleophile Y (OH, N- or SH), 2 equivalents of a halogenated aldehyde with a selected structure are mixed with 2 equivalents of formaldehyde or the corresponding acetal and 4 equivalents of pyrrole to synthesize an asymmetric halogenated porphyrin (Formula V to Formula VII) in which both positions (---) are in the form of a double bond. After hot NaY filtration, the porphyrin is purified by preparative flash chromatography to obtain an asymmetric photosensitizer of the PS2 family (Formula V to Formula VII):

[0047]

[0048] Represents a carbon-carbon single or double bond.

[0049] Symmetrical porphyrins (Formula VII, in which both positions (---) are double bonds) are synthesized using the nitrobenzene or nitrobenzene-NaY method. The method comprises mixing 4 equivalents of 1-methyl-2-imidazolecarboxaldehyde with 4 equivalents of pyrrole at a temperature between 100°C and 140°C in the presence of oxygen, using a mixture of acetic acid or propionic acid and nitrobenzene as a solvent, with or without a reusable zeolite-type NaY catalyst. After filtration of the hot NaY, the porphyrin (Formula VIII) is purified by flash chromatography.

[0050] An asymmetric porphyrin (Formula IX) containing double bonds at both (-) positions is synthesized using a nitrobenzene or nitrobenzene-NaY method. The method comprises mixing two equivalents of 1-methyl-2-imidazolecarboxaldehyde with two equivalents of formaldehyde or its acetal and four equivalents of pyrrole at a temperature between 100°C and 140°C in the presence of oxygen, using a mixture of acetic acid or propionic acid and nitrobenzene as a solvent, with or without a reusable NaY zeolite catalyst. After filtration through the hot NaY, the porphyrin (Formula IX) is purified by flash chromatography.

[0051]

[0052] Represents a carbon-carbon single or double bond.

[0053] Metal complexes of PS2- or PS3-type porphyrins (Formulas II to VII; VIII and IX) are prepared by mixing solutions of the corresponding porphyrins dissolved in a suitable solvent, preferably chloroform or dimethylformamide (DMF), at temperatures between 40°C and 150°C, to which a saturated solution of a suitable metal salt (Zn(OAc)2, Pd(OAc)2, or AlCl3) is added. After the complexation reaction is complete, the reaction mixture is purified by sequential washing with saturated aqueous solutions of sodium bicarbonate and water. If necessary, the metal complex is purified by flash chromatography.

[0054] The corresponding dihydrochlorins with a single bond at one position (---) and a double bond at the other position (---) were synthesized according to the method described by Pereira MM et al. One of the porphyrin-type precursors (Formulas II to IX) prepared according to the above method was mixed with a small excess of p-toluenesulfonylhydrazine (15 equivalents) in a Schlenk tube in the solid state and then placed under a vacuum of 0.1 bar for 1 hour. The mixture was then heated to 120°C to 140°C, with the heating time optimized for each substrate. The reaction mixture was dissolved in a very small amount of organic solvent and washed sequentially with sodium hydroxide and water. The resulting solid was dissolved in dimethyl ether (DME), FeCl3.6H2O (1 equivalent) was added to the solution, and then hydrogen peroxide (aqueous solution, 3%) was slowly added. The reaction was terminated when the bacterial dihydrochlorin absorption peak (approximately 750 nm) disappeared. The corresponding chlorins (Formula II to Formula VII; wherein one position (---) is in the form of a single bond and the other position (---) is in the form of a double bond) were washed and then purified by flash chromatography.

[0055] The corresponding bacterial chlorins PS2 and PS3, in which the (---) position is a single bond, were synthesized according to the method described by Pereira MM et al., wherein the porphyrins were mixed with an excess of p-toluenesulfonylhydrazine (40 equivalents) in a Schlenk tube and then placed under vacuum (0.1 bar) for 1 hour. The mixture was then heated at 140°C for a time optimized for each porphyrin (Formula II to Formula VII, in which (---) is a double bond). After cooling to room temperature, the corresponding bacterial chlorins (Formula II to Formula IX, in which (---) is a single bond) were purified by washing or flash chromatography.

[0056] Table 1 discusses all possible combinations of polymeric materials and photosensitizers (MPn-PSm) and their possible structures, where the D component is always an ether (O), thioether (S) or amine (N-) covalent bond (Table 1):

[0057] Table 1 - Possible combinations of polymer materials and photosensitizers (MPn-PSm) and their possible structures

[0058]

[0059]

[0060]

[0061]

[0062] Table 2 discusses all possible substituents (R1, R2, Z, Z' and M) for the MPn-PSm combination embodied in Table 1:

[0063] Table 2 - MPn-PSm combinations with possible substituents R1, R2, Z, Z' and M

[0064]

[0065]

[0066] Method for preparing MPn-PSm type functionalized polymer products using photosensitizer (PS)

[0067] All photosensitizers (PSs) (PS1, PS2, PS3) used in the present invention contain nucleophiles (OH, N-, or SH) with specific chemical properties in their structures. Therefore, a simple and efficient method can be developed to form covalent bonds with polymer materials (MP1 and MP2) containing halogen-type leaving groups in their structures through a nucleophilic substitution reaction through the following steps:

[0068] A photosensitizer (PS) selected from the PS1, PS2 or PS3 family, comprising an organic base (triethylamine, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), 1,5-diazabicyclo[4.3.0]non-5-ene (DBN) or pyridine) or preferably an inorganic substance (CaCO3, CeCO3, NaOH, KOH, Ba(OH)2, Al(OH)3, Mg(OH)2, Be(OH)2, Ca(OH)2) is dissolved in a solvent of the dimethylformamide (DMF), dimethoxyethane (DME), tetrahydrofuran (THF) and its derivatives, dimethylpyrrolidone, dichloromethane, ethyl acetate or preferably dimethyl sulfoxide (DMSO) type to form a solution referred to hereinafter as "Sol A".

[0069] The polymer material MP1 or MP2 is immersed in the Sol A solution and placed at a temperature between 0°C and 40°C for 0.5 to 48 hours, preferably in the absence of oxygen. The polymer material (MPn-PSm) covalently linked to the desired photosensitizer (PS) is then removed from the Sol A solution and washed 1 to 10 times with an organic solvent (preferably dimethyl sulfoxide (DMSO)) and then with a low-boiling point organic solvent (preferably ethanol).

[0070] The MPn-PSm is dried (preferably at room temperature under vacuum) and stored in the presence or preferably in the absence of oxygen and light.

[0071] 2 of the Examples, the functionalized polymer products prepared according to the present invention [hereinafter referred to as MP1-PS1, MP1-PS2 (Formula II to Formula VII), MP1-PS3 (Formula VIII and Formula IX), MP2-PS1, MP2-PS2 (Formula II to Formula VII) and MP2-PS3 (Formula VIII and Formula IX)] were characterized by Cary 5000 UV-Vis diffuse transmittance spectroscopy, confirming that the photosensitizer (PS) was bonded to the polymer material due to the presence of typical absorption bands of each photosensitizer (PS) in the polymer material.

[0072] The functionalized polymer materials of the types MP1-PS1, MP1-PS2, MP1-PS3, MP2-PS1, MP2-PS2 and MP2-PS3 prepared according to the present invention were characterized by infrared spectroscopy on a Nicolet 5700 (ThermoElectron Corporation) FTIR spectrometer (equipped with a Smart Orbit accessory) to observe the typical vibration bands of the polymer and the typical vibration bands of the specific functional groups of each photosensitizer (PS) bound to the polymer, as shown in Figure 3.

[0073] Functionalized polymeric materials based on MP1-PS1, MP1-PS2, MP1-PS3, MP2-PS1, MP2-PS2, and MP2-PS3 prepared according to the present invention were characterized by scanning electron microscopy (SEM) operating at a 15 kV electron beam, 2.82 A current, and a 200 pA1 probe. The samples were coated with 6 nm of gold and stored in a desiccator until analysis. The analysis revealed the presence of the photosensitizer (PS) on the polymer surface ( Figure 4 ).

[0074] The functionalized polymeric materials of the type MP1-PS1, MP1-PS2, MP1-PS3, MP2-PS1, MP2-PS2 and MP2-PS3 prepared according to the present invention were characterized by fluorescence spectra obtained on the exterior of the catheter. Figure 5 shows the typical fluorescence emission of curcumin at 550 nm on the exterior surface of the material MP1-PS1.

[0075] MPn-PSm polymer materials inhibit biofilm growth

[0076] The MP1-PS1, MP1-PS2, MP1-PS3, MP2-PS1, MP2-PS2 and MP2-PS3 polymer products prepared according to the present invention and functionalized by sensitizers inhibit the growth of Gram-positive bacteria (particularly Staphylococcus aureus) (Figure 6) biofilms or Gram-negative bacteria (particularly Escherichia coli and Pseudomonas aeruginosa) biofilms in the dark, or preferably in the presence of a device emitting light of an appropriate wavelength [i.e., visible light (370 to 700nm) or near-infrared light (700 to 850nm)]. The microorganisms forming the biofilm are incubated in a suspension for 24 hours and then exposed to light of an appropriate wavelength emitted from the device; the suspension (MPn-PSm material) comprises a polymer material (MP1 or MP2) covalently linked to a selected PS1, PS2, PS3 type photosensitizer (PS). In order to determine the efficiency of photodynamic inactivation, a method for recovering cells by colony forming units (CFU / mL) is adopted. The results show that for each photosensitizer (PS), the MPn-PSm polymer material effectively reduced microbial adhesion and inhibited microbial biofilm growth in the absence of light or, preferably, when irradiated with light of an appropriate wavelength. Figure 6 of Example 1 shows microbial inactivation in the dark (56%) and after irradiation with light of 450 nm (98%). This example demonstrates that the MPn-PSm material effectively promotes biofilm inactivation in a dark environment through photodynamic action.

[0077] Example 1: Covalent bonding of curcumin (PS1) to an endotracheal tube (ET) made of polyvinyl chloride (PVC) (MP1) Method for preparing polymer material MP1-PS1

[0078] A solution designated as "Sol A" in the present invention was prepared by dissolving curcumin (396 mg; 1.07 mmol) and Cs2CO3 (1.99 g; 6.01 mmol) in dimethyl sulfoxide (DMSO; 80 mL).

[0079] An endotracheal tube (ET) made of polymer material MP1 (11 g) was weighed and immersed in sol A at 30° C. to 40° C. for 4 to 8 hours in an environment with nitrogen or argon;

[0080] The functionalized endotracheal tube composed of MP1-PS1 was removed from the solution and initially washed with dimethyl sulfoxide (DMSO) (4 times, 20 mL) and finally with ethanol (4-10 times, 20 mL) until no curcumin was observed by UV-visible spectroscopy.

[0081] The functionalized endotracheal tube (MP1-PS1) is dried for 1 to 3 days, preferably at room temperature under vacuum.

[0082] Store at room temperature in an oxygen-free, light-free environment.

[0083] The curcumin-functionalized endotracheal tube (PS1), designated MP1-PS1 material, was characterized by various techniques as follows:

[0084] UV-Vis Absorption Spectra: Figure 2 shows the UV-Vis absorption spectrum of a curcumin-functionalized endotracheal tube (Figure 2, green curve); the spectrum displays a band at 430 nm typical of the absorption spectrum of non-immobilized curcumin in ethanol solution (Figure 2, blue curve). Figure 2 also shows the UV-Vis absorption spectrum of an endotracheal tube (TE), which does not absorb in this region (Figure 2, black curve).

[0085] Fourier Transform Infrared Spectroscopy (FTIR): In Figure 3, the binding of curcumin to the endotracheal tube (TE) is confirmed by infrared analysis of the curcumin-functionalized catheter (MP1-PS1, blue curve) compared to the endotracheal tube (TE, black curve) and curcumin (PS1, red curve). The spectrum of curcumin is at 3509 cm –1 (hydroxyl-OH), 1600-1650cm –1 (C=O), 1509cm –1 (C=C ethylene) (carbon=carbon ethylene), 1250cm –1 The spectrum of the curcumin-functionalized endotracheal tube (MP1-PS1, blue curve) compared with the spectra of non-immobilized curcumin (PS1) and endotracheal tube (TE) showed a peak at the following position: 3506 cm –1 (hydroxyl-OH), 1600-1650cm –1 (C=O), 1512cm –1 (C=C ethylene) (carbon=carbon ethylene), confirming the presence of curcumin covalently bonded to the endotracheal tube (TE).

[0086] Scanning electron microscopy: The bonding of curcumin to the endotracheal tube (ET) was also confirmed by scanning electron microscopy (SEM) of the endotracheal tube (ET) ( Figure 4 , a) and scanning electron microscopy (SEM) of the curcumin functionalized endotracheal tube (MP1-PS1) ( Figure 4 , b), in which the presence of curcumin was detected on the surface of the endotracheal tube ( Figure 4 , b).

[0087] Stability of curcumin-functionalized endotracheal tube (PS1) designated MP1-PS1

[0088] The stability of the curcumin-functionalized endotracheal tube (MP1-PS1) was confirmed by analyzing the UV-visible absorption spectra at different pH values ​​(2, 7, and 10) of the simulated biological system over time (Figures 7a-c). No release of curcumin from the endotracheal tube was observed at any of the pH values ​​evaluated.

[0089] Curcumin Catheter Microbiome

[0090] The microorganism used was Staphylococcus aureus (ATCC 25925). The inoculum was prepared in a 15 mL Falcon tube containing Brain Heart Infusion (BHI) growth medium and bacterial inoculum at a ratio of 9:1. The pre-inoculum was incubated in a rotary oven at 37°C and 140 rpm for 15 hours. For biofilm formation, the following steps were taken:

[0091] - Separate the microorganisms in the pre-inoculum from the culture medium by centrifugation (1500 rpm, 15 minutes);

[0092] - Wash twice with phosphate-buffered saline (PBS) by centrifugation (1500 rpm, 15 min);

[0093] — Sterile endotracheal tubes were cut into 1 cm long pieces in a sterile environment (laminar flow, air flow, Esco Class II bsc);

[0094] Insert each piece of the previously cut endotracheal tube into each well of a 24-well plate.

[0095] Next to the well containing the cut tube, add 900 μL of liquid culture medium and 100 μL of bacterial inoculum.

[0096] In each well of the plate, homogenize the solution six times using a 1000 μL pipette.

[0097] ——The formed biofilm was characterized by colony count (CFU / mL).

[0098] The experimental lighting was provided by Technological Support Laboratory-LAT / USP ( The light source used in the MP1-PS1 irradiation experiments emits 450 nm irradiation and is designed for irradiation in a 24-well plate at 70 mW / cm 2 Irradiation was carried out uniformly and continuously for 12 minutes. Irradiation was measured with a potentiometer; the collector radius was 0.4 cm and the total area was 0.5 cm. 2 In order to 2 To calculate irradiance for a unit, use the following formula:

[0099] I = P / A;

[0100] Where I = potentiometer irradiance; P = power measured by the potentiometer, and A = area of ​​the potentiometer.

[0101] To calculate the illumination time required to achieve the desired energy dose, the following formula is used:

[0102] T = D / I;

[0103] In the formula, T = lighting time, D = required energy dose, and I = calculated LED irradiance.

[0104] During the illumination period, the samples were protected with aluminum foil to avoid any possible external influences.

[0105] After incubating the samples at 37°C for 24 hours, count the bacterial colonies in the Petri dishes using solid BHI medium and visually observe the bacteria. Count each experimental group three times, counting 3 to 30 colonies. The mean value for each group is calculated in CFU / mL using the following formula:

[0106] CFU / mL = (number of colonies x number of dilutions) / volume;

[0107] The number of colonies was the average value obtained by counting the colonies on the experimental plates three times.

[0108] After removing planktonic cells by washing all tubes with PBS, they were divided into four experimental groups: biofilms formed on the surface of endotracheal tubes (ET); biofilms formed on the surface of curcumin-functionalized tubes (MP1-PS1); and biofilms formed on the surface of endotracheal tubes (ET) at 50 J / cm 2 Biofilm formed on ET surface under light dose irradiation; at 50J / cm 2 Biofilm formed on the surface of MP1-PS1 under light dose irradiation. Bacterial biofilm was removed from the catheter surface by mechanical agitation using phosphate-buffered saline (PBS), and the antimicrobial activity of each group was evaluated using the plate method on solid agar for colony count.

[0109] Therefore, this example is to prepare an endotracheal tube (ET) functionalized with curcumin (named MP1-PS1 material) for use in patients who require mechanical ventilation and are usually bedridden, with or without photodynamic therapy (PDT) to inactivate microorganisms and hinder the formation of microbial biofilms.

[0110] In a preferred embodiment of the present invention, the above-mentioned endotracheal tubes (ET) functionalized with curcumin (designated MP1-PS1 material) are able to reduce microbial adhesion and inactivate microorganisms; for this purpose, a light source of a specific wavelength (450 nm) is used that facilitates the decontamination process of these tubes.

Claims

1. A method for obtaining a functionalized polymer surface, comprising the following steps: a) dissolving a photosensitizer PS having a nucleophilic group (Y) in a base and an organic solvent selected from dimethylformamide (DMF), dimethoxyethane (DME), tetrahydrofuran (THF), dimethylpyrrolidone, dichloromethane, ethyl acetate or dimethyl sulfoxide (DMSO) to form a solution, wherein the photosensitizer PS having a nucleophilic group (Y) is curcumin; b) immersing the surface of a biomedical device comprising a polymer material having a leaving group (X) in the solution obtained in step (a), wherein the immersion is performed at a temperature in the range of 0° C. to 40° C., wherein the polymer material having a leaving group (X) is polyvinyl chloride (MP1); c) allowing a nucleophilic substitution reaction to occur between the leaving group (X) and the nucleophilic group (Y), the reaction occurring in an organic solvent for 0.5 to 48 hours to form a surface functionalized polymer; d) Drying the obtained surface.

2. The method according to claim 1, wherein the drying step (d) is carried out under vacuum at room temperature.

3. The method according to claim 1, wherein the base used in step (a) is selected from the group consisting of triethylamine, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), 1,5-diazabicyclo[4.3.0]non-5-ene (DBN), pyridine, CaCO3, CeCO3, NaOH, KOH, Ba(OH)2, Al(OH)3, Mg(OH)2, Be(OH)2 and Ca(OH)2.

4. The method according to claim 1, wherein the organic solvent used in step (a) is dimethyl sulfoxide (DMSO).

5. The method according to claim 1, further comprising, after step (c), a step of washing with an organic solvent having a boiling point lower than dimethyl sulfoxide (DMSO). The method according to claim 5 , wherein the organic solvent is ethanol.

7. A surface consisting of at least one polymer surface MP1 obtained by the method according to any one of claims 1 to 6, characterized in that The halogen leaving group on the surface is combined with the photosensitizer through a stable chemical bond (D) of ether type (O), thioether type (S) or amine type (NH).

8. The surface according to claim 7, wherein MP1 is polyvinyl chloride (PVC) having a leaving group (X), wherein X is a halogen atom.

9. The surface according to claim 7, characterized in that It exhibits antimicrobial effects in a lightless environment, and this effect is more pronounced under irradiation with light of a specific wavelength in the range of 400 to 850 nm.

10. The surface according to claim 7, characterized in that It is used to prepare and produce polymeric biomedical devices.

11. The surface of claim 10, wherein the biomedical device is a medical device selected from the following list: a catheter, a probe, a reservoir, a tracheostomy tube, an IV needle, packaging for transporting and preserving organs.

12. The surface according to claim 10, wherein the biomedical device is a medical device selected from the following list: endotracheal tube, nasal cannula for oxygen administration, hemodialysis catheter, rectal probe, urethral probe, probe for endotracheal suction.

13. The surface of claim 10, wherein the biomedical device is an endotracheal tube.

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