Classifications

• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/44—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating
  • C23C16/50—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges
  • C23C16/503—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges using DC or AC discharges
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M23/00—Constructional details, e.g. recesses, hinges
  • C12M23/20—Material Coatings
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B05—SPRAYING OR ATOMISING IN GENERAL; APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D—PROCESSES FOR APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D1/00—Processes for applying liquids or other fluent materials
  • B05D1/62—Plasma-deposition of organic layers
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  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M23/00—Constructional details, e.g. recesses, hinges
  • C12M23/02—Form or structure of the vessel
  • C12M23/10—Petri dish
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M3/00—Tissue, human, animal or plant cell, or virus culture apparatus
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/02—Pretreatment of the material to be coated
  • C23C16/0227—Pretreatment of the material to be coated by cleaning or etching
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/02—Pretreatment of the material to be coated
  • C23C16/0227—Pretreatment of the material to be coated by cleaning or etching
  • C23C16/0245—Pretreatment of the material to be coated by cleaning or etching by etching with a plasma
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/22—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the deposition of inorganic material, other than metallic material
  • C23C16/30—Deposition of compounds, mixtures or solid solutions, e.g. borides, carbides, nitrides
  • C23C16/40—Oxides
  • C23C16/401—Oxides containing silicon
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/44—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating
  • C23C16/455—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating characterised by the method used for introducing gases into reaction chamber or for modifying gas flows in reaction chamber
  • C23C16/45502—Flow conditions in reaction chamber
  • C23C16/4551—Jet streams
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/44—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating
  • C23C16/455—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating characterised by the method used for introducing gases into reaction chamber or for modifying gas flows in reaction chamber
  • C23C16/45563—Gas nozzles
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/44—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating
  • C23C16/50—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges
  • C23C16/505—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges using radio frequency discharges
  • C23C16/509—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges using radio frequency discharges using internal electrodes
  • C23C16/5093—Coaxial electrodes
• • C—CHEMISTRY; METALLURGY
  • C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
  • C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
  • C23C16/00—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes
  • C23C16/44—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating
  • C23C16/50—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges
  • C23C16/513—Chemical coating by decomposition of gaseous compounds, without leaving reaction products of surface material in the coating, i.e. chemical vapour deposition [CVD] processes characterised by the method of coating using electric discharges using plasma jets
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B05—SPRAYING OR ATOMISING IN GENERAL; APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D—PROCESSES FOR APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D2201/00—Polymeric substrate or laminate
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B05—SPRAYING OR ATOMISING IN GENERAL; APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D—PROCESSES FOR APPLYING FLUENT MATERIALS TO SURFACES, IN GENERAL
  • B05D2203/00—Other substrates
  • B05D2203/30—Other inorganic substrates, e.g. ceramics, silicon
  • B05D2203/35—Glass

Definitions

• PRO-BIOFILM COATING METHOD FOR ITS PRODUCTION AND SUBSTRATE
• the present invention relates to the field of bacterial control, more specifically the control of bacterial biofilms, and in particular to the promotion of the appearance of bacterial biofilms.
• Biofilm is a grouping of microorganisms surrounded by a matrix of extracellular polymeric substances that include water, polysaccharides, proteins, lipids, nucleic acids, and other biopolymers. This extracellular matrix favors the adhesion of microorganisms to surfaces, protects them from adverse environmental conditions and antimicrobial agents, and helps them capture nutrients from the environment to facilitate microbial proliferation.
• biofilm takes place in the following stages: an initial fixation stage in which bacterial cells adhere to and colonize solid surfaces. It is followed by the proliferation and maturation stages in which, through the activation of intercellular communication systems, microcolonies and the extracellular matrix begin to form, reaching an optimal population density and a mature biofilm structure. Finally, the dispersal stage occurs in which bacterial cells or components of the biofilm can separate from it, migrate and colonize other surfaces, which is a persistent source of dissemination and contamination.
• Biofilms deteriorate products or surfaces on which they develop, but also, when they are formed by microorganisms of a pathogenic nature, they are a cause of great concern in the food, pharmaceutical or clinical industries since they represent a source of contamination of products and a risk for public health.
• the elimination of pathogenic biofilm plays a vital role to guarantee an optimal state of quality microbiological food.
• the biofilm acts as a reservoir that can protect or release pathogenic bacteria in the free state (planktonic), ubiquitous organisms that are especially difficult to control, resulting in bacterial persistence in the production plant and episodes of cross-contamination of food, with obvious risks for food safety.
• JP2013173715A discloses a method of biofilm formation by plasma irradiation treatment of a surface such as a polycarbonate plate.
• the plasma treatment produces a chemical modification of the polycarbonate that favors the adhesion of microorganisms that can form a first biofilm.
• This first biofilm in turn favors the adhesion of new microorganisms that will form a new biofilm.
• the chemical modification of the surface by plasma treatment depends on the material that constitutes said surface, and therefore not all surfaces can be favorably modified by said method.
• the adhesive ability of species generated on the polycarbonate surface by plasma treatment gradually decreases over time.
• the present invention provides, in a first aspect thereof, a pro-biofilm coating applied by polymerization of a precursor by cold atmospheric plasma on a substrate.
• the coating according to the present invention has such a roughness that it promotes the creation of more than 100% of biofilm on the substrate, being 100% of the biofilm produced on the same substrate lacking said pro-biofilm coating.
• the present invention provides a method of producing a pro-biofilm coating as defined in the first aspect of the invention.
• the method comprises applying a flow of cold atmospheric plasma and precursor material on a surface of the substrate to be coated until obtaining a roughness such that it promotes the creation of more than 100% of biofilm on the substrate.
• the invention also relates to a substrate that is covered by a pro-biofilm coating according to the first aspect of the invention.
• Figure 1 is a schematic cross-sectional view of a device for carrying out a pro-biofilm coating production method according to the preferred embodiment of the present invention.
• Figure 2 is a graph showing the relationship between passes, roughness of the coating and% of biofilm produced with respect to the control, using polystyrene as substrate and (3-aminopropyl) triethoxysilane (APTES) as precursor material, according to a preferred embodiment of the present invention.
• APTES (3-aminopropyl) triethoxysilane
• Figures 3A and 3B show atomic force microscopy (AFM) and scanning electron microscopy (SEM) images, respectively, of coatings obtained according to a preferred embodiment of the present invention, in which the percentage of biofilm is also indicated. obtained with respect to the control.
• AFM atomic force microscopy
• SEM scanning electron microscopy
• Figure 4 is a graph that shows the relationship between the roughness of the coating and the% of biofilm obtained with respect to the control, using polystyrene as substrate and APTES as precursor material, according to a preferred embodiment of the invention.
• Figure 5 is a graph that shows the relationship between the roughness of the coating and the% of biofilm obtained with respect to the control according to another preferred embodiment of the invention in which a mixture of polyethylene glycol methyl ether methacrylate (PEGMA) and isopropanol (IPA) as a precursor material.
• PEGMA polyethylene glycol methyl ether methacrylate
• IPA isopropanol
• Figure 6 shows SEM images of coatings obtained according to another preferred embodiment of the present invention, in which the percentage of biofilm obtained with respect to the control is also indicated.
• Figure 7 is a graph showing the relationship of the atomic percentage of carbon (C), oxygen (O), silicon (Si) and nitrogen (N) with respect to the number of passes according to an embodiment of the present invention.
• Figure 8 shows graphs representing the deconvolution of the high resolution spectrum of carbon for different numbers of passes according to a preferred embodiment of the invention.
• Figure 9 is a graph representing the relationship between coating roughness and water contact angle of coatings according to a preferred embodiment of the invention.
• Figure 10 is a graph showing bacterial growth versus time with different coatings obtained in accordance with a preferred embodiment of the present invention.
• FIG 11 is a schematic of the biofilm over-production mechanism (PRO-Biofilm) that occurs in the coating of the present invention.
• Figure 12 represents SEM (Scanning Electron Microscopy) images of: [a] Uncoated Substrate (x2000), [b] 72p Coating (x2000) and [c] 72p Coating (x4000).
• Figure 13 shows a diagram of biofilm overproduction as roughness increases with passes: [a] No Op coating, [b] 4 passes 4p, [c] 12 passes, 12p and [d] 72 passes , 72p.
• Figure 14 shows a diagram of the operation of the APTES molecule.
• Figure 15 is a graph showing the amount of biofilm produced over time with a coating according to a preferred embodiment of the present invention compared to the control.
• Plasma is the state that a gas reaches when it is given an amount of energy that manages to ionize its molecules and atoms. That is, the passage of matter from a gaseous state to a plasma state occurs through a dissociation of molecular bonds, accompanied by an increase or decrease in the electrons of the atoms, which gives rise to the formation of ions with a positive charge or negative. Depending on whether or not there is a thermal equilibrium between the plasma particles, the thermal plasma is distinguished from the cold.
• a cold or unbalanced plasma is one in which the temperature of the electrons (105-5000 ° C) is much higher than that of the heavier particles (neutral particles and ions), which are at temperatures close to that of the ambient (25-100 ° C). In this way, the temperature of a cold plasma is generally kept below 100 ° C, which allows its use in surface treatments on a wide variety of materials without causing their deterioration due to excessive heating.
• cold plasma can be carried out at atmospheric pressure in an open environment, that is, it does not require the use of vacuum systems or chambers within which specific conditions are established. These characteristics provide cold atmospheric plasma technology with great versatility, relative simplicity and low cost. From the point of view of its industrial application, plasma has become an important tool for carrying out a multitude of surface treatments.
• plasma polymerization One of the main modifications that the surface of a plasma treated substrate can undergo is "plasma polymerization”. This modification consists of the deposition of thin coatings using monomers in liquid state as precursors through their exposure to plasma flow.
• the plasma polymerization method was used using unbalanced or cold atmospheric plasma equipment (APPJ).
• APPJ unbalanced or cold atmospheric plasma equipment
• the APPJ equipment used (see figure 1) consists of two coaxial electrodes (10, 12), between which the gas (14) circulates to generate the plasma (in this specific case, nitrogen (N2) was used with a flow of 80 slm).
• the inner electrode (10) is connected to ground, while the outer electrode (12) is excited with a certain frequency (high voltage current) by a generator (16) with a power of 300 W.
• the precursor material (18) (1.5 slm of N2 that atomizes and transports the liquid precursor material) is introduced to the plasma actuation zone (20).
• the pro-biofilm coating production method first comprises the step of activating the surface of the Petri dish (22) (or other substrate to be coated ) by a plasma jet (e.g. N2 plasma) without polymerizable precursor material.
• a plasma jet e.g. N2 plasma
• This activation and cleaning of the surface is preferably carried out by 4 passes of plasma.
• this previous step of surface activation may be omitted.
• the method preferably also comprises the step of performing, simultaneously with the application of the plasma flow, a relative displacement between the substrate to be coated and the plasma flow to coat the entire surface of the substrate.
• the flow of plasma (20) projected on the surface (which transforms, transports and projects the precursor material on the base of the Petri dish) has approximately 10mm diameter. Therefore, for the homogeneous application of the coating (preferred embodiment) throughout the base of the Petri dish (22), this jet must travel throughout the same (30 mm in diameter). Since the flow of plasma (80 slm of N2) and of the precursor (1.5 slm of N2 that atomizes and transports the precursor material) are constant, to apply a homogeneous coating, this displacement must be carried out at a constant speed.
• the Petri dish (22) rotates and moves (on an axis) simultaneously with the application of the plasma flow (20). Meanwhile, the plasma application equipment remains stationary.
• the Petri dish can remain stationary while a plasma application nozzle is moved.
• both the Petri dish and the plasma application nozzle can move simultaneously relative to each other.
• the linear (tangential) speed of coating (Vt) is constant (10 mm / s). Each time a turn is covered (at a certain radius) the turning speed (W) is modified (it is reduced at the edge, it is increased in the center) so that the linear speed (Vt) always remains constant. For each complete revolution made by the Petri dish (22), it moves along the axis a certain advance distance (24). Keeping the linear speed (Vt) constant allows the applied coating to be homogeneous.
• FDA fluorescein diacetate
• FIG. 2 shows the relationship between the roughness of APTES-based coatings and the% biofilm produced in each of them. Coatings that generate less biofilm than that produced in the control sample (the one that has not been coated, Op, equivalent to 100% biofilm) are defined as anti-biofilm ( ⁇ 100%), while those that generate more than 100 % biofilm are defined as pro-biofilm (> 100%).
• the “pro-biofilm limit” is defined as that roughness from which a biofilm greater than 100% is obtained (with respect to that generated in the Op control).
• the pro-biofilm limit is a minimum Ra roughness of approximately 10 nm (Ra: arithmetic mean roughness).
• the above method was then repeated using a glass Petri dish as a substrate, instead of PS.
• a glass plate was coated with 48 passes.
• the coating method was identical to that used with PS Petri dishes described above and APTES was used as the liquid precursor material.
• the number of passes was set to 2 and the percentage by weight of PEGMA was varied in a dilution of PEGMA and IPA (5, 10 and 100% by weight).
• Figure 5 shows the relationship between roughness and% biofilm of coatings applied on PS using APTES (previous embodiment) and PEGMA + IPA (present embodiment) as liquid precursor materials.
• APTES previously embodiment
• PEGMA + IPA present embodiment
• coatings based on PEGMA require a greater roughness to achieve the amount of biofilm generated with coatings based on APTES.
• the pro-biofilm limit of APTES coatings is approximately 10 nm (roughness from which a biofilm greater than 100% is obtained with respect to the control), in the case of coatings based on The PEGMA pro-biofilm limit is 160 nm ( Figure 5).
• the chemical characterization was carried out by means of X-ray photoelectron spectroscopy analysis (XPS) and the wettability was carried out by an analysis of the water contact angle (WCA) measurement.
• XPS X-ray photoelectron spectroscopy analysis
• WCA water contact angle
• Figure 7 shows the atomic percentage of the elements present (C, O, Si and N) in each of the coatings studied based on APTES (uncoated, Op, and coated with 2, 4, 12, 24, 48 , 72 and 96 passes).
• These elements come from the plasma polymerization of APTES (in the case of the coated samples), the surface of PS (in the case of the control uncoated Petri dish) and air surrounding during the veneering procedure.
• Figure 8 shows the deconvolution (or decomposition) of the high resolution “total” spectrum of carbon (C1s) from samples Op, 2p and 72p.
• Carbon deconvolution is a common practice in the chemical characterization of surfaces. This deconvolution makes it possible to identify the partial spectra that make up the total spectrum. Each partial spectrum corresponds to a specific bond to which the carbon is attached. The area of each partial spectrum (associated with a specific bond) allows quantifying its presence on the surface with respect to the total number of bonds.
• Figure 8 shows the percentage relative to each link (inscribed in a rectangle).
• Op sample deconvolution uncoated PS Petri dish
• the typical bonds to which carbon is bound are identified in a plasma treated PS sample (uncoated, plasma only).
• the PS Petri dishes used in this study had previously been treated with plasma by their supplier (Nunc TM Petri dishes / cell cultures from Thermo Scientific TM) for surface activation.
• Groups [A], [B] and [E] are derived from the PS molecule, while groups [C] and [D] are formed during plasma treatment (performed by the supplier of PS Petri dishes).
• [B] and [C] linkages are common to plasma activated PS and polymerized APTES.
• samples 2p and 72p representative of all samples coated from APTES
• the characterization was then carried out by analyzing the WCA, which refers to the angle that the surface of a liquid forms when it comes into contact with a solid.
• the value of the contact angle depends mainly on the relationship between the adhesive forces between the liquid and the solid and the cohesive forces of the liquid. The higher the adhesive forces, the lower the WCA.
• Figure 9 shows the relationship between roughness and WCA of the coatings studied based on APTES (uncoated, Op, and coatings with 2, 4, 12, 24, 48, 72 and 96 passes).
• APTES uncoated, Op, and coatings with 2, 4, 12, 24, 48, 72 and 96 passes.
• the increase in% biofilm does not depend on the "chemistry” or “wettability” (adhesion) of the coating, but depends on the modification of the "morphology” of the coating (roughness).
• Bacterial growth was then studied for 24 hours (0, 3, 6 and 24 hours) of the uncoated (Op) and coated samples with 2, 4, 12, 24 and 48 passes.
• the bacterial growth of each sample was determined using a Microplate Reader 680XR Bio-Rad spectrophotometer and measuring absorbance at a wavelength of 620 nm.
• Figure 10 shows that only in coating 12p more bacteria grew than in the uncoated sample (Op). That is, fewer bacteria grew in most of the coated samples than in the control. Therefore, the production of more biofilm by these samples (with respect to the uncoated sample, Op), is not due to an increase in the number of adhered microorganisms.
• NH2 amine groups
• Fig. 1 1 a diagram of the mechanism of overproduction of biofilm (PRO-Biofilm) that occurs in the coating of the present invention is shown. A diagram of the initial adhesion of bacteria to the coating is shown in Fig. 1 1a.
• Fig. 13 is shown a diagram of the production of biofilm as the roughness of the coating increases (PRO-Biofilm) with the passes.
• Fig. 13a represents the adhesion and biofilm creation phase of a bacterium (Pseudomonas aeruginosa) on the UNcoated substrate.
• Fig. 13b, c and d represent the formation of colonies and the increase in the amount of biofilm as the roughness of the coating increases (from 4 passes, 4p, to 72, 72p).
• Fig. 13a would correspond to the image of Fig. 12a
• Fig. 13d would correspond to the image of Fig. 12b and Fig. 12c.
• Fig. 14 a diagram is shown representing the "function" of each "part” of the APTES molecule after it has been decomposed in the plasma-polymerization process at atmospheric pressure.
• the amino groups (NH2) promote the adhesion of the bacteria
• the groups of silicon oxide (SiO x ) accumulate in different agglomerates (whose size depends on the number of passes) that promote the formation of larger colonies of bacteria with the consequent greater production of biofilm.
• Figure 15 shows the amount of biofilm (measured by FDA) generated in the Op and 48p samples during 24 hours (measurements performed at 6, 12 and 24 hours).
• the maximum amount of biofilm produced in the 48p sample is reached 6 hours after inoculation, being already at that time (6h) significantly higher than the biofilm produced in the uncoated sample (Op ).
• This speed is of vital importance when it is urgently necessary to determine which treatment is optimal for a patient in a given situation (most effective antibiotic and dose thereof).

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