MX2008010696A - Molecular plasma deposition of colloidal materials. - Google Patents

Abstract

A molecular plasma discharge deposition method for depositing colloidal suspensions of biomaterials such as amino acids or other carbon based substances onto metal or nonmetal surfaces without loss of biological activity and/or structure is described. The method is based on generating a charged corona plasma which is then introduced into a vacuum chamber to deposit the biomaterial onto a biased substrate. The deposited biomaterials can be selected for a variety of medical uses, including coated implants for <i>in</i> <i>situ</i> release of pharmaceuticals.

Description

DEPOSITION OF MOLECULAR PLASMA OF COLOIDAL MATERIALS FIELD OF THE INVENTION The invention relates to an apparatus and process for using corona discharge to deposit colloidally suspended molecules on substrate surfaces. The method is applicable to the deposition of organic or inorganic compounds, particularly to proteins and related biological compounds of interest on selected substrates with little or no loss of native structure or activity.

BACKGROUND OF THE INVENTION There is an increasing interest in the immobilization of biologically active substances on various substrates without significant alteration of the desired function or activity. Surfaces coated with antibiotics, for example, are typically prepared by dipping or painting process, which often result in poor adhesion, incomplete surface wetting or poor adhesion. Ion plasma deposition (IPD) methods have been extensively developed and used in ef.195516 coating procedures, predominantly with the aim of producing highly adhesive coatings and customized surface characteristics. Recently attention has been focused on the preparation of coated surfaces that are biocompatible, such as those suitable for medical implants where coatings improve cell adhesion or where anti-microbial coatings are important to avoid potential sepsis after surgery. Corona discharge is a well-known phenomenon that has long been observed in nature and traditionally used in a number of commercial and industrial processes. This is currently used in the production of ozone, the control of electric charges generated superficially, and in photocopying. The corona electric discharge has also been used to modify surfaces, particularly for plastic articles to improve surface characteristics, as described in U.S. Patent No. 3, 274, 089. An electrostatic coating process involving a discharge in crown of a liquid or powder material is described as a coating method in U.S. Patent No. 4, 520, 754. A corona is generated when the potential gradient is large enough at a point in the fluid to cause the ionization of the fluid, so that it becomes conductive. If a loaded object has a sharp or sharp point, the air around that point will be at a much higher gradient than in other places. The air near an electrode can become ionized (partially conductive) while the more distant regions do not. When the air near the point becomes conductive, this has the effect of increasing the apparent size of the conductor, since the new conductive region is less acute, the ionization can not extend beyond the local region. Outside the region of ionization and conductivity, slowly charged particles find their way to an opposite charged object, and are neutralized. The corona discharge usually involves two asymmetric electrodes; one highly curved, for example, a needle point or a wire of small diameter, and one of low curvature, for example a plate, or a ground. The high curvature ensures a high potential around the electrode, providing the generation of a plasma. If the geometry and gradient are such that the ionized region continues to grow rather than stop at a certain radius, a fully conductive path can be formed, resulting in a momentary spark or continuous arc. Crowns can be positive or negative. This is determined by the polarization of the voltage over the highly curved electrode. If the curved electrode is positive with respect to the flat electrode, there is a positive corona; otherwise the crown is negative. The physics of positive and negative crowns is surprisingly different. This asymmetry is a result of the large difference in mass between the electrons and the positively charged ions, only with the electron that has the ability to undergo a significant degree of non-elastic ionizing collision at ordinary temperatures and pressures. Corona discharge systems have been used to activate chemical compounds, in general to deposit polymerizable and polymerizable monomers formed within a corona discharge on the surfaces as protective coatings; as described in U.S. Patent No. 3,415,683. A corona discharge reactor for chemically activating the constituents of a gas stream; for example, sulfur and nitrogen oxides and mercury vapor, is described in U.S. Patent No. 5,733,360. The reactor is designed to generate pulses from a corona by applying high voltage pulses for up to 100 nanoseconds to a plurality of corona discharge electrodes. Document O 2006/046003 describes various methods for coating substrates, involving the use of a plasma, including the use of pulsed plasma at low pressure, to introduce monomers or monomers in combination with free radical initiators to initiate polymerization on a substrate suitable. A diffuse dielectric barrier discharge assembly, at atmospheric pressure, is used into which an atomized liquid containing the monomers is introduced, so that an atomized spray coating material of 10 to 100 μm is formed. An apparatus for generating effluvium plasma or luminescent discharge, at atmospheric pressure, using electrodes energized by radiofrequency, is described in WO 03/084682. A plasma coating apparatus and method are described in WO 92/28548. The atomized coating materials, liquid or solid, are introduced into a plasma discharge at atmospheric pressure, and are useful for organic coatings such as polyacrylic acid or perfluorinated compounds in addition to the silicon-containing monomers. The effects of the crown are not always considered beneficial and may in fact cause arcing, or the breaking of the crown. In addition to this disruption, the corona effect may be too strong to successfully successfully single out only one complete molecule. When molecules are ionized at a higher level, they can separate and lose structural and functional properties. An advantage of depositing materials generated in plasma from liquid solutions is that any solvent present is typically deposited along with the intended material, creating unintended structures. For most processes where corona discharge can occur during plasma generation, efforts are usually taken to reduce the corona effect instead of using this effect as a deposition technique. The loss of functional and / or physical properties of organic molecules deposited superficially with plasma points to the need to develop methods of maintaining the desirable biological activities of the immobilized materials. Attempts to engineer biological coatings on a range of substrates, such as plastics, metals, polymers and ceramics have met with limited success and have generally failed to deposit biologically active agents on surfaces without compromising the desired activity.

BRIEF DESCRIPTION OF THE INVENTION The present invention relates to a method of molecular plasma deposition for biological non-destructive coating agents on substrate surfaces. The method employs a modified IPD device that uses electric or vacuum fields to lay a biological coating on virtually any conductive surface, and many non-conductive surfaces. A molecular discharge by corona plasma is generated from a highly charged conductive tip. The method is applicable to the deposition of a wide range of organic and inorganic materials, which are dispersed as solutions or suspensions from the conductive tip. There are several characteristics of the described method that differ from the conventional uses of either corona discharge or ion plasma deposition. The method uses solutions or suspensions of the materials that are going to be deposited. This allows a wide range of organic as well as inorganic materials, including elements and compounds, to be used. The liquids are atomized through a small tip orifice maintained at a high voltage, so that an ionized plasma is supplied from the orifice. In a subsequent step of the method, the ionized plasma is directed towards an evacuated chamber where an opposingly deviated substrate is located, causing the material to be deposited where it is bound to the surface of the substrate. An important feature of the method is the deposition of biologically active agents on a surface with little or no alteration of structural or functional characteristics. The method is equally applicable to inorganic materials, elements and selected compositions, which are not otherwise suitable for the coating processes. Due in part to the wide range of materials that can be deposited by this method, the ability to modify or to bio-engineer different surfaces is significantly expanded. The known characteristics of the corona effect under atmospheric conditions and the advantages of the ionic plasma deposition (IPD) methods in the coating processes have been used to develop a new corona plasma deposition process and a coating method. An important aspect of the invention is the ability to use a corona generated from a liquid or colloidal composition to deposit a coating consisting solely of the desired component, without the solvent in which the material to be deposited is dissolved or dissolved. scattered. In addition, maintenance of the original structural properties of a wide range of materials deposited from a molecular plasma generated in corona was not expected, most notably as shown with a polypeptide enzyme, which maintained the catalytic activity after deposition by molecular plasma. The atomic bonds are not broken during the deposition process, which is a factor in the retention of the activity and / or the structural integrity of the deposited product. In one embodiment, the process is carried out partly under atmospheric or partial pressure, and partly under vacuum. The deposition apparatus is designed to generate a corona from a solution or suspension introduced through a narrow electrified opening, such that a plasma is produced in front of a small opening that opens into a vacuum chamber that houses a substrate. Depending on the charge produced on the material dispersed in the corona plasma, the substrate is wired as an oppositely charged electrode on which the plasma particulate materials will deposit. The basic structural characteristics of the tested deposited materials are not affected by the thickness of the deposit. This is in contrast to the results obtained by Storey (Breakup of Biomolecules through low-energy ion Bombardment, Master's thesis, University of Missouri, Rolla, 1998) where more than 40 or more glycine or arginine monolayers deposited by flooding a solution of the amino acids on gold, caused the loss of structure, as indicated by the increase in the difficulty in detecting the carboxyl groups of the amino acids deposited as the thickness of the sample increases. The molecular plasma method described allows the thickness to be controlled from a monolayer of desired material to micrometer thickness, for example, 2-200 micrometers in thickness while maintaining structure and activity. The apparatus for the deposition of molecular plasma can be deposited to accommodate a partial vacuum around the conducting tip where the crown is generated. This allows the most efficient volatilization of the solvent that suspends suspended dissolved material, so that only the material itself is extracted into the vacuum chamber, towards the housing of the vacuum chamber that is to be deposited on the substrate and little if it is that some solvent is present in the coating. This is necessary because, in biological applications, if the suspending solvent is also co-deposited, it can cause an adverse interaction with the deposited material. For example, if a protein that is desired to be deposited on a medical implant device is dissolved in methyl alcohol and deposited without volatilization of the alcohol before being placed in the body, the residual alcohol can cause serious physiological problems. Where little or no solvent is pulled into the chamber, it is convenient to generate the corona under conditions of atmospheric pressure. As discussed, the new method is a molecular plasma method for the deposition of a biomolecule on a substrate. A corona discharge plasma is generated under atmospheric or partial vacuum conditions from a liquid solution or suspension. The suspension is preferably a colloidal suspension for materials that have low solubility in organic or aqueous solvents. The deposited materials may be an element, a compound, or any of a number of biomolecules. The liquid solution or suspension is expelled from a source of the conductive point to a high potential gradient, and the resulting corona discharge is directed through an opening into an evacuated chamber where the ionized molecular plasma will be deposited on a substrate that is maintained to an opposite induced potential of the relatively high potential at the source of the point where the corona is generated. In general, the tip or conductive point from which the colloidal suspension is expelled provides a means for atomizing the liquid solution or suspension, so that there is ready formation of a corona discharge at the high voltage tip. In many applications, it will be preferred to introduce the liquid solution or suspension from the tip under atmospheric conditions, but a low or partial vacuum, preferably 100 mTorr or greater may also be used. The charged plasma then passes through a hole or orifice into an evacuated chamber, for example, at 40 mT or less, housing a substrate maintaining a voltage substantially opposite to the voltage at the conductive tip. The ionized molecules in the corona plasma will then be deposited on the substrate in the chamber, which is at a lower pressure than at the conducting tip where the corona is generated. It is important to recognize that the substrate is under a vacuum, typically less than 100 mTorr, preferably 40 mTorr or less, and that if the plasma corona is formed at a tip also under reduced pressure, the substrate must be in an atmosphere of reduced pressure , such that the plasma can be effectively extracted into the substrate housing and deposited on the substrate. The vacuum around the substrate is typically in the range of 40-0.1 mTorr. further, the substrate must be oppositely deviated in order to effect the deposition of the ionized molecular plasma, which can be positive or negative, depending on the material in the colloidal suspension. The ions formed in the discharge can be positive or negative. The discharge will determine the polarization of the substrate, for example, if a positive corona is used, the substrate must be negatively deflected. The amount of polarization imposed on the substrate will depend on the substrate and the area for deposition. In the examples provided herein, the substrates are approximately 4 cm. The polarization of the substrate is constant despite the size and deposition. However, the larger the area, the higher the current necessary to keep the voltage constant. The voltage applied to the substrate can be in the range as high as 60 kV, which can be positive or negatively offset, for example, to the voltage at the conducting tip, where the corona is generated. Typical voltages are in the range of + 15kv to - 15kV. Where the substrate is connected to ground, the voltage will be zero on the substrate. The method is partly based on the efficient generation of an ionized plasma. This is achieved by the atomization of a liquid solution or suspension of the desired material for deposition through a sharp orifice or sharp tip. This is typically a small diameter tube or needle that is imposed with a high voltage. An exemplary high voltage on an 18 gauge metal needle with an internal diameter of approximately 0. 83 MI, for example, can be approximately 5000 volts. In this example, the voltage applied to the substrate is typically in the range of about 5000 or less volts or is zero if it is connected to ground. Another aspect of the invention is the ability to control the corona effect for processing more efficiently; for example, for engineering surfaces. The method takes advantage of the physics of the corona effect to deposit ionized material on a substrate, so that the material is ionically or covalently bound to a surface of the substrate. This is an easy and quick way to produce an adherent coating and to structure a surface using a non-destructive technique. The described apparatus can be used to effectively deposit a wide range of materials without significantly altering the physical, functional or chemical characteristics, by generating material in a corona in plasma from a liquid solution or suspension. Materials such as proteins are preferably ionized from colloidal suspensions, due to their limited solubility in most solvents. Thymine, cytosine, adenine and guanine with solubilities in respective waters of 4.5 g / 1, insoluble, 0.5 g / 1, and insoluble, are more efficiently deposited as aqueous suspensions. Almost any organic or inorganic material can be dissolved or suspended in some solvent or mixture of solvents. The materials include proteins, amino acids, peptides, polypeptides, nucleic acids and nucleic acid bases, such as purines and pyrimidines, and the like. Examples of inorganic materials include compounds such as metals and metal oxides, for example, copper oxide, elements, including carbon. In general, biological and non-biological materials that can be prepared as a liquid solution or suspension will be suitable materials for deposition. While colloidal suspensions may be preferable for some biomolecules, microparticulate suspensions may also be suitable, depending on the material, thereby creating ionized molecular plasmas from liquids containing micron sized or larger particles. Examples of materials that are preferably deposited from molecular plasmas generated from colloidal suspensions include, but are not limited to: DNA, RNA, graphites, antibiotics, growth factors, growth inhibitors, viruses, inorganic compounds, catalysts , enzymes, organic compounds and elements. Catalase is an example of a polypeptide enzyme that can be deposited by this method without loss of catalytic activity. Other proteins that are expected to be deposited by this method without loss of biological activity, include the SNAP marker fusion proteins, hAGT fusion proteins, glucose binding proteins, glutamine binding protein, and the ctato-dehydrogenase. Oxidoreductases and oxidases such as glucose oxidase are also expected to be deposited without loss of activity. Nucleotide bases such as guanine, thymine, adenine, cytosine and uracil are examples of DNA and RNA bases that can be deposited on a substrate from a corona generated plasma. Liquid solutions and suspensions that are suitable for deposition can be formulated from any of a number of aqueous or organic solvents including pure water, alcohol and water / alcohol mixtures. Some materials can be prepared in less common solvents, such as DMSO or glycols. For the preferred practice of the deposition process, a solvent or combination of solvents in which the material can be dissolved or suspended, preferably as a colloidal suspension, and which does not cause problems such as toxic or explosive fumes, will be selected.

DEFINITIONS The biomolecules are intended to include compounds and agents that have some biological effect or use in the body and may include, without limitation, proteins, peptides, amino acids, nucleic acids and compounds having drug activity or related to pharmacological activity. As used herein, biomolecules also include carbon and carbon-based compounds, and elements and compounds such as copper oxide that can be used as coating materials on medical devices. Colloidal particles are finely divided particles of about 10 to 10,000 angstroms in size, dispersed within a continuous medium. The particles are not easily filtered and settle slowly over a period of time. The nanoparticles are approximately 100 nanometers or less in size. The ionic plasma deposition (IPD) is the vacuum deposition of the ionized material, generated in a plasma, in general by the application of high voltage or high current to a cathode target where the ionized plasma particles are deposited on a substrate that acts as an anode. A corona is produced by a process by which a current, perhaps sustained, develops from an electrode with a high potential in a neutral fluid, such as air, by ionizing the fluid to create a plasma around the electrode. The ions generated sooner or later transfer a charge to nearby areas of lower potential, or recombine to form neutral gaseous molecules. The polarization of the voltage is the potential, in relation to the earth, at which the substrate is maintained. The potential is in the range of zero to 15,000 volts and can be positive or negative. The potential on the substrate for polarization depends on the potential of the corona and is typically the same and opposite to this potential. The voltage can be as high as 60 kV but more typically it is in the 5-10 kV range. Where the corona plasma is about +5000 volts, the polarization voltage for the substrate will be -5000 volts, all in relation to the ground. As used herein, substantially or substantially means that an interval is involved, of the order of plus or minus ten percent and is not intended to be limited to an exact number, for example, the substantial function may include different or less than the original function.

BRIEF DESCRIPTION OF THE FIGURES Figure 1 is a sketch of the molecular plasma deposition apparatus: vacuum chamber 1; 2 high voltage power supply; substrate fastener 3; substrate 4; 5 high voltage power supply; needle 6; needle tube feeder 7; orifice 15 inside the tank 8; colloidal liquid suspension 9. Figure 2 is a sketch of a modification of the apparatus of Figure 1: vacuum chamber 1; 2 high voltage power supply; substrate fastener 3; substrate 4; 5 high voltage power supply; needle 6; feeder tube 7; orifice 15 inside the tank 8; liquid suspension 9; secondary chamber 10; 11 supply of secondary chamber gas; line 12 of secondary chamber gas supply; pressure regulator 13; Gas line from regulator 14. Figure 3 is a representation of the electric field equation for a point charge.

DETAILED DESCRIPTION OF THE INVENTION The present invention takes advantage of the corona effect and the effect of corona discharge in the creation of a charged plasma that can be directed to a substrate surface. The basic apparatus is shown in Figure 1 and Figure 2. In an exemplary procedure, it is applied in high voltage of 5 kV or greater to the needle or to another inductive, sharp-edged, hollow-core material. A liquid solution or suspension is passed through the hollow core. The high electric field of the needle tip causes atomization of the core as the result of the corona effect. The molecules in the solution or suspension are loaded, and they remain intact. The needle is placed in front of a differentially pumped high vacuum system, connected to ground, with a small hole in the chamber that houses the substrate. The substrate is placed inside the evacuated chamber at a potential opposite or almost opposite to that imposed on the needle, or is adjusted to ground (zero). The charged molecules within the corona travel through the opening to the substrate and become deposited or coupled to the substrate, becoming ionically or covalently bound. The complete apparatus is enclosed in an environmentally controlled chamber within which selected gases such as oxygen or nitrogen can be introduced. For example, if oxidation is desired, to control the rate of deposition, to perform the deposition in an inert atmosphere. Mixtures of gases can be introduced, including other inert gases such as xenon, argon, helium gas combinations. The molecular plasma generation process can also be run at pressures lower than atmospheric, for example, under reduced pressure, in the presence of gases other than atmospheric gases (for example, argon or previous oxygen atmosphere). When the molecular plasma in the conductive tip is generated under reduced pressure, the pressure in the chamber that houses the substrate must be less than that of the plasma discharge that passes easily through the opening in the chamber that houses the substrate, as is shown in Figure 1. As shown in Figure 1, the molecular plasma generating apparatus provides a system for producing a plasma discharge under atmospheric conditions by passing a liquid colloidal suspension 9 through a discharge needle. at a high voltage 5. The resulting atomized liquid forms an ionized plasma in the atmosphere. The plasma passes through a hole 15 in the vacuum chamber 1 which houses the substrate 4 on the holder 3 of the substrate. A power supply 2 provides voltage to the substrate 4 at a voltage opposite that provided by the power supply 5 to the discharge needle 6. Figure 2 illustrates an alternative embodiment of a system for producing an ionized plasma discharge. A reservoir 8 feeds a liquid solution or suspension of the material 9 through an orifice 15 for the deposition of the colloidal material on the substrate 4. The liquid is passed through the highly charged needle 6 from the power supply 5. In this mode, the feeder and the needle are housed in a second chamber 10, which can be regulated in pressure by a pressure control 13 through the opening 14 towards the secondary chamber. The atmosphere inside the secondary chamber 10 can be modified from a gas container 11 having a conduit 12 passing through the regulator 13. The vacuum chamber 1 is maintained at a lower pressure than in the chamber 10. The substrate 4 it is deflected or polarized using the power supply 2 at a voltage opposite to that supplied by the power supply 5 to the needle 6. The liquid suspensions can be prepared in organic or inorganic liquids, which must not be toxic or flammable. Most materials are preferably prepared as aqueous solutions or can be prepared in organic acids such as acetic acid, propionic acid, acetic acid substituted with halogen, oxalic acid, malonic acid and / or hydrocarboxylic acids alone or with water. The liquid mixtures may include salts or miscible organic / water preparations. Examples of alcohols include ethanol, methanol, and ketones such as acetone, D F, THF and methyl ethyl ketone. Amino acids, for example, can be soluble in water at low concentrations, but form colloidal suspensions at higher concentrations. Lysine and threonine are highly soluble in water whereas tyrosine has a limited solubility of approximately 0.045 g / 100 ml at 252C. Corona discharge of positive or negative variety is commonly characterized as the ionization of a neutral atom or molecule in a region of a strong electric field, typically in the high potential gradient near a curved electrode, creating a positive ion and a free electron. The electric field then separates and accelerates the charged particles, providing for recombination and imparting each particle with kinetic energy. Energized electrons, which have a much higher charge / mass ratio and are thus accelerated at a higher velocity, can create additional pairs of electron / positive ion by collision with neutral atoms. These then undergo the same separation process, giving rise to an avalanche of electrons. Positive and negative crowns rely on electron avalanches. Figure 3 illustrates a typical point charge formed in a strong electric field. The energy of these plasma processes is converted into initial electron dissociations to plant additional avalanches. An ionic species created in this series of avalanches, which differs between positive and negative crowns, is attracted to a non-curved electrode, for example, a flat surface, completing the circuit, and holding. the current flow. A corona is a process by which a current, whether sustained or not, develops from an electrode with a high potential gradient in a neutral fluid, usually air. When the potential gradient is large enough at a point in the fluid, the fluid at that point is ionized and becomes conductive. If a loaded object has a sharp or sharp point, the air around that point will be at a higher gradient than elsewhere, and may become conductive while other points in the air are not. When the air becomes conductive, it effectively increases the size of the conductor. If the new conductive region is less acute, the ionization may not extend beyond this local region. Outside this region of ionization and conductivity, the charged particles slowly find their way to an opposite charged object and are neutralized. On the other hand, if the geometry and gradient are such that the ionized region continues to grow rather than stop at a certain radius, a completely conductive path is formed, and a continuous spark or momentary arc occurs. The corona discharge usually involves two asymmetric electrodes, one highly curved such as the tip of a needle or a narrow wire, and one of low curvature, such as a plate, or the ground. The high curvature ensures a high potential gradient around an electrode in order to effectively generate a plasma. Crowns can be positive or negative. This is determined by the plurality of the voltage on the highly curved electrode. If the curved electrode is positive with respect to the flat electrode, the crown is positive; If the electrode is negative, there is a negative corona. The physics of positive and negative crowns is strongly different. This symmetry is a result of the large difference in mass between electrons and positively charged ions, only with the electron that has the ability to undergo a significant degree of non-elastic ionizing collisions at standard pressures and temperatures. A negative corona is manifested as a non-uniform crown, varying according to the characteristics and surface irregularities of the curved conductor. This often appears as crown plumes on sharp edges, the number of plumes changes with the strength of the field. The shape of the negative crowns is a result of its source of secondary avalanche electrons. This appears a bit larger than the corresponding positive corona, since the electrons are allowed to move out of the ionizing region, allowing the plasma to continue a certain distance beyond it. The total number of electrons and electron density is much higher than in the corresponding positive corona; however, the electrons are at a predominantly lower energy, due to being in a region of lower potential gradient. Therefore, while for many reactions the increased electron density will increase the reaction rate, the lower energy of the electrons means that reactions that require a higher electron energy can take place at a lower speed. A positive corona is manifested as a uniform plasma across the length of a conductor. This is often observed as a blue / white illumination, although much of the emission is in the ultraviolet range. The uniformity of the plasma is due to the homogeneous source of secondary avalanche electrons. With the same geometry and voltages, a positive corona appears somewhat smaller than the corresponding negative corona, due to the 'lack of a non-ionizing plasma region between the internal and external regions. There are much less free electrons in a positive corona, perhaps a thousandth of the electron density, and one hundredth of the total number of electrons, compared to a negative corona, with the exception of the area near the curved electrode, where the electrons are highly concentrated. This region has a high potential gradient, causing electrons to have higher energy. Most electrons in a negative corona are in more, lower, external energy field areas. In a positive corona, secondary electrons, which give rise to additional avalanches, are generated predominantly in the fluid itself, the region outside the plasma or avalanche region. These are created by the ionization caused by the photons emitted from that plasma in the various de-excitation processes that occur within the plasma after the collisions of electrons. The thermal energy released in these collisions creates photons that are radiated to the gas. The electrons that result from the ionization of a neutral gas molecule are then electrically attracted back to the curved electrode and to the plasma, making the process of creating additional avalanches within the plasma cyclical. The positive corona is divided into two regions, concentric around the acute electrode. The internal region contains ionizing electrodes, and the positive ions, which act like a plasma, the avalanche of electrons in this region, which creates many additional ion-electron pairs. The outer region consists almost entirely of the massive positive ions that migrate slowly, moving towards the uncurved electrode together with, close to the interface of this region, the secondary electrons, released by the photons that leave the plasma, which are accelerated within the plasma. The inner region is known as the plasma region, the external region is the unipolar region. A negative crown is manifested as a non-uniform crown, varying according to the characteristics and surface irregularities of the curved conductor. This often appears as crown plumes on sharp edges, the number of plumes changes with the strength of the field. The shape of the negative crowns is a result of its source of secondary avalanche electrons. The negative corona seems a little larger than the corresponding positive corona, due to the displacement of the electrons from the ionizing region, so that the plasma continues a certain distance beyond it. The total number of electrons, and consequently the electronic density, is much greater than the corresponding positive corona. The electrons are of lower energy than those in a positive corona because they are in a region of lower potential gradient. Negative crowns are more complex than positive crowns under construction. As with positive crowns, the establishment of a corona begins with an exogenous ionization event that generates a primary electron, followed by an avalanche of electrons. The difference between positive and negative crowns is in the generation of secondary electron avalanches. In a positive corona, the avalanches are generated by the gas surrounding the plasma region, the new secondary electrons traveling inward, while in a negative corona these are generated by the curved electrode itself, the new secondary electrons travel outward . An additional structural feature of negative crowns is the outward displacement of electrons, where electrons find neutral molecules and can combine with electronegative molecules such as oxygen and / or water vapor to produce negative ions. These negative ions are then attracted to a non-curved, positive electrode, completing the "circuit". A negative corona can be divided into three radial areas, around the acute electrode. In the inner area, the high-energy electrons collide inelasticly with the neutral atoms and cause avalanches, while the external electrons, usually at a lower energy, combine with neutral atoms to produce negative ions. In the intermediate region, the electrons combine to form negative ions, but typically have insufficient energy to cause avalanche ionization. This is still part of a plasma due to the different polarities of the species present, and the ability to participate in characteristic plasma reactions. In the external region, only a flow of negative ions and, to a lesser degree and decreasing radially, the movement of free electrons towards the positive electrode takes place. The two internal regions are known as the corona plasma. The internal region is an ionizing plasma, the intermediate region is a non-ionizing plasma. The external region is known as the unipolar region. As discussed, the main crown has been used to create an approximately infinite electric field at the point of a sharp needle. For practical reasons, it can be assumed that the tip of the device is atomically sharp and closely approximates a point load. This is because r goes to zero, E approaches infinity. A corona effect is initiated at the tip of the device. The energy of the electrons and the relation to the distance of the point source of the generation, is based on the electric field of a point charge derived from the Coulomb law. This law establishes that the electric field coming from any number of point charges can be obtained from a sum of vectors of the individual fields. A positive number is taken to be an external field; the field of a negative charge is towards it. This can be shown in equation 1 and illustrated in figure 3: font 9 _ ^^ font qr EXAMPLES It is intended that the following examples be only illustrations of the invention and in no way be considered limiting of what is described and taught herein.

Example 1 - Apparatus for Molecular Plasma Deposition An exemplary apparatus includes a vacuum chamber with a small opening, and a small hole, a small blade connected to a tube connected to a reservoir that holds a suspension or liquid solution of the material that is you want to deposit. The deposit is at atmospheric pressure. An energy supply with the ability to supply up to 60 kV can be used; however, as used in the examples herein, the voltage coupled to the needle is typically from -5000 volts to +5000 volts. A substrate inside the vacuum chamber is centered over the opening with a polarization from -60 kV to -60 kV, including earth. The apparatus is illustrated in figure 1 Example 2 - Apparatus for the Generation of Molecular Plasma under Selected Environments The apparatus illustrated in Figure 2 can be modified such that the needle, the tube and the reservoir, are placed in a housing that excludes air, but allows the controlled introduction of other gases. The optionally selected gases include argon, oxygen, nitrogen, xenon, hydrogen, krypton, radon, chlorine, helium, ammonia, fluorine and combinations of these gases.

Example 3 - Apparatus for the Generation of Crown Discharge Under Reduced Pressure In the apparatus illustrated in Figure 1, the pressure differential between corona discharge and the substrate is about one atmosphere. The external pressure of the vacuum chamber is approximately 760 Torr, while the pressure in the substrate area is approximately 0.1 Torr. The apparatus shown in Figure 2, on the other hand, may optionally be operated at a predetermined pressure above or below atmospheric pressure. While atmospheric pressure is generally preferred for plasma generation, reduced pressure of up to about 100 mTorr may in some cases provide satisfactory depositions.

Example 4 - Deposition of Molecular Plasma from Amino Acids This example illustrates the deposition of an amino acid suspension on a gold rod. A colloidal suspension of a mixture of the amino acids glycine (solubility of 20 g / 1 at 25 ° C), alanine (166.5 g / 1), valine (88.5 g / 1), leucine (24.26 g / 1) and arginine (235.8) g / 1) in water was deposited using the apparatus of Example 1 on a gold coated rod, of 3.17 mm (1/8 inch) in diameter and approximately 0.75 mm2. The power supply was coupled to an 18 gauge needle of 304 stainless steel and adjusted to -5000 V. The gold substrate was adjusted to a potential of 5000 V. The substrate was centered over the hole in the chamber and placed at 5 cm of the orifice The vacuum chamber was pumped at 40 mTorr and the flow of the colloidal suspension was started. The deposition was carried out for 30 minutes. The coated rod was placed in a time-of-flight secondary ion mass spectrometer (TOF-SIMS) and the components were analyzed for composition. The results showed that the amino acids were deposited intact and only bound to the substrate. The mass-on-load calculations in conjunction with the flight spectrometry time were used to calculate the masses of the incoming species. These calculations were used to interpret the spectra from the SIMS. The m / q data showed the amino acids that are expelled intact from the surface. In a control comparison experiment, the substrate was submerged in the amino acid mixture and analyzed by TOF-SIMS as described above. These spectra were subtracted from the amino acid spectra generated from the corona deposition in order to isolate any effects that occurred due solely to the deposition method. Fragmentation was observed in both spectra, and after subtraction, it was determined that the fragmentation was an effect of the analytical technique not the ce deposition technique because the fragmentation occurred equally in both spectra.

Example 5 - Deposition of Graphite by Molecular Plasma A colloidal suspension of graphite powder in isopropyl alcohol (10 g / 100 ml) was deposited on an aluminum oxide substrate using the apparatus shown in Figure 1. The energy supply was coupled to an 18 gauge needle made of 304 stainless steel and adjusted to 5000 V. The aluminum oxide substrate was grounded. The substrate was centered on the hole in the chamber and placed 5 cm from the hole. The vacuum chamber was pumped at 40 mTorr and the flow of the colloidal suspension was started. The deposition was carried out for 30 minutes. The substrate was removed from the chamber and a simple ohmic measurement resistance test was performed. The substrate resistance changed from infinity to 1 ohm in the 30 minute deposition period.

Example 6 - Deposition of Copper Oxide by Molecular Plasma A colloidal suspension of copper oxide powder in water (10 g / 100 ml) was prepared. Using the apparatus illustrated in Figure 2, the high voltage power supply was coupled to an 18 gauge stainless steel needle 304, adjusted to -10,000 V. The substrate was 304 stainless steel and adjusted to a potential of 5000 V. substrate was centered and placed 5 cm from the hole in the chamber. The chamber was pumped at 40 mTorr and the flow of the colloidal suspension was initiated. The deposition on the substrate was allowed to proceed for 10 minutes. At the end of the deposition process, the substrate was removed from the chamber and a simple tape test showed good adhesion of deposited copper oxide. The good addition between the substrate and the copper oxide was confirmed by the repetition of the tape test and by the observation that after sonication the coated sample for 10 minutes, there was no evidence of flaking or flaking.

Example 7 - Deposition of RNA and DNA Bases by Molecular Plasma A colloidal suspension of guanine, adenine, cytosine, uracil and thymine in water (each 5 g / 100 ml) was deposited on a gold-coated rod having a surface of approximately 0.75 cm2 area, a diameter of 3.17 mm (1/8 inch), using the apparatus of Example 1. The power supply was coupled to a 18 gauge needle of 304 stainless steel, and adjusted to -5000. V. The gold substrate was adjusted to a potential of 5000 V. The substrate was centered on the hole in the chamber and placed 5 cm from the hole. The vacuum chamber was pumped at 40 mTorr and the colloidal suspension was started. The deposition was carried out for 30 minutes. The coated rod was placed in a time-of-flight secondary ionic mass spectrometer (TOF-SIMS) and the components were analyzed for composition. The results showed that the DNA bases were deposited intact and ionically bound to the substrate. Mass-on-load calculations in conjunction with time-of-flight spectrometry were used to calculate the masses of the incoming species. These calculations were used to interpret the spectra from SI S. The m / q data showed the bases that are ejected from the surface while intact. The spectra, from another method of deposition (immersion of the substrate in a mixture containing the bases) were also analyzed as a control to the deposited bases using the corona effect. The spectra were subtracted from the corona spectrum to isolate any effects that occurred due solely to the deposition method. Fragmentation was observed in both spectra, and once subtracted, it was determined that this observation was a product of the analytical technique and not the deposition technique, because fragmentation occurred equally in both spectra.

Example 8 - Deposition of Catalase by Molecular Plasma 25 ml of a crystallized bovine hepatic catalase 2X (Sigma C100-58MG; 056K7010) in colloidal suspension in water with 0.1% thymol was prepared. The protein concentration was 33 mg / ml with an activity of 4.1 x 104 U / ml. Using an apparatus illustrated in figure 2, the high-voltage power supply was coupled to an 18 gauge needle of 304 stainless steel set to -5000 V. The substrate was a 6.35 mm (1/4 inch) thick aluminum oxide disk by 38.1 mm (1.5 inches) in diameter, having an area of approximately 11 cm2 and adjusted to a potential of 5000 V. The substrate was centered and placed 5 cm from the hole in the chamber. The chamber was pumped at 40 mTorr and the colloidal suspension flow started. The deposition on the substrate was allowed to proceed for 10 minutes. At the end of the deposition process, the substrate was removed from the chamber and the sample was placed in a 5% solution of hydrogen peroxide. The results showed the catalysis of hydrogen peroxide by catalase, producing oxygen bubbling from the surface, showing that the enzyme remained intact throughout the process of deposition.

The deposition was repeated twice under the same conditions, except that after the substrate was removed from the chamber, the samples were placed in an ultrasonic water bath for 10 minutes. In addition, one of the samples was kept at 10 ° C for 72 hours after the bath was removed. In each case, exposing the sample to a 5% solution of hydrogen peroxide caused bubbling of oxygen from the surface of the substrate. Ultrasonic treatment did not affect the deposited material, indicating that a stable adherent coating of catalase had been deposited.

It is noted that in relation to this date the best method known by the applicant to carry out the aforementioned invention, is that which is clear from the present description of the invention.

Claims (20)

1. CLAIMS Having described the invention as above, the content of the following claims is claimed as property: 1. A molecular plasma method for the deposition of a biomolecule on a substrate, characterized in that it comprises: the generation of a corona discharge plasma, under atmospheric conditions from a liquid solution or suspension of a biomolecule expelled from a conductive point source having a high potential gradient; and the direction of the plasma generated through an orifice of an evacuated chamber that houses a substrate, at an induced potential opposite the point source; wherein the biomolecule is deposited on the substrate.
2. 2. The method according to claim 1, characterized in that the biomolecule is a colloidal suspension.
3. 3. The method according to claim 1, characterized in that the liquid is water, alcohol or mixtures thereof.
4. 4. The method according to claim 1, characterized in that the generated corona discharge plasma comprises a positively or negatively charged plasma.
5. 5. The method according to claim 1, characterized in that the biomolecule is an amino acid, a nucleic acid base or a polypeptide.
6. 6. The method of compliance with the claim 1, characterized in that the biomolecule is selected from the group consisting of carbon, copper oxide, an amino acid, a DNA or RNA base, catalase and mixtures thereof.
7. The method according to claim 1, characterized in that the biomolecule is at least one of glycine, alanine, valine, leucine, arginine, or mixtures thereof.
8. 8. The method of compliance with the claim 1, characterized in that the biomolecule is at least one of guanine, adenine, thymine, cytosine or mixtures thereof.
9. 9. The method of compliance with the claim 2, characterized in that the colloidal suspension is comprised of particles having a size range between about 100 angstroms and about 10,000 angstroms.
10. The method according to claim 1, characterized in that the substrate is a metal, ceramic or a plastic.
11. 11. A molecular plasma method for the deposition of a biomolecule on a substrate, characterized in that it comprises: the atomization of a liquid solution or suspension of a selected biomolecule, within a low vacuum atmosphere from a conductive tip at high voltage to generate a charged plasma; the direction of plasma charged through an orifice to an evacuated chamber that houses a substrate grounded, or maintained at a voltage opposite to the voltage of the conductive tip; wherein the biomolecule is deposited on the substrate.
12. The method according to claim 11, characterized in that the biomolecule is selected from the group consisting of carbon, copper oxide, an amino acid, an RNA or DNA base, a catalase or mixture thereof.
13. The method according to claim 11, characterized in that the biomolecule is at least one of glycine, alanine, valine, leucine, arginine, or mixtures thereof.
14. The method according to claim 11, characterized in that the biomolecule is at least one of guanine, adenine, thymine, cytosine or mixtures thereof.
15. The method according to claim 11, characterized in that the liquid suspension is a colloidal suspension comprised of particles having a size range between about 100 angstroms and about 10,000 angstroms.
16. The method according to claim 11, characterized in that the solution of the liquid suspension comprises water, alcohol, glycol, or combinations thereof.
17. 17. The method according to claim 11, characterized in that the vacuum in the chamber is less than 100 mTorr.
18. 18. The method of compliance with the claim 11, characterized in that the voltage applied to the conductive tip is positive or negative in the range of 15,000 volts.
19. 19. The method according to claim 11, characterized in that the liquid is atomized in a vacuum in the range of 100 mTorr up to atmospheric pressure. The method according to claim 11, characterized in that the biomolecule deposited on the substrate substantially retains the original structure / function.