JP6312128B2 - Method for producing phosphate-type ceramic thin film containing osteogenesis promoting substance and method for producing bone tissue implant having the thin film as a surface layer - Google Patents

Description

  The present invention relates to a phosphate-type ceramic thin film containing an osteogenesis promoting substance that can be applied to surface treatment of an implant used for implantation into bone tissue, a bone tissue implant having the thin film as a surface layer, and a method for producing the same. .

Dental implants that have been subjected to various surface treatments to impart biocompatibility, antibacterial properties, or osteogenesis promoting properties have been studied. One of them is a method of coating hydroxyapatite (Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ ; HA) excellent in biocompatibility. For example, a dental implant in which HA is coated by a sputtering method or a plasma spraying method. Compared to the case without coating, bone formation occurs quickly in the surroundings, and high bone bond strength can be obtained early.

  However, this dental implant may not be applicable to a state in which the bone mass or bone quality is insufficient, for example, an elderly person with low bone density, and a material having excellent osteoinductive ability is required. Conventionally, as an osteogenesis promoting substance, various elements or compounds are known in addition to calcium and phosphate contained in the HA. In recent years, strontium has been actively acting on bone formation and osteoblasts. It has been attracting attention as having an effect of suppressing the activity of osteoclasts as well as promoting its activity.

  For example, Patent Document 1 proposes an implantable article (implant) having a bioactive ceramic coating made of strontium-substituted apatite. Similarly, Patent Document 2 proposes a biocompatible strontium-substituted apatite ceramic produced by mixing calcium phosphate powder and strontium phosphate powder. In the strontium-substituted apatite coating disclosed in these patent documents, the strontium contained as an osteogenesis promoting substance is 33 mol% or less or 10 mol% or less in terms of mol% with respect to the amount of elements added with strontium and calcium. is there.

  On the other hand, it is well known or known in the field of medical implants to form a coating layer containing an antibacterial agent. For example, Patent Document 3 discloses a coating layer containing a bone forming substance and an antibacterial agent as bioactive substances. Medical implants having the following have been proposed.

JP 2004-358249 A US Pat. No. 5,441,536 Special table 2011-512957 gazette

  However, although the strontium-substituted apatite coating described in Patent Documents 1 and 2 contains strontium as a bone formation promoting substance, it does not have a remarkable effect on the osteoinductive ability as compared with the conventional HA coating, and therefore can be applied. The current situation is that little progress has been made.

  Moreover, although the coating layer containing the antibacterial agent described in Patent Document 3 is also effective for antibacterial properties, it is a conventional HA coating for improving the osteoinductive ability in a state where bone mass and bone quality are insufficient. A significant improvement could not be achieved. As described above, the composition and composition of the hydroxyapatite coating and the conventional strontium-substituted apatite coating do not provide sufficient effects for the effect of improving the osteoinductive ability, and further improvement and improvement are strongly demanded. Yes.

  The present invention has been made in view of the above-described conventional problems, and is a bone guide that is more than conventional in order to develop a technical problem of developing an implant material that can be applied in a state where bone mass and bone quality are insufficient. It is an object of the present invention to provide a ceramic thin film capable of promoting bone formation and improving adhesion or adhesion to jaw bone and cells by having a function, a bone tissue implant having the thin film as a coating layer, and a method for producing the same.

  In the present invention, among elements contained as an osteogenesis promoting substance, in addition to calcium, a predetermined metal element having osteoinductive ability is contained at a higher content than conventional strontium-substituted apatite coatings and the like. The present inventors have found that the above-mentioned problems can be solved by the phosphate ceramic thin film, and have reached the present invention.

That is, the configuration of the present invention is as follows.
[1] The present invention is a method for producing a phosphate ceramic thin film containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance, wherein the phosphate ceramic thin film is a bone The element contained as a formation promoting substance contains more than 20 atomic% with respect to the total amount of elements constituting the cation of the phosphate ceramic thin film, and the remaining element constituting the cation is calcium. Ah it is, after being film thick 0.1~5μm by sputtering, strontium ions, in alkaline aqueous solution containing at least one cation of the group consisting of magnesium ions and zinc ions or in a state of coexisting with the alkali aqueous solution Bone formation promoting substance-containing phosphate ceramic, characterized by being produced by hydrothermal treatment To provide a method of manufacturing a click film.
[2] The present invention is characterized in that the content of the element contained as an osteogenesis promoting substance is 40 to 90 atomic% with respect to the total amount of elements constituting the cation of the phosphate ceramic thin film. The method for producing a bone formation promoting substance-containing phosphate ceramic thin film according to the above [1] is provided.
[ 3 ] The present invention provides the osteogenesis promoting substance-containing phosphate according to [1] or [2] above, wherein the element contained as an osteogenesis promoting substance further contains an element having an antibacterial action. A method for producing a thin film of a mold ceramic is provided.
[ 4 ] The osteogenesis promoting substance-containing phosphate according to [ 3 ], wherein the element having an antibacterial action is at least one element selected from the group consisting of silver, copper and zinc. A method for producing a thin film of a mold ceramic is provided.
[ 5 ] The present invention is a method for producing a bone tissue implant having, as a surface layer, a bone formation promoting substance-containing phosphate ceramic thin film obtained by the production method according to any one of [1] to [4]. The following (A), (B), (C), (D) and (E), that is, (A) an apatite containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance Powder (B) Apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance and an element having antibacterial action,
(C) a powder obtained by mixing an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as a bone formation promoting substance, and an apatite powder containing calcium,
(D) Prepared by a solution method using an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as a bone formation promoting substance, a compound containing calcium and a compound containing an element having an antibacterial action And a powder obtained by mixing calcium and an apatite powder containing an element having an antibacterial action, and (E) containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance and an element having an antibacterial action Powder obtained by mixing apatite powder and calcium-containing apatite powder,
Forming a phosphate ceramic thin film with a thickness of 0.1 to 5 μm by performing sputtering treatment on the surface of the implant with a sputtering apparatus using any one powder selected from the group consisting of: Bone characterized by comprising a step of hydrothermally treating an acid salt type ceramic thin film in an alkaline aqueous solution containing at least one cation of the group consisting of strontium ions, magnesium ions and zinc ions or in the state of coexisting with the alkaline aqueous solution A method for manufacturing a tissue implant is provided.
[ 6 ] The present invention is a method for producing a bone tissue implant having, as a surface layer, a bone formation promoting substance-containing phosphate ceramic thin film obtained by the production method according to any one of [1] to [4]. In addition, when an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as a bone formation promoting substance and calcium are contained, a powder obtained by mixing apatite powder containing calcium is implanted by a sputtering apparatus. Forming a phosphate type ceramic thin film with a thickness of 0.1 to 5 μm on the surface of the substrate , and a group consisting of strontium ions, magnesium ions and zinc ions on the phosphate type ceramic thin film In an alkaline aqueous solution containing at least one cation or the alkaline water In the case where the hydrothermal treatment is performed in the state of coexisting with the solution, and further including an element having an antibacterial action, before or after the hydrothermal treatment of the phosphate ceramic thin film. There is provided a method for producing a bone tissue implant, comprising performing a step of adsorbing the antibacterial element on the phosphate ceramic thin film using a solution containing the antibacterial element.
[ 7 ] The method for producing a bone tissue implant according to [ 5 ] or [ 6 ] above, wherein the element having antibacterial action is composed of at least one element selected from the group consisting of silver, copper and zinc. To do.
[ 8 ] The present invention provides the method for producing a bone tissue implant according to any one of [5] to [7] , wherein the implant contains titanium or a titanium alloy.
[Effect of the invention]

  According to the present invention, at least one element of the group consisting of strontium, magnesium, and zinc that exhibits a sufficient function as an osteogenesis promoting substance is added to the total amount of elements constituting the cation of the phosphate ceramic thin film. By forming a phosphate-type ceramic thin film containing a large amount in excess of atomic percent, preferably in the range of 40 to 90 atomic percent, as an implant surface layer, the effect of osteoinductivity can be sufficiently obtained. . Thereby, since bone formation on the implant is promoted, adhesion or adhesion with the jawbone and cells can be improved. In addition, the bone tissue implant obtained in this way can be used even in a state of insufficient bone mass and bone quality, which has conventionally been difficult to provide, for example, elderly people with low bone density. It greatly contributes to the improvement of the safety of treatment and the expansion of its application range.

It is the schematic which shows the measuring method of the bone formation area by the calcification performed by seed | inoculating an osteoblast on the ceramic thin film of this invention. It is a figure which shows the measurement result of the bone formation area by the calcification performed by seed | inoculating an osteoblast on the ceramic thin film of this invention.

  As a result of various investigations on the type and composition of raw materials and the manufacturing method and conditions for the reason why the osteoinductive ability as expected in the conventional strontium-substituted apatite coating cannot be obtained, the present invention has shown that bone induction of strontium and the like is performed in the conventional coating It can be understood that the element having the ability was not in a state where the properties and characteristics could be fully utilized, and the superior bone inducing ability can be expressed by darely increasing the content of the element having the ability to induce osteogenesis than before. It was made based on the new knowledge that has never existed. The strontium-substituted apatite coatings disclosed in Patent Documents 1 and 2 have a very low strontium content compared to that of calcium, and the technical idea is that strontium is used as a trace amount of additive elements to modify calcium. ing. On the other hand, the present invention has a great feature in that a bone formation promoting substance such as strontium is contained in a large amount in the phosphate ceramic coating.

  Furthermore, the present invention has been found that among various elements known as osteogenesis promoting substances, the element of magnesium or zinc has an excellent osteoinductive ability like strontium. As an osteogenesis promoting substance, fluoride, silicon, vanadium, lithium, or the like is well known or known. However, these elements have almost the same or less inferior ability to calcium than calcium, so that conventional hydroxyapatite ( HA) Properties and performance beyond the coating cannot be realized. The osteogenesis promoting substance used in the present invention may be any substance composed of at least one element of the group consisting of strontium, magnesium and zinc, and contains one or more of these elements. Of these three elements, strontium has the most excellent osteoinductive ability, so that at least one of the three bone formation promoting substances preferably contains strontium.

  In the present invention, at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance is contained in an amount exceeding 20 atomic% with respect to the total amount of elements constituting the cation of the phosphate type ceramic thin film. And the balance of the elements constituting the cation is calcium. When the content of at least one element of the group consisting of strontium, magnesium and zinc is 20 atomic% or less, bone formation by calcification is almost the same as that of the conventional hydroxyapatite (HA) coating, and the osteoinductive ability is observed. Almost no effect can be obtained for a significant improvement. The effect of the osteoinductive ability becomes significant after the content of these elements exceeds 20 atomic%, and is prominent at 40 atomic% or more. This effect is continuously obtained at a content of 40 atomic% or more. On the other hand, when the content exceeds 90 atomic%, the crystallinity of the phosphate ceramic is greatly changed, and the phosphate ceramic The water solubility of the thin film becomes high, and a phenomenon occurs in which bone formed by calcification is detached together with a part of the surface layer of the phosphate ceramic thin film. As a result, an apparent decrease in osteoinductive ability may be observed. Therefore, in the present invention, the content of at least one element of the group consisting of strontium, magnesium and zinc is preferably specified in the range of 40 to 90 atomic%.

  Identification of each main apatite compound contained in the phosphate-type ceramic thin film containing an osteogenesis promoting substance obtained in the present invention can be performed, for example, by X-ray diffraction (XRD). In the XRD crystal pattern, typical peaks attributed to strontium apatite and hydroxyapatite are observed separately at different crystal angles. In the thin film in which strontium apatite and hydroxyapatite are mixed, another peak is observed between the crystal angles indicating both peaks, and depending on the degree of shift from the peak caused by hydroxyapatite, It is possible to estimate the mixing ratio to some extent. Further, when a substrate or implant is used when forming a phosphate-type ceramic thin film containing an osteogenesis promoting substance on the surface, a peak due to the substrate or implant is simultaneously observed in the XRD crystal pattern. For example, when titanium or a titanium alloy is used as a substrate or an implant, a peak due to titanium is observed. On the other hand, magnesium apatite and zinc apatite depend on their contents, but it is difficult to observe a clear peak in the XRD method. Therefore, the quantitative analysis of the content of at least one element of the group consisting of strontium, magnesium and zinc is usually performed by inductively coupled plasma emission spectroscopy (ICP-AES) described below.

  Quantitative analysis of elements by inductively coupled plasma optical emission spectrometry (ICP-AES) is performed using a nitric acid solution in which a phosphate-type ceramic thin film containing an osteogenesis promoting substance is dissolved, and a group consisting of strontium, magnesium and zinc The content of at least one element can be accurately determined. For example, in the case of a phosphate type ceramic thin film containing strontium apatite and hydroxyapatite, strontium (Sr) and calcium (Ca) are included as all elements constituting the cation of the phosphate type ceramic thin film. The Sr ratio Sr / (Ca + Sr) is calculated and defined as the Sr content of the present invention. In the present invention, it was confirmed that the charged amounts of strontium apatite and hydroxyapatite substantially correspond to the Sr ratio of the phosphate ceramic thin film obtained by the ICP-AES method. Therefore, it is easy to adjust the Sr content so that both raw materials have a desired Sr ratio by changing the charged amounts.

  In the present invention, the osteoinductive ability of the phosphate-type ceramic thin film containing an osteogenesis promoting substance is evaluated as follows. First, mouse-derived osteoblasts are seeded on the phosphate-type ceramic thin film of the present invention having a predetermined area, and cultured according to predetermined conditions. Then, the phosphate-type ceramic thin film is phosphate-buffered saline. Next, the bone formation area is measured by analyzing the image obtained by observing the phosphate-type ceramic thin film surface after the washing with a CCD camera. Osteoblasts cultured on the phosphate-type ceramic thin film exhibit a white color and show a color different from that of the substrate. Therefore, image analysis can be performed from the difference in color between the two. It means that it is a phosphate type ceramic thin film which has the outstanding osteoinductive ability, so that this bone formation area is large. Here, when comparing osteoinductivity between phosphate ceramic thin films with different areas, use the ratio of the area where bone formation was observed to the initial thin film area, not the bone formation area. Compare with each other. Since the bone formation area varies depending on the conditions of seeding and culturing of mouse-derived osteoblasts, it is desirable to compare the osteoinductive ability according to the type or composition of the phosphate ceramic thin film under the same conditions.

The seeding and culturing performed in the present invention is carried out in a container containing five samples of the phosphate-type ceramic thin film formed on a substrate having an area of 100 mm ² , as will be described later in the examples. When 1 dish is used, a condition is employed in which osteoblasts are seeded at 1 × 10 ⁴ cells / dish on a phosphate ceramic thin film and cultured at 37 ° C. for 21 days. This condition is suitable not only for general evaluation as a simple evaluation but also because it can clearly contrast the difference in osteoinductive ability of the phosphate ceramic thin film. The phosphate-type ceramic thin film containing an osteogenesis promoting substance according to the present invention is found to have an osteogenesis area measured according to these conditions of an average value of 5 test samples exceeding 20 mm ² and 25 mm ² or more. It is characterized by a higher value than the hydroxyapatite thin film.

  The phosphate-type ceramic thin film containing an osteogenesis promoting substance according to the present invention can be imparted with antibacterial properties by containing an element having an antibacterial action in addition to the excellent osteoinductive ability.

  The antibacterial agent used in the present invention is higher in safety than the organic antibacterial agent, can exhibit a wide range of antibacterial properties, and is excellent in durability and heat resistance. Therefore, an inorganic antibacterial agent is preferable. It is. Among them, the compatibility with the phosphate-type ceramic thin film of the present invention is good, and since a strong microbicidal or antibacterial property can be obtained in a small amount, at least one element of the group consisting of metals, particularly silver, copper and zinc, is selected. More preferably it is used. In the phosphate ceramic thin film according to the present invention, these metals may include a part of a cation composed of at least one element of the group consisting of strontium, magnesium and zinc and calcium, silver ions, copper ions or zinc ions. And can be incorporated as a constituent of the phosphate ceramic thin film. In addition, when zinc is included as an osteogenesis promoter in the phosphate ceramic thin film of the present invention, since the zinc also has a function as an antibacterial agent, it is not necessary to add an antibacterial agent.

  The content of at least one element of the group consisting of silver, copper and zinc is preferably 10 atomic% or less with respect to the total amount of elements constituting the cation of the phosphate mineral thin film of the present invention. When the content of these elements exceeds 10 atomic%, as a result, the content of at least one element of the group consisting of strontium, magnesium, and zinc that functions as an osteogenesis promoter is reduced. It becomes difficult to improve. Further, the antibacterial action due to silver ions, copper ions or zinc ions tends to be saturated when the content exceeds 10 atomic%, and further improvement of the antibacterial effect cannot be expected. In the present invention, when priority is given to improving the osteoinductive ability, the content of at least one element of the group consisting of silver, copper and zinc is more preferably 5 atomic% or less.

  A bone tissue implant can be obtained by forming a phosphate-type ceramic thin film containing an osteogenesis promoting substance obtained as described above on the surface of the implant. Examples of the bone tissue implant in the present invention include an artificial bone such as an artificial joint or an artificial tooth root. A preferred bone tissue implant in the present invention is an artificial tooth root.

  As a material used for the bone tissue implant, a metal such as titanium, zirconium, hafnium, tantalum, or an alloy thereof is used. These metals or alloys can form a relatively strong bond with the bone tissue, and can form a bond equivalent to or stronger than the bone tissue itself. Among these metals or alloys, titanium or titanium alloys are suitable as an implant for use in the present invention because they are excellent in biocompatibility and have a track record of use.

Metals such as titanium, zirconium, hafnium, tantalum or their alloys may be oxidized and covered with a thin oxide layer. For example, in a titanium implant, a surface layer made of titanium dioxide (TiO ₂ ) containing a small amount of Ti ₂ O ₃ , TiO and Ti ₃ O ₄ is formed, and the phosphate ceramic thin film and bone tissue of the present invention are formed. The bond between and may increase.

  The metal used as a material for implants is used before forming the phosphate-type ceramic thin film in order to increase adhesion or adhesion with the phosphate-type ceramic thin film containing an osteogenesis promoting substance according to the present invention. The surface may be roughened. By increasing the surface roughness, the contact and adhesion area between the implant and the phosphate ceramic thin film is increased, and the effect of increasing the adhesion strength or adhesion strength between the two can be obtained. The surface roughness is processed by a known or well-known method such as plasma spraying, mechanical blasting with a shot material, or etching with acid or the like, and the surface roughness is adjusted to a desired surface roughness by adjusting the surface submerging conditions. Can be formed.

  The phosphate ceramic thin film of the present invention formed as a surface layer on the implant is preferably formed with a thickness of 0.1 to 5 μm, more preferably 1 to 2 μm. When the thickness is less than 0.1 μm, bone formation by the osteoinductive ability is not promoted, and the bonding strength between the implant and the bone tissue is not improved. On the other hand, when the thickness exceeds 5 μm, it becomes difficult to uniformly form the phosphate-type ceramic thin film containing the bone formation promoting substance according to the present invention in the depth direction. Since the properties and characteristics of the thin film cannot be precisely controlled due to the non-uniformity of the composition, the bonding strength between the implant and the phosphate ceramic thin film may be reduced. In addition, forming the phosphate-type ceramic thin film of the present invention thickly takes a long time in the process, and an increase in manufacturing cost is inevitable. In the present invention, by defining this thickness in the range of 1 to 2 μm, sufficient expression of osteoinductivity and a reduction in manufacturing cost can be achieved at the same time.

  Next, a description will be given of a phosphate type ceramic thin film containing an osteogenesis promoting substance and a method for producing a bone tissue implant having the thin film as a surface layer according to the present invention.

  The method for producing a bone tissue implant of the present invention is roughly divided into two steps, a coating step by a sputtering method and a hot water treatment step. That is, on the surface of the implant metal or alloy, a powder comprising a compound of phosphoric acid and at least one element of the group consisting of strontium, magnesium and zinc which are osteogenesis promoting substances, and, if necessary, hydroxyapatite powder, Alternatively, bone formation is promoted by using a solution containing calcium and a compound containing an antibacterial element and a powder containing calcium and an element having an antibacterial action. A coating process in which a mixture mixed so that the content of the element as a substance is in a desired ratio is coated by a sputtering method to form a phosphate type ceramic thin film, and hot water is added with an alkaline aqueous solution after the coating process. This is a hot water treatment process.

  Here, in the case where a phosphate ceramic thin film not containing calcium as a constituent element has an antibacterial action, a compound containing at least one element of the group consisting of strontium, magnesium and zinc, which are osteogenesis promoting substances, and an antibacterial An apatite powder containing strontium, magnesium, or zinc and an element having an antibacterial action produced in a solution method using a compound containing an element having an action in a desired ratio can be used as a target at the time of sputtering. A powder containing a desired ratio of strontium, magnesium or zinc and an element having antibacterial action is mixed with a hydroxyapatite powder containing calcium to form a phosphate-type ceramic thin film containing calcium. May be used as another target material.

  The coating process by said sputtering method is performed as follows, for example. First, in the case of containing a powder composed of a compound of phosphoric acid and at least one element of the group consisting of strontium, magnesium and zinc, which are bone formation promoting substances, and at least one of calcium and an element having an antibacterial action, Each has calcium and antibacterial activity, prepared by a solution method using apatite powder containing calcium, or a compound containing calcium and a compound containing at least one element of the group consisting of silver, copper and zinc as antibacterial agents A powder obtained by mixing a powder containing an element is used as a target, and the target is attached to a target support portion provided in a vacuum chamber of a sputtering apparatus. When a phosphate ceramic thin film not containing calcium as a constituent element has an antibacterial action, a powder containing strontium, magnesium or zinc and an element having an antibacterial action in a desired ratio is used as a target. . Next, the vacuum chamber is evacuated by an evacuation unit having a device such as a vacuum pump to achieve a predetermined degree of vacuum. A DC high voltage is applied between the substrate or the implant and the target while an inert gas such as argon gas is introduced into the vacuum chamber from the gas introduction unit, and ionized argon (Ar) collides with the target. The atoms or molecules constituting the target evaporate with high energy by the collision of argon ions, and collide with the substrate or implant to form a film. In this way, a thin film tank having a substantially uniform thickness is formed on the substrate or the implant, and becomes a layer firmly bonded or adhered to the substrate or the implant. The substrate or the implant can be sputter-coated only at a desired position by performing sputtering while exposing only the portion where the thin film layer is to be formed and covering the other portion.

  The above sputtering apparatus may be provided with a mechanism for rotating the substrate or implant, which is a coating, in order to coat the surface of the substrate or implant with atoms or molecules that constitute the target three-dimensionally and uniformly.

  The phosphate-type ceramic thin film containing an osteogenesis promoting substance according to the present invention undergoes a hydrothermal treatment step of hydrothermal treatment with an alkaline aqueous solution after the coating step described above, thereby improving the crystallinity of the phosphate-type ceramic coating. Can be improved. In particular, when calcium is used as a cation constituting the phosphate ceramic coating, this hydrothermal treatment step is essential to improve low crystallinity. In a thin film containing only at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance, for example, a thin film containing only strontium exhibits crystallinity without performing a hydrothermal treatment step. Furthermore, since the behavior of increasing the number of cells adhering to the film is clearly observed by performing the hydrothermal treatment step, the hydrothermal treatment step is performed after the coating step in the present invention. The hydrothermal treatment step has not only an improvement in crystallinity but also an effect of removing other crystal phases such as CaO generated during coating, which is considered to have contributed to an increase in the number of adherent cells. In the present invention, not only the crystallinity of the phosphate-type ceramic thin film is increased, but also by increasing the number of adherent cells, it has the effect of promoting osteogenesis by enhancing osteoinductivity. It is preferable to do.

  The hydrothermal treatment step performed in the production method of the present invention is performed at 100 ° C. to 150 ° C. for 3 hours to 24 hours. In that case, hydrothermal treatment can be performed under stable conditions by using a pressure-resistant sealed container such as an autoclave as the processing container.

In addition, the hydrothermal treatment step is preferably performed in an alkaline aqueous solution or in the coexistence of an alkaline aqueous solution in order to reduce the solubility of the phosphate-type ceramic thin film containing an osteogenesis promoting substance. When acidic, the solubility of the thin film is high, and it is easy to dissolve even with neutral purified water, and the thin film is easily peeled or detached. The alkaline aqueous solution in the hydrothermal treatment is adjusted to pH 9 to pH 11, but preferably pH 9.5 to pH 10.5. These alkali solutions can be usually adjusted by appropriately adding an alkali such as NaOH, KOH, NH ₄ OH, etc. In the present invention, an alkali compound containing at least one element of the group consisting of strontium, magnesium and zinc Is preferably used. For example, in the case of a phosphate type ceramic thin film containing strontium as an osteogenesis promoting substance, the effect of suppressing detachment of strontium contained in the thin film is obtained by using Sr (OH) ₂ as an alkali compound in the hydrothermal treatment process. The obtained phosphate-type ceramic thin film having a uniform composition can be stably formed. In addition, there is an advantage that the strontium ratio in the phosphate ceramic thin film can be controlled to a substantially desired value depending on the raw material charging ratio. In particular, in the production of a phosphate ceramic thin film having a high strontium ratio of 40 atomic% or more, the strontium ratio in the thin film is controlled with high accuracy by using an aqueous solution containing Sr (OH) ₂ during hydrothermal treatment. It becomes possible. Specifically, when a thin film having a strontium ratio of 40 atomic% and 45 atomic% is prepared, when an aqueous solution of NaOH alone is used during hydrothermal treatment, the strontium ratio after hydrothermal treatment is 37.9 atomic% and 38 respectively. It was as low as 4 atomic%. Further, when 100 atomic% of the strontium ratio is hydrothermally treated with an aqueous solution of NaOH alone, the thin film may disappear. On the other hand, when using a Sr (OH) ₂ aqueous solution, the strontium ratio is not limited to 40 atomic% and 45 atomic%, and even if it is 80 atomic% or more, no decrease in the strontium ratio due to hydrothermal treatment is observed, A thin film having a desired strontium ratio could be produced. On the other hand, when forming a phosphate type ceramic thin film containing magnesium or zinc as a bone formation promoting substance, examples of the alkali compound used in the hydrothermal treatment step include Mg (OH) ₂ or Zn (OH) ₂ . Since these compounds are hardly soluble in water, in order to increase the solubility in water, the temperature of the aqueous solution may be increased, or a mixture with other alkali compounds may be used. In addition to the above hydroxides, oxides (SrO, MgO, or ZnO) can be used in combination with other alkali compounds.

  When an element having an antibacterial action is contained in the method for producing a bone tissue implant of the present invention, a solution containing an element having an antibacterial action is used instead of using the compound containing the element having an antibacterial action described above. Then, a step of adsorbing the element having antibacterial action to the phosphate ceramic thin film of the present invention may be employed. Here, the solution containing an element having an antibacterial action is preferably a solution containing nanoparticles composed of at least one element of the group consisting of silver, copper and zinc. Among them, the antibacterial action in the adsorbed state is the most. Silver nanoparticles that can be made higher are more preferred. By using the solution containing nanoparticles, not only can the element having antibacterial action rapidly penetrate into the pores of the phosphate ceramic thin film of the present invention, but also uniform fixing can be performed. The particle diameter of the nanoparticles is in the range of 4 to 100 nm in terms of average primary particle diameter. When the average primary particle size is less than 4 nm, the nanoparticles are likely to aggregate and the handling property of the solution is inferior. When the average primary particle size exceeds 100 nm, the impregnation property and uniformity of the nanoparticles are deteriorated. The adsorption process of the element having antibacterial action may be performed either before or after the hydrothermal treatment of the phosphate ceramic thin film. Among them, it is preferable to perform the adsorption step after the hydrothermal treatment step, because the desorption of nanoparticles adsorbed on the phosphate ceramic thin film of the present invention can be reduced.

  EXAMPLES Next, although an Example demonstrates this invention further more concretely, this invention is not limited to these Examples.

<Examples 1-4, Comparative Examples 1-2>
[Sample preparation]
A strontium-containing phosphate compound [Sr ₁₀ (PO ₄ ) prepared by a solution method and baked at 800 ° C. as a target used for forming a phosphate-type ceramic thin film containing an osteogenesis promoting substance by sputtering. ₆ (OH) ₂ : SrAp] powder, hydroxyapatite [Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ : HA] powder [manufactured by Taihei Chemical Industrial Co., Ltd.], and SrAp and HA powder [Sr / SrAp thin films, HA thin films, and thin films containing Sr and Ca were prepared using powders mixed so that (Sr + Ca) was 20, 40, 60, and 80 atomic%, respectively. Film formation was performed with a sputtering apparatus SPF-210HD [Canon Anelva Co., Ltd.] using a titanium (Ti) plate [Nicola Co., Ltd.] as a substrate. The sputtering conditions were a gas pressure of 0.5 Pa and a discharge power of 100 W, and a thin film with a thickness of 1.0 μm was produced. Furthermore, hydrothermal treatment was performed in order to improve the crystallinity of the thin film. When performing the hydrothermal treatment, the SrAp thin film and the thin film containing Sr and Ca were used in a Sr (OH) ₂ aqueous solution using a stainless steel container, and the HA thin film was used in a NaOH aqueous solution using the same container. . The pH of both aqueous solutions was adjusted to 9.5. As for the conditions of the hydrothermal treatment, the temperature is 120 ° C., the pressure is 0.20 Mpa, and the treatment time is 24 hours.

  Samples are Sr0, Sr20, Sr40, Sr60, Sr80, and Sr100 in which the Sr ratio in the thin film subjected to hydrothermal treatment is 0 atomic%, 20 atomic%, 40 atomic%, 60 atomic%, 80 atomic%, and 100 atomic%, respectively. Various measurements and evaluations of the thin film were performed as shown below. Here, Sr0 and Sr20 are referred to as Comparative Examples 1 and 2, respectively, and Sr40, Sr60, Sr80 and Sr100 are referred to as Examples 1, 2, 3 and 4, respectively. In addition, a cell adhesion test is also performed on a Ti substrate alone on which no thin film is formed, and the sample is referred to as Reference Example 1.

[Thin film evaluation method]
(1) The produced thin film was subjected to XRD measurement using an X-ray diffractometer RINT2000 [Rigaku Corporation] to evaluate the product identification and the crystallinity of the film.
(2) The Sr ratio of the produced thin film was determined by measurement with an inductively coupled plasma atomic emission spectrometer [ICP-AES: ICPS-7510, Shimadzu Corporation].

[Measurement method of bone formation area]
The bone formation area by the calcification in which osteoblasts were seeded and cultured on the prepared thin film by the procedure shown in FIG. 1 was measured. The bone formation area is a physical quantity that reflects the osteoinductive ability of each thin film, and means that the thin film having a larger measured value has higher osteoinductive ability. As shown in FIG. 1, first, the osteoblasts were seeded on a phosphate ceramic thin film 1 formed on a Ti substrate using fresh rat primary culture osteoblasts. Here, seeding was performed using a 35 mm square container as 1 dish, and 5 sample plates of the phosphate ceramic thin film 1 were put in the container at a cell number of 1 × 10 ⁴ cells / dish. One sample plate has a square shape of 10 × 10 mm and an area of 100 mm ² . Subsequently, it culture | cultivated at 37 degreeC for 21 days using (alpha) -MEM (Gibco). After culturing, the thin film was washed with phosphate buffered saline, the sample was taken with a CCD camera, and the image 2 obtained by scanning was used for image analysis to calculate the bone formation area due to calcification. In the image 2 shown in FIG. 1, the white portion corresponds to the bone formation portion observed after the culture. Here, the measurement of the bone formation area is expressed as an average value of five sample plates.

  According to the XRD measurement results of each thin film after hydrothermal treatment, representative peaks of HA and SrAp corresponding to Sr0 and Sr100, respectively, were observed. In Sr20-Sr80, the peak of both Sr ratios was shifted to the lower angle side, and Sr solid solution was confirmed. This is presumably because Sr having a larger atomic radius than Ca enters the HA crystal and the interplanar spacing is increased.

  Further, when the Sr ratio [Sr / (Sr + Ca)] in the thin film calculated from the ICP measurement was compared with the Sr ratio calculated from the raw material charge amount of the target, the Sr ratio almost matched within 3%. From this result, it can be seen that by changing the Sr ratio in the target, it is possible to produce thin films having different Sr ratios, and the Sr ratio can be almost controlled.

The measurement result of the bone formation area mineralized using the osteoblast in FIG. 2 is shown. The bone formation area tends to increase after the Sr ratio exceeds 20 atomic%, a significant increase is observed after 40 atomic%, and the maximum is reached at 60 atomic%. When the Sr ratio is 40 atomic% or more (Examples 1 to 4), the bone formation area has a high value of 25 mm ² or more on average. On the other hand, when the Sr ratio is 20 atomic% or less (Comparative Examples 1 and 2), although the bone formation area is increased as compared with the Ti substrate alone (Reference Example 1), the average value of the bone formation area is 20 mm ^2. It is less than the thin film of Examples 1-4, and has shown the small value. Thus, it turns out that the phosphate ceramic thin film of this invention containing more Sr than before has a high effect with respect to osteoblast activation.

As shown in FIG. 2, even when the Sr ratio is 100 atomic% (Example 4), a high bone formation area of 25 mm ² or more is obtained. However, when the Sr ratio exceeds 90 atomic%, the crystallinity and the like of the phosphate-type ceramic is greatly changed, the water-solubility of the phosphate-type ceramic thin film is increased, and the bone formed by calcification is The phenomenon of desorption with part of the surface layer of the phosphate ceramic thin film tends to occur. Thereby, the measured value of the bone formation area is apparently 25 mm ² or less, and the osteoinductive ability may be reduced. Therefore, in the present invention, it is preferable to define the Sr ratio in the range of 40 to 90 atomic%.

  So far, for increasing the bone mass, complexing with a biomolecule such as growth factor (BMP) has attracted attention, but it has been difficult to put it to practical use in terms of the cost and the origin of BMP. If an inorganic substance such as strontium is effective for bone augmentation as in the present invention, a bone tissue implant can be expected to be obtained at a relatively low cost by mixing a strontium source with the raw material of an existing HA coating system. .

<Example 5>
The same SrAp powder as that used in Examples 1 to 4 and calcium nitrate tetrahydrate [Ca (Ca ( _{NO 3) 2 · 4H 2 O} ], and silver nitrate (powder containing Ca and Ag obtained by firing in prepared 800 ° C. in a solution method using _{AgNO 3) [(Ca + Ag} ) powder as the antimicrobial component And a thin film containing Sr, Ca and Ag using a powder mixed so that the ratio of each element is Sr: Ca: Ag = 60 atomic%: 35 atomic%: 5 atomic% did. Film formation was performed under the same sputtering conditions using the same sputtering apparatus as in Examples 1 to 4, and a thin film with a thickness of 1.0 μm was produced. Furthermore, hydrothermal treatment was performed in order to improve the crystallinity of the thin film. Hydrothermal treatment conditions are the same as those in Examples 1-4.

The thin film thus produced was measured for the area of bone formation due to calcification in which rat osteoblasts were seeded and cultured under the same method and conditions as in Examples 1-4. As a result, the calcified bone formation area was 27 mm ² as an average value of five samples, and it was confirmed that the bone inducing ability was almost equivalent to that of Example 2.

  In addition, in order to evaluate the antibacterial property of the thin film of this example, an antibacterial activity test using Escherichia coli was conducted in a simulated manner. The thin film of this example on which the Ti substrate was formed was placed in phosphate buffered saline together with the Ti substrate, and the antibacterial activity against E. coli was measured by adding the bacterial solution of E. coli. After the E. coli was allowed to stand at room temperature for 24 hours, it was below the detection limit value for counting the number of bacteria, and E. coli was not detected. Thus, it was confirmed that the thin film of this example has excellent antibacterial properties.

  In addition to Ag as an antibacterial agent, a thin film containing 5 atomic% of Cu or Zn was formed on a Ti substrate, and the bone formation area was measured and the antibacterial activity test was performed under the same method and conditions as described above. Here, when Cu or Zn is contained in a thin film, a powder of copper nitrate or zinc nitrate is used as a raw material, respectively, and a (Ca + Cu) powder or (Ca + Zn) powder is prepared by the same method as described above and mixed with SrAp powder. Sputtering is performed using the powder as a target. The ratio of Sr, Ca and Cu (or Zn) was adjusted to be the same as that of the thin film containing Ag. As a result, it was confirmed that the bone formation area due to calcification and the number of bacteria after standing at room temperature for 24 hours were almost the same as those of the thin film containing Ag.

<Example 6>
The same SrAp powder and HA powder as used in Example 2 were used as targets used to form a phosphate-type ceramic thin film containing an osteogenesis promoting substance by sputtering, and the Sr ratio was 60 atomic%. A thin film containing Sr and Ca was prepared. Film formation was performed using the same sputtering apparatus as in Example 2 under the same sputtering conditions to produce a thin film with a thickness of 1.0 μm. Furthermore, hydrothermal treatment was performed in order to improve the crystallinity of the thin film. The hydrothermal treatment conditions are the same as in Example 2. Thereafter, using a diluted citrate buffer solution in which silver nanoparticles having an average primary particle size of 60 nm are dispersed as an antibacterial agent, the thin film containing Sr and Ca after hydrothermal treatment is placed in the diluted citrate buffer solution. The silver nanoparticles were adsorbed while being stirred at room temperature for 24 hours, and then dried at 200 ° C. The thin film after drying is subjected to quantitative analysis by ICP-AES so that the ratio of each element of Sr, Ca and Ag is Sr: Ca: Ag = 57 atomic%: 38 atomic%: 5 atomic%. It was prepared by adjusting.

  The thin film thus produced was subjected to the measurement of the bone formation area by calcification and the antibacterial activity test using Escherichia coli under the same method and conditions as in Example 5. As a result, it was confirmed that the osteoinductive ability and antibacterial properties were almost the same as in Example 5.

<Example 7>
As a target used to form a phosphate-type ceramic thin film containing an osteogenesis promoting substance by sputtering, the same SrAp powder as used in Examples 1 to 4, HA powder, and a solution method were used. Magnesium-containing phosphate compound [Mg ₁₀ (PO ₄ ) ₆ (OH) ₂ : MgAp] powder calcined at 800 ° C. in a ratio of Sr: Mg: Ca = 55 atomic%: 5 atomic%: 40 atomic% A thin film containing Sr, Mg, and Ca was prepared using the powder mixed so as to be. Film formation was performed under the same sputtering conditions using the same sputtering apparatus as in Examples 1 to 4, and a thin film with a thickness of 1.0 μm was produced. Furthermore, hydrothermal treatment was performed in order to improve the crystallinity of the thin film. When performing the hydrothermal treatment, a small amount of Mg (OH) ₂ was mixed in the Sr (OH) ₂ aqueous solution using a stainless steel container, and the pH of the aqueous solution was adjusted to about 9.5. The conditions for the hydrothermal treatment are the same as in Examples 1 to 4.

The bone formation area by calcification after seeding and culturing rat osteoblasts with the same method and conditions as in Examples 1 to 4 was measured. As a result, a value of 29 mm ² was measured as an average value of five samples for the calcified bone formation area, and it was confirmed that the bone inducing ability was equal to or greater than that of Example 2.

<Example 8>
As a target used to form a phosphate-type ceramic thin film containing an osteogenesis promoting substance by sputtering, the same SrAp powder as used in Examples 1 to 4, HA powder, and a solution method were used. A zinc-containing phosphate compound [Zn ₁₀ (PO ₄ ) ₆ (OH) ₂ : ZnAp] powder calcined at 800 ° C. is used as Sr: Zn: Ca = 59.5 atomic%: 0.5 atomic%: 40. A thin film containing Sr, Zn and Ca was produced using powder mixed so as to have a ratio of atomic%. Film formation was performed under the same sputtering conditions using the same sputtering apparatus as in Examples 1 to 4, and a thin film with a thickness of 1.0 μm was produced. Furthermore, hydrothermal treatment was performed in order to improve the crystallinity of the thin film. When performing the hydrothermal treatment, a very small amount of Zn (OH) ₂ was mixed in the Sr (OH) ₂ aqueous solution using a stainless steel container, and the pH of the aqueous solution was adjusted to about 9.5. The conditions of the hydrothermal treatment are the same as in Examples 1 to 4.

The bone formation area by calcification after seeding and culturing rat osteoblasts with the same method and conditions as in Examples 1 to 4 was measured. As a result, the calcified bone formation area was 28 mm ² as an average value of five samples, and it was confirmed that the bone inducing ability was almost equivalent to that of Example 2.

  As described above, a phosphate ceramic thin film containing at least one element of the group consisting of strontium, magnesium and zinc that exhibits a sufficient function as an osteogenesis promoting substance more than a conventional strontium-substituted apatite coating or the like By forming as an implant surface layer, the effect of osteoinductive ability is sufficiently obtained. Thereby, since bone formation on the implant is promoted, adhesion or adhesion with the jawbone and cells can be improved. The bone tissue implant obtained in this way can be used even in a state in which the bone mass and bone quality, which has been conventionally difficult to provide, are insufficient, for example, elderly people with low bone density, etc. It greatly contributes to the improvement of safety and the expansion of its application range. In addition, the bone tissue implant of the present invention can be applied not only to artificial bones or artificial tooth roots such as artificial joints, but also to implants in various fields that need to promote bone regeneration, and its usefulness is extremely high. .

DESCRIPTION OF SYMBOLS 1 ... Phosphate ceramic capsule, 2 ... Image after bone formation by calcification.

Claims (8)

A method for producing a phosphate-type ceramic thin film containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance,
The phosphate ceramic thin film contains the element contained as an osteogenesis promoting substance in excess of 20 atomic% with respect to the total amount of elements constituting the cation of the phosphate ceramic thin film, and the Ri elements calcium der residual that make up the cation,
After forming a film with a thickness of 0.1 to 5 μm by a sputtering method, hydrothermal treatment is performed in an alkaline aqueous solution containing at least one cation of the group consisting of strontium ions, magnesium ions and zinc ions or in the coexistence with the alkaline aqueous solution. A method for producing a phosphate ceramic thin film containing an osteogenesis promoting substance, characterized in that it is produced by performing . The content of the element contained as an osteogenesis promoting substance is 40 to 90 atomic% with respect to the total amount of elements constituting the cation of the phosphate ceramic thin film. A method for producing a phosphate-type ceramic thin film containing an osteogenesis promoting substance. 3. The method for producing a phosphate-type ceramic thin film containing an osteogenesis promoting substance according to claim 1, wherein the element contained as the osteogenesis promoting substance further contains an element having an antibacterial action. 4. The method for producing a bone formation promoting substance-containing phosphate ceramic thin film according to claim 3 , wherein the element having antibacterial action comprises at least one element selected from the group consisting of silver, copper and zinc. A method of manufacturing a bone tissue implant having a bone formation promoting material containing phosphates ceramic thin film obtained by the process according to any one of claims 1 to 4 as a surface layer, the following (A), ( B), (C), (D) and (E), that is, (A) an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance,
(B) an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance and an element having an antibacterial action,
(C) a powder obtained by mixing an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance and an apatite powder containing calcium,
(D) Prepared by a solution method using an apatite powder containing at least one element of the group consisting of strontium, magnesium and zinc as a bone formation promoting substance, a compound containing calcium and a compound containing an element having an antibacterial action And a powder mixed with calcium and an apatite powder containing an element having an antibacterial action, and (E) at least one element of the group consisting of strontium, magnesium and zinc as an osteogenesis promoting substance and an element having an antibacterial action Powder obtained by mixing apatite powder and calcium-containing apatite powder,
Forming a phosphate ceramic thin film with a thickness of 0.1 to 5 μm by performing sputtering treatment on the surface of the implant with a sputtering apparatus using any one powder selected from the group consisting of: Bone characterized by comprising a step of hydrothermally treating an acid salt type ceramic thin film in an alkaline aqueous solution containing at least one cation of the group consisting of strontium ions, magnesium ions and zinc ions or in the state of coexisting with the alkaline aqueous solution A method for manufacturing a tissue implant. A method for producing a bone tissue implant having a bone formation promoting substance-containing phosphate ceramic thin film as a surface layer obtained by the production method according to any one of claims 1 to 4 , wherein the bone formation promoting substance is strontium, When apatite powder containing at least one element of the group consisting of magnesium and zinc and calcium are contained, a powder obtained by mixing the apatite powder containing calcium is sputtered on the surface of the implant by a sputtering apparatus to obtain phosphorus. A step of forming an acid salt ceramic thin film with a thickness of 0.1 to 5 μm , and an alkaline aqueous solution containing at least one cation of the group consisting of strontium ions, magnesium ions and zinc ions in the phosphate ceramic thin film water or condition that coexists with the alkali aqueous solution An element having an antibacterial action before or after the step of hydrothermally treating the phosphate-type ceramic thin film when an element having an antibacterial action is further included. A method for producing a bone tissue implant, comprising performing a step of adsorbing the element having an antibacterial action on the phosphate ceramic thin film using a solution containing. The method for producing a bone tissue implant according to claim 5 or 6 , wherein the element having an antibacterial action comprises at least one element of the group consisting of silver, copper and zinc. The method for producing a bone tissue implant according to any one of claims 5 to 7 , wherein the implant contains titanium or a titanium alloy.

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