A MEDICAL DEVICE HAVING A SURFACE COMPRISING ANTIMICROBIAL
METAL
Field of the invention
The present invention relates to a medical device having a surface
layer comprising an anti-microbial metal, and to methods of producing such a
device.
Background of the invention
For any type of medical device intended for contact with living tissue,
biocompatibility is a crucial issue. The risk for foreign body reaction, clot
formation and infection, among many other things, must be addressed and
minimized in order to avoid adverse effects, local as well as systemic, which
may otherwise compromise the health of the patient and/or lead to failure of
the device. This is particularly the case for permanent implants.
Healing or regeneration of tissue around an implant is often vital in
order to secure the implant and its long-term functionality. This is especially
important for load-bearing implants such as dental or orthopedic implants.
Dental implant systems are widely used for replacing damaged or lost
natural teeth. In such implant systems, a dental fixture (screw), usually made
of titanium or a titanium alloy, is placed in the jawbone of the patient in order
to replace the natural tooth root. An abutment structure is then attached to the
fixture in order to build up a core for the part of the prosthetic tooth protruding
from the bone tissue, through the soft gingival tissue and into the mouth of the
patient. On said abutment, the prosthesis or crown may finally be seated.
For dental fixtures, a strong attachment between the bone tissue and
the implant is necessary. For implants intended for contact with soft tissue,
such as abutments which are to be partially located in the soft gingival tissue,
also the compatibility with soft tissue is vital for total implant functionality.
Typically, after implantation of a dental implant system, an abutment is
partially or completely surrounded by gingival tissue. It is desirable that the
gingival tissue should heal quickly and firmly around the implant, both for
medical and aesthetic reasons. A tight sealing between the oral mucosa and
the dental implant serves as a barrier against the oral microbial environment
and is crucial for implant success. This is especially important for patients
with poor oral hygiene and/or inadequate bone or mucosal quality. Poor
healing or poor attachment between the soft tissue and the implant increases
the risk for infection and peri-implantitis, which may ultimately lead to bone
resorption and failure of the implant.
There are several strategies for increasing the chances of a successful
implantation of a medical device, for example enhancing the rate of new
tissue formation and/or, in instances where tissue-implant bonding is desired,
enhancing the rate of tissue attachment to the implant surface, or by
reducing the risk for infection. Enhancement of new tissue formation may be
achieved for example by various surface modifications and/or deposition of
bioactive agents on the surface.
The risk of infection in connection with dental implants is today
primarily addressed by preventive measures, such as maintaining good oral
hygiene. Once a biofilm is formed on the surface of a dental implant, it is
difficult to remove it by applying antibacterial agents. In the case of infection
in the bone or soft tissue surrounding a dental implant (peri-implantitis),
mechanical debridement is the basic element, sometimes in combination with
antibiotics, antiseptics, and/or ultrasonic or laser treatment.
Summary of the invention
It is an object of the present invention to overcome this problem, and to
provide a medical device, such as an implant, having a surface which reduces
the risk for infection upon contact of the medical device with living tissue.
According to a first aspect of the invention, this and other objects are
achieved by medical device intended for contact with living tissue, comprising
a substrate having a surface layer comprising one or more compound(s) of a
non-toxic post-transition metal.
The layer comprising one or more compound(s) of a non-toxic posttransition
metal, e.g. gallium compound(s), may have an atomic concentration
(at%) of post-transition metal, e.g. gallium and/or bismuth, of at least 5 at%. In
embodiments of the invention, the gallium concentration in said layer is at
least 10 at%, for example at least 15 at%, e.g. at least 20 at%. However in
other embodiments of the invention, the content of the post-transition metal
may be less than 5 %, for example at least 0.05 at%. The layer may have a
gallium content of up to 50 at%.
In embodiments of the invention, the compound of a non-toxic posttransition
metal constitutes the major part of the layer.
A medical device surface having a layer incorporating a non-toxic posttransition
metal such as gallium or bismuth has been shown to be effective
against various bacterial strains, and was shown to inhibit biofilm formation in
vitro. The medical device according to the invention may also be effective
against other microbes, such as fungi.
In embodiments of the invention said living tissue is soft tissue.
Alternatively, said living tissue may be cartilage or bone tissue.
In embodiments of the invention, said one or more compound(s) of a
non-toxic post-transition metal also comprises an additional metal, for
example a biocompatible metal such as titanium. Titanium is known to be well
tolerated by living tissue, and has been used as an implant material for many
years. By including a biocompatible metal, e.g. titanium, in the galliumcontaining
layer, a surface is obtained which is more similar to established
implant surfaces and which is even more likely to be well tolerated by living
tissue.
In embodiments of the invention, the surface layer may be a metallic
layer. The compound comprising a non-toxic post-transition metal may be a
metallic compound.
In embodiments of the invention, the non-toxic post-transition metal is
selected from bismuth and gallium. Hence, the compound(s) of the non-toxic
post-transition metal may be selected from bismuth compound(s) and gallium
compound(s).
A galliunn compound may be selected from the group consisting of
gallium-titanium oxide, gallium nitride, gallium-titanium nitride, gallium
carbide, gallium selenide, and gallium sulphide, gallium chloride, gallium
fluoride, gallium iodide, gallium oxalate, gallium phosphate, gallium maltolate,
gallium acetate and gallium lactate. A bismuth compound may be selected
from the group consisting of bismuth-titanium, bismuth-titanium oxide,
bismuth-titanium nitride, bismuth nitride, bismuth carbide, bismuth selenide,
and bismuth sulphide, bismuth chloride, bismuth fluoride, bismuth iodide,
bismuth oxalate, bismuth phosphate, bismuth maltolate, bismuth acetate and
bismuth lactate.These compounds of a non-toxic post-transition metals are in
general well tolerated by living tissue of a mammal, and can be deposited on
a surface using a thin film deposition technique.
In embodiments, the compound of a non-toxic post-transition metal is a
nitride of at least one non-toxic post-transition metal, such as gallium nitride
or bismuth nitride, optionally also comprising titanium. Nitrides containing
non-toxic post-transition metals such as gallium or bismuth are particularly
useful in the present invention, in particular for dental implant applications,
because such compounds may provide an aesthetically desirable surface
layer, in particular with respect to color. Such nitrides can be deposited using
thin film deposition techniques. The nitride of a non-toxic post-transition metal
may be selected from the group consisting of gallium-titanium nitride, gallium
nitride, bismuth-titanium nitride, bismuth nitride, and gallium-bismuth-titanium
nitride.
In embodiments of the invention, the layer comprising a compound of a
non-toxic post-transition metal may further comprise a salt of a non-toxic posttransition
metal, e.g. a gallium salt or a bismuth salt. The salt may be
deposited onto the surface layer comprising the compound of a non-toxic
post-transition metal. For example a gallium salt or a bismuth salt may be
deposited onto a first layer comprising a first gallium or bismuth compound. A
salt deposit may increase the release of antimicrobial post-transition metal
from the surface early after contact with living tissue, thus temporarily further
enhancing an antibacterial or antimicrobial effect of the layer.
Generally, the layer comprising the compound of a post-transition
metal may have a thickness in the range of from 10 nm to 1.5 mhh . A layer of
at least 10 nm may be sufficient to provide a desirable antibacterial effect,
whereas thick layers of up to 1 mhh may be desirable for aesthetic reasons,
having a color suitable for e.g. dental implants.
Typically, in embodiments of the invention, the layer comprising the
compound of a non-toxic post-transition metal may be a homogeneous layer.
The layer may also be a non-porous layer. A non-porous layer is typically less
susceptible of bacterial growth and biofilm formation compared to a porous
layer.
The substrate on which the layer comprising the at least one gallium
compound is provided may comprise a metallic material, typically a
biocompatible metal or alloy such as titanium or titanium alloy. Alternatively,
the substrate may comprise a ceramic material. In other embodiments, the
substrate may comprise a polymeric material, or a composite material.
The medical device of the invention is typically an implant intended for
long-term contact with, or implantation into, living tissue. In one embodiment
the medical device is an implant intended for implantation at least partially
into soft tissue. In yet other embodiments of the invention, the medical device
may be intended for short-term or prolonged contact with living tissue,
typically soft tissue.
For example, the medical device may be a dental implant, in particular
a dental abutment. In another embodiment, the medical device may be a
bone anchored hearing device. In another embodiment, the medical device
may be an orthopaedic implant. In another embodiment, the medical device
may be a stent. In another embodiment, the medical device may be a shunt.
As another example, the medical device may be a catheter adapted for
insertion into a bodily cavity such as a blood vessel, the digestive tract or the
urinary system.
In another aspect, the invention provides a method of producing a
medical device as described herein, comprising
a) providing a substrate having a surface; and
b) applying a compound of a non-toxic post-transition metal onto said
surface to form a layer.
Typically, the compound of a non-toxic post-transition metal is a nitride of a
non-toxic post-transition metal, such as a nitride of gallium and/or bismuth.
In some embodiments, step b) involves simultaneously or sequentially
applying a non-toxic post-transition metal and an additional metal or metal
compound onto said surface. The additional metal or metal compound may
comprise titanium. Applying the post-transition metal, and optionally also an
additional metal or metal compound, can be achieved using a thin film
deposition technique.
A medical device as described above may be used for preventing
biofilm formation and/or bacterial infection of a surrounding tissue, in
particular soft tissue. In particular the medical device of the invention may be
used for preventing bacterial infection of gingival tissue and/or periimplantitis.
It is noted that the invention relates to all possible combinations of
features recited in the claims.
Brief description of the drawings
Figure 1 is a side view of a medical device according to an
embodiment of the invention, wherein the medical device is a dental
abutment.
Figure 2 illustrates in cross-section part of a medical device according
to embodiments of the invention, showing a substrate material and a layer
comprising a gallium compound.
Detailed description of the invention
The present inventor has found that a medical device having a surface
layer comprising a compound of a post-transition metal, in particular a no n
toxic, antimicrobial post-transition metal, such as gallium and/or bismuth,
provides very advantageous effects in terms of reduced risk of infection,
improved tissue healing and/or aesthetic performance. The inventor has
demonstrated that a titanium body having a surface coating incorporating
gallium (Ga) can prevent the growth of bacteria on and around the surface
and thus may be useful in preventing detrimental infection around e.g. a
dental abutment implanted into the gingiva. It has also been demonstrated
that a titanium body having a surface coating incorporating bismuth (Bi) can
prevent the growth of bacteria on and around the surface and thus may be
useful in preventing detrimental infection around e.g. a dental abutment
implanted into the gingiva.
According to the present invention, a tissue contact surface of a
surface of a medical device comprises a compound of a post-transition metal.
Typically, the post-transition metal is non-toxic to mammalian cells at the
concentrations that have a lethal effect on bacterial cells.
The term "post-transition metal" generally refers to metal elements
found in groups 13-1 6 and periods 3-6 of the periodic table. Usually,
aluminium, gallium, indium, thallium, tin, lead, bismuth and polonium are
regarded as post-transition metals. In contrast, transition metals are formed of
group 3-1 2 elements. Germanium and antimony are considered not as posttransition
metals, but metalloid elements.
Group 16 of the periodic table only contains one post-transition metal:
polonium, which is toxic. Hence, in the present invention, post-transition
metals of groups 13-15 and periods 3-6 are preferred.
In embodiments of the invention, the post-transition metal used is non
toxic.
As used herein, "non-toxic" means that the substance (e.g. a
compound or element) in question does not damage mammalian cells at
concentrations that have a lethal effect on bacterial cells.
Examples of post-transition metals that are toxic include thallium (Tl),
lead (Pb) and polonium (Po). Other post-transitional metals (e.g. indium (In)
and tin (Sn)) may be considered non-toxic in the pure metal form, but may be
toxic in other forms or when they form compounds with other elements.
The non-toxic post transition metal used in the present invention is
typically non-toxic when present in elementary form, as metal ions, and/or as
one of the exemplary compounds given herein.
Furthermore, the post-transition metal used in the present invention
typically has an antimicrobial or antibacterial effect. Post-transition metals that
are considered to have some antimicrobial or antibacterial effect (including
any oligodynamic effect) include at least gallium, tin, lead, and bismuth.
In view of the above, the invention may employ at least one non-toxic,
antimicrobial post-transition metal, which is preferably selected from gallium
and bismuth.
For example, a gallium compound may be applied to a medical device
as a surface layer. Alternatively, a gallium compound could be incorporated
into at least part of a body forming a medical device, such that at least one
surface of the device comprises the gallium compound.
As another example, a bismuth compound may be applied to a medical
device as a surface layer. Alternatively, a bismuth compound could be
incorporated into at least part of a body forming a medical device, such that at
least one surface of the device comprises the bismuth compound.
Gallium has been used in medicine at least since the 1940's, primarily
as a radioactive agent for medical imaging. The antibacterial properties of
gallium have been investigated in several studies. In Kaneko et al. (2007) it
was established that gallium nitrate (Ga(NO3)3) inhibits growth of
Pseudomonas aeruginosa in batch cultures. Olakanmi et al (201 0) found that
Ga(NOs)3 inhibited the growth of Francisella novicida. Gallium acts by
disrupting iron metabolism. It may be assumed that gallium is also effective
against other microbes, e.g. fungi such as yeasts or moulds.
Bismuth is known to possess antibacterial activity. Bismuth compounds
were formerly used to treat syphilis, and bismuth subsalicylate and bismuth
subcitrate are currently used to treat peptic ulcers caused by Helicobacter
pylori. The mechanism of action of this substance is still not well understood.
Bibrocathol is an organic bismuth-containing compound used to treat eye
infections, and bismuth subsalicylate and bismuth subcarbonate are used as
ingredients in antidiarrheal pharmaceuticals.
Directive 2007/47/ec defines a medical device as: "any instrument,
apparatus, appliance, software, material or other article, whether used alone
or in combination, including the software intended by its manufacturer to be
used specifically for diagnostic and/or therapeutic purposes and necessary for
its proper application, intended by the manufacturer to be used for human
beings". In the context of the present invention, only medical devices intended
for contact with living tissue are considered, that is, any instrument, apparatus
appliance, material or other article of physical character that is intended to be
applied on, inserted into, implanted in or otherwise brought into contact with
the body, a body part or an organ. Furthermore, said body, body part or organ
may be that of a human or animal, typically mammal, subject. Preferably
however the medical device is intended for human subjects. Medical devices
included within the above definition are for example implants, catheters,
shunts, tubes, stents, intrauterine devices, and prostheses.
In particular, the medical device may be a medical device intended for
implantation into living tissue or for insertion into the body or a body part of a
subject, including insertion into a bodily cavity.
The present medical device may be intended for short-term, prolonged
or long-term contact with living tissue. By "short-term" is meant a duration of
less than 24 hours, in accordance with definitions found in ISO 10993-1 for
the biological evaluation of medical devices. Furthermore, "prolonged",
according to the same standard, refers to a duration of from 24 hours up to 30
days. Accordingly, by the same standard, by "long-term" is meant a duration
of more than 30 days. Thus, in some embodiments the medical device of the
invention may be a permanent implant, intended to remain for months, years,
or even life-long in the body of a subject.
As used herein the term "implant" includes within its scope any device
of which at least a part is intended to be implanted into the body of a
vertebrate animal, in particular a mammal, such as a human. Implants may be
used to replace anatomy and/or restore any function of the body. Generally,
an implant is composed of one or several implant parts. For instance, a dental
implant usually comprises a dental fixture coupled to secondary implant parts,
such as an abutment and/or a restoration tooth. However, any device, such
as a dental fixture, intended for implantation may alone be referred to as an
implant even if other parts are to be connected thereto.
By "biocompatible" is meant a material which, upon contact with living
tissue, does not as such elicit an adverse biological response (for example
inflammation or other immunological reactions) of said tissue.
By "soft tissue" is meant any tissue type, in particular mammalian
tissue types, that is not bone or cartilage. Examples of soft tissue for which
the medical device is suitable include, but are not limited to, connective
tissue, fibrous tissue, epithelial tissue, vascular tissue, muscular tissue,
mucosa, gingiva, and skin.
As used herein, the term "compound of a post-transition metal" refers
to a chemical entity comprising at least one post-transition metal and at least
one additional element. Non-limiting examples of such compounds include
oxides comprising post-transition metal, nitrides comprising post-transition
metal, alloys of at least one post-transition metal, and salts comprising posttransition
metal. The compound may comprise two or more post-transition
metals. The term "compound(s) of a post-transition metal" is intended to also
refer to compounds incorporating one or more other metals, in particular
biocompatible metals such as titanium, in addition to the one or more posttransition
metal(s). Hence, as used herein, the term "gallium compound"
refers to a chemical entity comprising gallium and at least one additional
element. Non-limiting examples of gallium compounds include gallium oxide,
gallium nitride, and gallium salts. The gallium and the at least one additional
element may be joined by covalent bonding or ionic bonding. The term
"gallium compound" is intended to also include compounds incorporating
other metals in addition to gallium, in particular titanium. Thus, galliumtitanium
oxides, gallium-titanium nitrides etc are included within the definition
"gallium compound".
By analogy, the term "bismuth compound" refers to a chemical entity
comprising bismuth and at least one additional element. Non-limiting
examples of bismuth compounds include bismuth oxide, bismuth nitride, and
bismuth salts. The bismuth and the at least one additional element may be
joined by covalent bonding or ionic bonding. The term "bismuth compound" is
intended to also include compounds incorporating other metals in addition to
bismuth, in particular titanium. Thus, bismuth-titanium oxides, bismuth -
titanium nitrides etc are included within the definition "bismuth compound".
As used herein, "metallic compound" refers to a compound which
comprises at least one metal and which has one or more properties which are
associated with metals, such as a metallic lustrous color. A metallic
compound may be formed of a metal and a non-metal, or of two or more
metals. Hence, a metallic compound need not comprise non-metallic
elements, but may be formed of metallic elements only. In the context of the
present invention, at least the following compounds are considered metallic
compounds: gallium nitride, gallium-titanium nitride, gallium-titanium, galliumbismuth-
titanium, gallium-bismuth-titanium nitride, bismuth nitride , bismuthtitanium
nitride, and bismuth-titanium.
As used herein, "homogeneous layer" refers to a layer having a
chemical composition that is uniform in all directions (three dimensions).
Figures 1 and 2 illustrate an embodiment according to the present
invention in which the medical device is a dental abutment. The dental
abutment 100 comprises a body of substrate material 102 coated with a layer
10 1 comprising a gallium compound. The layer 101 forms the surface of the
abutment intended to face and contact the gingival tissue after implantation.
The medical device of the invention may be made of any suitable
biocompatible material, e.g. materials used for implantable devices. Typically
the medical device comprises a substrate having a surface which comprises a
compound of a post-transition metal, such as a gallium compound or a
bismuth compound. The substrate may for example be made of a
biocompatible metal or metal alloy, including one or more materials selected
from the group consisting of titanium, zirconium, hafnium, vanadium, niobium,
tantalum, cobalt and iridium, and alloys thereof. Alternatively, the substrate of
the medical device may be made of a biocompatible ceramic, such as
zirconia, titania, shape memory metal ceramics and combinations thereof. In
embodiments where the medical device is used as or forms part of a dental
abutment, the substrate is preferably made of a metallic material.
In contact with oxygen, the metals titanium, zirconium, hafnium,
tantalum, niobium and their alloys instantaneously react to form an inert
oxide. Thus, the surfaces of articles of these materials are virtually always
covered with a thin oxide layer. The native oxide layer of a titanium substrate
mainly consists of titanium(IV) dioxide (T1O2) with minor amounts of T12O3,
TiO and Ti3O4.
Thus, in embodiments where the medical device comprises one or
more of titanium, zirconium, hafnium, tantalum, niobium or an alloy of any one
thereof, the medical device typically has a native metal oxide surface layer.
Such a native metal oxide layer may, in turn, be covered by a thin film
comprising the gallium compound. Alternatively, a native metal oxide layer
may be modified or replaced with a modified surface layer incorporating a
post-transition metal, such as gallium or bismuth, for example a titaniumgallium
oxide or titanium-bismuth oxide layer.
In other embodiments of the present invention, the medical device, in
particular the substrate, may be made of a biocompatible polymer, typically
selected from the group consisting of polyether ether ketone (PEEK), poly
methyl methacrylate (PMMA), poly lactic acid (PLLA) and polyglycolic acid
(PGA) and any combinations and copolymers thereof.
In embodiments of the invention, the medical device is intended for
short-term, prolonged or long-term contact with living tissue. For example, the
medical device of the invention may be an implant, typically intended to
temporarily or permanently replace or restore a function or structure of the
body.
Typically, at least part of the surface of the medical device is intended
for contact with soft tissue, and at least part of this soft tissue contact surface
has a layer comprising a compound of a post-transition metal, e.g. a gallium
compound or a bismuth compound. For example, the medical device may be
an implant intended for contact primarily or exclusively with soft tissue, for
example a dental abutment. Alternatively, the medical device may be an
implant to be inserted partially in bone and partially in soft tissue. Examples of
such implants include one-piece dental implants and bone-anchored hearing
devices (also referred to as bone anchored hearing aids). Where only part of
the implant is intended for contact with soft tissue, it is preferred that the layer
comprising the compound of a post-transition metal, e.g. gallium or bismuth
compound, is provided at least on a part of a soft tissue contact surface.
The medical device may also be suitable for contact with cartilage.
In other embodiments, the medical device may be intended for contact
with bone tissue, e.g. the jawbone, the femur or the skull of a mammal, in
particular a human. Examples of such medical devices include dental fixtures
and orthopedic implants.
In embodiments of the invention, the compound of a post-transition
metal may be a gallium compound and/or a bismuth compound. Suitable
compounds include in particular compounds that can be applied on a surface
using a thin film deposition technique.
Examples of suitable gallium compounds include gallium oxide, gallium
nitride, gallium carbide, gallium selenide, gallium sulphide, and other gallium
salts that can be deposited using a thin film deposition, for example gallium
chloride, gallium fluoride, gallium iodide, gallium oxalate, gallium phosphate,
gallium maltolate, gallium acetate and gallium lactate.
In embodiments of the invention, the gallium compound comprises at
least gallium oxide (Ga2Os). Gallium oxide may be present in amorphous or
crystalline form. Crystalline forms of gallium oxide include a-Ga 2O3, -Ga2O3,
y-Ga2O3, 5-Ga2O3, and s-Ga2O3.
Examples of suitable bismuth compounds include bismuth oxide,
bismuth nitride, bismuth carbide, bismuth selenide, bismuth sulphide, and
other bismuth salts that can be deposited using a thin film deposition, for
example bismuth chloride, bismuth fluoride, bismuth iodide, bismuth oxalate,
bismuth phosphate, bismuth maltolate, bismuth acetate and bismuth lactate.
The compound of a post-transition metal may optionally include at least
one further metal, such as titanium. Hence, in embodiments of the invention,
a gallium compound may be selected from the group consisting of gallium
oxide, gallium-titanium oxide, gallium nitride, gallium-titanium nitride, galliumtitanium,
gallium-bismuth-titanium, and gallium-bismuth-titanium nitride.
Similarly, a bismuth compound may be selected from the group consisting of
bismuth oxide, bismuth-titanium oxide, bismuth nitride , bismuth-titanium
nitride, bismuth-titanium, bismuth-gallium-titanium, bismuth-gallium-titaniumoxide,
and bismuth-gallium-titanium nitride.
Not wishing to be bound by any particular theory, it is believed that
upon contact with living tissue and/or body fluids, a layer of for example
gallium oxide or gallium nitride (optionally comprising an additional metal such
as titanium) exhibits slow, sustained release of gallium ions. Such release
may be slower and more sustained compared to the release of gallium ions
from a precipitated gallium salt, and may thus provide a more long-term effect
with respect to biofilm formation. In addition, a surface layer deposited using a
thin film deposition method as used in embodiments of the invention firmly
adheres to the underlying substrate and thus avoids problem related to
peeling and flaking of the surface layer. Peeling and flaking may give rise to
adverse inflammatory response of the surrounding tissue, and in addition may
undermine the biofilm prevention effect of the surface layer.
Depending on the intended use of the medical device, different release
properties may be desirable. For example, a higher release rate of
antimicrobial post-transistion metal may be more favorable for short term use,
i.e. for a medical device intended for short-term contact with living tissue,
compared to a device intended for prolonged or long-term contact. The
release rate may be affected by various factors, for example the crystallinity
of the compound.
Optionally, in embodiments of the invention the medical device may
additionally comprise a salt of a post-transition metal, e.g. a gallium salt
selected from the group consisting of gallium acetate, gallium carbonate,
gallium chloride, gallium citrate, gallium fluoride, gallium formate, gallium
iodide, gallium lactate, gallium maltolate, gallium nitrate, gallium oxalate,
gallium phosphate, and gallium sulphate. Alternatively the salt may be a
bismuth salt of formed of any of these counterions. Such a salt may be
provided as a deposit, e.g. precipitated, on the layer comprising the
compound of a post-transition metal.
As mentioned above, the compound of a post-transition metal, e.g.
gallium compound or bismuth compound, is typically contained in an applied
surface layer. In embodiments of the invention, the compound, optionally
including a further metal such as titanium, may constitute the major part of
said layer. The atomic concentration (at%) of the elements together forming
the compound of a post-transition metal constitute at least 50 at% of the
layer, preferably at least 70 at% and more preferably at least 80 at% of the
elements of the layer.
The atomic concentration of post-transition metal, e.g. gallium, in the
layer may be in the range of from 5 at% up to 50 at%, for example at least
10 at%, at least 15 at%, at least 20 at%, at least 35 at% or at least 30 at%,
and up to for example 50 at%, such as up to 45 at%, or up to 40 at%.
However in embodiments of the invention, the atomic content of the posttransition
metal may be less than 5 at%. For example, the atomic content of
the post-transition metal may be in the range of from 0.01 to 20 at%, e.g. from
0.05 to 15 at%, such as from 0.1 to 15 at%. For example, in embodiments
where the compound of post-transition metal comprises gallium, the content
of gallium in the layer may be at least 0.05 at% or at least 0.1 at%, for
example at least 0.3 at% or at least 0.5 at%. In embodiments where the
compound of post-transition metal comprises bismuth, the content of bismuth
in the layer may be for instance at least 0.1 at%, at least 0.2 at%, for example
at least 1 at% or at least 1.5 at%.
When the gallium compound is substantially pure gallium oxide
(Ga2Os), the maximum content of gallium in the layer is 40 at%, and the
maximum content of oxygen in the layer is 60 at%. However, impurities and
contamination, for example carbon, may be present at up to 20 at%. If the
gallium compound is gallium-titanium oxide, part of the gallium is replaced
with titanium and so the total content of gallium and titanium would be 40 at%.
Where the gallium compound is substantially pure gallium nitride
(GaN), the maximum content of gallium in the layer is 50 at%, and the
maximum content of nitrogen is 50 at%. Contaminations may be present as
described above. If part of the gallium is replaced with another metal, such as
titanium, the total content of gallium and said other metal may still be 50 at%,
or it may be higher, e.g. up to 75 %, if the nitrogen content is low. As an
example, the layer may contain 50 at% nitrogen, 25 at% titanium and 25 at%
gallium.
The atomic concentration may be measured for example to a depth of
40 nm or less, and preferably not more than the layer thickness. The atomic
concentration can be measured using X-ray photoelectron spectroscopy
(XPS).
As mentioned above, upon contact with living tissue, some posttransition
metal, such as gallium or bismuth, may be released from the
surface of the medical device over time. Hence after implantation the content
of post-transition metal and possibly also of other materials present on the
surface of the medical device may change over time.
In some embodiments, the layer may in addition to the compound of a
post-transition metal include one or more other elements or compounds (a
dopant), for example at a content of 10 at% or less, as will be described in
more detail below. The layer comprising the gallium compound or other
compound of a post-transition metal may also contain impurities or
contamination, for example carbon, typically in an amount of 20 at% or less,
and preferably 15 at% or less, or 10 at% or less. Such contamination may
originate from the packaging. It may be noted that wet packaging, in which
the surface may be protected by water, ethanol or the like, reduces the
amount of contamination by carbon, compared to dry packaging where the
surface is exposed to air which normally contains volatile hydrocarbons.
Contamination may also be present on the surface of the substrate before the
layer comprising the gallium compound is applied. The level of contamination,
typically represented by the atomic concentration of carbon, may be reduced
by cleaning the surface before applying the compound of post-transition
metal, and optionally after applying the compound of post-transition metal
and/or by avoiding further contaminating the surface before measuring the
atomic concentration of elements on the surface.
Table 1 summarizes possible atomic concentration ranges for a layer
comprising bismuth nitride, bismuth-titanium nitride, gallium oxide, galliumtitanium
oxide, gallium nitride or gallium-titanium nitride, respectively.
Table 1. Exemplary atomic concentrations of various elements of a gallium
compound.
In embodiments of the invention, the compound of a post-transition
metal may primarily comprise a post-transition metal and an additional metal,
such as titanium. For example, the surface layer may comprise titaniumgallium
or titanium-bismuth as the compound of a post-transition metal. In
such embodiments, the content of post-transition metal may be in the range
of from 0.01 to 20 at%, e.g. from 0.05 to 15 at%, such as from 0.1 to 15 at%,
as described above. Further, the titanium content may be in the range of from
10 to 99.9 at%, e.g. 10 to 60 at%, or 10 to 30 at%. The layer may additionally
contain nitrogen at a content of from 0 to 15 at%. For example, the nitrogen
content of the layer may typically be in the range of from 0 to 5 at%.
Furthermore, depending on the handling (e.g. cleaning steps) and/or
analytical technique used for determining the surface chemical composition,
the layer may additionally have a relatively high but superficial oxygen
content, e.g. from 0 up to about 60 at%.
In some embodiments, the surface layer consists essentially of one or
more of the compound(s) of a post-transition metal mentioned above. In
accordance with the above, "consists essentially of " here means that the
layer contains little or no other material (dopants, contaminants, etc) except
the one or more compound(s) of a post-transition metal, only for example up
to 10 at%, preferably up to 5 at%, more preferably up to 2 at% and even more
preferably up to 1 at% of other material.
In general, the layer comprising the compound of a post-transition
metal, e.g. a gallium compound or bismuth compound, is free of carrier
material such as polymers, solvents, etc.
The layer comprising the compound of a post-transition metal such as
a gallium compound or a bismuth compound may have a thickness in the
range of from 1 nm to 1.5 mhti , for example 0.1 to 1 miti , in particular from 0.3
to 1 miti . A layer having a thickness of at least 1 nm may provide sufficient
antimicrobial effect. Increasing layer thickness may provide a whiter color,
which may be desirable for dental applications. However, also a layer having
a thickness of from about 10 nm may be more aesthetically advantageous
than present commercial dental abutments. For example, a gallium oxide
layer of 40 nm has a deep bronze color which would be less visible through a
patient's gingiva than current grey-metallic titanium abutments.
Where mainly an antimicrobial effect is sought, the layer containing the
compound of a post-transition metal may optionally have a thickness of from
10 to 100 nm, or optionally up to 300 nm. On the other hand, where the
aesthetic appearance of e.g. a dental abutment is of high importance, a layer
thickness in the range of from 0.5 to 1.5 mhti , e.g. from 0.7 to 1.5 mhh or from
0.7 to 1 m ΐ may be preferred. However also thinner layers may provide an
acceptable color appearance and which at least may be more advantageous
than prior art dental abutments.
The layer may be a dense layer, i.e. a non-porous layer.
In embodiments of the invention, the surface of the medical device may
comprise a single layer. Alternatively, in other embodiments, the medical
device may comprise multiple layers, at least one comprising a compound of
a post-transition metal such as a gallium compound or a bismuth compound.
In embodiments of the invention, a salt of non-toxic post-transition
metal, optionally forming a further layer, may be provided on at least a portion
of a thin-film deposited layer comprising the above-mentioned compound of
non-toxic post-transition metal. For example, a solution of at least one gallium
salt may be applied onto a thin-film deposited layer of a compound of posttransition
metal, and allowed to evaporate. Such embodiments may provide a
high initial release of gallium upon contact with living tissue, which may be
advantageous in many instances, for short-term, prolonged as well as for
long-term tissue contact.
In embodiments of the invention, a surface layer of the medical device
may contain at least one additional element(s) or compound(s), for example a
bioactive element or compound that may further enhance tissue healing or
function. Such elements or compounds may be included as a dopant in the
layer comprising the compound of a post-transition metal, typically at a
content of 10 at% or less. Alternatively such element(s) or compound(s) may
be applied as a separate layer, e.g. on the layer comprising the compound of
a post-transition metal.
In embodiments of the invention, the substrate may have a rough
surface on which a layer comprising the compound of a post-transition metal
is arranged. Since the layer comprising the compound of a post-transition
metal may be thin, e.g. 100 nm or less, it may have good conformal step
coverage, meaning that the layer follows the underlying surface roughness
and substantially preserves it, without making it smoother. However, in
embodiments where the layer comprising the compound of a post-transition
metal is relatively thick, it may reduce the roughness of the underlying
substrate surface.
The substrate surface roughness, and hence optionally also the
surface of the medical device formed by the layer comprising the gallium
compound of a post-transition metal, may have an average surface
roughness Ra of at least 0.05 mhti , typically at least 0.1 mhti , for example at
least 0.2 mhh . Since surfaces having an average surface roughness (Ra) of at
least 0.2 mhh are believed to be more susceptible of biofilm formation, a layer
comprising a compound of a post-transition metal, e.g. a gallium compound or
a bismuth compounds as described herein may be particularly advantageous
for medical devices having a surface roughness of at least 0.2 mhti , and may
be increasingly useful for preventing biofilm formation on medical devices
having even higher surface roughness. As an example, a dental abutment
comprising a titanium substrate may have a surface roughness of about 0.2-
0.3 m ΐ . A surface layer of e.g. a gallium compound having a thickness of
about 40 nm may substantially preserve this surface roughness (which may
be desirable e.g. in order to facilitate a firm anchorage of the implant in the
surrounding tissue) but may prevent biofilm formation on the implant surface
and hence reduce the risk for infection and periimplantitis.
The layer comprising a compound of a post-transition metal may be
formed by applying the compound of a post-transition metal onto the surface
of a medical device, to form a surface layer. The compound of a posttransition
metal may be applied using known deposition techniques,
especially thin film deposition techniques. Suitable techniques may include
physical deposition, chemical deposition and physical-chemical deposition.
One example of such techniques is atomic layer deposition (ALD) which can
be used to provide e.g. a gallium oxide layer on a substrate surface
(Nieminen et al, 1996; Shan et al, 2005). Other techniques that may be used
for depositionare chemical vapor deposition (Pecharroman et al, 2003; Kim et
al, 201 0), spray pyrolysis deposition (Kim & Kim, 1987; Hao et al, 2004), arc
discharge deposition (Heikman et al, 2006), electron beam deposition (Al-
Kuhaili et al, 2003), pulsed laser deposition (Orita et al, 2002), thermal
evaporation deposition (Karaagac & Parlak, 201 1) , sputter deposition (Kim &
Holloway, 2004) and molecular beam deposition (Passlack et al, 1995).
Physical deposition, chemical deposition, and physical-chemical deposition
techniques may also be used for incorporating additional element(s) into a
layer of compound(s) of post-transition metal, e.g. as dopants, in order to
enhance healing or regeneration of the tissue contacting the medical device.
ALD and other thin film deposition techniques are associated with
several advantages for the deposition of the compound(s) of a post-transition
metal, such as controlled layer thickness, controlled composition, high purity,
conformal step coverage, good uniformity (resulting in a homogeneous layer),
and good adhesion. In particular, PVD may provide dense layers with
excellent adhesion to the underlying substrate.
Examples
Example 1A. Production of gallium oxide coated specimens
Coins of commercially pure (cp) titanium (grade 4) were manufactured
and cleaned before deposition of a 40 nm thick layer of amorphous Ga2O3
using atomic layer deposition (Picosun, Finland) with precursors of GaCb and
H2O, respectively. Specimens were thereafter packaged in plastic containers,
and sterilized with electron beam irradiation.
Example 1B. Surface characterization of gallium oxide coated
specimens
For all surface characterization experiments, eight specimens each of
commercially pure (cp) titanium, Ga2O3 coated cp titanium produced as
described above, and commercially available TiN coated cp titanium, were
prepared as described in Example 1 (cleaned, coating using ALD in the case
of the Ga2O3 coated specimens, packaged, and sterilized). The TiN coated
specimens were included for comparison since it is known that a TiN coating
provides a weakly antibacterial effect.
It was found that the surface morphology and surface roughness was
unaltered by the ALD coating, but there was a slight increase of hydrophobic
properties.
a) Surface chemistry
Surface morphology and surface chemistry was analyzed with
environmental scanning electron microscopy (XL30 ESEM, Philips,
Netherlands)/energy dispersive spectroscopy (Genesis System, EDAX Inc.,
USA) at an acceleration voltage at 10-30 kV. Elements detected on the
surface of the Ga2O3 coated specimens were oxygen (O), gallium (Ga), and
titanium (Ti). Ga concentrations varied between 4 to 9 atomic % (at%), as
measured with acceleration voltages at 30 kV and 10 kV, respectively. The
analytical depth with this technique is estimated to be approximately 1 mhti , i.e.
much deeper than the layer thickness. No differences in terms of surface
morphology could be detected between commercially pure titanium controls
and the Ga2O3 coated titanium.
Additionally, surface chemistry was analyzed with X-ray photoelectron
spectroscopy (XPS, Physical Electronics, USA), which is a more surface
sensitive technique than energy dispersive spectroscopy. As X-ray source
monochromatic AIKa was used. The beam was focused to 100 mhh . Elements
detected were oxygen (O), gallium (Ga), and carbon (C). Gallium
concentrations varied between 34 and 37 at%. Oxygen concentrations varied
between 47 and 50 at%. The analytical depth with this technique is estimated
to be approximately 5-1 0 nm. The results are summarized in Table 2 .
Table 2. Atomic concentration of detected elements using XPS
Element At% detected using XPS (depth 5-10 nm)
Ga 34-37 at%
O 47-50 at%
C 13-1 8 at%
b) Surface morphology
Surface roughness was measured with surface profilometry (Hommel
T 1000 wave, Hommelwerke GmbH, Germany). A vertical measuring range of
320 mhti , and an assessment length of 4.8 mm were used. Two specimens of
each type were included in the analysis, and three measurements per
specimen were performed. The surface roughness Ra was calculated after
using a filtering process, with cut-off at 0.800 mm. The results are presented
in Table 3.
Table 3. Surface roughness (Ra) ± standard deviations (SD)
c) Wettability
In order to investigate the wettability, the contact angle was measured
using a contact angle measuring system (Drop Shape Analysis System DSA
100, Kruss GmbH, Germany). Measurements were performed with deionized
water. The results indicate that all specimens were hydrophobic (>90°), see
Table 4 .
Table 4. Contact angles (°) ± standard deviations (SD)
Example 1C. Antimicrobial effect of gallium oxide-coated surfaces
It was found that a titanium body having a surface comprising gallium
(Ga) in the form gallium oxide can prevent the growth of Pseudomonas
aeruginosa and Staphylococcus aureus on and around a surface and thus
may be useful in preventing detrimental infection around e.g. a dental
abutment implanted into the gingiva.
a) Inhibition of bacterial growth on streak plate
In a first experiment commercially pure titanium coins (0 6,25 mm)
with or without a gallium oxide coating were placed on agar plates containing
homogeneously distributed colonies of Pseudomonas aeruginosa. After
incubation for 24 hours at 37°C there was a 4 mm wide visible colony free
zone surrounding the gallium oxide coins, in contrast to the titanium coins that
were surrounded by bacterial colonies.
b) Inhibition of bacterial growth using film contact method
In a second experiment, a film contact method (Yasuyuki et al, 201 0)
was used. Streak plates of Pseudomonas aeruginosa (PA01 ) or methillicin
resistant Staphylococcus aureus (MRSA) were made and 1 colony was
inoculated to 5 ml tryptic soy broth (TSB) in culture tubes and grown under
shaking conditions for 18 hours. Cell density was measured in a
spectrophotometer at OD 600 nm and counted using a cell counting chamber.
The cell culture was adjusted with sterile TSB to 1-5 X 106 cells/ml.
Specimens of commercially pure (cp) titanium coins (0 6.25 mm), cp titanium
coins with a gallium oxide coating, or cp titanium coins with a commercially
available titanium nitride (TiN) coating were aseptically prepared and put in
respective well of a 12 well plate. Thin transparent plastic film was punched,
and sterilized using 70 % ethanol and UV irradiation on each side. A 15m I
drop of bacteria in TSB was applied on each specimen. One thin plastic film
per specimen was placed over the bacteria on the specimens so that the
bacterial solution was evenly spread over the specimen surface, ensuring
good contact. After incubation for 24 hours at 30±1 °C, the film of each
specimen was aseptically removed and washed by pipetting 1ml PBS over
the surface into a separate 2 ml eppendorf tube per specimen. The
specimens were transferred to the same eppendorf tubes as used when
washing the film. First each specimen surface was washed by pipetting the
very same PBS as the film was previously washed with. Next, the specimens
were sonicated and for 1 minute and vigorously vortexed for 1 minute in the
very same tube as previously used when washing the film. Serial dilutions
and plate count were performed. Plates were incubated for 24 hours and
colony numbers counted and recorded. The antibacterial activity of gallium
oxide coated titanium was determined to 92 % reduction against PA01 and
7 1 % reduction against MRSA, compared to titanium, see Tables 5 and 6 .
Table 5. Viable counts (cfu/ml) ± standard deviations (SD) after 24 hours
incubation of test specimens against Pseudomonas Aeruginosa (PA01).
c) In situ effect on a biofilm
In a third experiment, the antibacterial activity of titanium discs, with or
without a gallium oxide coating, was evaluated in situ using Live/Dead®
BacLight™ stain (Life Technologies Ltd, UK). Streak plates of Pseudomonas
aeruginosa (PA01) were made and 1 colony was inoculated to 5 ml TSB in
culture tubes and grown under shaking conditions for 18 hours. Cell density
was measured in a spectrophotometer at OD 600 nm and adjusted with
sterile TSB to 1 x 106 cells/ml. 400 m I bacteria were aliquoted into 8-
chambered slides. The biofilm was allowed to be formed during 24 hours at
35±2°C. Specimens of commercially pure (cp) titanium coins (0 6.25 mm), cp
titanium coins with a gallium oxide coating, or cp titanium coins with a titanium
nitride (TiN) coating were aseptically prepared and applied onto the biofilm.
The antibacterial activity was analyzed in situ using the Live/Dead® stain.
In situ analyses indicated that both titanium nitride and gallium oxide
coatings have an anti-biofilm activity compared with uncoated titanium in
terms of viability. At 24 hour analysis, it was visualized that the typical
mushroom structure of biofilms had disappeared for titanium nitride and
gallium oxide. It was also found that more dead cells were seen on gallium
oxide than on titanium nitride.
Example 2A. Production of nitride or metal coated specimens
Coins of commercially pure (cp) titanium (grade 4) were manufactured
and cleaned before deposition of a 0.5-1 miti thick layer of either titanium
bismuth (TiBi), titanium nitride with bismuth (TiNBi), titanium gallium (TiGa) or
titanium nitride with gallium (TiNGa) using physical vapor deposition (PVD).
Targets of TiBi and TiGa were used for TiBi/TiNBi and TiGa/TiNGa,
respectively. The specimens were thereafter packaged in plastic containers,
and sterilized with electron beam irradiation.
Example 2B. Surface characterization of nitride or metal coated
specimens
For these surface characterization experiments, specimens of
commercially pure (cp) titanium, TiBi coated titanium, TiNBi coated titanium,
TiGa coated titanium and TiNGa coated titanium produced as described in
Example 2A were used.
It was found that the surface morphology and surface roughness was
unaltered by the PVD coating, and that both uncoated titanium and the coated
specimens had hydrophilic properties.
The nitride specimens (TiNGa, TiNBi) had a goldish color, similar to the
color of TiN surfaces on commercially available dental implants. It is expected
that a BiN or a GaN surface layer would have a goldish appearance as well.
In contrast, the TiBi and TiGa specimens had a grey-metallic color, similar to
that of the uncoated titanium specimens.
a) Surface morphology and surface chemistry
Surface morphology and surface chemistry were analyzed with
environmental scanning electron microscopy (XL30 ESEM, Philips,
Netherlands)/energy dispersive spectroscopy (Genesis System, EDAX Inc.,
USA) at an acceleration voltage at 10 kV. Concentrations of bismuth (Bi) and
gallium (Ga) on the coated specimens (TiBi, TiNBi, TiGa, TiNGa) were found
to be up to 12.6 at% for Bi and 1.1 at% for Ga, as measured with acceleration
voltages at 10 kV. Other elements detected on the surfaces were titanium
(Ti), nitrogen (N), oxygen (O), and carbon (C). The analytical depth with this
technique is estimated to be approximately 1 mhti , i.e. possibly deeper than
the layer thickness. No differences in terms of surface morphology could be
detected between commercially pure titanium controls and the coated
titanium (TiBi, TiNBi, TiGa, TiNGa).
Additionally, surface chemistry was analyzed with X-ray photoelectron
spectroscopy (XPS, Physical Electronics, USA), which is a more surface
sensitive technique than energy dispersive spectroscopy. As X-ray source
monochromatic AIKa was used. The beam was focused to 100 mhh . Elements
detected were titanium (Ti), nitrogen (N), carbon (C), and oxygen (O), as well
as bismuth (Bi) (for TiBi and TiNBi specimens) and gallium (Ga) (for TiGa and
TiNGa specimens). Bi and Ga concentrations were detected to be up to
3.3 at% and 0.4 at%, respectively. The analytical depth with this technique is
estimated to be approximately 5-1 0 nm. The results are summarized in
Table 7 .
Table 7. Summary of surface chemistry analyses of titanium controls and
coated surfaces (TiBi, TiNBi, TiGa, TiNGa). Values (at%) describe the ranges
of elementary concentrations (n=6). * The titanium signal (TiL) and the
nitrogen signal (NK) are closely located, which contributes to the variance in
nitrogen (N) concentration.
b) Surface roughness
The surface roughness of specimens produced according to Example
2A was measured with surface profilometry (Hommel T 000 wave,
Hommelwerke GmbH, Germany). A vertical measuring range of 320 mhh and
an assessment length of 4.8 mm were used. Three specimens of each type
were included in the analysis, and three measurements per specimen were
performed. The surface roughness in terms of arithmetic mean value of
vertical deviations of the roughness profile from a mean line (Ra) was
calculated after using a filtering process, with cut-off at 0.800 mm. The results
are presented in Table 8 .
Table 8. Surface roughness (Ra) ± standard deviations (SD), n=9.
c) Wettability
In order to investigate the wettability of specimens produced according
to Example 2A, the contact angle was measured using a contact angle
measuring system (Drop Shape Analysis System DSA 100, Kruss GmbH,
Germany). Measurements were performed with deionized water. The result
showed that all specimens were hydrophilic (<90°) although variations
between test specimens were observed, see Table 9 .
Table 9. Contact angles (°) ± standard deviations (SD)
Example 2C. Release of gallium and bismuth from surface coatings
Release of gallium (Ga) or bismuth (Bi) from all surface coatings (TiBi,
TiNBi, TiGa, TiNGa) was confirmed in experiments. At a pH of 7.4 low
amounts of Ga and Bi, respectively, were released, but at pH 5.0 the release
of Bi increased significantly.
For all release experiments, specimens of commercially pure (cp)
titanium, TiBi coated titanium, TiNBi coated titanium, TiGa coated titanium
and TiNGa coated titanium produced as described in Example 2A were used.
The release experiments were performed by immersing specimens in either
phosphate buffer (pH 7.4) or acetate buffer (pH 5.0). The pH of 7.4 was
intended to simulate a physiologic neutral condition (pH 7.4). The acidic pH
(5.0) was intended to simulate a condition that may occur locally in the tissue
the case of inflammation and/or infection. Glass bottles containing 10 ml
buffer were used with one specimen per bottle. The bottles were placed on a
shanking table at 37°C during the experiment. After 1, 4, 7, and 24 days liquid
samples were taken and the Bi and Ga concentrations were analyzed using
Inductively Coupled Plasma- Sector Field Mass Spectrometry (ICP-SFMS).
The results are presented in Table 10 .
Table 10. Release of bismuth (Bi) or gallium (Ga) to acetate buffer (pH 5.5) or
phosphate buffer (pH 7.4). Figures are mean values (n=2), standard
deviations within parentheses.
Time Release of Bi ( g/ l) Release of Ga (pg/l)
(days) TiBi coated Ti TiNBi coated TiGa coated TiNGa coated
Ti Ti Ti
pH 5.0 pH 7.4 pH 5.0 pH 7.4 pH 5.0 pH 7.4 pH 5.0 pH 7.4
1 2840 16.2 1150 3.6 0.9 1.3 8.4 11.9
(240) (0.1 ) (28) (0.2) (0.1 ) (0.1 ) ( 1 .4) ( 1 .2)
4 3225 25.6 1660 3.9 0.7 1.3 12.7 12.4
(21 ) (7.9) (806) ( 1 .2) (0.1 ) (0.2) (0.4) ( 1 .7)
7 2700 22.9 1470 2.6 0.7 1.6 13.3 12.4
(141 ) ( 1 .6) (438) (0.3) (0.1 ) (0.1 ) (2.1 ) (0.4)
24 - 10.3 - 8.3 2.1 14.7
(0.1 ) (2.0) (0.3) (0.9)
Example 2D. Antimicrobial effect of nitride or metal coated specimens
It was found that a titanium body having a surface comprising gallium
(Ga) or bismuth (Bi) in the form TiGa, TiNGa, TiBi or TiNBi can prevent the
growth of Pseudomonas aeruginosa and Staphylococcus aureus on a surface
and thus may be useful in preventing detrimental infection around e.g. a
dental abutment implanted into the gingiva.
a) Antimicrobial effect of bismuth (Bi) and gallium (Ga) against
different bacteria species
Streak plates and characterization by gram-staining were done. MRSA
and PA01 were grown on Tryptic Soy agar (TSA) plates and S. sanguinis on
horse blood plates for up to 24 hours whereas P. gingivalis was grown
between 4 and 5 days on Fastidious Anaerobic agar (FAA) plates. Fivefold
dilution in at least five steps of gallium nitrate (Ga(NOs)3) or bismuth chloride
(B1CI3) was done. Bismuth salt had to be suspended in DMSO in order to
enable salt to dissolve in culture media.
Equal amounts of bacterial solution and antimicrobial salt (in fivefold
dilution) were mixed and incubated at 35±2°C: MRSA, PA01 , and S.
sanguinis for 24 hours and P. gingivalis for 4 or 5 days. After incubation, the
concentration where no visual growth could be detected was documented.
Results are summarized in Table 11.
Table 11. Minimum Inhibitory Concentration (MIC) values in [jig/ml.
* The MIC values differed at the three different runs, hence MIC values from
all runs are included.
The MIC values indicate that both Ga and Bi exert an antibacterial
effect against various bacterial species, although the effect varies for different
species. Variation in MIC values for B1CI3 against MRSA and PA01 was
probably caused by inadequate dissolution of the B1CI3 (high values indicating
that the salt was not completely dissolved and was thus unavailable for
inhibition).
b) Inhibition of bacterial growth using film contact method
In another experiment, a film contact method (Yasuyuki et al, 201 0)
was used. Streak plates of Pseudomonas aeruginosa (PA01 ) were made and
1 colony was inoculated to 5 ml tryptic soy broth (TSB) in culture tubes and
grown under shaking conditions for 18 hours. Cell density was measured in a
spectrophotometer at OD 600 nm and counted using a cell counting chamber.
The cell culture was adjusted with sterile TSB to 1-5 X 106 cells/ml.
Specimens of commercially pure (cp) titanium coins (0 6.25 mm) and cp
titanium coins with coatings of TiBi, TiNBi, TiGa or TiNGa were aseptically
prepared and put in respective well of a 12 well plate. Thin transparent plastic
film was punched, and sterilized using 70 % ethanol and UV irradiation on
each side. A 15 m I drop of bacteria in TSB was applied on each specimen.
One thin plastic film per specimen was placed over the bacteria on the
specimens so that the bacterial solution was evenly spread over the
specimen surface, ensuring good contact. After incubation for 24 hours at
30±1 °C, the film of each specimen was aseptically removed and washed by
pipetting 1ml PBS over the surface into a separate 2 ml eppendorf tube per
specimen. The specimens were transferred to the same eppendorf tubes as
used when washing the film. First each specimen surface was washed by
pipetting the very same PBS as the film was previously washed with. Next,
the specimens were sonicated and for 1 minute and vigorously vortexed for 1
minute in the very same tube as previously used when washing the film.
Serial dilutions and plate count were performed. Plates were incubated for 24
hours and colony numbers counted and recorded. The antibacterial activity of
TiBi, TiNBi, TiGa, or TiNGa coated titanium specimens was determined to
>99,5% reduction against PA01 compared to titanium, see Table 12 .
Table 12. Antibacterial activity was calculated as percent reduction
(compared to uncoated titanium) after 24 hours incubation of test specimens
against Pseudomonas Aeruginosa (PA01).
Example 2E. Cytotoxicity of nitride or metal coated specimens
All surface coatings (TiBi, TiNBi, TiGa, TiNGa) were found to enhance
fibroblast mitochondrial activity, i.e. cell viability, compared to uncoated
titanium surfaces, using a cytotoxicity test.
Cytotoxicity was assessed with a (3-(4,5-dimethylthiazol-2-yl)-5-(3-
carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay using
human fibroblasts (MRC-5). An MTS assay measures the mitochondrial
activity in cells and correlates to the viability / the number of cells. For
cytotoxicity experiments, specimens of commercially pure (cp) titanium, TiBi
coated titanium, TiNBi coated titanium, TiGa coated titanium and TiNGa
coated titanium produced as described in Example 2A were used.
The cells were subcultured according to guidelines from the American
Type Culture Collection. They had reached approximately 80 % confluence
when they were used in the experiment. Six coins of each specimen were
aseptically placer in a 24 well plate, 1 coin per well. 100 m I of MRC-5 cells at
1x1 05 cells/ml were added to each disc and controls. The plates were then
incubated for one hour to allow cell attachment before 1 ml complete cell
culture media was added to each well. The plates were then incubated for
24 hours before analysis. For the MTS assay 4 coins of each specimen were
transferred from the 24 well plates to 48 well plates. 500 m I MTS assay
reagent was added to each well and were incubated for 4 hours. Absorbance
was then read at 490 nm. The absorbance values for each specimen were
normalized to the uncoated titanium control in percentage set to 100 % (Table
13).
The remaining 2 coins of each specimen were used for DAPI staining
and fluorescence microscopy, in order to verify the MTS assay result. DAPI is
a fluorescent stain that binds to cell nucleus and can thus be used to visualize
cells on metals.
Table 13. Mitochondrial activity (i.e. viability) as measured with an MTS assay
for human fibroblasts cultured on uncoated and coated titanium specimens.
Mean values, standard deviations within parentheses.
In conclusion, it has been experimentally demonstrated that a surface
layer comprising a compound of a post-transition metal, here a compound of
gallium, bismuth, or both, can be applied on a biocompatible metal substrate
(titanium) to provide an antimicrobial effect against several bacterial species.
It has also been shown that there is a release of post-transition metal from the
surface layer at physiologically neutral conditions. Furthermore, at acidic
conditions, which may occur very locally in the tissue in the case of
inflammation/infection, the release of post-transition metal may even be
significantly increased. Finally, the tested surface layers are non-toxic to
human fibroblast cells and may in fact enhance fibroblast mitochondrial
activity.
The person skilled in the art realizes that the present invention by no
means is limited to the preferred embodiments described above. On the
contrary, many modifications and variations are possible within the scope of
the appended claims.
Additionally, variations to the disclosed embodiments can be
understood and effected by the skilled person in practicing the claimed
invention, from a study of the drawings, the disclosure, and the appended
claims. In the claims, the word "comprising" does not exclude other elements
or steps, and the indefinite article "a" or "an" does not exclude a plurality. The
mere fact that certain measures are recited in mutually different dependent
claims does not indicate that a combination of these measured cannot be
used to advantage.
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CLAIMS
1. A medical device intended for contact with living tissue, comprising a
substrate having a surface layer comprising one or more compound(s) of a
non-toxic post-transition metal.
2 . The medical device according to claim 1, wherein said living tissue is
soft tissue.
3 . The medical device according to claim 1 or 2, wherein said
compound of a non-toxic post-transition metal also comprises an additional
metal, preferably titanium.
4 . The medical device according to any one of the preceding claims,
wherein said surface layer is a metallic layer and said compound of a no n
toxic post-transition metal is a metallic compound.
5 . The medical device according to any one of the preceding claims,
wherein said non-toxic post-transition metal is selected from bismuth and
gallium.
6 . The medical device according to any one of the preceding claims,
wherein said compound of a non-toxic post-transition metal is selected from
the group consisting of gallium-titanium, gallium-titanium oxide, galliumtitanium
nitride, gallium nitride, gallium carbide, gallium selenide, and gallium
sulphide, gallium chloride, gallium fluoride, gallium iodide, gallium oxalate,
gallium phosphate, gallium maltolate, gallium acetate and gallium lactate.
7 . The medical device according to any one of the preceding claims,
wherein said compound of a non-toxic post-transition metal is selected from
the group consisting of bismuth-titanium, bismuth-titanium oxide, bismuthtitanium
nitride, bismuth nitride, bismuth carbide, bismuth selenide, and
bismuth sulphide, bismuth chloride, bismuth fluoride, bismuth iodide, bismuth
oxalate, bismuth phosphate, bismuth maltolate, bismuth acetate and bismuth
lactate.
8 . The medical device according to any one of the preceding claims,
wherein said compound of a non-toxic post-transition metal is a nitride.
9 . The medical device according to claim 8, wherein said nitride is
selected from the group consisting of gallium-titanium nitride, gallium nitride,
bismuth-titanium nitride, bismuth nitride, and gallium-bismuth-titanium nitride.
10 . The medical device according to any one of the preceding claims,
wherein said compound of a non-toxic post-transition metal constitutes the
major part of said layer.
11. The medical device according to any one of the preceding claims,
further comprising a salt of a non-toxic post-transition metal deposited onto
said surface layer.
12 . The medical device according to any one of the preceding claims,
wherein said surface layer has a thickness in the range of from 10 nm to
1.5 mhh .
13 . The medical device according to any one of the preceding claims,
wherein said layer is a homogeneous layer.
14. The medical device according to any one of the preceding claims,
wherein said layer is a non-porous layer.
15 . The medical device according to any one of the preceding claims,
wherein said substrate comprises a metallic material, preferably titanium or
titanium alloy.
16 . The medical device according to any one of the claims 1 to 14,
wherein said substrate comprises a ceramic material.
17 . The medical device according to any one of the claims 1 to 14,
wherein said substrate comprises a polymeric material.
18 . The medical device according to any one of the claims 1 to 14,
wherein said substrate comprises a composite material.
19 . The medical device according to any one of the preceding claims,
which is an implant intended for long-term contact with living tissue.
20. The medical device according to any one of the claims 1 to 18,
which is intended for prolonged contact with living tissue.
2 1. The medical device according to any one of the claims 1 to 18,
which is intended for short-term contact with living tissue.
22. The medical device according to any one of the claims 1 to 20,
which is a dental implant.
23. The medical device according to claim 22, wherein said dental
implant is a dental abutment.
24. The medical device according to any one of the claims 1 to 20,
which is a bone anchored hearing device.
25. The medical device according to any one of the claims 1 to 2 1,
which is a stent.
26. The medical device according to any one of the claims 1 to 2 1,
which is a shunt.
27. The medical device according to any one of the claims 1 to 2 1,
which is an orthopaedic implant.
28. The medical device according to claim 20 or 2 1, which is a catheter
for insertion into a bodily cavity.
29. A method of producing a medical device according to any one of
the claims 1 to 28 comprising
a) providing a substrate having a surface; and
b) applying a compound of a non-toxic post-transition metal onto said
surface to form a layer.
30 The method according to claim 29, wherein said compound of a
non-toxic post-transition metal is a nitride of a non-toxic post-transition metal,
preferably a nitride of gallium and/or bismuth.
3 1. The method according to claim 29, wherein step b) involves
applying a compound of a non-toxic post-transition metal and an additional
metal or metal compound onto said surface.
32. The method according to claim 3 1, wherein said additional metal or
metal compound comprises titanium.
33. The method according to any one of the claims 29 to 32, wherein
step b) is performed using a thin film deposition technique.