Required Biocompatibility Training and Required Biocompatibility
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Plasma for Biomedical
Applications
Presented by
Prof. Sudarsan Neogi
Department of Chemical Engineering
Indian Institute of Technology
Kharagpur
Presentation outline
• Plasma – introduction
• Salient features of plasma
• Biomedical applications of polymers
• Plasma surface modification of polymers
• Specific biomedical applications of polymers using plasma
• Plasma sterilization
• Latest development
• Summary
Solid
Energy
Liquid
Energy
Gas
Energy
Plasma
Plasma• Plasma – a quasi neutral gas, referred to as fourth state of
matter
• Collection of particles consisting of electrons, ions and excited
atoms and molecules
• Moves in random directions
• Electrically neutral
• Ionization of neutrals sustains the plasma in the steady state
• Natural Plasma – Lightning, all stars including sun
Advantages of plasma treatment
• Alters the surface properties of material without affecting their
bulk property
• Surface modification in a controlled fashion
• Highly cross-linked films irrespective of the surface
geometries
• Formation of multilayer films
• Eco-friendly nature
• The prospect of scaling up
• No water and chemicals required
• Selection of desired chemical pathways
• Minimization of thermal degradation and rapid treatment
Applications of Plasma
• Microelectronics
• Chemical
• Biomedical
Surface properties
Hydrophobicity
Chemical structure
Roughness
Conductivity
Material properties modified by plasma
Effect of plasma on Material surface
Plasma Effect On Surface
Surface modification Induced Grafting Polymerization
Plasma surface activation
Plasma etching
Functionalization
Plasma Surface Activation
• Physical sputtering – Noble gas
• Chemical reaction - O2, H2
• Free radicals created on surface
• Coupling of free radicals with active species in
plasma
• By products - CO2, H2O and low molecular weight
hydrocarbons
Plasma etching
• For the removal of materials from surface
• Selective removal by chemical reactions and/or physical
sputtering
• Cleaning, polishing surfaces, processing plate edges
• Makes the surface rough
• Improves adhesion by increasing surface energy
Functionalization
Reactive plasma gases
Chemical groups
incorporated
Hydroxyl
Carboxyl
Carbonyl
Amino
Peroxyl
Plasma Induced grafting
• Two-step process
• Free radical formation
using inert gas plasma
• Introduction of an
unsaturated monomer
• Improves adhesion
Plasma gases and their applications
Plasma gases Application
Oxidizing gases (O2, air, H2O)
Reducing gases (H2, mixtures
of H2)
Noble gases (Ar, He)
Removal of organics and to
leave oxygen species
Replacement of F or O in
surfaces, removal of oxidation
sensitive materials, conversion
of contaminants to low
molecular weight species
To generate free radicals in
surfaces to cause cross linking
or to generate active sites for
further reaction
Plasma gases and their applications
Plasma gases Application
Active gases (NH3)
Fluorinated gases
Polymerizing gases
To generate amino groups
To make the surface inert and
hydrophobic
Polymerization of layers onto
substrates by direct
polymerization or by grafting
on Ar or He pretreated polymer
surface
Biomaterials
• Biomaterial: Non-viable material
used in a biomedical device intended
to interact with biological systems.
• Biocompatibility: Ability of a
material to perform with an
appropriate host response in a
specific application.
• Blood compatibility: A derivative of
biocompatibility, a complex function
of many parameters including
characteristics of the blood, material
and time.
Surface functionalities for biomaterials
• Biocompatibility
• Blood compatibility
• Reactive groups for attaching
biomolecules
• Compositions promoting cell growth
• Temperature sensitive coatings
• Non-fouling (protein resistant)
coatings
Antibacterial coatings
Controlled drug release
Wettability
Micropatterning
Surfaces strongly adsorbing
proteins
Surface functionalities for biomaterials
Biomedical applications of polymers
Polymers Biomedical applications
Polyethylene Tubes for various catheters, hip
joint, knee joint prostheses
Polypropylene Suture materials, hemodialysis,
blood transfusion bags
PolyTetrafluroethylene Vascular and auditory prostheses,
catheters, tubes
Polyacetals Hard tissue replacement
Polymers Biomedical applications
PMMA Bone cement, intraocular lenses, contact lenses,
fixation of articular prostheses, dentures
Polycarbonate Syringes, arterial tubules, hard tissue replacement,
hemodialyzers, blood pumps, oxygenators
PET Vascular, laryngeal, esophageal prostheses, surgical
sutures, knitted vascular prostheses
Biodegradable polymers Sutures, drug delivery matrix, adhesives, temporary
scaffolding, temporary barrier
Polyurethane Adhesives, dental materials, blood pumps, artificial
hearts and skin and blood contacting devices
Biomedical applications of polymers
Why plasma treatment for biomedical
applications?
• Lack of surface properties
• Surface properties influence cell adhesion
• RF plasma surface modification without affecting their bulk
properties
• Smooth, pinhole free ultra thin film
• Surface tuning
Surface modification of polymeric biomaterials
by RF plasma
• Modifies surface physical and chemical properties without
affecting bulk properties
• Advantageous for the design, development and manufacture of
biocompatible polymers
• Surface modification or by thin-film deposition - protein–
surface interaction and cell adhesion can be optimized for
improving biocompatibility
• RF plasma-treated polymeric biomaterials - Hindering
bacterial adhesion, found wide applications in antimicrobial
coatings.
• Antimicrobial coating on RF plasma-treated polymers can
prevent microbial adherence on the surface, thus preventing
biofilm formation.
Improving biocompatibility/blood compatibility
• Biocompatibility – not an inherent property of a material, but
results from complex interactions between an implant and the
surrounding tissues
• Polymer in biomedical application - should be
biocompatible, should have a low friction coefficient and
hydrophilicity
• Biomaterials selected for mechanical strength or stability in
the body suffer from problems associated with surface-
induced thrombosis
• Thrombosis - initiated by the deposition of a plasma protein
layer on the surface of the implanted biomaterial
• Platelets, fibrin and leukocytes adhere to the deposited protein
• Interaction between the plasma proteins and the surface of the
implant determines the adhesion, activation and spreading of
platelets, activation of coagulation, cell attachment and protein
deposition.
• Polymeric materials - modified to meet the needs of tissue
engineering
• Immobilization of protein with antithrombogenic or
thrombolytic qualities is a way of introducing the
antithrombogenic characteristics on blood-contact materials
• Antithrombogenic materials - heparinated high molecular
weight materials, urokinase immobilized high polymer
materials or plasma-treated high molecular weight materials
• Immobilization of various proteins with antithrombogenic
properties like recombinant hirudin (rHir), thrombomodulin
and human thrombomodulin on polymers
• Vascular grafts - Inner surface of the segmented polyurethane
tube modified by air-plasma treatment holds good as a suitable
substrate
• Improved blood compatibility and enhanced growth of
smooth muscle cells - Polyethyleneterephthalate (PET) films
grafted with acrylic acid using oxygen plasma, immobilized
with insulin, heparin and collagen
• Prevention of platelet aggregation - Polymer-coated encased
stents, facilitating endothelial cell growth on the inner lining of
the stent are used to prevent platelet aggregation
• Plasma treatment - improves the wettability, oxidizes the surface and
enhances endothelial cell growth and cell adhesion on polymer (eg.
polyurethane) surfaces
• Endothelial cell adhesion - also achieved by ion implantationand
carbon deposition
• PET - reinforced composite in prosthetics shows poor adhesion
• O2 plasma treated PET fibres - improved adhesion in fibre matrix
composite and increased the surface energy
Antimicrobial coating
• Adherence of bacteria to a polymer surface - results in biofilm
formation
• Biofilm - resistance to antibiotics makes the device-associated
infection difficult to treat and necessitates the removal and
replacement of the infected device
• Antibacterial agent is coated on medical polymers to prevent
biofilm formation
• Surface treatment prevents the initial adhesion of bacteria to
the polymer surface or kills the bacteria as they come in
contact with the surface
• Silver ions - possess good antimicrobial properties, anti-
inflammatory properties and enhances healing rates
• Surface properties influencing bacterial adhesion - hydrophobicity,
composition, mechanical properties and morphology
• Both metallic and ionic silver - incorporated into several
biomaterials such as polyurethane, hydroxyapatite, and bioactive
glasses
Tomato fruit at zero time, (a) similar fruit kept for 13 days into a polypropylene bag
fabricated from PP film covered by 12 alternating chitosan/pectin layers in
comparison to similar fruit kept for the same period of time in untreated PP bag (b),
and another one kept in open air (c).
(a) O2 plasma pattern; (b) N2 plasma pattern; (c) NH3-VUV pattern
Plasma for specific Biomedical applications
• Implants
• Bioseparation
• Plasma sterilization
• Biosensors
• Ophthalmology
• Plasma treatments are given to the polymers to achieve the
above-mentioned properties, namely antimicrobial properties
and bio/blood compatibility
Orthopedic implants
• Any material having desirable mechanical properties and
biocompatibility with the bone is used in bone replacement
• Mainly constructed using titanium alloys for strength and lined
with polymers that act as artificial cartilage
• The major obstacle to long-term use of metallic substrates is
bone resorption due to stress shielding, leading to their
degradation after 10–15 years
• Ultra high molecular weight
polyethylene (UHMWPE) and
polytetrafluoroethylene (PTFE) -
used in joint socket
• Polyurethane - in bone joint due
to their excellent wear,abrasion,
corrosion and fatigue resistance
• UHMWPE - used in surgical
replacement of damaged
cartilage in total joint/diseased
joint
• Biocompatibility of UHMWPE - modified by means of cross-
linking, functionalization using various plasmas such as Ar, C2F6,
C2H4, NH4, CH4 and HMDSO, and no cytotoxicity was observed
• Argon and Ar/CH4 plasma-treated samples showed little red
blood cell destruction and thus are more blood compatible
• Carbon-fibre-reinforced polyether ether ketone (PEEK) - treated
by oxygen plasmaand N2/O2 plasma to get better surface
activation for subsequent joining and coating processes, initiation
sites for the formation of calcium phosphate coatings in
supersaturated solutions
Cardiac implants
• Polymers in cardiac implants - as implant leads, artificial
hearts, stents and controlled drug release devices
• Non-biodegradable polymers - polyurethane, silicone rubber,
ethylene vinyl acetate
• Biodegradable polymers - poly(glycoliclactic acid), and high
molecular weight polyanhydride
• Polyurethane matrix synthesized with pore formers and loaded
with ciprofloxacin releases antibiotic at a controlled rate when
coated with n-butyl methacrylate by RF plasma deposition
Dental implants
• Polymethylmethacrylate (PMMA) - used in dental implants as
denture bases, artificial teeth, removable orthodontics, surgical
splinting and aesthetic filling in anterior teeth
• Many other polymers have been explored for several dental
applications such as dentures, crowns, bridges, fillings, mouth
protectors, sutures and implants
• Biofilm formation due to the adhesion of Candida albicans on
PMMA causes denture-induced stomatitis, which is a common
intraoral disease
• Surface loading of histatin 5 - either by
adsorption or chemical cross-linking,
reduces C. albicans biofilm formation
• Modification of PMMA by
copolymerization of methyl methacrylic acid
resulted in twofold increase of the
adsorption of the added amount of histatin 5
per unit surface area
• Amount of histatin adsorption on PMMA
increases more than six times when PMMA
is treated with O2 plasma compared to that
adsorbed onto untreated PMMA
Bio-separation
• Membranes used for biomedical applications should have high
ion/solute permeability, blood compatibility, mechanical
stability and dimensional stability upon swelling
• Hydrophilic composite membranes consisting of acrylic acid
polymer and porous polypropylene with high ion permeability
and dimensional stability were developed by plasma
interpenetrating polymer network techniques
• Deposition of hydrophilic monomers, namely 1-vinyl-2-
pyrrolidone, 2-hydroxyethyl methacrylate (HEMA) and methyl
methacrylate by plasma deposition onto chemical and O2
plasma-treated Nylon 4 membrane
• Plasma deposited polymer layer of dimethylaniline and acrylic
acid on the surface of PET track changed their transport
properties, especially water permeability
• Membranes used for bioseparation are fouled and clogged
when non-specific proteins are adsorbed on it reducing the
thrombogenicity of blood-contacting surfaces or
inhibiting/preventing the non-specific adsorption of protein
surfaces by polymerizing a phospholipid
• Plasma polymerization as pretreatment for phospholipids -
better reduction in platelet adhesion compared to that in
untreated polymer.
Biosensors
• Biosensors require two and three-dimensional microstructured
substrates with a chemically suited surface to mimic the basic
functions of natural tissue
• Chemical micro-patterning of cell culture by plasma
processing allows the introduction of functional groups on the
polymer surface, without affecting its bulk properties
• It enables covalent bonding for fixation and immobilization of
biomolecules on various substrates
• Pulsed RF plasma for polymerization of allylamine - for
successful DNA adsorption and hybridization.
• Micropatterning of amine groups, used in DNA array
technology was achieved with excellent thickness
controllability and uniformity in a relatively short time by
selective deposition of plasma polymerized ethylene diamine
on glass slides
• RF plasma treatment yields a more compatible interface with
biological fluids
• Hydrophobic polypropylene membrane has been made
hydrophilic on one side when treated with ammonia plasma
and coupled to urease to construct a urea sensor, and an
appreciable reduction in the response time has been achieved
Ophthalmology
• Contact lenses - should have high oxygen permeability, good
wettability by tears and resistance to deposition of protein, mucus,
lipid, microorganisms and other foreign substances on the lens
surface
• CF4 plasma-treated PMMA intraocular lens - reduces the
adhesion of proteins, the development of inflammatory cells and
the formation of cellular debris
• Surface modification of silicone with O2 and CO2 plasma - CO2
plasma more suitable for grafting functional groups on the surface
of poly(dimethylsiloxane), since CO2 could be used for a longer
period without causing surface damage, unlike O2 plasma
• Silicone rubber grafted with pHEMA by plasma-induced graft
polymerization - suitable for cell attachment and growth
• Argon RF plasma treatment of polyvinyl alcohol copolymer
hydrogel
Optimal for epithelial cell migration and proliferation
Allows migration, proliferation and synthesis of matrix
and adhesion molecules in vitro
No inflammatory response on the treated surface
PLASMA STERILIZATION
Sterilization
• Emerging application in medical, food and pharmaceutical
industries
• Complete elimination of all types of microorganisms
• Microorganisms normally sterilized
- Bacteria
- Fungi
- Yeast
- Molds
- Spores (most-resistant bacteria)
Importance of sterilization
• Design of medical devices
• To provide a sterile or contaminant-free environment
• Sterility Assessment: SAL (Sterility Assurance Level)
- the probability of a non-sterile product in million containing
a contaminant
• Surgical tools used in brain surgery needs high SAL
• Medical device manufacturers – 10-6 (one in a million devices
may be non-sterile)
• Decontamination of food storage devices
Controversy in commercial sterilizers
• 2 commercial sterilizers – Sterrad®, Plazlyte®
• Sterrad – Hydrogen peroxide + RF plasma(1987)
• Plazlyte – Per acetic acid + MW plasma(1993)
• Both gases have disinfectant properties
• Plasma – used as a detoxifying agent( Krebs et al., 1998)
Is it a plasma sterilizer or chemical sterilizer?
“…non-toxic gas mixture which does not have any disinfectant
property by themselves and they should sterilize only under the
applied electric field”
- Alexander Fridmann, Low temperature plasmas Vol 2
Requirements of a sterilization process
• Highly efficient
• Operate at/near ambient temperatures
• Short treatment times
• Non-toxic
• Minimal substrate damage
• Compatible to wide range of materials
Latest developments
IN
Plasma sterilization
Micro plasma
• Plasma needle – being confined in narrow
spaces (10 - 500 µm)
• Stable operation – non-thermal plasma
• Site-specific treatment – Dental, corneal
infections
FE-DBD – Floating electrode-Dielectric
Barrier Discharge
Floating electrode
(Human hand/ animal organ)
Dielectric Barrier Discharge
electrode
(Insulated electrode)
Gregory Fridman, 2006
Plasma-assisted blood coagulation
+ Saphenous vein cut for mouse
+ Continuous bleeding
+ 15 sec DBD treatment
+ Stops bleeding
+ Blood vessel remains sealed
+ No visible/microscopic damage
Plasma-assisted blood coagulation
(Continued)
• FE-DBD plasma promote blood clot formation for patients
suffering from clotting disorders
Biofilm treatment
• 10 min exposure kills 100% biofilm (hazardous) formation using
RF high-pressure cold plasma jet
Nina et al., IEEE Transactions on Plasma Science, (2006) 34, 1304 - 9
Other important applications
• In wound healing, the tissue damage should be minimized as
well as wounds need to be sterilized to prevent bacterial
invasion
• Treatment in melanoma skin cancer, ulcers, burns
• Treating corneal infections – In vivo test in rabbit eyes showed
strong bactericidal effect without tissue damage
• Treatment of tumor cells
RF plasma applied to human skin
• Atmospheric pressure RF plasma can be safely applied to
human skin treatment without electrical and thermal damage
S Y Moon et al., Thin solid films, (2009) 517, 4272-4275
Summary
• RF plasma treatment of polymeric biomaterials has been a topic of
extensive investigations pertaining to a wide range of applications
• Biomaterials, which have permanent contact with the body and
tissues, require unique surface properties like surface free energy,
hydrophilicity and specific surface morphology, for improved
cell/protein adhesion on the polymer surface
• Ability to modify a surface in a controlled way, deposition of cross-
linked films on complex geometries, formation of multilayer films,
rapidity, sterility and the prospect of scaling-up are the salient
features of RF plasma processing that makes it suitable for
biomedical applications
• RF plasma treatment produces three major effects on biomaterials:
surface modification, grafting and film deposition
• Surface modification improves adhesion, enhances surface
wettability and spreading, and reduces surface friction
• Antimicrobial coatings on RF plasma-treated biopolymers can
prevent microbial adherence on polymer surface, thus preventing
biofilm formation
• Either by surface modification or by thin-film deposition, protein–
surface interactions and cell adhesion can be optimized for
improving biocompatibility
• Modified polymeric biomaterials are widely used in implants,
bioseparation, biosensors and ophthalmology
• Possibility of fast and low temperature sterilization minimizing
the substrate damage especially for heat sensitive materials
• Atmospheric plasma sterilization for live tissue treatment
• Plasma needle – site-specific treatment
• A possible solution for the future medical industry
Plasma Treatment Operating conditions
• Gas (Ar) flow rate : 10 sccm
• Reactor Pressure : 12.5 mTorr
• Power : 30 –50 W
• Exposure time : 3 –7 mins
Various Doses
SL.No Etch
Time
Etch
Power
1 3 30
2 4 50
3 5 30
4 6 50
5 7 30
DEVELOPMENT OF AN ANTIMICROBIAL
IUD BY VACUUM PLASMA ETCH
METHOD
Deposition of Nanosize/ Picosize particles of
RISUG® over IUDs
by Vacuum Plasma Etch Method
Uncoated Copper T Coated Copper T
Surface morphology study
Uncoated Untreated Polyethylene Uncoated Plasma treated Polyethylene
RISUG® Coated T, 6 mins at 50 WPlasma Treated T, 6 mins at 50 W
ATOMIC FORCE MICROSCOPY IMAGES
Untreated polyethylene
Plasma treated RISUG®
drug coated polyethylene
Plasma treated PE
(6 mins, 50 W)
Microencapsulated Effervescent
Vaginal Drug Delivery System.
SMA acid
Effervescent mixture
Plasma treatment
Vaginal fluid
Effervescence
starts
CO2 liberationPressure increase
Our Idea
Plasma Treatment
Radio frequency plasma in M-PECVD-1A [S] instrument designed by MILMAN Thin Solid Films.
Flow rate of 10 sccm, Pressure 175 mT, DC bias 112V, Radio Frequency 13.56 MHz and Argon Plasma were maintained.
Sample Code Power (W) Initial Temp in C
Final temp in C
Time in mins
1 30 26.7 35.3 5
2 40 25.7 36.2 5
3 50 25.1 36.4 5
4 60 23.8 36.8 5
5 70 22.1 35.9 5
FT IR data
4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1800 1600 1400 1200 1000 800 600
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
1.1
1.2
1.3
1.4
1.5
absorb
ance
wave number
encapsulated effervescence product
plasma treated encapsulated effervescence product
styrene maleic acid
citric acid
4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1800 1600 1400 1200 1000 800 600
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
Y A
xis
Title
X Axis Title
encapsulated effervescence product
plasma 30 W
plasma 40 W
plasma 50 W
plasma 60 W
plasma 70 W
1) Negligible change in chemistry on plasma treatment.
2) O-H stretch of the alcohols in citric acid have disappeared in the encapsulated product suggesting encapsulation.
SEM Data
No plasma treatment Power 30 W
Power 40 W Power 50 W
Power 60 W Power 70 W
Plasma treatment causes surface roughness in the form of pits crevices and pores.
Honeycomb arrangement has been observed in certain situations.
Pulsing Plasma Treatment
Pulsing plasma in M-PECVD-1A [S] instrument designed byMILMAN Thin Solid Films.
Flow rate of 25 sccm, Pressure of 275 mT, Frequency of 20kHz, 70 % duty cycle and Argon Plasma were maintained.
Sample Code
Voltage (V) Initial Temp in C
Final temp in C
Time in mins
1 400 22.8 24.4 5
2 500 25.1 26.5 5
FT IR data
4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1800 1600 1400 1200 1000 800 600
0.00
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
absorb
ance
wave number
Encapsulated no plasma
plasma 500 V
plasma 400 V
No change in the chemistry of the sample
Differential Scanning Calorimetry
No change on plasma treatment.
Different from the pure drug molecule
Position [°2Theta]
20 30 40 50 60 70 80 90
Counts
0
20
40
60 ENCAP
Position [°2Theta]
20 30 40 50 60 70 80 90
Counts
0
20
40
60
plasma
Peak
no
2 θ d spacingCount
1 30.741 2.9061 47
2 31.937 2.8000 61
3 45.626 1.9867 35
Peak
no
2 θ d spacingCount
1 30.475 2.9309 46
2 31.671 2.8229 71
3 45.626 1.9867 44
Encapsulated product
Plasma treated Encapsulated product
XRD data
Plasma treatment does not cause any change to the crystal structure.
SEM Data
Well defined pits and pores on the surface.
Formation of arrays of pits.