BIOFILM TECHNOLOGIES COMMON TERRITORY OF BIOTECHNOLOGY AND NANOTECHNOLOGY Ludmil Nikolov Biological

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BIOFILM TECHNOLOGIES COMMON TERRITORY OF
BIOTECHNOLOGY AND NANOTECHNOLOGY Ludmil Nikolov
Biological Faculty of Sofia University St.
Kl. Ohridski
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INSTEAD OF INTRODUCTION

• Activity 4.1. Nanosciences and Nanotechnologies
• 4.1.1 Nanosciences and converging sciences
• 4.1.2 Nanotechnologies and converging
  technologies
• 4.1.3 Health, Safety and Environmental Impacts

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4.1.2. Nanotechnologies and converging
technologies

• Pilot lines to study, develop and up-scale
  nanotechnology-based processes from laboratory
• Equipment and methods for nanotechnology
• Analysis of the ethical, regulatory, social and
  economic environment of nanomedicine
• Coordination in nanometrology
• Examining capacity building in nanobiotechnology

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The biofilms

• They are well spread in the nature self
  organizing formations consisted of
• Spontaneously fixed cells of microorganisms,
  plants or animals on carriers
• Produced by themselves substances (EPS -
  exopolysaccharides, DNA, precipitated salts, etc)
  
• Inclusions of nano- and micro-particles in their
  structures
• Circulating free cells in their structure

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BIOFILMS IMPACTS

• Biofilms in ecosystems
• positive role in acceleration of natural cycles
  of biogenic elements - C, N2, P, S, O2
• Biofilms in higher organisms plants, animals
  and humans
• controversial roles
• negative dental plaques, diseases prolongation,
  
• positive acceleration of digestion processes in
  high organisms.
• Biofilms in artificial systems
• negative role metal protection biocorrosion,
  decreasing of the working characteristics of
  equipment and apparatus water transport
  decrease of the transport equipment speed, heat
  exchangers - decrease of heat transfer
  coefficients, artificial organs, nano-membranes
  plug formations.
• positive role biofilm reactors, metal leaching
  acceleration.

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BIOFILM TECHNOLOGIES

• 1. PROCESSING TECHNOLOGIES
• BIOFILM AS BIOAGENTS (BIOCATALISTS) IN BIOFILM
  REACTORS IN
• ENVIRONMENTAL PROTECTION (WASTEWATER TREATMENT,
  TAIL GAS PURIFICATION)
• POTABLE WATER CONDITIONING (BIODENITRIFICATION),
• BIOMETALLURGY METAL LEACHING
• BIOACTIVE SUBSTANCES PRODUCTION ETC.
• 2. PROTECTING TECHNOLOGIES (ALSOPRESERVING,UMP
  ERMISSIBLE)
• TO AVOID THE BIOFILM FORMATION (BIOCORROSION OF
  METALS, DENTAL PLAQUES)
• TO DESTROY BIOFILMS IN ARTIFICIAL SYSTEMS (LIKE
  IN POTABLE WATER SUPLYING SYSTEMS
• IN HEALTH CARE ARTIFICIAL ORGANS WITHOUT
  BIOFIILMS
• WATER TRANSPORT EQUIPMENT PROTECTION,
  NANO-MEMBRANES WITHOUT BIOFILMS ETC).

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THE AIMS

• To draw attention to the analogies of mechanisms
  of biofilms formation and development with some
  subjects of nanotechnology.
• To show similarities of their exploitation or
  prevention of development as well as of research
  methodology with some nanotechnology.
• To illustrate one of the positive rolls, which
  the biofilms can play in industry for high
  performance biofilm reactors design and
  development
• To provoke discussion about the possibility of
  transfer of knowledge from one to another
  scientific fields.
• To give examples of so called converging
  technologies with nanotechnologies.

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METODOLOGYComparative analysisSIMILARITIES AND
DIFFERENCES OF BIOFILM TECHNOLOGIES
(BIOTECHNOLOGY)WITH NANOTECHNOLOGIES
(NANOSCIENCE)

• The similarities
• They are coming into being due to amalgamation of
  physical, chemical, physico-chemical (and
  biological phenomena), accepted as summarizing
  notion of spontaneous fixation.
• Mechanisms of biofilm formation are similar to
  those of thin layers formation in
  nanotechnologies but in addition they have all
  the pluses and minuses of their biological
  character due to the cells, which are both
  participants and reason of the formation of
  these specific living systems.
• Diffusion character of structure of biofilm in
  the terms of system approach.
• Some subsystems of biofilm large systems are
  consisted of macromolecules with nano
  characteristic parameters exoploysaccharides,
  enzymes, nonstoichiomentric compounds, nucleic
  acids (DNA, RNA etc), parts of cells (organelles,
  protoplasts etc).

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METODOLOGYComparative analysis SIMILARITIES AND
DIFFERENCES OF BIOFILM TECHNOLOGIES
(BIOTECHNOLOGY)WITH NANOTECHNOLOGIES
(NANOSCIENCE)

• The differences
• The sizes of the participants of biofilms
  microbial cells and moreover - cells of higher
  organisms (plants or animals) are bigger than the
  characteristic parameters of subjects of
  nanotechnologies
• Self-organizing character of biofilms as large
  system
• Complicated self-created structures of biofilms
• Living matter in processing systems
• Necessity of intervene into the processes of self
  organization

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PECULIARITIES OF BIOFILMS AS SELF-ORGANIZING
LIVIG SYSTEMS 1. BIOFILM STRUCTURES Fig. 1a.
Stylized scheme of biofilm system formed on
spherical carrier
a)

• A carrier
• B biofilm
• C fluid
• 1 clusters
• 2 exopolysaccharides
• 3 pores
• 4 closed area
• 5 cells
• 6 microparticles
• 7 inert macroparticles
• 8 stream lines

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1. BIOFILM STRUCTURES Fig. 1b. Stylized scheme
of biofilm system formed on plane surface carrier

• 1. Carrier
• 2. Biofilm
• 3. Strongly linked cells in the biofilm
• 4. Weakly linked cells in the biofilm
• 5. Dead cells
• 6. Stream lines
• 7. Substrate
• 8. Suspended cells
• 9. Pores
• 10. Closed area
• 11. Exsopolysaccharides
• 12. Jarosite.

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1. BIOFILM STRUCTURESFig. 2. Formation of
biofilm structures on the frontier with liquid
phase at different scales of liquid flow linear
velocity

• ? - low velocities
• B middle velocities
• C high velocities.
• 1 piques on the biofilm surface
• 2 pores (channels)
• 3 - streamers
• 4 - microorganisms
• 5 liquid flow lines
• 6 closed empty room.

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1. BIOFILM STRUCTURESFig. 3. Various states of
fixed cells at the beginning of biofilm formation

• Structures A and B
• 1 - active cells
• 2 - inactive cells
• Structure C
• 3 - weakly linked cells
• 4 - strongly bounded microbial cells

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2. DYNAMICS OF BIOFILM FORMATION and FUNCTIONING

• Admissions
• 1. Biofilm formation and functioning is carried
  out in open systems
• 2. No mass- and heat transport limitations in the
  liquid phase
• 3. No limitation regarding the constant cells
  concentrations in the liquid phase

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Stages of biofilm system developmentStage 1

• Fig. 4?. Stage 1. Adaptation of cells to the
  carrier surface
• ? carrier S substrate l distance from the
  surface of the carrier
• 1- swimming cells (suspended cell culture) 2-
  adhered cell
• Start of the stage cell adaptation
• End of the stage single attached cells

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BIOLOGICAL PHENOMENA

• swimming cells
• Cell adaptation
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - EPS on cell surface need for their future
  fixation
• - specific compounds for carrier surface
  preparation for cell fixation (adsorption)
• - byproducts
• fixed cells only sporadic single attached cells

• PHYSICO-CHEMICAL PHENOMENA
• Adsorption of nutrients from the liquid phase
• Adsorption of specific compounds excreted by
  cells
• Adsorption of cells on the carrier surface
  (spontaneous fixation)
• (Cell fixation on base of electrostatic
  interactions)

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• RESULTS
• Carrier surface prepared for cell fixation
• First adhered cells
• Needed products
• Prepared surface of cell surface for fixation
  (adsorption)
• BIOFILM SYSTEM STATE homogeneous mechanism of
  bioconversion (HMBC)
• No biofilm formation, isolated cases of attached
  cells
• Domination of swimming cells productivity
• NECESSITY OF INTERVENTION INTO SELF ORGANIZATION
• In the case of processing technologies -
  artificial carrier surface modification for
  acceleration of cell fixation
• In the case of protective (impermissible)
  technologies deliberated carrier surface
  treatment to avoid biofilm formation

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Stages of biofilm system developmentStage
2Formation of cell monolayer

• Fig.4b. Stage 2. Formation of cell monolayer
• 3 - adhered cells enveloped by exopolysacharides
• Start of the stage single attached cells
• End of the stage monocell layer.

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• BIOLOGICAL PHENOMENA
• swimming cells
• Cell adaptation
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to - products (BAC biological active
  compounds) according the aim of the technology
  - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc) -
  byproducts
• fixed cells
• Beginning of the fixed cells reproduction on the
  carrier surface isolated cases
• Bioconversion of substrates by fixed cells to
  - products (BAC biological active compounds)
  according the aim of the technology -
  substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc) -
  byproducts
• PHYSICO-CHEMICAL PHENOMENA
• Adsorption of nutrients from the liquid phase
• Adsorption of specific compounds excreted by
  cells
• Adsorption of cells on the carrier surface
  (spontaneous fixation)
• (Cell fixation on base of electrostatic
  interactions)

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• RESULTS
• Envelopment of cells with substances for biofilm
  formation mainly EPS.
• First cell configurations on the carrier (cell
  monolayer formation)
• Fixed cell reproduction
• Needed products
• BIOFILM SYSTEM STATE homogeneous-heterogeneous
  mechanism (HHMBC)
• Domination of swimming cells productivity
• Fixed cells start to participate in bioconversion
• INTERVENTION INTO SELF ORGANIZATION
• In processing technologies - no need
• In protective (impermissible) technologies
  mechanical means high shear stress around
  carrier surface, scraping off biofilm formed,
  biocides introduction in the liquid phase etc.

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Stages of biofilm system development Stage
3Biofilm structure formation

• Fig. 4 c. Stage 3. Formation of the biofilm
  structure. First critical thickness of the
  biofilm - d1Cr
• Start of the stage monocell layer
• End of the stage polycell layer, first critical
  biofilm thickness -d1Cr.

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• BIOLOGICAL PHENOMENA
• swimming cells
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• fixed cells
• Fixed cells reproduction on the biofilm surface
  and in the biofilm volume
• Bioconversion of substrates by fixed cells to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• PHYSICO-CHEMICAL PHENOMENA
• Adsorption of cells on the biofilm surface
  (spontaneous fixation)

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• RESULTS
• First cell configurations on the carrier (cell
  monolayer formation)
• Fixed cell reproduction
• Biofilm structure formation
• Needed products
• BIOFILM SYSTEM STATE mixed homogeneous-heterogen
  eous mechanism
• Domination of fixed cells productivity
• Bioconversion without internal diffusion
  limitations
• Conditions for pure biofilm kinetics
  investigation
• INTERVENTION INTO SELF ORGANIZATION
• In processing technologies - artificial biofilm
  thickness control at the first critical value -
  d1Cr (by the biofilm reactor design)
• In protective (impermissible) technologies
  mechanical means high shear stress around
  carrier surface, scraping off biofilm formed,
  chemical means biocides introduction in the
  liquid phase etc.

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Stages of biofilm system development Stage
4Stable growth of biofilm system

• Fig. 4 d. Stage 4. Stable biofilm growth. Second
  critical biofilm thickness - d2Cr
• Start of the stage first critical biofilm
  thickness -- d1Cr
• End of the stage second critical biofilm
  thickness -- d2Cr

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• BIOLOGICAL PHENOMENA
• swimming cells
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring - EPS on cell surface
• - byproducts
• fixed cells
• Fixed cells reproduction in the biofilm surface
  and volume
• Bioconversion of substrates by fixed cells to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• - new products obtained due to change of
  metabolism with substrate limited kinetics
• PHYSICO-CHEMICAL PHENOMENA
• Adsorption of cells on the biofilm surface
• Internal diffusion limitation in the zone behind
  d1Cr of biofilm

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• RESULTS
• Well developed biofilm
• Fixed cell reproduction in the biofilm volume and
  surface
• Needed products according the technology aim
• New products (usually undesired, but sometimes
  quite possible to be useful)
• Linear dependence of biofilm thickness
  development on time
• Second critical biofilm thickness - d2Cr S 0
  at the carrier surface
• BIOFILM SYSTEM STATE mixed homogeneous-heteroge
  neous mechanism (HHM)
• Domination of fixed cells productivity
• Bioconversion under internal diffusion
  limitations
• Conditions for biofilm kinetics investigation
  with internal diffusion limitations
• New products due to bioconversion under changed
  condition in the biofilm
• Biofilm system is still stable
• INTERVENTION INTO SELF ORGANIZATION
• In processing technologies - biofilm thickness
  control at desired values - higher than d1Cr (if
  necessary)
• In protective (impermissing) technologies -
  mechanical means high shear stress around
  carrier surface, scraping off biofilm formed,
  chemical means biocides introduction in the
  liquid phase etc.

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Stages of biofilm system development Stage
5Uncontrolled and unstable biofilm growth

• Fig.4 e. Stage 5. Uncontrolled and unstable
  biofilm growth. Cavities formation
• 4 cavity
• Start of the stage second critical biofilm
  thickness -- d2Cr
• End of the stage cavities formation

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• BIOLOGICAL PHENOMENA
• swimming cells
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• fixed cells
• Fixed cells reproduction in the biofilm surface
  and volume
• Bioconversion of substrates by fixed cells to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• - new products obtained due to change of
  metabolism with substrate limited kinetics
• Cell decay and lysis nearby the carrier surface
  due to lack of substrates
• Products of the decay and lysis in the biofilm
  volume (possibly also in the liquid)
• Cavities formation

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• RESULTS
• Thick biofilm with cavities
• Fixed cell reproduction in the biofilm volume and
  surface
• Needed products according to technology aim
• New products (usually undesired, but sometimes
  quite possible to be useful)
• Non linear dependence of biofilm thickness
  development on time
• Lack of substrates in the deeper biofilm layers
• Products of cells decay and lysis
• BIOFILM SYSTEM STATE beginning of destruction,
  HHM
• Domination of fixed cells productivity
• Bioconversion under internal diffusion
  limitations
• Conditions for biofilm kinetics investigation
  with internal diffusion limitations
• Production of new products due to changed
  condition in the biofilm
• Biofilm system is in the state of the beginning
  of its destruction
• INTERVENTION INTO SELF ORGANIZATION
• In processing technologies - no need - biofilm
  system destruction is starting and this process
  is uncontrollable
• In protective (impermissible) technologies
  mechanical means high shear stress around
  carrier surface, scraping off biofilm formed,
  chemical means biocides introduction in the
  liquid phase etc.

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Stages of biofilm system development Stage
6Biofilm destruction

• Fig. 4 f. Stage 6. Biofilm destruction. Third
  critical biofilm thickness- d3Cr
• 5 detached part of the biofilm structure
• Start of the stage cavities formation
• End of the stage detachment of parts of biofilm
  volume

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• BIOLOGICAL PHENOMENA
• swimming cells
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring - EPS on the cell surface
• - byproducts.
• fixed cells
• Fixed cells reproduction in the biofilm surface
  and volume
• Bioconversion of substrates by fixed cells to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• - new products obtained due to change of
  metabolism with substrate limited kinetics
• Cell decay and lysis nearby the carrier surface
• Products of the decay and lysis in the biofilm
  volume (possibly also in the liquid)

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• RESULTS
• Destroyed biofilm
• Fixed cell reproduction in the biofilm volume and
  surface
• Needed products according to the technology aim
• New products (usually undesired, but sometimes
  quite possible to be useful)
• Non linear dependence of biofilm thickness on
  time, decrease of biofilm volume
• Lack of substrates in the deeper biofilm layers
• Cavities formation
• Products of cells decay and lysis
• Detached biofilm parts swimming in the liquid
  phase
• Third critical biofilm thickness d3Cr
• BIOFILM SYSTEM STATE system destruction HHM
• Domination of fixed cells productivity
• - in the biofilm
• - in the detached biofilm parts swimming in the
  liquid phase
• Bioconversion under internal diffusion
  limitations
• Conditions for biofilm kinetics investigation
  with internal diffusion limitations
• Production of new products due to changed
  condition in the biofilm

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Stages of biofilm system development Stage
7Restart of new biofilm formation. Simultaneous
realization of all the stages.
1 - swimming cells (suspended cell culture) 2-
adhered cell 3 - adhered cells enveloped by
exopolysacharides 4 - cavity 5 - detached part
of the biofilm structure 6 - new attached cells

• Fig. 4 g. Stage 7. Restart of new biofilm
  formation
• Start of the stage cavities formation
• End of the stage detachment of parts of biofilm
  volume

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• BIOLOGICAL PHENOMENA
• swimming cells
• Swimming cells reproduction
• Bioconversion of substrates by swimming cells
  to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring - EPS on the cell surface
• - byproducts
• fixed cells
• Fixed cells reproduction in the biofilm surface
  and volume
• Bioconversion of substrates by fixed cells to
• - products (BAC biological active compounds)
  according the aim of the technology
• - substances for biofilm formation and its
  structuring (EPS, precipitated salts, etc)
• - byproducts
• - new products obtained due to change of
  metabolism with substrate limited kinetics
• Cell decay and lysis nearby the carrier surface
• Products of the decay and lysis in the biofilm
  volume (possibly also in the liquid)

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• RESULTS
• Destroyed biofilm
• Fixed cell reproduction in the biofilm volume and
  surface
• Needed products according to the technology aim
• New products (usually undesired, but sometimes
  quite possible to be useful)
• Non linear dependence of biofilm thickness on
  time, decrease of biofilm volume
• Lack of substrates in the deeper biofilm layers
• Cavities formation
• Products of cells decay and lysis
• Detached biofilm parts swimming in the liquid
  phase
• Restart of biofilm formation
• All the phases of biofilm dynamics are performing
  simultaneously
• Repeated biofilm growth
• - on the carrier using places of detached
  biofilm parts
• - on the outer biofilm surface
• BIOFILM SYSTEM STATE system renovation, HHM,
  all the stage existence
• Domination of fixed cells productivity
• - in the biofilm

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APPLICATIONS inBIOFILM REACTOR DESIGN AND
DEVELOPMENT HIGH PERFORMANCE INVERSE
FLUIDIZED BED BIOFILM REACTORS
Stage 1
Stage 2
Stage 3
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APPLICATIONS inBIOFILM REACTOR DESIGN AND
DEVELOPMENT HIGH PERFORMANCE INVERSE
FLUIDIZED BED BIOFILM REACTORS
Stage 1
Stage 2
Stage 3
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APPLICATIONS inBIOFILM REACTOR DESIGN AND
DEVELOPMENTHIGH PERFORMANCE BIODISK REACTOR
Stage 7
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CONCLUSIONSCOMON TERITORYOF BIOTECHNOLOGY AND
NANOTECHNOLOGY IS IN ACTIVE INTERVENTION IN THE
DINAMICS OFBIOFILM SELFORGANIZATION AS FOLLOW

• 1. PROCESSING TECHNOLOGIES
• To accelerate the first stage of biofilm
  formation by means of introduction of specific
  compounds for modification of the carrier surface
• 2. PROTECTING TECHNOLOGIES
• To find new appropriate approaches to eliminate
  any possibility of biofilm formation on the
  material surfaces
• To develop new nanotechnologies for
  biofilm-defended membranes production for ultra
  filtration under industrial conditions