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Chitin deacetylases: new, versatile tools in
Iason Tsigos, Aggeliki Martinou, Dimitris Kafetzopoulos and Vassilis Bouriotis
Chitin deacetylases have been identified in several fungi and insects. They catalyse the hydrolysis of N-acetamido bonds of chitin,
converting it to chitosan. Chitosans, which are produced by a harsh thermochemical procedure, have several applications
in areas such as biomedicine, food ingredients, cosmetics and pharmaceuticals. The use of chitin deacetylases for the
conversion of chitin to chitosan, in contrast to the presently used chemical procedure, offers the possibility of a controlled,
non-degradable process, resulting in the production of novel, well-defined chitosan oligomers and polymers.
hitin, a homopolymer comprising ␤-(1-4)-linked various fractions of acetylated units. It is biodegradable,
-acetyl-D-glucosamine residues is one of the
non-toxic to animals (in mice, the LD was Ͼ16 g kgϪ1),
most abundant, easily obtained and renewable
soluble in acidic solutions, available in various physical
natural polymers, second only to cellulose. It is com-
forms and much more tractable than chitin2,3. Thus,
monly found in the exoskeletons or cuticles of many
chitosan offers properties with great potential for many
invertebrates and in the cell walls of most fungi1.
Because of its high crystallinity, chitin is insoluble in
Today, several companies are producing chitin and
aqueous solutions and organic solvents2.
chitosan products on a commercial scale; the majority
Chitosan is a polycationic biopolymer that occurs
are located in Japan, where Ͼ100 billion tons of
naturally or is obtained by the N
chitosan are manufactured each year from the shells of
chitin; its name does not refer to a uniquely defined
crabs and shrimps, an amount that accounts for ~90%
compound but rather to a family of copolymers with
of the global chitosan market (approximately four tril-lion yen). The major areas of application include watertreatment, biomedical applications (including wound
I. Tsigos, A. Martinou and D. Kafetzopoulos are at the Institute of
dressing and artificial skin) and personal-care products2–13.
Molecular Biology and Biotechnology, Foundation of Research and
In addition, oligomers of chitin and chitosan have
Technology, Crete, Greece. V. Bouriotis (email@example.com) isat the Division of Applied Biology and Biotechnology, Department of
also attracted considerable attention because they have
Biology, University of Crete, Crete, Greece.
been reported to exhibit certain interesting physiological
0167-7799/00/$ – see front matter 2000 Elsevier Science Ltd. All rights reserved. PII: S0167-7799(00)01462-1
The catalytic action of chitin deacetylases; chitin is deacetylated by the enzyme chitin deacetylase to form chitosan and acetate.
activities, such as antitumour and antimicrobial activ-
ity4,11 and elicitor activity for plants4. Furthermore,
Chitin deacetylases have been purified and character-
chitin and chitosan oligomers are soluble in aqueous
ized from several fungi. The most well-studied enzymes
solutions, and can be easily characterized by a variety
are those from the fungi Mucor rouxii
18–20, Absidia coerulea
of analytical procedures (e.g. nuclear magnetic resonance
22 and two strains of Colletotrichum
spectroscopy14 and mass spectroscopy15).
23,24. The most important characteristics
Presently, chitosan is produced from chitin via a harsh
of the enzymes are summarized in Table 1. All the
thermochemical procedure. This process shares most of
enzymes are glycoproteins and are secreted either into
the disadvantages of a severe chemical procedure; it is
the periplasmic region or into the culture medium.
environmentally unsafe and not easily controlled, lead-
Furthermore, all enzymes exhibit a remarkable thermal
ing to a broad and heterogeneous range of products16.
stability at their optimum temperature (50ЊC), and
Similarly, chitosan oligomers are prepared by partial
exhibit a very stringent specificity for water-soluble
acid-hydrolysis of chitosan polymers. A chemo-enzym-
atic method using lysozyme and tri-N
Nevertheless, they vary considerably in their molecu-
derivatives as substrates has also been reported for the
lar weight and carbohydrate content, and display a wide
preparation of specific chitosan oligomers17. However,
range of pH optima. One interesting property with a
the resulting products from both methods are mixtures
potential biotechnological application for the enzymes
of randomly deacetylated chitosan oligomers with from C. lindemuthianum
and A. nidulans
is that, apartvarious degrees of polymerization.
from their thermal stability, they are not inhibited by
The use of chitin deacetylase for the preparation of
acetate, a product of the deacetylation reaction22–24.
chitosan polymers and oligomers offers the possibility
The respective genes of chitin deacetylases from the
of the development of an enzymatic process that could
fungi M. rouxii
26, C. lindemuthianum
27 and Saccharomyces
potentially overcome most of these drawbacks. Chitin
28,29 have been cloned, sequenced and charac-
deacetylase (CDA; EC 188.8.131.52) catalyses the hydrolysis
terized. Furthermore, the expression of chitin deacetyl-
-acetamido bonds in chitin to produce chitosan
ase genes from C. lindemuthianum
and M. rouxii
(Fig. 1). The presence of this enzyme activity has been
30 and Pichia pastoris
31 respectively, have
reported in several fungi18–24 and insect species25. This
article reviews the most important characteristics ofchitin deacetylases, and highlights some initial studies
exploiting their potential use in the deacetylation of
Two different biological roles have been suggested
chitinous substrates for the production of oligomer and
for fungal CDAs, namely their involvement in cell-
polymer products with novel characteristics.
wall formation and plant–pathogen interactions. The
Table 1. Characteristics of chitin deacetylases from different sources
Mucor Absidia Aspergillus
Abbreviations: Min. DP, minimum degree of polymerization of a chitin oligomer required for catalysis; NA, not available.
involvement of CDA in cell-wall chitosan biosynthesiswas demonstrated for the first time during studies to
investigate chitin and chitosan biosynthesis in fungi. Inthe case of the fungus M. rouxii
, it was revealed that
The enzyme forms an active enzyme–poly-
chitin synthase operates in tandem with chitin deacety-
mer complex and catalyses the reaction in a ‘zipper’
lase; chitin synthase synthesizes chitin by the polymeriz-
fashion towards one end of the chitin chain; it does notform an active complex with another substrate until it
ation of N
-acetyl-D-glucosamine residues from uridine
Multiple chain or random type
The enzyme forms an
and chitin deacetylase hydrolyses the N
active enzyme–polymer complex and catalyses the
bonds in the chitin chains, acting more efficiently on
hydrolysis of only one acetyl group before it dissociates
nascent rather than on microfibrillar chitin32. Similar
results were also obtained for the fungus Absidia coerulea
The enzyme forms an enzyme–polymer
where it was also found that CDA was localized near
complex and further catalyses the hydrolysis of several
the inner face of the cell wall (periplasmic region)21.
acetyl groups before it dissociates and forms a new
This spatial arrangement of CDA is in agreement with
active complex with another polymer chain
previous studies, which reported the presence of chitin
Degree of multiple attack
The maximum number of
acetyl groups that can be hydrolysed by the enzyme
synthase activity and chitin biosynthesis in the plasma
membrane fraction of fungi33. A similar biological rolehas been reported for the two CDAs (Cda1p andCda2p) from S. cerevisiae
. It was shown that these
high-pressure liquid chromatography (HPLC) and the
enzymes are required for correct ascospore wall for-
results were further verified using 1H-NMR spec-
mation28. Furthermore, it was found that Cda2p was
troscopy. It was observed that the length of the oligomer
the most active enzyme and performed most of the
was important for enzyme action (Fig. 2). The enzyme
deacetylation tasks in the ascospore cell wall29.
could not effectively deacetylate chitin oligomers with
An alternative biological role, namely the involve-
a degree of polymerization lower than three. Tetra-N
ment of the enzyme in plant–pathogen interactions, has
acetylchitotetraose and penta-N
been suggested for the CDA from Colletotrichum linde-
were fully deacetylated by the enzyme, however, in
, considering that the fungus is a plant
pathogen, and the enzyme is extracellular and active
-acetylchitoheptaose, the reducing end-residue
on chitin oligomers. Taking into account that chitin
remained intact. Furthermore, the enzyme initially
oligomers (tetramer to hexamer) elicit plant-defence
removed an acetyl group from the non-reducing end-
mechanisms (callose formation, lignification and residue of all chitin oligomers with a degree of poly-synthesis of coumarin derivatives)34,35, whereas their
merization greater than two, and further catalysed the
deacetylated forms do not36,37, it has been proposed that
hydrolysis of the next acetamido groups in a progres-
CDA might deacetylate chitin oligomers that arise from
sive fashion. This mechanism resembles the mode of
the fungus cell wall subsequent to the activity of plant
action that the enzyme exhibits on polymeric substrates39.
chitinases, and thereby diminish their elicitor activity23.
Chitin oligosaccharides (DP2–4) have also been used
Finally, a role for CDA during the penetration pro-
as model substrates to study the mode of action of CDA
cess of the fungus hypha (which contain a considerable
from C. lindemuthianum
41. Using HPLC and fast-atom-
amount of chitin) in plant tissues has been proposed.
bombardment mass spectrometry (FAB-MS), it was
Because decreasing levels of acetylation result in less-
found that in a similar manner to the M. rouxii
efficient hydrolysis of chitin by plant endochitinases, it
CDA from C. lindemuthianum
could fully deacetylate
was suggested that the penetrated hypha might be tetra-N
-acetylchitotetraose. However, it could alsoprotected by enzymatic deacetylation38.
fully deacetylate tri-N
-acetylchitotriose and the non-reducing end-residue of di-N
Mode of action
thermore, it was shown that under specific conditions
The mode of action of CDA has been studied in both
(in the presence of 3 M sodium acetate), this enzyme
chitosan polymers39 and chitin oligomers40,41. The mode
could also catalyse the reverse deacetylation reaction42.
of action of CDA from M. rouxii
has been investigated
Using FAB-MS and NMR it was demonstrated that
on an ~32% randomly deacetylated water-soluble chito-
CDA from C. lindemuthianum
could acetylate chito-
san substrate, with an average degree of polymerization
biose (GlcNGlcN) and convert it into 2-acetamido-2-
of 30. Using 1H- and 13C-NMR spectroscopy, it was
found that the enzyme hydrolysed the acetyl groups of
the substrate according to a multiple-attack mechanism
In summary, enzymatic deacetylation of both chitin
(see Glossary) with a degree of multiple attack of three.
oligomers and chitosan polymers is a well-defined reac-
This is the maximum number of successive deacetyl-
tion, in contrast to chemical deacetylation, in which
ations that could be achieved by the enzyme because
the hydrolysis of the N
-acetamido bonds of N
the maximum number of the consecutive N
D-glucosamine residues is performed in a random fash-
glucosamine residues that were found in this substrate
ion. Therefore, enzymatic deacetylation involving chitin
deacetylases offers a possibility for the preparation of
The mode of action of CDA from M. rouxii
specific novel chitosan oligomers and polymers.
oligosaccharides (DP 2–7) has also been studied40. Thesequence of chitin oligomers following enzymatic
deacetylation was identified by the alternative use
The CDA genes from M. rouxii
, C. lindemuthianum
of two specific exoglycosidases in conjunction with
and S. cerevisiae
have been cloned and characterized26–28.
Mode of action of chitin deacetylase from Mucor rouxii on chitin oligomers. Blue spheres correspond to N -acetyl-D-glucosamine residues;green spheres to their deacetylated counterparts (D-glucosamine). Numbered arrows indicate the order and the site of deacetylation. Theenzyme initially deacetylates the non-reducing end-residue of the oligomers, and further catalyses the hydrolysis of the next acetamidogroups in a progressive fashion. Tetra-N-acetylchitotetraose and penta-N-acetylchitopentaose are fully deacetylated; in tri-N-acetylchitotriose,hexa-N -acetylchitohexaose and hepta-N-acetylchitoheptaose, the reducing end-residue remains intact.
The enzymes are highly homologous and, furthermore,
peptidoglycan deacetylases, although this function has
there is a universal conserved region that exhibits a
significant similarity to the rhizobial nodulation pro-
Thus, the similarities between the CDAs highlights
teins43,44 (NodB proteins), certain regions in microbial
the catalytic domain in the CDA sequences and can
acetyl xylan esterases and xylanases45,46, and several
direct the design of an enzyme with improved efficiency
uncharacterized open reading frames (ORFs) in Bacillus
in the deacetylation of chitinous substrates.
(Fig. 3). This conserved region has been assigned asthe ‘nodB homology domain’ because of its similarity
Applications of chitosan
to NodB proteins. Apart from this region, no other
The following major characteristics of chitosan make
homologies were found between chitin deacetylases
this polymer advantageous for numerous applications:
(1) it has a defined chemical structure; (2) it can be
NodB proteins are chitooligosaccharide deacetylases
chemically and enzymatically modified; (3) it is physi-
that are essential for the biosynthesis of bacterial nodu-
cally and biologically functional; (4) it is biodegradable
lation signals, termed Nod factors43. Nod factors from
and biocompatible with many organs, tissues and cells;
different rhizobial species share a common basic struc-
and (5) it can be processed into several products includ-
ture: they are all N
-acetylglucosamine oligomers, with
ing flakes, fine powders, beads, membranes, sponges,
-acyl substitution at the non-reducing end-residue.
cottons, fibres and gels2–4. Furthermore, its main source
-acyl substitution is required for the biological
(crab and shrimp shells) is produced as a byproduct of
activity of all Nod factors because N
the seafood industry. Consequently, chitosan has found
oligomers fail to elicit any nodulation-specific responses
considerable application in various industrial areas
in host plants. NodB proteins specifically remove the
-acetyl group from the non-reducing terminal residue of
As a result of its high molecular weight, cationic char-
-acetylglucosamine oligomers, thus providing the nec-
acter and gel-forming ability, chitosan has been exten-
essary free amino group for the subsequent N
sively used in industry, foremost as a flocculent in the
The nodB homology domain present in Cellulomonas
clarification of waste-water and the detoxification of
and Clostridium thermocellum
xylanases was found to
hazardous waste2,4,5. It was found that the adsorption
be functional, exhibiting deacetylase activity against
capacity of chitosan varies with the amino-group con-
xylan, and thus contributing to the efficient hydrolysis
tent, and that polymers with ~50% deacetylation were
of acetylated xylan by xylanases45,46. Finally, the ORFs
the most effective5. The United States Environmental
sp., which also exhibit significant similar-
Protection Agency has also approved the use of com-
ity with CDAs have been suggested to correspond to
mercially available chitosan in potable-water purification
The NodB homology domain. Chitin deacetylases from Mucor rouxii, Saccharomyces cerevisiae and Colletotrichum lindemuthianum arealigned with NodB from Rhizobium melitoti, Xylanase D from Cellulomonas fimi, and acetyl xylan esterase A from Streptomyces lividans. Theblack segment indicates the NodB homology domain. Abbreviations: CDA, chitin deacetylase; NodB, nodulation protein B; XylD, xylanase D;AxeA, acetyl xylan esterase A; aa, amino acids.
systems4 up to a maximum level of 10 mg lϪ1. Fur-
pharmaceutical product (TegasorbTM, a wound-healing
thermore, chitosan has been used as a paper-coating
product), several other important applications of chito-
material, increasing the physical strength of cellulose
san as an excipient in pharmaceutical products are cur-
paper and improving the printing quality with anionic
rently being investigated6. Interestingly, in all cases the
degree of polymerization and N
-acetylation of the
Chitosan has also been used for the clarification of
oligomers and polymers proved to be extremely impor-
beverages (such as fruit juices and beers) and in the
tant, because both these parameters influence not only
agricultural sector2,4. Furthermore, chitosans (Ͼ70%
their biochemical characteristics but also their bio-
deacetylated) have recently been introduced to the
compatibility and immunological activity3. For exam-
nutritional-supplement market as a weight-loss aid and
ple, it was shown that chitin oligomers (DP 4–7) dis-
a cholesterol-lowering agent6,47. They are also used as
played a strong immuno-enhancing effect, inhibiting
a constituent of many food products, particularly in
the growth of various tumour mice cells, whereas
Japan2, and are presently approved as a food additive in
chitosan oligomers (DP 2–6) did not exhibit such an
effect4. Furthermore, high molecular weight chitosans
Chitosan polymers and oligomers have recently
with a high degree of deacetylation were more effec-
attracted considerable attention in the pharmaceutical
tive in inhibiting bacterial growth than chitosans with
and biomedical fields because of their favourable char-
a lower molecular weight and degree of deacetylation11.
acteristics, such as biocompatibility, biodegradability,
Owing to its cationic character and the presence of
antimicrobial action and non-toxicity to animals.
reactive functional groups, chitosan has also been used
Although chitosan has only been used in one registered
in the development of controlled-release technologies.
Table 2. Applications of chitosan polymers and oligomers
Recovery of metal ions and pesticides, removal of phenols, proteins, radioisotopes, 2,4,5
PCBs and dyes, recovery of solid materials from food-processing wastes
Seed- and fruit-coating, fertilizer and fungicide
Clarification and de-acidification of fruits and beverages, colour stabilization,
reduction of lipid adsorption, natural flavour extender, texture-controlling agent, food preservative and antioxidant, emulsifying, thickening and stabilizing agent, livestock and fish-feed additive, and preparation of dietary fibres
Treating major burns, preparation of artificial skin, surgical sutures, contact lenses, 3,7–13
blood dialysis membranes and artificial blood vessels, as antitumour, blood anticoagulant, antigastritis, haemostatic, hypocholesterolaemic and anti-thrombogenic agents, in drug- and gene-delivery systems, and in dental therapy
Immobilization of enzymes, as a matrix in affinity and gel permeation
Synthetic fibres, chitosan-coated paper, manufacturing material for fibres,
In such systems, the rate of drug administration is con-
chitin, has been used as a substrate. The degrees of
trolled while prolonging the duration of the therapeu-
deacetylation obtained were 0.5% and 9.5%. These
tic effects6,8. Chitosans used for these purposes have
relatively low degrees of deacetylation indicate that the
been fabricated in the form of gels, microspheres and
enzyme is not very effective in deacetylating insoluble
microcapsules, and it was found that the products were
chitin substrates. Similar results were also obtained
more stable and the rate of drug release was more sus-
using chitin deacetylases isolated from different
tained when chitosans with a high molecular weight
sources21,23,24. Pretreatment of crystalline chitin sub-
and degree of deacetylation were used6.
strates before enzyme addition seems to be necessary in
In addition, chitosan polymers have a moisturizing
order to improve the accessibility of the acetyl groups
effect on the skin, offer protection from mechanical
to the enzyme and therefore to enhance the yield and
hair damage and exhibit an anti-electrostatic effect on
the rate of the deacetylation reaction. Experiments have
hair. Their moisturizing effect (which is proportional
also been performed using chitin deacetylase from
to their molecular weight and degree of deacetylation)
and partially deacetylated water-soluble chi-
is comparable to that of an aqueous 20% propylene-
tosans as substrates39,52. It was shown that the enzyme
glycol or hyaluronic acid solution. They also protect
was effective in deacetylating polymers, with up to 97%
the skin from microbial infections. Consequently, they
deacetylation52, although this procedure has not yet
have been used widely in the cosmetic industry for skin-
been optimized. Moreover, because enzymatic
and hair-care products2. Finally, chitosan oligomers and
deacetylation is not a random process like chemical
polymers, as well as their derivatives have been used
deacetylation, new polymers with potentially different
extensively as analytical reagents, for example, in various
physical and chemical characteristics can be produced.
enzymatic reactions as substrates for chitinases, chito-
The advantages of an enzymatic process are more
sanases and lysozymes48,49. However, a broader use of
evident in the deacetylation of chitin oligomers. These
chitosan has been significantly restricted, primarily as a
compounds, in contrast to their corresponding pol-
result of the current methods of preparing these poly-
ymers, are soluble in aqueous solution and are there-
mers, which lead to products exhibiting non-uniform
fore more accessible for enzyme action. Furthermore,
enzymatically deacetylated oligomers, compared withchemically deacetylated oligomers, can easily be pro-
Enzymatic vs chemical deacetylation
duced and have a different distribution of N
Presently, chitin and chitosan are produced from crab
ated residues. A variety of well-defined chitosan oligomers
and shrimp shells by a thermochemical procedure. can be produced from a single enzymatic deacetylationFirst, shells are deproteinized under alkaline conditions,
step (Fig. 2). Another example indicating the specificity
and are subsequently demineralized to remove CaCO
of the enzymatic method and the facile preparation
under acidic conditions to produce pure chitin. The
of products is the selective N
-deacetylation of p
-deacetylation of chitin is either performed hetero-
Ј-diacetyl-␤-chitobioside [(GlcNAc) -p
geneously16 or homogeneously50. In the heterogeneous
by CDA from Colletotrichum lindemuthianum
method, chitin is treated with a hot, concentrated solu-
enzyme specifically deacetylates the non-reducing end
tion of NaOH and chitosan is produced as an insolu-
of the glycoside, converting it to p
ble precipitate. Chitosan prepared by this method is
~85–93% deacetylated. According to the homogeneous
method, chitin is also treated with NaOH, but under
This disaccharide derivative can be used to further clas-
milder conditions. This method results in a water-
sify chitinases degrading [(GlcNAc) -p
NP]. This reac-
soluble chitosan with an average degree of deacetylation
tion can also be performed chemically, but a number
of synthetic and purification steps would be required.
However, both methods have three critical disadvan-
Finally, the reverse deacetylation reaction catalysed
tages: (1) they consume considerable amounts of energy;
by CDA from C. lindemuthianum
also has several inter-
(2) they waste a large amount of concentrated alkaline
esting applications. One example is in the synthesis
solution, resulting in an increase in the level of envi-
of a triacetylated chitosan tetramer [(GlcNAc) GlcN],
ronmental pollution; and (3) they lead to products with
which can be used to explore the physiological activ-
a broad range of molecular weights and a hetero-
ity of partially deacetylated chitin oligomers53. The
geneous extent of deacetylation. However, a uniform
enzymatic reaction exhibits a regioselectivity that is
material with specific physical and chemical properties
hard to achieve by chemical methods.
is required for many high-value biomedical appli-
In conclusion, the development of a controllable
cations. Furthermore, the degree and distribution of
process using the enzymatic deacetylation of chitinous
deacetylation has been found to influence the physical
substrates presents an attractive alternative process that
and chemical properties, as well as the biological activ-
can, in principle, result in the preparation of novel
ities of this polymer and subsequently the properties of
the pharmaceutical or other industrial formulations thatare based on chitosan. In order to overcome these
drawbacks in the preparation of chitosan, an alternative
There is an increasing interest in chitin deacetylases
enzymatic method exploiting chitin deacetylases has
and as new primary structures become available, the
application of molecular biological techniques are
The effectiveness of chitin deacetylase for the prepa-
anticipated to provide the tools to tailor and manipu-
ration of chitosan has been tested using an enzyme iso-
late chitin deacetylases, resulting in enzymes with novel
lated from the fungus M. rouxii
51. Chitin, both in its
properties that can be used for the preparation of
crystalline and its chemically modified form, amorphous
chitosan polymers and oligomers. Future developments
in both basic research and biotechnological applications
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The authors’ work was funded by the EPET II pro-
Kafetzopoulos, D. et al.
(1993) The primary structure of a fungal
chitin deacetylase reveals the function for two bacterial gene prod-
gram (97EKBAN2-1.2-104) of the European Union,
ucts. Proc. Natl. Acad. Sci. U. S. A.
supported by the Hellenic General Secretariat of Research
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Particle-based biofilm reactor technology
Cristiano Nicolella, Mark C.M. van Loosdrecht and Sef J. Heijnen
Particle-based biofilm reactors provide the potential to develop compact and high-rate processes. In these reactors, a large
biomass content can be maintained (up to 30 g lϪ1), and the large specific surface area (up to 3000 m2 mϪ3) ensures that
the conversions are not strongly limited by the biofilm liquid mass-transfer rate. Engineered design and control of particle-
based biofilm reactors are established, and reliable correlations exist for the estimation of the design parameters. As a result,
a new generation of high-load, efficient biofilm reactors are operating throughout the world with several full-scale applications
for industrial and municipal waste-water treatment.
iofilm reactors are used in situations wherein the operated at high biomass concentrations to treat the
reactor capacity obtained by using freely sus-
large volumes of dilute aqueous solutions that are typi-
pended organisms is limited by the biomass cal of industrial and municipal waste-waters without
concentration and hydraulic residence time. This can
the need for separating the biomass and the treated
be the case either for slow-growing organisms (e.g.
nitrifiers, methanogens), whose growth in suspension
Although the use of biofilms overcomes limitations
requires long residence times, or for diluted feed
caused by a low reactor-biomass concentration, for
streams (often present in waste-water treatment pro-
high reactor capacities, a new bottleneck has to be con-
cesses), in which only a very low biomass concentration
sidered because the delivery of poorly soluble substrates
can be achieved without biomass retention. In these
(e.g. oxygen) to the biofilm surface might become lim-
cases, biofilms are an effective solution to successfully
iting. Systems with static biofilms (e.g. trickling filters)
retain biomass in the reactors and to improve the vol-
have small specific biofilm surface areas (typically less
umetric conversion capacity. Biofilm reactors are not
than 300 m2 biofilm mϪ3 reactor) available for substrate
particularly useful when fast-growing organisms (i.e.
transport and reaction, and thus a limited reactor
with a maximum specific growth rate Ͼ0.1 hϪ1) or
capacity (the oxygen-transfer rate is typically less than
concentrated feed streams are used1 (e.g. in industrial
3 kg mϪ3 dϪ1 for trickling filters). Therefore, static
fermentation processes). In these situations, sufficient
biofilm reactors can be useful if the biomass retention
biomass will be formed to metabolize the substrate with
and not the mass transfer is the main requirement, for
relatively short residence times without the need for
example, when large volumes of liquid with very low
any form of retention; it is the oxygen supply to the
substrate concentrations have to be treated (e.g. the
liquid phase, not the biomass concentration, which is
removal of xenobiotics from ground water). For more-
often the limiting factor. For this reason, in the major-
concentrated streams, the enlargement of the biofilm
ity of industrial fermentation processes where high specific surface area can lead to a substantial reductionsubstrate concentrations are used, biofilm formation is
in the reactor volume and the area requirements of the
either unnecessary or even disadvantageous, and the
process. A dramatic increase in biofilm surface area can
range of applications of immobilized-cell systems in
be obtained by growing biofilms as small particles. The
industry is mainly limited to waste-water treatment
choice of the optimal particle size is a compromise
processes2,3. Biofilms are extensively used in environ-
between the conversion rate and the particle sedi-
mental biotechnology because biofilm reactors can be
mentation rate. If the particles become too small (i.e. their settling velocity is too small), the process mightagain be limited by the biomass concentration that is
C. Nicolella (firstname.lastname@example.org) is at the Department
achievable in the reactor, as for cell suspensions.
of Food Science and Technology, University of Reading, PO Box 226,
Gravity separation can be enhanced by growing the
Reading, UK RG6 6AP. M.C.M. van Loosdrecht (email@example.com) and S.J. Heijnen are at the Kluyver Institute for Biotech-
biomass in the form of dense spherical aggregates.
nology, Delft University of Technology, Julianalaan 67, 2628 BC
These aggregates can either form spontaneously as large,
dense granules4,5, or attached to suspended carriers6
0167-7799/00/$ – see front matter 2000 Elsevier Science Ltd. All rights reserved. PII: S0167-7799(00)01461-X
ARTIGOS DE OPINIÃO OPINION ARTICLES Prevenção da Infecção Nosocomial da Infecção Nosocomial - ponto de vista do especialista. Prevention of Nosocomial Infection Sousa Dias, C* Serviço de Medicina Intensiva, Hospital São João ABSTRACT A infecção nosocomial ou infecção associada a Nosocomial infection or healthcare-associated cuidados de saúde (IACS) é um pro
500 MG POLVERE PER SOLUZIONE ORALE paracetamolo PRIMA DELL’USO LEGGERE CON ATTENZIONE TUTTE LE INFORMAZIONI CONTENUTE NEL FOGLIO ILLUSTRATIVO Questo è un medicinale di AUTOMEDICAZIONE che potete usare per curare disturbi lievi e transitori facilmente riconoscibili e risolvibili senza ricorrere all’aiuto del medico. Può quindi essere acquistato senza ricetta ma va usato correttamen