637377

research-article2016
JBC0010.1177/0883911516637377Journal of Bioactive and Compatible PolymersLiu and Chen

JOURNAL OF

Bioactive
and
Compatible
Polymers
Communication

Journal of Bioactive and

Enzyme immobilization on
Compatible Polymers
1­–15
© The Author(s) 2016
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DOI: 10.1177/0883911516637377
jbc.sagepub.com

Yue Liu1,2 and Jonathan Y Chen1

Abstract
Enzymes are excellent catalysts in many applications due to their biocompatibility, low energy
consumption, unique selectivity, and mild reaction condition. However, some disadvantages
limit the usage of enzymes in end uses, such as low stabilities and difficult recovery. In order to
overcome these disadvantages, enzyme immobilization was developed. Among various kinds of
substrates for attaching enzyme, cellulose and its derivatives are one of the ideal matrixes because
they are low cost, nontoxic, renewable, biodegradable, and biocompatible. In this review, we
summarize recent progress in the research of enzyme immobilization on cellulose matrixes.

Keywords
Cellulose, cellulose derivative, enzymes, immobilization

Introduction
Enzymes are macromolecular biological catalysts that promote chemical reactions in living sys-
tems. They are biocompatible, biodegradable, and derivable from renewable resources.1 Enzyme
products have been widely used in both basic research and industrial applications because of their
low energy consumption, unique selectivity for substrates, mild reaction conditions, and water
solubility.2 With development of biotechnology, most of the enzymes can be produced with com-
mercially acceptable prices.3
Notwithstanding all these advantages, some drawbacks limit enzyme uses, such as instability
outside physiological condition and difficult recovery and re-use of enzymes.4 Reusability
of catalysts is a key requirement in catalytic process. Reusing enzymes requires separation and
purification which are costly and may cause contamination and loss of activity.5 Technologies of
enzyme immobilization were developed to improve the stability of enzymes and reduce the cost of

1School of Human Ecology, The University of Texas at Austin, Austin, TX, USA
2Department of Chemistry, School of Science, Tianjin University, Tianjin, China

Corresponding author:
Jonathan Y Chen, School of Human Ecology, The University of Texas at Austin, Austin, TX 78712, USA.
Email: jychen2@austin.utexas.edu

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2 Journal of Bioactive and Compatible Polymers 

enzyme separation and purification. Immobilizing enzyme on insoluble supports allows not only
the re-use of enzymes, but also the modulation of the catalytic properties.6
Enzyme immobilization is a confinement of enzyme to a phase (matrix/support) different from
that of substrates and products.7 Characteristics of the phase are of paramount importance in deter-
mining performance of an immobilized enzyme system.8 An ideal matrix for enzyme immobiliza-
tion should be inert, stable, affordable, resistant to mechanical force, and biocompatible without
compromising the protein structure.9 In general, there are three types of matrixes: inorganic
matrixes, synthetic polymers, and natural polymers. Among many inorganic matrixes, silica mate-
rials have drawn most attention.10 Mesoporous silica SBA-15, silica gels, and vesicular and fumed
silica are typical silica matrixes for the enzyme immobilization. Silica SBA-15 is able to be used
for large-scale enzyme immobilizations.10 Synthetic polymers form a large and varied group of the
enzyme matrixes which are insoluble supports for enzyme trapping.11 In addition, functionalized
resins are widely used as commercial matrixes for enzyme immobilization. An example of the
commercial matrixes is ECR8205 epoxy methacrylate (Purolite Life Sciences Co., USA), one of
the commercial epoxy-activated resins.12 It is mechanically stable and finally immobilized biocata-
lysts can be used in stirred tank or bed reactor. Similarly, amino-activated and octadecyl-activated
resins are commonly used commercial matrixes. However, most of the resins are not biodegradable
and environmentally friendly. Natural polymers are of great interest because of their biocompati-
bility, biodegradability, nontoxicity, and chemical stabilization.13 The natural polymers most com-
monly used as enzyme matrixes include chitosan, alginate, agarose, and cellulose.14 Agarose was
used as commercial matrixes (Streamline DEAE, GE Healthcare Life Sciences Co).10
As one of the renewable biomaterials, cellulose has been extensively studied through theoretical
and empirical approaches due to its biocompatibility, biodegradability, chemical stabilization, and
low contamination risk to environment.15 Cellulose is one of the most abundant biopolymers on
earth, existing in wood, cotton, hemp, and other plant-based materials and serving as a dominant
reinforcing phase in plant structures.16 Cellulose is also synthesized by algae, tunicates, and some
bacterias.17 Cellulose is robust, dual hydrophilic/hydrophobic, nontoxic, and chemically inert
under physiological conditions, which is helpful for the survival of enzymes. In addition, a number
of hydroxyl groups on cellulose surface are capable of chemical reactions. Therefore, cellulose
materials are suitable for enzyme immobilization. Apart from uses of cellulose, the cellulose can
be processed into its derivatives via chemical, enzymatic, or microbiological methods.18 Cellulose
derivatives, such as carboxymethylcellulose (CMC), cellulose acetate (CA), and cellulose nitrate,
are large-scale commercial raw materials in chemical and biological industries because they are
inexpensive, nontoxic, renewable, biodegradable, and biocompatible.19 More importantly, cellu-
lose derivatives have functional groups to attach enzymes. Cellulose derivatives are also ideal
substrates of enzyme immobilization.20
Various immobilization techniques have been used in immobilizing enzymes onto cellulose and
its derivatives matrixes. These techniques can be broadly divided into covalent and physical meth-
ods according to the molecular forces between enzymes and matrixes.21 This article provides an
overview on existing techniques used for immobilizing enzymes onto cellulose matrixes. Collected
data are summarized in Table 1.

Covalent immobilization method

The method of covalent immobilization is that enzymes are immobilized onto matrixes through
chemical interactions between enzymes and matrixes (Figure 1). Covalent binding of enzymes is an
effective immobilization and has been demonstrated as the most stable interaction for the immobili-
zation of enzyme molecules to the matrixes.46 However, covalent binding is a relatively complicated

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Liu and Chen 3

Table 1.  Cellulose matrixes for enzymes immobilization.

Matrix Enzyme Method Application References

Cellulose powder Candida rugose Covalent binding Biocatalyst Girelli et al.22
lipase
Cellulose microfiber Laccase, urease Adsorption Biocatalyst Xing et al.23
Tissue C. rugose lipase Covalent binding Biocatalyst Girelli et al.22
Cellulose membrane Glucose oxidase Entrapment Biosensor Yabuki et al.24
Electrospun cellulose C. rugose lipase Covalent binding Biocatalyst Huang et al.25
nanofiber membrane
Cellulose hydrogel Peroxidase Covalent binding Biocatalyst Isobe et al.26
Cellulose macroporous Galactose Covalent binding Biocatalyst Bílková et al.27
beads oxidase
Cotton yarn Trypsin Covalent binding Biomedicine Nikolic et al.28
Nonwoven cotton fabric Lysozyme Covalent binding Biomedicine Edwards et al.29
Cotton gauze bandage Trypsin Covalent binding Biomedicine Seabra and Gil30
Make-up remover pads C. rugose lipase Covalent binding Biocatalyst Girelli et al.22
Loofah sponge Lipase Covalent binding Biocatalyst Gong et al.31
Cotton nanocrystal Lysozyme Covalent binding Biomedicine Edwards et al.32
Nanofibrillated cellulose Alkaline Covalent binding Biocatalyst Arola et al.33
phosphatase
Cellulose acetate beads Catalase Covalent binding Biosensor Yildiz et al.34
Cellulose acetate films Lipase Adsorption Biocatalyst Kosaka et al.35
Carboxymethylcellulose Polyphenol Covalent binding – Arica36
beads oxidase
Carboxymethylcellulose film Avidin Adsorption – Orelma et al.37
Cellulose–chitosan beads Penicillin G, Covalent binding Biocatalyst Luo and Zhang38
acylase
Carboxymethylcellulose– Amylase Adsorption Biocatalyst Singh and
silver nanoparticles–silica Ahmad39
hybrid
Cellulose acetate–TiO2 gel Glucose oxidase Entrapment – Kurokawa and
fiber Ohta40
Cellulose acetate–TiO2 gel Urease, lipase Entrapment Biocatalyst Hatayama et al.41
fiber
Cellulose acetate–ZrO gel β-Galactosidase Entrapment Biocatalyst Ikeda et al.42
fiber
Cellulose-coated magnetite Alpha-amylase Covalent binding Biocatalyst Namdeo and
nanoparticles Bajpai43
  Urease Entrapment Biomedicine Mahlicli and
Altinkaya44
Wood mimetic hydrogel Lipase Entrapment – Kim et al.45
beads

method because of some additions of chemical coupling agents.47 The covalent attachment involves
binding amino acid residues (–NH2, –COOH, –SH) to support matrixes.48 A requirement for the sup-
port matrixes is availability of a specific functional group of enzymes.49 Hydroxyl groups on cellu-
lose surface can be interacted weakly with an enzyme directly. Therefore, to get a strong covalent
immobilization, further functionalization steps are required. Functional groups that can react with
amino acid residues were introduced onto cellulose surfaces by a modification of cellulose matrixes.

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4 Journal of Bioactive and Compatible Polymers 

Figure 1.  Illustration of enzymes immobilized on a matrix through covalent binding.

There are two strategies for the functionalization of cellulose. The first strategy is to introduce amino
onto cellulose surfaces to react with –COOH of amino acids.32 The second strategy is to introduce an
aldehyde group, carboxyl group, or epoxy group to react with –NH2.13,50 Many studies have been
carried out on modifying cellulose to couple with enzymes.

Introduction of amino group

To immobilize enzymes on cellulose matrixes, carboxyl groups of enzymes were used to react with
functional groups of the matrixes, such as –NH2. There are no nitrogen-containing groups on cel-
lulose, but amino groups can be introduced through reactions between hydroxyl groups of cellu-
lose and functional groups of amino compounds. Esterification is one of the common and easy
methods to introduce the amino groups onto cellulose surface.51–53 Taking glycine for instance, as
shown in Figure 2(a), a carboxyl group of glycine reacts with a hydroxyl group of cellulose to
introduce an amino group on cellulose. Then, the amino group is able to couple with a carboxyl
group of enzymes. Edwards et al.54 reported that lysozyme was immobilized onto glycine-esteri-
fied cotton cellulose through a carbodiimide reaction. Antimicrobial activity observed on the
lysozyme-bounded cotton fibers was greater than that of lysozyme in solution due to an enhanced
stability of the enzyme afforded by the conjugation to cellulose.54 Employing the same method,
different glycine-esterified cotton fabrics (cotton twill, cotton spunlace nonwoven fabric, and
woven print cloth) were used as substrates for attachment of lysozyme.51 In addition, through an
amide linkage between the enzyme’s functional group and glutamate residues, lysozyme was
attached to the amino-glycine-cellulose, which was prepared by esterification of glycine in a for-
mation of cotton nanocrystals.32 Apart from enzymes, peptides were immobilized on esterified
cellulose. A human neutrophil elastase (HNE) substrate, n-succinyl-alanine-alanine-valine-para-
nitroanilide, was covalently attached to the glycine-esterified cotton cellulose nanocrystals with
HNE sensor activity.52
Other methods to introduce amino groups onto cellulose matrixes were studied. Figure 2(b)
demonstrates a process that an enzyme is attached to amino-propyloxy ether of cellulose (cell-
alkaline phosphatase (AP)). Cellulose was treated with 3-aminopropyltriethoxysilane to form cell-
AP. Cell-AP was then reacted with HNE chromogenic para-nitroanilide peptide substrates to form
a covalently linked conjugate of cellulose through amide bond between the Cell-AP amine and
succinyl carboxylate of the substrate.55

Introduction of aldehyde group and carboxyl group

Aldehyde and carboxyl groups are activated groups which can bind with amino groups of enzymes.
To obtain aldehyde or carboxyl groups, hydroxyl groups of cellulose were oxidized by different

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Liu and Chen 5

Figure 2.  Strategies for enzyme immobilization on (a) glycine-esterified cellulose and (b) cell-AP.

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6 Journal of Bioactive and Compatible Polymers 

chemical methods. Periodate oxidation is one of the most commonly used oxidation method. As
shown in Figure 3(a), it is a reaction converting 1, 2-dihydroxyl (glycol) groups to a pair of alde-
hyde groups at the positions C2 and C3 of an anhydroglucose unit which can couple with enzymes.50
This method has been widely used in structural analysis of carbohydrates since it is a highly spe-
cific reaction without significant side reactions.56
Many different cellulose matrixes are activated by periodate oxidation used in immobilization of
enzymes.57–59 Isobe et al. reported a technique of immobilizing an enzyme using cellulose hydrogel as
a matrix.26 The cellulose hydrogel was prepared from an aqueous alkali–urea solvent and activated by
partial oxidization to introduce aldehyde groups.26 Then, peroxidase and AP were covalently bound to
the cellulose hydrogel by a Schiff base formation between the aldehyde and amino groups of enzymes.26
The immobilized enzymes retained high enzymatic activities and were potential functional materials
for industrial and biomedical applications.26 Various enzymes were immobilized on periodate-
oxidized cellulose matrixes. A glucoamylase was immobilized on periodate-oxidized cellulose fiber
from bagasse with a good stability, resulting in a 36% relative remaining activity for the immobilized
enzyme.59 Lipase was immobilized on different cellulose materials, such as cellulose powder, cotton
buds, and cotton tissue.22 Sodium periodate-oxidized pulp fiber was used as a new carrier for pectinase
immobilization.60 Galactose oxidase was also immobilized on magnetic cellulose bead through perio-
date oxidation.61 Employing the similar method, cotton yarn, viscose yarn, and cotton gauze bandage
were oxidized by periodate oxidation and used as substrates for trypsin immobilization.30,62
Some natural cellulose materials can be used as matrixes for enzyme immobilization. A loofah
sponge was studied as a carrier for covalent immobilization of lipase. Cellulose in loofah sponge
was oxidized by sodium periodate to introduce aldehyde groups and then coupled with lipase.31
The immobilized lipase presented improved thermal stability and reusability.31 Periodate oxidation
was also used in modifications of cellulose derivatives.63–65 Periodate-oxidized CA membrane was
reported as a matrix for immobilization of cholesterol oxidase, and enzyme attached membrane
could be used to construct a fiber-optic fluorescent biosensor.64
Besides the above cellulose materials, nanoscale cellulose materials are ideal matrixes for
enzyme immobilization. However, it is difficult to recover nanoscale materials. Therefore, for an
easy recovery, some modifications were made using magnetic cellulose nanoparticles (MCN) pre-
pared by introducing magnetite. Covalent attachment of alpha-amylase to MCN resulted in the
formation of a novel starch degrading system.43
Recently, various enzymes (e.g. hen egg lysozyme and galactose oxidase) were immobilized onto
cellulose nanocrystals prepared from microcrystalline cellulose through a 2, 2, 6, 6-tetramethylpyper-
idin-1-oxyl (TEMPO)-mediated oxidation.66 TEMPO-mediated oxidation is a method to selectively
oxidize hydroxyl groups of polysaccharides into carboxylate and aldehyde groups, thus allowing for
a direct amide coupling.67 TEMPO-meditated oxidation converts hydroxyl groups to aldehyde or
carboxyl groups at positions C6 of an anhydroglucose unit (Figure 3(b)), which is different from
periodate oxidation. Barazzouk and Caneault reported an immobilization of tryptophan (Trp) and
Trp-based peptides onto oxidized nanocellulose (ONC) obtained by the TEMPO-mediated oxidation.
The ONC-amino acid and ONC-peptides could be used as biosensors because of their fluorescence
properties.68,69 Bioactive films with long-term stability and activity were prepared by conjugating AP
onto TEMPO-oxidized nanofiber cellulose (NFC).33 The activity of free AP decreases by 90% after
16 h at 21°C, but the conjugated AP-NFC shows no decrease in AP activity after 168 h at 21°C.33

Introduction of epoxy group

Epoxy groups are able to react with amino groups of enzymes. Therefore, introducing epoxy
groups on cellulose matrixes is an effective way to couple with enzymes. Halogen-containing

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Liu and Chen 7

Figure 3.  Strategies for enzyme immobilization on (a) periodate-oxidized cellulose and
(b) TEMPO-cellulose.

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8 Journal of Bioactive and Compatible Polymers 

Figure 4.  Strategy for enzyme immobilization on epoxy-cellulose.

epoxides are always used to react with cellulose. Figure 4 demonstrates a strategy of enzyme
immobilization on cellulose through the epoxy method. Epoxy groups were introduced onto cel-
lulose via a nucleophile reaction between chloride groups of epoxy chloropropane and hydroxyl
groups of cellulose.38 Then, penicillin G acylase (PGA) was attached on cellulose microspheres
activated with epoxy chloropropane.38 The immobilized PGA exhibited a highly effective catalytic
activity, thermal stability, and an enhanced tolerance to pH variations.38 Roy and Gupta studied an
immobilization of lactozyme (a commercially available product of β-galactosidase from
Kluyveromyces fragilis) on cellulose beads via an epoxy chloropropane coupling.70 The immobi-
lized lactozyme showed a lower activity than free enzyme in relatively low temperature
(30°C–50°C) and neutral conditions, but it had a better tolerance to pH and temperature, and
showed a good removability.65 Employing the same reaction principle, polyphenol oxidase was
immobilized on CMC hydrogel beads.36 By comparing to free polyphenol oxidase, a higher ther-
mal stability and heat storage capacity were observed.36
In addition, some complex epoxides also were used in epoxidation of cellulose. An example is
1.4-buntanediol diglycidyl ether often used to activate bacterial cellulose. As a result, glucoamyl-
ase was immobilized on epoxy-activated bacterial cellulose beads.51 Graft copolymerization of
glycidyl methacrylate (GMA) onto cellulose filter paper (CFP) was carried out by a free-radical
initiating process. Urease was immobilized on CFP-g-GMA discs through a covalent between the
amino groups of urease and epoxide groups of the matrix.71 A slight decrease in the immobilized
urease activity was recorded when compared with soluble enzymes due to the structural changes in
urease induced by the interaction of the macromolecule with the matrixes.71
Overall, the covalent attachment is an effective and stable method to immobilize enzymes onto
cellulose matrixes. However, the stable chemical bonding tends to weaken the enzymatic activity.

Physical immobilization methods

Different from the covalent attachment, physical methods use physical interactions to immobilize
enzymes onto cellulose substrates, in order to retain catalytic activities of enzymes. Physical methods
are simple and feasible compared to the covalent coupling methods. Therefore, physical methods
have a wide range of applications, especially in some industrial sectors where low cost and simple
process are needed. Primary physical immobilization methods include entrapment and adsorption.

Entrapment method
Entrapment can be defined as a physical restriction of an enzyme with a confined polymer
network.72 This method differs from the chemical coupling methods described above, in that
enzymes are not chemically bound to the matrix.73 As illustrated in Figure 5, enzyme entrapment

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Liu and Chen 9

Figure 5.  Illustration of enzymes immobilized on matrix through entrapment.

is typically achieved using a polymer network such as an organic polymer and is usually performed
in suit.21 The entrapment method is based on the occlusion of an enzyme within a polymeric net-
work that allows the substrate and products to pass through but retains the enzyme.74
A traditional enzyme entrapping method is sol-gel method.75 Various polysaccharide hydrogels
(alginate, chitosan, agarose, and carrageenan) have been employed for entrapments of different
enzymes such as lipases, lactase, invertase, and peroxidase.76–78 Cellulose is one of the ideal
matrixes for enzymes entrapment. However, the development of cellulose hydrogel has been ham-
pered by the difficulty of dissolving cellulose, because cellulose is highly crystalline.45 Therefore,
soluble cellulose derivatives such as CA were always used as matrixes for enzyme immobilization.
CA-based hemodialysis membranes entrapped with urease were produced and protein adsorption
capacity of the membranes was investigated.40 CA and metallic oxide composites were another
type of commonly used matrixes for enzyme immobilization.79–81 Various enzymes were entrap-
immobilized on CA and metal alkoxide composite gel. CA–ZrO2 fibers were prepared from CA
and zirconium alkoxide onto which invertase was entrap-immobilized.82
With the development of ionic liquids (IL) used in dissolving cellulose, a new method of
enzymes entrap-immobilization into cellulose matrixes has been developed using a cellulose-in-IL
dissolution and regeneration process. Cellulose and laccase were dissolved in 1-butyl-3-methylim-
idazalium chloride to obtain homogeneous solution, and then the solution was cast into thin films
and reconstituted into distilled water.83 Enzymes were physically entrapped within a cellulose
matrix via solution processing demonstrating a sustained activity. Lipase from Candida rugosa
was entrapped in cellulose and cellulose-biopolymer composite hydrogels by dissolving cellulose
and lipase in biocompatible IL 1-ethyl-3-methylimidazolium acetate ([EMIM]Ac), followed by
reconstitution.45 Employing similar strategy and same IL, the same enzyme was successfully
entrapped into wood mimetic beads containing cellulose, xylan, and lignin.84 Yabuki et al. to
develop a novel method for preparing enzyme immobilized cellulose membranes. Glucose oxidase
was attached onto the electrode surface, then the electrode surface was coated with a cellulose
[EMIM]Ac solution, and the IL was removed by immersing the electrode into water.24 Enzyme
activity was retained in the membrane and the duration of stability of the enzyme electrode was
lengthened to be at least 6 months.24 In addition, Cai and Zhang studied on rapid dissolution of

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10 Journal of Bioactive and Compatible Polymers 

Figure 6.  Illustration of enzymes immobilized on matrix through adsorption.

cellulose in LiOH/urea and NaOH/urea aqueous solutions, which would give a new strategy to
form cellulose gel used for enzyme immobilization.85

Adsorption method
Adsorption is another approach for enzyme physical immobilization. Among the enzyme immobi-
lization techniques, adsorption onto a solid material is easiest to perform.37 It is different from the
entrapment method. Generally, cellulose matrixes were synthesized first and then enzymes were
immobilized onto the matrixes by adsorbing. Figure 6 is an illustration of enzymes immobilized on
a matrix through adsorption. Adsorption makes use of physical interactions taken place between
the matrix and enzyme that include van der Waals forces, ionic interactions, and hydrogen
bonding.86 The physical interactions typically do not change native structure of enzymes. This
prevents active sites of enzymes from disturbing and allows the enzymes to retain its activity.87
However, the bonds between matrixes and enzymes are rather weak so that immobilized enzymes
are easy to be washed out from matrixes.
Many studies reported that enzymes were immobilized on cellulose matrixes by adsorption.
Avidins were immobilized on regenerated cellulose beads and cellulose nanofiber, which was per-
formed using a physical adsorption, and the adsorption was irreversible.37 Immobilization of
enzymes by adsorption is a simple and inexpensive method to improve stabilities of enzymes and
does not change the enzymes chemically. Comparing with a free pectinase, an adsorb-immobilized
pectinase on regenerated cellulose beads showed higher thermal and pH stabilities.88 An immobi-
lization of proteolytic enzymes trypsin and α-chymotrypsin was studied. Enzymes were reversibly
adsorbed on microcrystalline cellulose matrixes.89
Some modifications on cellulose would help enzyme immobilizations through adsorption.
Kosaka et al. prepared ultrathin films of lipase immobilized cellulose esters (CA, CA propionate,
and CA butyrate). Lipase as an adsorbate immobilized on the films exhibited a higher activity
compared with free lipase and lost ca. 30% of original enzymatic activity after recycled three
times.35 CMC–Ag nanoparticles–silica hybrids were synthesized as an efficient adsorbent matrix
for diastase alpha-amylase.39 An immobilization was done by adsorption. Catalytic ability of the
immobilized enzyme was improved significantly, capable of retaining 74% of its initial activity
after six cycles.39
A layer-by-layer (LbL) technique appeared to be an advantageous method to immobilize bioactive
molecules such as enzymes adsorbed on top or inside of the LbL polyelectrolyte film, due to the pos-
sibility of maintaining structures and functionalities of the enzymes.90 As compared to other adsorption
methods of immobilizing enzymes, LbL is more versatile without assistances of carbohydrate-bonding
modules or cellulose-binding polysaccharides.23 Lipase was assembled into multiple layers on a poly-
ethylenimine-treated cotton flannel cloth, utilizing the enzyme property of forming bimolecular aggre-
gates via the LbL immobilization technique.91 Cellulose microfibers–enzyme composites were

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Liu and Chen 11

fabricated through the LbL nanoassembly by alternate adsorption and enzymatic catalytic activity was
retained proportional to the number of coated enzyme layers.23

Conclusion
Enzymes as effective and environmentally friendly catalysts play an important role in research
laboratories and industries. There are various studies on enzyme immobilizations to improve
capacities of enzymes. Cellulose and its derivatives are good matrixes for enzyme immobilization
due to their biocompatibility, biodegradability, bioactivity, and low contamination risk to the envi-
ronment. There are two types of immobilization methods. The covalent immobilization method is
that enzymes are immobilized onto matrixes through chemical interactions between enzymes and
matrixes. It provides the most stable interaction between enzymes and matrixes, but it generally is
a more complicated method comparing with physical immobilization methods and results in a
decrease in catalytic activity because stable chemical bonding changes the structure of enzymes.
The use of physical interactions to immobilize enzymes on cellulose matrixes is a second method
which would retain enzyme catalyst activities. But immobilized enzymes are easy to be washed out
from matrixes because of no chemical bonding between enzymes and matrixes. Therefore, the
durability of enzyme immobilization needs to be improved.

Declaration of conflicting interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publi-
cation of this article.

Funding
The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publica-
tion of this article: The author(s) thank the China Scholarship Council to sponsor Yue Liu as a visiting scholor
at The University of Texas at Austin.

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