Biochemical Engineering Journal 72 (2013) 96–101

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Biochemical Engineering Journal

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Regular article

Co-immobilization of oxalate oxidase and catalase in films for scavenging

of oxygen or oxalic acid
Sandra Winestrand a,b,∗,1 , Kristin Johansson c,1 , Lars Järnström c , Leif J. Jönsson b
a
Department of Chemistry and Biomedical Sciences, Karlstad University, SE-651 88 Karlstad, Sweden
b
Department of Chemistry, Umeå University, SE-901 87 Umeå, Sweden
c
Department of Chemical Engineering, Karlstad University, SE-651 88 Karlstad, Sweden

a r t i c l e i n f o a b s t r a c t

Article history: Oxalate oxidase has potential to act as an oxygen scavenger in active packaging to increase the shelf-life
Received 5 October 2012 of food and beverages, while simultaneously producing the protective packaging gas carbon dioxide. This
Received in revised form study shows that oxalate oxidase from barley can be immobilized with retained catalytic activity through
14 December 2012
entrapment in a latex polymer matrix. Conditions for formation of film containing oxalate oxidase have
Accepted 9 January 2013
been evaluated as well as effects of storage and latex on enzyme activity, migration of enzyme in films,
Available online 18 January 2013
and the ability of the latex films to resist higher temperatures. Drying of enzyme-containing latex films at
75 ◦ C prior to conditioning at 30 ◦ C resulted in higher activity than drying solely at 30 ◦ C, or drying at 95 ◦ C
Keywords:
Oxalate oxidase
or 105 ◦ C followed by conditioning at 30 ◦ C. Storage of films in air at 4 ◦ C for 14 days did not negatively
Active packaging affect the enzymatic activity. Inclusion of catalase in films with oxalate oxidase effectively prevented
Oxygen scavenger release of hydrogen peroxide. The results suggest that the immobilized enzyme can successfully be used
Oxalic acid both as an oxygen scavenger and as an oxalic-acid scavenger.
Latex © 2013 Elsevier B.V. All rights reserved.
Entrapment

1. Introduction
Oxalate oxidase (EC 1.2.3.4) catalyzes the conversion of oxalic
acid and molecular oxygen to carbon dioxide and hydrogen perox-
ide (I).
HOOC COO− + H+ + O2 → 2CO2 + H2 O2
Oxalate oxidase has been found in fungi, bacteria, and plants
like barley and wheat. Oxalate oxidase belongs to a large family
of germin-like proteins that have been termed cupins because of
their conserved ␤-barrel fold [1]. The active site, which contains a
manganese ion, is found within the center of the ␤-barrel. The pH
optimum of oxalate oxidase has been reported to be 3.8 [2] and
the KM value has been determined to 0.28 mM [3]. The sensitivity
of oxalate oxidase towards inhibitory compounds has been studied
previously [2–5].
Oxalate oxidase is mainly used in the analysis of the levels of
oxalic acid in clinical samples, such as blood plasma and urine [1].
Analysis of oxalic acid in clinical samples is of relevance for patients
with kidney stones, which mainly consist of crystallized calcium
∗ Corresponding author at: Department of Chemistry, Umeå University, SE-901
87 Umeå, Sweden. Tel.: +46 90 7865467; fax: +46 90 7867655.
E-mail address: Sandra.Winestrand@chem.umu.se (S. Winestrand).
1
These authors contributed equally to this work.
oxalate. Patients suffering from reoccurring kidney stones are often
recommended a diet with low content of oxalic acid. Another
approach is to treat patients suffering from primary and secondary
hyperoxaluria with the oxalate-degrading bacterium Oxalobacter
formigenes [6]. There are also problems with calcium oxalate pre-
cipitations in industry, for example in the pulp and paper industry
(1) [7]. Thus, one potential application for oxalate-degrading enzymes
and microorganisms is to prevent formation and precipitation of
calcium oxalate.
Another potential application for oxalate oxidase is as an oxygen
scavenger. The most widely used type of active packaging today is
based on the scavenging of oxygen. Oxygen-scavenging enzymes
suggested for active packaging application today include glucose
oxidase, laccase and ethanol oxidase (Table 1). The fact that oxalate
oxidase could potentially scavenge both oxalic acid (to make the
food healthier) and oxygen (to increase the shelf-life of food) makes
it a very interesting enzyme in the field of active packaging.
For the production of enzyme-based active packaging, it is desir-
able to immobilize the enzyme onto any of the components of
the packaging material. The simplest immobilization technique for
active packaging would be entrapment of the enzyme in a disper-
sion coating. Glucose oxidase was recently successfully entrapped
in a polymer-based dispersion coating for the use as an oxy-
gen scavenger in active packaging [8]. In this work, we have
investigated immobilization of oxalate oxidase in films through
entrapment in a latex polymer matrix. The possibility to utilize

1369-703X/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.bej.2013.01.006
 S. Winestrand et al. / Biochemical Engineering Journal 72 (2013) 96–101
Table 1
Oxygen scavenging enzymes suggested for packaging applications.
Oxygen-scavenging enzyme Reaction catalyzed
Glucose oxidase glucose + O2 → gluconic acid + H2 O2
Laccase 4Ar–OH + O2 → 4Ar–O. + 2H2 O
Ethanol oxidase ethanol + O2 → acetaldehyde + H2 O2
oxalate-oxidase-based systems either as oxygen scavengers or as
oxalic acid scavengers was explored. Furthermore, we also inves-
tigated the possibility to co-immobilize catalase to degrade the
hydrogen peroxide produced in the reaction catalyzed by oxalate
oxidase. The reaction catalyzed by catalase generates oxygen (II),
but when both reactions (I and II) are combined they result in a net
consumption of oxygen (III):
H2 O2 → H2 O + (1/2)O2
HOOC COO− + H+ + (1/2)O2 → 2CO2 + H2 O
To our knowledge this is the first study of oxygen-scavenging
films based on oxalate oxidase and co-immobilization of catalase
to prevent release of hydrogen peroxide.
2. Materials and methods
2.1. Preparation of free films
Oxalate oxidase from barley seedlings (0.71 U/mg; one unit (U)
forms one ␮mol of hydrogen peroxide per min at pH 3.8 and 37 ◦ C)
(Sigma–Aldrich, Steinheim, Germany) was mixed with HPU70 latex
(styrene butadiene co-polymer; Tg 6 ◦ C; dry solids content 50.6%)
(Styron Europe GmbH, Horgen, Switzerland), hereafter denoted as
SB-latex, in a ratio of 416 g dry latex/g enzyme preparation. The
mixture was incubated for 20 min at 4 ◦ C with stirring to ensure
homogeneous dispersion. Oxalic acid was added to a portion of the
enzyme/latex dispersion to evaluate the need for having oxalic acid,
one of the substrates of the enzyme, present directly in the film. For
this experiment, 2.033 g of the enzyme/latex dispersion was mixed
with oxalic acid to a final concentration of 20 mM. The pH of the
dispersion without oxalic acid was 5.5, while the pH was 4.9 in the
dispersion containing oxalic acid. The mixture was left to stir for
an additional 15 min at 4 ◦ C. The two dispersions with and with-
out oxalic acid were coated onto the backside of silicone-treated
release papers using a wire-wound bar (K202 Control Coater, RK
Print Coat Instruments Ltd., Royston, UK) resulting in a nominal
wet deposit of 60 ␮m. Unless otherwise indicated, the drying proce-
dure [9] consisted of an incubation for 30 s at 105 ◦ C in a ventilated
oven followed by an incubation for 24 h at 30 ◦ C and 50% relative
humidity (RH) to ensure complete film formation. When substrate
was present the extended drying was performed under nitrogen
atmosphere to prevent pre-oxidation of the substrate. In order to
evaluate the influence of the drying conditions on the enzyme activ-
ity, free films without oxalic acid were prepared also by (i) drying
for 24 h at 30 ◦ C and 50% RH, (ii) drying for 30 s at 75 ◦ C followed by
incubation for 24 h at 30 ◦ C and 50% RH, and (iii) drying for 30 s at
95 ◦ C followed by incubation for 24 h at 30 ◦ C and 50% RH.
As a negative control, SB-latex without enzyme was coated onto
the backside of a release paper as described above. The drying pro-
cedure for the negative control was 30 s at 105 ◦ C followed by 24 h
at 30 ◦ C and 50% RH.
The films were stored at 4 ◦ C until further analyzed. Prior to
analysis, the films were peeled off from the release paper.
2.2. Enzyme activity measurements using oxygen electrode
The specific activity of the enzyme-containing films was eval-
uated using an oxygen electrode (Oxygraph system, Hansatech
97
Reference
Nestorson et al. [8]
Johansson et al. [28]
Labuza and Breene [29]
Instruments, Kings Lynn, UK). A small piece of film was weighed
and added to the reaction chamber that contained 3 ml pre-heated
(25 ◦ C) reaction mixture. The reaction mixture for the films without
substrate contained 2 mM oxalic acid pH 3.8 (the pH was adjusted
to 3.8 with a solution of sodium hydroxide) and 20 mM succinic
acid buffer (pH 3.8). The reaction mixture for the films with oxalic
acid contained only 20 mM succinic acid buffer (pH 3.8). The weight
of the films with oxalic acid corresponded to a maximum the-
(2)
oretical concentration of oxalic acid in the reaction chamber of
(3) 2 mM. The loss of activity due to immobilization was determined
by comparing the specific activity of the enzyme before and after
immobilization.
All measurements were performed in triplicates and the
degradation of oxalic acid was measured during 10 min. The
specific activity was calculated as ␮mol O2 min−1 mg enzyme
preparation−1 .
2.3. Film stability
The film prepared as negative control was used to investigate the
stability of the latex films at higher temperatures when incubated
in aqueous solutions. Three pieces of film were peeled off from the
release paper and weighed. Two pieces of films were incubated,
each separately, in 3 ml of deionized water; one at 65 ◦ C and the
other one at room temperature. The third piece was used as a con-
trol and was not incubated in water. The films were subsequently
allowed to dry during 24 h at 30 ◦ C and 50% RH. The weight of the
films was determined before and after the treatment.
2.4. Migration of the enzyme
A film without substrate was used to evaluate if the enzyme
migrates out from a film incubated in an aqueous solution for a
short period of time. A small piece of free film was weighed prior
to incubation for 10 min in 3 ml 20 mM succinic acid buffer (pH
3.8). The film was thereafter removed and the 3 ml succinic acid
buffer solution was transferred to the oxygen electrode chamber.
A volume of 12 ␮l 0.5 M oxalic acid pH 3.8 (the pH was adjusted to
3.8 with a solution of sodium hydroxide) was added to the oxygen
electrode chamber resulting in an initial oxalic acid concentration
of 2 mM. The concentration of oxygen was then monitored during
10 min. As a positive control, the activity of the same amount of
film without substrate placed in the reaction chamber was mea-
sured according to the section “Enzyme activity measurements
using oxygen electrode”.
2.5. Storage of enzyme-containing films
The storage stability of enzyme-containing films without sub-
strate was studied by measuring the activity (as described in
Section 2.2) after 1, 6 and 14 days of storage at 4 ◦ C in air atmo-
sphere.
2.6. The effect of latex on enzyme activity
The potential inhibitory effect of SB-latex on the activity of
oxalate oxidase was measured using the oxygen electrode. In all
experiments, the reaction mixture contained 20 mM succinic acid,
98 S. Winestrand et al. / Biochemical Engineering Journal 72 (2013) 96–101

2 mM oxalic acid, and 0.1 mg/ml of the oxalate oxidase prepara-

tion. A portion of the SB-latex was diluted to 27% solids content
and the serum phase was obtained by centrifugation according to
the method described by Krishnan et al. [10]. The enzymatic activ-
ity was measured in the presence of SB-latex (with solids content
50% or 27%) and in the serum phase of the SB-latex (27% solids
content before the centrifugation). Of the total reaction volume
(3 ml), 200 ␮l was either any of the two SB-latex dispersions or the
serum phase solution. As a positive control, the enzyme activity was
measured in the buffered aqueous solution without any SB-latex or
serum phase present.

2.7. Co-immobilization of oxalate oxidase and catalase

The effect of co-immobilizing oxalate oxidase and catalase was

investigated by preparing three sets of films containing (i) SB-latex,
(ii) SB-latex and oxalate oxidase [in a ratio of 416 g dry latex/g
oxalate oxidase preparation (from barley, Sigma–Aldrich)], and (iii)
SB-latex, oxalate oxidase [416 g dry latex/g oxalate oxidase prepa-
ration], and catalase [Catazyme from Novozymes, applied in a ratio
of 50 U catalase/U oxalate oxidase (where 1 U of catalase degrades
1 ␮mol hydrogen peroxide per min at 23 ◦ C)]. The mixtures were
incubated for 20 min at 4 ◦ C with stirring to ensure homogeneous
dispersion. The coating and drying procedure was performed as
described above, except that in this case the mixtures were coated
onto the backside of Skultuna fridge and freezer plastic reinforced
aluminum foil (Cuki Cofresco S.p.A, Västerås, Sweden).
A spectrophotometric assay was used to evaluate the effect of
co-immobilizing catalase and oxalate oxidase. The total volume
of the assay was 5 ml and the temperature was 23 ◦ C. The assay
mixture contained the following constituents (final concentrations
indicated): 50 mM succinate buffer (the pH of a solution of sodium
succinate was adjusted to 3.8 using 5 M HCl), 1 mM ABTS (Fluka,
Buchs, Switzerland), 2 mM oxalic acid (pH adjusted to 3.8 using
5 M NaOH) and 24 U/ml peroxidase (type VI from horseradish,
Sigma–Aldrich). One piece (5 cm × 5 cm) of coated foil was added
to each reaction. The reaction time was 20 min and the absorbance
at 414 nm was measured every two min using a spectrophotometer
(UV-1800, Shimadzu, Kyoto, Japan). The reaction was started (t = 0)
by adding the coated foil to the reaction mixture. The reactions
were performed in triplicates, and the mean values with standard
deviations were calculated.
3. Results and discussion
3.1. Immobilization of oxalate oxidase in latex film
Oxalate oxidase was successfully immobilized in active form
through entrapment into SB-latex film (Fig. 1). Oxalic acid can
either be present directly in the film or, alternatively, be supplied
externally to the enzyme-containing film. The specific activity of
the film containing both oxalic acid and enzyme was determined
to 0.045 ± 0.017 ␮mol O2 min−1 mg enzyme preparation−1 and the
specific activity of the film containing enzyme but no added oxalic
acid was determined to 0.045 ± 0.008 ␮mol O2 min−1 mg enzyme
preparation−1 . As there was no significant difference between the
two approaches, subsequent experiments were performed with
films containing enzyme but no oxalic acid.
The enzyme lost activity during immobilization and the
remaining specific activity was determined to 6% of that of the ini-
tial. This result can be compared with other reports on attempts
to immobilize oxalate oxidase. When oxalate oxidase was immobi-
lized using glutaraldehyde on a PDMS silicone elastomer, 47.5% of
the activity was retained [11]. Immobilization of oxalate oxidase on
a PVC sheet, also by using glutaraldehyde, resulted in 65% remaining
Fig. 1. Immobilization of oxalate oxidase in latex film. The immobilized enzyme is
functional either with or without its substrate oxalic acid as a component of the
film. Error bars indicate standard deviation.
activity. When comparing with glucose oxidase entrapped in silica
gel, the remaining activity varied from 3.14% to 66.3% depending on
the ageing and drying procedure [12]. However, it is important to
note that these attempts relied on fundamentally different immobi-
lization methods and that there was no high-temperature drying at
105 ◦ C, as in our study. Using procedures that were more similar to
the ones used in the present study, glucose oxidase was entrapped
in latex with 6% remaining activity [8]. Laccases from Trametes
versicolor, Myceliophthora thermophila and Rhus vernicifera were
immobilized by entrapment in a latex/clay matrix with a remaining
activity between 18 and 53% [13].
Since experimental conditions vary between studies, it is gen-
erally difficult to say whether differences in remaining activity
between different studies depend on the enzymes or on the exper-
imental conditions chosen. However, the differences observed for
the three laccases in the same study [13] suggest that the proper-
ties of the enzymes are of importance for the fraction of activity
retained.
3.2. The effect of latex on the oxalate oxidase activity
The SB-latex partially inhibited the enzyme activity, as can be
seen by comparing the results of the experiments with deionized
water, 27% SB-latex, and 50% SB-latex (Fig. 2). The results of the
experiments with 27% SB-latex and serum phase (Fig. 2) show that
the serum phase gave the strongest inhibitory effect. This sug-
gests that the inhibitory effect is due to surfactants or remaining
monomers present in the serum phase of the latex.
Fig. 2. The effect of latex on the oxalate oxidase activity. Error bars indicate standard
deviation.
 S. Winestrand et al. / Biochemical Engineering Journal 72 (2013) 96–101 99

Fig. 3. Comparison of different drying conditions during immobilization of oxalate

oxidase in latex. All films were incubated for 24 h at 30 ◦ C (as in the experiment Fig. 5. The specific activity in the film after one, six and 14 days of storage at 4 ◦ C.
represented by the bar to the left), but before that some films were dried for 30 s Error bars indicate standard deviation.
at either 75 ◦ C, 95 ◦ C or 105 ◦ C (experiments represented by the other three bars).
Error bars indicate standard deviation.
solution after 10 min incubation of the film corresponds to 33%
of the activity in the control reaction. It is probably the enzyme
3.3. Evaluation of drying conditions
molecules that are positioned close to the surface or even on the
surface of the film that migrate or loosen from the film.
The results from the evaluation of different drying conditions
Immobilization techniques are often used to avoid migra-
during immobilization of oxalate oxidase in latex are shown in
tion of enzymes to food, and also to increase the stability of
Fig. 3. After drying only at 30 ◦ C, the activity in the film was deter-
enzymes towards changes in pH and temperature, the presence
mined to 0.011 ± 0.002 ␮mol min−1 mg enzyme preparation−1 .
of surfactants, and mechanical force. Some of the most common
When the incubation at 30 ◦ C was preceded by a rapid drying step
immobilization techniques are cross-linking, adsorption and cova-
at a higher temperature, the activity (given in ␮mol min−1 mg
lent attachment to carrier support, entrapment in a gel lattice,
enzyme preparation−1 ) was determined to: 75 ◦ C, 0.016 ± 0.003;
and microencapsulation [14,15]. Out of these methods covalent
95 ◦ C, 0.009 ± 0.001, and 105 ◦ C, 0.009 ± 0.001. Drying at 75 ◦ C for
attachment to a support is considered the most effective in terms
30 s thus resulted in the highest activity and seems to be the best
of thermal stability [16]. Oxalate oxidases have previously been
alternative among the conditions studied. No decrease in oxygen
immobilized using various supports, such as for example nylon
concentration was detected for the negative control. These results
tubing [17], pig intestine membrane [18], nylon and collagene
can be compared to reported results of drying of films with laccases
membranes [19], polyethylene glycol [20], glass beads [21], PDMS
from Trametes versicolor, Myceliophthora thermophila and Rhus ver-
silicone elastomer [11], a multilayer biosensor [22], PVC membrane
nicifera for 30 s at 75 ◦ C, 90 ◦ C or 105 ◦ C. When comparing the results
[23], a gold electrode [24], and egg-shell membrane [25].
from 30 s drying, the remaining activity was reported to be highest
after drying at 75 ◦ C for all three laccases [13], which agrees with
3.5. Storage of enzyme-containing films
the results from the present study of oxalate oxidase.

The possibility to store the films was investigated and the result
3.4. Stability of latex film and migration of the enzyme
does not indicate any loss of enzymatic activity after storing the
film at 4 ◦ C during a period of up to two weeks (Fig. 5). The activ-
The test of the stability of the latex films showed that there was
ity (given as ␮mol min−1 mg enzyme preparation−1 ) after storage
no sign of disintegration after 10 min of incubation either at room
was determined to: 1 day, 0.0316; 6 days, 0.0363; and 2 weeks,
temperature (23 ◦ C) or 65 ◦ C, since the remaining weight of the film
0.0385. The weak tendency, although not statistically significant,
was determined to 100.0% in all cases studied. Since entrapment
that the activity increases with time could indicate that the enzyme
represents a form of immobilization where the enzyme molecules
migrates towards the surface during storage. To investigate storage
are embedded rather than physically attached to the solid car-
for a longer period of time would be of interest in the future.
rier, the possibility that the enzyme molecules actually migrate
Pundir et al. noticed a loss of 10% of the enzymatic activity after
out from the film cannot be excluded. An experiment was con-
180 days of storage of oxalate oxidase immobilized on a chemically
ducted to address this issue, and the results suggest that the enzyme
activated egg-shell membrane using glutaraldehyde [25]. It has also
can migrate from the film into the solution (Fig. 4). The activity in
been reported that oxalate oxidase immobilized on a PVC sheet
using glutaraldehyde lost 15% of its activity when stored at 4 ◦ C
during 100 days [23]. Another enzyme, glucose oxidase, was immo-
bilized on a cellulose acetate-polymethylmethacrylate membrane
using glutaraldehyde and was reported to have a remaining activity
of 94% after 1 month of storage [26]. After entrapment in latex, glu-
cose oxidase showed no decrease in activity after 14 days of storage
at 8 ◦ C [8]. Both the immobilization technique and the storage sta-
bility result of the study of Nestorsson et al. [8] correspond well
with the result presented in this study.

3.6. Co-immobilization of oxalate oxidase and catalase

The results from the experiment on co-immobilization of

Fig. 4. Migration of oxalate oxidase. Error bars indicate standard deviation. oxalate oxidase and catalase are presented in Fig. 6. The increase
100 S. Winestrand et al. / Biochemical Engineering Journal 72 (2013) 96–101
Fig. 6. Co-immobilization of oxalate oxidase and catalase. A coupled assay with
horseradish peroxidase and ABTS was used to detect hydrogen peroxide production
by films containing oxalate oxidase, a combination of oxalate oxidase and catalase,
and a control film without enzymes. Error bars indicate standard deviations.
in absorbance observed for the film containing oxalate oxidase but
no catalase (Fig. 6) indicates that hydrogen peroxide was produced
by oxalate oxidase and was then consumed by horseradish peroxi-
dase, which catalyzed the oxidation of ABTS resulting in an increase
in absorbance at 414 nm. For the film containing both oxalate oxi-
dase and catalase, the hydrogen peroxide formed by oxalate oxidase
was successfully degraded by catalase as indicated by the absence
of an increase in absorbance, which was also noticed for the con-
trol film without enzymes (Fig. 6). The results indicate that oxalate
oxidase and catalase can be successfully co-immobilized in active
form, and that the inclusion of catalase efficiently prevents release
of hydrogen peroxide from the film.
As hydrogen peroxide is potentially harmful, it would be crucial
to prevent its release in an active-packaging system. Furthermore,
too high concentrations of hydrogen peroxide may inhibit the
enzyme used for oxygen scavenging [27]. Among enzymes stud-
ied for scavenging of oxygen, glucose oxidase is the one that has
so far been investigated most thoroughly. Although the use of
catalase to prevent the release of hydrogen peroxide has previ-
ously been discussed as a possibility, this is to our knowledge the
first time that this approach has been investigated experimentally
after co-immobilization of catalase in films together with another
enzyme.
3.7. The potential of oxalate oxidase in active packaging
Our investigation indicates that it is possible to utilize oxalate
oxidase in active packaging, for example as an oxygen scavenger,
and that catalase can be added to take care of the hydrogen
peroxide that is generated in the reaction catalyzed by oxalate oxi-
dase. Large amounts of oxalic acid are generated as a by-product
from some pulping processes, and could potentially be used as a
component of an oxygen-scavenging system based on oxalate oxi-
dase. There is an added advantage of oxalate oxidase compared to
other oxygen-scavenging enzymes that are being considered for
this purpose, such as glucose oxidase [8]. When oxygen is con-
sumed by oxalate oxidase, carbon dioxide is formed as one of
the products. This has the potential to improve modified atmo-
sphere packaging (MAP), a process used by the packaging industry.
In MAP, oxygen is replaced by other gases, such as carbon diox-
ide, which inhibits microbial growth and increases the shelf-life
of the food. By its ability to generate carbon dioxide, oxalate
oxidase therefore has a theoretical advantage over many other
oxygen-scavenging enzymes for increasing the shelf-life of food
and beverages.
4. Conclusions
The investigation shows that catalytically active oxalate oxidase
can be entrapped in latex films and that the oxalic acid can be sup-
plied either as one of the original constituents of the film or as a
later addition to the system. This opens up two alternative ways
to exploit oxalate oxidase in active packaging; either as a part of
an oxygen-scavenging system or as an agent that can be used to
remove unwanted oxalic acid from food and beverages. The inves-
tigation also shows that co-immobilization of catalase in films is
an efficient approach to degrade hydrogen peroxide produced in
a reaction catalyzed by an oxidase. In this study we have focused
on active-packaging applications using oxalate oxidase as an oxy-
gen scavenger. In the future it might also be possible to use oxalate
oxidase for intelligent-packaging applications to indicate the pres-
ence of oxygen in the headspace of a package. Further investigations
of alternative potential applications for oxalate oxidase have been
initiated in our laboratories.
Acknowledgements
This work was supported by grants from the Knowledge Foun-
dation (KK-stiftelsen), the Knut and Alice Wallenberg Foundation
and through the Bio4Energy program (www.bio4energy.se).
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