Functionally-modified egg white albumen hydrogels

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					Polymer International                                                                           Polym Int 53:1994–2000 (2004)
                                                                                                          DOI: 10.1002/pi.1611




Functionally-modified egg white
albumen hydrogels
GVN Rathna, J Li and S Gunasekaran∗
Food and Bioprocess Engineering Laboratory, University of Wisconsin-Madison, 460 Henry Mall, Madison, WI 53706, USA



    Abstract: Hydrogels were prepared by using egg white albumen (EWA) before and after chemical
    modification of its lysyl residues with ethylenediamine tetraacetic dianhydride (EDTAD) to incorporate
    carboxylic groups. This resulted in an increase in swelling ratio of the EWA hydrogels. The swelling
    ratio increased dramatically in deionized water substantially, more than in pH 7.4 buffer solution. The
    effects of medium pH, temperature and swelling were investigated, along with crosslinking of the gel
    network by glutaraldehyde (GLA), as well as acetone treatment. The gels denatured by acetone showed
    an insignificant increase in swelling ratio for the gels crosslinked with GLA during gel preparation, which
    is in contrast to the gels crosslinked subsequent to gel formation. The swelling behavior was positively
    affected by temperature and time. However, an insignificant effect of pH was observed due to electrostatic
    screening of the carboxylic groups by sodium ions in the buffer solution. Availability of various functional
    groups on EWA has resulted in adsorption of metals (Cu+2 ions) and non-metals (PO4 −2 anions).
     2004 Society of Chemical Industry

Keywords: egg white albumen; hydrogel; chemical modification; swelling ratio; chelation



INTRODUCTION                                                        contains various functional groups such as –NH2 ,
Hydrogels are three-dimensional hydrophilic networks                –COOH, –SH and –OH. Therefore, modification of
with an ability to absorb large quantities of water. A              these functional groups without altering the structural
considerable amount of research has been performed                  properties could enable us to develop a novel
on the properties of hydrogels made from synthetic and              hydrogel with improved properties. The lysyl residues
natural polymers. These investigations have focused                 (–NH2 ) of proteins can be modified with carboxylic
on the effects of composition,1 temperature,2,3 pH,2 – 5            groups by reacting with ethylenediamine tetraacetic
ionic strength, magnetic properties,6 electric fields,               dianhydride (EDTAD).8 – 10 Such a modification
added additives, etc. Many synthetic polymers have                  of the lysyl residues of proteins can improve
limited structural and functional properties, while the             the swelling properties and transparency of the
natural polymers are unique, with various functional                hydrogel formed. Therefore, the developed gels
and structural properties.7 The mechanical properties               may be suitable for several potentially ‘value-
and swelling properties of hydrogels made by using                  added’ biomedical or biotechnological applications.
natural polymers can be improved by chemical or                     Additionally, an interesting aspect of EDTA-like
physical modification of the functional groups.8 – 10                chelating groups in the modified polymers has
Modification does not alter the biodegradable and                    provided an ability to form anionic complexes which
biocompatible characteristics of the proteins. There-               show adsorption to metal cations.12,13
fore, if suitable functionalities can be imparted, the                 Our objectives were to (1) chemically modify the
natural polymers could replace some potentially toxic               lysyl residues of EWA, (2) prepare various EWA
synthetic polymers, which are unsuitable for many                   hydrogels, (3) determine the swelling behavior of
biological applications.                                            different EWA hydrogels, and (4) study the ability
   Egg white albumen (EWA) protein is widely used                   of EWA hydrogels to adsorb metal and non-metal
for various food applications.11 However, the use of                ions.
EWA for ‘value-added’ non-food applications has
received little attention. The EWA has an ability
to form a non-reversible gel at temperatures above                  EXPERIMENTAL
80 ◦ C due to covalent disulfide (S–S) bond formation.               The EWA powder was obtained from the Oskaloosa
Nonetheless, the so-formed EWA gels are opaque with                 Food Products Corporation (Oskaloosa, IA, USA),
minimum swelling ability. The EWA, being a protein,                 while EDTAD, cupric sulfate, glutaraldehyde (GLA),

∗ Correspondence to: S Gunasekaran, Food and Bioprocess Engineering Laboratory, University of Wisconsin-Madison, 460 Henry Mall,
Madison, WI 53706, USA
E-mail: guna@wisc.edu
(Received 18 July 2003; revised version received 11 February 2004; accepted 9 March 2004)
Published online 13 October 2004
 2004 Society of Chemical Industry. Polym Int 0959–8103/2004/$30.00                                                       1994
                                                                                                   Modified egg white albumen hydrogels

sodium phosphate (monobasic), trinitrobenzenesul-                          spectrophotometer (UV/Vis-1601PC, Shimadzu), at a
fonic (TNBS) acid, sodium potassium tartarate                              wavelength of 415 nm against a blank. The percentage
and dialysis membranes (molecular weight cutoff,                           of lysyl residues modified (PLRM) in the EWA was
6000–8000 g mol−1 ) were obtained from the Sigma-                          determined as follows:14
Aldrich Company (St Louis, MO, USA). All reagents                                               OD × number of dilutions
were of analytical grade. Synthesis of EDTAD-                              PLRM =
                                                                                         1.5 × 107 × number of moles of protein
modified EWA was performed according to the
method of Hwang and Damodaran.10 Samples of                                            × 100                                  (1)
100 ml of 5 % EWA solution were adjusted to pH                             where OD is the optical density of the protein solution.
12, and measured amounts of EDTAD were then
added incrementally while stirring and maintaining                         Preparation of EWA hydrogels
the solution at pH 12. After complete addition of                          The EWA and modified EWA powders were used for
the EDTAD, the stirring was continued for 3 h. The                         hydrogel preparation. Additional experimental vari-
reaction mixture was dialysed overnight and finally                         ables during gel preparation were GLA addition (for
freeze-dried to obtain the dry modified EWA product.                        crosslinking) and acetone treatment (for denatura-
The experimental steps for EWA modification and                             tion). The latter treatment can denature the protein,
hydrogel formation are illustrated in Fig 1.                               leading to conformational changes of the protein and
                                                                           thus enhance the rate and extent of swelling of the
Estimation of modified lysyl residues                                       hydrogels.8,9
The lysyl contents of the modified and unmodified                               The gels were prepared by dissolving the calculated
EWA were determined by the TNBS method in                                  amounts of protein in water (15 % (w/v)), and the
triplicates as described by Hall et al.14 According                        solutions were heated to, and held at, 80 ◦ C for
to this procedure, 0.5 ml of protein is added to                           1 h. The gels were then cooled to room temperature
1.0 ml of 4 % NaHCO3 solution, followed by the                             (∼25 ◦ C) and finally further cooled at −20 ◦ C for 1 h.
addition of 0.2 ml TNBS solution (12.5 mg ml−1 ).                             For crosslinking the gels, GLA was added either
This solution mixture was incubated at 40 ◦ C for 2 h,                     during gelation (ie to the 15 % protein solution—a
followed by the addition of 3.5 ml of concentrated                         1 % GLA solution was added before it was heated)
HCl, and further incubated at 110 ◦ C for 3 h. The                         or subsequently to gel formation (ie by immersing
solution was cooled and made up to 10 ml, extracted                        thin gel disks in 1 % GLA solutions overnight). The
twice with equal amounts of ether, from which the                          crosslinked gels were washed thoroughly and finally
aqueous solution was separated and left at 40 ◦ C to                       dried at 30 ◦ C to constant mass. Prior to drying,
remove traces of ether. The absorbency of the resulting                    some of the uncrosslinked gels and gels crosslinked
yellow solution was measured by using a UV–visible                         with GLA during gelation were also treated with
                                                                           acetone for 3 h. All gel samples were dried to constant
                                  Protein solution
                                                                           mass in a desiccator. The various hydrogels prepared
        NaOH solution
                                                                           were denoted differently for easy reference in the
                                                                           subsequent tables and figures, as listed in Table 1.
                                  PH 12, protein
                                                                           Swelling experiments
 Incubate for 30 min at 60 °C                                              The hydrogel samples were swollen in deionized water
                                                     EDTAD
                                                                           and/or in phosphate buffer solution at 37 ◦ C. Their
                                  Modified protein
                                                                           Table 1. Abbreviations used to refer to the different EWA hydrogels
               Dialysis                                                    prepared for investigation
                                                     Lyophilization
                                                                           Abbreviation       Description
                                  EDTAD modified
                                                                           AC                 Albumen control
                    Water                                                  AA                 Acetone-treated albumen
                                                                           AG                 Crosslinked albumen during gel preparation
                                  Modified protein                         ACG                Cross-linked albumen after gel preparation
                                                                           MAC                Modified-albumen control
   Incubate for 60 min at 80 °C
                                                                           MAA                Acetone-treated modified albumen
                                                                           MAG                Crosslinked modified albumen during gel
                                     Hydrogel
                                                                                                preparation
                                                                           MAAG               Acetone-treated crosslinked modified
     Dried hydrogel                                   Acetone treatment                         albumen during gel preparation
                                                                           MACG               Crosslinked modified albumen after gel
                                                                                                preparation
                                                          Dried hydrogel
                                                                           MAAC               Modified-acetone-treated-albumen control
                                                                           MAACG              Acetone-treated crosslinked modified
Figure 1. Flowchart for lysyl residue modification and preparing                                 albumen after gel preparation
different modified EWA hydrogels.


Polym Int 53:1994–2000 (2004)                                                                                                              1995
GVN Rathna, J Li, S Gunasekaran

masses were measured periodically until equilibrium                       Adsorption of phosphate anions and copper ions
was achieved (ie the mass change between two                              The hydrogels of modified and unmodified EWA were
consecutive measurements over 6 h was within                              used to determine their chelating abilities of phosphate
±0.1 g). Since we suspected a possible interaction                        anion (PO4 −2 ) and copper ion (Cu+2 ).
between the phosphate buffer and the gel, for a selected                     Stock solutions of sodium phosphate (monobasic)
condition the hydrogel samples were also swollen in                       were made in water at different concentrations of
Tris buffer solution at various pH levels at 25 ◦ C.                      5, 10, 20, 40, 80 and 160 mg l−1 . Dry pellets of
  The swelling experiments were performed in                              48 %-modified and unmodified EWA were weighed
duplicate. The swollen masses of the hydrogels were                       and immersed in the stock solutions in duplicate for
measured gravimetrically after wiping off any excess                      2 h. The amount of phosphate anion adsorbed by the
water on the surfaces with filter paper. The swelling                      swollen hydrogels was estimated by using a Lachat
ratios (SR, g g−1 of dry gel mass) were calculated by                     UV/Visible spectrophotometer operating at 880 nm
using the following equation:                                             (Zellweger Analytics, Lachat Instrument Division,
                                                                          Milwaukee, WI, USA).
                                   mt − mf                                   A limited study was carried out to investigate
                          SR =                                      (2)   the adsorption of copper by the EWA gels. Copper
                                      mf
                                                                          sulfate (CuSO4 ) solution was prepared in water at a
                                                                          concentration of 5 mg l−1 : 20 ml samples of CuSO4
where mt is the mass of swollen gel at time t and mf is                   solutions were taken in duplicate. Dry pellets of
the mass of final dried gel after swelling and drying to                   48 %-modified and unmodified EWA dry pellets
constant mass.                                                            were immersed in these solutions. The EWA pellets
  A ‘24 ’ unreplicated factorial experimental design                      were removed from the CuSO4 solutions after 0.5,
was used for evaluating the effects of PLRM, pH,                          2 and 24 h of reaction time and the amounts of
temperature and swelling time on the swelling behavior                    Cu+2 adsorbed by the hydrogels were estimated by
of modified albumen control (MAC) hydrogels in                             using a flame atomic absorption spectrophotometer.
phosphate buffer solution. The experimental variables                     A ‘standard curve’ was established by using copper
are described in Table 2. The initial factorial design                    solutions with concentrations of 1, 3 and 5 mg l−1
considering all of these effects assumes the following                    to estimate the amounts of Cu+2 adsorbed by the
model:                                                                    EWA gels.

               A         B       C         D       AB
    y=η+          x1 + x2 + x3 + x4 +                   x1 x2
                2        2        2        2        2                     RESULTS AND DISCUSSION
            AC           AD            BC           BD                    Modification of lysyl residues
        +       x1 x3 +       x1 x4 +       x2 x3 +      x2 x4
             2             2             2            2                   The extent of lysyl residue modification increased
            CD           ABC                ABD                           linearly (R2 = 0.9769) with increasing EDTAD to
        +       x3 x4 +         x1 x2 x3 +        x1 x2 x4
             2              2                 2                           EWA ratio, as shown in Fig 2. Based on the reaction
            ACD               BCD                                         conditions, EDTAD has introduced about three
        +         x1 x3 x4 +         x2 x3 x4                             carboxylic groups for each lysyl residue.10 The reaction
              2                 2
                                                                          scheme is shown in Fig 3.
            ABCD
        +           x1 x2 x3 x4 + ε                            (3)
               2
                                                                          Swelling ratios
                                                                          The swelling ratios of MAC hydrogels prepared
where y is the swelling ratio, η the grand mean, A the
                                                                          under different ‘24 ’ unreplicated factorial experimental
PLRM, B the pH, C the temperature and D the time.
Multiple parameters (AB, AC, . . ., ABCD) indicate
interaction effects. The analysis was performed by                                   120
using Yates algorithm in the ‘XLISPSTAT’ statistical
                                                                                     100
package, and the significant effects were determined by
using Loh’s method for the treatment of unreplicated                                  80
                                                                          PLRM (%)




factorial experimental data.15
                                                                                      60
                                                                                                                 y = 483.56x + 1.1191
                                                                                      40
Table 2. The unreplicated ‘24 ’ factorial design used for investigating                                             R 2 = 0.9769
the swelling behavior of EWA hydrogels
                                                                                      20
Treatment factor              Variable Low-level (−) High-level (+)
                                                                                       0
                                                                                           0   0.05     0.1    0.15         0.2         0.25
Modified lysyl residue (%)        A             0              28
pH                               B            4.6             9.7                                     EDTAD/EWA (wt/wt)
Temperature (◦ C)                C            25              37
Time (min)                       D            30              60          Figure 2. Percentage of lysyl residues modified (PLRM) in EWA as a
                                                                          function of the EDTAD/EWA ratio.


1996                                                                                                      Polym Int 53:1994–2000 (2004)
                                                                                                                            Modified egg white albumen hydrogels


                                                                                                                       CH2COO−
                                                                    at pH 12
                                       H 2N         +   EDTAD                    HN                                    N     COO−
                                                                   Room             C                             N
                                                                               NH2
                                              NH2               temperature                                       CH2COO−
                                                                                    O

                                         Egg white albumen                                              EDTAD− egg white albumen

Figure 3. Lysyl residue modification reaction scheme in EWA.


conditions are listed in Table 3. The results of the                              The swelling kinetics of uncrosslinked hydrogels
statistical analysis of these data are presented in                            (in deionized water), prepared from EWA after
Table 4. From the statistical analysis, the model for the                      different lysyl residue modifications, is presented
swelling ratio of EWA hydrogels in 0.1 M phosphate                             in Fig 4. The swelling ratios of these hydrogels
buffer solution can be written as follows:                                     increased with the PLRM. Due to the competition
                                                                               between expansion and contraction (disulfide bonds
                          A     C    D                                         and hydrogen bonds), the EWA hydrogels prepared
               y=η+         x1 + x3 + x4 + ε                         (4)
                          2     2    2                                         with a PLRM of 100 % disintegrated after 3 h, while
                                                                               those prepared by using EWA with a PLRM of 83 %
In the above model, all of the coefficients (x1 , x3                            collapsed after 5 h.
and x4 ), η (grand mean) and ε (distributed normal                                The profiles displayed in Fig 5 represent the swelling
variable) are positive, hence indicating the positive                          behavior in phosphate buffer of pH 7.4 at 37 ◦ C for
effects of the different treatments, ie PLRM (A),
temperature (C) and time (D). The positive value
                                                                               Table 4. Effects calculated by the Yates algorithm and Loh’s
of x1 associated with the PLRM clearly indicates that
                                                                               method15 (the conditions having a significant effect are indicated by
introducing carboxylic groups into EWA increases the                           ‘bold script’)
gel SR. In addition, judging from the relative values of
x1 , x3 and x4 it is clear that the effect of the PLRM on                      Effect                                          Value                        Significant?
the SR is about twice that of the effects of temperature
                                                                               η                                               15.08                            No
and time. The positive effect of temperature on the                            A                                               18.14                            Yes
SR of the hydrogel may be related to an increase                               B                                                0.11                            No
in the degree of expansion. When the levels of all                             AB                                               1.95                            No
three factors are high (ie modification ratio, 28.5 %,                          C                                                8.12                            Yes
at 37 ◦ C over 60 min), a maximum SR of EWA will be                            AC                                               3.48                            No
expected. It is worth noting that the results indicate an                      BC                                              −0.49                            No
insignificant effect of pH (see Table 4), which may be                          ABC                                              0.19                            No
due to electrostatic screening of the carboxyl groups                          D                                                7.27                            Yes
by sodium ions in the buffer solution.                                         AD                                               1.40                            No
                                                                               BD                                              −0.41                            No
                                                                               ABD                                              0.47                            No
Table 3. Swelling ratio values obtained by the unreplicated ‘24 ’              CD                                               2.24                            No
experimental design                                                            ACD                                              0.19                            No
                                                                               BCD                                              0.41                            No
                                   Treatment factora                           ABCD                                             0.22                            No
                                                              Swelling
Number        Run order        A       B        C       D     ratio (%)

1                   9          −       −        −       −        1.83                                   180
2                   5          +       −        −       −       13.77                                                                                  Unmodified
                                                                                                        160
3                   1          −       +        −       −        1.73                                                                                  36%
4                  11          +       +        −       −       16.69                                   140                                            56%
                                                                               Swelling ratio (wt/wt)




                                                                                                                                                       83%
5                  15          −       −        +       −        5.29                                   120                                            100%
6                  12          +       −        +       −       23.87
                                                                                                        100
7                   6          −       +        +       −        3.44
8                   2          +       +        +       −       24.93                                    80
9                   7          −       −        −       +        6.70                                    60
10                 10          +       −        −       +       20.58
                                                                                                         40
11                  8          −       +        −       +        4.49
12                  3          +       +        −       +       22.36                                    20
13                 16          −       −        +       +       13.89                                     0
14                 13          +       −        +       +       34.29                                         0        2            4              6       8          10
15                  4          −       +        +       +       10.67                                                                   Time (h)
16                 14          +       +        +       +       36.73
                                                                               Figure 4. Swelling behavior of hydrogels made from unmodified EWA
aA, percentage-modified lysyl residue; B, pH; C, temperature; D, time:          and EWA with different extents of percentage of lysyl residues
‘+’ indicates ‘high-level’ and ‘−’ indicates ‘low-level’ (see Table 2).        modified (PLRM) in deionized water as a function of time.


Polym Int 53:1994–2000 (2004)                                                                                                                                       1997
GVN Rathna, J Li, S Gunasekaran

                          3.5                                                                                   14

                          3.0                                                                                   12




                                                                                      Swelling ratio (wt/wt)
                                                                                                                10
 Swelling ratio (wt/wt)




                          2.5
                                                                                                                 8
                          2.0
                                                                                                                 6
                          1.5                                                                                    4                                                         MAC
                                                                                                                                                                           MAA
                          1.0                                                                                    2                                                         MACG
                                                                              AC                                                                                           MAAG
                                                                                                                                                                           MAG
                                                                              AA                                 0
                          0.5                                                 AG                                     0   20            40         60    80     100   120     140
                                                                              ACG
                                                                                                                                                    Time (h)
                          0.0
                                0   20      40     60        80   100   120     140
                                                                                      Figure 7. Swelling behavior of 48 %-modified EWA hydrogels in a pH
                                                    Time (h)
                                                                                      7.4 buffer solution (see Table 1 for legend description).

Figure 5. Swelling behavior for unmodified EWA hydrogels (see
Table 1 for legend description).
                                                                                                                20
                                                                                                                18
unmodified EWA hydrogels with or without GLA
                                                                                                                16
crosslinking, during and after gel preparation, and

                                                                                       Swelling ratio (wt/wt)
with acetone treatment. For a given PLRM, the SR is                                                             14
higher in deionized water than in the buffer solution                                                           12
by several orders of magnitude. An increase in SR                                                               10
for the acetone-treated EWA hydrogels was expected,
as per earlier reports on other proteins.8 However, in                                                           8
                                                                                                                                                                      MAC
                                                                                                                                                                      MAAC
this case the acetone treatment did not improve the                                                              6                                                    MACG
SR, possibly due to the effect of temperature (80 ◦ C)                                                           4
                                                                                                                                                                      MAACG
                                                                                                                                                                      MAG
at which the gels were prepared. At this temperature,                                                                                                                 MAAG
                                                                                                                 2
the EWA is completely denatured, hence countering                                                                    0        5                    10        15       20
any positive effects of the acetone treatment. The SR                                                                                               Time (h)
decreased when the gels were crosslinked with GLA
after the gels were formed. However, the SR increased                                 Figure 8. Swelling behavior of 63 %-modified EWA hydrogels in a
                                                                                      pH7.4 buffer solution (see Table 1 for legend description).
when GLA was added during gelation. The higher
swelling for gels crosslinked during gelation may be
due to the hydrolysis of EWA in the presence of GLA at                                   Figures 7 and 8 present the swelling behavior for
80 ◦ C. The scheme for this hydrolysis is given in Fig 6.                             modified EWA, with and without cross-linking, and
   The aldimine bond (–HC=N–) is unstable at high                                     after acetone treatment in a pH 7.4 buffer at
temperatures, and tends to hydrolyse. Free EWA                                        37 ◦ C, respectively, for PLRMs of 48 and 63 %.
chains are formed after hydrolysing, and the crosslink-                               When comparing Figs 5, 7 and 8, it is clear that
ing density is decreased, and, therefore, the swelling                                the lysyl-residue modification of the EWA resulted
of EWA is increased.                                                                  in an increase in swelling by several orders of


                                                                        C HO                                                  H               N
                                                                                                                              C
                                                 NH2 +

                                                                        C HO                                                  H               N
                                                                                                                              C

                                         Egg white albumen          Glutaraldehyde
                                                                                                                              Hydrolysis




                                                                                                                                           80 °C




                                                                                                                                  C HO H2N


                                                                                                                                  C HO H2N
                                                                                                                                            Free albumen chains

Figure 6. Scheme of the proposed hydrolysis of the EWA hydrogel prepared with GLA addition during gelation.


1998                                                                                                                                               Polym Int 53:1994–2000 (2004)
                                                                                                                    Modified egg white albumen hydrogels

magnitude, regardless of the gel preparation protocol,                                           100
when compared to their unmodified-EWA-hydrogel                                                          Deionized water
                                                                                                 90
counterparts. The increase in swelling is due to an                                                    phosphate buffer solution (pH 7.4)
increase in number of carboxylic groups, which do                                                80    Tris buffer solution (pH 7.2)
not participate in the crosslinking step. In addition,
                                                                                                 70




                                                                          Swelling ratio (w/w)
as stated previously, the higher the PLRM, then
the higher the SR, regardless of the gel preparation                                             60
protocol, because at a higher PLRM the crosslink                                                 50
density is lower. Figure 9 illustrates the swelling
behavior for 36 %-modified EWA hydrogels and                                                      40
unmodified EWA hydrogels in Tris buffer solutions.                                                30
There is no significant difference for the unmodified
EWA hydrogels in the pH range 7.0–8.9, while                                                     20
swelling of the modified EWA hydrogels increased                                                  10
when the pH increased in the Tris buffer solution.
Figure 10 clearly illustrates these effects for MAC gels,                                         0
                                                                                                         36%               48%              56%        63%
both in deionized water and in pH 7.4 buffer. There
                                                                                                                                  PMRL
is no significant difference in swelling between the
Tris buffer solution (pH 7.2) and the phosphate buffer                    Figure 10. Comparison of the swelling ratios of EWA hydrogels as a
solution (pH 7.4), while the swelling in deionized                        function of percentage of lysyl residues modified (PLRM), in deionized
water is much higher than those in buffer solutions.                      water, in pH 7.4 buffer solution and in Tris buffer solution (pH 7.2)
                                                                          (bars represent standard deviations).

Adsorption of phosphorous anions and copper
                                                                          Table 5. Phosphorous adsorption over 2 h by hydrogels made with
ions                                                                      63 %-lysyl-residue-modified egg white albumen
Table 5 shows the amounts of phosphorous anion
adsorbed onto the modified and unmodified EWA                               Initial                                           Modified               Unmodified EWA
hydrogels. The adsorption of phosphorous anion is                         concentration (mg l−1 )                         EWA (mg g−1 )              (mg g−1 )
not affected by lysyl-residue-modification of EWA.
                                                                          5                                                      4.5                   4.08
This is because, apart from amino groups, there are                       10                                                     9.00                  9.47
other positive sites, such as hydroxyl (–OH) and                          20                                                    16.79                 15.36
–SH groups, which are involved in adsorption of the                       40                                                    18.93                 21.18
phosphorous anions. Hence, an increase in the PLRM                        80                                                    32.97                 34.92
has no effect on the phosphorous anion adsorption.                        160                                                   69.94                 70.8
Theoretically, there should not be any difference in
the phosphorous anion adsorption ability between
the modified EWA hydrogel and the unmodified                                5 to 160 mg l−1 of sodium phosphate resulted in
EWA hydrogel, because the modifying protein is                            an increase in phosphorous adsorption from 4 to
actually adding more carboxylic groups into the                           70 mg g−1 of dry hydrogel for both modified and
protein. (PO4 −2 is an anion and so is the carboxylic                     unmodified EWA proteins. It seems that the gels have
group (COO− )). An increase in concentration from                         not reached the maximum extent of possible PO4 −2
                                                                          absorption but only that of ‘slowly saturating’ available
                                                                          sites—the binding was 80 %, equivalent to a solution
                         25
                                                                          concentration of 5 mg l−1 , compared to about 45 % at
                                                                          a concentration of 160 mg l−1 .
                         20                                                  The results obtained from the limited studies on
                                                                pH 7.2
                                                                          copper ion adsorption indicated that 63 %-PLRM
Swelling ratio (wt/wt)




                                                                pH 8.0    gels adsorbed 2.25 mg g−1 , compared to 1.87 mg g−1
                         15
                                                                pH 8.9    of the unmodified gel. This increase, due to EWA
                                                                pH 7.2    modification, is attributed to an increase in carboxylic
                         10                                     pH 8.0    groups, which are the negative sites for adsorption of
                                                                pH 8.9    positively charged copper (Cu+2 ) ions.
                          5

                                                                          CONCLUSIONS
                          0                                               The swelling ratios (SR) of gels made from egg white
                              0   5   10     15       20   25        30
                                                                          albumen (EWA) can be increased by modifying its
                                           Time (h)
                                                                          lysyl residues and incorporating carboxylic groups. An
Figure 9. Swelling behavior of 36 %-modified EWA hydrogels
                                                                          increase in the extent of lysyl residue modification
(closed symbols) and unmodified EWA hydrogels (open symbols) in            resulted in an increase in the SR, both in deionized
Tris buffer solution (at pH 7.2).                                         water and in pH 7.4 buffer solutions. In addition,

Polym Int 53:1994–2000 (2004)                                                                                                                                 1999
GVN Rathna, J Li, S Gunasekaran

the temperature (25 ◦ C versus 37 ◦ C) also had a                        5 Wang T, Turhan M and Gunasekaran S, Selected properties of
positive effect on the SR. For a given amount of                             a pH-sensitive, biodegradable chitosan–PVA hydrogel. Polym
                                                                             Int 53:911 (2004).
modification, the SR is higher in deionized water                         6 Xulu PM, Filipcsei G and Zrinyi M, Preparation and responsive
than in the buffer solutions by several orders of                            properties of magnetically soft poly(N-isopropylacrylamide)
magnitude. The SR decreased when the gels were                               gels. Macromolecules 33:1716 (2000).
crosslinked with glutaraldehyde after the gels were                      7 Amiya T and Tanaka T, Phase transitions in crosslinked gels of
formed, but increased when glutaraldehyde was added                          natural polymers. Macromolecules 20:1162 (1987).
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2000                                                                                                  Polym Int 53:1994–2000 (2004)

				
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Description: Egg per day is helpful for weight loss, because the egg white in the development of amino acids to help muscle tissue, thereby enhancing the body's metabolism function. Add a little time to eat egg white cheese, not only add flavor. And more conducive to weight loss.