Hydrogen peroxide alteration of whey proteins in whey and concentrated whey systems


Cooney, C.M.; Morr, C.V.

Journal of Dairy Science 55(5): 567-573

1972


The extent of denaturation and physical aggregation of individual and total whey proteins, produced by treating whey and concentrated whey systems with H2O2 under experimental conditions that simulate pressure membrane and gel filtration processes for preparing an undenatured whey protein concentrate, was studied by preparative ultracentrifugation, Sephadex G-150 gel filtration chromatography and zone electrophoresis. Proteose-peptones were most susceptible, immunoglobulins, beta -lactoglobulin and bovine serum albumin were intermediately susceptible, and a-lactalbumin was least susceptible to alteration by H2O2v Variations in pH of the reaction mixture had only a minor influence upon whey protein alteration, whereas all of the other variables, i. e., peroxide concn., temp., and reaction time, greatly affected the extent of total and individual whey protein alteration produced by the peroxide-catalase treatment.

RESEARCH
PAPERS
Hydrogen
Peroxide
Alteration
of
Whey
Proteins
in
Whey
and
Concentrated
Whey
Systems'
C.
M.
COONEY
and
C.
V.
MORR
Department
of
Food
Science
and
Industries
University
of
Minnesota,
St.
Paul
55101
Abstract
The
extent
of
denaturation
and
physical
aggregation
of
individual
and
total
whey
proteins,
produced
by
treating
whey
and
concentrated
whey
systems
with
hydro-
gen
peroxide
under
experimental
conditions
that
simulate
pressure
membrane
and
gel
filtration
processes
for
preparing
an
un-
denatured
whey
protein
concentrate,
was
studied
by
preparative
ultracentrifugation,
Sephadex
G-150
gel
filtration
chromatog-
raphy
and
zonal
electrophoresis.
Proteose
peptones
were
most
susceptible;
immuno-
globulins,
,8-lactoglobulin
and
bovine
serum
albumin
were
intermediately
susceptible;
and
a-lactalbumin
was
least
susceptible
to
alteration
by
hydrogen
peroxide.
Variations
in
pH
of
the
reaction
mixture
had
only
a
minor
influence
upon
whey
protein
altera-
tion,
whereas,
all
of
the
other
variables,
i.e.,
peroxide
concentration,
temperature,
and
reaction
time,
greatly
affected
the
extent
of
total
and
individual
whey
protein
alteration
produced
by
the
peroxide-catalase
treat-
ment.
Introduction
Considerable
effort
is
being
expended
within
the
food
industry
to
develop
processes
for
recovering
undenatured
proteins
from
cheese
whey
(11).
Such
processes
are
needed
to
alleviate
pollution
and
disposal
problems
and
to
recover
these
nutritional
and
functional
proteins
for
human
use.
Several
processes
which
are
receiving
much
attention
for
preparing
an
undenatured
whey
protein
concentrate
are
re-
verse
osmosis,
ultrafiltration
and
gel
filtration.
However,
these
processes
involve
treating
whey
at
ambient
temperature
or
above
for
extended
times
during
which
microbial
contamination
and
growth
is
a
potential
hazard
(5).
The
extreme
susceptibility
of
whey
proteins
to
heat
de-
naturation
is
a
serious
obstacle
to
controlling
Received
for
publication
November
27,
1971.
1
Scientific
Journal
Series
Paper
7823,
Min-
nesota
Agricultural
Experiment
Station.
this
problem
by
heating
processes.
Hydrogen
peroxide
offers
considerable
promise
as
an
alternative
antimicrobial
agent
in
milk
systems
(12).
Preliminary
studies
in
our
laboratory
con-
firmed
that
hydrogen
peroxide
used
in
suf-
ficient
amounts,
produced
gross
aggregation
of
up
to
50%
of
the
total
whey
proteins
(3).
The
present
study
was
to
determine
the
effect
of
hydrogen
peroxide
upon
the
denaturation
and
aggregation
of
whey
proteins
under
conditions
that
simulate
pressure
membrane
and
gel
filtra-
tion
processes
for
preparing
whey
protein
concentrate.
The
findings,
which
represent
an
extension
of
those
of
others
(4,
6)
dealing
with
the
major
changes
in
milk
proteins
by
treating
skimmilk
with
hydrogen
peroxide,
should
be
useful
in
selecting
optimum
con-
ditions
for
preventing
microbial
growth
without
substantial
protein
alteration
during
the
prep-
aration
of
whey
protein
concentrate
by
pressure
membrane
and
gel
filtration
processes.
Experimental
Procedure
Whey
preparation
and
concentration.
The
experimental
scheme
to
prepare
and
handle
whey
is
presented
in
Figure
1.
pH
4.6
acidified
skimmilk
was
centrifuged
at
1,000
X
g
to
remove
lipid
and
fine
casein
particles
and
the
resulting
whey
was
concentrated
by
the
ultra-
filtration
apparatus
of
Tessier
and
Rose
(14)
using
a
pressure
of
1
atm
or
by
a
rotary-film
evaporator
with
a
bath
temperature
of
40
C.
For
the
latter
procedure,
whey
was
dialyzed
for
24
to
48
hr
against
two
changes
of
20
volumes
of
0.1
az
NaC1
solution
to
remove
lactose
and
milk
salts
before
being
concentrated
by
the
rotary-film
evaporator,
and
the
concentrate
was
redialyzed
for
24
to
48
hr
against
two
changes
of
20
volumes
of
pH
4.6
whey
at
0
to
5
C
to
return
the
proteins
to
their
normal
whey
envi-
ronment.
This
treatment
prevented
precipita-
tion
of
phosphate
salts
and
crystallization
of
lactose
during
the
5:1
and
10
:1
concentration
of
whey
and
was
designed
to
simulate
the
whey
system
composition
by
the
ultrafiltration
process.
Calcium-free
whey
was
prepared
by
dialyzing
p11-4.6
whey
against
Ca-free
simulated
milk
ultrafiltrate
(8)
in
which
the
calcium
and
567
568
COONEY
AND
MORR
SKIMMILK
pH
4.6
WITH
N
HCI
AT
35-40
C
WHEY
pH
ADJUSTED
TO
4.6,
5.6,
or
6.6
WITH
N
NaOH
CONCENTRATION
I
UP
TO
10:1
BY
ULTRAFILTRATION
HYDROGEN
PEROXIDE
TREATMENT:
OR
ROTARY—FILM
EVAPORATOR
H
2
0
2
:
0
to
2%
(WM
TEMPERATURE:
25
or
50
C
TIME:
4
to
24
HOURS
Xs
CATALASE
TO
REMOVE
RESIDUAL
H2
0
2
DENATURATION
AGGREGATION
READJUST
pH
TO
4.6
WITH
pH
NOT
READJUSTED
N
HC1
TO
4.6
CENTRIFUGE
206,000
x
g
1
HOUR
SUPERNATANT
EXAMINE
FOR:
TOTAL
NITROGEN
BY
MICROKJELDAHL
POLYACRYLAMIDE
GEL
ELECTROPHORESIS
SEPHADEX
G-150
GEL
FILTRATION
FIG.
1.
Experimental
procedure
for
preparing
and
treating
whey
and
for
tein
denaturation
and
aggregation.
determining
whey
pro-
magnesium
were
replaced
by
equivalent
amounts
of
sodium
and
potassium.
N-Ethylmaleimide
2
was
dissolved
in
whey
and
Cs-free
whey
systems
to
block
sulfhydryl
and
potential
sulfhydryl
groups
prior
to
the
peroxide
reaction.
Hydrogen
peroxide
treatment.
Hydrogen
peroxide,
30%
(w/w),
3
was
added
to
whey
to
a
final
concentration
of
up
to
2%
(w/w).
After
completion
of
the
treatment,
residual
peroxide
was
removed
by
adding
an
excess
of
catalase
enzyme.
4
Whey
protein
alteration.
Whey
systems
were
centrifuged
at
206,000
X
g
for
1
hr
to
sediment
aggregated
and
denatured
whey
proteins.
Pro-
tein
aggregation
was
determined
with
the
pH
5.6
and
6.6
wheys
and
denaturation
was
determined
with
wheys
adjusted
to
pH
4.6
(9).
Protein
sedimentation
was
determined
by
comparing
the
protein
concentration
(micro-Kjeldahl
N)
for
2
Purchased
from
Calbiochem.
3
Purchased
from
Merck
and
Company.
4
Crude
beef
liver
catalase
purchased
from
Nutritional
Biochemicals
Corp.
JOURNAL
OF
DAIRY
SCIENCE
VOL.
55,
No.
5
the
resulting
supernatant
fractions
with
that
of
the
total
whey.
Aliquots
of
the
treated
whey
ultracentrifugate
supernatant
fractions
were
also
examined
for
protein
alteration
by
Sepha-
dex
G-150
gel
filtration
using
2.5-
x
30-cm
columns
eluted
with
pH
6.98
phosphate
buffer
(9)
and
by
vertical
polyacrylamide
gel
electro-
phoresis
(9).
The
peroxide-denatured
whey
proteins
recovered
in
the
ultracentrifugate
sediment
fractions
were
examined
by
urea
starch
gel
electrophoresis
in
the
presence
of
2-
mercaptoethanol
(10).
Results
The
effect
of
varying
peroxide
concentration,
time,
temperature,
and
pH
upon
the
extent
of
individual
whey
protein
denaturation
in
whey
is
shown
in
Figure
2.
Proteose-peptones
were
most
susceptible
to
peroxide
denaturation,
i.e.,
they
were
essentially
removed
under
the
mildest
condition
of
1%
peroxide
at
25
C
(Fig.
2
g).
Additional
experiments
not
shown
demonstrated
that
proteose
peptones
were
completely
de-
natured
by
treating
whey
with
as
low
as
0.1%
HYDROGEN
PEROXIDE
AND
PROTEINS
569
abcdef
g
FIG.
2.
Effect
of
peroxide
concentration,
time,
temperature
and
pH
upon
whey
protein
denatura-
tion
in
whey
shown
by
polyaerylamide
gel
elm-
trophoresis.
pH
of
the
reaction
was
adjusted
to
4.6
before
ultracentrifugation
and
electrophoresis.
Volume
added to
all
slots,
50
µliters.
Electro-
phoresis
conditions:
8%
polyacrylamide
gel;
pH
8.6
veronal
buffer
(F
=
.02)
in
the
gel
and
cell;
and
constant
voltage
of
180
v
for
3
to
4
hr
(9).
Slot
Per-
oxide
Time
Temp
Reac-
tion
pH
(%
w/w)
(hr)
(C)
a
0
4.6
b
1
4
50
6.6
c
1
4
50
5.6
d
1
4
50
4.6
e
1
6
50
4.6
f
2
6
50
4.6
g
1
24
25
4.6
peroxide
at
25
C.
This
high
degree
of
lability
towards
peroxide
denaturation
of
the
proteose
peptones
is
in
direct
contrast
to
their
high
degree
of
stability
to
heat
denaturation
(9).
The
immunoglobulins
(I
g
)
were
second
in
sus-
ceptibility
to
peroxide
denaturation.
For
ex-
ample,
1%
peroxide
at
50
C
for
4
hr
denatured
substantial
amounts
of
the
immu-
noglobulins
and
the
extent
of
denaturation
was
progressively
greater
at
the
lower
pH
values
(Fig.
2
h-d).
Bovine
serum
albumin
is
also
more
susceptible
to
peroxide
denaturation
at
pH
4.6
than
at
pH
5.6
and
6.6
(Fig.
2
h-d).
On
the
other
hand,
p-lactoglobulin
(i3-Lg)
exhibited
a
greater
extent
of
peroxide
denaturation
at
pH
6.6
and
5.6
than
at
pH
4.6.
a-Lactalbumin
(a-La)
was
least
susceptible
to
peroxide
denaturation
and
was
not
totally
denatured
at
the
most
drastic
conditions,
i.e.,
2%
peroxide
for
6
hr
(Fig.
2
f).
a-Lactalbumin
denaturation
was
ir-
regular
with
respect
to
pH,
i.e.,
it
was
more
susceptible
to
peroxide
denaturation
at
5.6
than
at
pH
4.6
or
6.6.
More
peroxide
de-
naturation
of
a-lactalbumin
and
$-lactoglobulin
was
produced
at
6
than
at
4
hr
(Fig.
2
d
and
e).
Increasing
peroxide
to
2%
resulted
in
denaturation
of
all
of
the
whey
proteins
except
a-lactalbumin
and
bovine
serum
albumin.
The
varying
effect
of
temperature
and
pH
upon
the
individual
whey
proteins
suggests
that
each
protein
has
a
different
peroxide
denaturation
mechanism.
Urea
starch
gel
electrophoresis
data
(Fig.
3)
confirm
that
maximum
protein
de-
naturation
was
produced
at
50
C
(condition
d)
and
that
the
denatured
proteins
were
only
FIG.
3.
Urea
starch
gel
electrophoresis
patterns
of
peroxide-denatured
whey
proteins.
Reaction
conditions:
time,
16
hr;
pH
of
the
reaction
was
adjusted
to
4.6
before
ultracentrifugation
and
electrophoresis.
Volume
added
to
all
slots,
30
to
35
µliters.
Electrophoretie
conditions:
11.4%
starch
gel
in
pH
8.6
Tris-citrate
buffer
(.076
30
containing
7
as
urea
and
0.1%
(w/w)
2-mercapto-
ethanol;
pH
8.6
borate
buffer
(0.3
xt)
in
electrode
compartments;
16
hr
at
a
constant
current
of
30
mamp
at
0
to
5
C
(10).
Slot
Per-
oxide
Temp
Reac-
tion
pH
(%
w/w)
(C)
0
4.6
b
1
5
4.6
1
25
4.6
d
1
50
4.6
e
1
25
5.0
f
1
25
5.5
g
1
25
6.0
h
1
25
6.5
PP
BSA
a-
La
13-
Lg
13-L
g
a-
La
4461
a
bcdef
g
h
JOURNAL
OF
DAIRY
SCIENCE
VOL.
55,
No.
5
Slot
Whey
Per-
oxide
Temp
(%
w/w)
(0)
a
Original
0
b
5:1
0
10:1
c
0
d
Original
1
50
e
5:1
1
50
10:1
f
1
50
g
Original
1
25
h
10
:1
1
25
Ig
PP
BSA
a-La
13-L
g
MD
MO
41•16
I
570
COONEY
AND
MORR
TABLE
1.
Effect
of
experimental
factors
upon
total
whey
protein
denaturation
by
peroxide
in
pH
4.6
whey.
Peroxide
Time
Temp
Dena-
turation.
(%
w/w)
(hr)
(C)
(%)
BSA
1
24
25
4
a
-
La
1
4
50
18
1
6
50
28
2
4
50
38
-Lg
partially
resolubilized
and
fractionated
in
the
presence
of
excess
2-mercaptoethanol
and
7
M
urea.
The
effect
of
the
aforementioned
factors
upon
total
whey
protein
denaturation
is
shown
in
Table
1.
The
reaction
temperature
is
ex-
tremely
important,
i.e.,
only
4%
protein
was
denatured
by
treating
whey
with
1%
peroxide
a
b
c
d
e
U
g
h
FIG.
5.
Effect
of
protein
concentration
and
temperature
upon
whey
protein
denaturation
by
1%
peroxide
for
3
hr
at
pH
4.6
shown
by
poly-
acrylamide
gel
electrophoresis.
Whey
was
con-
centrated
by
pressure
ultrafiltration.
Volume
added
to
all
slots,
75
µliters.
Electrophoresis
conditions:
polyacrylamide
gel;
pH
8.6
veronal
buffer
(I'
=
.02)
in
gel
and
cell;
and
constant
voltage
of
180
v
for
3
to
4
hr
(9).
abcdefgh
FIG.
4.
Effect
of
peroxide
concentration,
pH
and
whey
protein
concentration
(concentrated
by
ultrafiltration)
on
peroxide
alteration
of
whey
proteins
shown
by
polyacrylamide
gel
electro-
phoresis.
Reaction
conditions:
time,
4
hr;
tem-
perature,
50
C;
pH,
held
at
indicated
values
throughout
peroxide
reaction
and
ultracentrifuga-
tion.
Volume
added
to
all
slots,
50
µliters.
Elec-
trophoresis
conditions:
polyacrylamide
gel;
pH
8.6
veronal
buffer
(I'
=
.02)
in
the
gel
and
cell;
and
constant
voltage
at
180
v
for
3
to
4
hr
(9).
Per-
Reac-
tion
Slot
oxide
Whey
PH
(%
w/w)
a
0
Original
4.6
b
1
Original
4.6
1
Original
5.6
d
1
Original
6.6
e
0
3:1
4.6
f
1
3:1
4.6
g
h
0
6:1
1
6:1
4.6
4.6
for
24
hr
at
25
C
whereas
28%
protein
was
denatured
by
treating
whey
at
50
C
for
only
6
hr.
The
direct
relationship
between
peroxide
concentration
and
time
with
extent
of
protein
denaturation,
shown
here,
is
consistent
with
the
electrophoretic
data
in
Figure
2.
Electrophoretic
data
in
Figure
4
demonstrate
the
effect
of
pH
upon
the
extent
of
physical
aggregation
of
individual
whey
proteins
by
peroxide
as
well
as
the
relationship
between
whey
protein
concentration
and
peroxide
de-
naturation
of
these
individual
whey
proteins.
As
in
Figure
2,
peroxide
treatment
of
whey
at
progressively
higher
pH
values
produced
suc-
cessively
greater
amounts
of
physical
aggrega-
tion
and
greater
electrophoretic
mobility
shifts
in
p-lactoglobulin,
but
caused
successively
smaller
changes
in
bovine
serum
albumin
and
the
immunoglobulins.
a-Lactalbumin
was
rela-
tively
unaffected
by
any
peroxide
treatment.
These
data
further
confirm
that
not
all
of
the
$-lactoglobulin
denatured
at
pH
5.6
and
6.6
(Fig.
2
b
and
c)
are
aggregated
to
sufficient
JOURNAL
OF
DAIRY.
SCIENCE
VOL.
55,
No.
5
HYDROGEN
PEROXIDE
AND
PROTEINS
571
TABLE
2.
Effect
of
protein
concentration
upon
extent
of
whey
protein
denaturation
by
1%
peroxide
in
pH
4.6
whey.
Time
Protein
Protein
denaturation
Per
100
ml
whey
Of
total
whey
proteins
(hr)
(
%)
(g)
(
%)
3
0.66
0.12
18
2.75
0.70
25
5.80
0.80
14
4
0.93
0.18
19
2.69
0.52
19
5.36
0.62
12
size
to
sediment
at
206,000
X
g
(Fig.
4
c
and
d).
Comparable
changes
in
electrophoretic
patterns
were
produced
by
peroxide
treatment
of
whey
and
3:1
and
6:1
concentrated
whey.
Eleetrophoretic
data
in
Figure
5
reveal
that
total
whey
protein
concentration
affects
the
relative
susceptibility
of
individual
whey
pro-
teins
to
peroxide
denaturation
at
50
C
but
not
at
25
C.
For
example,
$-lactoglobulin
is
less
susceptible
to
peroxide
denaturation
at
high
protein
concentration
(Fig.
5
d—f),
whereas,
the
reverse
is
true
for
bovine
serum
albumin
and
the
immune
globulins.
Again,
a-lactalbumin
denaturation
is
not
a
function
of
protein
con-
centration.
Data
in
Table
2
demonstrate
a
direct
relationship
between
protein
concentra-
tion
and
extent
of
total
whey
protein
denatura-
tion.
However,
the
percentage
of
total
whey
protein
denaturation
is
not
linear
with
total
whey
protein
concentration.
Sephadex
G-150
gel
filtration
experiments
43
51
CONTROL
38,4
5
..•••••••••..
23
71
HP2
2
&4
5
ELUTION
VOLUME
FIG.
6.
Effect
of
peroxide
upon
Sephadex
G-150
gel
filtration
patterns
of
whey
proteins
by
treat-
ing
pH
4.6
whey
with
1%
peroxide
at
50
C
for
6
hr.
Small
numbers
designate
peaks
and
large
numbers
indicate
percentage
of
total
pattern
area
by
planimeter.
TABLE
3.
Role
of
calcium
and
sulfhydryl
groups
on
denaturation
and
physical
aggrega-
tion
of
whey
proteins
produced
by
treating
whey
and
Ca-free
whey
with
1%
(w/w)
per-
oxide
at
50
C
for
4
hours.
See
procedure
for
details
of
Ca-free
whey
preparation.
Dena-
Aggre-
turation
gation
Sample
(pH
4.6)
(pH
5.6)
(percent
of
total
proteins)
Whey
Control
9
25
+
NEM
9
8
Ca-free
whey
Control
19
11
+
NEM
19
9
demonstrated
that
peroxide
produces
significant
alteration
of
whey
protein
molecular
species,
i.e.,
peroxide
treatment
reduced
the
amount
of
the
major
whey
protein
components
that
remained
in
the
ultracentrifugal
supernatant
fraction
(Fig.
6).
Additional
experiments
demonstrated
that
the
peroxide
treatment
pro-
duced
a
small
amount
of
breakdown
of
proteins
to
low
molecular
weight
peptides
and
amino
acids
in
Peak
5.
Results
in
Table
3
demonstrate
that
calcium
increases
the
susceptibility
of
whey
proteins
to
peroxide-induced
physical
aggregation
at
pH
5.6,
but
that
blocking
sulfhydryl
groups
on
the
proteins
with
N-ethylmaleimide
eliminates
the
ability
of
calcium
to
promote
protein
aggrega-
tion
at
this
pH.
However,
the
extent
of
protein
denaturation
produced
by
treating
both
whey
and
Ca-free
whey
with
peroxide
at
pH
4.6,
was
not
affected
by
sulfhydryl
group
blocking,
and
unexpectedly
a
significantly
greater
amount
of
whey
protein
was
denatured
at
pH
4.6
in
the
Ca-free
whey
system
than
in
the
presence
of
calcium.
These
observations
suggest
that
blocking
sulfhydryl
groups
stabilizes
the
whey
proteins
against
conformational
changes
re-
quired
to
permit
aggregation
via
ionized
side
chains
with
calcium
cross-links.
The
reason
for
greater
amounts
of
whey
protein
denatura-
tion
at
pH
4.6
in
the
absence
of
calcium
than
in
the
presence
of
calcium
is
not
known.
The
higher
degree
of
protonation
of
ionized
groups
on
the
protein
at
this
lower
pH
would
be
expected
to
lower
the
ability
of
calcium
to
par-
ticipate
in
intermolecular
protein
interactions.
Discussion
Hydrogen
peroxide-catalase
which
has
been
recommended
for
treating
milk
intended
for
ABSO
R
B
ANC
E,
280
n
m
JOURNAL
Or
DAIRY
SCIENCE
VOL.
55,
No.
5
572
COONEY
AND
MORR
cheese
manufacture
(see
reference
13),
offers
an
alternative
approach
for
controlling
micro-
bial
growth
during
extended
times
required
for
preparing
a
whey
protein
concentrate
by
pres-
sure
membrane
and
gel
filtration
processes.
Others
have
used
peroxide-catalase
in
processes
for
preparing
protein
concentrates
from
whey
(1,
7)
and
from
peanuts
(2).
A
major
advan-
tage
of
peroxide-catalase
as
an
antimicrobial
agent
is
that
residual
peroxide
can
be
quantita-
tively
converted
to
water
and
oxygen
upon
completion
of
the
process.
Admittedly,
the
concentrations
of
peroxide
in
this
study
were
higher
than
the
.05%
limit
permitted
for
treating
milk
intended
for
Cheddar
cheese-
making
(15).
However,
the
present
peroxide
concentrations
are
comparable
to
those
used
by
others
studying
changes
in
milk
proteins
in
skimmilk
(1,
4,
6,
13).
Results
in
this
study
indicate
that
minimal
whey
protein
de-
naturation
and
aggregation
are
produced
when
peroxide
and
temperature
are
maintained
below
0.5%
and
in
the
region
of
25
C.
Goldsmith
et
al.
(5)
strongly
recommended
that
ultra-
filtration
for
recovering
whey
proteins
from
acid
whey
be
at
about
50
C
for
optimum
membrane
flux
characteristics
and
for
secondary
bacterio-
static
benefits.
Roundy
(12)
reported
that
0.06
and
0.3%
hydrogen
peroxide
at
55
C
provided
99%
reduction
of
common
fermenta-
tive
and
pathogenic
bacteria
and
bacterial
spores
in
milk,
respectively.
However,
the
present
study
shows
that
the
peroxide-catalase
treat-
ment
(-1%
H
2
0
2
)
produces
high
amounts
of
whey
protein
alteration
at
these
high
temper-
atures.
Further
work
is
required
to
determine
the
optimum
combination
of
temperature
and
peroxide
treatment
for
preparing
an
unde-
natured
whey
protein
concentrate
with
minimal
microbial
growth.
The
effects
of
peroxide
upon
whey
proteins
in
whey
and
concentrated
whey
systems,
in
this
study,
agree
generally
with
those
reported
for
whey
proteins
treated
with
peroxide
in
skimmilk
(4,
6).
However,
the
nature
of
the
whey
protein
reactions
and
interactions
in
peroxide-treated
whey
are
undoubtly
different
than
those
in
peroxide-treated
skimmilk,
where
the
greater
probability
exists
for
interaction
between
denatured
whey
proteins
and
the
pre-
dominant
casein
components.
Disappearance
of
/3-lactoglobulin
by
treating
skimmilk
with
per-
oxide
has
been
well
documented
(4,
6),
however,
it
was
not
known
whether
the
reaction
proceeded
through
interactions
with
caseins
or
through
/3-
lactoglobulin
self
interaction.
The
present
study
provides
new
information
on
the
denaturation-
aggregation
reactions
of
/3-lactoglobulin
and
the
other
whey
proteins
and
provides
a
basis
for
predicting
and
avoiding
protein
alteration
where
peroxide
is
utilized
as
an
antimicrobial
agent
during
whey
protein
concentrate
prep-
aration.
Additional
findings
in
our
laboratory
confirm
that
proper
control
of
peroxide
levels
and
operating
conditions
completely
eliminates
the
problem
of
whey
protein
alteration
during
the
pressure
membrane
process.
5
5
Whey
protein
concentrate
samples
supplied
by
A.
A.
Nieland
and
Dean
Spatz,
Osmonics,
Inc.,
Minneapolis.
References
(1)
Bechtle,
R.
M.,
and
T.
J.
Claydon.
1971.
Glucose-residue
polymers
as
protectants
against
heat
denaturation
of
whey
proteins.
J.
Dairy
Sci.,
54:
1410.
(2)
Chandrasekhara,
M.
R.,
B.
R.
Ramanna,
K.
S.
Jagannath,
and
P.
K.
Ramanthan.
1971.
Miltone
vegetable
toned
milk.
Use
of
peanut
protein
expands
supply
of
milk.
Food
Technol.,
25
:
596.
(
3
)
Cooney,
C.
M.,
C.
V.
Morr,
M.
A.
Nielsen,
and
R.
H.
Schmidt.
1971.
Hydrogen
peroxide-induced
denaturation-aggregation
of
whey
proteins.
J.
Dairy
Sci.,
54:
753.
(4)
Fish,
Nancy
L.,
and
R.
Mickelsen.
1967.
Effect
of
hydrogen
peroxide
on
whey
pro-
tein
nitrogen
value
of
heated
skimmilk.
J.
Dairy
Sci.,
50:
1045.
(
5
)
Goldsmith,
R.
L.,
C.
H.
Amundson,
B.
S.
Horton,
and
S.
R.
Tannenbaum.
SOS
70
Proceed.
Third
Int.
Congr.
Food
Science
and
Technol.,
Washington,
D.C.,
August,
1970.
(6)
Grindrod,
Jean,
and
T.
A.
Nickerson.
1967.
Changes
in
milk
proteins
treated
with
hydrogen
peroxide.
J.
Dairy
Bei.,
50:
142.
(
7
)
Hansen,
P.
M.
T.,
and
Lois
Crauer.
1971.
Production
of
complexes
between
whey
pro-
teins
and
food
stabilizers.
J.
Dairy
Sci.,
54:
756.
(8)
Jenness,
R,
and
J.
Koops.
1962.
Prep-
aration
and
properties
of
a
salt
solution
which
simulates
milk
ultrafiltrate.
Nether-
lands
Milk
Dairy
J.,
16:
153.
(
9
)
Morr,
C.
V.
1969.
Protein
aggregation
in
conventional
and
ultra
high-temperature
heated
skimmilk.
J.
Dairy
Sci.,
52:
1174.
(10)
Morr,
C.
V.
1971.
Comparison
of
protein
preparation
procedures
and
starch
versus
polyacrylamide
gel
electrophoresis
for
ex-
amining
casein
degradation
products
in
cheese.
J.
Dairy
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54:
339.
JOURNAL
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VOL.
55,
No.
5
HYDROGEN
PEROXIDE
AND
PROTEINS
573
(11)
Proceedings
of
Whey
Utilization
Conference.
U.S.
Department
of
Agriculture,
College
Park,
Maryland,
June
1970.
(12)
Roundy,
Z.
D.
1958.
Treatment
of
milk
for
cheese
with
H202.
J.
Dairy
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41:
1460.
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Schmidt,
R.
H.,
H.
A.
Morris,
and
C.
V.
Morr.
1969.
Aetion
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rennet
on
casein
as
influenced
by
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Tessier,
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41:
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Title
21.
Code
of
Federal
Regulations,
Section
19.500.
Revised
January
1971.
JOURNAL
OF
DAIRY
SCIENCE
Vol..
55,
No.
5