1
P. FEILLET* J.C. AUTRAN O. AIT MOUH
ROLE OF LOW MOLECULARWEIGHT GLUTENINS IN DETERMINING COOKING QUALITY OF PASTA PRODUCTS**
It
is
the
cooking
the
baking
clearly
established
quality
of
quality
durum
that
gluten
wheat
pasta
of conunon wheat
proteins
controlled
products
as
well
as
flour.
Figure
1
LOW MOLECULARWEIGHT GLUTENINS : LARGE PROTEIN AGGREGATES WHICH YIELD UPON REDUCTION SUBUNITS WITH APPARENT MWOF 12,000 TO 60,000
Among
glut en
( LMWG ) are
those
aggregates
and
proteins pro tein s
yield
weight
*Institut Technologie Cédex . **Presented
National de la des Céréales, the
molecular
weight
which
correspond
to
upon
molecular
at
of
low
12,000
72nd
reduction to
subunits
glutenins
large with
prote
in
apparent
60 , 000 .
Recherche 9 Place
Agronomique, Viala, 34060
AACC meeting,
Nashville
Laboratoire Montpellier (USA),
de 1987.
2
Figure
2
GLUTEN PROTEINS TYPE
---------
GLIADINS HMWGLUT. LMW GLUT.
MW (K)
LOCI
PROPERTIES
------------- MONOMERIC -----------
-------
GLI - l**;GLI-2 GLU-1 GLI-1
25-70 65-130* 12-60*
POLYMERIC AGGREGATIVE
*AFTER REDUCTION **OMEGA AND GAMMAGLIADINS
They glutenins
{HMWG) by
bec a use short
different
are
they
are
from
the
of
chromosomes
on
the
long
located
smaller
genetically
arms
polymeric
and
monomeric
materials.
arm
the
high
size
of
controlled 1 of
aggregative
their
by loci
while
HMWG have
groupe
1
proteins
Figure
molecular subunits located
and on the
several
chromosomes. while
weight
They
gliadins
3
------------------------------------------281 566 622 750 806 872 901 928 - 316
--------
-776
-847
-1071
-----
HMWG LMWG VLMWG f GLIAD ~ GLIAD fGLIAD ~GLIAD ALB-GLOB
-------3
are are
SDS PAGE SUBUNITS OF TOTAL PROTEINS
PROTEINS
MW ( xl0
loci
)
110
68
62.5
51.5
44.5
43
41.5
----35
3
the y
However polyacrylamide protein
gel
in
belonging
the
chromatography identify
to
Bef ore
in
the
the
known
relationship ty,
and
protein
how
cooking
occuring
between
the
of
pasta
genetic
gliadin
Nevertheless
it and
of
is
to
by
exchange possible
appreciate
to their
pool.
qua li ty on
omega
ion
LMWG could
findings
on
zone.
SDS
occurence
and
mobilities, ically
by
the
solubility,
specif
total
explaining
determining
quali
subuni t
and · SDS-PAGE
contribution
of
gamma gliadîn
LMW glutenin
LMWG subuni ts
well
because
to
differences
combining
identified
easily
not
electrophoresis
fractions
proteins
are
have
pas ta
let
cooking gamma
control
a
major
· us
recall
quality,
gliadins of
role
three on
and
gamma
in
the
cooking
gliadins
and
LMWG:
Figure
a)
4
PASTA COOKING PROPERTIES 1 - MATTER LOSSES, SWELLING 2 - VISCOELASTICITY, FIRMNESS 3 - CONDITIONS OF SURFACE
cooking
Pas ta
.
parameters
mat ter (or
absorption behaviour
and
condition
of
It surface
is are
quality
is
los ses
in
swelling)
firmness surface
now well
after
boiling
during
cooking
cooking
.,
of cooked
pasta.
documented
that
two independant
related
parameters.
pas ta
firmness
to water ;
three and
water
viscoelastic
desaggregation
and
main
condition
or
of
4
b)
Figure
PASTA COOKING QUALITY MATTER LOSSES, SWELLING VISCOELASTICITY: PROTEIN CONTENT "! 45/LMWG1.. CONDITIONS OF SURFACE
1
2 3
It
is
also
viscoelasticity but
also
lacis
gamma 45
· well is
not
genetically
: glutens
their
high
rated
as good
documented only
are
that
correlated
determined
extracted
allele
5
.
or very
good
in
varieties
their
high
LMWGare loci,
under
which
has
the
two
gamma 45. LMWG subuni ts 42 group
of
loci
gamma 45 group
genetic of
loci.
the
firmness
and
pasta
are
cooked
LMW-1 (4) LMW-2 (3)
control
of
allelic
types
( a quadruplet)
; LMWGsubunits of
with
of
LOCUS
6l-33-35-38 W-35
group
group
LMWGLUTENINS
GLIADINS '(-42 ~-45
42 45
content
6
ALLELIC TYPES OF GLI-Bl ALLELE
protein
gamma 42/ 45
following
pasta
firmness.
Figure
c)
the
cooked
wheat
by
, the
ty
the
durum
characterized
viscoelastici
to
by
from
the
(a
are triplet)
the
GLI-Bl
..
coded are
group
gamma by
the
coded
42
of and
gamma by the
0
5
Figure
7
GENETIC CONTROL OF GLUTENINS GLUTENINS
---------
LOW MW HIGH MW
Much more in
durum
It
is
the
HMWGallelic
not
easy
of
gluten
only
This
properties
of
gamma differences
type
5 8
(5 x 8) have
of
these
been
between
differences
from
the
presence
two
reasons
two
do
gliadins
the
extracted
not
LMWG, which 45
explain
for
contents
proteins.
to
basis
gliadins.
their
------------2 (42/45)
GLI-Bl GLU-Al GLU-B1
identified
wheat.
properties on
ALLELIC TYPES
LOCI
------
42 or of
are
are
genetically
could
be
42 and
45 type
the
or
42
more
the
of
the
total
linked
to
gluten.
8
LMWGALLELIC TYPES AND GLUTEN RECOVERY GLI AD
----- 42 45 42
GLI AD
-----42 45 45
LMWG 42 45 45
*PROVI DED BY N . POGNA
N n Il
GLUTEN RECOVERY
--------
1
1it
45
ganuna
LOW HIGH HI GH
same
;
pasta
gamma 42
responsible
durum wheat
Figure
wheats
physicochemical like
%
physical
durum
very
2-3
the
45 type
the
proteins excess
in
for
and the
•
6
The discovery and
of
1(42 is
which only
of
the
a variety
gluten
a genetic
which
recovery
is
is
~42/w
45/LMWG 45 type
high\Jll;J.ngs
the
proof
that
marker.
Figure
9
VARIETAL DIFFERENCES IN LMWGLUTENINS - PHYSICOCHEMICAL, FUNCTIONAL? - QUANTITATIVE: % GLUTEN PROTEINS?
Then between in
one
more
glutens
due
their
to
Let
us try
and to
.
arises
differences
physicochemical
parameters.
I.
question
in
the
functional
answer
this
are
the
quantity
properties
differences of
LMWGor
or
to
both
question.
LMWGCONTENT
The
amount
co mbi n i n g sol vant SOS-PA GE.
of
LMWG in extraction,
semolina
was
io n ex change
first
determined
ch r omatography
by and
7
Figure
l 'e-
PROTEIN COMPOSITION OF DURUMWHEAT PROTEINS
CALVINOR (42)
AGATHE (45)
SALT SOLUBLE GLIADINS GLUTENINS INSOLUBLE
23.5 33.4* 33.5 9.6
19.2 22.2** 49.5 9.1
* '6"-42:
Two
varieties
AGATHE ( type in
gliadins,
but
good
(at
** ~45
were
45) .
glutenins
gliadins
3.9
least
examined
We first quality
only
2.6
:
conf irmed durum
minor
CALVINOR ( type the
wheat
differences
in absolute
42)
and
higher
amount
of
lower
co n tent
in
and
a
in
'(42
and
~45
gamma
value).
Figure
11
GLUTENIN COMPOSITION OF DURUMWHEAT GLUTENINS
CALVINOR ( 4 2)
AGATHE (45)
HIG H MW LOW MW VERY LOW MW
15 . 1 8.2
27 . 7 10.3
----------- ----------------------10 .2 11 . 5
PER CENT TOTAL PROTEINS
The contributions which are
we have 10.2
and
(VLMWG)
for
of the
included 11.5
respectively
.
indications
on the
VLMWG-
(HMWG ),
gamma These
different
42
15.1 and
data
content
to and
the
these
total
27.7
gamma certainly
in
typ e s of glu te n in s - in gl u t e nin
(LMWG),
45 give
glutenins
type
8.2
content and
durum no
families.
more
10 . 3 wheat than
8
Figure
12
GLUTENIN COMPOSITION OF DURUMWHEAT GLUTENINS
CALVINOR (42)
AGATHE (45)
HIGH MW LOW MW VERY LOW MW
45.1 24.4
56.0 20.8
----------- ----------------------30.5 23.2 PER CENT TOTAL GLUTENINS
Nevertheless, demonstrate gluten
that
is
glutenin higher
expressed the
to
phosphate mainly frationate
second
the
a
the
approach
buffer LMWG,
in
( that
high
total
glutenin,
glutenin
of 42 type
extract
we have
shown
salt
soluble exclusion
Figure
SE-HPLC
of
HMWG and
to
by size
of
percentage
both
was
VLMWG and solubles
of
viscoelasticity
Conversely,
concentration
Our
strong
simultaneous fraction.
in percent
to
the
they AGATHE
LMWG in
the
VLMWG are
in
durum wheat.
proteins
with
be
to
able
proteins)
a SOSextract and
to
chromatography.
13
Peak N°
PM ( KO)
800
SE HPLCOFSOS-PHOSPHATE BUFFER EXTRACT
2
250
3
43
4
13
9
Four
peaks
from
800
KD to
and
2
weight
are
were
13 KD. It
mainly
glutenins
satisfactory
that to
with
can
be
constituted but
Assuming
identified
this
this
quote
postulated by
has
molecular
still
to
following
that
aggregated
ranging
both low
peaks
1
molecular
be proved.
hypothesis
the
weight
is
true
it
is
quite
results.
Figure
14
CORRELATION BETWEEN HPLC PEAKS SURFACE AND GLUTEN PROPERTIES GLUTEN* HPLC PEAKS
FIRMNESS
Pl P2 P3
RECOVERY
0.81 0.75 - 0.85
0.88 0.82 - 0.86
*HEAT SHAPED
Having locations) the
compared we
contents
firmness
* Heat
and
shaped
found in
the
.
24 a
Pl, gluten*
durum highy
P2,
( Pl
elastic
wheat . samples signif +
icant P2)
recovery.
(6
varieties,
correlation
peaks
and
between the
gluten*
4
0
10 Figure
15
VARIANCE IN HPLC PEAKS SURFACE HPLC PEAKS
% OF VARIABILITY
----------------------------------LOCATION VARIETY LOCATION
VARIETY
-----
-------
Pl P2 P3
in
genetically
--------
91.0 91.6 93.8
Furthermore, variability
F TEST
--------
** ** **
of
P2
P3
contents
is
almost
the
location
has
no effect.
Pl,
and
while
variance
NS NS NS
analysis
dependant
Figure
r 42 TYPE ~-----14
-------
1.4 4.1 1.2
has
shown
that
the
exclusively
16
---'~"--4
~-Y_P E__ _
% Peak 1
12 10
8
0 0
6
Elastic Recovery 0
1.0
2.0 mm
BETI-/EEN PEAK1 SE HPLCOF SOS-PHOSPHATE RELATIONSHIP BUFFER SOL UB LE PROTEINS ANDGLUTEN ELASTIC RECOVERY
0
11
This
relation
( in
% of
total
the
gluten
wheat
are
durum
wheat
consti
of
the
tuted
content
which
temp~ ·rature ei ther
the
of
is
mainly
of
pasta
amount
of
proteins)
Pl
versus
in
Pl,
two
groups
the
42-type
of and
durum 45-type
OF LMWG
an
the
left
in
the
shape),
important
drying
performed
behavior
hydrothermic
The
the
soluble
over1ap
is
during
frequently
beeing
( depending on
buffer
FUNCTIONAL PROPERTIES
more
by plotting
groups.
technology
pasta
visible
recovery.
basis
Because
and
more
SOS-phosphate
elastic
On the
II.
is
of
operation
above
boiling we pasta
parameter
70°C
water
which or
for
concentrated proteins
of
our under
are
during
about
pasta more
cooking, 10
minutes
investigations well
defined
treatment.
first
finding
responsable proteins
is
that
(but during
heat
not
the
formation
only) treatments.
for
of the
disulfide insolubilization
bonds
12
Figure
17
EFFECT OF HEAT TREATMENTAT 30 % MOISTURE CONTENT ON PASTA PROTEIN SOLUBILITY TREATMENT
SEQUENTIAL
EXTRACTION
MIN.
T°C
MSH
SDS
SDS
MSH
30 120
120 120
42 20
58 68
43 31
67 57
-----------------------------------52 100 1 51 CONTROL
MSH SDS
Pasta and in
with
1 % MERCAPTO ETHANOL 0.5 % SDS
30 % moisture
content
120 minutes
. By comparison
SDS solubility
was observed
"insoluble
SDS"
mercapto when in
.
ethanol
using
proteins (at
least
was not
to
were
the
after
left
heat
the
treatment
the
first
a sharp
in is
not
solvant,
so important.
Figure
18
..
SOLUBLE PROTEI NS ( %NT) 100
C14 /MSH
50
0
C14 CH3 COOH H o 2
------~--~-L------•---• s
c
Pl3
P18
120°C
treatment;
extracted
in
at
control
were if
mercaptoethanol
solubility
(?)
P24
EFFECTS OF HEAT TREATMENT (2HRS-90°C) AT 13 % (Pl3), 18 % (Pl8) AND 24 % (P24) MOISTURE CONTENT. S: SEMOLINA; PC: CONTROL PASTA
for
30
decrease then
the
presence too the
long) decrease
of ;
0
13
In
another
moisture was
content
the
the
experiment
heat
lost
were
disrupting were
aggregate
in
with
to
during
18
%,
at
higher
the
sodium
myristate,
% and
90°C.
humidity),
of
more
accuretely
was
hydrophobie
the
residual
the
proteins
which
treatments.
Figure
19
....
r.. •· • t .. . •••••• C f ' "lf1~f ' ...-, t I Il I
-. ..,~...
.
w~·
DT
PAGE (PH=3.2) OF CHLOR0-2-ETHANOL SOLUBLE PROTEINS A: SEMOLINA; B: PASTA DRYED AT 55°C; C, D ANDE: PASTA LEFT FOR 2 HRS AT 90°C AT 13 %, 18 % AND 24 % MOISTURE CONTENT Beside which to
have
outline
the
well
a very the
proteins
( still
phoresis)
after
known
low
content
disappearance staying heat
%
Stronger larger
a
most
12
mercaptoethanol.
identify
heat
24
2 hours
Simultaneously,
soluble
PAGE allows
wi th
during
(i.e.
solubility agent.
proteins
left
treatment
of
pas ta
heat
resistan in
of in
treatment.
the
ce
sulphur, the
of it
streaks
sample
wells
omega is
and
gliadins,
most of after
the
val~able "well" electro-
()
14
To identify millimeter the
of
proteins
the the
with
"well"
gel
pro teins
after
the
we sliced
well,
of
which
a mercaptoethanol-phosphate
Figure
94
out
the
f irst
we dissolved
buffer.
20
67
43
30
20
14
'
i ··...,
Most
of
this
molecular
weight
the
of
range
the
SDS-PAGE of the
same
conclusions
products
material ranging
is from
LMWGsubunits
SDS phosphate after
or
composed 35. 000
to
with
. -
proteins
50. 000 .
with
That
say
.
soluble not
proteins
reduction
extracted lead
from
to
similar
.
Fig ure P24 P18 P13
in
'
21
.
!.i
'
c
s SDS-PAGE OF SDS PHOSPHATE BUFFER SOLUBLE PROTEINS . S: SEMOLINA; C: CONTROL PASTA; Pl3, Pl8, P24 : PASTA LEFT AT 90°C FOR 2 HRS AT 13, 18, 24 % MOISTURE
•
•
:j ·• .. ..1
c 15
Figure
22
1
P24
1'
•
P18
1
P13
•
..
c s
SOS-PAGE OF REDUCED SDS PHOSPHATE SOLUBLE CONTROL PASTA; PROTEINS. S: SEMOLINA; C: Pl3, Pl8, P24: PASTA LEFT AT 90°C FOR 2HRS AT 13, 18 , 24 % H 0 2
S.E. results.
HPLC of
the
same
(unreduced)
extracts
confirm
these
16 Figure
23
43 KD
.,~13KD KD }c>2 GEL PERMEATION HPLC CHROMATOGRAPHY OF SOS-PHOSPHATE EXTRACTS-SEMOLINA: A; PASTA DRIED AT 55°C: B; PASTA LEFT FOR 2 HRS AT 90°C AT 13 % (C), 18 % (D) AND 24 % (E) MOISTURE CONTENT
Figure
MW(l0 3 )
-------
>800
s c
24 3
P13 P18 P24
1 2 0
24
250
43
13
18 11 0 0 0
38 70 69 38 37
11 14 22 29 31
S E HPLC OF SOS-PHOSPH ATE SOLUBLE PROTEINS . S : SEMOLINA ; C: CONTROL PASTA; Pl3 , P l8 AND P2 4: PASTA LEFT FOR 2 HRS AT 90 °C AT 13 , 18, 24 % MOISTURE
Pl, curves The heat
P2 and while
P3 progressively
increases
sensitivity
the of
inten
disappeared s i ty
LMWGaggregates
of
the
was
from
the
heat
treatments.
so confirmed
elution
.
17 ROLE OF VERY LOW MOLECULARPROTEINS
The discovery sulfur
glutenin"
DSG
flour
(DSG) allows
of
They
then
in
content higher
proteins
0.5
with
NaCl
wi th
other
proteins
in
wheat
varieties
very ty
durum good
because
candidates of
abili
to
low
flour)
from
and
N acetic
140
acid
but
LMWG). 180
have ty
a
to
Their
mol/g.
and these
major
create
%
60
consequently
to
their
g.
(mainly
from
rich
by
solution
O. 01
ranges
good
form
mg per
SH + SH groups
are quali
by
solubilized
mixture in
pasta
also
(3.5
wheat
further.
purified
a
myristate
extracted
a "durum
go a little
in
sodium
previously
ethanol.
to
solubilized
are
concentration
is
by KOBREHEL and ALARY of
role
in
disulfide
linkages. Figure
25
DURUMWHEAT SULFUR RICH GLUTENINS . SH+SS CONTENT: 180-140 MOL/G • SOLUBLE IN PURIFIED FORM BY Cl4 . SOLUBLE WITH LMWGBY CH3COOH . MOLECULARWEIGHT : 14,000-17,000
As
a
matter
of
fact
positive
correlation
glutenin
and
the
condition
to
a
functional
of
the
concluded desaggregation
KOBREHEL and
betwe en
surface
the of
content
surface
role of
ALARY found
of the
in of DSG
cooked
SS+
cooked in pasta.
a
highly
SH group pasta
preventing
;
in they the
18
Figure
1 2
3
There true
is
(what
linked
pro teins proteins
and,
PASTA COOKING QUALITY MATTER LOSSES, SWELLING VISCOELASTICITY CONDITIONS OF SURFACE: DSG (SULPHUR RICH GLUTENIN)
no special
glutenins
26
interest does
in
that
through
discussing
means
or
?)
hydrophobie
by consequence,
wether
DSG are
soluble
bonds
to
like
insoluble
insoluble.
Figure
27
DURUMWHEAT SULFUR RICH GLUTENINS DSG: LOW MOLECULARWEIGHT MONOMERIC PROTEINS, HIGH IN CYSTEIN, LINKED TO LMWGBY HYDROPHOBIC BONDS, ABLE TO ENTER INTO S-S BONDS DURING PASTA PROCESSING
Our weight t o link
hypothesis
monomeric together
wh ic h they
is
that
p r oteins by
ar e a l ready
the and
d i sulfide linked
so
high bond,
called in
DSG ( lo w molecular
cystein)
a nd
by hydrophobie
t h en
would to
b onds.
link
be
able
LMWG to
19
Figure
28
LOW M.W. GLUTENINS ~
î
~
{ ~
\ ;J.
SH ••• HS
l
' DSG
:1
SH
1
}
HYPOTHETIC ROLE OF DS-GLUTENIN IN THE AGGREGATION OF LMW-GLUTENINS
This
hypothesis
is
scherned
on Figure
28 .
CONCLUSION
Figure
29
HYPOTHESIS 1-LMWG CONTENT IS RESPONSIBLE FOR THE VISCOELASTIC PROPERTIES OF HEAT SHAPED GLUTEN IN DURUMWHEAT 2-LMWG AGGREGATE THROUGH HEAT TREATMENT AND THEREFORE CONTRIBUTE TO THE PASTA FIRMNESS AND VISCOELASTICITY 3-THE LMWGCONTENT OF DURUM WHEAT IS GENETICALLY CONTROLLED TWO FAMILIES OF DURUMWHEAT HAVE BEEN IDENTIFIED 4-DSG CONTRIBUTE TO THE AGGREGATION OF LMWG , THROUGH HYDROPHOBIC AND DISULFIDE BONDS 5-THE LMWG-DSG BONDS ARE TIGHT ENOUGH TO PREVENT STARCH LEACHING DURING PASTA COOKING AND TO KEEP SATISFACTORY THE STATE OF SURFACE OF COOKED PASTA 6- "
