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RESEARCH
Baking performance of
glutathione in yeasts
Effect of yeast-equivalent amounts of glutathione on the baking process.
+The quality impairment of dough and baked products
due to the amounts of glutathione in the yeast itself is
taken into account only for certain processes, e.g. freezing
and thawing dough pieces. Therefore the aim was to measure
yeast equivalent glutathione quantities typically exist in
commercially available fresh and dry yeasts, and to shed
light on their technological effect during the baking process.
As the link between the mixing and baking processes, the
fermentation of dough pieces is considered to be an essential
production step in controlling the leavening attributes of
baked goods. As a rule the production of bread and bread
rolls employs biological methods in which the addition of
yeasts in particular (as a rule Saccharomyces cerevisiae), to-
gether with the use of starter cultures, determines the pore
structure and final volume of the products. As well as the de-
sired leavening of the dough pieces through the production
of CO, other organic substances enter the dough medium
via the organism, e.g. acids, flavor precursors or reducing
Relationship between the water-soluble glutathione content
(mg/g of yeast (IDM)) in commercial fresh () and dry () yeasts
and the proportion of dead cells. A linear relationship exists
between the water-soluble glutathione content and the percentage
proportion of dead yeast cells (R .)
agents, e.g. glutathione (GSH). Glutathione is a so-called
pseudo-tripeptide formed from the amino-acids glutamic
acid, cysteine and glycine in the yeast cell. Whereas GSH
performs a multitude of physiologically important tasks in
the yeast cell, the free thiol group (-SH) of the cysteine im-
pairs the formation of the gluten network. Although the in-
tentional addition of rather small amounts of glutathione
leads to softer, more plastic doughs, the diffusion of uncon-
trollable quantities out of yeast cells can cause a sustained
reduction in product properties, e.g. restricted machinability
or volume losses. For example dough freeze-thaw processes
are implicated in this connection. The large temperature
gradient can be expected to cause damage to the yeast cells,
from which glutathione can accordingly merge into the dough
medium and damage the product quality. Therefore, the aim
was to determine the content of water-soluble glutathione in
commercial compressed and instant dry yeast, and to adjust
these amounts to the dough and end product.
The relationship between the proportion of dead cells
and the amount of water-soluble glutathione in yeast
An initial prerequisite to elucidate the effect of yeast-equiva-
lent amounts of glutathione on the properties of dough and
baked products was the use of a spectrophotometric method
to determine the water-soluble glutathione content of 27
commercial compressed (6) and instant dry (21) yeasts. As a
supplement, the percentage proportion of dead cells was
analyzed by using methylene blue to stain the dead yeast cells,
thus enabling a relationship to be derived between the pro-
portion of dead cells and the amount of glutathione. As can
be seen in Figure 1, a linear relationship exists between the
proportion of dead yeast cells and the glutathione content
(R = 0.8483). The slight departure from linearity is not sur-
prising, since although the amount of glutathione in the
yeast cell rises with increasing stress (critical temperatures,
oxidation, hyper-osmolality), dissolution/destruction of the
cell wall (lysis) is the primary mechanism for the release of
glutathione into the aqueous phase of the sample solutions
© Wikipedia
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RESEARCHRESEARCH
or dough. This means that in spite of a stress-mediated in-
activation of the metabolism, cell lysis need not necessarily
follow, and thus no GSH is available to weaken the dough.
The differences between fresh and dry yeasts are conspicuous.
Whereas the percentage proportion of dead cells was between
1.8 and 8.8 % for the fresh yeasts typically used in the manu-
facture of baked products, the percentage proportion of
dead cells determinable for dry yeasts was up to 75.0 %. As
a result of this, fresh yeast products yielded glutathione
amounts up to a maximum of 10.5 mg/g of yeast (dry base;
db) and for the dry samples a maximum value of 81.2 mg/g
of yeast (db). Interestingly, there are significant differences
not only between fresh and dry yeasts but even between
samples of the same brands produced on different days. The
percentage deviations for the proportion of dead cells reached
a maximum value of 75 %, and for the glutathione content a
difference of 91 %. Although these results point to fluctua-
tions in the production process, a very simple and quick de-
termination of the proportion of dead cells allows a good as-
sessment of the glutathione content in the yeast batch in
question. e analyses of the eect of yeast-equivalent amounts
of glutathione on dough development and end product quality
shown below were carried out with the following quantities
of GSH: 0.0; 7.3; 27.8; 58.6 and 76.2 mg per 100 g of flour.
Yeast-equivalent glutathione contents impair dough
stability during mixing/kneading
The effect of yeast-equivalent glutathione quantities on
dough development time (A), dough stability (B) and dough
softening (C) were analyzed during a 20-minute mixing time
in the 50 g bowl of a spiral mixing system (DoughLab;
Perten, Hägersten, Sweden). The dough recipe was composed
of 50 g of flour (moisture content corrected to 14 %), 29.4 g
of demineralized water and various amounts of glutathione
(0.0; 3.65; 13.9; 28.8 and 38.1 mg).
All three of the measured variables illustrated in Figure 2
(A–C) show that destabilization of the structure is obser vable
as the amounts of yeast-equivalent glutathione increase.
Whereas the dough development time of GSH-free doughs
needs approx. 7.4 minutes, this decreases by 67 % to approx.
2.5 minutes as the amount of GSH increases. Although a
The effect of yeast-equivalent amounts of glutathione (.; .; .; .; . mg/ g of flour) on (A) dough development time (), (B)
dough stability () and (C) dough softening (
) in the DoughLab. Dough development time (R .) and dough stability (R .)
show an asymptotic decrease. On the other hand dough softening rises as the amount of glutathione increases (R .). The average
value with the standard deviation (n ) is shown in each case
Influence on the gluten network of glutathione diffusing into the dough
medium from lysed yeast cells during blending/kneading. Illustration of the
dough system consisting of the gluten network with free thiol groups (-SH)
and disulfide bonds (R-SS-R) together with active/viable (oval-shaped, pale
blue filled, closed objects with yellow GSH molecules) and lysed (oval-shaped,
blue-filled, dashed objects) yeast cells. GSH molecules from the lysed yeast
cells enter the aqueous phase of the dough system, where they interfere with
the formation of new, and/or cause cleavage of existing disulfide bonds (see
schematic illustration with a magnifying glass)
shortening of the dough development time appears benefi-
cial at first, the extent of the destabilizing effect is apparent
especially in the asymptotic drop in dough stability (B) from
17.5 min to 1.8 min and in the increase in dough softening
(C) from 26 to 282 FE (Farinograph Units). As a rule, ex-
ceeding a dough softening of 70 FE is associated with moist,
very sticky doughs that makes further processing almost im-
possible. As already assumed by Pyler and Gorton (1988),
the quantities of glutathione introduced via dry yeasts lead
to significant impairment of the dough stability during mix-
ing/kneading [1]. In contrast, the amounts of GSH from fresh
yeasts have no significant influence on dough development.
Furthermore, it can be inferred from the asymptotic shape of
the three measurement values that the GSH-induced cleav-
age and/or inhibition of disulfide bridge bond formation is
limited by the gluten content of the flour [2].
Figure 3 shows schematically how individual glutathione
molecules (yellow objects) from lysed yeast cells (dashed oval -
shaped objects) enter the aqueous phase of the dough during
the blending / kneading process, thus impairing dough for-
mation. This does not apply for intact vital cells (oval- shaped
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RESEARCH
objects with unbroken lines), and the GSH molecules remain
in the cells during the blending / kneading process.
Reduction of gas retention capacity due to the input
of yeast-equivalent amounts of glutathione
e eect of yeast-equivalent amounts of glutathione on the gas
retention properties during proofing/fermentation was elu-
cidated by using a chemical leavening agent system consisting
of the leavening acid SAPP 10 (Sodium Acid Pyrophosphate;
provided by Chemische Fabrik Budenheim KG) and the car-
bon dioxide carrier NaHCO (Sodium hydrogen carbonate,
sodium bicarbonate). e chemical leavening method enabled
effects on the structure due to yeast metabolic products to be
avoided. Moreover, the influence of GSH in the presence of a
defined amount and formation rate of CO can be analyzed
through the chemical reaction between SAPP 10 and NaHCO.
Dough development during fermentation was measured in an
F3 Rheofermentometer (Chopin, Villeneuve- La-Garenne
Cedex, France). The quantities of added water and amounts
of glutathione relative to 300 g of flour (14 % flour moisture
content) were adjusted to correspond to dough preparation
in a spiral mixer. e chemical leavening agent system consists
of 9.9 g of SAPP 10 and 7.3 g of NaHCO resulting from the
neutralization value (NV) of the acid carrier. Following the
kneading process in the spiral kneader, 315 g of dough was
transferred into the Rheofermentometer, after which it was
stored at 30 °C for 3 hours. e maximum dough height Hm and
the height h at the end of the measuring cycle, among other
things, were determined during the dough resting period, and
the dough weakening coecient WC was calculated from them.
Effect of yeast-equivalent amounts of glutathione (.; .; .;
.; . mg/ g flour) on the dough Weakening coefficients WC
in the Rheofermentometer. To avoid structure-influencing effects
due to the yeast metabolism, a chemical leavening agent system
consisting of SAPP (Sodium Acid Pyrophosphate) and NaHCO₃
(sodium hydrogen carbonate, sodium bicarbonate) was used
instead of S. cerevisiae. Based on g of wheat flour, . g of
NaHCO₃ and . g of SAPP (neutralization value: ) were applied
to form an average CO volume of . ml (n ). The findings
indicate that an exponential increase (R .) in the dough
weakening coefficient of up to . is recorded as the amount of
added glutathione increases. The mean value with standard
deviation (n ) is plotted in each case
Approx. 670 ml of CO was formed during the three-hour
resting time in the Rheofermentometer. This corresponds
approximately to the volume released by 0.5 % of dry yeast or
1.5 % of fresh yeast. Thus the application of chemical leaven-
ing agents is a suitable way to simulate the gas-forming
properties of yeast [3]. It is clear from Figure 4 that there is
only a minimal increase in the dough weakening coefficient
from 0.0 % to 6.0 % for glutathione amounts of between 0.0–
58.6 mg per 100 g, and thus the dough system’s gas retention
properties are affected only slightly but nonetheless signifi-
cantly. Therefore this indicates that there is already a
pre-weakening of the dough structure, which possibly no
longer withstands the oven rise during the baking phase. In-
creasing the glutathione content to 76.2 mg per 100 g flour,
similar to the level that can enter the dough through yeasts
with a dead yeast cell proportion of approx. 60 %, causes the
dough weakening coefficient to increase to 20.5 %. This means
that the dough loses approx. on fifth of its maximum volume
as proong progresses. In addition to the loss of CO associated
with this, it indicates in particular a significant impairment
of the gluten network.
Impairment of the specific bread volume due to
yeast-equivalent amounts of glutathione
The effect of glutathione on the specific volume of wheat
bread was also studied on chemically leavened products. The
leavening agent system consisted of the acid carrier gluco no-
delta-lactone (neutralization value NV: 30) and the carbon
dioxide carrier sodium hydrogen carbonate (sodium bicar-
bonate, NaHCO). The effect of various different gas forma-
tion rates during the dough resting period in the presence of
GSH was also analyzed. Varying amounts of glucono-delta-
lactone, 1.5–12.9 g per 100 g of flour, were added to control
the gas formation rate (280–2160 ml of CO per hour). There-
fore the proportion of NaHCO needed to neutralize the acid
carrier varied between 0.45–3.87 g per 100 g of flour. After the
mixing/kneading process, dough pieces each weighing 250 g
were prepared, transferred into baking pans and stored for a
30-minute dough resting period in a proofing chamber. The
subsequent baking process took place at 230 °C for 18 min-
utes. After a cooling down period of 1 hour, the weight and
volume of the products were detected by means of a Volscan
(TexVol Instruments, Viken, Sweden) and the specific bread
volume (volume per unit weight) was calculated.
The curve profiles presented in Figure 5 show the effect on
the final product volume of yeast-equivalent amounts of
glutathione (0.0; 7.3, 27.6 and 76.2 mg per 100 g of flour) in
combination with various gas formation rates. Whereas a
maximum specific volume of 3.8 ml/g is attained for samples
without glutathione, the maximum amount of glutathione
leads to a 24 % reduction in baked product volume (2.9 ml/g).
Moreover it is evident that, apart from the samples with the
highest amount of GSH, an increase in the gas formation rate
causes maximization of the baked product volume. The
cause of this is the greater volume of CO, which logically in-
creases as the gas formation rate rises while the duration of
© TUM
www.bakingbiscuit.com 01/2016
RESEARCHRESEARCH
the dough resting period remains constant. Furthermore,
these experiments confirm the supposition by numerous re-
search groups that the structural impairment of baked goods
due to the freeze-thaw processes of dough pieces favors the
lysis of yeast cells, and the glutathione content in the dough
increases as a result [4-7].
Summary and Conclusion
The structure of wheat dough is permanently damaged by
the amount of yeast-equivalent glutathione. The quantities
of glutathione detected particularly in dry yeasts (5.4–81.2
mg per g of yeast (IDM)) cause a reduction in dough stability
(
–
67 %) and an increase in dough softening (+91 %). In par-
ticular the largest amount of GSH, 76.2 mg per 100 g of flour,
leads to very sticky/adhesive, moist doughs resulting in re-
stricted processability. Furthermore, it was possible to show
that the dough-weakening action of glutathione is limited
due to the finite number of thiol groups in the gluten. More-
over, the simulation of the fermentation/proofing process in
the Rheofermentometer shows that, measured in terms of the
dough weakening coefficient, pre-weakening of the dough
structure can occur as a result of the combination of GSH
and CO. The oven spring, i.e. the rapid CO release, steam
and ethanol vapor at the start of the baking phase, no longer
withstands this, leading to the collapse of the as yet un-
developed crumb structure. As a result the large amounts of
lysed cells, particularly in commercial dry yeasts, lead to a
severe reduction (max. 20 %) in the specific volume. On the
other hand the use of liquid or compressed yeast has no effect
on the structure of either the dough or the end product. The
only situation in which GSH levels like those in dry yeasts
can be reached is in the case of freeze-thaw processes and the
Presentation of the specific bread volume (ml/g) as a function of
the CO₂ formation rate. Wheat doughs were prepared using
yeast-equivalent quantities of glutathione: . mg (), . mg (),
. mg (
) and . mg () per g of flour. Various gas formation
rates were achieved by using a chemical leavening agent system
consisting of glucono-delta-lactone (.–. g per g of flour,
neutralization value: ) and sodium hydrogen carbonate (.–
.g per g of flour). The gas formation rates were calculated
on the basis of Rheofermentometer data. The curve shapes
indicate that the specific bread volume has a polynomial profile
(R. mg GSH .; R. mg GSH .; R. mg GSH .;
R. mg GSH .). The mean value with standard deviation is
plotted in each case (n )
associated dying-off of the cells, and so an appropriate pro-
cess control, the use of baking agents and/or cold-tolerant
yeasts is to be recommended. The principal finding from the
studies is that, in addition to the formation of CO as the pri-
mary function of yeasts in the manufacture of baked products,
other metabolites can also shape the structural properties of
doughs and of baked goods.
Literature references
1. Pyler, E.J. and L.A. Gorton, Yeasts, Molds and Bacteria, in
Baking Science and Technology: Fundamentals and Ingre-
dients 1988, Sosland Publishing Company: Kansas City.
2. Grosch, W. and H. Wieser, Redox Reactions in Wheat
Dough as Affected by Ascorbic Acid. Journal of Cereal
Science, 1999. 29(1): p. 1-16.
3. Verheyen, C., M. Jekle, and T. Becker, Effects of Saccharo-
myces cerevisiae on the structural kinetics of wheat dough
during fermentation. LWT – Food Science and Technolo-
gy, 2014. 58(1): p. 194-202.
4. Autio, K. and E. Sinda, Frozen Doughs: Rheological
Changes and Yeast Viability. Cereal Chemistry, 1992. 69(4):
p. 409-413.
5. Bayrock, D. and W.M. Ingledew, Fluidized bed drying of
baker’s yeast: moisture levels, drying rates, and viability
changes during drying. Food Research International,
1997. 30(6): p. 407-415.
6. Meziani, S., et al., Influence of yeast and frozen storage on
rheological, structural and microbial quality of frozen
sweet dough. Journal of Food Engineering, 2012. 109(3):
p. 538-544.
7. Wolt, M.J. and B.L. D’Appolonia, Factors Involved in the
stability of Frozen Dough. I. The Influence of Yeast Re-
ducing Compounds on Frozen-Dough Stability. Cereal
Chemistry, 1984. 61(3): p. 209-212. +++
— Authors: Christoph Verheyen (Photo),
Mario Jekle, Thomas Becker;
Technische Universität München (TUM),
Chair of Brewing and Beverage Technology,
Cereals Process Technology Work Group,
85354 Freising, Germany,
E-mail: christoph.verheyen@tum.de
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