International Journal of Food Science and Technology 2018, 53, 1597–1602 1597

Original article
Wheat preharvest herbicide application, whole-grain flour
properties, yeast activity and the degradation of glyphosate in
bread

Pierre Gelinas,* Fleur Gagnon & Carole McKinnon

Saint-Hyacinthe Research and Development Centre, Agriculture and Agri-Food Canada, Saint-Hyacinthe, Quebec J2S 8E3, Canada
(Received 7 December 2017; Accepted in revised form 8 January 2018)

Summary The aim of this study was to determine the effects of wheat preharvest application of a glyphosate-based
herbicide (Roundup WeatherMaxÒ with TransorbÒ 2 Technology) on whole-grain flour composition and
properties, including yeast activity. The effect of dough fermentation on the degradation of herbicide resi-
dues was also estimated. Grain samples from two hard red spring wheat varieties exceeded the maximum
residue limits (5 mg kg 1) in Canada. Glyphosate had minor effects on wheat kernels composition and
properties, including fructans content and yeast gassing power. No degradation of RoundupÒ or pure gly-
phosate was seen after dough fermentation for up to 4 h and baking. These results call for more scientific
studies on glyphosate residues in wheat.
Keywords Bread, glyphosate, herbicides, wheat, yeast.

currently available on the market (Castro et al., 2014;

Introduction
Glyphosate-based herbicides may be applied shortly
before wheat harvesting to allow plant drying in the
field so kernels would be easier to harvest without
compromising the yield when the crop has 30% or less
moisture or typically 3–5 days before normal swath
(Darwent et al., 1994; Monsanto Canada, 2012). When
used according to label directions, glyphosate would
not present risks to human health or the environment
(Williams et al., 2000; Harris & Gaston, 2004; Sviridov
et al., 2014; Health Canada, 2017). However, Cuhra
et al. (2016) recommended that, in the future and
whenever possible, glyphosate-based herbicides should
play a reduced role in agriculture. According to CFIA
(2017), in wheat, the maximum residue limit (MRL)
set by Health Canada is 5 mg kg 1 based on the sum
of glyphosate and its primary degradation product
aminomethylphosphonic acid (AMPA). As stressed by
Blackshaw & Harker (2016), glyphosate degradation in
soils would result from microbial activity. Wheat ker-
nels harvested after glyphosate application would be
safe because they contained less than 5 mg kg 1 resi-
dues (Cessna et al., 1994; Wigfield et al., 1994). How-
ever, the latter studies were performed on earlier
versions of commercial glyphosate that contained dif-
ferent adjuvants or surfactants compared to products
*Correspondent: E-mail: pierre.gelinas@agr.gc.ca
Rissoli et al., 2016).
Glyphosate residues in mature wheat would have lit-
tle effects on the bread-making potential of white
flour, including dough mixing tolerance and bread vol-
ume (Darwent et al., 1994; Manthey et al., 2004; Cra-
ven et al., 2007). However, whole-grain flour would be
more contaminated than white flour because pesticides
residues accumulate on the external portions of wheat
kernels (Amvrazi, 2011). Glyphosate preharvest appli-
cation on wheat crops might result in changes in ker-
nels constituents due to the sudden stop of the plant
growth. Bresnahan et al. (2003) reported that glypho-
sate-treated kernels contained higher levels of shikimic
acid, a phenolic acid showing high antioxidant activity.
After glyphosate application, there is a possibility that
water-soluble fibre constituents such as fructans would
be retained in wheat kernels instead of migrating
towards other plant tissues (Pollock & Cairns, 1991;
Li et al., 2013). As prebiotics, fructans would stimulate
the growth of probiotic micro-organisms although it
might also cause gastrointestinal discomfort to some
individuals (Gelinas et al., 2016).
In liquid media, inhibition assays involving baker’s
yeast, Saccharomyces cerevisiae, have been proposed as
a cheap alternative to toxicity bioassays to determine
the degradation of weed killers, fungicides and insecti-
 c & Durovic-Pejcev,
cides (Esteve et al., 2009; Dordevi

doi:10.1111/ijfs.13741
© 2018 Her Majesty the Queen in Right of Canada International Journal of Food Science and Technology © 2018 Institute of Food Science and Technology
Reproduced with the permission of the Minister of Agriculture and Agri-Food Canada.
1598  linas et al.
Glyphosate in wheat and yeast fermentation P. Ge
2015; Ehrampoush et al., 2017). Inhibition of baker’s
yeast cells growth would vary much according to the
herbicide (Sharma et al., 2005; Regueiro et al., 2015).
Compared to pure glyphosate, commercial prepara-
tions of RoundupÒ may contain as much as 20% sur-
factants and would have higher inhibitory potential on
wine yeast and dairy starters (Braconi et al., 2006;
Clair et al., 2012). According to Low et al. (2005),
yeast would metabolise about 20% of pure glyphosate
in dough fermented for only 1 h. To our knowledge,
no scientific evidence has been reported on the effect
of commercial glyphosate preparation on baker’s yeast
fermentative activity. Little information is also avail-
able on the impact of yeast fermentation and bread
baking on the degradation of glyphosate.
The aim of this study was to determine the effects of
wheat preharvest glyphosate application on the com-
position and properties of whole-grain flour, especially
the fermentative activity of baker’s yeast and the
degradation of pure glyphosate and RoundupÒ in
dough. Due to the current awareness for pesticides in
foods, this study was designed to advance scientific
knowledge on glyphosate residues in wheat flour and
bread.
Materials and methods
Whole-grain wheat flour samples
Two hard red spring wheat varieties (Major; Walton)
for bread making were grown in a series of 1 m2 par-
cels (450 plants per m2) at different farming sites in the
province of Quebec, respectively, in Alma (Lac-Saint-
Jean) and Saint-Anicet (Monteregie-Ouest). Nitrogen
(100 kg ha 1) was applied in the field. Concentration
(1.67 L ha 1 corresponding to 0.82 kg ha 1) and tim-
ing of glyphosate-based herbicide (RoundupÒ) applica-
tion (hard-dough Zadock growth stage 87) were based
on recommendations of the manufacturer (Roundup
WeatherMaxÒ with TransorbÒ 2 Technology; Mon-
santo Canada, Winnipeg, MB, Canada). For Major,
RoundupÒ was sprayed on 11 September 2015, and
kernels were harvested on 16 September. For Walton,
RoundupÒ was applied on 7 August and harvesting
was performed on 20 August. Wheat samples were
stored in bags at room temperature for about
4 months and, in preparation for analyses, later kept
at 10 °C. Whole-grain flour (wholemeal) was prepared
by milling 500 g of grain with a Perten disc mill (disc
#2; Model 3301; Perten Instruments, Huddinge, Swe-
den) equipped with a mill feeder.
Whole-grain analysis (except glyphosate)
Protein content (dry basis; N 9 5.7) was measured
with the combustion method (Vario Max cube;
International Journal of Food Science and Technology 2018
Elementar Analysensysteme GmbH, Hanau, Ger-
many). Falling number was determined according to
AACC Method 56-81.03 (AACC, 2009). Total dry
gluten content in wholemeal was assayed with the
Glutomatic based on AACC Method 38-12.02 (AACC,
2009). Flour moisture was measured according to
AACC Method 44-15.02 (AACC, 2009). As described
in Gelinas et al. (2016), fructans analysis was per-
formed with the Fructan Assay Procedure (K-Fruc;
Megazyme International Ireland, Wicklow, Ireland)
with pretreatment of samples with a-galactosidase to
eliminate galactosyl-sucrose oligosaccharides. Analyses
were performed in duplicate, except protein and gluten
contents which were determined in triplicate.
Dough rheology
Farinograms were performed in duplicate according to
AACC Method 54-21.02 (AACC, 2009), except analy-
ses were performed at 25 °C instead of 30 °C (Gelinas
& McKinnon, 2013).
Bread making
As described in Gelinas et al. (2016), bread was pre-
pared in duplicate (two mixed doughs) according to
AACC Method 10-10.03 (AACC, 2009), with a 3-h
fermentation period at 35 °C. Formulation included
wholemeal (100 g), water (variable), shortening (3 g),
salt (1.5 g), dry yeast (1.2 g) and ascorbic acid
(50 mg kg 1 flour). Dough water absorption (14%
moisture, flour basis) was determined according to
farinogram plus 0.5%. After cooling for 1 h, bread
volume was determined with a laser volumeter (Vols-
can Profiler VSP300; Stable Micro Systems Ltd.,
Godalming, Surrey, UK) based on AACC Method 10-
14.01 (AACC, 2009).
Gas production
In duplicate, yeast gassing power (gas pressure) was
determined with the Risograph (National Manufactur-
ing, TMCO, Inc., Lincoln, NE, USA) using RisoSmart
version 1.0.233 from 2005, the software provided with
the equipment. Dough was prepared from 100 g whole-
meal as described in bread-making assays. Immediately
after mixing, 2 9 25-g dough portions were placed into
Risograph jars. Equipment was programmed to get 1
measure per min for 3 h at 38 °C, except for assays on
glyphosate degradation which were performed for 4 h.
Means were calculated from 4 data.
Glyphosate degradation
In duplicate, experiments were performed as reported
for gas production assays, except dough prepared from
© 2018 Her Majesty the Queen in Right of Canada International Journal of Food Science and Technology
© 2018 Institute of Food Science and Technology

200 g flour was spiked with Roundup WeatherMaxÒ
with TransorbÒ 2 Technology (13.5 or 27 mg kg 1
flour); wholemeal was replaced by commercial white
flour (Robin HoodÒ Best for Bread Homestyle White
flour; Smucker Foods of Canada Corp., Montreal,
QC, Canada). In the Risograph, 8 9 25-g dough por-
tions were fermented at 38 °C and gas production was
determined after 1, 2, 3 or 4 h. Additional experiments
were performed with yeasted or nonyeasted dough
supplemented with pure glyphosate (13.5 mg kg 1;
Sigma) instead of RoundupÒ. At the completion of
each fermentation period, doughs were removed from
the Risograph jar, placed in a pan, baked at 204 °C
for 19 min (Fisher & Paykel Aero Tech USA, Hunt-
ingdon Beach, CA, USA), cooled on a grid for 1 h,
packaged into plastic bags and frozen at 45 °C until
glyphosate analysis. After thawing, bread samples were
comminuted by hand, left to dry for 48 h at room
temperature and milled with a coffee mill (Brau Mex-
ico, Naucalpan de Juarez, Mexico).
Glyphosate analysis
In duplicate, 1 g wholemeal or dehydrated bread was
extracted for 2 h with 10 mL water at room tempera-
ture, with stirring at 200 rpm. After centrifugation
(5750 g), 50 lL of supernatant was combined with
1 mL of 0.2 M borate buffer (pH 9) and 200 lL of a
fresh solution of FMOC.Cl in acetone (2 mg mL 1),
vortexed and filtered after 30 min. Adapted from a
procedure recommended by Phenomenex Inc. (Ter-
rance, CA, USA), glyphosate and AMPA were deter-
mined with a Knauer Smartline HPLC system
(Berlin, Germany) using a 250 9 4.6 mm Gemini C18
column (Phenomenex Inc.) with 20 mM ammonium
bicarbonate/acetonitrile/methanol 70/15/15 to 65/17.5/
17.5 for 12 min, then 10/45/45 for 10 min at
0.6 mL min 1. Detection was performed with a 2475
fluorescence detector (Waters, Milford, MA, USA)
with excitation and emission at 360 and 410 nm,
respectively. Calibration curve was prepared with gly-
phosate and AMPA standards (0.1–4 lg mL 1) and
detection limits were 0.05 lg mL 1 and 0.1 lg mL 1,
respectively. As determined in triplicate, recovery of
glyphosate on dry basis in wholemeal and bread was
Table 1 Effect of wheat preharvest RoundupÒ application on wholemeal composition
Wheat variety Roundupâ application Glyphosate residues (mg kg 1)
c a
Major No 0 404
Yes 11.1a
Walton No 0c
Yes 6.1b
Glyphosate concentrations are reported on flour weight basis; no AMPA was detected. Mean data (dry basis) were obtained from two repetitions,
except protein and gluten (three repetitions). Within columns, means followed by the same letter are not significantly different at the P < 0.05 level.
© 2018 Her Majesty the Queen in Right of Canada International Journal of Food Science and Technology
© 2018 Institute of Food Science and Technology
 linas et al.
Glyphosate in wheat and yeast fermentation P. Ge 1599
92  3% and 95  11%, respectively; for AMPA,
recovery was 94  1% and 80  1%.
Statistical analyses
One-way analysis of variance procedure was per-
formed with SAS (version 9.3, TS1M2, 2012, SAS
Institute Inc., Cary, NC, USA). The least significant
difference (LSD) test was used to determine the differ-
ences between treatments (P < 0.05).
Results
Effect of wheat preharvest Roundupâ application
Total glyphosate residues in wheat kernels harvested
after RoundupÒ treatment were 11.1 and 6.1 mg kg 1,
respectively, for Major and Walton (Table 1). What-
ever the sample, AMPA was below the detection limit.
With Major, the wheat variety with the highest poten-
tial for pan bread, preharvest application of
RoundupÒ had little effect on the composition of the
kernels, including falling number as well as protein,
gluten and fructans contents (Table 1), and whole-
grain flour water absorption, dough mixing stability
and yeast gassing power (Table 2). Similar results were
seen with Walton, a variety with lower bread-making
potential, except preharvest RoundupÒ application
had a negative effect on yeast fermentative activity.
Depending on the wheat variety, bread volume varied
significantly, but no trend could be seen according to
RoundupÒ treatment (Table 2).
Effect of yeast fermentation on glyphosate degradation
To eliminate the possibility that whole-grain flour inhi-
bits yeast activity (Gelinas et al., 2016), additional
experiments on gassing power were performed with
commercial white flour, free of bran. The degradation
of glyphosate was also checked in dough spiked with
RoundupÒ at 13.5 or 27 mg kg 1 flour (Table 3).
After dough fermentation for up to 4 h, yeast gassing
power was not inhibited by glyphosate and no degra-
dation of the herbicide was seen. Compared to spiked
dough, about 15% less glyphosate was recovered in
Falling number (s) Protein (%) Gluten (%) Fructans (%)
b a
15.8 14.0 1.08a
402a 16.2a 13.8a 0.94a
412a 13.2d 11.3b 1.05a
406a 13.8c 11.0b 1.15a
International Journal of Food Science and Technology 2018
1600  linas et al.
Glyphosate in wheat and yeast fermentation P. Ge

Table 2 Effect of wheat preharvest RoundupÒ application on wholemeal dough mixing properties, gas production and bread volume

Dough

â
Wheat variety Roundup application Water absorption (%) Mixing stability (min) Gas production (mL) Bread specific volume (mL g 1)

Major No 71.0a 8.2a 205.5a 6.0a

Yes 71.4a 7.8a 201.1a 5.7b
Walton No 62.6b 8.6a 202.5a 3.6d
Yes 63.0b 8.1a 181.5b 4.2c

Mean data were obtained from 2 data (2 repetitions 9 1 sample), except total gas production after 3 h at 38 °C (2 repetitions 9 2 samples). Within
columns, means followed by the same letter are not significantly different at the P < 0.05 level.

Table 3 Effect of the addition of glyphosate preparation Table 4 Effect of the addition of pure glyphosate (13.5 mg kg 1
(RoundupÒ) in dough made with white commercial flour on yeast flour) in yeasted or nonyeasted dough on the degradation of glypho-
gassing power and the degradation of glyphosate after fermentation sate after fermentation and baking
and baking
Yeasted Dough fermentation Residual glyphosate
Dough Added Residual glyphosate Dough time (h) after baking (mg kg 1)
fermentation glyphosate after baking Gas
time (h) (mg kg 1) (mg kg 1) production (mL) No 0 12.9a
1 12.6a
0 0 0d 0e 2 14.2a
13.5 12.7c 0e 3 13.3a
27 23.5ab 0e 4 11.9a
1 0 0d 66.8d Yes 0 11.4a
13.5 11.5c 62.3d 1 11.8a
27 21.3bc 67.4d 2 12.0a
2 0 0d 167.4c 3 12.5a
13.5 11.9c 161.3c 4 12.2a
27 23.1ab 168.7c
3 0 0d 274.1b Glyphosate concentrations are reported on flour weight basis; no
13.5 10.2c 267.1b AMPA was detected. Doughs were prepared with commercial white
27 24.2a 275.6b flour. Mean data were obtained from 2 data (2 repetitions 9 1 sample).
4 0 0d 363.7a Within column, means followed by the same letter are not significantly
13.5 11.7c 357.3a different at the P < 0.05 level.
27 21.6ab 364.9a

Glyphosate concentrations are reported on flour weight basis; no

AMPA was detected. Mean data for glyphosate were obtained from 2
data (2 repetitions 9 1 sample); data for total gas production at 38 °C
were obtained from 4 data (2 repetitions 9 2 samples). Within col-
umns, means followed by the same letter are not significantly different
at the P < 0.05 level.
bread. The latter was attributed to losses during
extraction because no AMPA, the main degradation
product of glyphosate, was found. In yeasted or non-
yeasted dough spiked with pure glyphosate instead of
RoundupÒ, no herbicide degradation was seen after
fermentation and baking (Table 4); no AMPA was
found, and yeast gassing power was not inhibited by
pure glyphosate (data not shown).
Discussion
In Canada, MRL for pesticides corresponds to the
maximum amount of residues that will not be a con-
cern to human health (Health Canada, 2015).
According to our assays, wheat harvested after appli-
cation of a recent RoundupÒ commercial formulation
had 6 to 11 mg glyphosate kg 1, which is higher than
MRL or 5 mg kg 1 (CFIA, 2017). There is a small
possibility that the highest value was found in grains
where RoundupÒ was used closer to the harvest date,
due to lower degradation of glyphosate. Such concen-
trations are higher than those reported in the literature
with earlier and different commercial glyphosate for-
mulations (Cessna et al., 1994; Wigfield et al., 1994).
Recently, high levels of glyphosate residues, up to
15 mg kg 1, were found in grain samples others than
wheat, specifically in genetically modified soya bean
(Bøhn et al., 2014). In several countries, glyphosate
tolerance levels have been periodically increased to
compensate for higher residue levels found in grain,
including wheat (Benbrook, 2016; Cuhra et al., 2016).
Recently, wheat preharvest application of glyphosate
has been prohibited in Austria (European Glyphosate
Task Force, 2014). According to Cuhra (2015), very

International Journal of Food Science and Technology 2018 © 2018 Her Majesty the Queen in Right of Canada International Journal of Food Science and Technology
© 2018 Institute of Food Science and Technology

little scientific reports have been published on glypho-
sate residues in glyphosate-tolerant crops. We consider
that our data on glyphosate residues in wheat kernels
should be interpreted with caution because they have
been obtained from only two independent assays in
the field.
Baker’s yeast has been proposed as a model in toxi-
cological studies because it is sensitive to toxic sub-
stances such as herbicides (Braconi et al., 2016).
Compared to pure glyphosate, commercial RoundupÒ
preparations would have much higher potential for
weed control because they contain highly specific sur-
factants (Green & Beestman, 2007; Castro et al.,
2014). In our hands, the fermentative activity of baker’
yeast in dough containing 13.5 or 27 mg RoundupÒ
kg 1 flour was not inhibited, confirming observations
by others with wild-type wine yeast (Braconi et al.,
2006) and dairy food starters such as yeast-like Geotri-
chum candidum (Clair et al., 2012). Contrary to data
reported by Low et al. (2005), our results also suggest
that dough fermentation would not be an effective
technique to degrade glyphosate residues in wheat.
Despite some differences seen depending on the
wheat sample, the presence of 6–11 mg kg 1 of gly-
phosate in whole-grain flour had minor effects on
dough and bread properties, confirming results
obtained by others with lower levels of glyphosate in
bran-free flours (Darwent et al., 1994; Manthey et al.,
2004). Not reported previously, glyphosate application
in the field did not change the level of fructans in
wheat kernels. More data are needed on the levels of
glyphosate residues in a larger set of wheat samples
treated with different formulations of glyphosate-based
herbicides.
Conclusion
Preharvest Roundup WeatherMaxÒ with TransorbÒ 2
Technology application on two bread wheat varieties
had minor effects on whole-grain composition and
dough properties, and no specific trend could be seen.
High levels of glyphosate residues, 6–11 mg kg 1, were
found in the treated wheat kernels, but the latter
results must be confirmed with more field assays.
Yeast gassing power was not inhibited in dough spiked
with RoundupÒ or pure glyphosate, and no degrada-
tion of the herbicide was seen in dough fermented for
up to 4 h.
Acknowledgments
The authors would like to thank Elisabeth Vachon
(Moulins de Soulanges, Saint-Polycarpe, Quebec,
Canada) and Jean Cantin (MAPAQ, Saint-Hyacinthe,
Quebec, Canada) for supervising field assays and pro-
viding grain samples.
© 2018 Her Majesty the Queen in Right of Canada International Journal of Food Science and Technology
© 2018 Institute of Food Science and Technology
 linas et al.
Glyphosate in wheat and yeast fermentation P. Ge 1601
Conflict of interest
There is no conflict of interest to declare.
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