International Journal of Food Microbiology 112 (2006) 75 – 80

www.elsevier.com/locate/ijfoodmicro

Development of a new oat-based probiotic drink

Angel Angelov a,⁎, Velitchka Gotcheva a , Radoslava Kuncheva a , Tsonka Hristozova b
a
Department of Biotechnology, University of Food Technologies, 26 Maritza Blvd, 4002 Plovdiv, Bulgaria
b
Institute of Microbiology, Bulgarian Academy of Sciences, 26 Maritza Blvd, 4002 Plovdiv, Bulgaria

Received 31 May 2005; received in revised form 8 February 2006; accepted 27 May 2006

Abstract

In the present work, a whole-grain oat substrate was fermented with lactic acid bacteria to obtain a drink, combining the health benefits of a
probiotic culture with the oat prebiotic beta-glucan. The levels of several factors, such as starter culture concentration, oat flour and sucrose
content, affecting the fermentation process, were established for completing a controlled fermentation for 8 h. The viable cell counts reached at the
end of the process were about 7.5 × 1010 cfu ml− 1. It was found that the addition of sweeteners aspartame, sodium cyclamate, saccharine and
Huxol (12% cyclamate and 1.2% saccharine) had no effect on the dynamics of the fermentation process and on the viability of the starter culture
during product storage. Beta-glucan content in the drink (0.31–0.36%) remained unchanged both throughout fermentation and storage of the
drink. The shelf life of the oat drink was estimated to 21 days under refrigerated storage.
© 2006 Elsevier B.V. All rights reserved.

Keywords: Oat fermentation; Oat drink; Probiotic; Lactic acid bacteria; Beta-glucan

1. Introduction
Continuous development of new functional foods is the
response of science and industry to the increased consumer
awareness regarding health and the role of foods for improving
quality of life (Bland and Medcalf, 1994; Blades, 2000;
Verschuren, 2002). Long known for its benefits, oats is be-
coming popular as part of a healthy diet and new oat products
emerge at the functional food market.
Most probiotic foods at the markets worldwide are milk-
based and very few attempts are made for development of
probiotic foods using other fermentation substrates such as
cereals. Their large distribution and important nutritive value
have focused the attention on their use as raw materials for the
development of new fermented functional foods. Oats and
barley are major sources of beta-glucan, recognized as the main
functional component of cereal fibers. Studies have indicated
the hypocholesterolaemic effect of this compound, leading to
20–30% reduction of LDL-cholesterol, and to an expected
overall effect of reduced cardiovascular disease risk (Stark and
⁎ Corresponding author. Fax: +359 32 644 102.
E-mail address: angelov@uft-bio.com (A. Angelov).
0168-1605/$ - see front matter © 2006 Elsevier B.V. All rights reserved.
doi:10.1016/j.ijfoodmicro.2006.05.015
Madar, 1994; Wrick, 1994; Gallaher, 2000). The cereals with
highest beta-glucan content are oats and barley (Wood and Beer,
1998; Manthey et al., 1999). In 1997, the FDA of US has
officially acknowledged as functional the products made of
whole-grain oats or oat fiber with a minimum of 0.75 g beta-
glucan/serving size. The low glycaemic index of oat products is
especially important for diabetics, and the ingestion of beta-
glucan-containing viscous foods is reported to affect the level of
fat emulgation in the gastro-intestinal tract and reduces lipase
activity (Anderson and Bridges, 1993; Wrick, 1993, 1994; Stark
and Madar, 1994). The release of low-molecular fatty acids
during beta-glucan fermentation in the colon preconditions its
potential anti-carcinogenic effect (Oku, 1994; Salminen et al.,
1998; Gallaher, 2000). In addition, beta-glucan is known as
prebiotic, stimulating the growth of some beneficial residential
colon microorganisms such as bifidobacteria (Jaskari et al.,
1998; Wood and Beer, 1998).
Marklinder and Lonner (1992), as well as Johansson et al.
(1998) reported that after appropriate processing, oats is a
suitable substrate for fermentation with lactic acid bacteria.
Enzymatically treated oat bases have been developed and
applied by Màrtensson et al. (2001, 2002) as substrates for lactic
acid fermentation with dairy starter cultures.
76 A. Angelov et al. / International Journal of Food Microbiology 112 (2006) 75–80
The aim of the present work was to develop a synbiotic
functional drink from oats by combining a probiotic starter
culture and whole-grain oat substrate.
2. Materials and methods
2.1. Starter culture
A probiotic strain — Lactobacillus plantarum B28 was used
in the study. The strain was isolated from a cereal-based fermented
drink and selected as probiotic in previous studies (Gotcheva
et al., 2000, 2002). The strain was maintained on MRS-agar (E.
Merck, Darmstadt, Germany) at 4–6 °C. Starter culture was
obtained by overnight incubation at 37 °C in MRS-broth. The
culture was centrifuged (4500 g, 10 min, 4 °C), washed in distilled
water and re-suspended in distilled water to its original volume.
The cell count of the culture was 1011 cfu ml− 1.
2.2. Oat substrate
The oat mash used as a substrate was prepared from whole-
grain oat flour “Mina Prim” (oat cultivar “Mina”, Blagopoluchie,
Bulgaria) and tap water in different concentrations 4.0, 5.5 and
7.0% (w/v). Food grade sucrose in concentrations of 1.0, 1.5 and
2.0% (w/v) was added to the slurry. Test was performed also with
adding combinations of sucrose and sweeteners as follows: A)
1.5% sucrose (control), B) 1.5% sucrose + 0.5% (v/v) Huxol
(Veelman, Germany) and C) 1.5% sucrose + 0.04% (w/v) sodium
cyclamate (E952) + 0.005% (w/v) saccharine (E954) + 0.008%
(w/v) aspartame (E951). All sweeteners are products of Esarom,
Essenzanfabrik G.m.b.H., Austria. Huxol contains 12% cycla-
mate and 1.2% saccharine. The slurry was then heated at 95 °C
for 10 min and cooled to 37 °C.
2.3. Fermentation and storage
The oat mash was inoculated with 1%, 5% and 10% (v/v)
starter culture, which gave inoculation levels of 9.6 × 107,
2.8 × 108 and 7.1 × 108 cfu ml− 1, respectively. Fermentation was
carried out at 37 °C for 6–10 h. Storage observations were
carried out at 4–6 °C for 24 days.
2.4. Viable cell enumeration
Enumeration of viable cells of L. plantarum B28 was
performed by estimation of colony forming unit number on
MRS-agar plates (medium pH 5.7) after incubation at 37 °C for
48 h.
2.5. Determination of pH, titratable acidity and dry matter
For the measurement of pH ORION 420A pH-meter
(Thermo Electron Corp., Waltham, MA, USA) was used, and
titratable acidity (TA) was determined by titrating 10-ml
samples with 0.1 N NaOH, phenolphthalein used as indicator.
TA was expresses as °N (degrees Neuman) (Vangelov and
Karadjov, 1993).
2.6. Determination of beta-glucan
Beta-glucan content was evaluated by an enzymatic kit
according to the manufacturer's instructions (Megazyme
International Ireland Ltd., Co. Wicklow, Ireland). Absorbance
was measured by spectrophotometer Anthelie Light (Secomam,
Ales, France) at λ = 510 nm.
2.7. Estimation of shelf life
The shelf life of the fermented oat drink was defined as the
period of refrigerated storage (4–6 °C) during which pH
remained above 4.0 and the number of viable cell counts was
above 106 cfu ml− 1. Refrigerated storage was carried out for
24 days with periodical observations of pH, TA, beta-glucan
content and the viability of the starter culture.
2.8. Statistical analysis
Each experiment was performed in four separate trials.
Results represent the means with standard deviations. Data was
submitted to one-way analysis of variance (ANOVA) with a
least significant difference of 95%.
3. Results and discussion
Based on previous research, the probiotic strain L. plantarum
B28 was selected for fermentation of the heat treated oat mash.
The appropriate concentrations of oat flour, starter culture and
sucrose had to be estimated, as well as sweetener effect on
fermentation and storage, and the shelf life of the drink.
3.1. Starter culture concentration
The oat mash was inoculated with 1.0, 5.0 and 10% starter
culture suspension of L. plantarum B28 aiming to achieve the
required levels of viable cells in probiotic products — at least
106–107 cfu ml− 1 (Mattila-Sandholm et al., 1999). Short
fermentation time was preferable in order to minimize the risk
of contamination. Results from the fermentations are presented
in Fig. 1. The application of 5.0% or 10% inoculum resulted in
reaching the desired pH levels of 4.0–4.5 in significantly shorter
time — 8 and 4 h, respectively, compared to 1.0% inoculum.
Values of TA around 1.5°N were reached for 6 and 8 h with 10.0
and 5.0% inoculum, respectively. These results show that in
terms of acid formation 8 h is an appropriate fermentation time.
Marklinder and Lonner (1992) achieved pH 4.3–4.5 for 6 h
when fermenting oatmeal soups with L. plantarum strains,
while Martensson et al. (2002) reported pH 3.9–4.5 after much
longer fermentation (16 h) of an oat base with commercial
mixed dairy cultures. The different fermentation rates could be
attributed both to strain specificities and differences of oat
media. With 5.0 and 10% inoculum concentrations applied, in
6 h the levels of viable cell counts reached 9.3 × 109 and
7.5 × 1010 cfu ml− 1, respectively. As these levels were above the
required for probiotic products, it was more reasonable to use
5.0% starter culture concentration.
 A. Angelov et al. / International Journal of Food Microbiology 112 (2006) 75–80 77

Fig. 1. Effect of starter culture concentration on oat mash fermentation. Changes of pH (A), titratable acidity (B) and viable cell counts (C).

3.2. Concentration of oat flour
Concentration of whole-grain oat flour in the mash is
important for the fermentation and the beta-glucan content in
the oat products. Results from testing three oat flour concentra-
tions (4.0, 5.5 and 7.0%) are presented in Fig. 2. Acid production
was lowest when oat flour consisted of 7.0% mash and pH values
reached 4.52 in 8 h. With the lower flour concentrations applied,
pH values below 4.5 were registered in about 6 h of fermen-
tation. Other authors reported pH 4.0–4.5 after 16 h fermentation

Fig. 2. Effect of the oat flour concentration on oat mash fermentation (average inoculum level — 2 × 108 cfu ml− 1). Changes of pH (A), viable cell counts (B) and beta-
glucan content (C).
78 A. Angelov et al. / International Journal of Food Microbiology 112 (2006) 75–80

Fig. 3. Effect of sucrose concentration on oat mash fermentation (average inoculum level — 2 × 108 cfu ml− 1). Changes of pH (A), titratable acidity (B) and viable cell
counts (C).

of different oat substrates (Màrtensson et al., 2000) probably due was 2.65 and 2.41 log orders, respectively. These results show that
to the higher dry matter content of the medium — 16–18%. The 4.0–5.5% oat flour content in the mash is more appropriate for
increase of flour content influenced the viable cell counts as well. intensive fermentation. Marklinder and Lonner (1992) fermented
When 4.0 and 5.5% oat flour were used, the increase of cell counts 5% w/w oatmeal soups for 6 h to reach pH 4.0–4.5 and 15 h to

Fig. 4. Effect of sweeteners on oat mash fermentation (average inoculum level — 2 × 108 cfu ml− 1). Changes of pH (A), titratable acidity (B), viable cell counts (C).
Variant A: sucrose, Variant B: sucrose + Huxol, Variant C: sucrose + aspartame + sodium cyclamate + saccharine.
 A. Angelov et al. / International Journal of Food Microbiology 112 (2006) 75–80
Fig. 5. Laboratory model for the production of an oat drink. Changes of pH and titratable acidity (A), viable cell counts and beta-glucan concentration (B).
register similar cell counts. The initial beta-glucan content in the
oat mashes obtained with different flour concentrations was 0.23%
for 4.0% oat flour, 0.36% for 5.5% oat flour and 0.41% for 7.0%
oat flour, respectively. No significant changes of these concentra-
tions were found during fermentation, which indicated that the
starter culture did not ferment beta-glucan. Since differences
between pH and cell count changes at oat flour concentrations 4.0
and 5.5% were not significant, 5.5% was preferred in order to
obtain higher beta-glucan content in the fermented products.
3.3. Sucrose concentration
Sucrose in concentrations of 1.0, 1.5 and 2.0% was added
into the oat mash to increase fermentation rate — results are
presented in Fig. 3. With 2.0% sugar added, pH 4.5 was reached
on the 6th hour, and with 1.5% sucrose such value was
registered 0.5 h later. Acid production was quite slower with
1.0% sucrose in the medium and pH 4.54 was reached after 8 h.
The initial viable cell counts in the three product variants were
within 1.3 × 108–2.1 × 108 cfu ml− 1. At the end of the fermen-
tation, significant differences were observed between cell count
increases in the variants, with the highest of 2.81 log orders for
2.0% sucrose. The insignificant differences in pH dynamics
between the variants with 1.5 and 2.0% sucrose and the ob-
served viable cell counts increase support the conclusion that
the addition of 1.5% sucrose to the oat mash is sufficient for
obtaining a probiotic product.
Fig. 6. Establishment of the oat drink shelf life under refrigerated storage (4–6 °C).
79
3.4. Effect of sweeteners
The addition of 1.5% sucrose to the oat mash was not sufficient
to provide acceptable sweetness of the oat products. Therefore,
two variants of oat mashes with the addition of different sweet-
eners (Var. B and Var. C) were fermented. Variant A was used as
control. Results of the experiment are presented in Fig. 4.
Changes of pH were similar for all three variants, reaching values
of 4.48 to 4.55 at the 6th hour. TA changed correspondingly, with
final levels of 1.4–1.6°N. Viable cell counts in the three mashes
reached 4.3 × 1010–8.5 × 1010 cfu ml− 1 at the end of the fermen-
tation. Results showed that sweeteners applied had no effect on
the fermentation. Similarly, Vinderola et al. (2002) found no effect
of aspartame on the growth of probiotic lactic acid bacteria. Of the
three combinations, variant C was preferred for application in oat
drinks because of the better taste and suitability for the diet of
diabetics.
3.5. A laboratory model for the production of an oat-based
drink
The laboratory model for oat drink production was based on
the above estimated conditions: concentration of L. plantarum
A28 starter culture — 5.0%; oat flour — 5.5%; sucrose —
1.5%; combination of aspartame, sodium cyclamate and
saccharine, fermentation time — 8 h. The aim was to achieve
fast fermentation for a limited risk of microbial contamination,
80 A. Angelov et al. / International Journal of Food Microbiology 112 (2006) 75–80
high viable cell concentration to ensure the probiotic effect of the
oat drinks and appropriate beta-glucan content for a functional
effect in the consumer's system. Results are presented in Fig. 5.
Dynamics of pH and TA complied with those observed in the
previous stages of the work. The appropriate fermentation time
was confirmed to be 6–8 h, ensuring pH of 4.0–4.5. Viable cell
count increase of 2.47 log orders ensured about 7.5 × 1010 cfu
ml− 1 in the drink. Beta-glucan content in the oat mash remained
within 0.31–0.36%, which proved that this functional ingredient
was not fermented by the starter culture.
3.6. Estimation of shelf life
Results from the shelf life study are presented in Fig. 6.
Titratable acidity and pH of the drink remained within the
desired ranges for 21 days. During this period, viable cell counts
decreased with about 1 log order (from 7.3 × 10 10 to
9.3 × 109 cfu ml− 1). Sweeteners did not affect the viability of
the starter culture during storage. Viable cell counts at the end of
the storage were 106–107 cfu ml− 1, which ensured the probiotic
characteristics of the drink for 21 days. Beta-glucan content
remained unchanged within the storage time.
4. Conclusions
The study on several factors affecting the obtaining of a
functional oat drink established the appropriate starter culture
concentration, oat flour and sucrose content for completing a
controlled fermentation for 8 h. It was found that the addition of
aspartame, sodium cyclamate, saccharine and Huxol had no
effect on the dynamics of the fermentation process and on the
viability of the starter culture during product storage. The
chosen oat flour concentration contributed to 0.31–0.36% beta-
glucan in the drink, which remained unchanged both during
fermentation and storage. This would ensure its beneficial
health effect at regular consumption of the drink. The shelf life
of the synbiotic oat drink was estimated to 21 days under
refrigerated storage.
Acknowledgments
The study was carried out under project CC 1004/2000
funded by the Bulgarian Ministry of Education and Science.
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