World Journal of Microbiology and Biotechnology (2022) 38:133

https://doi.org/10.1007/s11274-022-03322-4

ORIGINAL PAPER

Inulin addition improved probiotic survival in soy‑based fermented

beverage
Carolina Iraporda1 · Irene A. Rubel1 · Nicole Managó1 · Guillermo D. Manrique1 · Graciela L. Garrote2 ·
Analia G. Abraham2,3

Received: 11 February 2022 / Accepted: 22 May 2022

© The Author(s), under exclusive licence to Springer Nature B.V. 2022

Abstract
Currently, the growing demand for non-dairy functional foods leads to the constant development of new products. The objec-
tive of the present work was to obtain a soy-based fermented beverage employing the strains Lactiplantibacillus plantarum
CIDCA 8327 or Lacticaseibacillus paracasei BGP1 and to analyze the effect of post-fermentation addition of inulin of low
or high average polymerization degree on the bacterial resistance. Also, the antimicrobial and antioxidant activity of the
fermented soy-based beverages were analyzed. The soy-based matrix was shown to be a suitable substrate for the growth of
both lactic acid bacteria, and the fermented beverages obtained presented bioactive properties such us antioxidant activity
and bactericidal effect against pathogen microorganisms. The addition of inulin after the fermentation process avoid the
hydrolysis and so, preserve its polymerization degree and thus the potential prebiotic effect. The incorporation of inulin to
the soy-based fermented beverages increased the bacterial count after 30 days of refrigerated storage up to 8.71 ± 0.15 and
8.41 ± 0.10 log CFU/mL for L. paracasei and L. planatrum respectively. The resistance to the gastrointestinal conditions of
the strain L. paracasei BGP1 in the fermented beverage was improved up to 70% when inulin of high polymerization degree
was added. Meanwhile the strain L. plantarum CIDCA 8327 showed a survival of 97 and 94% in the fermented beverage
added with inulin of low or high polymerization degree, respectively. These results contribute to the development of non-
dairy products containing inulin and probiotics and the diversification agri-based functional foods.

* Carolina Iraporda
ciraporda@fio.unicen.edu.ar
1
Departamento de Ingeniería Química y Tecnología de los
Alimentos, Facultad de Ingeniería, Universidad Nacional del
Centro de la Provincia de Buenos Aires, Av. Del Valle 5737,
7400 Olavarría, Argentina
2
Centro de Investigación y Desarrollo en Criotecnología de
Alimentos (CIDCA, UNLP-CIC-CONICET), Calle 47 y 116,
1900 la Plata, Argentina
3
Área Bioquímica y Control de los Alimentos, Facultad de
Ciencias Exactas, Universidad Nacional de La Plata, Calle 47
y 115, 1900 La Plata, Argentina

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133 Page 2 of 11 World Journal of Microbiology and Biotechnology (2022) 38:133

Graphical abstract

Soy-based
formula

Fermentaon
37 °C - 8 h
pH 4,0

 Actimicrobial activity against pathogens  ↑ bacterial viability during refrigerated storage

 Antioxidant activity  Inulin with high DPn ↑ L. paracasei BGP1
tolerance to simulated gastrointestinal conditions

Keywords Bioactive properties · Fermented soy-based beverage · Inulin · Lactic acid bacteria · Non-dairy products

Introduction benefits that could replace cow milk when its inclusion
in the diet is restrained (Bernat et al. 2014). In this sense,
Although the primary sector involved in the production of ‘soymilk’ refers to the traditional oriental beverage that
functional foods is represented nowadays by the dairy indus- consists of a water extract of soy beans. This product pro-
try, other sectors that employ alternative matrices as carriers vides a rich supply of nutrients and functional components,
of bioactive compounds such as probiotics and prebiotics including proteins, peptides, isoflavones, saponins, oligo-
are emerging. This fact is in line with the worldwide trend saccharides, dietary fiber, plant sterols, protease inhibitors,
of consumption of “lactose-free” beverages and foods, espe- lectins, phytic acid, and polyamines (Fukuda et al. 2017).
cially with the increase in the diagnosis of lactose intoler- Moreover, soy-based formula for infant containing soy
ance and allergies and the increase in the number of vegetar- protein isolate and carbohydrates are found in the market
ians, among others (Granato et al. 2010). as an alternative for cow-milk based infant formula. Also,
Probiotics, generally including species of Lactobacilli products obtained by the fermentation of soymilk with lac-
and Bifidobacteria, are defined as live microorganisms that, tic acid bacteria (LAB) have emerged as a viable alterna-
when administered in adequate amounts, are thought to con- tive to traditional yoghurts, with a particular interest in the
fer health benefits on the host (Hill et al. 2014). In order to enhanced health-promoting effects of these products (Bao
provide these effects, Lactobacillus strains have to be deliv- et al. 2012; Marazza et al. 2012; Wang et al 2006; Zhao
ered in a food system with a minimum level of ­106 CFU/mL and Shah, 2014). Interest in soy-based beverage production
at the time of consumption. They also must resist physical has led to developing strategies to improve its nutritional
and chemical barriers along the gastrointestinal tract, and value and the health benefits that they confer. So, it has been
have to confer specific beneficial health effects (Bao et al. shown that the soymilk fermentation by LAB contributes to
2012). increasing the nutritional value while reducing the off-flavor
The so-called vegetable milk refers to a water vegeta- associated with soy products, though improving the general
ble extract and represents an attractive option for probiotic acceptability of the beverages (Bergsveinson et al. 2017;
delivery because of its intrinsic nutritional value and health Rekha and Vijayalakshmi 2011). In addition, several studies

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World Journal of Microbiology and Biotechnology (2022) 38:133 Page 3 of 11 133

have evidenced that soymilk represents a suitable matrix broth (NB) (Britania, Argentina) was used for all pathogen
to be employed as a driver for probiotic delivery (Bedani growth.
and Rossi 2013; Donkor et al. 2007a, b). In addition to the LAB strains were stored at − 20 °C in MRS with skim
health benefits, the incorporation of prebiotics, like inulin, milk, and pathogen strains were kept in NB with glycerol
has been reported to provide also technological advantages 20% w/v. To activate microbial cultures, incubations in MRS
on physicochemical properties in addition to exercing a pro- (for LAB) or NB (for pathogens) at 37 °C for 24–48 h were
tective effect on probiotics survival during processing and conducted.
storage (de Souza et al. 2009). Inulin is a component of the
water-soluble fiber fraction that can preferentially stimulate Soy‑based formula fermentation
the growth and activity of a limited number of desirable
bacteria in the colon, which improves host health (Kolida The growth of each LAB strain in reconstituted commercial
et al. 2002). Moreover, it was demonstrated that incorporat- soy-based formula powder (Nutrilon Soya, Nutricia Bago,
ing inulin in combination with probiotics to soy yoghurt Argentina) was evaluated, determining the bacterial count
enhanced its textural and rheological characteristics (Mishra in MRS agar, pH (pHmeter HI 2211, HANNA, USA), and
and Mishra 2013). titratable acidity (as % lactic acid) at different incubation
The strain Lactiplantibacillus plantarum CIDCA8327 is times. For this, the sterile soy-based formula sample was
a facultative heterofermentative lactic acid bacteria isolated inoculated at 1% v/v (~ ­106–7 CFU/mL) with the correspond-
from kefir grains (Garrote et al. 2001). This strain showed ing LAB culture (grown 24 h, in MRS broth at 37 °C), and
inhibitory activity against some pathogens (Golowczyc et al. then it was incubated at 37 or 42 °C for 24 h. Aliquots every
2008) and promisory probiotic potential for the develop- 2 h, during 24 h, were withdrawn for the determinations. The
ment of functional foods (Gangoiti et al. 2017; Londero et al. experiments were done by triplicate.
2015). The commercial strain Lacticaseibacillus paracasei
BGP1 is a recognized probiotic microorganisms employed
for functional food processing (Céspedes et al. 2013). This Bioactive propeties
work aimed to obtain a functional soy-based beverage by
fermentation with Lactiplantibacillus plantarum CIDCA Antimicrobial activity of the fermented soy‑based
8327 or Lacticaseibacillus paracasei BGP1 and to analyze beverages
the effect of different polymerization degrees inulin addi-
tion to the fermented beverages, on the bacterial viability Fermented soy-based beverages with L. paracasei or L.
during storage as well as the bacterial tolerance to gastroin- plantarum, unfermented acidified soy-based formula (ASF)
testinal conditions. In addition, the antimicrobial and anti- adjusted to pH 4 with lactic acid or unfermented soy-based
oxidant activity of the fermented soy-based beverages were formula (Control), were inoculated at 1% with the pathogen
analyzed. strain grown in NB at 37 °C for 24 h. The plate count viable
microorganism analysis was performed immediately after
the inoculation (initial) and after 24 h of incubation at 37 °C
Materials and methods (final) in selective agar plates for the pathogen microorgan-
isms studied. The results were expressed as CFU/mL. The
Bacterial strains and culture media growth degree was calculated as the difference between the
final and the initial log CFU/mL. Three independents experi-
For soy-based formula fermentation, the commercial pro- ments were done.
biotic strain Lacticaseibacillus paracasei BGP1 (Clericci,
SACCO, Italy) and the strain Lactiplantibacillus plantarum Antioxidant activity of the fermented soy‑based beverages
CIDCA 8327 isolated from kefir grains (Garrote et al. 2001;
Golowczyc et al. 2008) were used. For LAB culture, MRS Antioxidant activity of fermented soy-based beverage
broth (Britania, Argentina) was employed. was evaluated by DPPH radical scavenging activity meas-
The pathogen strains Escherichia coli, Salmonella sp., ured according to Brand-Williams et al. (1995). Aliquots
and Staphylococcus aureus isolated from clinical animal (100 μL) of fermented beverage supernatant (obtained by
samples were identified and provided by Laboratorio 9 de centrifugation 10 min, 10,000 ×g) were added to 1.9 mL
Julio (Buenos Aires, Argentina). The media Mac Conkey of a methanolic solution of DPPH radical (100 μM). After
Agar (Britania, Argentina), Salmonella-Shigella Agar (Bri- agitation, the mixture was left in the dark for 30 min, and
tania, Argentina), and Salted Mannitol (Brizuela, Argentina) the absorbance was measured at 517 nm employing a spec-
were employed for the enumeration of viable E. coli, Salmo- trophotometer (UV-1800PC, Mapda Instruments, China).
nella sp. and Staphylococcus aureus, respectively. Nutrient The antioxidant activity was expressed as a percentage of

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133 Page 4 of 11 World Journal of Microbiology and Biotechnology (2022) 38:133

DPPH radical scavenging calculated according to the fol- 45 mM ­NaHCO3, and 0.3% w/v pepsin (SIGMA-Aldrich,
lowing formula: USA), and the final pH was adjusted to 2.5 with concen-
trated HCl. The second was in an intestinal solution con-
(AC −AS)
% DPPH radical scavenging = 100 × taining 22 mM NaCl, 3.2 mM KCl, 7.6 mM N ­ aHCO3, 0.1%
AC w/v pancreatin (SIGMA-Aldrich, USA), and 0.15% w/v bile
where AC corresponds to the absorbance of DPPH solution, sales (SIGMA-Aldrich, USA) and the final pH adjusted to 8
and AS is the absorbance of DPPH solution after 30 min of with 4 N NaOH. For the tolerance test, 2.5 mL of fermented
reaction with the supernatants of the fermented soy-based beverage with or without inulin, were mixed with 2.5 mL of
beverages. Three independents experiments were done. gastric solution (2×), the pH adjusted to 2.5 with HCl, and
the final mixture incubated for 90 min at 37 °C. Then, 5 mL
of intestinal solution (2×) was added, the pH adjusted to 8
Effect of the inulin addition to the fermented
with NaOH, and the final mixture was incubated for 180 min
soy‑based beverages
at 37 °C. Bacterial viability was assessed by plate count in
MRS agar (37 °C, 48 h), and the percentage of survival was
Formulation of the beverage with inulin addition
calculated as follows:
A commercial infant formula powder, based on isolated soy (log CFU∕mL)after
protein and glucose syrup (Nutrilon Soya, Nutricia Bago, % Survival = 100 ×
(log CFU∕mL)before
Argentina) was used. The powder was reconstituted with dis-
tilled water according to manufacture instructions and then where log CFU/mL represents cell viability at the beginning
sterilized. The strains L. plantarum CIDCA 8327 or L. para- (before) and after incubation (after) in the simulated gastro-
casei BGP1 were grown in MRS at 37 °C for 24 h, harvested intestinal conditions, as described above. Three independent
by centrifugation (10 min, 7000 ×g), washed twice, and assay were performed.
resuspended in PBS buffer (pH 7). Soy-based formula was
inoculated at 1% v/v with bacterial suspension in PBS and Statistical analysis
incubated at 37 °C for 8 h. Then, the pH of the fermented
soy-based beverage was adjusted to 4 with lactic acid, and Results were expressed as mean ± standard deviation of three
its final lactic acid concentration was measured employing a independent experiments with duplicates on each assay.
commercial kit (Lactate, Wiener Lab., Argentina). After the One-way analysis of variance was performed. Mean differ-
fermentation process, inulin with different average polym- ences were statistically tested using the LSD Fisher multiple
erization degrees: Low, DPn ≥ 10 (GR, Beneo® Orafti, Bel- comparison test conducted by the InfoStat® Software (Ver-
gium) or high, DPn ≥ 23 (HP, Beneo® Orafti, Belgium) was sion 2008, Argentina). A p value ≤ 0.05 indicated significant
added at 3% w/v. Finally, samples (with and without inulin) differences.
were placed in hermetically sealed plastic containers and
stored at 4 °C for 30 days.
Results
Bacterial viability in fermented soy‑based beverage
during the storage Fermentation of soy‑based formula by LAB strains
The bacterial viability (CFU/mL) in the fermented soy-based The ability of the probiotic strains Lacticaseibacillus para-
beverages samples after 0, 10, 20, and 30 days of storage at casei BGP1 and Lactiplantibacillus plantarum CIDCA 8327
4 °C was assayed by plate count in MRS agar. The effect of to grow and to ferment the soy-based formula, was evalu-
post-fermentation addition of inulin on the bacterial viability ated. It was observed that both LAB grew in the soy-based
during refrigerated storage of the fermented products was formula at the two temperatures evaluated (Fig. 1). Besides,
also evaluated, by triplicate. the growth rate in the soy-based formula at the two different
temperatures is strain dependent. It was observed that the
Bacterial tolerance to simulated gastrointestinal conditions temperature had no significant effect on the growth of L.
plantarum in the soy-based formula; meanwhile the growth
In order to evaluate the resistance of the LAB strains pre- of L. paracasei at 37 °C was significantly higher concerning
sent in the fermented soy-based beverages to simulated the growth observed at 42 °C.
gastrointestinal conditions, two serial incubations were The reconstituted soy-based formula (unfermented)
used according to Grimoud et al. (2010). The first was in presented a pH value of 6.80 ± 0.27. Fig. 2a shows the pH
a gastric solution containing 125 mM NaCl, 7 mM KCl, values during fermentation of soy-based formula at 37 and

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World Journal of Microbiology and Biotechnology (2022) 38:133 Page 5 of 11 133

Fig. 1  Growth of LAB in a b

soy-based formula at different 10 10
temperatures. Growth (log CFU/
mL) of a L. paracasei BGP1 9 9
and b L. plantarum CIDCA

Log CFU/mL

Log CFU/mL
8327, during 24 h at 37 °C
(open symbols) and 42 °C (full 8 8
symbols)
7 7

6 6
0 6 12 18 24 0 6 12 18 24
Time (h) Time (h)

Fig. 2  Acidification of soy-

based formula fermented with
LAB. a pH and b Titratable
acidity (% lactic acid) of soy-
based formula fermented during
24 h at 37 °C (open symbols)
and 42 °C (full symbols) with
L. paracasei BGP1 (□,■) or L.
plantarum CIDCA 8327 (○,●)

42 °C for 24 h with L. paracasei BGP 1 and L. plantarum Table 1  Characterization of the soy-based beverage obtained after
CIDCA8327. It was observed that the pH remained constant 8 h fermentation at 37 °C with L. paracasei BGP1 and L. plantarum
CIDCA 8327
during soy-based formula fermentation at 42 °C with both
strains throughout the assay period. Instead, at 37 °C, the fer- L. paracasei BGP1 L. plantarum
mentation with L. paracasei produced a significant decrease CIDCA 8327
in the pH reaching a minimum (4.22 ± 0.22) after 24 h, while Bacterial count 8.48 ± 0.08 8.53 ± 0.15
L. plantarum reached the minimum pH (4.12 ± 0.07) after (log CFU/mL)
12 h. During the soy-based formula fermentation with both pH 5.32 ± 0.06 4.56 ± 0.11
LAB at 37 °C, the titratable acidity increased (Fig. 2b) as Lactic acid 0.12 ± 0.01 0.16 ± 0.02
the pH decreased. (%w/v)
The titratable acidity of the soy-based formula, expressed Values in each row are the mean ± standard deviation
as percentage of lactic acid during the first 8 h of fermenta-
tion at 37 °C, was in the range between 0.07 and 0.15%, and
after 24 h the amount of lactic acid produced by L. plan- plantarum CIDCA 8327 was evaluated, considering the fact
tarum was higher compared with L. paracasei (0.64 and that these pure cultures exerted inhibitory effect when facing
0.25%, respectively). the pathogenic bacteria Salmonella sp., Escherichia coli, and
Finally, the fermentation process for producing the soy- Staphylococcus aureus.
based fermented beverages with both LAB was conducted at The growth of pathogens after incubation 24 h at 37 °C
37 °C during 8 h, then the beverages were bottled and stored in the soy-based fermented beverages and the unfermented
at 4 °C for further analysis. The Table 1 shows the results beverage (control) is shown in Fig. 3. The three pathogens
about bacterial concentration (log CFU/mL), pH and titable microorganisms were able to grow in unfermented soy-based
acidity (lactic acid %w/v) obtained in these conditions. formula, reaching at 24 h concentrations significantly higher
than the initial values, with a growth increase respect to the
Soy‑based fermented beverages: antimicrobial initial values of 0.86, 1.60, and 1.63 log CFU/mL cycles for
and antioxidant activities E. coli, Salmonella sp., and S. aureus, respectively. In order
to evaluate the effect of acidity on the pathogen growth,
The antimicrobial activity of the soy-based fermented bev- a sample of unfermented soy-based formula acidified with
erages obtained with the strains L. paracasei BGP1 and L. lactic acid (pH 4) was used to inoculate the pathogen strains.

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133 Page 6 of 11 World Journal of Microbiology and Biotechnology (2022) 38:133

a b c
10 10 d 10
* *c
8 *c c
8 c 8
Initial Initial Initial
b *b
Log CFU/mL

Log CFU/mL

Log CFU/mL
6 6 6
*b
4 *b 4 4

2 2 2 *a
*a a
*
0 0 0
Control ASF SFB BGP1 SFB 8327 Control ASF SFB BGP1 SFB 8327 Control ASF SFB BGP1 SFB 8327

Fig. 3  Antimicrobial activity of fermented soy-based beverages. Via- CIDCA 8327 (SFB 8327) of A Escherichia coli, B Salmonella sp and
ble concentration (log CFU/mL) initial (dotted line) and after 24 h C Staphylococcus aureus. Bars with different letters indicate signifi-
(bars) incubation in unfermented soy-based formula (Control), artifi- cant differences among the concentrations obtained at 24 h, while the
cially acidified soy-based formula (ASF), fermented soy-based bev- asterisk (*) indicate significant differences with the initial value (p
erage with L. paracasei BGP1 (SFB BGP1), and with L. plantarum value ≤ 0.05). ANOVA followed by LSD Fisher

The artificially acidified soy-based formula presented a bac- degree was evaluated (Fig. 4). For both LAB strains, the
teriostatic effect against the pathogens; moreover the con- soy-based beverage with post-fermentation addition of
centrations obtained for Salmonella and S. aureus after 24 h inulin presented significantly higher viable counts than
were significantly lower than those obtained in the unfer- in the soy-based beverage without inulin (control) after
mented control soy-based formula. The soy-based fermented storage. Moreover, the post-fermentation addition of low
beverages with each LAB strain presented bactericidal effect or high DPn inulin to the beverages contributed to main-
against the three pathogens. In the case of soy-based bev- taining the viability of both LAB strains during storage,
erage fermented with L. plantarum, no growth of E. coli since the final counts (30 days) did not present significant
and Salmonella sp. was detected (< ­102 CFU/mL), while S.
aureus presented a decrease of 4.47 log CFU/mL cycles. The
concentrations found after 24 h of incubation in soy-based
beverage fermented with L. paracasei were significantly a
10
* * Initial
lower than the initial values showing a decrease of 2.90; a
a a a a a a a a
10 days
b
2.83 and 0.54 log CFU/mL cycles for E. coli, Salmonella 8 20 days
Log CFU/mL

c c
30 days
sp., and S. aureus, respectively.
The percentage of inhibition of the DPPH radical found 6
in the soy-based beverage fermented with L. plantarum was
25.3 ± 1.8% and did not show significant differences from 5
the unfermented control (27.3 ± 1.0%). However, the per- Control Inulin Low DPn Inulin High DPn

centage of inhibition values found for the soy-based bev-

erage fermented with L. paracasei were 24.3 ± 1.8% and b 10
Initial
resulted significantly lower than the unfermented control. a a a a *
a a
a
*
a 10 days
a b b
In addition, no significant differences were found between 8
b
20 days
Log CFU/mL

the fermentation with both strains. 30 days

At this point the antimicrobial and antioxidant activities 6

of the fermented beverages were described, and this product
was employed as basis for the final product formulations.
5
Control Inulin Low DPn Inulin High DPn

Storage of soy‑based fermeted beverage

with addition of inulin Fig. 4  LAB viability in fermented soy-based beverages during stor-
age. Counts of viable (log CFU/mL) in fermented soy-based bever-
ages with the addition of inulin (3% w/v) of different average polym-
After 30 days of storage at 4 °C the soy-based fer- erization degree ­(DPn ≥ 10 low or ­DPn ≥ 23, high), during storage at
mented beverages presented viable counts of 2 × ­1 0 7 4 °C of A L. paracasei BGP1 and B L. plantarum CIDCA 8327. Bars
and 6 × ­107 CFU/mL for L. paracasei and L. plantarum, with different letters indicate significant differences among the viable
respectively. Also, the viability of LAB strains in the fer- counts at different storage times of each fermented soy-based bever-
age (p value ≤ 0.05). The asterisk (*) indicate significant differences
mented soy-based beverage with inulin (3% w/v) of low with the control after 30 days of storage. ANOVA followed by LSD
(DPn ≥ 10) or high (DPn ≥ 23) average polymerization Fisher

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World Journal of Microbiology and Biotechnology (2022) 38:133 Page 7 of 11 133

differences with respect to the initial counts, regardless of Discussion

the polymerization degree of the inulin added.
The variation of the pH values of the fermented bever- Many challenges arise when attempting to incorporate
age is another factor that should be taken into account probiotic bacteria in foods matrices, mainly related to
as an indicative parameter of the product performance their growth, survival, viability, stability and functional-
during refrigerated storage. After 30 days of storage at ity in food processing, storage and consumption as well as
4 ºC, the fermented soy-based beverage without inulin changes of sensory characteristics of probiotic foods (Min
presented pH values of 4.02 ± 0.10 and 3.44 ± 0.3 for L. et al. 2018). It is known that many LAB strains possess
paracasei and L. plantarum, respectively. It is worth to the metabolic tools to degrade oligosaccharides through
mention that no significant differences between the pH α-galactosidase activity (Silvestroni et al. 2002) and are
values of the fermented beverage with or without inulin capable of adapting their metabolism and growing in
stored were obtained for each strain. The pH values of the soy-based products (Aguirre et al. 2014; Farnworth et al.
fermented soy-based beverages with inulin of low and high 2007; Granato et al. 2012). The LABs might present dif-
polymerization degree were 3.74 ± 0.2 and 3.79 ± 0.3 for ferent metabolic activities, evidencing differences in their
L. paracasei, or 3.25 ± 0.3 and 3.31 ± 0.3 for L. plantarum, nucleotide sequence described by technologies that have
respectively. the power to rapidly reveal important genetic detail for
the LAB involved in these fermentations, as reported by
Bergsveinson et al. (2017). In this sense, in this work, it
Survival of LAB in soy‑based fermented beverages was observed a satisfactory growth of L. paracasei BGP1
with inulin after gastrointestinal simulation and L. plantarum CIDCA 8327 in the soy-based formula.
For both strains, a low pH and high titratable acidity was
The soy-based beverages fermented with L. paracasei found in the soy-based formula fermented at 37 °C, similar
and L. plantarum without or with inulin addition, were to the results reported by Wang et al. (2002) in soymilk
subjected to simulated gastrointestinal conditions, and fermented with combinations of bifidobacterias and lactic
the results concerning the bacterial survival are shown acid bacteria and dos Santos et al. (2019) for soymilk fer-
in Table 2. The survival of L. plantarum in fermented mented with kefir grains. Other work showed higher lac-
soy-based beverage submitted to in vitro gastrointestinal tic acid concentrations for soy beverages containing cane
conditions was higher than the survival of L. paracasei, juice fermented with L. johnsonii NCC533 and L. rhamno-
in the respective fermented soy-based beverage, suggest- sus ATCC 53103 (Farnworth et al. 2007). The variability
ing that this characteristic is microorganism dependent. in the lactic acid content as a metabolic product detected
Moreover, it was observed that the post-fermentation addi- in the fermented beverages is dependent on the metabolic
tion of inulin (either of low or high DPn) contributed to and adaptive characteristics of each particular strain or
increase the percentage of survival of L. plantarum after starter and also the fermentation conditions.
simulated gastrointestinal treatment. Meanwhile, the post- Many studies have reported that soy fermented bever-
fermentation addition of inulin with high DPn produced ages containing probiotics present good general accept-
a higher increase of the survival of L. paracasei in fer- ability by potential consumers since the fermentation
mented soy-based beverage than when inulin of the lower improves the product’s flavor and texture (Donkor et al.
DPn was added. 2007a, b; Granato et al. 2012; Shimakama et al. 2003).
The preliminary promissory results obtained in the present
work leads to complete characterization of this new func-
tional non-dairy beverage considering the sensory analysis
Table 2  Percentage of survival of LAB strains in fermented soy- for the future studies. Among other parameters, the pH
based beverages with addition of inulin (3% w/v) after simulated gas- influences the stability, aroma, flavor, texture, and shelf-
trointestinal treatment
life of the fermented soymilk products.
Percentage of survival (%) Both bacteria and also their spent culture supernatants
Control Inulin low DPn Inulin high DPn employed in the present work, showed inhibitory effect
against E. coli, Salmonella sp. and S. aureus. In general,
L. paracasei BGP1 52.4 ± 3.1a 48.1 ± 2.2a 70.6 ± 8.6b the antimicrobial activity of Lactobacillus strains is well
L. plantarum 87.2 ± 3.2a 97.9 ± 1.2b 94.6 ± 2.0b documented; for instance, in coincidence with the results
CIDCA8327
obtained in this work, Bao et al. (2012) demonstrated that
DPn: average polymerization degree (Low: ≥ 10; High: ≥ 23). Values the strain Lactobacillus plantarum IMAU70004 presented
in each row with the same superscript are not significantly different antagonistic activity against the foodborne pathogens
(p > 0.05). ANOVA followed by LSD Fisher Test

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133 Page 8 of 11 World Journal of Microbiology and Biotechnology (2022) 38:133

Salmonella typhimurium S50333, Escherichia coli O157 and bioactive characteristics of the product, a compromise
882364, and Staphylococcus aureus AC1.2465; mean- relationship is generated between these parameters.
while other studies show different degrees of antagonism The soy-based fermented beverages were characterized
against S. aureus and E. coli exerted by Lactobacillus and employed as a basis for the development of the func-
strains (Ebhodaghe et al. 2012; Shokryazdan et al. 2014). tional beverage with addition of inulin. In most of the studies
The spent culture supernatant contains bacterial metabolic inulin is mainly added before the soymilk fermentation, so it
products that might exert inhibitory effects, as suggested is available to be hydrolyzed and employed as fermentable
by Bian et al. (2011) for L. reuteri DPC16 spent culture substrate by bacteria (Battistini et al. 2018; Bedani et al.
against food-borne pathogens, due to acidic effect in a pH- 2013; Saarela et al. 2006). In the present work, to avoid
dependent way. Other research concluded that the inhibi- inulin from being hydrolyzed during the fermentation and
tory effect of Lactobacillus supernatants against pathogens to analyze the effect that inulin polymerization degree has
was due to their organic acid productions (Shokryazdan on the bacterial survival, this compound was added after
et al. 2014). the bacterial soy-based formula fermentation. During the
The fermented beverages obtained also exibited anti- storage of the beverages, the probiotics are exposed to a
microbial activity against the three pathogens analyzed in low pH values that may damage both the cell wall and the
this work. In line with these results, Kumari and Vij (2015) cell membrane, thus influencing ΔpH and the membrane
reported antimicrobial activity of soymilk fermented with potential, acidifying of the cytosol and affecting in conse-
L. rhamnosus C6 against S. aureus and E. coli 0157:H7 quence the bacterial viability (Papadimitriou et al. 2016).
ATCC 35150. Moreover, soymilk fermented with L. hel- The soy-based beverage with inulin presented significantly
veticus V3 inhibited the growth of S. aureus and S. typhi, higher viable counts than in the soy-based beverage without
while fermentation with S. thermophilus MD2 inhibited inulin after storage, regardless of the polymerization degree.
the growth E. coli (Hati et al. 2018). The antimicrobial These results can be attributed to the better environment
activity against pathogens exhibited by the fermented bev- and the direct interactions between inulin polymer struc-
erages could be attributed either to the presence of organic ture with bacterial proteins and lipids from the membranes,
acid produced by the strains (Ebhodaghe et al. 2012) and contributing to their survival during storage, as was also
also to bioactive peptides produced during soymilk fer- described for different food applications (Khorasany and
mentation (Singh et al. 2015). Shahdadi 2021; Schaw et al. 2007). The protective effect
The antioxidant capacity of the soy-based formula and of inulin during during freeze-drying is attributed to direct
the fermented products obtained in this work was mod- interactions with proteins and membranes. Other authors
erate and no increase of this activity was obtained after showed that the addition of inulin in fermented soymilk
fermentation. In contrast, Marazza et al. (2012) and Rani exerted a protective effect on LAB viability and activity
and Pradeep (2015) informed that the antioxidant activity during fermentation (Donkor et al. 2007a, b; Oliveira et al.
increase after fermentation of soymilk with L. rhamnosus 2011; Valero-Cases and Frutos 2017). One of the advantages
and L. paracasei KUMBB005. Also, other works reported of the post-fermentation addition of inulin can be related to
higher antioxidant activity of soymilk after the fermen- keeping the chemical structure of this ingredient, since it is
tation with other LAB and bifidobacteria (Wang et al. not hydrolyzed nor consumed by bacteria during the stor-
2006). Meanwhile the antioxidant activity obtained in the age at 4 °C, as was evidenced by thin layer chromatography
present work is in accordance to the results reported for analysis (data not shown).
soy protein extract fermented with L. acidophilus CSCC The metabolic activity of the LAB cultures employed for
24, L. paracasei CSCC 279, L. zeae ASCC 15820, and L. soymilk fermentation is reduced during refrigerated stor-
rhamnosus WQ2 (Zhao and Shah 2014). The antioxidant age, however some variation of the pH of the final products
activity seems to have a positive correlation either with may take place. The difference between the pH values after
fermentation time and the bacterial proteolysis capacity 30 days of storage of each fermented soy-based beverage
(Monajjemi et al. 2012; Singh et al. 2015), and so it is obtained, could be related to the metabolic activity intrin-
possible that 8 h of fermentation were not enough to evi- sic of each strain. This was evidenced in the present work
dence an increase in the DPPH radical scavenging activity since in one hand, no significant differences were obtained
of the beverage developed in the present work. However, between the initial and final pH of the soy-based beverage
the antioxidant activity of the fermented soymilks could fermented with L. paracasei with or without inulin. On
increase by the gastrointestinal conditions due to larger the other hand, the beverage fermented with L. plantarum
hydrolysis of soy proteins and the release of bioactive pep- showed a significant decrease in the pH values after the
tides (Capriotti et al. 2015). Finally, given that the fermen- storage, and no signifficant differences were obtained in
tation time and proteolysis degree affect the organoleptic the final pH values between the samples. As suggested by
other authors, the acidity could be maintained or decreased

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during storage in fermented soy-products (Capriotti et al. and improved the bacterial tolerance to simulated gastroin-
2015; dos Santos et al. 2019; Saito et al. 2014). In addi- testinal conditions. Finally, this study demonstrate that the
tion, inulin-supplementation of fermented soymilk did not fermented soy-based beverages with inulin represents a good
produce changes in pH values of after 28 days of storage at matrix for probiotic delivery, as alternative to the traditional
4 °C, as reported by Mishra and Mishra (2018). It should be dairy probiotic beverages also contributing to diversify the
taken in account that there is a negative correlation between type of agri-based food products.
the titable acidity and pH with the consummers acceptability
(Raja et al. 2009; Souza et al. 1991). However, fermented Acknowledgements CI, IAR, GGL and AGA are researchers of the
National Council for Scientific and Technical Research (CONICET,
beverages with a pH under 4 showed higher stability against Argentina), GDM is member of the research career from the Scientific
spoilage and resulted a refreshing product with a mild acidic Research Commission (CIC, Argentina), and NM is graduated of the
taste (Simova et al. 2002). National University of the Center of Buenos Aires Province (UNCPBA,
Probiotic foods must demonstrate the ability to preserve Argentina). Authors are grateful to Laboratorio 9 de Julio (Argentina)
for pathogen microbial cultures supply.
the viability of the strains after passage through the gastro-
intestinal tract (Buriti et al. 2010). It was determined that Author contributions CI and IAR have made substantial contributions
the soy matrix contributed to improve the survival of both to conception and design, acquisition, analysis and interpretation of
strains when compared with their survival as free cells, as data, and manuscript writing. NM performed the experimental assays.
described by Iraporda et al. (2019). In accordance other GDM, GLG and AGA have been involved in drafting the manuscript
and revising it critically for important intellectual content and final
authors reported that the survival rate of different LAB approval of the version to be published.
after simulated gastrointestinal conditions was higher in a
fermented soy beverage than in culture media (Bedani et al. Funding Financial support was provided by the National Council for
2013; Guo et al. 2009). Moreover, it was observed that the Scientific and Technical Research (CONICET, Argentina) and National
post-fermentation addition of inulin increase the percentage Agency fos Scientific and Technological Promotion (ANPCyT, Argen-
tina) PICT-2019-1220 and PICT-2019–0211.
of survival of L. plantarum after simulated gastrointestinal
treatment. Meanwhile the post-fermentation addition of inu-
lin with high DPn increased the survival of L. paracasei in
Declarations
fermented soy-based beverage. Therefore, the mean molecu- Conflict of interest The authors declared no potential conflict of inter-
lar weight, and hence the DPn of the inulin, could be cor- est with respect to the research, authorship, and/or publication of this
related with the degree of protection that this polysaccharide article.
exerts on bacterial tolerance to gastrointestinal conditions,
as suggested by Valero-Cases and Frutos (2017). This result
leads to highlighting a differential action with regards of the
inulin structure; thus, post-fermentation inulin incorporation References
could be advantageous.
Aguirre L, Hebert EM, Garro MS, de Giori GS (2014) Proteolytic
activity of Lactobacillus strains on soybean proteins. LWT-Food
Sci Technol 59(2):780–785. https://​doi.​org/​10.​1016/j.​lwt.​2014.​
Conclusion 06.​061
Bao Y, Zhang Y, Li H, Liu Y, Wang S, Dong X, Su F, Yao G, Sun T,
In this work, it was shown that the soy-based formula is a Zhang H (2012) In vitro screen of Lactobacillus plantarum as
probiotic bacteria and their fermented characteristics in soymilk.
suitable matrix for the growth of both Lacticaseibacillus Ann Microbiol 62(3):1311–1320. https:// ​ d oi. ​ o rg/ ​ 1 0. ​ 1 007/​
paracasei BGP1 and Lactiplantibacillus plantarum CIDCA s13213-​011-​0377-4
8327, obtaining a fermented product that has a bactericidal Battistini C, Gullón B, Ichimura ES, Gomes AMP, Ribeiro EP, Kunigk
effect against pathogen microorganisms, while the antioxi- L et al (2018) Development and characterization of an innovative
synbiotic fermented beverage based on vegetable soybean. Braz
dant activity of the products was maintained. Moreover, the J Microbiol 49(2):303–309. https://​doi.​org/​10.​1016/j.​bjm.​2017.​
beverages’ appearances were homogeneous and remained 08.​006
stable during storage. In particular, the strain L. plantarum Bedani R, Rossi EA, Isay Saad SM (2013) Impact of inulin and okara
CIDCA 8327 exhibited better performance with respect on Lactobacillus acidophilus La-5 and Bifidobacterium animalis
bb-12 viability in a fermented soy product and probiotic survival
to L. paracasei BGP1. In order to avoid inulin from being under in vitro simulated gastrointestinal conditions. Food Micro-
hydrolyzed during the fermentation process, and though biol 34(2):382–389. https://​doi.​org/​10.​1016/j.​fm.​2013.​01.​012
preserving its chemical structure, this bioactive ingredient Bergsveinson J, Kajala I, Ziola B (2017) Next-generation sequenc-
was incorporated after the bacterial fermentation. As a rel- ing approaches for improvement of lactic acid bacteria-fermented
plant-based beverages. AIMS Microbiol 3(1):8. https://d​ oi.o​ rg/1​ 0.​
evant result of this work, the post-fermentation addition of 3934/​micro​biol.​2017.1.8
inulin to the fermented soy-based beverages contributed to Bernat N, Cháfer M, Chiralt A, González-Martínez C (2014) Hazel-
increasing the bacterial viability during refrigerated storage, nut milk fermentation using probiotic Lactobacillus rhamnosus

13
 133 Page 10 of 11 World Journal of Microbiology and Biotechnology (2022) 38:133

GG and inulin. Int J Food Sci Technol 49(12):2553–2562. concepts, and products. Compr Rev Food Sci F 9(3):292–302.
https://​doi.​org/​10.​1111/​ijfs.​12585 https://​doi.​org/​10.​1111/j.​1541-​4337.​2010.​00110.x
Bian L, Molan AL, Maddox I, Shu Q (2011) Antimicrobial activity Granato D, Masson ML, Ribeiro JCB (2012) Sensory acceptability
of Lactobacillus reuteri DPC16 supernatants against selected and physical stability evaluation of a prebiotic soy-based dessert
food borne pathogens. World J Microbiol Biotechnol 27(4):991– developed with passion fruit juice. Food Sci Technol 32(1):119–
998. https://​doi.​org/​10.​1007/​s11274-​010-​0543-z 126. https://​doi.​org/​10.​1590/​S0101-​20612​01200​50000​04
Brand-Williams W, Cuvelier ME, Berset CLWT (1995) Use of a free Grimoud J, Durand H, Courtin C, Monsan P, Ouarné F, Theodorou
radical method to evaluate antioxidant activity. LWT-Food Sci V, Roques C (2010) In vitro screening of probiotic lactic acid
Technol 28(1):25–30. https://​doi.​org/​10.​1016/​S0023-​6438(95)​ bacteria and prebiotic glucooligosaccharides to select effective
80008-5 synbiotics. Anaerobe 6(5):493–500. https://​doi.​org/​10.​1016/j.​
Buriti FC, Castro IA, Saad SM (2010) Effects of refrigeration, freez- anaer​obe.​2010.​07.​005
ing and replacement of milk fat by inulin and whey protein con- Guo Z, Wang J, Yan L, Chen W, Liu XM, Zhang HP (2009) In vitro
centrate on texture profile and sensory acceptance of symbiotic comparison of probiotic properties of Lactobacillus casei Zhang,
guava mousses. Food Chem 123(4):1190–1197. https://​doi.​org/​ a potential new probiotic, with selected probiotic strains. LWT-
10.​1016/j.​foodc​hem.​2010.​05.​085 Food Sci Technol 42(10):1640–1646. https://​doi.​org/​10.​1016/j.​
Capriotti AL, Caruso G, Cavaliere C, Samperi R, Ventura S, Chi- lwt.​2009.​05.​025
ozzi RZ, Laganà A (2015) Identification of potential bioactive Hati S, Patel N, Mandal S (2018) Comparative growth behaviour and
peptides generated by simulated gastrointestinal digestion of biofunctionality of lactic acid bacteria during fermentation of soy
soybean seeds and soy milk proteins. J Food Comp Anal Aci milk and bovine milk. Prob Antimicrob 10(2):277–283. https://​
44:205–213. https://​doi.​org/​10.​1016/j.​jfca.​2015.​08.​007 doi.​org/​10.​1007/​s12602-​017-​9279-5
Céspedes M, Cárdenas P, Staffolani M, Ciappini M, Vinderola Hill C, Guarner F, Reid G, Gibson GR, Merenstein DJ, Pot B, Morelli
G (2013) Performance in nondairy drinks of probiotic L. L, Berni Canani R, Flint HJ, Salminen S, Calder PC, Sanders ME
casei strains usually employed in dairy products. J Food Sci (2014) The international scientific association for probiotics and
78(5):756–762 prebiotics consensus statement on the scope and appropriate use
de Souza Oliveira RP, Perego P, Converti A, De Oliveira MN (2009) of the term probiotic. Nat Rev Gastroenterol Hepatol 11(8):506–
The effect of inulin as a prebiotic on the production of probiotic 514. https://​doi.​org/​10.​1038/​nrgas​tro.​2014.​66
fibre-enriched fermented milk. Int J Dairy Tech 62(2):195–203. Iraporda C, Rubel IA, Manrique GD, Abraham AG (2019) Influence
https://​doi.​org/​10.​1111/j.​1471-​0307.​2009.​00471.x of inulin rich carbohydrates from Jerusalem artichoke (Helian-
Donkor ON, Henriksson A, Vasiljevic T, Shah NP (2007a) Rheologi- thus tuberosus L.) tubers on probiotic properties of Lactobacillus
cal properties and sensory characteristics of set-type soy yogurt. strains. LWT-Food Sci Technol 101:738–746. https://​doi.​org/​10.​
J Agric Food Chem 55(24):9868–9876. https://​doi.​org/​10.​1021/​ 1016/j.​lwt.​2018.​11.​074
jf071​050r Khorasany S, Shahdadi F (2021) Improvements in survival of probiotic
Donkor ON, Henriksson A, Vasiljevic T, Shah NP (2007b) bacteria, rheology and sensory characteristics of yogurts during
α-Galactosidase and proteolytic activities of selected probiotic and storage. Nutr Food Sci Res 8(1):35–43
dairy cultures in fermented soymilk. Food Chem 104(1):10–20. Kolida S, Tuohy K, Gibson GR (2002) Prebiotic effects of inulin and
https://​doi.​org/​10.​1016/j.​foodc​hem.​2006.​10.​065 oligofructose. Br J Nutr 87(2):193–197. https://​doi.​org/​10.​1079/​
dos Santos DC, de Oliveira Filho JG, Santana ACA, de Freitas BSM, BJN/​20025​37
Silva FG, Takeuchi KP, Egea MB (2019) Optimization of soymilk Kumari P, Vij S (2015) Growth and antimicrobial activity of proteo-
fermentation with kefir and the addition of inulin: physicochemi- lytic probiotic Lactobacillus rhamnosus C6 in soymilk and whey.
cal, sensory and technological characteristics. LWT-Food Sci Indian J Dairy Sci 68(3):229–238
Technol 104:30–37. https://​doi.​org/​10.​1016/j.​lwt.​2019.​01.​030 Londero A, Iraporda C, Garrote GL, Abraham AG (2015) Cheese whey
Ebhodaghe SO, Abiose SH, Adeniran HA (2012) Assessment of fermented with kefir micro-organisms: antagonism against Sal-
physico–chemical characteristics, viability and inhibitory effect monella and immunomodulatory capacity. Int J Dairy Technol
of Bifidobacteria in soymilk. J Food Res 1(2):159. https://d​ oi.o​ rg/​ 68(1):118–126
10.​5539/​jfr.​v1n2p​159 Marazza JA, Nazareno MA, de Giori GS, Garro MS (2012) Enhance-
Farnworth ER, Mainville I, Desjardins MP, Gardner N, Fliss I, Cham- ment of the antioxidant capacity of soymilk by fermentation with
pagne C (2007) Growth of probiotic bacteria and bifidobacteria in Lactobacillus rhamnosus. J Funct Foods 4(3):594–601. https://​
a soy yogurt formulation. Int J Food Microbiol 116(1):174–181. doi.​org/​10.​1016/j.​jff.​2012.​03.​005
https://​doi.​org/​10.​1016/j.​ijfoo​dmicro.​2006.​12.​015 Min M, Bunt CR, Mason SL, Hussain MA (2018) Non-dairy probiotic
Fukuda M, Kobayashi M, Honda Y (2017) Functional components and food products: an emerging group of functional foods. Crit Rev
health benefits of fermented soymilk in soft chem. Food Ferm. Food Sci Nutr 59(16):2626–2641
https://​doi.​org/​10.​1016/​B978-0-​12-​811412-​4.​00006-0 Mishra S, Mishra HN (2013) Effect of synbiotic interaction of fruc-
Gangoiti MV, Puertas AI, Hamet MF, Peruzzo PJ, Llamas MG, tooligosaccharide and probiotics on the acidification profile,
Medrano M, Abraham AG (2017) Lactobacillus plantarum textural and rheological characteristics of fermented soy milk.
CIDCA 8327: An α-glucan producing-strain isolated from kefir Food Bioproc Tech 6(11):3166–3176. https://​doi.​org/​10.​1007/​
grains. Carbohyd Polym 170:52–59 s11947-​012-​1021-4
Garrote GL, Abraham AG, De Antoni GL (2001) Chemical and micro- Mishra S, Mishra HN (2018) Comparative study of the synbiotic effect
biological characterisation of kefir grains. J Dairy Res 68(4):639– of inulin and fructooligosaccharide with probiotics with regard to
652. https://​doi.​org/​10.​1017/​S0022​02990​10052​10 the various properties of fermented soy milk. Food Sci Technol
Golowczyc MA, Gugliada MJ, Hollmann A, Delfederico L, Garrote Int 24(7):564–575. https://​doi.​org/​10.​1177/​10820​13218​776529
GL, Abraham AG, Semorile L, De Antoni G (2008) Characteriza- Monajjemi M, Aminin AN, Ilkhani AR, Mollaamin F (2012) Nano
tion of homofermentative lactobacilli isolated from kefir grains: study of antioxidant activities of fermented soy whey prepared
potential use as probiotic. J Dairy Res 75(2):211–217. https://​doi.​ with lactic acid bacteria and kefir. Afr J Microbiol Res 6(2):426–
org/​10.​1017/​S0022​02990​80031​17 430. https://​doi.​org/​10.​5897/​AJMR11.​1249
Granato D, Branco GF, Nazzaro F, Cruz AG, Faria JA (2010) Func- Oliveira RPDS, Perego P, De Oliveira MN, Converti A (2011) Effect
tional foods and non-dairy probiotic food development: trends, of inulin as a prebiotic to improve growth and counts of a

13
World Journal of Microbiology and Biotechnology (2022) 38:133 Page 11 of 11 133

probiotic cocktail in fermented skim milk. LWT-Food Sci Technol Silvestroni A, Connes C, Sesma F, De Giori GS, Piard JC (2002) Char-
44(2):520–523. https://​doi.​org/​10.​1016/j.​lwt.​2010.​08.​024 acterization of the melA locus for α-galactosidase in Lactobacillus
Papadimitriou K, Alegría Á, Bron PA, De Angelis M, Gobbetti M, plantarum. Appl Environ Microbiol 68(11):5464–5471. https://​
Kleerebezem M, Kok J (2016) Stress physiology of lactic acid doi.​org/​10.​1128/​AEM.​68.​11.​5464-​5471.​2002
bacteria. Microbiol Mol Biol Rev 80(3):837–890. https://​doi.​org/​ Simova E, Beshkova D, Angelov A, Hristozova T, Frengova G, Spasov
10.​1128/​MMBR.​00076-​15 Z (2002) Lactic acid bacteria and yeasts in kefir grains and kefir
Raja A, Gajalaksshmi P, Raja MMM, Imran MM (2009) Effect of Lac- made from them. J Ind Microbiol Biotechnol 28(1):1–6. https://​
tobacillus lactis cremoris isolated from kefir against food spoilage doi.​org/​10.​1038/​sj/​jim/​70001​86
bacteria. Am J Food Technol 4(5):201–209. https://​doi.​org/​10.​ Singh BP, Vij S, Hati S, Singh D, Kumari P, Minj J (2015) Antimicro-
3923/​ajft.​2009.​201.​209 bial activity of bioactive peptides derived from fermentation of
Rani VU, Pradeep BV (2015) Antioxidant properties of soy milk fer- soy milk by Lactobacillus plantarum C2 against common food-
mented with Lactobacillus paracasei KUMB B005. Int J Pharm borne pathogens. Int J Ferm Foods 4(1):91–99
Sci Rev Res 30(1):39–42 Souza G, De OL, Vetra AJ (1991) Acceptability of the flavour of soy
Rekha CR, Vijayalakshmi G (2011) Isoflavone phytoestrogens in bean milk yoghurt with added cow milk. Dairy Sci Abst 13(9):729
soymilk fermented with β-glucosidase producing probiotic lactic Valero-Cases E, Frutos MJ (2017) Effect of inulin on the viability of L.
acid bacteria. Int J Food Sci Nutr 62(2):111–120. https://​doi.​org/​ plantarum during storage and in vitro digestion and on composi-
10.​3109/​09637​486.​2010.​513680 tion parameters of vegetable fermented juices. Plant Foods Hum
Saarela M, Virkajarvi I, Nohynek L, Vaari A, Matto J (2006) Fibres Nutr 72(2):161–167. https://​doi.​org/​10.​1007/​s11130-​017-​0601-x
as carriers for Lactobacillus rhamnosus during freeze-drying and Wang YC, Yu RC, Chou CC (2002) Growth and survival of bifidobac-
storage in apple juice and chocolate-coated breakfast cereals. Int teria and lactic acid bacteria during the fermentation and stor-
J Food Microbiol 112(2):171–178. https://​doi.​org/​10.​1016/j.​ijfoo​ age of cultured soymilk drinks. Food Microbiol 19(5):501–508.
dmicro.​2006.​05.​019 https://​doi.​org/​10.​1006/​fmic.​2002.​0506
Saito VST, Dos Santos TF, Vinderola CG, Romano C, Nicoli JR, Wang YC, Yu RC, Chou CC (2006) Antioxidative activities of soymilk
Araújo LS et al (2014) Viability and resistance of lactobacilli fermented with lactic acid bacteria and bifidobacteria. Food
isolated from cocoa fermentation to simulated gastrointestinal Microbiol 23(2):128–135. https://​doi.​org/​10.​1016/j.​fm.​2005.​01.​
digestive steps in soy yogurt. J Food Sci 79(2):M208–M213. 020
https://​doi.​org/​10.​1111/​1750-​3841.​12326 Zhao D, Shah NP (2014) Changes in antioxidant capacity, isoflavone
Schwab C, Vogel R, Gänzle MG (2007) Influence of oligosaccharides profile, phenolic and vitamin contents in soymilk during extended
on the viability and membrane properties of Lactobacillus reuteri fermentation. LWT-Food Sci Technol 58(2):454–462. https://​doi.​
TMW1.106 during freeze-drying. Cryobiology 55(2):108–114 org/​10.​1016/j.​lwt.​2014.​03.​029
Shimakama Y, Matsubara S, Yuki N, Ikeda M, Ishikawa F (2003) Eval-
uation of Bifidobacterium breves strain Yakult fermented soymilk Publisher's Note Springer Nature remains neutral with regard to
as a probiotic food. Int J Food Microbiol 81(2):131–136. https://​ jurisdictional claims in published maps and institutional affiliations.
doi.​org/​10.​1016/​S0168-​1605(02)​00224-6
Shokryazdan P, Sieo CC, Kalavathy R, Liang JB, Alitheen NB, Faseleh
Jahromi M, Ho YW (2014) Probiotic potential of Lactobacillus
strains with antimicrobial activity against some human pathogenic
strains. BioMed Res Int. https://​doi.​org/​10.​1155/​2014/​927268

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