In vitro stability of bioactive peptides derived from fermented soy milk against heat treatment, pH and gastrointestinal enzymes

doi:10.1016/j.lwt.2018.01.066

LWT

Volume 91, May 2018, Pages 303-307

https://doi.org/10.1016/j.lwt.2018.01.066 Get rights and content

Highlights

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Antioxidative and ACE-inhibitory activity of peptides enhanced after heating treatments.
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ACE-inhibitory peptides was resistant to pH and in vitro gastrointestinal digestion.
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Antioxidant activity was increased after trypsin, pepsin and pancretin treatments.
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Soy bioactive peptides from fermented soy milk have potential as ingredients for the development of functional foods.

Abstract

In this study, bioactive peptides (antioxidative and ACE-inhibitory), which had previously been identified in soy milk fermented by a Lactobacillus plantarum strain C2, were examined for stability during processing and after in vitro digestion. Results indicate that, antioxidative and ACE-inhibitory activity of peptides was either retained or enhanced after applying diverse heating conditions (from 25 to 121 °C). Conversely, the peptides retained almost the same ACE-inhibitory activity before and after treatment with pH from 2 to 7, but their ABTS radical scavenging activity was slightly decreased. Moreover, the significant increment (11–35%) in antioxidant activity was documented with different concentrations of trypsin, pepsin and pancretin, whereas ACE-inhibitory peptides showed some resistance to gastrointestinal digestion. This study indicates the high stability of ACE-inhibitory peptides against temperature of processing and gastrointestinal digestion. The ability of soy peptides to resist in various thermal and in vitro gastrointestinal conditions may be useful for their application as an ingredients for the development of functional foods.

Introduction

Several in vitro and in vivo studies have demonstrated that bioactive dietary peptides positively influence human health via a wide range of biological functions including antioxidative, antihypertensive, antimicrobial, antiobesity, hypocholesterolemic, anticancer, immunomodulatory etc. (Nongonierma and FitzGerald, 2017, Singh et al., 2014). However, it is necessary that bioactive peptides remain intact during food processing and gastrointestinal (GI) digestion before reaching to their target sites to accomplish physiological effects. Therefore, delivery and bioavailability of peptides offers many challenges to the food technologist, pharmacist and clinicians, because a bioactive peptide does not always imply a physiological effect in vivo after oral administration (Vermeirssen, Van Camp, & Verstraete, 2004).

Biologically active peptides can be hydrolyzed during different stages of the GI digestion before being transferred and absorbed to the intestinal epithelium (Picariello et al., 2010). The GI tract is known to be one of the major barriers in the human body. The conditions in the GI tract, such as digestive enzymes and pH values in the stomach might influence the structures and functions of the peptides (Ao & Li, 2013).

Similarly, peptides can be hydrolyzed by high temperature treatment applied during food processing (Wang et al., 2017). Therefore, resistance of bioactive peptides to GI barriers and processing temperatures must be evaluated along with their health promoting properties. In this context, several studies have proved that in vitro digestion methods are useful tools to analyze structural changes, bioavailability and digestibility of bioactive compounds (Chen and Li, 2012, Hur et al., 2011).

Previously, we have characterized 51 peptides from 10 kDa fraction of soy milk fermented by Lactobacillus plantarum strain C2. Among them, 17 peptides were showing both antioxidant and ACE-inhibitory activities (Singh & Vij, 2017). This study was designed to assess the effect of temperature and simulated GI conditions on the integrity of this 10 kDa bioactive peptides fraction. A two-stage in vitro GI environment (pH and digestive enzymes) was used to assess the potential changes in the biofunctional activities (antioxidative and ACE-inhibitory) of peptides.

Section snippets

Preparation of peptide fraction

Lyophilized Peptides fraction (10 kDa), which had previously been prepared from the cell free supernatant of fermented soy milk using molecular weight cut-off (MWCO) membrane, was reconstituted in sterile double distilled water. The peptide content of the reconstituted peptide fraction was 0.628 ± 0.02 mg/ml, determined by OPA method and mentioned in our previous article (Singh & Vij, 2017).

Treatments of peptides

Thermal stability of peptide fraction was analyzed by treatment with different temperatures (25, 37, 55,

Thermal stability

Thermal stability of bioactive peptides are important because food products underwent several heat treatments before reach to the market. Usually, thermal treatment can cause protein denaturation, association, and aggregation. The significant improvement (P < 0.01) was observed in antioxidant activity (ABTS) at 25 °C (12.67%), 75 °C (13.21%) and 100 °C (12.58%) (Fig. 1A). Similarly, the DPPH radical scavenging activity was also increased (P < 0.01) at 75 °C (17.09%) and 100 °C (17.52%) (Fig. 1

Conclusion

In conclusion, this work has demonstrated the stability of antioxidative and ACE-inhibitory bioactive peptides against thermal treatment and in vitro GI conditions. However, both the activities were retained or enhanced against most of the treatments, suggesting that these bioactive peptides fractions can be incorporated in functional foods. On the other hand, soy food fermented with specific lactobacilli having these bioactive peptides can be used for specific health benefits followed by

Conflicts of interest

No conflict of interest declared.

Acknowledgements

BPS wish to thank “Ministry of Social Justice and Empowerment” and “University Grants Commission, Government of India for providing fellowship during Ph.D.

References (23)

J. Ao et al.
Stability and antioxidative activities of casein peptide fractions during simulated gastrointestinal digestion in vitro: Charge properties of peptides affect digestive stability
Food Research International
(2013)
W. Brand-Williams et al.
Use of a free radical method to evaluate antioxidant activity
LWT-Food Science and Technology
(1995)
M. Chen et al.
The effect of molecular weights on the survivability of casein-derived antioxidant peptides after the simulated gastrointestinal digestion
Innovative Food Science & Emerging Technologies
(2012)
D.W. Cushman et al.
Spectrophotometric assay and properties of the angiotensin-converting enzyme of rabbit lung
Biochemical Pharmacology
(1971)
E. Escudero et al.
Stability of ACE inhibitory ham peptides against heat treatment and in vitro digestion
Food Chemistry
(2014)
P. García-Mora et al.
Identification, functional gastrointestinal stability and molecular docking studies of Lentil peptides with dual antioxidant and Angiotensin I converting enzyme inhibitory activities
Food Chemistry
(2017)
S.J. Hur et al.
In vitro human digestion models for food applications
Food Chemistry
(2011)
J.S. Hwang
Impact of processing on stability of angiotensin I-converting enzyme (ACE) inhibitory peptides obtained from tuna cooking juice
Food Research International
(2010)
M. Maeno et al.
Identification of an antihypertensive peptide from casein hydrolysate produced by a proteinase from Lactobacillus helveticus CP790
Journal of Dairy Science
(1996)
A.B. Nongonierma et al.
Strategies for the discovery and identification of food protein-derived biologically active peptides
Trends in Food Science & Technology
(2017)

G. Picariello et al.

Peptides surviving the simulated gastrointestinal digestion of milk proteins: Biological and toxicological implications

Journal of Chromatography B

(2010)

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¹: Present address: Department of Dairy Microbiology, College of Dairy Science and Technology, Guru Angad Dev Veterinary and Animal Sciences University (GADVASU), Ludhiana – 141004, Punjab, India.

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Short communicationIn vitro stability of bioactive peptides derived from fermented soy milk against heat treatment, pH and gastrointestinal enzymes

Highlights

Abstract

Introduction

Section snippets

Preparation of peptide fraction

Treatments of peptides

Thermal stability

Conclusion

Conflicts of interest

Acknowledgements

Food Research International

LWT-Food Science and Technology

Innovative Food Science & Emerging Technologies

Biochemical Pharmacology

Food Chemistry

Food Chemistry

Food Chemistry

Food Research International

Journal of Dairy Science

Trends in Food Science & Technology

Journal of Chromatography B

Short communication
In vitro stability of bioactive peptides derived from fermented soy milk against heat treatment, pH and gastrointestinal enzymes