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Production of an antihypertensive peptide from milk by the brown rot fungus Neolentinus lepideus

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Abstract

Fermented milk has been shown to have beneficial effects on human health exerted through its bioactive peptides. The lactose-fermenting brown rot fungus Neolentinus lepideus has been shown to ferment milk; however, its ability to produce bioactive peptides has not been explored. This study, therefore, aimed to investigate the ability N. lepideus for the production of antihypertensive peptides in fermented milk and to characterize the peptides. Here we have used 9% skim milk as the substrate and fermented it with the mycelia of N. lepideus. The fermented milk exhibited higher angiotensin I-converting enzyme (ACE) inhibitory activity as compared to the commercially available sour milk containing Ile-Pro-Pro and Val-Pro-Pro. As a result of isolation and purification by reverse-phase high-performance liquid chromatography, followed by peptide sequencing and MS analysis, the dipeptide Tyr-Pro (YP) was identified as an active peptide. The concentration of YP in the fermented milk reached up to 450 μg/ml, which was > twofold higher than the IC50 for ACE inhibition. Subsequently, the antihypertensive effect of YP was validated through intravenous injection of the peptide, which exhibited a significant lowering of blood pressure in a rat model of genetic hypertension. In addition, the transcriptomic analysis revealed the probable role of the carboxypeptidases for the increased yield of YP. These results together indicated that the fermentation with basidiomycete fungus could be a viable approach to produce bioactive peptides, and may, therefore, be applicable in the development of a cost-effective mass production system for functional peptides.

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Abbreviations

ACE:

Angiotensin I-converting enzyme

RP-HPLC:

Reverse-phase high-performance liquid chromatography

HPLC:

High-performance liquid chromatography

ESI–MS:

Electrospray ionization mass spectrometry

YP:

Dipeptide Tyr-Pro

RAAS:

Renin-angiotensin aldosterone system

TFA:

Trifluoroacetic acid

IPP:

Ile-Pro-Pro

VPP:

Val-Pro-Pro

FPKM:

Fragments per kilobase of transcript per million mapped reads

SHR:

Spontaneously hypertensive rat

SHRSP:

Stroke-prone spontaneously hypertensive rat

MAP:

Mean arterial pressure

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Acknowledgements

This work was supported by the Project of the NARO Bio-oriented Technology Research Advancement Institution (Research program on development of innovative technology) [Grant number 28010A]. We would like to thank Editage (https://www.editage.jp) for English language editing.

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Authors and Affiliations

  1. Department of Chemistry and Biotechnology, Graduate School of Engineering, Tottori University, 4-101 Koyama, Tottori, 680-8552, Japan

    Kenji Okamoto, Sayo Kawamura, Mizuki Tagawa & Toshifumi Mizuta

  2. Department of Functional Pathology, Shimane University School of Medicine, Izumo, 693-8501, Japan

    Hasan M. Zahid & Toru Nabika

Authors
  1. Kenji Okamoto
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  2. Sayo Kawamura
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  3. Mizuki Tagawa
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  4. Toshifumi Mizuta
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  5. Hasan M. Zahid
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  6. Toru Nabika
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Contributions

KO conceptualized and designed the study, performed the experiments, drafted the initial manuscript, reviewed the manuscript, and revised the manuscript. SK and MT performed the experiments and analyzed the data. TM contributed to the amino acid analysis, peptide sequencing, and MS analysis. HMZ performed the animal experiments. TN designed animal experiments and reviewed the manuscript. All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work.

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Correspondence to Kenji Okamoto.

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The authors declare no conflict of interest.

Compliance with ethics requirements

The procedures of animal experiments were reviewed and approved by the local committee for animal research at Shimane University. The animal experiments were performed in accordance with relevant guidelines and regulations.

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Okamoto, K., Kawamura, S., Tagawa, M. et al. Production of an antihypertensive peptide from milk by the brown rot fungus Neolentinus lepideus. Eur Food Res Technol 246, 1773–1782 (2020). https://doi.org/10.1007/s00217-020-03530-y

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  • DOI: https://doi.org/10.1007/s00217-020-03530-y

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