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. 2024 Jun 27;22(7):297.
doi: 10.3390/md22070297.

Generation, Characterisation and Identification of Bioactive Peptides from Mesopelagic Fish Protein Hydrolysates Using In Silico and In Vitro Approaches

Maria Hayes  1 , Azza Naik  1 , Leticia Mora  2 , Bruno Iñarra  3 , Jone Ibarruri  3 , Carlos Bald  3 , Thibault Cariou  4 , David Reid  4 , Michael Gallagher  5 , Ragnhild Dragøy  6   7 , Jorge Galino  8 , Alba Deyà  8 , Sissel Albrektsen  9 , Lars Thoresen  9 , Runar G Solstad  6
Affiliations

Generation, Characterisation and Identification of Bioactive Peptides from Mesopelagic Fish Protein Hydrolysates Using In Silico and In Vitro Approaches

Maria Hayes et al. Mar Drugs. .

Abstract

This study generated bioactive hydrolysates using the enzyme Alcalase and autolysis from mesopelagic fish, including Maurolicus muelleri and Benthosema glaciale. Generated hydrolysates were investigated for their bioactivities using in vitro bioassays, and bioactive peptides were identified using mass spectrometry in active hydrolysates with cyclooxygenase, dipeptidyl peptidase IV and antioxidant activities. In silico analysis was employed to rank identified peptide sequences in terms of overall bioactivity using programmes including Peptide Ranker, PrepAIP, Umami-MRNN and AntiDMPpred. Seven peptides predicted to have anti-inflammatory, anti-type 2 diabetes or Umami potential using in silico strategies were chemically synthesised, and their anti-inflammatory activities were confirmed using in vitro bioassays with COX-1 and COX-2 enzymes. The peptide QCPLHRPWAL inhibited COX-1 and COX-2 by 82.90% (+/-0.54) and 53.84%, respectively, and had a selectivity index greater than 10. This peptide warrants further research as a novel anti-inflammatory/pain relief peptide. Other peptides with DPP-IV inhibitory and Umami flavours were identified. These offer potential for use as functional foods or topical agents to prevent pain and inflammation.

Keywords: anti-diabetic; anti-inflammatory; circular economy; heart health; hydrolysates; in silico analysis; mesopelagic fish; peptides; type-2-diabetes.

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Conflict of interest statement

Ragnhild Dragøy is employed by Aker BioMarine. All other authors declare that there are no potential conflicts of interest. Aker BioMarine has no role in the study design, collection, analysis, interpretation of data, the writing of this article or the decision to submit it for publication.

Figures

Figure 1
Figure 1
Distribution of oil, hydrolysate, solids and bones generated using different enzyme combinations applied to Spanish mesopelagic trawls consisting of the species M. muelleri. Error bars represent SD (n = 3). Different letters indicate where a significant difference exists between samples at 95% confidence.
Figure 2
Figure 2
(a) COX-1 inhibition by whole mesopelagic hydrolysates generated with different proteolytic enzymes. Samples were assayed at a concentration of 1 mg/mL and compared to a commercial control. Assays were performed in triplicate (n = 9). (b) COX-2 inhibition by whole mesopelagic hydrolysates generated with different proteolytic enzymes. Samples were assayed at a concentration of 1 mg/mL and compared to a commercial control, Resveratrol. Assays were performed in triplicate (n = 9). Irish hydrolysates: 14 corresponds to CE21004 Haul4 (Code 14) Benthosema glaciale biomass hydrolysed with Alcalase®; 2 corresponds to CE21004 Haul2 (Code 2) Notocopelus elongtus kroyeri hydrolysed with Alcalase®; 23 corresponds to CE21009 Haul23 Maurolicus muelleri biomass hydrolysed with Alcalase®; 13 corresponds to CE21004 Haul13 (Code 13) Maurolicus muelleri mixed biomass hydrolysed with Alcalase®. Norwegian hydrolysates: Bromelain corresponds to M. muelleri biomass hydrolysed with Bromelain; MaxiPro corresponds to M. muelleri biomass hydrolysed with MaxiPro enzymes; Corolase corresponds to M. muelleri biomass hydrolysed with Corolase and Roholase corresponds to M. muelleri hydrolysed with the Roholase enzyme. Spanish hydrolysates: MMD02 and MME02 correspond to M. muelleri hydrolysed with Alcalase 2.4 LG; MMD06 and MME06 correspond to M. muelleri hydrolysed with Papain; MMD10 and MME10 correspond to M. muelleri hydrolysed with Bromelain; MMB010 and MMD18 correspond to M. muelleri hydrolysed with Protamex; MMA058 and MMD14 correspond to M. muelleri hydrolysed with Papain and Bromelain and MMB034 and MMC019 correspond to hydrolysates generated from M. muelleri using endogenous enzymes.
Figure 3
Figure 3
Antioxidant activity of whole, mesopelagic hydrolysates generated with different proteolytic enzymes. Samples were assayed using the ABTS antioxidant assay at a concentration of 1 mg/mL and compared to a commercial control Resveratrol. Assays were performed in triplicate (n = 9). Samples are represented by code, as shown in Figure 2.
Figure 4
Figure 4
Renin inhibition by whole, mesopelagic hydrolysates generated with different proteolytic enzymes. Samples were assayed using the renin inhibition assay method described in the materials and methods section at a concentration of 1 mg/mL and compared to commercial and internal controls SB, SP and Captopril. Assays were performed in triplicate (n = 9). Samples are represented by codes, as shown in Figure 2.
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