In Vitro and In Silico Approaches to Generating and Identifying Angiotensin-Converting Enzyme I Inhibitory Peptides from Green Macroalga Ulva lactuca

Abstract

A protein extract was generated from the macroalga Ulva lactuca, which was subsequently hydrolysed using the food-grade enzyme papain and angiotensin-converting Enzyme I and renin inhibitory peptides identified using a combination of enrichment strategies employing molecular weight cutoff filtration and mass spectrometry analysis. The generated hydrolysates with the most promising in vitro activity were further purified using preparative RP-HPLC and characterised. The 1 kDa hydrolysate (1 kDa-UFH), purified and collected by preparative RP-HPLC at minutes 41‒44 (Fr41‒44), displayed statistically higher ACE-I inhibitory activities ranging from 96.91% to 98.06%. A total of 48 novel peptides were identified from these four fractions by LC-MS/MS. A simulated gastrointestinal digestion of the identified peptide sequences was carried out using in silico enzyme cleavage simulation tools, resulting in 86 peptide sequences that were further assessed for their potential activity, toxicity and allergenicity using multiple predictive approaches. All the peptides obtained in this study were predicted to be non-toxic. However, 28 out of the 86 novel peptides released after the in silico gastrointestinal digestion were identified as potential allergens. The potential allergenicity of these peptides should be further explored to comply with the current labelling regulations in formulated food products containing U. lactuca protein hydrolysates.

Soybean (Glycine max) Protein Hydrolysates as Sources of Peptide Bitter-Tasting Indicators: An Analysis Based on Hybrid and Fragmentomic Approaches

“…Moreover, it is the least expensive source of protein, which may be helpful in solving feeding and agronomical problems [44]. However, hydrolysis of proteins, also these from soybean, contributes to the production of peptides that taste bitter and are not accepted by Western consumers, even if they offer a health-beneficial value [45,46]. Our initial bioinformatic predictions also confirmed the potential of soybean proteins as the richest sources of bitter-tasting sequences when comparing them to the other sequences of proteins derived from grains, oil, and leguminous plants (21 sequences in total; data not shown).…”

“…The BIOPEP-UWM database was applied to predict the umami taste of peptides derived from clam M. meretrix Linnaeus [64]. Garcia-Vaquero et al [45] applied the hybrid approach to assess the bitter taste properties of novel peptide ACE inhibitors derived from U. lactuca (green algae). To this end, green algae proteins were in vitro (enzyme used: papain) and in silico (tool used: PeptideCutter) hydrolyzed.…”

“…Such a correlation is defined as the Q-rule, according to which the peptide likely to be bitter should have Q over 1400 cal/mole [66]. Based on the Q-rule, it was found that the following green algae-originating sequences: SAGVLPWK (GV, VL), GAAPTPPSPPPATKPSTPPKPPT (PK, KP, PPP), IECCLLFALV (LL), PVGCLPK (PK), DAVEIWRVK (VE, EI), DEVIPGAL (VI), PKPPALCN (PK, KP), and PPNPPNPPN, were characterized with Q-values ranging from 1440 to 1743.33 cal/mol, which suggested they could taste bitter [45]. Defining these sequences as "parent peptides" and implementing the fragmentomic idea, we found that seven out of eight such peptides contained motifs (in bold) described as bitter-tasting indicators (provided in brackets).…”

Current knowledge on the extraction, purification, identification, and validation of bioactive peptides from seaweed

“…Recently, membrane filtration was applied with three MWCO, i.e., 10, 3, and 1 kDa, when processing an U. lactuca protein hydrolysate at laboratory-scale. The 3 kDa permeate fraction had the highest renin inhibitory activity [71]. On the other hand, Beaulieu et al [85] reported that the >10 kDa (retentate) of Saccharina longicruris hydrolysates exhibited higher antibacterial activity compared to <10 kDa permeate fraction.…”

“…A total of 21 potential ACE inhibitory peptides from a thermolysin hydrolysate of Grateloupia asiatica, aqueous extracts were identified using the ExPASy PeptideCutter tool (https://web.expasy.org/ peptide_cutter/) [108]. In a study by Garcia-Vaquero et al [71], the ExPASy PeptideCutter tool was used to simulate the in silico digestion of 48 peptides identified in a hydrolysate of U. lactuca resulting in the identification of 16 peptides with potential ACE and renin inhibitory activity based on information provided in the BIOPEP database and PeptideRanker. The BIOPEP database was also used to indicate that AVVK and VIAE identified in a hydrolysate of Mastocarpus stellatus had previously been reported as antibacterial peptides [100].…”