Purification and identification of angiotensin I-converting enzyme inhibitory peptides from fermented walnut residues

Abstract

Defatted walnut residues (DWR) were hydrolysed using Bacillus subtilis to produce angiotensin-I-converting enzyme (ACE) inhibitory peptides. Fermented walnut residue extract (FWRE) was fractionated using ultrafiltration methods to identify the ACE-inhibitory peptides. The fraction with a molecular weight of <1 kDa (WPs-I) presented the greatest ACE-inhibitory activity. Reverse-phase high-performance liquid chromatography was used to separate WPs-I from the most active fractions, and the amino acid sequence of WPs-I was confirmed as VQTL (459.351 Da) and LGYEN (594.276 Da) by UPLC-ESI-MS/MS. ARTICLE HISTORY

Bioactive Peptides from Walnut Residue Protein

“…It was found that the 50% inhibitory concentration (IC50) values of LPGRPPIKPWP before and after digestion were 0.136 and 0.173 µM/mL, respectively, which confirmed that the peptide was relatively stable during digestion and had the potential of ACE inhibitory activity in vivo [20]. Zheng, Li and Ding reported that the component with molecular weight less than 1kDa in the walnut residue extract fermented by Bacillus subtilis had the largest ACE inhibitory activity [36]. In a word, more and more ACE inhibitory peptides from walnut residue have been successfully prepared, separated and purified ( Table 3).…”

“…After centrifugation and microfiltration, a walnut peptide solution could be obtained [35]. Zheng, Li and Ding successfully prepared an ACE inhibitory peptide by solid-state fermentation of defatted walnut residue with Bacillus subtilis [36]. It should be pointed out that the solid fermentation system has certain advantages over the liquid fermentation system, specifically in the higher enzyme productivity [37], more stable fermentation process [38], higher environmental friendliness [39], which is more suitable for extracting walnut residue resource active peptide.…”

“…The hydrolysate of walnut protein passes through the ultrafiltration membrane with a certain pore size under pressure driving, and WBP can pass through the ultrafiltration membrane smoothly because of its small particle size, while insoluble substrate, enzyme and protein with larger molecular weight remain outside the membrane to achieve the purpose of preliminary purification of walnut peptide. The preliminary purified walnut peptide can pass through the ultrafiltration membrane with different pore sizes in turn, and can also complete the separation of active peptides with different molecular weight ranges [12,19,36,40]. However, with ultrafiltration technology is difficult to achieve the effective separation of peptides with small or very similar molecular weights.…”

“…At present, they are widely used in the analysis of amino acid sequence of walnut peptide, such as electrospray ionization mass spectrometry (ESI-MS) [24,36], matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) [40,41], quadrupole time-of-flight mass spectrometry (Q-TOF-MS) [21,42], determination of clenbuterol by liquid chromatography-tandem mass spectrometry (LC-MS/MS) [19] and so on. In recent years, de novo sequencing method has shown significant advantages in the structural identification of walnut peptides [24,36,43,44]. This technology can complete the de novo sequencing of unknown walnut peptide without any protein database, with high accuracy and sensitivity.…”

“…A large number of literatures have reported that the chain length, amino acid composition, amino acid sequence and spatial structure have a great influence on the activity of ACE inhibitory peptide. Long peptides are difficult to bind to the active sites of ACE, so most of the high active ACE inhibitory peptides have a chain length of 2-12 amino acids, the lower the molecular weight (<3kDa), the higher the inhibitory activity [36,60,61]. However, in many cases, the influence of amino acid type and sequence on ACE inhibitory activity of peptide is much greater than that of peptide chain length.…”