236
Enzymatic Hydrolysis (In Vitro)
The most significant approach for the production of bioactive peptides is the hydrolysis of parent proteins by using multiple proteases or using a single protease. The
enzymatic hydrolysis is reported to have profound advantages over fermentation
approaches. Unlike fermentation enzymatic hydrolysis is usually rapid and controllable. Also, the optimization of the hydrolysis factors like pH, composition of buffer, temperature and time of reaction results in reproducible profiles of molecular
weight and composition of peptides (Clemente 2000; Rizzello et al. 2016). Thus
enzymatic hydrolysis may be considered as one of the most suited methods for the
generation of target bioactive peptides. Traditionally at laboratory scale, the protein
hydrolysis is carried in solution format, but other methods are also available like
enzyme immobilization approach that allows the reduction in production cost and
scale up of the process (Piovesana et al. 2018).
The enzymes like pepsin, trypsin, alcalase, chymotrypsin, etc., are being used in
the enzymatic hydrolysis. These enzymes can mimic gastrointestinal digestion
effects on the food peptides (Tavano 2013). Enzymatic hydrolysis process is also
much more valuable and milder compared to chemical treatment in generating bioactive peptides. The enzymes can be site unspecific or specific, since enzymes are
not equivalent to one another. Particularly, the site-specific enzymes like trypsin
allows peptide identification using the established proteomics technologies, as the
space for search is strongly reduced when the site of cleavage is known (Piovesana
et al. 2018). On the contrary, the use of site-specific enzymes in large-scale productions is not mostly affordable due to high costs. Thus site-unspecific enzymes like
alcalase can be used. However, the specificity of the information obtained is lost and
thus complicating the finishing step of data management. In recent past, number of
studies has been carried out in this direction and the peptidiomics based on modern
approaches has been found of great help in identification of peptides in such cases
(Rizzello et al. 2016).
ACE-inhibitory peptides from enzymatic digestion have been extensively studied (Hernendez-Ledesma et al. 2007; Murray and FitzGerald 2007; Otte et al.
2007a, b). Studies where trypsin was used to digest β-lactoglobulin found whey
protein hydrolysates to show strong antihypertensive activity (da Costa et al. 2007;
Ferreira et al. 2007; Roufik et al. 2006). Other proteolytic enzymes such as alcalase,
subtilisin, and thermolysin are used in conjunction with pepsin and trypsin to generate peptides with known biological activities (Agyei and Danquah 2011). Enzymes
from fungal and bacterial sources has also been utilized to achieve the same
(Mohanty et al. 2016).
Liberation of peptides with a wide range of actions can thus be released using
different enzymes. Although the bioactivity of peptides has been extensively
reported in literature, the mechanism of action is not well understood. Few studies
hypothesize it as a structure-activity relationship, while others suggest that the
S. Maqsood et al.
Enzymatic Hydrolysis (In Vitro)
The most significant approach for the production of bioactive peptides is the hydrolysis of parent proteins by using multiple proteases or using a single protease. The
enzymatic hydrolysis is reported to have profound advantages over fermentation
approaches. Unlike fermentation enzymatic hydrolysis is usually rapid and controllable. Also, the optimization of the hydrolysis factors like pH, composition of buffer, temperature and time of reaction results in reproducible profiles of molecular
weight and composition of peptides (Clemente 2000; Rizzello et al. 2016). Thus
enzymatic hydrolysis may be considered as one of the most suited methods for the
generation of target bioactive peptides. Traditionally at laboratory scale, the protein
hydrolysis is carried in solution format, but other methods are also available like
enzyme immobilization approach that allows the reduction in production cost and
scale up of the process (Piovesana et al. 2018).
The enzymes like pepsin, trypsin, alcalase, chymotrypsin, etc., are being used in
the enzymatic hydrolysis. These enzymes can mimic gastrointestinal digestion
effects on the food peptides (Tavano 2013). Enzymatic hydrolysis process is also
much more valuable and milder compared to chemical treatment in generating bioactive peptides. The enzymes can be site unspecific or specific, since enzymes are
not equivalent to one another. Particularly, the site-specific enzymes like trypsin
allows peptide identification using the established proteomics technologies, as the
space for search is strongly reduced when the site of cleavage is known (Piovesana
et al. 2018). On the contrary, the use of site-specific enzymes in large-scale productions is not mostly affordable due to high costs. Thus site-unspecific enzymes like
alcalase can be used. However, the specificity of the information obtained is lost and
thus complicating the finishing step of data management. In recent past, number of
studies has been carried out in this direction and the peptidiomics based on modern
approaches has been found of great help in identification of peptides in such cases
(Rizzello et al. 2016).
ACE-inhibitory peptides from enzymatic digestion have been extensively studied (Hernendez-Ledesma et al. 2007; Murray and FitzGerald 2007; Otte et al.
2007a, b). Studies where trypsin was used to digest β-lactoglobulin found whey
protein hydrolysates to show strong antihypertensive activity (da Costa et al. 2007;
Ferreira et al. 2007; Roufik et al. 2006). Other proteolytic enzymes such as alcalase,
subtilisin, and thermolysin are used in conjunction with pepsin and trypsin to generate peptides with known biological activities (Agyei and Danquah 2011). Enzymes
from fungal and bacterial sources has also been utilized to achieve the same
(Mohanty et al. 2016).
Liberation of peptides with a wide range of actions can thus be released using
different enzymes. Although the bioactivity of peptides has been extensively
reported in literature, the mechanism of action is not well understood. Few studies
hypothesize it as a structure-activity relationship, while others suggest that the
S. Maqsood et al.