Digestive aspartic proteases from sábalo (Prochilodus lineatus): Characterization and application for collagen extraction

Abstract

Acid proteases from sábalo stomach mucosa were recovered using salting-out procedure. This single step produced an enzyme extract purified 1.8-fold over the crude extract with a recovery of 45.1% of its initial proteolytic activity. Sábalo proteases exhibited the highest activity at 45 °C-pH 2.0, showed pH stability between 2.0 and 5.0 and retained more than 70% of its activity after incubation at pH 7.0 for 2 h. Fish extract was unstable at temperatures greater than 45 °C. Its activity was inhibited by pepstatin A but not by PMSF, while EDTA and SDS showed partial inhibitory effects. Presence of CaCl and MgCl increased the proteolytic activity, while increasing concentrations of NaCl strongly decreased it. In addition, compared to the acid extraction method, the use of sábalo enzymatic extract increased 1.7 times the yield of collagen extraction.

Obtainment and characterization of digestive aspartic proteases from the fish Caranx hippos (Linnaeus, 1766)

“…Thus, the result of this test indicates that EE acidic proteases belong to the class of aspartic proteases, as well as pepsins, and because they are stomach enzymes, they are probably of the pepsin-like type. Similar results were found in studies using crude, or partially purified, stomach extracts from fish, in which a high inhibition of acidic proteolytic activity was observed (Bkhairia et al, 2016;Acevedo Gomez et al, 2018). Fish pepsins are strongly inhibited by pepstatin A (Klomklao et al, 2007;Wu et al, 2009;Nalinanon et al, 2010;Cao et al, 2011).…”

“…However, at pH 4.0 the proteolytic activity was greater than 80% demonstrating an optimum pH range between pH 1.5 and 4.0. Similar results were described by Miura et al (2015) for two M. Salmoides pepsins (pH 1.5 and 2.0), for P. lineatus (Acevedo Gomez et al, 2018) and P. orbignyanus (Candiotto et al, 2018) digestive acidicproteases (pH 2.0), Bougatef et al (2008) for M. mustelus pepsins (pH 2.0), Nalinanon et al (2010) for T. alalunga pepsins (pH 2.0) and Tanji et al (2007) for Latimeria chalumnae (Smith, 1939) pepsins (pH 2.0). Bkhairia et al (2016) determined the optimum pH for L. aurata digestive acidic protease activity as pH 3.0, similarly to the report by Vannabun et al (2014) for P. gigas digestive acidic proteases and by Khaled et al (2011) for S. aurita pepsin.…”

“…The resistance to the presence of these elements is an interesting characteristic of C. hippos acidic proteases that could be used without the need to remove metals present in the process. Acevedo Gomez et al (2018) reported that the addition of Mg 2+ stimulated the activity of P. lineatus digestive aspartic proteases, increasing the activity by approximately 10%, while in crevalle jack this ion inhibited 13% of the EE acidic proteolytic activity. Klomklao et al (2007) showed that the activity of C. pectoralis pepsins was potentiated by the divalent cations Ca 2+ and Mg 2+ .…”

“…However, many industrial applications, such as those mentioned above, do not need to use such pure enzymes (Freitas-Júnior and Bezerra, 2015). Recently, some researchers have shown the potential of using crude or partially purified enzymatic extracts, obtained from the fish stomach, to obtain gelatin and fish collagen (Bkhairia et al, 2016;Acevedo Gomez et al, 2018).…”

Digestive Proteases From Fish Processing Wastes: Their Partial Characterization and Comparison

“…The enzymes of Ub were stable at 10 and 30 °C, and these extracts stand out for maintained a high activity at 30 °C, from 11.78 U/mg protein (time 0 min) to 11.6 U/mg protein (time 150 min). Similar results were reported for Sardinella aurita (Ben Khaled et al 2011), S. sagax (Castillo-Yañez et al 2004), P. lineatus(Gomez et al 2018) and P. gigas(Vannabun et al 2014) where optimum temperatures were between 33-55 °C.…”