Milk-clotting and hydrolytic activities of an aspartic protease from Salpichroa origanifolia fruits on individual caseins.

In recent years, many attempts have been made to find new plant proteases to make artisan cheeses. The global increase in cheese consumption, together with a lower supply and increasing cost of calf rennet, religious factors (Islam and Judaism) and food choices (vegetarianism) have led to the search for suitable rennet substitutes for milk clotting. This study describes the milk-clotting and hydrolytic activities of an aspartic protease from Salpichroa origanifolia fruits (SoAP) on individual caseins to explore its potential use as an alternative to animal rennet. The milk-clotting index obtained for SoAP was 8.4 times lower than that obtained for chymosin. SoAP showed a higher degree of hydrolysis on α-casein than on the other fractions under the proposed conditions. RP-HPLC, mass spectrometry analyses and sequencing of the hydrolysates allowed identifying five peptides from α-casein, one peptide from ß-casein, and three peptides from k-casein. In silico analysis showed that the peptides identified may display a wide variety of potential biological activities. These results demonstrate the possibility of using SoAP for the manufacture of new types or artisan cheeses, with the simultaneous added value of the potential health-promoting benefits of the bioactive peptides generated during the hydrolysis.

Adding value to the chia (Salvia hispanica L.) expeller: Production of bioactive peptides with antioxidant properties by enzymatic hydrolysis with Papain.

Chia expeller is a by-product of the extrusion process of chia seeds generated during oil production. Typically, this material is non-utilized or used for non-valuable applications. In the present work, the chia expeller was hydrolysed with Papain and the antioxidant properties of the resultant peptides were evaluated. Papain treatment of the chia seed expeller demonstrated an enrichment of low molecular weight peptides (molecular weight <15 kDa) as determined by SDS-PAGE and MALDI-TOF/MS analyses. Such peptides showed a potent radical scavenging effect in vitro against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) radicals in comparison with those non-hydrolysed samples. Taken together our results demonstrate the generation of functional peptides from the chia expeller by enzymatic hydrolysis with Papain. This value-added hydrolysate can be potentially included as a supplement in functional food and nutraceutical products.

Potential antioxidant peptides produced from whey hydrolysis with an immobilized aspartic protease from Salpichroa origanifolia fruits.

An aspartic protease from Salpichroa origanifolia fruits was successfully immobilized onto an activated support of glutaraldehyde agarose. The immobilized enzyme presented higher thermal stability than the free enzyme from 40°C to 50°C and high reusability, retaining 54% of the initial activity after ten cycles of the process. Whey protein concentrates (WPC) were hydrolyzed with both free and immobilized enzyme, reaching a similar degree of hydrolysis of approximately 6-8% after 20h. In addition, the immobilized derivate hydrolyzed α-lactalbumin protein with a higher affinity than ß-lactoglobulin. The hydrolysate was ultra-filtrated, and the fractions were evaluated for antioxidant activities with the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity method. The fraction containing peptides with a molecular mass below 3kDa demonstrated a strong radical quenching effect (IC50: 0.48mg/ml). These results suggest that hydrolyzed WPC could be considered as a promising source of natural food antioxidants for the development of functional food.

Cyclodextrin production by cyclodextrin glycosyltransferase from Bacillus circulans DF 9R.

Cyclodextrins (CD) are cyclic oligosaccharides with multiple applications in the food, pharmaceutical, cosmetic, agricultural and chemical industries. In this work, the conditions used to produce CD with cyclodextrin glycosyltransferase from Bacillus circulans DF 9R were optimized using experimental designs. The developed method allowed the partial purification and concentration of the enzyme from the cultural broth and, subsequently, the CD production, using the same cassava starch as enzyme adsorbent and as substrate. Heat-treatment of raw starch at 70 degrees C for 15 min in the presence of adsorbed cyclodextrin glycosyltransferase allowed the starch liquefaction without enzyme inactivation. The optimum conditions for CD production were: 5% (w/v) cassava starch, 15 U of enzyme per gram of substrate, reaction temperature of 56 degrees C and pH 6.4. After 4h, the proportion of starch converted to CD reached 66% (w/w) and the weight ratio of alpha-CD:beta-CD:gamma-CD was 1.00:0.70:0.16.