Assaâd Sila’s research while affiliated with University of Gafsa and other places

The industrial processing of Argentine shortfin squid to obtain rings generates a significant amount of protein-rich waste, including the skin, which is rich in collagen and attached myofibrillar proteins. This waste is generally discarded. In this study, skin was used as a source of proteins that were hydrolysed using Trypsin, Esperase® or Alcalase®, which released peptides with antioxidant potential and, in particular, antihypertensive (ACE inhibition), hypoglycemic (DPP-IV inhibition) and/or nootropic (PEP inhibition) potential. Among the three enzymes tested, Esperase® and Alcalase produced hydrolysates with potent ACE-, DPP-IV- and PEP-inhibiting properties. These hydrolysates underwent chromatography fractionation, and the composition of the most bioactive fractions was analysed using HPLC-MS-MS. The fractions with the highest bioactivity exhibited very low IC50 values (16 and 66 µg/mL for ACE inhibition, 97 µg/mL for DPP-IV inhibition and 55 µg/mL for PEP inhibition) and were mainly derived from the hydrolysate obtained using Esperase®. The presence of Leu at the C-terminal appeared to be crucial for the ACE inhibitory activity of these fractions. The DPP-IV inhibitory activity of peptides seemed to be determined by the presence of Pro or Ala in the second position from the N-terminus, and Gly and/or Pro in the last C-terminal positions. Similarly, the presence of Pro in the peptides present in the best PEP inhibitory fraction seemed to be important in the inhibitory effect. These results demonstrate that the skin of the Argentine shortfin squid is a valuable source of bioactive peptides, suitable for incorporation into human nutrition as nutraceuticals and food supplements.

Phycocyanin (PC), a primary light-harvesting pigment derived from Spirulina platensis, finds extensive application owing to its biological properties. In this study, we present an efficient procedure for the extraction of PC pigment, followed by a thorough exploration of its biological activities. A novel extraction method, involving a four-step process including cell lysis through freeze-thawing, maceration, centrifugation, and tangential microfiltration, was employed, resulting in purity index of 1.32. The crude PC was analyzed by UV–visible and FTIR spectroscopy. SDS-PAGE analysis showed two bonds corresponding to α (17 kDa) and β (20 kDa) subunits of PC. The crude PC was examined for its antioxidant activity through diverse antioxidants assays, including assessments of DPPH radical-scavenging capacity, reducing power, and ABTS assay. In addition, assays assessing antimicrobial activity revealed significant inhibitory effects of the crude PC against various bacterial and fungal strains. The inhibitory effect of crude PC on the proliferation of human breast cancer cells (MCF-7) was detected by MTT assay. Interestingly, crude PC was most active with 59.2% of cell proliferation inhibition at 4 mg/mL, while it exhibits greater safety and lower toxicity on normal cell lines (HEK-293). Moreover, the application of crude PC in a dose-dependent manner to MCF-7 cells led to the inhibition of cell migration as observed in the wound healing assay. In summary, these findings suggest that crude PC holds promise for its utilization as valuable functional and bioactive components in the food and nutraceutical sectors.

The objective of this work was to purify the small (705 Da) and hydrophilic antimicrobial Arg-Asp-Arg-Phe-Leu peptide from RuBisCO protein hydrolysate and to evaluate its effect on the microbiological and oxidative stability of beef mince during refrigeration. RuBisCO was obtained from alfafa green juice. The peptide extract was fractionated using RP-HPLC, and the active fractions were analyzed by liquid chromatography, electrospray ionization, and tandem mass spectrometry (LC-ESI-MS). Beef mince was analyzed in the following treatments: negative control, meat with two different BHT concentrations of 0.1 and 0.5% (w/w), and meat with two different Arg-Asp-Arg-Phe-Leu peptide concentrations of 0.1 and 0.5% (w/w). Lipid oxidation using the thio-barbituric acid-reactive substance (TBARS) values were significantly affected by the storage period and the concentration of bioactive peptide. Arg-Asp-Arg-Phe-Leu, a small antibacterial peptide from RuBisCO, can be isolated and purified by HPLC from alfafa green juice with retention time between 10 and 50 min, which corresponds to antimicrobial peptides. RuBisCO peptide Arg-Asp-Arg-Phe-Leu 0.5% increases oxidative stability of beef mince during refrigeration. RuBisCO peptide Arg-Asp-Arg-Phe-Leu inhibit microbial growth under refrigeration for 11 days.

Glycosaminoglycans (GAGs) play a crucial role due to their significant biomedical functions. Chondroitin sulfate (CS) and dermatan sulfate (DS), the main representative family of GAGs, were extracted and purified from garfish (Belone belone) by-products, i.e., skin (GSB), bones (GCB), and heads (GHB), and their composition and anticoagulant activity were investigated. CS/DS were purified by ion-exchange chromatography with yields of 8.1% for heads, 3.7% for skin, and 1.4% for bones. Cellulose acetate electrophoresis was also explored for analyzing the extracted CS/DS. Interestingly, GHB, GSB, and GCB possessed sulfate contents of 21 ± 2%, 20 ± 1%, and 20 ± 1.5%, respectively. Physico-chemical analysis showed that there were no significant differences (p > 0.05) between the variances for sulfate, uronic acid, and total sugars in the GAGs extracted from the different parts of fish. Disaccharide analysis by SAX-HPLC showed that the GSB and GCB were predominately composed of ΔDi-4S [ΔUA-GalNAc 6S] (74.78% and 69.22%, respectively) and ΔDi-2,4S [ΔUA2S-GalNAc 4S] (10.92% and 6.55%, respectively). However, the GHB consisted of 25.55% ΔDi-6S [ΔUA-GalNAc 6S] and 6.28% ΔDi-2,6S [ΔUA2S-GalNAc 4S]. Moreover, classical anticoagulation tests were also used to measure their anticoagulant properties in vitro, which included the activated partial thromboplastin time, prothrombin time, and thrombin time. The CS/DS isolated from garfish by-products exhibited potent anticoagulant effects. The purified CS/DS showed exceptional anticoagulant properties according to this research and can be considered as a new agent with anticoagulant properties.

Tub gurnard is a highly abundant fishery species caught as a discard in the Mediterranean Sea. This work proposes its valorisation through the release of potential antihypertensive peptides and glycosaminoglycans (GAGs) through the controlled hydrolysis of tub gurnard skin proteins. Four proteases (Esperase, Alcalase, Trypsin and Pronase E) were used to obtain potent angiotensin converting enzyme I (ACE)-inhibitory hydrolysates. Peptides and GAGs were separated and evaluated for their antihypertensive potential by fluorometry. The peptide-rich fractions derived from the Esperase and Alcalase hydrolysates showed very low IC50 values (47 and 68 μg/mL, respectively). Only the GAGs from the Trypsin and Esperase hydrolysates were relevant ACE inhibitors (63 and 52% at 1 mg/mL, respectively). The peptide composition of the most potent ACE-inhibitory fractions derived from the Esperase and Alcalase hydrolysates (IC50 values of 33 and 29 μg/mL, respectively) was analysed by RP-LC-ESI-MS/MS. The analysis suggests that the ACE-inhibitory activity is related to the peptide hydrophobicity, as well as to the presence of specific residues at any of the last four C-terminal positions. The in silico gastrointestinal digestion of these fractions yielded small peptides with antihypertensive potential.

An efficient synthesis of new γ,δ‐insaturated δ‐lactam and glutarimide derivatives bearing a phosphonomethyl group from a common allylphosphonate precursor is described. Our approach is based on a two‐step procedure involving the preparation of phosphonated‐1,5‐ketoester and ‐1,5‐diester followed by an amidation‐heterocyclization sequence. The first step proceeds via Michael's addition of ethyl acetoacetate and diethyl malonate on an allylphosphonate starting material. The second step consists of a base‐promoted intramolecular amidation‐cyclization sequence with primary amines, which accounts for the construction of δ‐lactams and glutarimide skeletons. We performed the evaluation of angiotensin I‐converting enzyme (ACE) inhibition using an in vitro enzyme assay on six new compounds. Five compounds showed potent ACE inhibitory activity, with IC50 values ranging from 0.02 to 0.27 mg/mL. Compared with Captopril, used as a reference drug, two new glutarimide derivatives exhibited higher efficiency ACE inhibition activity.

Pistacia vera by-products are discarded by the pistachio processing industries, notably leading to environmental problems. By-products such as external green hull represent a major part of the nut. Thus, there is an urgent need to find suitable applications for this by-product. In this context, this study was designed to promote the management of these by-products rich of biologically active molecules. The present study reports the extraction method of pistachio external hull polysaccharides (PHP) and their protective effects against the hepatotoxicity and nephrotoxicity in vivo caused by CCl4 and cisplatin in rats, respectively. Results showed that CCl4 and cisplatin administration altered the hepatic and renal functions. Whereas PHP treatment restored the increased plasmatic biomarkers of hepatotoxicity through deacreasing the percentage of aspartate transaminase (AST), alanine transaminase (ALT), and lactate dehydrogenase (LDH) by 23.23, 98.97, and 81.39%, respectively. PHP reestablished also plasmatic biomarkers of nephrotoxicity (creatinine, urea, and uric acid) and their levels were reduced by 17.5, 14.59 and 14.81%, respectively. PHP normalized plasmatic lipid profile, in so far as total cholesterol (TC), triglyceride (TG), low-density lipoprotein-cholesterol (LDL-C), and high-density lipoprotein-cholesterol (HDL-C), were altered by CCl4. In addition, PHP pre-co-treatment markedly improved the oxidative stress biomarkers, as evidenced by the reduction of lipid peroxidation by 50 and 67.56% in both liver and kidney tissues. It was also marked by the elevation of the superoxyde dismutase (SOD), glutathion peroxydase (GSH-Px), and catalase activities by 19.89, 31.15, and 27.57% compared with CCl4-treated rats and by 20.49, 40.55, and 61.82%, respectively, compared with cisplatin-treated group. The detected potential role of PHP in liver and kidney tissues were supported by histopathologic examination. Due to their antioxidant activity, PHP’s monosaccharides could be considered as powerful agents able to prevent the hepatic and renal tissue. The obtained results confirmed the medicinal benefits of pistachio external hull and highlighted the potential beneficial reuse of hulls as a source of bio-antioxidants for the pharmaceutical industries. Graphic Abstract