Purification and identification of hypocholesterolemic peptides from freshwater clam hydrolysate with in vitro gastrointestinal digestion

Abstract

The muscles of freshwater clams were extracted separately using hot water. Subsequently, the edible muscle part was freeze‐dried, hydrolyzed at 50°C using Protamex to obtain the freshwater clam hydrolysate (PX), and then digested with pepsin. The bile‐acid‐binding capacity and inhibition of cholesterol micelle formation were subsequently investigated using pepsin‐digested PX (PXP) through ultrafiltration (UF) fractionation or size exclusion chromatography. After the UF treatment, soluble protein (171.0 mg/g) and peptide (109.4 mg/g) contents in fraction III were found to be higher than those of all other membrane fractions. Assuming that bile acid binds to PXP at 100%, the relative bile‐acid‐binding capacities of fractions I, II, and III were 161.2%, 64.3%, and 55.1%, respectively. Fraction f showed the highest inhibitory efficiency ratio (IER), and its inhibition‐peptide content percentage was 831.5% mg/mL. The amino acid sequences of two hypocholesterolemic peptides were Val–Lys–Pro and Val–Lys–Lys, with IERs of 64.8% and 10.2% mg/mL, respectively. Practical applications The edible muscle part of freshwater clam after hot water extraction can be recovered as value‐added by‐product. After hydrolysis by commercial and digestive proteases, the edible muscle part of clams, including the viscera, showed higher bile acid binding capacity and exhibited in vitro inhibition percentages of cholesterol micellar solubility. Novel hypocholesterlemic peptides (Val‐Lys‐Pro and Val‐Lys‐Lys) were identified from the freshwater clam hydrolysate. Notably, this hydrolysate was confirmed to have hypocholesterolemic and hypolipidemic effects in vivo. Additionally, in vivo antihypertensive effects were confirmed in spontaneously hypertensive rats. This freshwater clam hydrolysate is expected to be a useful ingredient in physiologically functional foods for the prevention of hypertension and hypercholesterolemia.

Purification and Identification of Cholesterol Micelle Formation Inhibitory Peptides of Hydrolysate from High Hydrostatic Pressure-Assisted Protease Hydrolysis of Fermented Seabass Byproduct

“…In recent years, numerous studies have investigated protein hydrolysates or peptides as natural compounds for novel hypocholesterolemic activities. To obtain hypocholesterolemic peptides, many studies have focused on the protein hydrolysates of various food proteins such as β-lactoglobulin [ 6 ], beans [ 9 , 10 , 11 ], Atlantic horse mackerel [ 12 ], and clams [ 13 , 14 , 15 ]. Nagaoka et al [ 6 ] demonstrated that the trypsin hydrolysate of β-lactoglobulin can inhibit CMF and purified and identified the first hypocholesterolemic peptide, Ile-Ala-Glu-Lys.…”

“…In addition, the protein hydrolysate of clam meat (16.6% in the diet) was fed to Sprague–Dawley rats with hyperlipidemia induced by high cholesterol for 4 weeks, and the results signified that the hydrolysate accompanied a significant reduction in cholesterol (cholesterol content in plasma and the liver were reduced by 26.1% and 50.5%, respectively) and blood lipids [ 14 ]. After further purification of the clam meat hydrolysate through column chromatography and identification of protein sequence, new peptide sequences that inhibited CMF were obtained, namely Val-Lys-Pro and Val-Lys-Lys [ 15 ]. Nonetheless, information on the hypocholesterolemic effects of protein hydrolysates and peptides is still scarce.…”

New Lignanamides with Antioxidant and Anti-Inflammatory Activities Screened Out and Identified from Warburgia ugandensis Combining Affinity Ultrafiltration LC-MS with SOD and XOD Enzymes

“…Based on the diversities of affinity capabilities between ligand-enzyme complexes after incubation, those potential ligands in groups with the active enzymes displayed bigger peak areas than those of with inactivated enzymes. For further evaluation of the affinity binding strength between the potential ligands and the target enzymes, the relative binding affinity (RBA) was employed to compare the variation of the correlated peak areas in the UF-HPLC-UV chromatograms before and after activation [ 33 ]. The RBA formula is expressed as RBA = A active /A inactivated , where the A active and A inactivated represent the peak areas obtained from the WUE-A4 samples with activated and inactivated SOD or XOD, respectively.…”

Amino Acid, Mineral, and Degree of Hydrolysis of Vinegar-Egg and Its Lipid Lowering and Antioxidant Effects in vitro and in vivo

“…Functional food exerts hypolipidemic and antiatherogenic effects via a promotion of cholesterol catabolism and inhibition of intestinal absorption of cholesterol to improve lipid profiles (Alhaj et al 2010;Lin et al 2017). The results of cholesterol micellar solubility inhibition ability showed that the inhibition of vinegaregg concentrates (at 2, 4, 6, 8, 10, 12 and 14 mg/mL), were 11.3, 23.5, 34.4, 51.7, 51.2, 50.6 and 52.1%, respectively (Figure 1(B)).…”