Novel antihypercholesterolemic bioactive peptides (BAP) from peptic camel whey protein hydrolysates (CWPH) were generated at different time, temperature, and enzyme concentration (%). Hydrolysates showed higher pancreatic lipase- (PL; except 3 CWPH) and cholesterol esterase (CE)-inhibiting potential, as depicted by lower half-maximal inhibitory concentration values (IC50 values) compared with nonhydrolyzed camel whey proteins (CWP). Peptide sequencing and in silico data depicted that most BAP from CWPH could bind active site of PL, whereas as only 3 peptides could bind the active site of CE. Based on higher number of reactive residues in the BAP and greater number of substrate binding sites, FCCLGPVPP was identified as a potential CE-inhibitory peptide, and PAGNFLPPVAAAPVM, MLPLMLPFTMGY, and LRFPL were identified as PL inhibitors. Molecular docking of selected peptides showed hydrophilic and hydrophobic interactions between peptides and target enzymes. Thus, peptides derived from CWPH warrant further investigation as potential candidates for adjunct therapy for hypercholesterolemia.
Novel bioactive peptides from camel milk protein hydrolysates (CMPH) were identified and tested for inhibition of cholesterol esterase (CEase), and their possible binding mechanisms were elucidated by molecular docking. Papain-generated CMPH showed the highest degree of hydrolysis. All CMPH produced upon enzymatic degradation demonstrated a dramatic enhancement of CEase inhibition compared with intact camel milk proteins, with papain-generated hydrolysate P9 displaying the highest inhibition. Peptide identification and their modeling through PepSite 2 revealed that among 20 potential bioactive peptides in alcalase-generated hydrolysate A9, only 3 peptides, with sequences KFQWGY, SQDWSFY, and YWYPPQ, showed the highest binding toward CEase catalytic sites. Among 43 peptides in 9-h papain-generated hydrolysate P9, 4 peptides were found to be potent CEase inhibitors. Molecular docking revealed that WPMLQPKVM, CLSPLQMR, MYQQWKFL, and CLSPLQFR from P9 hydrolysates were able to bind to the active site of CEase with good docking scores and molecular mechanics-generalized born surface area binding energies. Overall, this is the first study reporting CEase inhibitory potential of peptides generated from milk proteins.
The development of free and total cholesterol nanobiosensors based on a single step electrochemical integration of gold nanoparticles (AuNPs), cholesterol oxidase (COx), cholesterol esterase (CE) and a mediator with polypyrrole (PPy) films is described. The incorporation of the various components in the PPy films was confirmed by chronopotentiometry, cyclic voltammetry (CV), scanning electron microscopy, energy dispersive X-ray analysis (SEM-EDX), and Fourier transformed infrared (FTIR) spectroscopy. The free cholesterol, PPy-NO3--Fe(CN)64--AuNPs-COx, nanobiosensor achieved a minimum detectable concentration of 5 μM, a linear concentration range of 5-25 μM and a sensitivity of 1.6 µA cm-2 µM-1 in 0.05 M phosphate buffer (pH 7.00). For the total cholesterol, PPy-NO3--Fe(CN)64--AuNPs-COx-CE, nanobiosensor which also involved the co-incorporation of cholesterol esterase (CE) with the other components, the achieved performances include a minimum detectable total cholesterol concentration of 25 μM, a broader linear concentration range of 25-170 μM and a lower sensitivity of 0.1 µA µM-1 cm-2. Owing to its high selectivity, the presence of common interferants did not affect the total cholesterol measurement with the PPy-NO3--Fe(CN)64--AuNPs-COx-CE nanobiosensor. Both nanobiosensors were successfully used for direct and indirect determination of total cholesterol in human blood serum samples.
Leucaena leucocephala had been traditionally used to treat diabetes. The present study was designed to evaluate in vitro "insulin-like" activities of Leucaena leucocephala (Lam.) deWit. aqueous fruit extract on lipid and glucose metabolisms. The ability of the extract to stimulate adipogenesis, inhibit lipolysis, and activate radio-labeled glucose uptake was assessed using primary rat adipocytes. Quantitative Real-Time RT-PCR was performed to investigate effects of the extract on expression levels of genes (protein kinases B, AKT; glucose transporter 4, GLUT4; hormone sensitive lipase, HSL; phosphatidylinositol-3-kinases, PI3KA; sterol regulatory element binding factor 1, Srebp1) involved in insulin-induced signaling pathways. L. leucocephala aqueous fruit extract stimulated moderate adipogenesis and glucose uptake into adipocytes when compared to insulin. Generally, the extract exerted a considerable level of lipolytic effect at lower concentration but decreased gradually at higher concentration. The findings concurred with RT-PCR analysis. The expressions of GLUT4 and HSL genes were upregulated by twofold and onefold, respectively, whereas AKT, PI3KA, and Srebp1 genes were downregulated. The L. leucocephala aqueous fruit extract may be potentially used as an adjuvant in the treatment of Type 2 diabetes mellitus and weight management due to its enhanced glucose uptake and balanced adipogenesis and lipolysis properties.
The nutraceutical benefits of β-sitosterol (SIT) are well documented. The present study investigated the in vitro effects of SIT on adipogenesis, glucose transport, and lipid mobilization in rat adipocytes. Primary cultures of rat preadipocytes and differentiated adipocytes were used in this study. Glucose uptake was measured by the uptake of radio-labeled glucose. Adipogenesis and lipolysis were measured by oil-red-O and glycerol quantification methods, respectively. The expression of protein kinase B (Akt), glucose transporter 4 (GLUT4), hormone sensitive lipase (HSL), and phosphatidylinositol-3-kinase (PI3 K) genes in SIT-treated adipocytes were assessed by real-time reverse transcription polymerase chain reaction (RT-PCR). The data showed that SIT induced glucose uptake in adipocytes. It also stimulated adipogenesis in differentiating preadipocytes. Interestingly, although SIT displayed general insulin-mimetic activity by stimulating glucose uptake and adipogenesis, it also induced lipolysis in adipocytes. Furthermore, the SIT-induced lipolysis was not attenuated by insulin and co-incubation of SIT with epinephrine improved epinephrine-induced lipolysis. GLUT4 gene expression was highly down-regulated in SIT-treated adipocytes, compared to insulin-treated adipocytes, which was up-regulated. Insulin- and SIT-treated adipocytes showed similar levels of Akt, HSL, and PI3 K gene down-regulation. These observations suggest that the elevation of glucose uptake in SIT-treated adipocytes was unrelated to de novo synthesis of GLUT4 and the SIT-induced lipolysis is associated with the down-regulation of Akt and PI3K genes. The unique effects of SIT on the regulation of glucose uptake, adipogenesis, and lipolysis in adipocytes show that it has potential to be utilized in diabetes and weight management.
Mushrooms have been used in folk medicine for thousands of years. In this study, the effect of β -glucan-rich extract of P. sajor-caju (GE) on lipid lowering and antioxidant potential was assessed in C57BL/6J mice fed on a high-fat diet. Obesity was induced in C57BL/6J mice by feeding a high-fat diet. The control groups in this study were ND (for normal diet) and HFD (for high-fat diet). The treated groups were ND240 (for normal diet) (240 mg/kg b.w) and HFD60, HFD120, and HFD240 (for high-fat diet), where the mice were administrated with three dosages of GE (60, 120, and 240 mg GE/kg b.w). Metformin (2 mg/kg b.w) served as positive control. GE-treated groups showed significantly reduced body weight, serum lipid, and liver enzymes levels. GE also attenuated protein carbonyl and lipid hydroperoxide levels by increasing the enzymic antioxidants (SOD, CAT, and GPx) activities in the mice. GE-treated groups induced the expression of hormone sensitive lipase (HSL) and adipose triglyceride lipase (ATGL) while downregulated the expression of peroxisome proliferator-activated receptor gamma (PPAR- γ ), sterol regulatory binding protein-1c (SREBP-1c), and lipoprotein lipase (LPL). Hence, GE prevented weight gain in the mice by inducing lipolysis and may be valuable in the formulation of adjuvant therapy for obesity.
The present study investigated the influence of berberine (BBR) supplementation in normal and high-lipid (HL) diets on lipid metabolism and accumulation in black sea bream (Acanthopagrus schlegelii). BBR was supplemented at 50 mg/kg to control (Con, 11·1 % crude lipid) and high-lipid (HL, 20·2 % crude lipid) diets and named as ConB and HLB, respectively. After the 8-week feeding trial, fish body length and specific growth rate were significantly reduced by HL diets (P < 0·05). Muscle and whole-body crude lipid contents were significantly influenced by both BBR supplementation and dietary lipid level. Fish fed the HLB diet had significantly lower serum TAG, LDL-cholesterol contents and alanine aminotransferase activity compared with the HL group. The HL group presented vast lipid accumulation in the liver, and hypertrophied hepatocytes along with large lipid droplets, and translocation of nuclear to the cell periphery. These abnormalities in black sea bream were alleviated in the HLB group. BBR supplementation in the HL diet significantly down-regulated the hepatic expression levels of acetyl-CoA carboxylase α, sterol regulatory element-binding protein-1, 6-phosphogluconate dehydrogenase, glucose 6-phosphate dehydrogenase and pparγ, whereas the lipoprotein lipase, hormone-sensitive lipase and carnitine palmitoyltransferase 1a expression levels were significantly up-regulated. However, the expression levels of these genes showed opposite trends in muscle (except for pparγ). In conclusion, dietary BBR supplementation in the HL diet reduced hepatic lipid accumulation by down-regulating lipogenesis gene expression and up-regulating lipolysis gene expression, and it increased muscle lipid contents with opposite trends of the mechanism observed in the liver.