Effects of Silk Fibroin Enzyme Hydrolysates on Memory and Learning: A Review.

Silk protein products have been used for a wide range of applications. This review focuses on the studies conducted relative to cognitive functions with silk fibroin enzyme hydrolysates (FEH) in humans and animals. All known studies reported in PubMed and Google Scholar have been included. Studies have been conducted on children, high school and college students, adults and seniors, ranging in ages from 7-92 years. Doses of 200-600 mg silk FEH per day for three weeks to 16 weeks have been used. Based on these studies, it can be concluded that silk FEH exhibit beneficial cognitive effects with respect to memory and learning, attention, mental focus, accuracy, memory recall, and overall memory and concentration. These conclusions are supported by studies in rats and mice. Mechanistic studies that have been conducted in animals and cell culture systems are also reviewed. These studies indicate that silk FEH exerts its positive effects on memory and learning by providing neuroprotection via a complex mechanism involving its potent antioxidant and inflammation-inhibiting activities. Acetylcholine (ACh) is secreted by cholinergic neurons, and plays a role in encoding new information. Silk FEH were shown to decrease the levels of the pro-oxidant and pro-inflammatory mediators interlukin-1 (IL-1ß), IL-6 and tumor necrosis factor-alpha (TNF-α), protecting the cholinergic system from oxidative stress, thus enhancing ACh levels in the brain, which is known to promote cognitive functions. In addition, the expression of brain-derived neurotrophic factor (BNDF), which is involved in the survival of neurons, is enhanced, and an increase in the expression of the phosphorylated cAMP response element-binding protein (p-CREB) occurs, which is known to play a positive role in cognitive functions. No adverse effects have been reported in association with the use of silk FEH.

Protein Hydrolysate from Pterygoplichthys disjunctivus, Armoured Catfish, with High Antioxidant Activity.

Pterygoplichthys disjunctivus, locally named the armoured catfish, is a by-catch of tilapia fishing that accounts for up to 80% of total captured fish in the Adolfo Lopez Mateos dam, in Michoacán, México, affecting the economy of its surrounding communities. This invasive fish is discarded by fishermen since native people do not consume it, partly due to its appearance, yet it is rich in protein. The aim of this study was to produce hydrolysates from armoured catfish using food-grade proteases (neutrases HT and PF and alcalase PAL) and investigate the processing conditions (pH and temperature) that lead to a high degree of hydrolysis, antioxidant activity, and Angiotensin I-Converting Enzyme (ACE) Inhibitory activity. No other similar research has been reported on this underutilized fish. The antioxidant activity was measured by three different methods, ABTS, FRAP and ORAC, with relevance to food and biological systems in order to obtain a more comprehensive assessment of the activity. In addition, the main peptide sequences were identified. All enzymes produced hydrolysates with high antioxidant activity. In particular, the protease HT led to the highest antioxidant activity according to the ABTS (174.68 µmol Trolox equivalent/g fish) and FRAP (7.59 mg ascorbic acid equivalent/g fish) methods and almost the same as PAL according to the ORAC method (51.43 µmol Trolox equivalent/g fish). Moreover, maximum activity was obtained at mild pH and temperature (7.5; 50 °C). Interestingly, the ORAC values obtained here were higher than others previously reported for fish hydrolysates and similar to those reported for fruits such as blueberries, apples and oranges. The peptide sequence IEE(E) was present in several peptides in both hydrolysates; this sequence may be partly responsible for the high antioxidant activity, particularly the one based on iron-reducing power. These findings will be relevant to the valorization of other fish/fish muscle discards and could contribute to the production of food supplements and nutraceuticals.

Nutrient recycling of lipid-extracted waste in the production of an oleaginous thraustochytrid.

Improving the economics of microalgae production for the recovery of microbial oil requires a comprehensive exploration of the measures needed to improve productivity as well as to reduce the overall processing costs. One avenue for cost reduction involves recycling the effluent waste water remaining after lipid extraction. This study investigates the feasibility of recycling those wastes for growing thraustochytrid biomass, a heterotrophic microalgae, where wastes were generated from the enzymatic extraction of the lipids from the cell biomass. It was demonstrated that secondary cultures of the tested thraustochytrid grown in the recycled wastes performed favorably in terms of cell and oil production (20.48 g cells L(-1) and 40.9 % (w/w) lipid) compared to the control (13.63 g cells L(-1) and 56.8 % (w/w) lipid). Further, the significant uptake of solubilized cell material (in the form of amino acids) demonstrated that the recycled waste has the potential for offsetting the need for fresh medium components. These results indicate that the implementation of a nutrient recycling strategy for industrial microalgae production could be possible, with significant added benefits such as conserving water resources, improving production efficiency, and decreasing material inputs.

Recycling of lipid-extracted hydrolysate as nitrogen supplementation for production of thraustochytrid biomass.

Efficient resource usage is important for cost-effective microalgae production, where the incorporation of waste streams and recycled water into the process has great potential. This study builds upon emerging research on nutrient recycling in thraustochytrid production, where waste streams are recovered after lipid extraction and recycled into future cultures. This research investigates the nitrogen flux of recycled hydrolysate derived from enzymatic lipid extraction of thraustochytrid biomass. Results indicated the proteinaceous content of the recycled hydrolysate can offset the need to supply fresh nitrogen in a secondary culture, without detrimental impact upon the produced biomass. The treatment employing the recycled hydrolysate with no nitrogen addition accumulated 14.86 g L(-1) of biomass in 141 h with 43.3 % (w/w) lipid content compared to the control which had 9.26 g L(-1) and 46.9 % (w/w), respectively. This improved nutrient efficiency and wastewater recovery represents considerable potential for enhanced resource efficiency of commercial thraustochytrid production.

Identification of New Angiotensin-Converting Enzyme Inhibitory Peptides Isolated from the Hydrolysate of the Venom of Nemopilema nomurai Jellyfish.

Recently, jellyfish venom has gained attention as a promising reservoir of pharmacologically active compounds, with potential applications in new drug development. In this investigation, novel peptides, isolated from the hydrolysates of Nemopilema nomurai jellyfish venom (NnV), demonstrate potent inhibitory activities against angiotensin-converting enzyme (ACE). Proteolytic enzymes-specifically, papain and protamex-were utilized for the hydrolysis under optimized enzymatic conditions, determined by assessing the degree of hydrolysis through the ninhydrin test. Comparative analyses revealed that papain treatment exhibited a notably higher degree of NnV hydrolysis compared to protamex treatment. ACE inhibitory activity was quantified using ACE kit-WST, indicating a substantial inhibitory effect of 76.31% for the papain-digested NnV crude hydrolysate, which was validated by captopril as a positive control. The separation of the NnV-hydrolysate using DEAE sepharose weak-anion-exchange chromatography revealed nine peaks under a 0-1 M NaCl stepwise gradient, with peak no. 3 displaying the highest ACE inhibition of 96%. The further purification of peak no. 3 through ODS-C18 column reverse-phase high-performance liquid chromatography resulted in five sub-peaks (3.1, 3.2, 3.3, 3.4, and 3.5), among which 3.2 exhibited the most significant inhibitory activity of 95.74%. The subsequent analysis of the active peak (3.2) using MALDI-TOF/MS identified two peptides with distinct molecular weights of 896.48 and 1227.651. The peptide sequence determined by MS/MS analysis revealed them as IVGRPLANG and IGDEPRHQYL. The docking studies of the two ACE-inhibitory peptides for ACE molecule demonstrated a binding affinity of -51.4 ± 2.5 and -62.3 ± 3.3 using the HADDOCK scoring function.

Porcine Brain Enzyme Hydrolysate Enhances Immune Function and Antioxidant Defense via Modulation of Gut Microbiota in a Cyclophosphamide-Induced Immunodeficiency Model.

This study examined how consuming porcine brain enzyme hydrolysate (PBEH) affects the immune function and composition of the gut microbiota in an immunodeficient animal model. Male Wistar rats aged 6 weeks were fed casein (control), 100 mg/kg body weight (BW), red ginseng extract (positive-control), and 6, 13, and 26 mg PBEH per kg BW (PBEH-L, PBEH-M, and PBEH-H, respectively) daily for 4 weeks. At 30 min after consuming assigned compounds, they were orally administered cyclophosphamide (CTX; 5 mg/kg BW), an immunosuppressive agent, to suppress the immune system by inhibiting the proliferation of lymphocytes. The normal-control rats were fed casein and water instead of CTX. Natural killer cell activity and splenocyte proliferation induced by 1 µg/mL lipopolysaccharide were lower in the control group than the normal-control group, and they significantly increased with PBEH consumption, particularly at high doses. The PBEH consumption increased dose-dependently in the Th1/Th2 ratio compared to the control. The lipid peroxide contents were lower in the PBEH group than in the control group. Moreover, PBEH m and PBEH-H consumption mitigated white pulp cell damage, reduced red pulp congestion, and increased spleen mast cells in the histological analysis. Intestinal microbiota composition demonstrated differences between the groups at the genus levels, with Akkermansia being more abundant in the control group than the normal-control group and the PBEH-H group showing a decrease. However, Bifidobacterium decreased in the control group but increased in the PBEH-H group. The ß-diversity revealed distinct microbial communities of PBEH and positive-control groups compared to the control group (p < 0.05). The metagenome predictions revealed that PBEH-H influenced amino acid metabolism, antioxidant defense, insulin sensitivity, and longevity pathways. In conclusion, PBEH-H intake boosted immune responses and reduced lipid peroxides by modulating gut microbiota composition. These findings suggest that PBEH-H has the potential as a dietary supplement for improving immune function and gut health in individuals with immunodeficiency.

Unlocking the bioactivity of meat proteins: Comparison of meat and meat hydrolysate via simulated gastrointestinal digestion.

Understanding the functional attributes of meat proteins is crucial for determining their nutritional benefits. Depending on the form in which meat proteins are available, the digestive process can release peptides which are valuable for nutrition and may also possess bioactive properties, affecting physiology. Liquid chromatography - mass spectrometry (LC-MS) was used to quantitatively compare the molecular peptide features (representing non-redundant peptides), during the different stages of a simulated gastrointestinal digestion process of a minimally processed powdered meat and its enzymatically produced hydrolysate. Results from a principal component analysis (PCA) indicated that the hydrolysate did not undergo extensive additional digestion whereas the powdered meat was digested both at the gastric and in the intestinal phases. Bioactive peptide sequence prediction identified the meat hydrolysate but not the meat powder as the only source of exact and partial bioactive matches in the angiotensin-I converting enzyme and dipeptidyl peptidase IV inhibition categories. Also, a higher source of cryptides (encrypted bioactive peptides), indicated that meat hydrolysates are potentially a better substrate for the release of these enzyme inhibitory peptides. These observations thus suggest that pre-digestion of a complex food matrix such as meat, may enhance its bioavailability following oral consumption early in the digestion process. SIGNIFICANCE: This work highlights enzymatic hydrolysis of meat proteins prior to ingestion allows for potentially higher bioavailability of bioactive peptides that inhibit angiotensin-I converting enzyme and dipeptidyl peptidase IV, thus possibly aiding high blood pressure and type 2 diabetes management.

Preparation and Characterization of Flavored Sauces from Chinese Mitten Crab Processing By-Products.

To achieve high-value utilization of Chinese mitten crab processing by-products, different types of sauces were prepared using crab legs (CLs) and crab bodies (CBs). Two processing methods, enzymatic hydrolysis and enzymatic hydrolysis coupled with the Maillard reaction, were employed to prepare these sauces. An electronic nose (E-nose) and electronic tongue (E-tongue) were used to measure the changes in the taste and odor, an automatic amino acid analyzer was used to measure the amino acid content, and a headspace solid-phase microextraction GC/MS (HS-SPME-GC/MS) was used to analyze the volatile compounds, qualitatively and quantitatively. The results showed that the sour, bitter, and other disagreeable odors of the enzymatic hydrolysis solution (EHS) were reduced following the Maillard reaction; meanwhile, the umami and saltiness were considerably enhanced. The quantity of free acidic amino acids with an umami and sweet taste in the CL and CB sauces after enzymatic hydrolysis and the Maillard reaction was substantially higher than that in the homogenate (HO). The Maillard reaction solution (MRS) produced more volatile compounds than the HO and EHS, such as aldehydes, pyrazines, ketones, etc. These compounds not only impart a unique flavor but also have antioxidant capabilities, making them a prototype for the high-value utilization of crab processing by-products.

The feasibility of enzyme hydrolysate gross energy for formulating duck feeds.

Two experiments were designed to investigate the feasibility of enzyme hydrolysate gross energy (EHGE) for formulating duck feeds. In experiment 1, six mixed diets and 6 experimental diets (compound feeds) with 20% CP were formulated, and their EHGE, AME, and TME were determined so as to analyze the correlation between EHGE and AME, TME. In experiment 2, six experimental diets with different EHGE levels were further arranged to determine the EHGE requirement for Pekin ducks from hatch to 21 D of age. A total of 384 freshly hatched ducklings was randomly divided into 6 experimental treatments, each treatment containing 8 replicates with 8 ducks per replicate. The results showed that there were a linear correlation between EHGE and AME (r = 0.998, P < 0.01), TME (r = 0.997, P < 0.01) for 6 mixed diets, and the regression models were AME = 0.996 × EHGE-1.062 (R2 = 0.996, P < 0.01), TME = 0.997 × EHGE+0.304 (R2 = 0.995, P < 0.01). For the 6 experimental diets, EHGE was also positively correlated with AME (r = 0.983, P < 0.01), TME (r = 0.984, P < 0.01), and the regression models were AME = 1.2054 × EHGE-3.180 (R2 = 0.967, P < 0.01), TME = 1.2054 × EHGE-1.783 (R2 = 0.967, P < 0.01). According to the broken-line model and optimal BW, the EHGE requirement for ducks from hatch to 21 D of age was 2,937 kcal/kg (calculated value), 3,182 kcal/kg (determined value). In conclusion, EHGE could be used to predict the AME and TME values for mixed diets and compound feeds based on established regression models, and the simulated digestion method in vitro has the potential for effective energy evaluation and formulation for duck feeds.

Evaluation of the prebiotic effects of citrus pectin hydrolysate.

Citrus pectin enzyme hydrolysate (PEH) of different hydrolysis time intervals (6 hours, PEH-6; 12 hours, PEH-12; 24 hours, PEH-24; or 48 hours, PEH-48) or concentrations (1%, 2%, and 4%) was tested for its growth stimulation effect on two probiotics, Bifidobacterium bifidum and Lactobacillus acidophilus. Higher monosaccharide concentrations and smaller molecular weights of PEHs were obtained by prolonging the hydrolysis time. In addition, higher PEH concentrations resulted in significantly higher (p < 0.05) probiotic populations, pH reduction, and increase in total titratable acidity than the glucose-free MRS negative control. Furthermore, significantly higher populations in the low pH environment and longer survival time in nonfat milk (p < 0.05) were observed when the two probiotics were incubated in media supplemented with 2% PEH-24, than in glucose and the negative control. In comparison with other prebiotics, addition of 2% PEH-24 resulted in a more significant increase in the probiotic population (p < 0.05) than in the commercial prebiotics. This study demonstrated that PEH derived from citrus pectin could be an effective prebiotic to enhance the growth, fermentation, acid tolerance, and survival in nonfat milk for the tested probiotics.

Sequential recycling of enzymatic lipid-extracted hydrolysate in fermentations with a thraustochytrid.

This study extends the findings of prior studies proposing and validating nutrient recycling for the heterotrophic microalgae, Thraustochytrium sp. (T18), grown in optimized fed-batch conditions. Sequential nutrient recycling of enzymatically-derived hydrolysate in fermentors succeeded at growing the tested thraustochytrid strain, with little evidence of inhibition or detrimental effects upon culture health. The average maximum biomass obtained in the recycled hydrolysate was 63.68±1.46gL(-1) in 90h the first recycle followed by 65.27±1.15gL(-1) in 90h in the subsequent recycle of the same material. These compared to 58.59gL(-1) and 64.92gL(-1) observed in fresh media in the same time. Lipid production was slightly impaired, however, with a maximum total fatty acid content of 62.2±0.30% in the recycled hydrolysate compared to 69.4% in fresh control media.