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.

Effects of 12-Week Multivitamin and Omega-3 Supplementation on Micronutrient Levels and Red Blood Cell Fatty Acids in Pre-menopausal Women.

The purpose of this study was to validate the efficacy of a customized vitamin-mineral supplement on blood biomarkers in pre-menopausal females. Women (21-40 years old) who were apparently healthy were recruited from the local community (ClinicalTrials.gov trial registration NCT03828097). Pretesting (PRE) occurred in the morning 5 ± 2 days following each participant's menses and involved a fasted blood draw, body mass assessment, and blood pressure assessment. Participants were then randomly assigned in a double-blinded fashion to either the multivitamins (MV) (n = 43) or placebo group (n = 51). Participants consumed two capsules per day with breakfast for 12 weeks. Following the trial, participants reported to the laboratory for POST assessments, which replicated PRE procedures. Red blood cell fatty acid and serum micronutrient analyses were performed in a blinded fashion at hematology laboratories. A group × time interaction was observed for serum vitamin D levels (p < 0.001). MV increased levels from PRE to POST (+43.7%, p < 0.001), whereas no change occurred in the placebo group. Additionally, 78% of MV participants at PRE exhibited inadequate vitamin D levels (<40 ng/dl), whereas only 30% exhibited levels below this threshold at POST. An interaction was also observed for serum folate levels (p < 0.001). MV increased serum folate from PRE to POST (p < 0.001), whereas no change occurred in the placebo group. Red blood cell omega-3 fatty acid content increased from PRE to POST in the MV group (p < 0.001) and placebo group (p < 0.05), although POST values were greater in the MV group (p < 0.001). An interaction was observed for serum HDL cholesterol levels (p = 0.047), and a non-significant increase in this variable from PRE to POST occurred in the MV group (p = 0.060). Four-day food recalls indicated MV increased intake of omega-3 fatty acids, vitamin D, folate, and other micronutrients. In summary, MV supplementation increased serum vitamin D, serum folate, and red blood cell omega-3 fatty acid levels. However, these data are limited to healthy females, and more research is needed to examine if MV can affect metabolic disturbances in individuals with micronutrient deficiencies.

Highly Bioavailable Forms of Curcumin and Promising Avenues for Curcumin-Based Research and Application: A Review.

Curcumin exerts a wide range of beneficial physiological and pharmacological activities, including antioxidant, anti-amyloid, anti-inflammatory, anti-microbial, anti-neoplastic, immune-modulating, metabolism regulating, anti-depressant, neuroprotective and tissue protective effects. However, its poor solubility and poor absorption in the free form in the gastrointestinal tract and its rapid biotransformation to inactive metabolites greatly limit its utility as a health-promoting agent and dietary supplement. Recent advances in micro- and nano-formulations of curcumin with greatly enhanced absorption resulting in desirable blood levels of the active forms of curcumin now make it possible to address a wide range of potential applications, including pain management, and as tissue protective. Using these forms of highly bioavailable curcumin now enable a broad spectrum of appropriate studies to be conducted. This review discusses the formulations designed to enhance bioavailability, metabolism of curcumin, relationships between solubility and particle size relative to bioavailability, human pharmacokinetic studies involving formulated curcumin products, the widely used but inappropriate practice of hydrolyzing plasma samples for quantification of blood curcumin, current applications of curcumin and its metabolites and promising directions for health maintenance and applications.

The fallacy of enzymatic hydrolysis for the determination of bioactive curcumin in plasma samples as an indication of bioavailability: a comparative study.

BACKGROUND: Numerous health benefits have been demonstrated for curcumin which is extracted from turmeric (Curcuma longa L). However, due to its poor absorption in the free form in the gastrointestinal tract and rapid biotransformation, various formulations have been developed to enhance its bioavailability. Previous studies indicate that the free form of curcumin is more bioactive than its conjugated counterparts in target tissues. Most curcumin pharmacokinetics studies in humans designed to assess its absorption and bioavailability have measured and reported total (free plus conjugated) curcumin, but not free, bioactive curcumin in the plasma because enzymatic hydrolysis was employed prior to its extraction and analysis. Therefore, the bioavailability of free curcumin cannot be determined. METHODS: Eight human subjects (4 male, 4 female) consumed a single dose of 400 mg curcumin in an enhanced absorption formulation, and blood samples were collected over 6 h. Plasma was treated either with or without glucuronidase/sulfatase prior to extraction. Curcumin and its major metabolites were analyzed using HPLC-tandem mass spectrometry. In addition, the literature was searched for pharmacokinetic studies involving curcumin using PubMed and Google Scholar, and the reported bioavailability data were compared based on whether hydrolysis of plasma samples was used prior to sample analysis. RESULTS: Hydrolysis of blood plasma samples prior to extraction and reporting the results as "curcumin" obscures the amount of free, bioactive curcumin and total curcuminoids as compared to non-hydrolyzed samples. As a consequence, the data and biological effects reported by most pharmacokinetic studies are not a clear indication of enhanced plasma levels of free bioactive curcumin due to product formulations, leading to a misrepresentation of the results of the studies and the products when enzymatic hydrolysis is employed. CONCLUSIONS: When enzymatic hydrolysis is employed as is the case with most studies involving curcumin products, the amount of free bioactive curcumin is unknown and cannot be determined. Therefore, extreme caution is warranted in interpreting published analytical results from biological samples involving ingestion of curcumin-containing products. TRIAL REGISTRATION: ClinicalTrails.gov, trial identifying number NCT04103788 , September 24, 2019. Retrospectively registered.

Effect of a Fibroin Enzymatic Hydrolysate on Memory Improvement: A Placebo-Controlled, Double-Blind Study.

The consumption of a specifically prepared silk fibroin protein enzymatic hydrolysate (FPEH) has been reported to improve cognitive function in healthy humans. The objective of the current study is to evaluate the dose-dependent effects of the FPEH on memory. Healthy adults with an average age of approximately 55 years were administered doses of 0, 280, 400 and 600 mg of FPEH per day in two divided doses for 3 weeks. The Rey-Kim Auditory Verbal Learning Test and the Rey-Kim Complex Figure Test of the Rey-Kim Memory Test were used to evaluate memory at baseline and after 3 weeks. The scores for each test were combined into the memory quotient score (MQ). Learning gradient, memory maintenance, retrieval efficacy, and drawing/recall scores were also compared. After 3 weeks of FPEH, dose-dependent increases were observed for the MQ, the learning gradient, the numbers of words remembered, the retrieval efficiency, and drawing/recall. The optimal dose for FPEH was 400 or 600 mg, depending on the end point measured. No adverse effects were reported. FPEH significantly improved measurements of memory in healthy adults by 3 weeks at doses over 280 mg daily, with an apparent plateau effect at 400-600 mg daily.

A Comparative Pharmacokinetic Assessment of a Novel Highly Bioavailable Curcumin Formulation with 95% Curcumin: A Randomized, Double-Blind, Crossover Study.

OBJECTIVE: Curcumin exhibits many beneficial health-promoting characteristics. However, its poor oral absorption precludes its general use. This study assessed the bioavailability of a novel curcumin formulation compared to 95% curcumin and published results for various other curcumin formulations. METHODS: A randomized, crossover, double-blind, comparator-controlled pharmacokinetic study was performed in 12 healthy adult subjects to determine the appearance of free curcumin and its metabolites curcumin sulfate and curcumin glucuronide in plasma after a single dose of a novel proprietary curcumin liquid droplet micromicellar formulation (CLDM) and unformulated 95% curcumin powder in capsule form. An equivalent 400-mg dose of each product was administered. The 95% curcumin contained 323 mg curcumin, and the CLDM contained 64.6 mg curcumin. Blood samples were drawn and plasma was analyzed for curcumin and its 2 conjugates without enzymatic hydrolysis by liquid chromatography-tandem mass spectroscopy. RESULTS: Plasma levels of curcumin sulfate and curcumin glucuronide after 1.5 hours from CLDM were approximately 20 and 300 ng/mL, respectively, whereas the levels for 95% curcumin were near baseline. Free curcumin reached a maximum level of 2 ng/mL for CLDM and 0.3 ng/mL for 95% curcumin at 1.5 hours. For the CLDM, a small secondary free curcumin peak occurred at 12 hours and a tertiary 1.5-ng/mL peak occurred at 24 hours. The total curcumin absorbed as represented by the area under the curve (AUC)/mg administered curcumin for CLDM was 522 times greater than for the 95% curcumin. CONCLUSIONS: The novel CLDM formulation facilitates absorption and produces exceedingly high plasma levels of both conjugated and total curcumin compared to 95% curcumin. A comparison of the Cmax/mg curcumin and AUC/mg of administered curcumin for CLDM with data from pharmacokinetic studies of various enhanced absorption formulations indicate that the greatest absorption and bioavailability are produced with the novel CLDM formulation.

A long journey to effective obesity treatments: is there light at the end of the tunnel?

As the obesity epidemic continues, more Americans are getting fatter, having more weight-related problems such as cardiovascular disease, and are experiencing new metabolic dysfunctions. For over 50 years, the adipose tissue (AT), commonly referred to as fat, has been of interest to academic and clinical scientists, public health officials and individuals interested in body composition and image including much of the average public, athletes, parents, etc. On one hand, efforts to alter body shape, weight and body fat percentage still include bizarre and scientifically unfounded methods. On the other hand, significant new scientific strides have been made in understanding the growth, function and regulation of anatomical and systemic AT. Markers of transition/conversion of precursor cells that mature to form lipid assimilating adipocytes have been identified. Molecular 'master' regulators such as peroxisome proliferator-activated receptor gamma and CCAAT-enhancer-binding proteins were uncovered and regulatory mechanisms behind variables of adiposity defined and refined. Interventions including pharmaceutical compounds, surgical, psychosocial interventions have also been tested. Has all of the preceding research helped alleviate the adverse physiologies of overweight and/or obese people? Does research to date point to new modalities that should be the focus of efforts to rid the world of obesity-related problems in the 21st century? This review provides a general overview of scientific efforts to date and a provocative view of the future for adiposity.

The Effect of Nutritional Supplements on Muscle-Derived Stem Cells in vitro.

Postnatal muscle stem cells, recognized as myogenic satellite cells, were isolated from sheep skeletal muscle and used in these experiments. Forty-one different metabolic compounds that are commonly found in commercially-available oral supplements were exposed to primary muscle stem cell cultures, in an effort to ascertain whether any one compound could alter satellite cell proliferation or differentiation (a first step towards elucidating the metabolomics or nutrigenomics of these stem cells). These compounds included energetic moieties, amino acid analogs, fatty acids and analogs including different forms of conjugated linoleic acid, minerals and mineral conjugates, insect hormones, caffeine, plant extracts, and extracts from over-the-counter supplements, and were obtained by key manufacturers in a form that would be commercially available. The compounds were sterilized and then exposed to myogenic satellite cell cultures at different levels (ranging from toxic to physiologic) to ascertain if there would be an effect. The results suggested that exposure of satellite cells to only a few compounds resulted in any measurable effect(s). Ten compounds elicited increases in proliferation, and four compounds promoted increases in differentiation. These results suggest avenues for the exploration of enhancing muscle stem cell activity of interest for muscle wasting disorders, sarcopenia of aging and physical performance.

Absorption, uptake and tissue affinity of high-molecular-weight hyaluronan after oral administration in rats and dogs.

The purpose of this study was to determine the absorption, distribution and excretion of (99m)technetium-labeled, high-molecular-weight hyaluronan (((99m)Tc-HA) and (99m)technetium pertechnetate ((99m)Tc-P) after single dose, oral administration to Wistar rats and Beagle dogs. A pilot study utilized (99m)Tc-HA alone, and a second confirmatory study compared uptake of labeled (99m)Tc-HA with (99m)Tc-P. Urinary and fecal excretion after (99m)Tc-HA ingestion by rats showed 86.7-95.6% of radioactivity was recovered, almost all in feces. All tissues examined showed incorporation of radioactivity from (99m)Tc-HA starting at 15 min and persisting for 48 h, in a pattern significantly different from (99m)Tc-P. Whole-body scintigraphs and close-ups of the ventral chest region showed nonalimentary radioactivity from (99m)Tc-HA concentrated in joints, vertebrae and salivary glands four hours after administration. Autoradiography of skin, bone and joint tissue pieces after 24 h showed incorporation of radioactivity from (99m)Tc-HA, but not from (99m)Tc-P. Conversely, absorption, distribution and excretion of (99m)Tc was completely different from (99m)Tc-HA, showing an expected pattern of rapid absorption and excretion in urine, with accumulation in thyroid glands, stomach, kidney and bladder. This report presents the first evidence for uptake and distribution to connective tissues of orally administered, high-molecular-weight HA.

The effects of ergogenic compounds on myogenic satellite cells.

PURPOSE: A series of studies were conducted in which compounds commonly shown to be ergogenic aids for strength athletes if taken orally were evaluated for their ability to directly induce postnatal muscle stem cell proliferation or differentiation/fusion in vitro. METHODS: Compounds tested were creatine monohydrate, creatine pyruvate, L-glutamine, dehydroepiandrosterone (DHEA), androstenedione, Ma Huang (Ephedra sinensis) extract, and Zhi Shi (Citrus aurantium) extract. Dulbecco's modified eagle medium, supplemented with minimal levels of serum and antibiotics, was used as the initial vehicle for the test compounds. Subsequently, a defined treatment medium termed ITTC was used. Satellite cells were exposed to the test compounds for the indicated times and then evaluated by counting mononucleated and multinucleated (fused) cells. RESULTS: In serum-containing media, none of the treatment groups displayed increased proliferation over that of the control. However, in the differentiation cultures, 0.10% creatine monohydrate increased differentiation over that of the control cultures. When 0.10% creatine monohydrate was added to defined media formulations, all treatments but one demonstrated increased differentiation over the 0.5% serum control. Time course experiments, which followed the effect of 0.10% creatine monohydrate contained in ITTC defined media over 120 h, suggested that cells exposed to this treatment differentiated earlier and to a greater level than cells exposed to ITTC alone. CONCLUSIONS: Creatine in the monohydrate form induced differentiation of myogenic satellite cells. Other agents examined did not increase satellite cell proliferation or differentiation. These results provide initial evidence for a mechanistic understanding of observed effects in vivo of increased muscular size and strength from creatine supplementation.

Randomized, double-masked, placebo-controlled clinical study of the effectiveness of zinc acetate lozenges on common cold symptoms in allergy-tested subjects.

This report of a randomized, double-masked, placebo-controlled clinical study demonstrates the effectiveness of zinc acetate lozenges on common cold symptoms in allergy-tested subjects. Subjects in the zinc and placebo groups were evenly matched with respect to sex, race/ethnicity, allergy test status, and age. Overall symptom duration was significantly less in the zinc group than in the placebo group (mean, 3.8 day vs 5.1 days). The mean severity rating for all symptoms was lower in the zinc group than in the placebo group; this difference, however, was not statistically significant. Allergy-positive subjects who used zinc had a statistically significant shorter duration of nasal symptoms than allergy-negative subjects (3.5 days vs 7.6 days). In conclusion, we propose that zinc acetate lozenges may significantly shorten the duration of common cold symptoms and relieve symptoms associated with allergies.