Enzymatically modified isoquercitrin in soy protein temporarily enhanced the plasma amino-acid concentrations, antioxidant index, and plasma hormone levels: a randomized, double-blind cross-over trial.

This study investigated the effects of a dietary protein supplement containing enzymatically modified isoquercitrin (EMIQ) on plasma amino-acid levels in healthy people. A randomized double-blind cross-over trial (UMIN000044791) was conducted with a sample of nine healthy individuals. These participants ingested soy protein with or without 42 mg EMIQ for 7 days after performing mild exercise. Plasma amino-acid levels were measured before ingestion and at 15, 30, 45, 60, 90, 120, 180, and 240 min after ingestion on the last day. The concentrations of total amino acids at 0 and 120 min and easily oxidized amino acids at 120 min were significantly higher in the plasma of individuals who consumed 42 mg EMIQ. Oxidative stress levels were lower and plasma testosterone levels were higher in participants who ingested soy protein with 42 mg EMIQ than in those who did not. These results suggest that daily ingestion of soy protein with 42 mg EMIQ can be useful for effective protein absorption.

Passion fruit seed extract protects beta-amyloid-induced neuronal cell death in a differentiated human neuroblastoma SH-SY5Y cell model.

Alzheimer's disease (AD) is a progressive neurodegenerative disease with accompanying perceptive disorder. We previously reported that decreasing levels of brain-derived neurotrophic factor (BDNF) promoted beta-amyloid (Aß)-induced neuronal cell death in neuron-like differentiated SH-SY5Y (ndSH-SY5Y) human neuroblastoma cells in an AD mimic cell model. We investigated the neuroprotective effects of passion fruit seed extract (PFSE) and one of the main stilbene compounds, piceatannol, in an AD cell model using ndSH-SY5Y cells. Both PFSE and piceatannol were found to protect Aß-induced neurite fragmentation in the cell model (protection efficacy; 34% in PFSE and 36% in piceatannol). In addition, both PFSE and piceatannol suppress Aß-induced neuronal cell death in the cell model (inhibitory effect; 27% in PFSE and 32% in piceatannol). Our study is the first to report that piceatannol-rich PFSE can repress Aß-induced neuronal cell death by protecting against neurite fragmentation in the AD human cell model. These findings suggest that piceatannol-rich PFSE can be considered a potentially neuroprotective functional food for both prevention and treatment of AD.

Effect of Sucrose on Amino Acid Absorption of Whey: A Randomized Crossover Trial.

Protein intake has been reported to secrete insulin and lower glucose levels, but the effect of carbohydrate and protein co-ingestion on amino acid absorption has not been well documented. A randomized, placebo-controlled, single-blinded, crossover trial was conducted to evaluate the effect of sucrose on blood amino acid levels. Eleven volunteers (both sexes aged 20-60 years with body mass index 21.4 ± 2.4 kg/m2) randomly received one of four test solutions: water (P-group), 10 g sucrose (S-group), 10 g whey protein (W-group), or 10 g whey protein + 10 g sucrose (W-S-group), and blood amino acid concentration, glucose levels, and insulin levels were monitored over 180 min. Following the wash-out period, randomized treatment and blood parameter monitoring were repeated. Consequently, amino acid concentration was significantly lower in the S-group than in the P-group, showing that single ingestion of sucrose decreased blood amino acid levels in a fasted state. However, there was no significant difference between blood amino acid levels of the W- and W-S-groups, suggesting that co-ingestion of sucrose does not affect blood amino acid concentration. Insulin levels were significantly higher in the W-S than in the S-group, and glucose levels were significantly lower in the W-S- than in the S-group, suggesting positive impact on glycotoxicity by reducing blood glucose levels. Therefore, whey protein co-ingestion with sucrose suppresses glucose levels and increases insulin levels as opposed to the sucrose ingestion, but does not affect amino acid absorption of whey protein, indicating that this co-ingestion may not be a problem for protein supplementation.

Effect of antioxidant supplementation on skeletal muscle and metabolic profile in aging mice.

Aging induces drastic changes in muscle mass and function (sarcopenia); however, the detailed mechanisms underlying sarcopenia remain poorly understood. Recent studies suggested that age-related increases in oxidative stress induce muscle atrophy. In this study, we investigated the effect of 6-month supplementation of antioxidants, specifically piceatannol (PIC) and enzymatically modified isoquercitrin (EMIQ), on age-related physiological changes, including skeletal muscle weight and quality, in 25-month-old (OLD) mice, compared to in 4-month-old (young, YNG) C57BL/6J mice. Muscle weight corrected by body weight significantly declined in OLD mice, compared to in YNG mice. The control OLD mice also showed changes in the expression of genes related to muscle fiber type, reduced locomotor activity, and increased oxidative stress markers in blood. Consistent with the muscle weight and quality changes, whole-body fat oxidation during sedentary conditions and exercise periods in control OLD mice was significantly lower than that in YNG mice. Interestingly, compared to the control OLD mice, the PIC- or EMIQ-fed OLD mice showed higher fat oxidation. Furthermore, EMIQ, but not PIC, increased locomotor activity, the expression of genes encoding antioxidant enzymes, and suppressed the carbonylated protein in the skeletal muscle of OLD mice. These results suggested that chronic antioxidant intake could alleviate aging-related muscle function changes.

Amazake made from sake cake and rice koji suppresses sebum content in differentiated hamster sebocytes and improves skin properties in humans.

Amazake is a traditional Japanese health drink. Here, we examined the effects of amazake on skin in cells and humans. Treatment with sake cake or rice koji suppressed intracellular lipid accumulation in differentiated hamster sebocytes, likely through the reduced expression of peroxisome proliferator-activated receptor-gamma (PPARγ) mRNA. In double-blind, placebo-controlled trial, seventeen Japanese women ingested either amazake or placebo for 4 weeks. Ingestion of the amazake decreased the sebum content compared to the placebo. The questionnaires showed improvements in "face color," "dark circles under the eyes," "glossy hair," and "waking up well", only in the amazake. In accordance with the questionnaires, additional analysis revealed the change in the L* values under the eyes was statistically increased in the amazake compared to the placebo. These results indicate that amazake may decrease sebum content in cells and humans and increase the L* values under the eyes, with some additional beneficial effects in humans.

Intake of a Mixture of Sake Cake and Rice Malt Increases Mucin Levels and Changes in Intestinal Microbiota in Mice.

Amazake is a traditional Japanese beverage. Its main ingredients are sake cake and rice malt. In this study, we examined the effect of sake cake and rice malt on the intestinal barrier function and gut microbiota. BALB/c mice were fed a control diet or a diet containing a mixture of sake cake and rice malt powder (SRP) for four weeks. Fecal IgA values did not change between groups, but the fecal mucin level was significantly greater in the SRP-fed group. Gene expression analysis in the ileum by real-time PCR demonstrated Muc2 expression did not change, while the Muc3 expression was upregulated in the SRP-fed group. Furthermore, microbiota analysis demonstrated a change by SRP intake at the family level, and the proportion of Lactobacillaceae significantly increased in the SRP-fed group. At the genus level, the proportion of Lactobacillus also significantly increased in the SRP-fed group. These results suggest that the intake of a mixture of sake cake and rice malt improves intestinal barrier function by increasing mucin levels and inducing changes in intestinal microbiota.

Correction to: Effects of enzymatically modified isoquercitrin in supplementary protein powder on athlete body composition: a randomized, placebo-controlled, doubleblind trial.

The original article [1] contained a typesetting error in Table 3 which was mistakenly introduced by the production team handling this article; this error has now been corrected.

Suppressive effects of whey protein hydrolysate on sucrose-induced hyperglycemia in silkworms.

Silkworms are useful for evaluating substances that suppress postprandial hyperglycemia by oral administration. In this study, orally administered whey protein hydrolysate (WPH), obtained by enzymatic treatment of whey protein, suppressed sucrose-induced hyperglycemia in silkworms in a dose-dependent manner. WPH also inhibited glucose-induced hyperglycemia in silkworms. These findings suggest that WPH contains a bioactive peptide that inhibits glucose uptake from the intestinal tract and thereby suppresses sucrose-induced hyperglycemia.

Effect of piceatannol-rich passion fruit seed extract on human glyoxalase I-mediated cancer cell growth.

Passion fruit seed extract (PFSE), a product rich in stilbenes such as piceatannol and scirpusin B, has various physiological effects. It is unclear whether PFSE and its stilbene derivatives inhibit cancer cell proliferation via human glyoxalase I (GLO I), the rate-limiting enzyme for detoxification of methylglyoxal. We examined the anticancer effects of PFSE in two types of human cancer cell lines with different GLO I expression levels, NCI-H522 cells (highly-expressed GLO I) and HCT116 cells (lowly-expressed GLO I). PFSE and its stilbenes inhibited GLO I activity. In addition, PFSE and its stilbenes supressed the cancer cell proliferation of NCI-H522 cells more than HCT116 cells. These observations suggest that PFSE can provide a novel anticancer strategy for prevention and treatment.

Effects of enzymatically modified isoquercitrin in supplementary protein powder on athlete body composition: a randomized, placebo-controlled, double-blind trial.

BACKGROUND: Enzymatically modified isoquercitrin (EMIQ), a water-soluble quercetin, has been shown to intensify muscle hypertrophy in mice. We investigated the effect of EMIQ in supplementary protein powder on athlete body composition. METHODS: Forty Japanese males who played American football (age: 19.8 ± 1.4 years; body height: 174.1 ± 6.0 cm; body mass: 75.5 ± 10.7 kg) were assigned to a randomized, placebo-controlled, double-blind trial of parallel group. Participants received either EMIQ in whey protein (EW, n = 19) or contrast whey protein (W, n = 20) 6 days per week over 4 months. Body composition was assessed using dual-energy X-ray absorptiometry. Markers of oxidative stress, derivatives of reactive oxygen metabolites (d-ROMs) and biological antioxidant potential (BAP), were assessed using a free radical analytical system. Data were analyzed using a univariate and repeated measures general model statistics. RESULTS: After 4 months, changes in lower limb fat-free mass and muscle mass were significantly greater in the EW group than in the W group (mean change ±95% CI; W: 324.1 ± 284.3, EW: 950.3 ± 473.2, p = 0.031, W: 255.7 ± 288.6, EW: 930.9 ± 471.5, p = 0.021, respectively). Moreover, the EW group exhibited a significantly higher BAP/d-ROMs ratio, antioxidation index, than the W group after 4 months (mean change ± SD; W: 8.8 ± 1.1, EW: 10.3 ± 2.8; p = 0.028). No significant differences in body mass, lean body mass, fat mass, or lower limb fat mass were observed between the groups. CONCLUSION: Ingestion of EMIQ in supplementary protein powder for 4 months exerts antioxidant effects and increases muscle mass among American football players. TRIAL REGISTRATION: University Hospital Medical Information Network Clinical Trial Registry, UMIN000036036 . Retrospectively registered in 2019.

Piceatannol markedly upregulates heme oxygenase-1 expression and alleviates oxidative stress in skeletal muscle cells.

Piceatannol (PIC), a phytochemical, is abundant in passion fruit (Passiflora edulis) seeds. In this study, we investigated the effects of PIC on the expression levels of antioxidant enzymes in C2C12 skeletal muscle cells and compared its effects with those of PIC analogues and polyphenols. We also evaluated its effects on hydrogen peroxide-induced accumulation of reactive oxygen species in C2C12 myotubes. Treatment with PIC led to dose-dependent upregulation of heme oxygenase-1 (Ho-1) and superoxide dismutase 1 (Sod1) mRNA expression in C2C12 myotubes. PIC was the most potent inducer of Ho-1 among the PIC analogues and major polyphenols tested. In addition, treatment with PIC suppressed the hydrogen peroxide-induced increase in intracellular reactive oxygen species levels. Our results suggest that PIC protects skeletal muscles from oxidative stress by activating antioxidant enzymes such as HO-1 and SOD1 and can therefore help prevent oxidative stress-induced muscle dysfunction such as muscle fatigue and sarcopenia.

Effect of Passion Fruit Seed Extract Rich in Piceatannol on the Skin of Women: A Randomized, Placebo-Controlled, Double-Blind Trial.

Piceatannol has been reported to have a wide variety of effects on the skin, including promoting collagen production, inhibiting melanin synthesis, inducing the antioxidant glutathione, and eliminating reactive oxygen species. In this study, a randomized, placebo-controlled, double-blind trial was conducted to clinically evaluate the effects of piceatannol-rich passion fruit seed extract on the skin of healthy Japanese women (age, 35-54 y). Thirty-two women with dry skin received either passion fruit seed extract (5 mg piceatannol) or a placebo (dextrin) for 8 wk. Skin hydration and other parameters on the face were assessed at 0, 4, and 8 wk by using specialized equipment. Furthermore, questionnaire interviews were conducted regarding the physical condition of subjects at 0, 4, and 8 wk. The results showed that consumption of passion fruit seed extract led to significant increases in the moisture content of human skin after 4 and 8 wk compared with that before the trial. The amount of transepidermal water loss decreased over time, although the differences were not significant. Moreover, a stratified analysis of subjects with moisture values of ≤200 µS revealed increased moisture content in the passion fruit seed extract group as compared with the placebo group. Furthermore, the results of questionnaires showed significant reductions in "perspiration" and "fatigue" in the passion fruit seed extract group as compared with the placebo group. These results indicate that oral intake of passion fruit seed extract that is rich in piceatannol could improve the moisture of dry skin and reduce fatigue.

Investigation of epigallocatechin-3-O-caffeoate and epigallocatechin-3-O-p-coumaroate in tea leaves by LC/MS-MS analysis.

(-)-Epigallocatechin-3-O-gallate (EGCG), the major catechin present in green tea, exhibits potent antioxidant activity. We thereby investigated the presence of unknown components bearing the (-)-epigallocatechin (EGC) moiety in fresh tea leaf samples. Initially, liquid chromatography tandem mass spectrometry (LC-MS/MS) was employed to examine fresh tea leaves of the Yabukita, the most popular tea cultivar in Japan, which suggested the presence of the EGC phenylpropanoid derivatives, (-)-epigallocatechin-3-O-p-coumaroate (EGCpCA) and (-)-epigallocatechin-3-O-caffeoate (EGCCA). The structures of the detected EGCpCA and EGCCA were then confirmed by LC-MS/MS using synthesized EGCpCA and EGCCA as standards. In addition, EGCpCA and EGCCA were evaluated for their antioxidant activity in the ORAC (oxygen radical antioxidant capacity) and DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging assays, where EGCCA (8.60µmolTE/µmol, TE=Trolox equivalents) exhibited a stronger antioxidant activity than EGCG (5.52µmolTE/µmol) in the ORAC assay. Finally, EGCpCA and EGCCA were quantitated in several tea leaf samples using LC-MS/MS, and it was found that these compounds were present in lower quantities (EGCpCA, 16.8-345.8µg/g, EGCCA, 4.3-75.1µg/g in the dry tea leaves) than the major catechins. In this study, we found the potent antioxidant EGCCA using LC-MS/MS and revealed its wide existence in various tea leaves.

The Effect of Piceatannol from Passion Fruit (Passiflora edulis) Seeds on Metabolic Health in Humans.

Animal studies have shown the beneficial effects of piceatannol on metabolic health; however, there is a lack of human studies designed to examine these effects. The objective of this study was to investigate the effects of piceatannol on metabolic health in humans. This randomized, placebo-controlled study was conducted on 39 subjects, including 10 overweight men and 9 overweight women (BMI ≥ 25), as well as 10 non-overweight men and 10 non-overweight women (BMI < 25). Subjects received piceatannol (20 mg/day) or placebo capsules for eight weeks in a random order. The primary outcome was the effect of piceatannol on glucose-metabolism, including insulin sensitivity. The secondary outcomes were the effects on other parameters, including blood pressure (BP), heart rate (HR), endothelial function, lipids, inflammation, oxidative stress, mood status, and Sirt1 and phospho-AMP-activated kinase (p-AMPK) expression in isolated peripheral blood mononuclear cells (PBMNCs). Supplementation with piceatannol in overweight men reduced serum insulin levels, HOMA-IR, BP and HR. Other groups, including non-overweight men, as well as overweight and non-overweight women, showed no beneficial effects on insulin sensitivity, BP and HR. Furthermore, piceatannol is not associated with other data, including body weight (BW), body composition, endothelial function, lipids, inflammation, oxidative stress, mood status, and Sirt1/p-AMPK expression in PBMNCs. In conclusion, supplementation with piceatannol can improve metabolic health, including insulin sensitivity, BP and HR, in overweight men.

Evaluation of the innate immune-stimulating activity of amazake using a silkworm muscle contraction assay.

We evaluated the innate immune-stimulating activity of amazake using a silkworm muscle contraction assay. Sake cake, a raw material used to make amazake, had high innate immunity-stimulating activity, whereas rice malt, another raw material used to make amazake, did not, even after fermentation. These results suggest that the silkworm muscle contraction assay is a useful tool to screen foods with high innate immune-stimulating activity and that amazake made from sake cake has immunomodulatory potential.

Enzymatically modified isoquercitrin supplementation intensifies plantaris muscle fiber hypertrophy in functionally overloaded mice.

BACKGROUND: Enzymatically modified isoquercitrin (EMIQ) is produced from rutin using enzymatic hydrolysis followed by treatment with glycosyltransferase in the presence of dextrin to add glucose residues. EMIQ is absorbed in the same way as quercetin, a powerful antioxidant reported to prevent disused muscle atrophy by targeting mitochondria and to have ergogenic effects. The present study investigated the effect of EMIQ on skeletal muscle hypertrophy induced by functional overload. METHODS: In Study 1, 6-week-old ICR male mice were divided into 4 groups: sham-operated control, sham-operated EMIQ, overload-operated control, and overload-operated EMIQ groups. In Study 2, mice were divided into 3 groups: overload-operated whey control, overload-operated whey/EMIQ (low dose), and overload-operated whey/EMIQ (high dose) groups. The functional overload of the plantaris muscle was induced by ablation of the synergist (gastrocnemius and soleus) muscles. EMIQ and whey protein were administered with food. Three weeks after the operation, the cross-sectional area and minimal fiber diameter of the plantaris muscle fibers were measured. RESULTS: In Study 1, functional overload increased the cross-sectional area and minimal fiber diameter of the plantaris muscle. EMIQ supplementation significantly increased the cross-sectional area and minimal fiber diameter of the plantaris muscle in both the sham-operated and overload-operated groups. In Study 2, EMIQ supplementation combined with whey protein administration significantly increased the cross-sectional area and minimal fiber diameter of the plantaris muscle. CONCLUSION: EMIQ, even when administered as an addition to whey protein supplementation, significantly intensified the fiber hypertrophy of the plantaris muscle in functionally overloaded mice. EMIQ supplementation also induced fiber hypertrophy of the plantaris in sham-operated mice.

Piceatannol exhibits anti-inflammatory effects on macrophages interacting with adipocytes.

Piceatannol (PIC), a natural analog of resveratrol (RES), is a phytochemical found in passion fruit seeds. To clarify the effects of PIC on obesity-induced inflammation in adipose tissue, we investigated the anti-inflammatory activity of PIC-related compounds (PIC, RES, and metabolites from PIC) in culture models of obese adipose tissue. Lipopolysaccharide (LPS) and conditioned medium from 3T3-L1 adipocytes (3T3-L1-CM) enhanced proinflammatory gene expression and synthesis of nitric oxide (NO), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) in RAW264.7 macrophages. Although each compound inhibited the mRNA expression of iNOS (inducible NO synthase), TNF-α, and IL-6, PIC potently inhibited them, and 30 µmol/L PIC suppressed the LPS- and 3T3-L1-CM-induced mRNA expression of iNOS (70.4% and 69.2% suppression, respectively), TNF-α (42.6% and 47.0% suppression), and IL-6 (27.3% and 42.1% suppression). PIC also significantly suppressed production of NO (80.3% suppression) and inflammatory cytokines (TNF-α; 33.7% suppression, IL-6; 66.5% suppression). Furthermore, PIC was found to rescue the uncoupling protein 1 mRNA expression induced by isoproterenol in 10T1/2 adipocytes, which was suppressed by LPS-activated macrophages. These results suggest that PIC may attenuate the pathologic inflammation triggered by adipose tissues.

Piceatannol is superior to resveratrol in promoting neural stem cell differentiation into astrocytes.

Piceatannol (3,3',4',5-trans-tetrahydroxystilbene) is a polyphenolic compound abundant in the seeds of passion fruit (Passiflora edulis). Piceatannol is an analogue of resveratrol (3,4',5-trans-trihydroxystilbene) and shares the structural motif and biological activities such as activation of SIRT1. Several studies have shown that piceatannol is more potent than resveratrol. In this study, we examined the effects of piceatannol on neural stem cell differentiation into astrocytes compared with those of resveratrol. At a concentration of 2.5 µM, piceatannol promoted astrocyte differentiation, while resveratrol had no effect at this concentration. Furthermore, we found that oral administration of piceatannol increased the number of astrocytes in the brains of adult mice, while resveratrol administration showed no effects. These results suggest that piceatannol has a superior effect to resveratrol in promoting astrocyte differentiation.

Vascular- and hepato-protective effects of passion fruit seed extract containing piceatannol in chronic high-fat diet-fed rats.

The effects of chronic administration of piceatannol-enriched (9.5% w/w) passion fruit seed extract (PFSE) on the cardiovascular damage induced in a high-fat (HF) diet-fed model of Fischer 344 rats were evaluated. Rats were fed the control, HF, or HF diets containing PFSE (0.5% w/w) for 16 weeks, and the effects of the various diets on the tissue weight, serum lipid profile, hepatic fibrosis, hepatic ductular reaction, cardiac function and aortic ring reactivity were examined. HF diet-fed rats developed signs of cardiovascular disease with abnormal serum profiles compared to control diet-fed rats. PFSE supplementation improved the liver hypertrophy and hepatic histology of the HF diet-fed rats. In addition, the triglyceride and cholesterol levels, platelet aggregation, cardiac function, and acetylcholine-mediated relaxation of the aortic ring were improved. These results suggest that the chronic intake of PFSE containing piceatannol prevents HF diet-induced cardiovascular disease in rats.

Piceatannol and its metabolite, isorhapontigenin, induce SIRT1 expression in THP-1 human monocytic cell line.

Piceatannol is a phytochemical that is present in large amounts in passion fruit (Passiflora edulis) seeds, and is an analog of resveratrol. Recently, the absorption and metabolism of piceatannol were investigated in rats, and isorhapontigenin, O-methyl piceatannol, was detected as a piceatannol metabolite in rat plasma. To elucidate the function of piceatannol and its metabolites, we investigated the expression of sirtuin 1 (SIRT1) in THP-1 monocytic cells after treatment with piceatannol and its metabolites, and compared their effects with those of resveratrol and its metabolites. Piceatannol and resveratrol upregulated the expression levels of SIRT1 mRNA and SIRT1 protein. An extract of passion fruit seeds, which contained high levels of piceatannol, also upregulated SIRT1 mRNA expression. As for the metabolites, isorhapontigenin upregulated SIRT1 mRNA expression, whereas resveratrol glucuronides and sulfate did not affect SIRT1 expression. These findings indicate that after intake of piceatannol, not only piceatannol itself, but also its metabolite, isorhapontigenin, contributed to the upregulation of SIRT1 expression.