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The immunomodulatory properties of exopolysaccharides (EPSs) produced by Streptococcus thermophilus have not been explored in depth. In addition, there are no comparative studies of the functional properties of EPSs produced by streptococci in different food matrices. In this work, EPSs from S. thermophilus SBC8781 were isolated after soy milk (EPS-s) or cow milk (EPS-m) fermentation, identified, and characterized in their abilities to modulate immunity in porcine intestinal epithelial cells. Fresh soy milk and cow milk were inoculated with S. thermophilus SBC8781 (7 log CFU/mL) and incubated at 37 °C for 24 h. The extraction of EPSs was performed by the ethanol precipitation method. Analytical techniques, including NMR, UV-vis spectroscopy, and chromatography, identified and characterized both biopolymer samples as polysaccharides with high purity levels and similar Mw. EPS-s and EPS-m had heteropolysaccharide structures formed by galactose, glucose, rhamnose, ribose, and mannose, although with different monomer proportions. On the other hand, EPS-s had higher quantities of acidic polymer than EPS-m. The biopolymer production of the SBC8781 strain from the vegetable culture broth was 200–240 mg/L, which was higher than that produced in milk, which reached concentrations of 50–70 mg/L. For immunomodulatory assays, intestinal epithelial cells were stimulated with 100 µg/mL of EPS-s or EPS-m for 48 h and then stimulated with the Toll-like receptor 3 agonist poly(I:C). EPS-s significantly reduced the expression of IL-6, IFN-β, IL-8, and MCP-1 and increased the negative regulator A20 in intestinal epithelial cells. Similarly, EPS-m induced a significant reduction of IL-6 and IL-8 expressions, but its effect was less remarkable than that caused by EPS-s. Results indicate that the structure and the immunomodulatory activity of EPSs produced by the SBC8781 strain vary according to the fermentation substrate. Soy milk fermented with S. thermophilus SBC8781 could be a new immunomodulatory functional food, which should be further evaluated in preclinical trials.

A critical factor for preventing osteoporosis after menopause is attenuation of the accelerated turnover rate of bone metabolism. The present randomized controlled study was conducted to clarify the effects of a lemon beverage with calcium (Ca) supplementation that makes use of the chelating action of citric acid. Comprehensive evaluations of bone were performed by assessments of bone mineral density (BMD) and biomarkers related to bone turnover. Seventy-nine postmenopausal women were enrolled and asked to participate in an 11-month continuous intake of the test beverages. The subjects were divided into three groups: those who consumed a lemon beverage containing citric acid with Ca supplementation (LECA group), those who consumed a lemon beverage containing citric acid without Ca supplementation (LE group), and those who consumed no test beverage (control group). Using a double-blind protocol, subjects in the LECA and LE groups consumed one bottle containing 290 mL of the test beverage each day. The ratio of change in BMD after 11 months was significantly higher in the LECA group as compared to the control and LE groups. The LECA group also showed significant decreases in concentrations of tartrate-resistant acid phosphatase 5b (TRACP-5b), a bone resorption marker, and bone alkaline phosphatase (BAP) as compared to the other groups, as well as a significant decrease in concentration of osteocalcin (OC), a bone formation marker, as compared to the LE group. Based on our findings, we speculated that bone resorption and bone formation in postmenopausal women might be suppressed along with an increase in Ca resorption caused by chelation of citric acid in association with continuous ingestion of a Ca-supplemented lemon beverage containing citric acid, resulting in suppression of high bone metabolic turnover. In addition, the results provide information regarding BMD maintenance in the bones of the trunk, including the lumbar spine and proximal femur.

Background Key approaches to preventing the onset of osteoporosis are to maximize peak bone mass (PBM) in early adulthood and to minimize the inevitable loss of bone mineral density (BMD) after menopause. Consequently, osteoporosis attracts a great deal of social as well as medical attention in Japan, a country experiencing population aging. Methods The present randomized, controlled study aimed to elucidate how bone metabolism and bone mineral density (BMD) are affected by the consumption of a lemon juice-containing, calcium-enriched beverage. The effectiveness of this investigational product was evaluated in 79 postmenopausal women. The subjects were assigned to three groups. The first two groups received calcium-supplemented (LECA group) and unsupplemented lemon (LE group) beverages, respectively. The third group (control) received no intervention. The LECA and LE groups consumed one bottle (290 ml) of the appropriate investigational product every day for 11 consecutive months. Results The results showed that the regular consumption of this lemon beverage promoted Ca absorption through the chelating function of citric acid, attenuating bone resorption. The results also suggest that the suppression of bone resorption inhibits bone formation mediated by the differentiation and activity of osteoblasts and leads to the prevention of high-turnover bone metabolism. Conclusion This hypothesis is further supported by the novel finding that the regular consumption of the lemon beverage increases the BMD of both the lumbar spine and femur. We believe that the Ca-supplemented lemon beverage used in the present study will be instrumental in developing effective strategies for the prevention of osteoporosis.

The aim of the present study was to elucidate how bone metabolism and bone mineral density are affected by the consumption of a lemon juice-containing calcium (Ca)-enriched beverage. The efficacy of this investigational product was evaluated in postmenopausal women during five months of continuous intake (intervention). This was a randomized, controlled trial. Eighty-three subjects were assigned to three groups. Using a double-blind format, the first two groups received a Ca-supplemented lemon-juice (lemon) beverage (LECA) or a Ca-unsupplemented lemon-juice (lemon) beverage (LE). The third group (control) received no intervention. Each subject in the LECA and LE groups consumed one bottle (290 mL) of their assigned investigational product every day for five consecutive months. After five months of intervention, the gain in bone mineral density at the lumbar spine was significantly larger in the LECA and LE groups than in the control group. In the femur, subjects in the LECA group gained significantly more bone mineral density than the control subjects. The largest gain in bone mineral density at the lumbar spine was observed in the LECA group. As for the concentrations of the bone resorption marker tartrate-resistant acid phosphatase 5b (TRACP-5b), subjects in the LECA group had significantly lower values than those in the control group. Similarly, when compared with the LE and control groups, a significant decrease was detected in the LECA group in the concentrations of the bone formation markers, bone alkaline phosphatase (BAP) and osteocalcin (OC). In postmenopausal women, continuous consumption of Ca-supplemented lemon beverages improved the absorption of Ca and inhibited bone resorption. This likely blocked the function of osteoblasts and led to the suppression of bone formation, resulting in the attenuation of high-turnover bone metabolism.

Polyphenols have been examined for their beneficial effects on health, particularly in rodents, but their lifelong effects are unclear. Lemons (Citrus limon), containing lemon polyphenols (LPP), are widely consumed but the effects of LPP on aging are unknown. Therefore, we examined the effects of LPP on aging such as aging-related scores, locomotor activity, cognitive functions, and intestinal microbiome using senescence-accelerated mouse prone 1 (SAMP1) and senescence-accelerated resistant mouse 1 (SAMR1). All mice had ad libitum access to water (P1_water group, SAMR1) or 0.1% LPP (P1_LPP group). In the P1_LPP group, LPP intake prolonged the lifespan by approximately 3 weeks and delayed increases in aging-related scores (e.g., periophthalmic lesions) and locomotor atrophy. The P1_water group showed large changes in the intestinal microbiome structure, while the R1 and P1_LPP groups did not. The phylum Bacteroidetes/Firmicutes, which is associated with obesity, in the P1_water group was significantly lower and higher than that in the P1_LPP and R1 groups, respectively. Although the relative abundance of Lactobacillus significantly increased in both P1 groups with aging, the P1_LPP group showed a significantly lower increase than the P1_water group. Thus, lifelong intake of LPP may have anti-aging effects on both phenotypes and the intestinal environment.

Coffee consumption reduces the risk of type 2 diabetes in humans, but the mechanism remains unclear. In this study, we investigated the effect of coffee on pancreatic β-cells in the induction of diabetes by streptozotocin (STZ) treatment in mice. We examined the effect of coffee, caffeine, or decaffeinated coffee ingestion on STZ-induced hyperglycemia. After STZ injection in Exp. 1 and 2, serum glucose concentration and water intake in coffee ingestion (Coffee group) tended to be lowered or was significantly lowered compared to those in water ingestion (Water group) instead of coffee. In Exp. 1, the values for water intake and serum glucose concentration in caffeine ingestion (Caffeine group) were similar to those in the Water group. In Exp. 2, serum glucose concentrations in the decaffeinated coffee ingestion (Decaf group) tended to be lower than those in the Water group. Pancreatic insulin contents tended to be higher in the Coffee and Decaf groups than in the Water group (Exp. 1 and 2). In Exp. 3, subsequently, we showed that coffee ingestion also suppressed the deterioration of hyperglycemia in diabetic mice which had been already injected with STZ. This study showed that coffee ingestion prevented the development of STZ-induced diabetes and suppressed hyperglycemia in STZ-diabetic mice. Caffeine or decaffeinated coffee ingestion did not significantly suppress STZ-induced hyperglycemia. These results suggest that the combination of caffeine and other components of decaffeinated coffee are needed for the preventive effect on pancreatic β-cell destruction. Coffee ingestion may contribute to the maintenance of pancreatic insulin contents.

Objectives We examined the factors considered to change body composition and blood biochemistry indicators in menopausal middle-aged and elderly women. These changes result from exercise by walking as part of their daily activities and lemon consumption by women who live on the small islands of the Seto Inland Sea, Japan's largest citrus fruit (lemon)-producing region. Methods Between September 2011 and March 2012, we recorded the daily lemon consumption and the number of steps taken by 101 middle-aged and elderly female lemon farmers. We also measured their body dimensions, body compositions, and blood pressure pulse wave velocity and conducted blood tests before and after the survey period. The results before and after the survey period were compared by the t-test and associations were determined on the basis of Pearson's correlation coefficient. Covariance structural analysis was carried out to determine causal associations. Results From the results of covariance structure analysis, lemon intake did not have a direct impact on each item examined. The third item, i.e., "the factors related to arteriosclerosis," was affected indirectly via citric acid and fatigue, and anticoagulation was shown. The fourth item, i.e., "the factors related to maintenance of muscle mass," which is affected by menopausal years and the change in walking speed, was shown to be associated with the second item, i.e., "the factors related to lipid metabolism." Menopausal years affected the first, third and fourth items. Conclusions Lemon intake did not have a direct impact on each item. Lemon has been shown to indirectly affect the third item through citric acid. Walking affected the second item, the level of total cholesterol, such as HDL cholesterol, through the fourth item. The importance of providing services that lead to sustained physical activity and a well-balanced metabolism between lipids and carbohydrates has been shown.

Coffee has an anti-diabetic effect, specifically the amelioration of both hyperglycemia and insulin resistance, in KK-A(y) mice, a type 2 diabetes animal model. To investigate coffee's effect on insulin signaling in liver, skeletal muscle, and adipose tissue (epididymal fat), we assayed the tyrosine phosphorylation of insulin receptor (IR) and serine phosphorylation of Akt. In Expt. 1, we assayed insulin signaling under nonfasting conditions in KK-A(y) mice that ingested water or coffee for 4 wk. Coffee ingestion ameliorated the development of hyperglycemia but did not affect insulin signaling in liver or skeletal muscle under such conditions. In Expt. 2, we assayed insulin signaling under basal and insulin-stimulated conditions in KK-A(y) mice that ingested water or coffee for 3 wk. The levels of tyrosine phosphorylation of insulin receptor in response to insulin injection in insulin-sensitive tissues were not different between mice that drank water and those that drank coffee. Coffee ingestion significantly increased the insulin-induced serine phosphorylation of Akt in liver and skeletal muscle, but not in epididymal fat, of KK-A(y) mice. Our results also indicated that coffee ingestion may contribute to the improvement of insulin resistance and hyperglycemia in KK-A(y) mice via the activation of Akt in insulin signaling in liver and skeletal muscle.

We have previously demonstrated that coffee and caffeine ameliorated hyperglycemia in spontaneously diabetic KK-A(y) mice. This present study evaluates the antidiabetic effects of coffee and caffeine on high-fat-diet-induced impaired glucose tolerance in C57BL/6J mice. C57BL/6J mice fed a high-fat diet were given regular drinking water (control group), or a 2.5-fold-diluted coffee or caffeine solution (200 mg/L) for 17 weeks. The ingestion of coffee or caffeine improved glucose tolerance, insulin sensitivity, and hyperinsulinemia when compared with mice in the control group. The adipose tissue mRNA levels of inflammatory adipocytokines (MCP-1 and IL-6) and the liver mRNA levels of genes related to fatty acid synthesis were lower in the coffee and caffeine groups than those in the control group. These results suggest that coffee and caffeine exerted an ameliorative effect on high-fat-diet-induced impaired glucose tolerance by improving insulin sensitivity. This effect might be attributable in part to the reduction of inflammatory adipocytokine expression.

Lemon flavonoid (LF) prepared from lemon peel predominantly contains eriocitrin as an antioxidant. It is indicated to have low bioavailability compared with lemon flavonoid aglycone (LFA), which predominantly contains eriodictyol. This study attempted to prepare LFA which has high bioavailability, using enzymes that are commonly used in the citrus industry, such as cellulase, naringinase, hesperidinase, and pectinase. LFA containing the highest amount of eriodictyol (19.4%) was prepared with naringinase, a debittering enzyme for citrus juice. Ten male normolipidemic subjects ingested LFA (3.7 g) after an overnight fast, and low-density lipoprotein (LDL) was prepared from 0-4 h plasma after intake of LFA. The LDL oxidizability was measured with lag time of the conjugated diene formation induced by an oxidative inducer. LDL in 0.5 h plasma after ingestion of LFA was shown to have a significantly longer lag time for oxidation than that before ingestion (P

The aim of this study was to investigate the effect of dietary lemon polyphenols on high-fat diet-induced obesity in mice, and on the regulation of the expression of the genes involved in lipid metabolism to elucidate the mechanisms. Mice were divided into three groups and fed either a low fat diet (LF) or a high fat diet (HF) or a high fat diet supplemented with 0.5% w/w lemon polyphenols (LP) extracted from lemon peel for 12 weeks. Body weight gain, fat pad accumulation, the development of hyperlipidemia, hyperglycemia, and insulin resistance were significantly suppressed by lemon polyphenols. Supplementation with lemon polyphenols also significantly up-regulated the mRNA level of the peroxisome proliferator activated receptor-alpha (PPARalpha) compared to the LF and HF groups in the liver. Furthermore, the mRNA level of acyl-CoA oxidase (ACO) was up-regulated in the LP group compared to the LF group, but not HF group in the liver, and was also significantly increased in the epididymal white adipose tissue. Thus, feeding with lemon polyphenols suppressed body weight gain and body fat accumulation by increasing peroxisomal beta-oxidation through up-regulation of the mRNA level of ACO in the liver and white adipose tissue, which was likely mediated via up-regulation of the mRNA levels of PPARalpha.

Eriocitrin (eriodictyol 7-O-β-rutinoside) is the main flavonoid in lemon fruit. In this study, eriocitrin was investigated for its lowering effect on serum and hepatic lipids in high-fat and high-cholesterol fed rats. Rats in the control group (N= 6) were fed a 20% lard and 1% cholesterol diet for 21 d, and rats in the 0.35% eriocitrin group (N= 6) and 0.70% eriocitrin group (N= 6) were fed a diet supplemented with eriocitrin 0.35% and 0.70%, respectively. The content of hepatic total cholesterol and triglyceride in the eriocitrin group was no different from that of the control group. The total cholesterol, VLDL+LDL, triglyceride, and phospholipid in the serum of the 0.35% eriocitrin group showed significantly lower concentrations than the control group (P < 0.05), although there was no difference in the HDL concentrations among the groups. The lowering effect of eriocitrin for serum total cholesterol was thought to be caused by a decrease in VLDL+LDL. The 0.35% eriocitrin group was shown to have a significant increase in excretion of fecal bile acid (P < 0.05) and a tendency for enhanced hepatic m-RNA levels of LDL receptor in comparison with the control group.

Lemon (Citrus limon) contains various bioactive flavonoids, and prevents obesity and obesity-associated metabolic diseases. We focused on eriocitrin (eriodictyol 7-rutinoside), a powerful antioxidative flavonoid in lemon with lipid-lowering effects in a rat model of high-fat diet. To investigate the mechanism of action of eriocitrin, we conducted feeding experiments on zebrafish with diet-induced obesity. Oral administration of eriocitrin (32 mg/kg/day for 28 days) improved dyslipidaemia and decreased lipid droplets in the liver. DNA microarray analysis revealed that eriocitrin increased mRNA of mitochondrial biogenesis genes, such as mitochondria transcription factor, nuclear respiratory factor 1, cytochrome c oxidase subunit 4, and ATP synthase. In HepG2 cells, eriocitrin also induced the corresponding orthologues, and reduced lipid accumulation under conditions of lipid loading. Eriocitrin increased mitochondrial size and mtDNA content, which resulted in ATP production in HepG2 cells and zebrafish. In summary, dietary eriocitrin ameliorates diet-induced hepatic steatosis with activation of mitochondrial biogenesis.

Background. Recent studies suggest that the daily intake of lemon (Citrus limon) has a good effect on health, but this has not been confirmed in humans. In our previous studies, it was observed that people who are conscious of their health performed more lemon intake and exercise. An analysis that took this into account was required.Methodology. For 101 middle-aged women in an island area in Hiroshima, Japan, a record of lemon ingestion efforts and the number of steps walked was carried out for five months. The change rates (Δ%) of the physical measurements, blood test, blood pressure, and pulse wave measured value during the observation period were calculated, and correlations with lemon intake and the number of steps walked were considered. As a result, it was suggested that daily lemon intake and walking are effective for high blood pressure because both showed significant negative correlation to systolic blood pressure Δ%.Conclusions. As a result of multiple linear regression analysis, it was possible that lemon ingestion is involved more greatly with the blood citric acid concentration Δ% and the number of steps with blood pressure Δ%, and it was surmised that the number of steps and lemon ingestion are related to blood pressure improvement by different action mechanisms.

Phenolic glucosides having radical scavenging activity were examined from the fraction eluted with 20% methanol on Amberlite XAD-2 resin applied to lemon (Citrus limon) juice by using reversed phase chromatography. Four phenolic glucosides were identified as 1-feruloyl-beta-D-glucopyranoside, 1-sinapoyl-beta-D-glucopyranoside, 6,8-di-C-glucosylapigenin and 6,8-di-C-glucosyldiosmetin by (1)H-NMR, (13)C-NMR, and MS analyses. They exhibited radical scavenging activity for 1,1-diphenyl-2-picrylhydrazyl (DPPH) and superoxide, although the activity was low in comparison with eriocitrin, a potent antioxidant in lemon fruit, and the eriodictyol of its aglycone. The phenolic compounds in lemon juice were examined for their suppressive effect on the expression of blood adhesion molecules by measuring the expression of intercellular adhesion molecule-1 (ICAM-1) in human umbilical vein endothelial cells (HUVECs) induced by necrosis factor-alpha (TNF-alpha). 6,8-Di-C-glucosylapigenin, apigenin, and diosmentin of the flavones were found to significantly suppress the expression of ICAM-1 at 10 muM (P0.05). The phenolic glucosides isolated in this study were contained in comparative abundance in daidai (Citrus aurantium) and niihime (Citrus unshiu x Citrus tachibana) among the sour citrus juices.

application/pdf, 学術雑誌論文, The concentrations of metabolites in human plasma after ingestion of flavanone glycosides (FG) and their aglycones (FA) in lemon were examined. FG consisting abundantly of eriocitrin were prepared from lemon peel and FA consisting abundantly of eriodictyol were prepared from FG by treatment with β-glucosidase. Eriodictyol, homoeriodictyol, and hesperetin in plasma up to 4 h after ingestion of FG with water or FA with water by subjects were not detected in plasma of non-enzyme treatment but in plasma after treatment with β-glucronidase and sulfatase. Metabolites in plasma after ingestion of FG and FA in humans were shown to exist as the glucuro- and/or sulfo-conjugates of eriodictyol, homoeriodictyol, and hesperetin. After ingestion of FA, the concentration of metabolites in plasma exhibited a high maximum peak at 1 h. The AUC (area under the blood concentration time curve) level of metabolites of FA was higher than that of FG. FA were suggested to be absorbed faster and in higher amounts than FG. The AUC of metabolites in subject plasma after ingestion of FG with flavonoid-depleted lemon juice was shown to change\ to a low level in comparison with that of FG with water. The maximum concentration peak of metabolites in plasma was faster at 0.5 h than FA with water but the AUC level was similar to FA with water, when subjects ingested FA with vodka (40% ethanol). The absorption hour of FG and FA was shown to be affected by the co-existing solution.

The effects of gallic acid and its esters on H2O2-induced cytotoxicity, mutagenicity and SOS response were investigated in bacterial assay systems, i.e., the Ames test with Salmonella typhimurium TA104 and the SOS chromotest with E. coli PQ37. In the Ames test, gallic acid esters showed protective effects against H2O2-induced cytotoxicity and no effects on the number of revertant colonies. In the SOS chromotest, gallic acid esters lowered the SOS induction factor raised by H2O2. Throughout the study, the effects of gallic acid itself were weak or negligible, and lauryl gallate was most effective among the three gallic acid esters. This structure-activity relationship indicates the similarity of the protective effects of gallic acid esters on the H2O2-induced damages to both bacterial and mammalian cells.