LC-QTOF/MS-Based Profiling of the Phytochemicals in Ice Plant (Mesembryanthemum crystallinum) and Their Bioactivities.

Recent assessments of the correlations between food and medicine underscore the importance of functional foods in disease prevention and management. Functional foods offer health benefits beyond basic nutrition, with fresh fruits and vegetables being particularly prominent because of their rich polyphenol content. In this study, we elucidated the phytochemicals in ice plant (Mesembryanthemum crystallinum), a globally consumed vegetable, using an LC-QTOF/MS-based untargeted detection method. The phytochemicals were clustered based on their structural similarity using molecular networking and annotated using the in silico tool for network annotation propagation. To identify the bioactive compounds, eight compounds were isolated from ice plant extracts. These compounds were identified using extensive spectroscopic methods, including 1H and 13C nuclear magnetic resonance (NMR) spectroscopy. Additionally, we evaluated the antioxidant and anti-inflammatory activities of all the isolates. Among the tested compounds, three showed antioxidant activity and all eight showed anti-inflammatory activity, demonstrating the potential of ice plant as a functional food.

Anti-Metastatic Effects of Standardized Polysaccharide Fraction from Diospyros kaki Leaves via GSK3ß/ß-Catenin and JNK Inactivation in Human Colon Cancer Cells.

A polysaccharide fraction from Diospyros kaki (PLE0) leaves was previously reported to possess immunostimulatory, anti-osteoporotic, and TGF-ß1-induced epithelial-mesenchymal transition inhibitory activities. Although a few beneficial effects against colon cancer metastasis have been reported, we aimed to investigate the anti-metastatic activity of PLE0 and its underlying molecular mechanisms in HT-29 and HCT-116 human colon cancer cells. We conducted a wound-healing assay, invasion assay, qRT-PCR analysis, western blot analysis, gelatin zymography, luciferase assay, and small interfering RNA gene silencing in colon cancer cells. PLE0 concentration-dependently inhibited metastasis by suppressing cell migration and invasion. The suppression of N-cadherin and vimentin expression as well as upregulation of E-cadherin through the reduction of p-GSK3ß and ß-catenin levels resulted in the outcome of this effect. PLE0 also suppressed the expression and enzymatic activity of matrix metalloproteinases (MMP)-2 and MMP-9, while simultaneously increasing the protein and mRNA levels of the tissue inhibitor of metalloproteinases (TIMP-1). Furthermore, signaling data disclosed that PLE0 suppressed the transcriptional activity and phosphorylation of p65 (a subunit of NF-κB), as well as the phosphorylation of c-Jun and c-Fos (subunits of AP-1) pathway. PLE0 markedly suppressed JNK phosphorylation, and JNK knockdown significantly restored PLE0-regulated MMP-2/-9 and TIMP-1 expression. Collectively, our data indicate that PLE0 exerts an anti-metastatic effect in human colon cancer cells by inhibiting epithelial-mesenchymal transition and MMP-2/9 via downregulation of GSK3ß/ß-catenin and JNK signaling.

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution.

Efficient utilization of galactose by microorganisms can lead to the production of valuable bio-products and improved metabolic processes. While Bacillus subtilis has inherent pathways for galactose metabolism, there is potential for enhancement via evolutionary strategies. This study aimed to boost galactose utilization in B. subtilis using adaptive laboratory evolution (ALE) and to elucidate the genetic and metabolic changes underlying the observed enhancements. The strains of B. subtilis underwent multiple rounds of adaptive laboratory evolution (approximately 5000 generations) in an environment that favored the use of galactose. This process resulted in an enhanced specific growth rate of 0.319 ± 0.005 h-1, a significant increase from the 0.03 ± 0.008 h-1 observed in the wild-type strains. Upon selecting the evolved strain BSGA14, a comprehensive whole-genome sequencing revealed the presence of 63 single nucleotide polymorphisms (SNPs). Two of them, located in the coding sequences of the genes araR and glcR, were found to be the advantageous mutations after reverse engineering. The strain with these two accumulated mutations, BSGALE4, exhibited similar specific growth rate on galactose to the evolved strain BSGA14 (0.296 ± 0.01 h-1). Furthermore, evolved strain showed higher productivity of protease and ß-galactosidase in mock soybean biomass medium. ALE proved to be a potent tool for enhancing galactose metabolism in B. subtilis. The findings offer valuable insights into the potential of evolutionary strategies in microbial engineering and pave the way for industrial applications harnessing enhanced galactose conversion.

Anti-Colitic Effect of an Exopolysaccharide Fraction from Pediococcus pentosaceus KFT-18 on Dextran Sulfate Sodium-Induced Colitis through Suppression of Inflammatory Mediators.

We previously reported the immunostimulatory effect of an exopolysaccharide fraction from Pediococcus pentosaceus KFT18 (PE-EPS), a lactic acid bacterium, in macrophages and primary splenocytes, as well as in cyclophosphamide-induced immunosuppressed mice. In this study, the anti-colitic activity of PE-EPS was investigated in a dextran sulfate sodium (DSS)-induced colitis animal model. PE-EPS relieved DSS-induced colitis symptoms, such as stool blood, decreased colon length, crypt disruption, and mucus layer edema. Regarding the molecular mechanism, PE-EPS reduced the enhanced expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and pro-inflammatory cytokines (TNF-α, IL-6, and IL-1) in the colon tissue of colitis-induced mice. Additionally, PE-EPS protected against DSS-induced phosphorylation of p65 and signal transducer and activator of transcription 1 (STAT1). These findings suggested that the exopolysaccharide fraction from Ped. pentosaceus KFT18 can be used to treat inflammatory bowel disease by alleviating colonic inflammation.

Ginsenoside Rf Enhances Exercise Endurance by Stimulating Myoblast Differentiation and Mitochondrial Biogenesis in C2C12 Myotubes and ICR Mice.

Ginsenoside Rf (G-Rf) is a saponin of the protopanaxatriol family and a bioactive component of Korean ginseng. Several ginsenosides are known to have a positive effect on exercise endurance, but there is not yet a report on that of G-Rf. Forced swimming tests were performed on G-Rf-treated mice to evaluate the effect of G-Rf on exercise endurance. Subsequently, the expression of markers related to myoblast differentiation and mitochondrial biogenesis in murine skeletal C2C12 myotubes and tibialis anterior muscle tissue was determined using Western blotting, quantitative real-time polymerase chain reaction, and immunofluorescence staining to elucidate the mechanism of action of G-Rf. The swimming duration of the experimental animal was increased by oral gavage administration of G-Rf. Moreover, G-Rf significantly upregulated the myoblast differentiation markers, mitochondrial biogenesis markers, and its upstream regulators. In particular, the mitochondrial biogenesis marker increased by G-Rf was decreased by each inhibitor of the upstream regulators. G-Rf enhances exercise endurance in mice, which may be mediated by myoblast differentiation and enhanced mitochondrial biogenesis through AMPK and p38 MAPK signaling pathways, suggesting that it increases energy production to satisfy additional needs of exercising muscle cells. Therefore, G-Rf is an active ingredient in Korean ginseng responsible for improving exercise performance.

Strain-Specific Identification and In Vivo Immunomodulatory Activity of Heat-Killed Latilactobacillus sakei K040706.

We previously reported that the immunostimulatory activity of heat-killed Latilactobacillus sakei K040706 in macrophages and cyclophosphamide (CTX)-treated mice. However, identification of heat-killed L. sakei K040706 (heat-killed LS06) using a validated method is not yet reported. Further, the underlying molecular mechanisms for its immunostimulatory effects in CTX-induced immunosuppressed mice remain unknown. In this study, we developed strain-specific genetic markers to detect heat-killed L. sakei LS06. The lower detection limit of the validated primer set was 2.1 × 105 colony forming units (CFU)/mL for the heat-killed LS06 assay. Moreover, oral administration of heat-killed LS06 (108 or 109 CFU/day, p.o.) effectively improved the body loss, thymus index, natural killer cell activity, granzyme B production, and T and B cell proliferation in CTX-treated mice. In addition, heat-killed LS06 enhanced CTX-reduced immune-related cytokine (interferon-γ, interleukin (IL)-2, and IL-12) production and mRNA expression. Heat-killed LS06 also recovered CTX-altered microbiota composition, including the phylum levels of Bacteroidetes, Firmicutes, and Proteobacteria and the family levels of Muribaculaceae, Prevotellaceae, Tannerellaceae, Christensenellaceae, Gracilibacteraceae, and Hungateiclostridiaceae. In conclusion, since heat-killed L. sakei K040706 ameliorated CTX-induced immunosuppression and modulated gut microbiota composition, they have the potential to be used in functional foods for immune regulation.

Protective effect of exopolysaccharide fraction from Bacillus subtilis against dextran sulfate sodium-induced colitis through maintenance of intestinal barrier and suppression of inflammatory responses.

We previously reported that an exopolysaccharide-enriched fraction from Bacillus subtilis J92 (B-EPS) could improve immune functions by regulating the immunological parameters of IFN-γ-primed macrophages, CD3/CD28-stimulated splenocytes, and in cyclophosphamide-induced immunosuppressed mice. In the present study, we investigated whether B-EPS contributes to the maintenance of intestinal barrier integrity in a dextran sodium sulfate (DSS)-induced colitis mouse model that mimics human inflammatory bowel disease (IBD). B-EPS treatment improved histological characteristics and common features including a high disease activity index (DAI), an increased spleen weight, and colon shortening in DSS-induced colitis. B-EPS also effectively restored intestinal barrier function by modulating tight junction-related proteins (claudin-1, claudin-2, and occludin) and epithelial-mesenchymal transition (EMT) marker proteins (E-cadherin, N-cadherin, and vimentin). Moreover, B-EPS downregulated immune cell infiltration and inflammatory responses including the production of inflammatory cytokines, such as IL-6 and IL-1ß, and activation of nuclear factor-kappa B (NF-κB) and signal transducer and activator of transcription 3 (STAT3). Taken together, these results suggest that B-EPS could serve as a functional food ingredient for improving intestinal barrier function and alleviating colonic inflammation in IBD.

Ganoderma lucidum Rhodiola compound preparation prevent D-galactose-induced immune impairment and oxidative stress in aging rat model.

Aging is an irreversible process. This research aims to study the anti-aging effects of GRCP, a compound preparation made by Ganoderma lucidum and Rhodiola rosen, in aging rats. Rats were subcutaneously injected with 400 mg/kg of D-galactose daily, and aging could be induced after 8 weeks. The aging rats were treated with GRCP. This experiment was divided into 6 groups. Rats were randomly divided into the model group, positive control group, low-dose GRCP group (25 mg/kg body weight), medium-dose GRCP group (50 mg/kg body weight), and high-dose GRCP group (100 mg/kg body weight), healthy and normal rats were used as blank controls. After the end, the results show that the use of GRCP at a dose of 100 mg/kg is the best treatment for improving aging rats. Rats gained weight, spleen and thymus indexes, and splenocyte proliferation improved, and inflammatory cytokine levels decreased. Besides, biochemical indicators show that GRCP can improve the antioxidant enzyme activity and reduce the content of lipofuscin and TGF-ß in aging rats (P < 0.05). GRCP can also inhibit the activation of the MyD88/NF-κB pathway in rat hippocampus. These results seem to suggest that GRCP can be used as a potential natural supplement or functional food to prevent aging.

Selective activation of the estrogen receptor-ß by the polysaccharide from Cynanchum wilfordii alleviates menopausal syndrome in ovariectomized mice.

The menopausal syndrome caused by rapid changes in hormone levels greatly influences the quality of life of women. Though hormone replacement therapy (HRT) is widely used to treat the menopausal syndrome, it exhibits many side effects, including the risk of thrombosis, cardiovascular diseases, and increased incidence of breast cancer; thus, diversifying the interest for phytotherapy-based materials as alternatives to HRT. Here, we isolated a crude polysaccharide fraction (CWPF) from Cynanchum wilfordii root that alleviated the ovariectomy-induced uterine atrophy and bone loss without changes in plasma estradiol concentration in mice. Increased plasma levels of follicle-stimulating hormone (FSH), alkaline phosphatase (ALP), osteocalcin (OC) in ovariectomized mice were also reduced to normal levels by CWPF administration. We found that the inhibitory effects of CWPF on menopausal symptoms were mediated by the estrogen receptor ß (ER-ß) specific activation, not ER-α. Moreover, CWPF treatment suppressed the phosphorylation of Akt, suggesting that CWPF alleviates post-menopausal symptoms by regulating ER-ß related Akt signaling pathway. These results demonstrate that the polysaccharides corresponding to CWPF among the water-soluble extracts of CW could be used as a beneficial herbal alternative for the development of therapeutic agents to prevent menopausal syndrome in women.

Polysaccharide isolated from persimmon leaves (Diospyros kaki Thunb.) suppresses TGF-ß1-induced epithelial-to-mesenchymal transition in A549 cells.

In the present study, to verify the effect of polysaccharides derived from persimmon leaves (PLE) at epithelial-to-mesenchymal transition (EMT), A549 cells were treated with TGF-ß1 alone or co-treated with TGF-ß1 and PLE (50 and 75 µg/mL). PLE-treated cells showed higher expression of E-cadherin and lower expression of N-cadherin and vimentin compared to TGF-ß1-treated cells by inhibiting the levels of transcription factors, including Snail, Slug, and ZEB1, all associated with EMT. PLE also significantly decreased migration, invasion, and anoikis resistance through TGF-ß1 mediated EMT suppression, whereby PLE inhibited the levels of MMP-2 and MMP-9 while cleaving PARP. These inhibitory effects of PLE against EMT, migration, invasion, and anoikis resistance were determined by activating the canonical SMAD2/3 and non-canonical ERK/p38 signaling pathways. Therefore, these results suggest that PLE could be used as a potential chemical therapeutic agent for early metastasis of lung cancer in vitro.

Ginseng-Induced Changes to Blood Vessel Dilation and the Metabolome of Rats.

Ginseng consumption has been shown to prevent and reduce many health risks, including cardiovascular disease. However, the ginseng-induced changes in biofluids and tissue metabolomes associated with blood health remain poorly understood. In this study, healthy rats were orally administered ginseng extracts or water for one month. Biofluid and tissue metabolites along with steroid hormones, plasma cytokines, and blood pressure factors were determined to elucidate the relationship between ginseng intake and blood vessel health. Moreover, the effect of ginseng extract on blood vessel tension was measured from the thoracic aorta. Ginseng intake decreased the levels of blood phospholipids, lysophosphatidylcholines and related enzymes, high blood pressure factors, and cytokines, and induced vasodilation. Moreover, ginseng intake decreased the level of renal oxidized glutathione. Overall, our findings suggest that ginseng intake can improve blood vessel health via modulation of vasodilation, oxidation stress, and pro-inflammatory cytokines. Moreover, the decrease in renal oxidized glutathione indicated that ginseng intake is positively related with the reduction in oxidative stress-induced renal dysfunction.

Acute Oral Toxicity and Genotoxicity of Polysaccharide Fraction from Young Barley Leaves (Hordeum vulgare L.).

Polysaccharides isolated from various plants are considered precious bioactive materials owing to their potent biological activities. Previously, we prepared a polysaccharide fraction (BLE0) isolated from young barley leaves (Hordeum vulgare L.), demonstrating its anti-osteoporotic and immunostimulatory activities. However, data regarding BLE0 toxicity is lacking. To establish its safety, in vitro genotoxicity (chromosomal aberration and bacterial reverse mutation assays) and acute oral toxicity assays were conducted. In the in vitro genotoxicity assays, bacterial reverse mutation and chromosomal aberration assays showed that BLE0 possessed no mutagenicity or clastogenicity. Furthermore, the median lethal dose (LD50) of BLE0 was higher than 5000 mg/kg in female and male Sprague-Dawley (SD) rats and no adverse effects were observed in terms of mortality and abnormal changes in clinical signs (body weight and necropsy). Based on these results, BLE0 was found to be safe with regards to genotoxicity under our test conditions, demonstrating no acute oral toxicity up to 5000 mg/kg in SD rats.

Polysaccharides isolated from lotus leaves (LLEP) exert anti-osteoporotic effects by inhibiting osteoclastogenesis.

Nelumbo nucifera, more commonly known as the Indian lotus, is an important plant that has been incorporated into traditional herbal remedies along the years. Even today, lotus leaves are considered reservoirs for bioactive compounds that can be used as nutritional supplements to treat various human diseases. However, despite the wide ranging biological activities of lotus polysaccharides, limited information is available regarding the anti-osteoporotic effects of these substances. The aim of this study was to investigate the beneficial effects of pectinase-assisted extractable polysaccharides from lotus leaves (LLEP) on estrogen deficiency-induced bone loss and osteoclast differentiation in bone marrow-derived macrophages. We found that LLEP markedly inhibited receptor activator of the nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation in a dose-dependent manner. It also revoked RANKL-induced activation of osteoclastogenic signals such as the expression of key transcription factors (i.e., c-Fos and nuclear factor of activated T cells cytoplasmic 1), resulting in a decrement in osteoclast-specific marker gene expressions. Microcomputed tomography and morphometric analysis revealed that a four-week oral administration of LLEP notably decreased trabecular bone loss. Taken together, our results suggest that LLEP can mitigate estrogen deficiency-induced bone loss by suppressing osteoclastogenesis, which makes it an excellent candidate for combating osteoporosis.

Effect of enzyme-assisted extraction on the physicochemical properties and bioactive potential of lotus leaf polysaccharides.

Lotus leaf polysaccharides were extracted by enzyme-assisted extraction using α-amylase (LLEP-A), cellulose (LLEP-C), pectinase (LLEP-P) or protease (LLEP-PR). Their physicochemical properties and immunostimulatory activities were compared with those of hot-water extracted polysaccharides (LLWP). HPAEC-PDA and HPSEC-RI profiles indicated that variations in their molecular weight patterns and chemical compositions. Moreover, their effects on proliferation, phagocytic activity, and cytokine production in macrophages could be ordered as LLEP-P > LLEP-C > LLEP-A > LLWP > LLEP-PR, suggesting that LLEP-P by pectinase-assisted extraction was the most potent enhancer of macrophage activation. LLEP-P was further purified by gel filtration, and the main fraction (LLEP-P-І) was obtained to elucidate the structural and functional properties. LLEP-P-І (14.63 × 103 g/mol) mainly consisted of rhamnose, arabinose, galactose, and galacturonic acid at molar percentages of 15.5:15.8:20.1:32.8. FT-IR spectra indicated the predominant acidic and esterified form, suggesting the pectic-like structure. Above all, LLEP-P-І exerted greater stimulation effects on NO and cytokines production and the phagocytic activity in macrophages. Transcriptome analysis also demonstrated that LLEP-P and LLEP-P-І could upregulate macrophage immune response genes, including cytokines, chemokines, and interferon via TLR and JAK-STAT signaling. Thus, these results suggest that pectinase application is most suitable to obtain immunostimulatory polysaccharides from lotus leaves.

Red Ginseng Improves Exercise Endurance by Promoting Mitochondrial Biogenesis and Myoblast Differentiation.

Red ginseng has been reported to elicit various therapeutic effects relevant to cancer, diabetes, neurodegenerative diseases, and inflammatory diseases. However, the effect of red ginseng on exercise endurance and skeletal muscle function remains unclear. Herein, we sought to investigate whether red ginseng could affect exercise endurance and examined its molecular mechanism. Mice were fed with red ginseng extract (RG) and undertook swimming exercises to determine the time to exhaustion. Animals fed with RG had significantly longer swimming endurance. RG treatment was also observed to enhance ATP production levels in myoblasts. RG increased mRNA expressions of mitochondrial biogenesis regulators, NRF-1, TFAM, and PGC-1α, which was accompanied by an elevation in mitochondrial DNA, suggesting an enhancement in mitochondrial energy-generating capacity. Importantly, RG treatment induced phosphorylation of p38 and AMPK and upregulated PGC1α expression in both myoblasts and in vivo muscle tissue. In addition, RG treatment also stimulated C2C12 myogenic differentiation. Our findings show that red ginseng improves exercise endurance, suggesting that it may have applications in supporting skeletal muscle function and exercise performance.

Immunostimulatory effects of polysaccharides isolated from young barley leaves (Hordeum vulgare L.) with dual activation of Th1 and Th2 in splenic T cells and cyclophosphamide-induced immunosuppressed mice.

Botanical polysaccharides have been widely known to possess immunological activity. The objective of this study was to investigate the molecular mechanisms underlying the immunostimulatory properties of polysaccharides isolated from barley leaf (Hordeum vulgare L.) (BLE0) in splenocytes and cyclophosphamide (CYP)-induced immunosuppressed mice. BLE0 showed cell proliferative activity and markedly increased the secretion of both Th1-cytokines (IFN-γ and IL-2) and Th2-cytokines (IL-4 and IL-10) in CD3/CD28-activated splenocytes. Molecular data revealed that BLE0 up-regulated the expression of T-bet with enhanced phosphorylation of Janus kinase (JAK)-signal transducer and activator of transcription (STAT) 1 signaling pathway. BLE0 also increase the phosphorylation of GATA3 via toll-like receptor (TLR) 2-mediated signaling pathway with nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein 1 (AP-1) activation. Oral administration of BLE0 effectively improved CYP-induced decrease of body weight, splenocyte proliferation, and natural killer (NK) cell cytotoxic activity and significantly increased Th1 and Th2 cytokines, T-bet, and GATA3 mRNA expression. Dietary intake of BLE0 improves the immunological manifestations by stimulating both Th1 and Th2 responses via JAK/STAT1/T-bet and TLR2/GATA3, respectively.

Isolation, purification, and characterization of novel polysaccharides from lotus (Nelumbo nucifera) leaves and their immunostimulatory effects.

In this study, a crude water-soluble polysaccharide (LLWP-C) was extracted from lotus leaves and further purified by size exclusion chromatography, to obtain the two main polysaccharides, LLWP-1 and LLWP-3. Physical and chemical analyses showed that they were homogeneous polysaccharides in ß-type glycosidic linkage. LLWP-1 was devoid of helical conformation, had a molecular weight of 85.1 kDa and was mainly composed of Rha, Ara, Gal, Glu, and GalA in a molar ratio of 7.0:24.8:28.0:6.0:26.4. LLWP-3 showed a helical conformation, had a molecular weight of 12.5 kDa and consisted mainly of Rha, Ara, Gal, Glu, Man, and GalA in a molar ratio of 6.6:9.8:15.0:8.9:6.1:47.2. It was demonstrated that LLWP-C and both purified LLWP-1 and LLWP-3 could effectively enhance the proliferation, phagocytosis, nitric oxide (NO), and cytokine secretions by activating corresponding mRNA expression in macrophages, via MAPK and NF-κB pathways. LLWP-3 displayed the greatest immunostimulatory potential, followed by LLWP-1 and LLWP-C. These findings suggest that polysaccharides extracted from lotus leaf exert immunostimulatory activity that could be further investigated to develop functional foods and natural immunopotentiating therapeutic agents.

Discrimination of Structural and Immunological Features of Polysaccharides from Persimmon Leaves at Different Maturity Stages.

In this study, we investigated changes in the structural and immunological features of polysaccharides (S1-PLE0, S2-PLE0, and S3-PLE0) extracted from persimmon leaves at three different growth stages. Physicochemical analyses revealed that their chemical compositions, molecular weight distributions, and linkage types differed. High-performance size-exclusion chromatograms showed that the molecular weights of the polysaccharides increased during successive growth stages. In addition, seasonal variation of persimmon leaves affected the sugar compositions and glycosidic linkages in the polysaccharides. S2-PLE0 was composed of comparatively more galactose, arabinose, rhamnose, xylose, and galacturonic acid, showing the presence of ß-glucopyranoside linkages. Significant differences also occurred in their immunostimulatory effects on RAW264.7 macrophages, with respect to which their activities could be ordered as S2-PLE0 > S3-PLE0 > S1-PLE0. Evidently, S2-PLE0 showed the greatest immunostimulatory activity by enhancing the phagocytic capacity and promoting nitric oxide (NO) and cytokines secretion through the upregulation of their gene expression in macrophages. These results suggest that differences in the structural features of polysaccharides according to the different maturity of persimmon leaves might impact their immunostimulatory properties. The results also provide a basis for optimizing persimmon leaf cultivation strategies for food and medical uses of the polysaccharides.

Protective effects of a polysaccharide BLE0 isolated from barley leaf on bone loss in ovariectomized mice.

Potent anti-osteoporotic drugs have been developed over the past decades; however, the substantial need for treatments that can effectively and safely manage osteoporosis remains unmet. Barley leaf-derived products are one of best functional foods that can be used as nutritional supplements and detoxifiers in humans and are beneficial in improving bone disease. However, little information is available regarding the anti-osteoporotic effects of polysaccharides as the main component of barley leaf. This study aimed to clarify the beneficial effects of barley leaf (BLE0) polysaccharides on bone loss in ovariectomized mice and osteoclast differentiation in bone marrow-derived macrophages. BLE0 remarkably inhibited receptor activator of the nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation in a dose-dependent manner. It also blocked RANKL-induced activation of osteoclastogenic signals including ERK and p38 and the expression of nuclear factor of activated T cells cytoplasmic 1, as a master regulator of osteoclast differentiation, leading to decreased expression of osteoclast-specific marker genes such as Atp6v0d2, DC-STAMP and cathepsin K. Micro-computed tomography revealed that a seven-week oral administration of BLE0 dramatically improved ovariectomy-induced trabecular bone loss. Anti-osteoporotic effects were confirmed using morphometric analysis. Taken together, BLE0 is a bioactive polysaccharide; it mitigates estrogen deficiency-induced bone loss by directly inhibiting osteoclast differentiation.

The Effect of Pectinase-Assisted Extraction on the Physicochemical and Biological Properties of Polysaccharides from Aster scaber.

The edible and medicinal perennial herb Aster scaber is known to have anticancer, antioxidant, and immunomodulatory properties. However, the biological effects of its polysaccharides are not well understood. Here, we aimed to extract novel polysaccharides with enhanced biological properties from Aster scaber using enzyme-assisted methods. Amylase, cellulase, and pectinase were used to extract enzyme-assisted polysaccharide (ASEP)-A, ASEP-C, and ASEP-P, respectively. The yields, physicochemical properties, and immunostimulatory activities of the polysaccharides were investigated and compared with those of hot water extracted polysaccharide (ASWP). The highest yield (3.8%) was achieved for ASEP-P extracted using pectinase digestion. Fourier-transform infrared spectroscopy (FT-IR) and chemical composition analysis revealed that ASWP and three ASEPs were typical acidic heteropolysaccharides, mainly comprising rhamnose, arabinose, galactose, glucose, and galacturonic acid. Immunostimulatory activity assays on RAW264.7 macrophages showed ASEP-P to have the greatest immunostimulatory potential in terms of nitric oxide (NO) and cytokine productions and phagocytic activity. ASEP-P administration improved immune-enhancing effects in normal mice by improving the spleen index and splenic lymphocyte proliferation, and in immunosuppressed mice by modulating lymphocyte proliferation, natural killer (NK) cell activity, and leukocyte counts. The ASEP-P derived from pectinase hydrolysate of Aster scaber demonstrated efficacious immunostimulatory properties and has potential applications as an immune stimulator.