The objective of this study was to investigate the anticancer effects of EMPS (edible mushroom mycelium polysaccharide: Tremella fuciformis) in animal models with colorectal cancer induced by AOM/DSS. The experimental groups consisted of Nor (normal), NC (AOM/DSS), EMPS (EMPS 50, EMPS 100), and PC (Fluorouracil). The NC group had the highest number of colon tumors, whereas it was observed that tumor occurrence was significantly reduced in the EMPS consumption group. The expression of Bcl-2, an apoptosis inhibitor, was significantly lower in the EMPS 50 & 100 and PC groups. On the other hand, the mRNA gene expression of Bax, a factor that induces apoptosis, was significantly higher in the EMPS 50 & 100 and PC groups compared to the NC group. The mRNA expression levels of TNF-α and COX-2 significantly increased in the NC group, but showed a significant decrease in the EMPS and PC groups, indicating inhibition of the cancer-promoting response of cells. At the phylum level of the mice's intestinal microbial composition, the proportion of Bacteroidetes tended to decrease, while the proportion of Firmicutes tended to increase with EMPS administration. This suggests that changes in the gut microbiota caused by inflammation can be influenced by dietary intake.

Lytic polysaccharide monooxygenases (LPMOs) catalyze the oxidative cleavage of glycosidic bonds in crystalline polysaccharides including chitin and cellulose. The recent discovery of LPMO family proteins in many insect species suggests that they presumably play a role in chitin degradation in the cuticle/exoskeleton, tracheae and peritrophic matrix during insect development. Insect LPMOs belong to auxiliary activity family 15 (AA15/LPMO15) and have been classified into at least four groups based on phylogenetic analysis. In this study, we identified and investigated the physiological functions of group I LPMO15 (MaLPMO15-1 and PhLPMO15-1) in two longhorn beetle species, Monochamus alternatus and Psacothea hilaris. In both species, depletion of LPMO15-1 transcripts by RNAi resulted in a lethal pupal-adult molting defect. The insects were unable to shed their old pupal cuticle and died entrapped in their exuviae. Furthermore, TEM analysis revealed a failure of degradation of the chitinous procuticle layer of their old cuticle, retaining intact horizontal laminae and vertical pore canals containing perpendicularly oriented chitin fibers (pore canal fiber, PCF) in their core. These results indicate that MaLPMO15-1 and PhLPMO15-1 are required for turnover of the chitinous old cuticle, which is critical for insect molting.

To increase industrial applicability of Astragalus membranaceus (AM) as immunostimulating materials, hot-water extract (AME) was prepared from AM and fermented with Kimchi-lactic acid bacteria (Lactobacillus sakei & Leuconostoc mesenteroides) to prepare fermented AM-postbiotics (FAME). Although FAME prepared from AM-postbiotics did not show a significant enhancement in macrophage stimulating activity compared to non-fermented AME, crude polysaccharide (FAME-CP) fractionated by EtOH precipitation from FAME showed significantly higher macrophage stimulating activity than AME-CP. Compared to AME-CP, FAME-CP showed dramatic changes in component sugar and molecular weight distribution. FAME-CP was a polysaccharide with a major molecular weight distribution of 113.4 kDa containing Man (44.2%), Glc (19.3%), Gal (10.2%), GalA (10.2%), and Ara (7.4%) as sugar components. FAME-CP with enhanced macrophage stimulatory activity not only increased expression levels of mRNA genes encoding macrophage-activated factors (iNOS, TNF-α, MCP-1, IL-6, and COX-2), but also led the nuclear translocation of activated p65 and c-Jun. In conclusion, crude polysaccharide from AM-postbiotics fermented with lactic acid bacteria could increase industrial applicability as a functional material with enhanced immunostimulating activity than AME-CP.

Lytic polysaccharide monooxygenases (LPMOs) catalyze the oxidative cleavage of glycosidic bonds in crystalline polysaccharides including chitin and cellulose. The recent discovery of LPMO family proteins in many insect species suggests that they presumably play a role in chitin degradation in the cuticle/exoskeleton, tracheae and peritrophic matrix during insect development. Insect LPMOs belong to auxiliary activity family 15 (AA15/LPMO15) and have been classified into at least four groups based on phylogenetic analysis. In this study, we identified, characterized and investigated the physiological functions of group I LPMO15 (MaLPMO15-1 and PhLPMO15-1) in two longhorn beetle species, Monochamus alternatus and Psacothea hilaris. In both species, depletion of LPMO15-1 transcripts in last instar larvae by RNAi had no effect on subsequent larval-pupal molting and the resulting pupae developed normally. However, adverse effects on their development were observed during the pupal-adult molting period. The pharate adults were unable to shed their old pupal cuticle and died entrapped in their exuviae probably due to a failure of degradation of the chitin in their old cuticle, which is critical for completion of the insect molting and continuous growth.

To investigate the anti-inflammatory activity of submerged culture using Cordyceps militaris mycelium, culture-including mycelia was extracted and lyophilized into postbiotics (hot-water extract; CM-HW). HW was fractionated into crude polysaccharide (CM-CP) by ethanol precipitation, and CM-CP was further dialyzed into CM-DCP by dialysis with running water using 12~14 kDa dialysis tube. When the cytotoxicity of subfractions against cells was assessed, no subfraction had a cytotoxic impact that was substantially different from the control groups. In an inflammatory model using LPS-stimulated RAW 264.7 cells, CM-DCP significantly decreased IL-6 and MCP-1 production levels compared to the LPS-control group. CM-DCP also inhibited IL-6 and IL-8 secretion in HaCaT keratinocytes stimulated with TNF-α and IFN-γ. In the meanwhile, the neutral sugar content and mannose ratio of anti-inflammatory CM-DCP were higher than the other fractions, and CM-DCP contained β-1,3/1,6-glucan of 216.1 mg/g. High pressure size exclusion chromatography revealed that CM-DCP contained molecules with a molecular weight range of 5.6 to 144.0 kDa. In conclusion, postbiotics of C. militaris mycelium significantly promoted anti-inflammatory activity, suggesting that neutral polysaccharides including Glc and Man contribute to the anti-inflammation in RAW 264.7 or HaCaT cells.

본 연구는 발효노니 다당체 추출물(Vitalbos)을 건강기능 식품 소재로 활용하기 위해 DAA, 총당 함량, 단당류 3종 (galacturonic acid, glucose 및 galactose)을 지표성분으로 설정하고, 지표성분에 대한 효과적인 분석법 설정 및 검 증을 위해 수행되었다. 기존에 보고된 분석법 검증 방법 을 수정하여 특이성, 직선성, 정밀성, 정확성, 검출한계 (LOD) 및 정량한계(LOQ)를 고성능 액체크로마토그래피와 페놀-황산법을 이용하여 측정하였다. 그 결과 DAA 및 단 당류 3종의 표준용액과 Vitalbos의 머무름 시간이 일치하 였으며 스펙트럼 또한 동일하여 분석법의 특이성을 확인 하였다. 지표성분의 검량선 상관계수(R2)는 0.9995-0.9998 범위로 0.99 이상의 우수한 직선성을 나타냈다. Intra-day 및 inter-day 정밀도는 0.14-3.01%의 범위로 5% 미만의 우 수한 정밀도를 나타냈고 회수율은 95.13-105.59% 범위에 서 우수한 정확도를 보였다. DAA 분석의 LOD와 LOQ는 각각 0.39 μg/mL 및 1.18 μg/mL이었으며 총당 함량의 LOD 및 LOQ는 각각 0.84 μg/mL 및 2.55 μg/mL로 측정 되었다. 단당류 3종에 대한 LOD는 0.48-0.81 μg/mL의 범 위였으며, LOQ는 1.45-2.44 μg/mL 범위에서 정량분석이 가능한 것으로 나타났다. 분석법 검증 결과, 특이성, 직선 성, 정밀성 및 정확성 모두 우수한 분석법임을 검증하였 으며, LOD와 LOQ 또한 Vitalbos 분석에 적합하였음을 확 인하였다. 검증된 분석법을 이용하여 Vitalbos의 지표 성분 함량을 측정하였을 때, DAA, 총당 함량, galacturonic acid, glucose 및 galactose의 함량은 각각 2.31±0.06 mg/dry weight g, 475.92±5.95 mg/dry weight g, 72.83±1.05 mg/dry weight g, 71.63±2.44 mg/dry weight g 및 67.30±2.31 mg/dry weight g으로 측정되었다. 본 연구에서 검증된 분석법을 사용했을 때 Vitalbos의 지표성분 3종에 대하여 우수한 재현성으로 정 량분석이 가능하였으며, 건강기능식품 소재로의 품질관리에 기여할 수 있을 것으로 판단된다.

The purpose of this study was to investigate the biological activity of fucoidan, a sulfur-containing polysaccharide, on cytotoxicity and apoptosis in the human HT-29 colorectal cancer cell line using cell viability, Flow cytometry, Western blot, and RT-PCR analyses. Fucoidan inhibited the proliferation of HT-29 cells by 39.6% at a concentration of 100 μg/mL for 72 h. The inhibition was dose-dependent and accompanied by apoptosis. Flow cytometric analysis showed that fucoidan increased early apoptosis and late apoptosis by 65.84% and 72.09% at concentrations of 25 and 100 μg/mL, respectively. Analysis of the mechanism of these events indicated that fucoidan-treated cells exhibited increases in the activation of caspase-3, caspase-8, and PARP in a dose-dependent manner. These results suggest that fucoidan may inhibit the growth of human colorectal cancer cells by various apoptosis-promoting effects, as well as by apoptosis itself.

Maca roots (Lepidium meyenii) are an important medicinal herb that have long been used by the Andes-indigenous peoples and South Americans. In Korea, recently, it has attracted attention as a health food material because of nutritional values and physiological activities. The purpose of this study was to investigate the industrial applicability of maca (red and golden varieties; R&G) as immunostimulating materials. In the macrophage stimulating assay using RAW 264.7 cells at 125~500 μg/mL of non-cytotoxicity doses, G-HW showed the most potent production of TNF-α, IL-6 and nitric oxide compared to red maca or the other extracts. The general component analysis results showed that all extracts comprised more than 90% neutral sugars with small amounts of uronic acid and protein. Meanwhile, component sugar analysis showed the difference in the content of uronic acids of cold- and hot-water extract. Additionally, the further fractionation of G-HW into crude polysaccharide (G-CP) greatly enhanced the macrophage stimulating activity, and G-CP contained macromolecules over 144 kDa, comprised mainly of glucose and galacturonic acid (51.0 and 34.9%). In conclusion, crude polysaccharide from maca showed industrial applicability as immunostimulating material, and especially golden maca showed higher macrophage stimulating activity than red maca.

After ethanol (BM-E and RW-E) and hot-water (BM-HW and RW-HW) extracts were fractionated from two herbal mixtures (BM and RW), their physiological activities were investigated. All extracts consisted of more than 50% of neutral sugar, with their total polyphenol levels higher than flavonoid levels. Radical scavenging activities of EtOH extracts remained significantly higher compared to that of hot-water extracts, and in particular, RW-E showed consistently higher antioxidant activity than BM-E. When anti-inflammatory activities of the extracts were evaluated by LPS-stimulated RAW 264.7 cells at 10~500 μg/mL non-cytotoxicity doses, BM-E showed significantly higher levels of TNF-α, IL-1β, IL-6, and nitric oxide inhibitory activity than those of hot-water extracts and RW-E. Murine peritoneal macrophage cells were shown to be enhanced in crude polysaccharides (BM-CP and RW-CP fractionated from BM-HW and RW-HW) compared to hot-water extracts and polysaccharide K (PSK, positive control). Especially, RW-CP exhibited higher activity than BM-CP, and component sugar analysis showed that BM-CP mainly contained galacturonic acid, glucose, arabinose, galactose, and xylose (34.5%, 33.9%, 16.1%, 7.1%, and 6.3%, respectively), whereas RW-CP showed different measurements (29.5%, 59.2%, 5.0%, 4.5%, and 0.2%). In conclusion, two herbal mixtures could contain varying sets of physiological activities dependent on different extraction and fractionation methods.

The purpose of this study was to observe the effects of the polysaccharide (GLP) obtained from the liquid cultured Ganoderma lucidum on the lipidperoxidation in a rat liver microsome and hepatotoxicity in the primary cultured rat hepatocytes. It is well known that the polysaccharide of Ganoderma lucidum exhibits hepatoprotective activity, antitumor activity etc., which many suggest a relationship to lipidperoxidation. The effect of GLP on CCl4- and galactosamineintoxicated cytotoxicity in the primary cultured rat hepatocytes were reduced the GPT value. In order to the estimate the effects of anti-lipidperoxidation of the polysaccharide, enzymatic and nonenzymatic reaction assays were performed, in vitro, in the rat liver microsome. An enzymatic lipidperoxidation reaction by ADP＋FeCl3＋NADPH and CCl4＋NADPH, GLP (1 mg/mL) inhibited 77.4% and 39.4%, respectively, and the nonenzymatic reaction displayed a 97.4% strongly inhibition. In the enzymatic and nonenzymatic inducers treated with GLP, the formation of malondialdehyde (MDA) progressively decreased by raising the GLP concentration. These results suggest that the anti-lipidperoxidation and radical scavenging activity of GLP may play an important part in the liver protection action.