正交试验优选桑黄多酚超声提取工艺

目的:优化桑黄多酚的超声提取工艺。方法:以乙醇浓度、超声时间、超声温度和料液比为考察因素,以桑黄总多酚提取率为评价指标,采用单因素试验和正交试验优选提取工艺。结果:最佳提取工艺为乙醇浓度60%,超声时间30 min,超声温度50℃,料液比1∶25。在此条件下,桑黄总多酚提取率可达26.4 mg·g-1。结论:采用超声辅助提取桑黄多酚,时间短、得率高,是一种高效、快捷的方法。     

裂蹄木层孔菌乙酸乙酯萃取物对人肝癌细胞SMMC-7721的抑制作用研究

目的：探讨裂蹄木层孔菌乙酸乙酯萃取物在体外的抗肝癌活性。方法：采用磺酰罗丹明染色法（SRB法）,对人肝癌细胞SMMC-7721进行体外抗肿瘤实验。结果：裂蹄木层孔菌乙酸乙酯萃取物对人肝癌细胞SMMC-7721表现出比较明显的高浓度杀死和中低浓度抑制的效果,且抑制率随萃取物浓度的升高而增大,呈剂量效应关系;其半数抑制浓度（IC50）为21.5μg.mL-1。结论：裂蹄木层孔菌乙酸乙酯萃取物对人肝癌细胞SMMC-7721具有比较明显的抑制作用。     

Comparison of intraperitoneal anti-adhesive polysaccharides derived from Phellinus mushrooms in a rat peritonitis model

AIM: To assess the adhesion- and abscess-reducing capacities of various concentrations of polysaccharides derived from fungus, Phellinus gilvus (PG) or Phellinus linteus(PL) in a rat peritonitis model. METHODS: In 96 SD rats, experimental peritonitis was induced using the cecal ligation and puncture model (CLP). Rats were randomly assigned to 8 groups; Ringer's lactate solution (RL group), hyaluronic acid (HA group), 0.025%, 0.25%, and 0.5% polysaccharides from PG (PG0.025, 0.25, and 0.5 groups), and PL (PL0.025, 0.25, and 0.5 groups). Adhesions and abscesses were noted at 7 d after CLP. RT-PCR assay was performed to assess the cecal tissue. RESULTS: Adhesion formation was significantly reduced in PG0.25, 0.5, PL0.25, 0.5, and HA groups (2.5±0.7, 2.4±0.7, 3.8±1.0, 3.6±0.8, and 2.7±1.1, P<0.05). The incidence of abscesses was significantly reduced in all treated groups compared to RL group (58%, P<0.05). The urokinase-type plasminogen activator (uPA) gene expression was greatly up-regulated by increasing the concentration of polysaccharides. The urokinase-type plasminogen activator receptor (uPAR) and tumor necrosis factor (TNF)-α mRNA were highly expressed in PG0.25, 0.5, PL0.25, and 0.5 groups. CONCLUSION: We concluded that 0.5% polysaccharide derived from PG and PL was the optimal concentration in preventing adhesion and abscess formation and may act by modulating activity of uPA and TNF-α in a rat peritonitis model.     

Effects of mycelial culture of Phellinus linteus on ethanol-induced gastric ulcer in rats

The gastroprotective effects of a mycelial culture of Phellinus linteus (MCPL) were evaluated by determining the ulcer index, gastric mucus content, histopathological observation and histochemical properties of mucin in an ethanol-induced ulcer model of rats. Preadministration with MCPL at doses of 20 and 60 mg/kg, showed a significant decrease of bleeding and ulcer index and alleviated the histopathological changes induced by ethanol such as hemorrhage and necrosis. Ethanol treatment decreased the gastric adhesion mucus content, but a higher level of gastric mucus persisted after preadministration of MCPL. As for the histochemical properties of mucins, marked changes were observed in both the surface and gland mucous cells in ethanol-treated rats, but these changes were detected only in the surface mucous cells in rat preadministered with MCPL. Using conventional methods for mucins, ethanol-treated rats revealed a decrease of neutral and acid mucin in the surface epithelium and mucous neck cells compared with normal rats. A marked decrease of BSL-1 by lectin histochemistry was also revealed in the ethanol-treated rats. But the MCPL preadministered rats showed similar stainabilities and lectin affinity patterns for mucins as the normal rats. These results indicate that pretreatment with MCPL provided protection of the gastric mucosa from ethanol-induced injury by maintaining the mucus barrier in rats.     

Identification of an anticancer compound against HT-29 cells from Phellinus linteus grown on germinated brown rice

Objective

To isolate and identify the anticancer compound against proliferation of human colon cancer cells from ethyl acetate (EtOAC) extract of Phellinus linteus grown on germinated brown rice (PB).

Methods

EtOAC extract of PB was partitioned with n-hexane, EtOAC, and water-saturated n-butanol. Anticancer compound of n-hexane layer was isolated and identified by HPLC and NMR, respectively. Cytotoxicity against HT-29 cells was tested by SRB assay.

Results

The n-hexane layer obtained after solvent fractionation of PB EtOAC extracts showed a potent anticancer activity against the HT-29 cell line. Atractylenolide I, a eudesmane-type sesquiterpene lactone, a major anticancer substance of PB, was isolated from the n-hexane layer by silica gel column chromatography and preparative-HPLC. This structure was elucidated by one- and two-dimensional NMR spectroscopic data. Atractylenolide I has not been reported in mushrooms or rice as of yet. The isolated compound dose-dependently inhibited the growth of HT-29 human colon cancer cells.

Conclusions

Atractylenolide I might contribute to the anticancer effect of PB.     

桑黄菌发酵过程中酶活性的变化

目的 研究桑黄菌发酵过程中酶活性的变化及其和胞外多糖量的关系。方法 在发酵过程中测定原始菌株和变异菌株的羧甲基纤维素酶、滤纸酶、木质素过氧化物酶（LiP）、锰过氧化物酶（MnP）、漆酶（Lac）活性变化及培养基中胞外多糖量变化。结果 2菌株的羧甲基纤维素酶和滤纸酶的活性变化呈单峰曲线,分别在第4天和第6天达到峰值;原始菌株和变异菌株的LiP、锰过氧化物酶和漆酶活性的变化呈双峰曲线,且原始菌株和变异菌株LiP的酶促反应米氏常数（K_m）分别为25.96、27.07 μg/mL;MnP的K_m分别为13.28、13.49 mmol/mL;Lac的K_m分别为0.12、0.21 mmol/mL;培养基中多糖量前期快速下降,10 d后趋于稳定。结论 桑黄菌发酵时能产生较全面的分解木质素和纤维素的酶系统,原始菌株和变异菌株产生的LiP和MnP为同种酶,Lac为同功酶。桑黄菌胞外酶活性变化和培养基中胞外多糖量变化有一定关系。     

桑黄多糖缓解日本血吸虫感染小鼠氧化应激及 肝纤维化的实验研究

目的　探讨桑黄醇提多糖（PPI）对日本血吸虫感染小鼠氧化应激、肝肉芽肿和肝纤维化的改善作用。方法　采用日本血吸虫尾蚴玻片贴腹感染法建立日本血吸虫肝病小鼠模型。设健康对照组（A组）、感染对照组（B组）、PPI单独治疗组（C组）、吡喹酮单独治疗组（D组）和PPI加吡喹酮混合治疗组（E组）,每组各10只小鼠;除A组外,其他各组每只小鼠感染（30 ± 2）条尾蚴。自感染后42 d开始,D、E组小鼠灌胃给予500 mg/kg吡喹酮,连续2 d;C、E组给予400 mg/kg PPI灌胃,连续给药30 d。HE染色观察小鼠肝组织病理学改变,测定小鼠血清丙氨酸氨基转移酶（ALT）、天门冬氨酸氨基转移酶（AST）、透明质酸（HA）、层黏连蛋白（LN）及小鼠肝组织匀浆中丙二醛（MDA）、超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH?PX）、谷胱甘肽还原酶（GSH?R）、谷胱甘肽（GSH）含量,采用免疫组化技术检测小鼠肝组织中转化生长因子?β（TGF?β）、α?平滑肌肌动蛋白（α?SMA）表达水平,采用实时荧光定量PCR检测Nrf2、Gsta4基因表达水平。结果　日本血吸虫感染但未予治疗小鼠出现典型血吸虫病肝病病理改变,PPI干预后能有效减轻小鼠肝虫卵肉芽肿及胶原沉积。血吸虫病肝病小鼠肝脏脂质过氧化加剧,诱导了氧化应激,小鼠血清中MDA含量增加,GSH和各种抗氧化酶含量下降。与B组相比,PPI治疗抑制了脂质过氧化,提高了GSH含量,恢复了抗氧化酶活性。此外,PPI治疗可抑制TGF?β信号通路,提升Nrf2、Gsta4基因表达水平。结论　PPI在治疗血吸虫病肝纤维化方面发挥重要作用,其内在机制可能是通过上调Nrf2和Gsta4基因表达、改善氧化应激损伤,从而抑制肝脏虫卵肉芽肿形成和肝纤维化。     

桑黄正丁醇提取物对白念珠菌生物膜形成的抑制作用

目的 探讨桑黄正丁醇提取物（butyl alcohol extract of Phellinus igniarius decoction,BAEP）体外对白念珠菌（Candida albicans）标准株SC5314生物膜形成的影响。 方法 采用二倍稀释法测定BAEP对白念珠菌的最低抑菌浓度（minimal inhibitory concentration,MIC）和抑制50%生物膜形成的最低浓度(sessile minimal inhibitory concentration,SMIC50);采用XTT还原法测定BAEP对白念珠菌生物膜代谢的影响;固体培养基上观察BAEP对白念珠菌菌落形态的影响;倒置显微镜与荧光显微镜下分别观察BAEP对白念珠菌生物膜形态与活性的影响;扫描电镜下观察BAEP对白念珠菌生物膜形态结构的影响;实时荧光定量PCR（quantitative real-time PCR,qRT-PCR）检测生物膜相关基因ALS1和HWP1的转录水平变化。结果 BAEP对白念珠菌生物膜的SMIC50为1 024 μg/mL;菌落形态实验观察结果显示,1 024 μg/mL BAEP可影响白念珠菌菌落;倒置显微镜、荧光显微镜下显示1 024 μg/mL BAEP可抑制白念珠菌生物膜的形态和活性;扫描电镜下显示1 024 μg/mL BAEP对白念珠菌生物膜有明显的抑制作用;qRT-PCR检测结果显示BAEP可明显下调HWP1的表达水平和上调ALS1基因的表达水平。结论 BAEP对白念珠菌SC5314生物膜的形成有一定的抑制作用。     

HPLC profiling of Phellinus linteus

HPLC chromatograms of MeOH extracts from a fruit body of the wild-grown P. linteus (natural fruit body), from cultivated fungus (cultivated fruit body), and from the cultured mycelia were compared. The extract prepared from the natural fruit bodies revealed a typical HPLC profile referred to as type 1 with a major peak corresponding to meshimakobnol A (1) together with two minor peaks of hypholomine B (3) and inoscavin A (4); the cultivated fruit bodies exhibited a profile referred to as type 2 with major peaks corresponding to 3 and 4 and a minor peak of 1, and the cultured mycelia showed a profile referred to as type 3 without any of these peaks. We also analyzed HPLC chromatograms of commercial products of P. linteus obtained in the markets. Most of the products claimed to be natural fruit bodies exhibited type 1 profiles, except for one product having an intermediate HPLC profile between type 1 and type 2. The products claimed to be cultivated fruit bodies and cultured mycelia revealed type 2 and type 3 profiles, respectively. The present results indicate that the HPLC chromatogram of the methanol extract of P. linteus can be used as a fingerprint to identify whether the product is from natural fruit bodies, cultivated fruit bodies, or cultured mycelia.     

桑黄对四氯化碳致大鼠肝损伤的保护作用

目的 :研究桑黄对肝损伤的保护作用。方法 :以四氯化碳诱导大鼠肝损伤 ,观察桑黄对血清学和组织学指标的影响。结果 :桑黄治疗组大鼠肝细胞变性明显减轻 ,肝组织结构完好 ;血清氨基酸转移酶水平显著降低 ,蛋白合成能力增强 ;血清活性氧显著减少 ,肝组织超氧化物歧化酶活力提高 ;血清白细胞介素 -4水平降低 ,γ -干扰素显著增高。结论 :桑黄具有保护肝细胞功能 ,抗脂质过氧化和调节炎症因子水平为其作用机理。