Aqueous-soluble components of sporoderm-removed Ganoderma lucidum spore powder promote ferroptosis in oral squamous cell carcinoma

Objective: Ferroptosis is a novel cell death process which displays a promising role in cancer treatment.However, clinically available drugs targeting ferroptosis are rarely used, and yet there are no studies reporting on inducing ferroptosis via Chinese herbal extracts. Here we explored the tumor inhibition effects of Ganoderma lucidum(G. lucidum) on oral squamous cell carcinoma(OSCC). Specifically, we aimed to clarify the biological mechanism of components in the dietary, aqueous-soluble sporoderm-removed G. lucidum spore powder(A-GSP).Methods: Preliminary transcriptome analysis revealed the significant enrichment of the ferroptosis pathway.Cellular Fe2+, glutathione(GSH), malondialdehyde(MDA), reactive oxygen species(ROS) and lipid peroxide levels were measured to identify ferroptosis occurrence. Western blotting was used to measure ferroptosis-related proteins. Changes in mitochondria morphology and function were observed with transmission electron microscopy(TEM) and ATP detection assays. Ferroptosis inhibitor ferrostatin-1 was then used to verify the anti-tumor effects of A-GSP. Finally, nude mice xenograft models of oral cancer confirmed that A-GSP inhibited tumor growth.Results: A-GSP promoted ferroptosis in oral cancer cells by inducing Fe2+influx, GSH depletion, as well as lipid peroxide and ROS accumulation. Ferroptosis-related proteins exhibited corresponding changes, particularly Acylco A synthetase long chain family member 4(ACSL4) increase and glutathione peroxidase 4(GPX4) decrease. A-GSP considerably lowered mitochondrial volume and ridge number, while significantly decreasing ATP production. Ferrostatin-1 reversed all of these A-GSP-induced changes. In vivo, A-GSP exerted a ferroptosismediated tumor-suppressing effect without observable adverse reactions.Conclusions: Our findings demonstrate the therapeutic potential of A-GSP for treating patients with OSCC by targeting ferroptosis.

Effects of Ganoderma lucidum spore powder on astrocyte expression and glutamine synthetase activity in the hippocampal region of epileptic rats

BACKGROUND： Recent studies have demonstrated that astrocyte dysfunction plays a central role in inhibiting epileptic seizures and that regulation of astrocyte function may be a new target for treatment of epilepsy. OBJECTIVE： To observe the effects of Ganoderma lucidum spore powder （GLSP） on astrocyte morphology and glutamine synthetase （COS） activity in the hippocampal region of epileptic rats. DESIGN, TIME AND SETTING： A randomized, controlled animal experiment was performed at the Function Laboratory, College of Basic Medicine, Jiamusi University between October and December 2006. MATERIALS： A total of 30 Sprague Dawley （SD） rats were randomized to three groups （n = 10）： control, model, and GLSP. GLSP was sourced from Jiamusi Wild Ganoderma Lucidum Planting Base and prepared to 30 g/L with physiological saline before use. Pentylenetetrazol （PTZ） （10 g/L） was provided by Sigma Company, USA. METHODS： The control group received intraperitoneal （i.p.） and intragastric （i.g.） physiological saline. Following epilepsy induction by i.p. administration of PTZ （35 mg/kg）, rats from the mode/and GLSP groups were ig injected with physiological saline and GLSP （300 mg/kg）, respectively. Each compound was administered once per day, for a total of 28 successive days. Epileptic seizure convulsions were graded 0-5. A higher grade indicated more severe epilepsy. Only those rats showing stage 2 or higher convulsions at least 5 times successively were included in further experiments. MAIN OUTCOME MEASURES： Immediately alter injection, seizure activity was monitored for 30 minutes to determine the latent period and seizure duration; simultaneously, astrocyte numbers and GS activity in the hippocampal region of rats with epilepsy were detected by immunohistochemistry. RESULTS： All 30 rats were included in the final analysis. On day 28, following PTZ administration epileptic seizures were not found in the control group. In the GLSP group, rats exhibited rhythmic head nodding or facial spasms, and the latent period was significantly longer than that of the model group （P 〈 0.05）. In the model group, the majority of rats exhibited myoclonus or generalized convulsions, and epileptic seizure duration was slightly （but not significantly） longer than that in the GLSP group （P 〉 0.05）. Within 2-3 minutes of PTZ injection, the model group exhibited grade 4 or 5 epileptic seizures, and the GLSP group mostly showed grade 2 or 3, and occasionally grade 4 or 5, epileptic seizures. All rats recovered within 30 minutes. The model group exhibited significantly increased astrocytes （P 〈 0.05）, with thicker cellular processes and a higher number of cellular processes, and significantly decreased GS activity （P 〈 0.05） tban the control group. The astrocyte count was significantly decreased but GS activity was significantly increased in the GLSP group when compared with the model group （P 〈 0.05）; however, astrocyte appearance was similar in both groups （P 〈 0.05）. CONCLUSION： GLSP can effectively inhibit astrocyte numbers and elevate GS activity in the hippocampal region of rats with epilepsy, thereby reducing epileptic seizures.

Fermentation Production of Ganoderma lucidum by Bacillus subtilis Ameliorated Ceftriaxone-induced Intestinal Dysbiosis and Improved Intestinal Mucosal Barrier Function in Mice

Objective This study aimed to develop a type of Ganoderma lucidum(G.lucidum)-probiotic fermentation broth that can effectively improve the intestinal mucosal barrier function of mice with ceftriaxone-induced intestinal dysbiosis.Methods By means of absorbance of optical density(OD)value and phenol-concentrated sulfuric acid measurement of polysaccharide content,the probiotic species can grow on the medium of G.lucidum were screened out,and the concentration of the medium of G.lucidum was determined,and the fermentation broth was prepared for subsequent experiments.Thirty-two SPF grade male BALB/c mice were randomly divided into four groups(eight mice in each group),namely control group(CON),intestinal mucosal barrier damage model group(CS),fermentation broth intervention group(FT)and G.lucidum medium intervention group(GL),respectively.The intestinal dysregulation model was induced by intra-gastric administration of 0.2 mL ceftriaxone sodium(twice a day for seven consecutive days).From day 8,the FT group and GL group were gavage with 0.2 mL fermentation broth and G.lucidum medium,respectively.On day 15,all mice were sacrificed.To draw the weight curve and measure the cecal index;pathological examination of colon tissues with HE staining;serum levels of LPS,IL-10,TNF and IL-6 were detected by ELISA.Flow cytometry was used to analyze the changes of CD3+T cells,CD4+T cells,CD8+T cells and macrophages in spleen.16S rRNA sequencing was performed to detect the intestinal microbiota structure of mice.Results Bacillus subtilis can decompose and utilize G.lucidum fruiting body medium,and the suitable concentration of G.lucidum fruiting body medium is 33.2 mg/mL.The effect of Bacillus subtilis-G.lucidum fermentation broth on the damage of intestinal mucosal barrier caused by ceftriaxone sodium was reduced,the body weight of mice recovered and colon swelling improved,colon histopathological injury was alleviated,inflammatory cell infiltration was alleviated,serum IL-10 increased significantly,LPS、TNF-αand IL-6 decreased significantly compared with model group,and the proportion of T cells and intestinal dysbiosis was improved.Conclusions The experimental results suggest that Bacillus subtilis-G.lucidum fermentation broth can effectively improve the intestinal barrier function damage,immune dysfunction and intestinal dysbiosis caused by antibiotic overdose,and has a certain regulatory effect on intestinal mucosal barrier function.

Effect of Ganoderma lucidum spore on expression of insulin-like growth factor-1, nuclear factor-kappa B, and neuronal apoptosis in the epileptic rat brain

BACKGROUND：It has been reported that Ganoderma lucidum spore powder, a very well known Chinese traditional medicine, can affect immunoregulation, free radical scavenging, and anti-hypoxia responses. OBJECTIVE： To investigate the effect of Ganoderma lucidum spore powder on expression of insulin-like growth factor-1 （IGF-1）, nuclear factor-κB （NF-κB） and neuronal apoptosis in rats with pentylenetetrazol （PTZ）-induced epilepsy. DESIGN, TIME AND SETTING： A cellular and molecular biology experiment with randomized controlled study design was performed at the Central Laboratory of Basic Medical College of Jiamusi University from June to August 2005. MATERIALS： Thirty healthy, adult, male, Wistar rats were selected and randomly divided into 3 groups （10 rats per group）： control, epilepsy model, and Ganoderma lucidum spore powder. A sub-eclampsia PTZ dose （35 mg/kg） was intraperitoneally injected to induce epilepsy in the latter two groups. Wild Ganoderma lucidum spore powder （30 g/L） was provided by the wild Ganoderma lucidum plant nursery at Jiamusi, China. Immunohistochemical detection and terminal deoxynucleotidyl transferase-mediate dUTP nick end-labeling （TUNEL） kits were purchased from Wuhan Boster Biological Technology Co., Ltd., China. METHODS： Ganoderma lucidum spore powder was intragastrically administered at a dose of 10.0 mL/kg, once a day for 28 days. In the epilepsy and control groups, an equivalent volume of normal saline was intragastrically administered. MAIN OUTCOME MEASURES： Immunoreactivity for IGF-1 and NF-κB/P65 were detected by immunohistochemical staining. Neuronal apoptosis was detected using TUNEL methods. RESULTS： The hippocampus and cerebral cortex of rats with PTZ-induced epilepsy exhibited a higher number of apoptotic cells at high magnification （×400）, compared with the control group. Expression of IGF-1 and NF-κB were higher in the epilepsy group, compared with the control group （P 〈 0.01）. In Ganoderma lucidum spore-treated rats, fewer apoptotic cells were observed in the hippocampus and cerebral cortex, expression of NF-κB/P65 was lower, and immunoreactivity to IGF-1 increased more distinctly, compared with the epilepsy group. In addition, seizure latency was longer on 17, 21, and 25 days post-PTZ treatment in the Ganoderma lucidum spore powder group, compared with the epilepsy group （P 〈 0.05-0.01）. CONCLUSION： Ganoderma lucidum spore powder down-regulated expression of NF-κB in brain tissues of rats with PTZ-induced epilepsy, increased immunoreactivity to IGF-1, and inhibited neuronal apoptosis. These results indicated that Ganoderma lucidum spore powder has a neuroprotective effect.

Ganoderma lucidum spore powder modulates Bcl-2 and Bax expression in the hippocampus and cerebral cortex,and improves learning and memory in pentylenetetrazole-kindled rats

We studied the effects of Ganoderma lucidum spore powder on Bax and Bcl-2 expression and neuronal apoptosis in pentylenetetrazole-kindled epileptic rats. Sixty adult rats were randomly divided into a control group, an epileptic group （kindled） and three medication groups （150,300 450 mg/kg given to kindled rats）. Bax and Bcl-2 immunohistochemistry and TUNEL labeling showed that the number of Bax- and TUNEL-positive cells in the hippocampus and cerebral cortex decreased significantly in the high-dose medication group, while the number of Bcl-2 immunoreactive cells increased. The Morris water maze test showed that high-dose treatment significantly shortened escape latency and increased spatial probe trial performance. Our findings indicate that a high dose of Ganoderma lucidum spore powder upregulates the expression of antiapoptotic Bcl-2 protein in the hippocampus and cerebral cortex, inhibits proapoptotic Bax expression, and decreases seizure-induced neuronal apoptosis. Further, Ganoderma lucidum appears to protect against epilepsy-related learning and memory impairments.

Treatment Effect of Fermentation of Ganoderma lucidum to Immune and Antioxidant Functions in Immunosuppressed Mice

The immune regulatory and antioxidant roles of Ganoderma lucidum were investigated using cyclophosphamide(CTX)-induced immunosuppressed mice. Mice were randomly divided into groups: untreated(groupⅠ), immunosuppressed(groupⅡ), unfermented G. lucidum polysaccharide(groupⅢ) and fermented G. lucidum polysaccharide(group Ⅳ). After seven consecutive days of treatments, the serum concentration of IL-4, IFN-gamma, IgG, IgA and IgM and the liver activity of GSH-Px, SOD, CAT and MDA enzymes were analyzed. The contents of IL-4, IFN-γ in serum and GSH-Px, SOD and CAT in liver tissues were significantly reduced in groupⅡ compared with those in group I, indicating successful CTX-induced immunosuppression. Interestingly, the results showed that the above immune and antioxidant indicators were significantly improved after G. lucidum polysaccharide treatment, regardless of fermentation. However, fermentation caused changes in polysaccharide structure, which might have a significant impact on immune regulation and antioxidant functions in immunosuppressed mice.

Study on Key Techniques for High-quality and High-yield Cultivation of Ganoderma lucidum Spore Powder

Comparative test of 4 Ganoderma lucidum varieties from different sources showed that the mycelium of Chizhi 1 grew fast with thick and dense hyphae,round and solid cap,and high spore powder yield.Chizhi 1 was proved to be an excellent variety because of its strong resistance and high spore powder yield.Different cultivation materials were chosen and combined to form 3 cultivation formulations.The results showed that Formulation(3),in which basswood was soaked in nutrient solution for 24 h,presented fast mycelial growth and high spore powder output,and therefore was proved to be a high-yield formulation of Ganoderma lucidum spore powder.

固态发酵三七渣产灵芝三萜的工艺条件优化

该研究通过单因素试验考察发酵温度、接种量、发酵时间对固态发酵三七渣生产灵芝三萜的影响,并采用响应面法对其发酵工艺条件进行优化。结果表明,发酵温度、接种量、发酵时间对灵芝三萜含量影响均极显著(P<0.01),这3个因素对于发酵培养物中灵芝三萜含量影响程度排序为:接种量>发酵时间>发酵温度。发酵温度和接种量之间的交互作用对灵芝三萜含量的影响极显著(P<0.01);而接种量和发酵时间、发酵时间和发酵温度之间的交互作用对灵芝三萜含量的影响不显著(P>0.05)。在实验室小试条件下最佳发酵工艺条件为:发酵温度30℃,发酵时间16 d,接种量为11%。在此优化条件下,灵芝三萜含量为(4.110±0.186)mg/g,比优化前提高了12.0%。

破壁灵芝孢子粉中多种重金属元素及硒含量检测方法探讨

采用微波消解进行前处理,三重四极杆-电感耦合等离子体质谱法测定破壁灵芝孢子粉中镉、铬、镍、砷、铅、汞及硒元素含量,对方法条件进行优化和讨论,并对20批次破壁灵芝孢子粉进行测定。结果表明,镉、铬、镍、砷、铅和汞元素在氦气碰撞模式,硒元素在氧气反应模式下,采用内标校正定量,可达到良好的测定效果。在优化的条件下,各元素线性关系良好,检出限均可达μg·kg^(-1)级别;镉、铬、镍、砷、铅及硒元素的加标回收率在90.3%~105.0%,汞的加标回收率在82.1%~92.5%;方法精密度在5%以内。此方法前处理操作简便,检测灵敏度高,稳定性好,给灵芝孢子粉多元素测定提供了良好的技术支撑。

灵芝菌发酵枇杷叶茶的研制

以枇杷叶为主料,研究利用灵芝菌来发酵枇杷叶制成养生茶,并测定其营养成分与重金属、农残含量。结果表明:美芝、大红芝可作为发酵枇杷叶的灵芝菌株;美芝菌丝生长速度较大红芝的快,生长速度配方B>配方C>配方A;添加少量的玉米粉、甜叶菊能够促进灵芝菌丝的生长,添加少量的甜叶菊有助于提升发酵茶的口感;发酵后的枇杷叶中多糖、总蛋白、游离氨基酸含量均有显著增加,但三萜酸含量有所下降;不同灵芝菌株发酵后的枇杷叶营养成分有所差别;发酵后的枇杷叶的重金属、农残含量均符合检测标准。

灵芝室内栽培模块化收粉装置的结构设计

灵芝是一种具有多种功效的中药材,近年室内人工栽培灵芝得以广泛应用。该技术采用代料栽培方法,逐步形成工厂化栽培。灵芝孢子粉的收集是灵芝栽培的重要工序之一,设计一种灵芝孢子粉收集装置,能有效地避免因受外境影响而导致孢子粉逃逸,提高孢子粉产量,降低劳动强度和用工用料成本。该装置可重复使用,以提高经济效益,促进灵芝工厂化栽培的规模化发展。

灵芝孢子粉咀嚼片的研发与制作

以灵芝孢子粉为原料,研制灵芝孢子粉咀嚼片。通过配方优化试验和感官品质评价分析,得出最佳配方组合为灵芝孢子粉14.4%、低聚果糖34.0%、全脂奶粉51.6%。按照此配方制作的灵芝孢子粉咀嚼片口感和形状俱佳,硬度和脆碎度均符合标准,入口易崩解,且无杂质产生。

酶法提取灵芝孢子粉胞壁水溶性糖的工艺优化及分子量分布

为提高灵芝(Ganoderma lucidum)孢子粉胞壁水溶性糖的提取效率,以灵芝孢子粉胞壁残渣为研究对象,用单因素实验、响应面实验优化酶法胞壁水溶性糖提取工艺,并比较酶法和不同物理法处理的灵芝孢子粉胞壁水溶性糖的分子量分布。结果表明:最佳提取工艺为溶壁酶加量2%,酶解时间5 h,料液比1︰10(g︰m L)、温度40℃、pH6.5,在此条件下提取的灵芝孢子粉胞壁水溶性糖含量达到17.86%。分子量分布分析结果表明,酶解后的水溶性糖以低聚糖为主。研究结果可为有效利用和开发灵芝孢子粉提供参考。

UPLC-Q-TOF-MS分析灵芝孢子粉中三萜类化合物

针对目前灵芝孢子粉中三萜类化合物的检测方法大多需要标准品且标准品稀少、昂贵问题,通过UPLC-Q-TOF-MS技术,对灵芝孢子粉中三萜类化合物进行定性检测和鉴别.结果表明,在灵芝孢子粉提取物中共鉴定25种灵芝三萜化合物.可为创建灵芝三萜指纹图谱库,灵芝三萜的检测、药效评价及品质测定提供参考.

不同菠萝叶渣添加量对赤灵芝发酵基质营养成分及体外抗氧化活性研究

【目的】揭示不同菠萝叶渣添加量对赤灵芝发酵基质营养成分和体外抗氧化活性的影响,为菠萝叶渣的开发利用提供了新的基础数据支撑。【方法】采用全自动氨基酸分析仪、OPLA-DA结合S-plot等仪器和方法分析发酵基质中粗纤维、多糖、总萜、水解氨基酸和游离氨基酸等营养成分,以及赤灵芝发酵基质水提物的羟基自由基清除能力差异、DPPH自由基清除能力和总抗氧化能力等体外抗氧化活性的变化规律进行分析。【结果】菠萝叶渣胁迫下促进赤灵芝发酵基质中粗纤维降解、释放多糖含量;对赖氨酸和甘氨酸代谢路径具有显著的调节作用含量;游离氨基酸中肌氨酸、a-氨基己二酸呈现出增加的趋势,其他氨基酸整体上呈现出下降的趋势。其中天冬氨酸和苏氨酸等8种游离氨基酸含量差异显著,且不同菠萝叶渣添加量下,存在特异性氨基酸成分;菠萝叶渣有助于提高赤灵芝发酵基质水提物的羟基自由基清除能力和总抗氧化能力,降低DPPH自由基清除能力。【结论】不同菠萝叶渣添加量对赤灵芝发酵基质粗纤维、氨基酸组成以及相关代谢路径具有不同的调节作用。

灵芝菌株深层发酵及发酵饮料的配方优化

灵芝(Ganoderma lucidum)是一种营养和药用价值极高的真菌。为提高灵芝深层发酵效益和促进发酵液的开发利用,本研究采用可食用原料配制培养基,以菌丝生物量为评价指标,采用正交试验优化培养基组成。同时将灵芝发酵液用于饮料制备,并使用模糊数学综合评价法优化发酵饮料的配方。结果表明,蔗糖添加量为30 g·L^(-1)和玉米粉添加量为15 g·L^(-1)条件下配制的培养基效果最佳,菌丝生物量可达6.86 g·L^(-1),所得发酵液中胞外多糖含量为430.21μg·mL^(-1)、多酚含量为26.07μg·mL^(-1)、黄酮含量为8.16μg·mL^(-1)。发酵液体积分数为60%、柠檬酸添加量为0.5%、黄原胶添加量为0.30%、蔗糖添加量为6%和柠檬黄添加量为0.05%的工艺条件下制备的发酵饮料的感官评分最高。

灵芝孢子粉复合饮料的加工工艺研究

为丰富功能性饮料市场,以灵芝孢子粉为原料制备一款功能性饮料,以灵芝孢子粉浸提液添加量、山楂枸杞浸提液添加量、甘草浸提液添加量、木糖醇添加量为考查因素进行单因素试验,采取正交试验筛选出最优工艺。结果表明,灵芝孢子粉浸提液添加量为10%,山楂枸杞浸提液添加量为7.5%,甘草浸提液添加量为2.5%,木糖醇添加量为14%时,感官评分最高。此时产品呈浅褐色,有灵芝孢子粉特有的气味与口感,澄清透明,无杂质沉淀,酸甜协调。

复合花粉灵芝发酵工艺优化及活性物质组分分析

对复合花粉灵芝液体发酵工艺进行优化并对发酵液中的活性物质进行分析,为花粉发酵加工提供一定的理论基础和数据支持。使用灵芝对复合花粉进行液体发酵,通过单因素及正交试验对发酵工艺进行优化,并采用高效液相色谱法分析发酵液中多糖的单糖组成,利用超高效液相色谱质谱联用法分析发酵液中的酚酸。结果表明,最优发酵工艺条件为花粉添加量1.5%、初始pH为7、发酵时间11 d,获得的复合花粉灵芝发酵液多糖含量为(393.69±4.09)mg/100 mL,总黄酮含量为(5.63±0.20)mg/100 mL,铁离子还原抗氧化能力达到(2.303±0.143)μmol/mL;发酵液中的多糖由甘露糖、核糖、鼠李糖、葡萄糖醛酸、半乳糖醛酸、葡萄糖、半乳糖、木糖、阿拉伯糖、岩藻糖10种单糖组成,其中葡萄糖含量(49.42%)最高;发酵液中含有5种酚酸类物质,分别为槲皮素、对羟基苯甲酸、木犀草素、芹菜酸和山奈酚,其中山奈酚含量(683.71 mg/kg)最高。综上,复合花粉灵芝发酵液富含多糖和黄酮类物质,抗氧化能力较高,具有潜在的保健功效。

党参茎叶液体发酵条件优化及成分变化分析

该试验以灵芝菌(Ganoderma lucidum)为菌种发酵党参茎叶,以多糖含量为响应值,通过单因素及响应面试验对党参茎叶液体发酵条件进行优化,并对发酵前后的党参茎叶成分变化进行分析。结果表明,党参茎叶液体发酵的最佳条件为发酵时间6.5 d、料液比7.5∶100(g∶mL)、培养基初始p H值8,在此条件下测得发酵党参茎叶粗多糖含量为37.59 mg/g。与发酵前相比,发酵党参茎叶中粗多糖含量增加115.32%,总皂苷含量降低56.22%,总黄酮含量降低77.33%。

不同干燥方式对灵芝孢子粉中灵芝孢子油过氧化值的影响

目的:比较不同干燥方式对灵芝孢子粉中灵芝孢子油过氧化值的影响。方法:以西藏灵芝孢子粉为例,采用不同的干燥方式将灵芝孢子粉干燥至合格,然后进行破壁,用超临界CO_(2)萃取法提取灵芝孢子油,检测灵芝孢子油的过氧化值,并比较不同干燥方式对灵芝孢子油的过氧化值的影响。结果:微波干燥和真空干燥方式得到的灵芝孢子油过氧化值均合格,分别为0.13%,0.14%;热风干燥和晾晒干燥方式得到的灵芝孢子油过氧化值不合格,分别为0.16%、0.17%。结论:水分变化率可能是灵芝孢子粉干燥过程中的重要影响因素,微波干燥方式水分变化率远高于其他方式,干燥效率高,耗能少,且不影响孢子油过氧化值,适合干燥大规模样品;其它干燥方式相对比较费时耗能,且灵芝孢子粉存在变质的风险。