Antioxidant activity in HepG2 cells,immunomodulatory effects in RAW 264.7 cells and absorption characteristics in Caco‐2 cells of the peptide fraction isolated from Dendrobium aphyllum

DA‐P, fraction of peptides with a molecular weight <1 kDa isolated from Dendrobium aphyllum, was analysed in three types of cell lines to verify its bioactivity and absorptivity. The cellular antioxidant activity of DA‐P in HepG2 cells was used and results revealed an EC50 of 2.88 ± 0.143 mg mL^?1 and a CAA unit of 63.46 ± 2.11 μm QE/100 g peptides. DA‐P treatment enhanced the secretion of cytokines in RAW 264.7 cells. After demonstrating the presence of tight junctions in Caco‐2 monolayers, the absorption was 25.57% ± 0.016% and 19.7% ± 0.012% from different sides. The relatively high absorption indicated that the antioxidant‐relevant immune functions of DA‐P had a greater possibility to be absorbed by Caco‐2 cells. Free amino acids and LC‐MS/MS analysis indicated the degradation and expulsion of components after the absorption of DA‐P, and Ser‐Ser‐Arg was able to come across the monolayers.     

Isolation,characterisation and sulphation of soluble polysaccharides isolated from Cucurbita maxima

Using hot water extraction, a large number of polysaccharides were obtained from Cucurbita maxima. A DEAE‐Sepharose CL‐6B chromatography column was used to isolate the major polysaccharides from C. maxima. Two fractions were obtained (LP2‐1 and LP2‐2). LP2‐1 and LP2‐2 consisted of neutral polysaccharides (MW: 1.02 × 10⁴ and 4.32 × 10⁸ g mol^?1, respectively) comprised mainly of galactose units. Analyses by FT‐IR spectrometry, partial acid hydrolysis, periodate oxidation, Smith degradation and GC‐MS indicated that LP2‐1 consisted of 85.3% (1→4) glycosidic linkages and 1.7% (1→3) or (1→6) glycosidic linkages. The LP2‐1 backbone consisted of (1→4)‐linked galactose units, which occasionally branched at O6 or O3. The branches were composed of (1→4)‐linked galactose and terminated with galactose (13%). Two sulphated derivatives (SLP2‐1 and SLP2‐2) with variable degrees of sulphation (DS) were obtained by the sulphur trioxide–pyridine method, without degradation of the polysaccharide. DS of PL2‐1 and PL2‐2 was 0.19 and 0.20, respectively.     

兜唇石斛发酵多肽Asp-Asp-Asp-Tyr和Asp-Tyr- Asp-Asp对LPS诱导RAW264.7细胞的抗炎作用

为了研究兜唇石斛发酵多肽Asp-Asp-Asp-Tyr（DDDY）、Asp-Tyr-Asp-Asp（DYDD）对LPS诱导的RAW264.7巨噬细胞抗炎活性。以合成的兜唇石斛发酵多肽为研究对象,采用噻唑蓝法筛选增殖活性最高的发酵多肽浓度,通过中性红吞噬实验和倒置显微镜观察发酵多肽对细胞的吞噬作用和分化形态变化,使用ELISA试剂盒测定细胞中NO及细胞因子IL-1β、IL-6、IL-10和肿瘤坏死因子TNF-α的分泌量。结果表明：12.5、25、50和100 μg/mL四组质量浓度的发酵多肽对细胞无毒性并有增殖作用;100 μg/mL的DDDY和DYDD和1 μg/mL的LPS处理可以激活细胞,增强细胞吞噬能力,相对吞噬率为2.05%、1.97%和2.19%;构建LPS诱导的RAW264.7细胞炎症模型,发现两种发酵多肽处理有效抑制细胞分化,使细胞恢复正常形态;抑制细胞NO分泌能力,100 μg/mL DDDY和DYDD处理组的NO分泌能力降低到LPS组的0.41倍和0.49倍;并且对抑炎细胞因子分泌能力的提高和促炎细胞因子的降低有显著效果,均表现出剂量反应关系。由此可知,DDDY和DYDD对LPS诱导的RAW264.7巨噬细胞炎症反应具有抗炎作用,为后面探究发酵多肽的炎症机制提供理论支持。     

霍山石斛多糖的闪式提取工艺优化及降血糖作用研究

目的 优化霍山石斛多糖的闪式提取工艺,并对其降血糖作用进行研究。方法 采用单因素试验和响应面试验优化霍山石斛多糖的提取工艺,并通过测定空腹血糖、血清胰岛素和糖化血清蛋白水平研究其降血糖功能。结果 霍山石斛多糖的最优提取工艺为料液比1:28 (g:mL),提取电压132 V,提取时间95 s,提取次数4次,在此条件下霍山石斛多糖提取率为(24.14±0.52)%。通过动物试验发现,霍山石斛多糖能极显著降低小鼠血糖值和糖化血清蛋白水平(P<0.01),提高小鼠血清胰岛素含量(P<0.01)。结论 霍山石斛多糖具有较好的降血糖功效。     

基于Caco-2单层细胞模型的兜唇石斛多糖的转运机制研究

以兜唇石斛多糖(DAP)为研究对象,借助人小肠上皮细胞Caco-2(the human colon carcinoma cell line)单层细胞模型,体外模拟DAP在小肠中的转运情况。在前期得到的DAP为基础,探究DAP的粘度对其转运情况的影响,同时建立Caco-2细胞的模型,采用MTT法测定DAP对Caco-2细胞存活率的影响,并考察了DAP在小肠的吸收转运的特征。实验表明:DAP的粘度与其在Caco-2细胞中的转运能力呈负相关趋势,粘度越大,转运速率越慢;在测定TEER值及荧光素钠渗透情况,观察Caco-2细胞的形态,建立了良好的细胞模型;DAP的存在下,Caco-2细胞的存活率为91.8%以上,因此DAP对Caco-2细胞存活无影响;DAP在Caco-2不同单边层的转移率分别为0.35%±0.02%和0.32%±0.02%,表观渗透率分别为(0.42±0.02)×10-6 cm/s和(0.19±0.18)×10-6 cm/s,表明DAP在小肠中不易被吸收。本研究所得结果为进一步研究DAP在肠道中的吸收机制奠定基础。     

铁皮石斛多糖化学修饰及其对免疫活性的影响

采用水提醇沉法结合冻融制备高纯度铁皮石斛多糖,并利用硫酸化、脱乙酰化及羧甲基化修饰方法对铁皮石斛多糖进行化学修饰,探讨不同方法修饰的铁皮石斛多糖对RAW264.7巨噬细胞免疫活性影响。结果表明,铁皮石斛纯多糖(purified polysaccharides of Dendrobium officinale,DOP)的中性糖含量较高,而糖醛酸及蛋白质含量则较低,分别为87.37%、3.19%和0.51%,羧甲基化及硫酸化显著降低了铁皮石斛多糖的中性糖含量。红外分析结果显示硫酸化、脱乙酰化及羧甲基化修饰衍生物均已成功制备。高性能凝胶渗透色谱法测定铁皮石斛多糖分子质量为960 k D,硫酸化及羧甲基化修饰显著增加了其分子质量。体外实验研究发现,硫酸化和脱乙酰化修饰的铁皮石斛多糖能够增强RAW264.7巨噬细胞的免疫活性,而羧甲基化修饰则表现为显著的抑制作用,表明不是所有的官能团接枝改性都可以改善铁皮石斛多糖对免疫活性的影响。     

铁皮石斛多糖调控线虫寿命及抗氧化作用的研究

摘 要: 目的 基于秀丽隐杆线虫自然衰老模型, 探讨铁皮石斛多糖(Dendrobium candidum polysaccharides, DOP)抗衰老作用及其机制。方法 将秀丽隐杆线虫野生型N2随机分为对照组和铁皮石斛多糖组(低、中、高), 观察铁皮石斛多糖对线虫N2寿命、运动能力和咽泵能力的影响; 通过检测氧化应激、抗氧化酶试剂盒以及活性氧评估铁皮石斛多糖对N2线虫抗氧化能力的影响。基于skn-1基因缺陷型突变株(EU1)线虫及带有荧光标记的转基因线虫CL2166分析铁皮石斛多糖延长健康寿限的可能作用途径; 采用实时荧光定量聚合酶链反应技术测定铁皮石斛多糖作用下线虫抗氧化、胰岛素信号途径相关基因转录水平。结果 铁皮石斛多糖中剂量组(250 μg/mL)可显著延长线虫N2的寿命, 平均延寿率达12.73%, 可显著提高运动、咽泵、抗氧化应激能力; 显著提高过氧化氢酶(catalase, CAT)、超氧化物歧化酶(superoxide disumutase, SOD)活性和谷胱甘肽(glutathione, GSH)含量, 降低活性氧(reactive oxygen species, ROS)水平和丙二醛(malondialdehyde, MDA)含量; 铁皮石斛多糖干预处理对skn-1基因缺陷线虫的延寿效果缺失; 但能够增强CL2166线虫的荧光强度; 同时上调sod-5、sod-3、ctl-1、ctl-2、daf-16和skn-1基因的表达水平, 下调daf-2基因的表达水平。结论 铁皮石斛多糖可能通过激活Nrf2/SKN-1和DAF-16途径拮抗氧化应激从而延缓线虫衰老。     

高压灭菌前后党参口服液中多糖的免疫活性研究

研究高压灭菌前后党参口服液中多糖的免疫活性变化。采用醇沉法、超滤膜分离法得到党参口服液中活性多糖部分,以RAW264.7巨噬细胞为模型,通过对细胞增殖指数、吞噬指数、NO释放量以及细胞因子（IL-6、IL-1β、TNF-α）的分泌量的测定,评价高压灭菌前后党参口服液中多糖的免疫活性。结果表明:超滤分离得到高压灭菌前以及高压灭菌后党参口服液中分子量小于30 k Da的多糖组分CPP-2和HCPP-2,其得率分别为22.41%和15.91%,纯度分别为69.03%和69.30%,且二者均能较好地促进巨噬细胞增殖（增殖指数=1.13¹.48）,因此CPP-2和HCPP-2是党参口服液中主要的活性多糖部分。与CPP-2相比,HCPP-2在25⁴00μg/m L浓度范围内作用于巨噬细胞的吞噬能力、TNF-α和IL-1β生成显著性降低（p<0.05）;在100⁴00μg/m L浓度范围内,NO和IL-6生成显著性降低（p<0.05）。说明高压灭菌会导致党参口服液中多糖的免疫活性降低。      

铁皮石斛多糖液态发酵工艺及其抗氧化活性研究

探讨液态发酵铁皮石斛多糖的最佳工艺及其抗氧化活性。采用单因素及正交实验研究了发酵温度、转速、p H和发酵时间对发酵工艺的影响,得到了液态发酵的最优条件:发酵温度28℃,p H4.5,转速170 r/min,发酵时间54 h;发酵后铁皮石斛水提多糖的总抗氧化能力、清除DPPH及ABTS自由基能力均有所提高,而且均具有很强的量效关系;发酵后石斛多糖的平均分子量由4.234×10~5降为1.077×10~5,推测发酵多糖抗氧化能力的提高可能与微生物发酵过程导致石斛多糖分子量的减小有关。      

轮叶党参粗多糖对体外培养小鼠脾淋巴细胞及RAW 264.7细胞的免疫活性

目的:研究轮叶党参多糖对小鼠脾淋巴细胞及巨噬细胞(RAW 264.7)作用以探究其免疫增强的功效。方法:浓度为1000,500,250,125 μg/mL多糖体外分别与脾脏细胞、RAW 264.7共培养,采用ELISA法检测细胞上清中细胞因子IL-2、IFN-γ、IL-6和TNF-α浓度的变化。采用MTT法,研究轮叶党参粗多糖对RAW 264.7增殖的影响。Griess法测定细胞上清液中NO的含量变化。通过RT-PCR法检测iNOS mRNA的生成情况。结果:与正常组相比,轮叶党参粗多糖组IL-2、IFN-γ、IL-6、TNF-α和NO的分泌量极显著增加(p<0.01),显著促进RAW 264.7增殖(p<0.05)。与正常组比较,iNOS mRNA明显增加。结论:轮叶党参粗多糖可与ConA协同增强T细胞活性,也可参与活化单核-巨噬细胞对特异性免疫与非特异性免疫活性起到正向调节的作用。     

石斛多糖的研究进展

石斛多糖作为石斛的重要功能活性物质,具有抗肿瘤、抗氧化、提高免疫力等多种功能活性。本文综述了石斛多糖的研究进展,主要从石斛多糖的提取纯化、结构解析及功能活性等方面对近年来石斛多糖研究所取得的成果进行综述。     

Health-promoting effects of dietary polysaccharide extracted from Dendrobium aphyllum on mice colon,including microbiota and immune modulation

The health-promoting effects of a polysaccharide (DAP) purified from Dendrobium aphyllum, on mice colon were investigated. The results indicated that consumption of DAP facilitated the differentiation degree of CD4⁺ T cells to Th₁, Th₁₇ and Treg cells; and depressed the differentiation degree to Th₂ cells, since DAP up-regulated IL-6 expression (from 1.08 ± 0.24 to 3.40 ± 0.36). Besides, oral administration of DAP reduced the gastrointestinal transit time from 255 ± 1.21 to 164 ± 0.46 min, decreased the pH from 6.70 ± 0.27 to 5.54 ± 0.31, improved faecal water-binding capability from 59.4% ± 0.35 to 63.5% ± 0.22. Additionally, the increase in four health-promoting short chain fatty acids were observed, which might result from the enrichment of the relative abundance of health-promoting microbiota genera, including Porphyromonadaceae and Ruminococcaceae. The DAP-induced up-regulation of the genus Erysipelotrichaceae might be related to the homeostasis in both Th₁/Th₂ and Treg/Th₁₇.     

Superfine grinding improves the bioaccessibility and antioxidant properties of Dendrobium officinale powders

In herbal markets, an overwhelming majority the D. officinale are sold in spiral or spring form, and this kind of product would lead to the incomplete release of some nutrients. To increase the bioaccessibility of D. officinale, we report a method of pulverisation treatments including general and superfine grinding to reduce the particle size. In comparison with coarse D. officinale, the superfine ground D. officinale with particle sizes of <18 μm resulted in significant higher bulk density, better flowability and greater hydration properties. The protein and polysaccharide solubility of D. officinale increased significantly as its particle size decreased. The extract of D. officinale with a particle size of <18 μm showed that highest total phenolic and total flavonoid contents accompanied with the best antioxidant activity through all antioxidant assays compared with coarse ground D. officinale and D. officinale products.     

铁皮石斛多糖对高脂饮食小鼠肠腔菌群的影响

目的 探究铁皮石斛多糖(Dendrobium officinale polysaccharides,DOP)对高脂饮食小鼠肠腔菌群的影响,为其功能性食品的开发提供参考.方法 以水提醇沉法提取DOP,分别以高脂饮食(high fat diet,HFD)、高脂饮食联合DOP对昆明小鼠干预8周,收集小鼠小肠段的内容物进行16...     

石斛多糖的研究进展

现代药理学研究表明,石斛多糖具有抗氧化、抗肿瘤、调节免疫及降低血糖等功能,其诸多应用价值引起了众多专家学者的广泛关注。文中概述了近年来国内外在石斛多糖的结构分析、分离提取以及功能作用领域的研究进展,为石斛多糖的进一步研究及其产业化应用提供依据。