匐枝马尾藻的化学成分研究

首次报道从中国北海匐枝马尾藻Sargassum polycystcum分离得到3个纯化合物 ，经过MS，IR，~1H NMR，~(13)C NMR（DEPT），HMQC和HMBC等波谱技术鉴定为： 豆甾-3β-羟基-5，23，25-三烯(1)，3-二下五烷氧基-1，2-丙二醇(2)，24ξ-甲 基胆甾-3β，5α，6β，25-四醇-25-乙酸酯(3)，其中化合物1为新化合物。     

羊栖菜多糖的提取和分离

羊栖菜多糖是从马尾藻科植物羊栖菜（Sargassum fusiforme(Harv.)Setchel)中提取到的，具有抗肿瘤，抗凝血，降血糖，提高机体免疫功能等生理活性，本文研究用水提法提取羊栖菜多糖，考虑到提取温度，时间，加水量等因素的影响，优化提取条件，确定水提法的最佳温度80℃，提取时间3h，水加量为羊栖菜样品的3倍，并采用乙醇沉淀，透析及凝胶层析等方法对羊栖菜提取物中多糖进行沉淀，提取和分离获得4个组分，并用光谱扫描法，凝胶色谱和旋光度法鉴定其纯度，进一步研究其分离获得的组分对生物体免疫功能指标的影响。     

Simultaneous analysis of ten phytohormones in Sargassum horneri by high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry

Phytohormones have attracted wide attention due to their important biological functions. However, their detection is still a challenge because of their complex composition, low abundance and diverse sources. In this study, a novel method of high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry was developed and validated for the simultaneous determination of ten phytohormones including indole‐3‐acetic acid, isopentenyladenine, isopentenyl adenosine, trans‐zeatin riboside, zeatin, strigolactones, abscisic acid, salicylic acid, gibberellin A3, and jasmonic acid in Sargassum horneri (S. horneri). The phytohormones were extracted from freeze‐dried S. horneri with methanol/water/methanoic acid (15:4:1, v/v/v) analyzed on a Hypersil Gold C₁₈ column and detected by electrospray ionization tandem triple quadrupole mass spectrometry in the multiple reaction monitoring mode. The experimental conditions for the extraction and analysis of phytohormones were optimized and validated in terms of reproducibility, linearity, sensitivity, recovery, accuracy, and stability. Distributions of the phytohormones in the stems, blades, and gas bladder of the S. horneri in drift, fixed, and semi‐fixed growing states were investigated for the first time. The observed contents of the phytohormones in S. horneri range from not detected to 5066.67 ng/g (fresh weight). Most phytohormones are distributed mainly in the stems of S. horneri in drift and semi‐fixed states.     

Bioassay‐guided isolation of an active compound with protein tyrosine phosphatase 1B inhibitory activity from Sargassum fusiforme by high‐speed counter‐current chromatography

A rapid and efficient method using high‐speed counter‐current chromatography was established for the bioassay‐guided separation of an active compound with protein tyrosine phosphatase 1B inhibitory activity from Sargassum fusiforme. Under the bioassay guidance, the ethyl acetate extract with the best IC₅₀ value of 0.37 ± 0.07 μg/mL exhibited a potential protein tyrosine phosphatase 1B inhibitory activity, which was further separated by high‐speed counter‐current chromatography. The separation was performed with a two‐phase solvent system composed of n‐hexane/methanol/water (5:4:1, v/v). As a result, dibutyl phthalate (19.7 mg) with the purity of 95.3% was obtained from 200 mg of the ethyl acetate extract. Its IC₅₀ was 14.05 ± 0.06 μM, which was further explained by molecular docking. The result of molecular docking showed that dibutyl phthalate enfolded in the catalytic site of protein tyrosine phosphatase 1B. The main force between dibutyl phthalate and protein tyrosine phosphatase 1B was the hydrogen bond interaction with Gln266. In addition, hydrogen bond, van der Waals force and hydrophobic interaction with the amino acids (Ala217, Ile219, and Gly220) were also responsible for the stable protein‐ligand complex.     

Sargassum fusiforme Polysaccharide-Based Hydrogel Microspheres Enhance Crystal Violet Dye Adsorption Properties

In this study, a polysaccharide-based hydrogel microsphere (SFP/SA) was prepared using S. fusiforme polysaccharide (SFP) and sodium alginate (SA). Fourier transform infrared spectroscopy (FT-IR) demonstrated that SFP was effectively loaded onto the hydrogel microsphere. Texture profile analysis (TPA) and differential scanning calorimetry (DSC) showed that, with the increase of SFP concentration, the hardness of SFP/SA decreased, while the springiness and cohesiveness of SFP/SA increased, and the thermal stability of SFP/SA improved. The equilibrium adsorption capacity of SFP/SA increased from 8.20 mg/g (without SFP) to 67.95 mg/g (SFP accounted 80%) without swelling, and from 35.05 mg/g (without SFP) to 81.98 mg/g (SFP accounted 80%) after 24 h swelling. The adsorption of crystal violet (CV) dye by SFP/SA followed pseudo-first order and pseudo-second order kinetics (both with R² > 0.99). The diffusion of intraparticle in CV dye was not the only influencing factor. Moreover, the adsorption of CV dye for SFP/SA (SFP accounted 60%) fit the Langmuir and Temkin isotherm models. SFP/SA exhibited good regenerative adsorption capacity. Its adsorption rate remained at > 97% at the 10th consecutive cycle while SFP accounted for 80%. The results showed that the addition of Sargassum fusiforme polysaccharide could increase the springiness, cohesiveness and thermal stability of the hydrogel microsphere, as well as improve the adsorption capacity of crystal violet dye.     

Preparation of New Sargassum fusiforme Polysaccharide Long-Chain Alkyl Group Nanomicelles and Their Antiviral Properties against ALV-J

Avian leukosis virus subgroup J (ALV-J) is an immunosuppressive virus which has caused heavy losses to the poultry breeding industry. Currently, there is no effective medicine to treat this virus. In our previous experiments, the low-molecular-weight Sargassum fusiforme polysaccharide (SFP) was proven to possess antiviral activity against ALV-J, but its function was limited to the virus adsorption stage. In order to improve the antiviral activity of the SFP, in this study, three new SFP long-chain alkyl group nanomicelles (SFP-C12M, SFP-C14M and SFP-C16M) were prepared. The nanomicelles were characterized according to their physical and chemical properties. The nanomicelles were characterized by particle size, zeta potential, polydispersity index, critical micelle concentration and morphology. The results showed the particle sizes of the three nanomicelles were all approximately 200 nm and SFP-C14M and SFP-C16M were more stable than SFP-C12M. The newly prepared nanomicelles exhibited a better anti-ALV-J activity than the SFP, with SFP-C16M exhibiting the best antiviral effects in both the virus adsorption stage and the replication stage. The results of the giant unilamellar vesicle exposure experiment demonstrated that the new virucidal effect of the nanomicelles might be caused by damage to the phospholipid membrane of ALV-J. This study provides a potential idea for ALV-J prevention and development of other antiviral drugs.     

Determination of amino acids in Sargassum fusiforme by high performance capillary electrophoresis

High performance capillary electrophoresis (HPCE) was developed for quantitative determination of 18 phenylthiohydantoin (PTH)-amino acids. The influence of electrolyte concentration, pH, organic modifier and applied voltage on HPCE performance was investigated. The HPCE separation of a PTH-amino acids mixture was much improved by adding organic modifier and Tris-boric acid buffer to the run buffer. After optimization of the method, 17 PTH-amino acids in a solution containing 18 PTH-amino acids could be separated using 400 mmol l⁻¹ Tris-boric acid, 1.0 mmol l⁻¹ diethylamine at pH 9.5 adjusted with 0.1 mol l⁻¹ NaOH as a run buffer, voltage of 25 kV was applied, temperature was maintained at 25 °C, detection wavelength was 254 nm. The precision (n = 7) of this method is less than 3.2% (peak area) and 1.1% (migration time) of relative standard deviation (R.S.D.). Linearity was established over the concentration range 50-1000 μM of each derivative, with correlation coefficients (r) ranging between 0.9904 and 0.9993. The detection limits (S/N = 3) range from 2 to 48 μmol l⁻¹. The method was applied to determine amino acids in Sargassum fusiforme, a marine algae collected from Tongtou County of Zhejiang Province in China with satisfactory results.     

高效毛细管电泳分析羊栖菜多糖对肿瘤机体红细胞合淌度的影响

为了分析羊栖菜多糖（SFPS）对肿瘤机体红细胞合淌度的影响,建立了肿瘤动物模型,分高、中、低剂量腹腔给予羊栖菜多糖7 d,采集并制备红细胞悬液,应用高效毛细管电泳法检测红细胞的合淌度。实验条件:毛细管为75 μm×50 cm,电泳缓冲液为含2 g/L羟丙基甲基纤维素的磷酸盐溶液(0.1 mol/L,pH 7.4),压力进样为3.448 kPa×10 s,分离电压为20 kV,柱温为25 ℃,电渗淌度为2.16×10-4 cm2·V-1·s-1。实验结果表明：羊栖菜多糖能降低肿瘤机体红细胞的电泳迁移时间(阴性对照组为15.76 min,SFPS低剂量组为13.96 min,SFPS中剂量组为12.90 min,SFPS高剂量组为13.51 min,正常对照组为11.51 min),增加红细胞的合淌度(阴性对照组为1.06×10-4 cm2·V-1·s-1,SFPS低剂量组为1.19×10-4 cm2·V-1·s-1,SFPS中剂量组为1.29×10-4 cm2·V-1·s-1,SFPS高剂量组为1.23×10-4 cm2·V-1·s-1,正常对照组为1.45×10-4 cm2·V-1·s-1),SFPS 3个剂量组红细胞的合淌度与阴性对照组比较均有非常显著的差异(P＜0.01)。羊栖菜多糖能够改变肿瘤机体红细胞的合淌度,并使之趋向于正常机体水平,这可能与其改变红细胞表面的电荷密度有关。高效毛细管电泳法可以作为检测红细胞生理状态和功能的一种辅助工具。     

半叶马尾藻粗糖中单糖的离子色谱法分析

以60～80 ℃的水从半叶马尾藻干粉中提取粗多糖,用Sevage溶剂去蛋白纯化后,将粗多糖用4.0 g/L 的三氟乙酸在 80 ℃下水解,水解液在CarboPacTM PA10离子色谱柱(2 mm i.d.×250 mm)上以14.0 mmol/L NaOH溶液为流动相进行 分离,以电化学检测器检测半叶马尾藻粗糖水解产生的单糖成分及含量。结果表明,半叶马尾藻粗糖中木糖、半乳糖、阿 拉伯糖、葡萄糖、鼠李糖和果糖的含量分别为2200,820,98,4560,358和740 mg/kg,加标回收率范围为86.0%~108.0%,检出限范围为5.6～89.6 μg/kg。该方法具有灵敏度高、精密度好、样品不需 要衍生化处理等优点,适合藻类样品中单糖的分析。     

超高效液相色谱串联质谱法同时测定马尾藻中4种内源性植物激素

建立了利用超高效液相串联质谱法(UPLC-MS/MS)同时测定马尾藻中4种植物激素(吲哚乙酸、吲哚丁酸、脱落酸、玉米素)的方法.马尾藻样品经过70％甲醇提取后,经PCX+PAX固相小柱净化,使用反相C18色谱柱分离.以5 mmol/L乙酸铵(含0.01％甲酸)和甲醇作为流动相进行梯度洗脱,采用多反应离子监测模式(MRM)分析测定.在0.01 ～ 1.0 μg/mL内,各种植物激素的相关系数均大于0.9990.4种植物激素的回收率为84.3％～102.1％,相对标准偏差为1.1％～6.4％.本方法的检出限为0.025～0.2μg/kg.