Controlled release of urea encapsulated by starch-g-poly(L-lactide)

This paper presented a new approach for preparing a new type of slow-release membrane-encapsulated urea fertilizer with starch-g-PLLA as biodegradable carrier materials. By solution-casting and washing rapidly with water the urea was individually encapsulated within the starch matrix modified by L-lactide through in situ graft-copolymerization.

Poly(L-lysine)-graft-chitosan copolymers: Synthesis, characterization, and gene transfection effect

Polypeptide/polysaccharide graft copolymers poly(L-lysine)-graft-chitosan (PLL-g-Chi) were prepared by ring-opening polymerization (ROP) of epsilon-benzoxycarbonyl L-lysine N-carboxyanhydrides (Z-L-lysine NCA) in the presence of 6-O-triphenylmethyl chitosan. The PLL-g-Chi copolymers were thoroughly characterized by H-1 NMR, C-13 NMR, Fourier transform infrared (FT-IR), and gel permeation chromatography (GPC). The number-average degree of polymerization of PLL grafted onto the chitosan backbone could be adjusted by controlling the feed ratio of NCA to 6-O-triphenylmethyl chitosan. The particle size of the complexes formed from the copolymer and calf thymus DNA was measured by dynamic light scattering (DLS). It was found in the range of 120 similar to 340 nm. The gel retardation electrophoresis showed that the PLL-g-Chi copolymers possessed better plasmid DNA-binding ability than chitosan. The gene transfection effect in HEK 293T cells of the copolymers was evaluated, and the results showed that the gene transfection ability of the copolymer was better than that of chitosan and was dependent on the PLL grafting ratio. The PLL-g-Chi copolymers could be used as effective gene delivery vectors.

Purification and structure identification of hyaluronic acid

Polysaccharide produced by mutated strain of Streptococcus zooepidemicus was purified by the procedures including Savage method, quaternary ammonium compound precipitation, DEAE-cellulose(DE52) chromatography and Sephadex G-75 gel filtration. The structure of the purified polysaccharide has been characterized by means of chemical composition analysis, C-13 NMR spectrum, infrared spectrum and circular dichroism (CD). All the results showed that the purified polysaccharide was hyaluronic acid (HA). The single helix conformation of the purified HA was determined by Congo red experiment. The molecular weight of the HA was about 1.16x10(6)D, which was measured by viscosity method.

Synthesis of chitosan-stabilized gold nanoparticles in the absence/presence of tripolyphosphate

Gold nanoparticles were prepared by reducing gold salt with a polysaccharide, chitosan, in the absence/ presence of tripolyphosphate (TPP). Here, chitosan acted as a reducing/stabilizing agent. The obtained gold nanoparticles were characterized with UV-vis spectroscopy and transmission electron microscopy. The results indicated that the shape and size distribution of gold nanoparticles changed with the molecular weight and concentration of chitosan. More interestingly, the gelation of chitosan upon contacting with polyanion (TPP) can also affect the shape and size distribution of gold nanoparticles. By adding TPP to chitosan solution before the reduction of gold salt, gold nanoparticles have a bimodal size distribution, and at the same time, polygonal gold particles were obtained in addition to spherical gold nanoparticles.

Synthesis and characterization of the biodegradable polycaprolactone-graft-chitosan amphiphilic copolymers

A novel synthetic approach to biodegradable amphiphilic copolymers based on poly (epsilon-caprolactone) (PCL) and chitosan was presented, and the prepared copolymers were used to prepare nanoparticles successfully. The PCL-graft-chitosan copolymers were synthesized by coupling the hydroxyl end-groups on preformed PCL chains and the amino groups present on 6-O-triphenylmethyl chitosan and by removing the protective 6-O-triphenylmethyl groups in acidic aqueous solution. The PCL content in the copolymers can be controlled in the range of 10-90 wt %. The graft copolymers were thoroughly characterized by H-1 NAM, C-13 NMR, FT-IR and DSC. The nanoparticles made from the graft copolymers were investigated by H-1 NMR, DLS, AFM and SEM measurements. It was found that the copolymers could form spherical or elliptic nanoparticles it? water. The amount of available primary amines on the surface of the prepared nanoparticles was evaluated by ninhydrin assail, and it can be controlled by the grafting degree of PCL.

Medicated wound dressings based on poly(vinyl alcohol)/poly(N-vinyl pyrrolidone)/chitosan hydrogels

Poly(vinyl alcohol) /poly(N-vinyl pyrrolidone) (PVP)/chitosan hydrogels were prepared by a low-temperature treatment and subsequent Co-60 -gamma-ray irradiation and then were medicated with ciprofloxacin lactate (an antibiotic) and chitosan oligomer (molecular weight = 3000 g/mol). The gel content, swelling ratio, tensile strength, and crystallinity of the hydrogels were determined. The effects of the chitosan molecular weight, the low-temperature treatment procedure, and the radiation dosage on the hydrogel properties were examined. The molecular weight of chitosan was lowered by the irradiation, but its basic polysaccharide structure was not destroyed. Repeating the low-temperature treatment and gamma-ray irradiation caused effective physical crosslinking and chemical crosslinking, respectively, and contributed to the mechanical strength of the final hydrogels. The incorporation of PVP and chitosan resulted in a significant improvement in the equilibrium swelling ratio. and elongation ratio of the prepared hydrogels. The ciprofloxacin lactate and chitosan oligomer were soaked into the hydrogels. Their in vitro release behaviors were examined, and they were found to follow diffusion-controlled kinetics.

Chemical characteristic of bioactive polysaccharides isolated from Ornithogalum caudatum Ait.

A water-soluble crude extract prepared from Ornithogalum caudatum Ait. (OCA) showing a high immunomodulating activitiy was isolated and characterized by virtue of get filtration and column chromatography. The presence of the monosaccharides has been established by the chemical analysis. The quantitative analysis of the alditol acetate derivatives of them showed the ratios of the monosaccharides analyzed by means of GC respectively. The concentrations of protein(280 nm) and carbohydrate (496 nm) were detected respectively. The information of the molecular weight from the pure polysaccharide was obtained by several standard Dextrans from the Sephadex chromatography.

Arabinogalactan as a carrier for contrast agent in magnetic resonance imaging

Arabinogalactan-Gd-DTPA was synthesized by the reaction of diethylenetriaminepenta-acetic acid (DTPA) bisanhydride with polysaccharide in dry DMSO and characterized by FTIR, elemental analysis and ICP-AES. Its stability was investigated by competition with Ca2+, EDTA, DTPA. The t(1)-relaxivity is 8.06 mmol(-1) . L . s(-1) in D2O, 8.48 mmol(-1) . L . s(-1) in 0.725 mmol . L-1 BSA, respectively. t(1)-weighted MR imaging of rat kidney and liver showed a remarkable enhancement post injection of Arabinogalactan-Gd-DTPA. The results indicate that the arabinogalactan-Gd-DTPA is a potential contrast agent for MRI.

CDNA cloning and mRNA expression of the lipopolysaccharide- and beta-1,3-glucan-binding protein gene from scallop Chlamys farreri

Lipopolysaccharide and beta-1,3-glucan-binding protein (LGBP) play a crucial role in the innate immune response of invertebrates as a pattern recognition protein (PRP). The scallop LGBP gene was obtained from Chlamys farreri challenged by Vibrio anguillarum by randomly sequencing cDNA clones from a whole body cDNA library, and by fully sequencing a clone with homology to known LGBP genes. The scallop LGBP consisted of 1876 nucleotides with a canonical polyadenylation signal sequence AATAAA and a poly(A) tail, encoding a polypeptide of 440 amino acids with the estimated molecular mass of 47.16 kDa and a predicted isoelectric point of 5.095. The deduced amino acid sequence showed a high similarity to that of invertebrate recognition proteins from blue shrimp, black tiger shrimp, mosquito, freshwater crayfish, earthworms, and sea urchins, with conserved features including a potential polysaccharide-binding motif, a glucanase motif, and N-glycosylation sites. The temporal expression of LGBP genes in healthy and V. anguillarum-challenged C farreri scallop, measured by real-time semiquantitative reverse transcription polymerase chain reaction (PCR), showed that expression was up-regulated initially, followed by recovery as the stimulation cleared. Results indicated that scallop LGBP was a constitutive and inducible acute-phase protein that could play a critical role in scallop-pathogen interaction. (C) 2004 Elsevier B.V. All rights reserved.

羟基自由基降解琼胶及其产物的活性研究

琼胶是一种从石花菜等红藻中提取的，目前生产工艺和结构等方面研究比较成熟的海藻多糖，广泛应用于医药、仪器等行业。但是，海藻多糖因为具有分子量大，粘度大，溶解度较小的等特点，而使其应用范围受到限制。利用降解的手段对其进行修饰，降低分子量和粘度，改善溶解性，可以拓展其应用范围。并且根据文献报道，琼 胶寡糖具有一些特殊的生物活性，如抗氧化性，抗炎症等。因此，对琼胶降解的研究具有生要意义。本研究中，为了选择一种合适的降解方法，进行了几种水解方法的尝试，其中包括在不同湿度和酸度下盐酸水解，过氧化氢和醋酸催化水解，Fenton体系羟基自由基降解。对于酸水解和Fenton体系氧化还原降解方法，通过粘度法对反应的速度进行了比较，表明氧化还原降解反应中琼胶的粘度降低比较快，并且具有代表性和新意，确定为本实验的降解琼胶的方法并对氧化还原降解所得的产物进行了活性实验。通过模仿自然界普遍存在的氧化还原降解反应，利用Vc诱导的Fenton体系产生的羟基自由基氧化还原降解琼胶得到低分子量的琼胶。降解产物经过高速离心、60％乙醇沉淀，除去分子量比较大的降解产物和磷酸盐，得到可溶于60％乙醇的分子量估计小于3000的降争产物，其产率为85％。利用经Sephadex-G25凝胶色谱分离所香的不同分子量的级分进行分子量和α－葡萄糖苷酶抑制活性关系的实验。降解产物对α－葡萄糖苷酶的抑制率和各级分的浓度呈线性正相关，并且各级分的IC_(50)则随着分子量的降低而降低。另外，对所得的降解产物混合物进行了红外吸收光谱、质子去偶核磁共震碳谱和负离子基质辅助激光诱导－飞行时间质谱结构分析。结果表明，氧化还原降解反应的专一性差，在得到寡糖的同时，在光谱图中出现一些比较复杂的副产物的结构信息。最后，根据MTT法的原理，以有体皮肤成纤维细胞为材料，通过紫外线辐射产生自由基造成氧化损伤，研究降解产物对成纤维细胞的保护作用。当无紫外线辐射时，降解产物对成纤维细胞具有显著的促进生长增殖作用：当经UVa、UBb辐射时则可以显著地表现出对损伤的保护作用，并且这种促进生长和保护作用呈显著的量效关系，表明降解产物具有清除基自由基的作用。但是，因为氧化还原降解以应的机理尚不十分明的以及琼羟胶的特殊结构，使得反应的副产物很难预测，也就使得分离工作难以进行，所以，根据目前所得的信息，尚不能确定是降解产物的什么级分产生的以上两种生物活性。

海洋藻类功能性蛋白质的纯化、鉴定及其应用的研究

使用膨化柱和离子交换或羟基磷灰石柱层析相结合的方法，分别从多管藻、坛紫菜及钝顶螺旋藻中分离纯化了R-藻红蛋白溶液和C-藻蓝蛋白。光谱检测及电泳分析结果证明完全符合经典的藻胆蛋白纯度标准。彭化床最突出的优点是克服了常规分离方法堵塞色谱柱的难题，纯化速度快、产量高、不需要常规色谱方法所要求的填料的平衡及粗提液的预处理，仅需一步操作就可以得到满足一般食品添加剂纯度要求的藻胆蛋白，极大地简化了后续的纯化程序，减少了分离纯化的步骤和时间，而其产率及纯度均高于常规的藻胆蛋白分离方法。这同时也降低了藻胆蛋白分离纯化的成本。 本文通过戊二醛或环氧氯丙烷交联的方法，合成了四种壳聚糖-氨基酸共聚小球。并选取吸附性好的戊二醛交联孔球和戊二醛交联微球系统测定了其对R-藻红蛋白和C-藻蓝蛋白的吸附和缓释性能。 纯化了藓羽藻中与其细胞器团聚密切相关的一种凝集素并进行了部分性质的鉴定。N端前15个氨基酸序列及LC-ESI-MS质谱分析结果证明此凝集素属于一种新的蛋白质族。实验证明，凝血活性与细胞器团聚活性并不完全依赖于此凝集素分子相同的结构域。 通过异双功能试剂SPDP处理藓羽藻凝集素使之衍生化，DTT处理R-PE在其分子内引入外源巯基，然后将活化的R-藻红蛋白与凝集素进行交联反应。交联产物经凝胶过滤纯化并检测，但电泳及荧光显微镜检测结果并不能证明交联探针的成功制备。

注射用低分子量褐藻多糖硫酸酯的制备及理化性质研究

褐藻多糖硫酸酯（Fucoidan Polysaccharide of Sulfated, FPS）是褐藻细胞间的一类硫酸化多糖，组成复杂，在不同褐藻中变化很大，典型的结构是岩藻多糖硫酸酯，也有由甘露糖、半乳糖、糖醛酸、木糖、葡萄糖、褐藻糖等多种单糖组成的结构复杂的杂多糖。研究发现FPS具有抗凝血、降血脂、抗肿瘤和抗HIV的作用，成为当今天然药物主攻的重点之一。由中国科学院海洋研究所所主持研究的海洋新药——褐藻多糖硫酸酯，用于临床治疗肾病综合症和中、早期肾衰具有很好的疗效。本论文研究的注射用褐藻多糖硫酸酯是用过氧化氢-抗坏血酸（1：1）体系降解所得的低分子量FPS，并用琼脂糖凝胶（DEAE）色谱柱进行分级制得，在治疗肾脏疾病方面，具有意想不到的良好效果。 本论文模拟植物生长发育过程中的氧化还原反应，分别用抗坏血酸-过氧化氢和Fe2+-过氧化氢-抗坏血酸两种体系降解FPS。各降解试剂的比例和浓度是影响降解效果的主要因素。分别用抗坏血酸-过氧化氢体系和Fe2+-过氧化氢-抗坏血酸体系降解所得样品的硫酸根、褐藻糖和糖醛酸含量与原料多糖无明显差别，红外光谱与原料多糖一致，表明多糖在降解过程中化学组成和结构并未发生改变。而所得样品多分散指数在1.49-1.77之间，表明用此法能获得分子量分布范围较窄的低分子量多糖。 本论文建立了柱前衍生化高效液相色谱法分析褐藻多糖硫酸酯中的单糖组成，将多糖样品用2 mol•L-1三氟乙酸降解成单糖后，以核糖为内标，用1-苯基-3-甲基-5-吡唑啉酮（PMP）进行柱前衍生化，优化分析条件，采用反相高效液相色谱，245nm紫外检测和使用梯度洗脱，分离测定各单糖。方法学验证实验，在浓度0.10 mmol•L-1～2.0 mmol•L-1各单糖衍生物都有良好的线性关系（r2≥0.999）；除葡萄糖醛酸外，各单糖衍生物都有很好的重现性和稳定性；甘露糖、鼠李糖、葡萄糖醛酸、葡萄糖、半乳糖、木糖、褐藻糖平均加样回收率分别为86.2%，95.1%，62.5%，102.0%，94.8%，66.6%，105.1%。此法可以用于FPS中褐藻糖、半乳糖、葡萄糖主要单糖进行含量测定。 本论文对注射用褐藻多糖硫酸酯的理化性质进行研究，包括性状、鉴别、检查和含量测定,同时做了原料药的强制降解实验，为原料药质量标准的制定和制 剂研究提供了依据。

孔石莼多糖及其衍生物的研究

在中国，孔石莼应用于医药方面已有几千年的历史，其煎剂常被当地居民用来治疗中暑和泌尿方面的疾病。实验室以前所做的工作表明，孔石莼的水溶多糖具有很好的抗高血脂活性，本论文在此研究的基础上主要就多糖的长期毒性、一般药理、多糖的衍生化、不同分子量孔石莼多糖的抗氧化活性、衍生物的抗氧化活性和动物调血脂活性进行了研究。 水提醇沉制备多糖，多糖主要由糖醛酸、鼠李糖、木糖、葡萄糖和硫酸根组成，还含有微量的半乳糖、甘露糖和阿拉伯糖。多糖中主要的二糖重复单位为[β-D-Glcp A-(1->4)- α-L-Rhap 3S] 和 [α-L-Idop A-(1->4)- α-L-Rhap 3S]。急性毒性实验表明，KM小鼠对孔石莼多糖的最大耐受量(MTD)大于4000 mg/kg，其腹腔注射雌性小鼠的半致死量(LD50)为408.7 mg/kg，雄性动物为432.7 mg/kg。长期毒性（6个月）实验表明孔石莼多糖无毒反应剂量为1.2 g/kg。一般药理研究表明孔石莼多糖对小鼠中枢神经系统无明显影响，对麻醉犬心血管系统和呼吸系统均无明显影响。 采用过氧化氢降解的方法制备了不同分子量的孔石莼多糖，并且测定了体外抗氧化活性；制备了高硫酸根含量的孔石莼多糖、乙酰化和苯甲酰化孔石莼多糖衍生物，测定了体外抗氧化活性和动物调血脂试验。结果表明，不同分子量的孔石莼多糖其抗氧化活性是不同的，分子量低的孔石莼多糖表现出了较强的抗氧化活性。孔石莼多糖的衍生物其抗氧化活性要优于孔石莼多糖。调血脂动物试验表明，孔石莼多糖以及其衍生物都具有很好的调血脂效果。高硫酸根含量孔石莼多糖的中、低剂量组的调血脂活性要优于高剂量组（低剂量组降低低密度脂蛋白的能力要稍弱于原料），而且，中剂量组与原料组相比，小鼠血清TG明显降低（P＜0.05），LDL－C 明显降低（P＜0.01）。低剂量组乙酰化衍生物降低甘油三酯（TG）和低密度脂蛋白（LDL－C）的能力要优于中高剂量组。对于孔石莼多糖以及衍生物调血脂的机制还需进一步的研究。

Preparation and in vitro antioxidant activity of kappa-carrageenan oligosaccharides and their oversulfated, acetylated, and phosphorylated derivatives

In order to study the relationship between chemical structure and properties of modified carrageenans versus antioxidant activity in vitro, K-carrageenan oligosaccharides were prepared through mild hydrochloric acid hydrolysis of the polysaccharide, and these were used as starting materials for the partial synthesis of their oversulfated, acetylated, and phosphorylated derivatives. The structure and substitution pattern of the oligosaccharides and their derivatives were Studied using FTIR and C-13 NMR spectroscopy, and their in vitro antioxidant activities were investigated. Certain derivatives of the carrageenan oligosaccharides exhibited higher antioxidant activity than the polysaccharides and oligosaccharides in certain antioxidant systems. The oversulfated and acetylated derivatives, which scavenge superoxide radicals, the phosphorylated and low-DS acetylated derivatives, which scavenge hydroxyl radicals, and the phosphorylated derivatives, which scavenge DPPH radicals, all exhibited significant antioxidant activities it, the systems examined. The effect of the molecular weight of the carrageenan on antioxidant activities, however, is not obvious from these studies. (c) 2005 Elsevier Ltd. All rights reserved.

Study on cytocompatibility of carboxymethyl-chitosan membranes to skin fibroblasts

Chitosan and carboxymethl-chitosan (CM-chitosan) membranes with different molecular mass were prepared by a casting method. The cytocompatibility of two kinds of polysaccharide membranes to skin fibroblasts that cultured in vitro were studied. The methods were to culture the cells in soaking fluid of membranes and to culture the cells on the membranes directly. The results showed that the soaking fluid had no toxicity to fibroblasts and the biological security of lower molecular mass membranes were better than higher molecular mass membranes, and CM-chitosan membranes were better than chitosan membranes. In addition, the growth of fibroblasts on chitosan membranes was inhibited and the cells would fall off from chitosan membranes after a period of culture. However, the cells adhered and expanded well on CM-chitosan membranes. All these demonstrated that cytocompatibility of CM-chitosan membranes to skin fibroblasts was better than chitosan membranes.