南极产低温几丁质酶菌株的16S rDNA序列分析及其发酵条件及酶学性质研究

从南极深海沉积物中筛选到一株产低温几丁质酶的菌株AC444,经细菌形态观察及16S rDNA序列分析,该菌株属于假交替单胞菌属（pseudoalteromonas）,其最适与最高生长温度分别为15℃和30℃,属于兼性嗜冷菌。AC444菌株能利用多种碳源产酶,在含胶体几丁质和蛋白胨的培养基中产酶最高,达13．47U,最适产酶温度为20℃。酶的最适反应pH为7．0,最适作用温度为35℃,属中性低温酶。     

高活性几丁质酶产生菌的筛选及鉴定

从四川、陕西、浙江等地食用菌栽培的土壤样品中,筛选到一株性能优良、产几丁质酶活力较强的菌株BLC08609,并结合菌落形态、生理生化指标和16S rDNA序列分析对其进行鉴定.结果显示,菌株BLC08609能以食用菌细胞壁(以几丁质为主)为唯一碳源生长,在菌株BLC08609个体形态和生理生化特性鉴定的基础上,通过16S rDNA测序鉴定,确定BLC08609为地衣芽孢杆菌(Bacillus licheniformis).     

超声对黑曲霉产菊粉酶的影响

为了探索超声提高黑曲霉产菊粉酶的可能,考察了超声处理时间、超声频率和不同发酵时期超声处理等参数的影响。试验发现在黑曲霉的延滞期和对数增长期的初期阶段（〈30h）,40kHz和59kHz的超声处理均抑制了黑曲霉产菊粉酶的活力,高频超声的抑制效果更明显,菊粉酶活力降低了47.4%。在黑曲霉的对数增长期的后期阶段（〉36h）,这两种频率的超声处理均促进了黑曲霉产菊粉酶的活力,低频超声的促进作用更明显,菊粉酶活力提高了29.3%。结果表明,在一定条件下,超声能够促进微生物的发酵产酶。     

产蛋白酶菌株的筛选鉴定及产酶优化

为筛选出高产蛋白酶的菌株,通过干酪素培养基筛选到一株高产蛋白酶的菌株BJ-4,经鉴定该菌株为解淀粉芽孢杆菌（Bacillus amyloliquefaciens）。对菌株的产酶条件进行单因素及响应面优化,确定最佳产酶条件为：葡萄糖0.5%、牛肉膏1.5%、培养温度37.61 ℃、培养初始pH值 6.84、接种量1.36%,优化后酶活力同比增长了71.01%。菌株BJ-4在工业中具有一定的应用前景。     

豆鼓溶栓酶产生菌的筛选及其酶学性质的初步研究

利用纤维蛋白平板筛选，纤溶活性测定，SDS－PAGE分析和体外溶栓实验等相结合的方法，成功地从豆豉中筛选到一株纤溶活性高达520U／mL和具有良好体外溶栓效果的细菌菌体（编号为DC－4），经鉴定为解淀粉芽孢杆菌（Bacillus amyloliquefaciens)。SDS－PAGE和纤维蛋白自显影表明该酶的分子量为28kD，其最适反应温度和pH值分别为42℃和8．0，在pH6-10较稳定。PMSF和盐酸苯甲醚能抑制纤溶活性，而EDTA和EGTA不能抑制，表明该酶是一种丝氨酸蛋白酶。它溶解纤维蛋白的方式是直接溶解纤维蛋白，而不激活纤溶酶原，并且不水解血细胞。     

脂肪酶产生菌的筛选及酶学性质

采取含油脂泥土,通过富集培养,平板筛选,得到多种菌株,酶活测定表明一株地酶属菌株酶活较高,对其酶学性质进行了研究。     

豆豉溶栓酶产生菌的筛选及其酶学性质的初步研究

利用纤维蛋白平板筛选、纤溶活性测定、SDS PAGE分析和体外溶栓实验等相结合的方法 ,成功地从豆豉中筛选到一株纤溶活性高达 5 2 0U/mL和具有良好体外溶栓效果的细菌菌株 (编号为DC 4 ) ,经鉴定为解淀粉芽孢杆菌 (Bacillusamyloliquefaciens)。SDS PAGE和纤维蛋白自显影表明该酶的分子量为 2 8kD ,其最适反应温度和 pH值分别为 4 2℃和 8.0 ,在 pH6～ 10较稳定。PMSF和盐酸苯甲醚能抑制其纤溶活性 ,而EDTA和EGTA不能抑制 ,表明该酶是一种丝氨酸蛋白酶。它溶解纤维蛋白的方式是直接溶解纤维蛋白 ,而不激活纤溶酶原 ,并且不水解血细胞。     

甲壳胺复合物固定化酶的催化反应机理研究

本文以天然高分子甲壳胺与羧甲基纤维素钠、聚丙烯酸的两种氢键复合物为载体固定葡萄糖氧化酶和纤维素酶。对其酶促反应的米氏常数Ｋｍ进行了分析，提出了氢键复合物固定化酶的催化反应机制，还通过红外光谱和透射电镜研究了这两种复合物固定化酶的结构。     

产脂肪酶海洋嗜冷菌的鉴定及酶学性质研究

从东海的深海底泥中筛选出一株产脂肪酶的海洋细菌 EastSeaG5-1415,该菌为革兰氏阴性好氧菌,短杆状,0.9～1.3μm×1.5～3.8μm,无鞭毛,无芽孢,无色素,菌落光滑不透明,过氧化氢酶和氧化酶阳性,甲基红实验阴性,氧化葡萄糖产酸,可耐受10%的NaCl.此菌最适生长温度为18℃,最高生长温度为35℃,在0℃也能生长,是典型的嗜冷菌.菌株脂肪酸种类为C17:1(28.2%),C18:1 9c(49.7%),辅酶Q-8是其主要的异戊烯醌类,DNA中G C含量为45.2mol%.以16SrRNA同源性为基础构建了相关种属细菌在内的系统发育树,在系统发育树中,分离菌株 EastSeaG5-1415 与 Psychrobacter glacincola 在同一分支,二者的序列相似性为97.6%,DNA 杂交显示与 P.glacincola 最为相近,达到87%.结合形态和生理生化试验,将其鉴定为嗜冷杆菌 P.glacincola.同时对其所产脂肪酶的性质进行了初步研究.该菌所产碱性脂肪酶最适反应温度为35℃,在5℃时仍有较高酶活,酶活达到23%,最适 pH 值为9,属低温碱性脂肪酶.     

酶膜反应器研究进展

对酶膜反应器的国内外研究现状和发展趋势进行简要介绍。概述了酶膜反应器的特点及其分类，介绍了酶膜反应器的应用领域和一些应用实例。讨论了酶膜反应器目前存在的一些瓶颈问题及其解决办法。     

抗菌核病转基因油菜植株的获得

将携带菜豆几丁质酶基因的植物表达载体ｐＢｃｈ通过农秆菌介导法导入甘兰型油菜Ｈ１６５中。经卡那霉素对Ｔ０、Ｔ１和Ｔ２代植株进行连续的筛选和菌核病（ｓｃｌｅｒｏ－ｔｉｎｉａ ｓｃｌｅｒｏｔｉｎｏｒｉｕｍ）接种试验,共获得２２株抗菌核病的Ｔ２代植株。对其中的２２株进行ＰＣＲＳ检测,从６株扩增得到与菜豆几丁质酶基因大小对应的ＤＮＡ带型,Ｓｏｕｔｈｅｒｎ杂交证实其中２株呈现阳性结果,表明外源基因已整合到油菜     

Selectively Promoting or Preventing Osteoblast Growth on Titanium Functionalized with Polyelectrolyte Multilayers

Titanium plays an important role in medical applications, such as hip joint implants or fixation plates. These implants must perform differently depending on their clinical application. In particular, the osseointegrative properties required of the implant vary with clinical application. The present work is aimed at the functionalization of titanium surface using polyelectrolyte multilayers consisting of natural biopolymers and testing their cell adhesive properties with respect to the osseointegration capacity. Multilayered coatings were created from chitosan (Chi), hyaluronic acid (HA), and gelatine (Gel) through layer‐by‐layer deposition. Cell adhesion, proliferation, and viability were tested in vitro with the human osteoblast cell line CAL‐72 at timescales up to 7 d. Two multilayer coatings consisting of alternated chitosan/gelatin or chitosan/hyaluronic acid layers with the outmost layer of gelatin (Chi/Gel) or hyaluronic acid (Chi/HA), respectively, were tested. The experimental results showed that surfaces functionalized with Chi/Gel and Chi/HA multilayers demonstrated a good initial adhesion of osteoblasts. After 4 d culturing, osteoblast cells were almost completely detached from the substrates functionalized with Chi/HA multilayers. In contrast to Chi/HA, the proliferation of osteoblasts on substrates with Chi/Gel multilayer coatings was statistically significantly higher compared to the control titanium. We have shown that the growth of osteoblasts can be enhanced or completely prevented on a titanium surface functionalized with polyelectrolyte multilayers consisting of natural biopolymers, as desired. Both multilayer coatings, Chi/Gel and Chi/HA, have potential for applications in the field of titanium implants, where rapid osseointegration is essential, and/or where no ingrowth of the implant is desired, respectively.     

Polysaccharide surface engineering of poly(D, L-lactic acid) via electrostatic self-assembly technique and its effects on osteoblast growth behaviours

The objective of this study was to surface modify the poly (D, L-lactic acid) (PDLLA) films and assess the effects of the modified surfaces on the functions of osteoblasts cultured in vitro. A layer-by-layer (LBL) self assembly technique, was used leading to the formation of multilayers on the PDLLA film surfaces. Chitosan (Chi) and poly (styrene sulfonate, sodium salt) (PSS) were utilized as polycation and polyanion in this study, respectively. The layer structure was investigated by using X-ray photoelectron spectroscopy (XPS) and water contact angle measurement, respectively. XPS analysis displayed the presence of chitosan on PDLLA surface. A full coverage of coating with PSS/Chi layers was achieved on the PDLLA surface only after the deposition layers of PEI/(PSS/Chi)₂. These results showed that PDLLA films could be modified with PSS/Chi pairs which may affect the biocompatibility of the modified PDLLA films. To confirm this hypothesis, cell proliferation, cell viability as well as alkaline phosphtase activity of osteoblasts on layer-by-layer modified PDLLA films as well as control samples were investigated in vitro. The proliferation of osteoblasts on modified PDLLA films was found to be greater than that on control (p < 0.05 and p < 0.01) after 1, 4 and 7 days culture, respectively. Cell viability measurement showed that the PSS/Chi modified PDLLA films have higher cell viability (p < 0.01) than control. Osteoblast differentiation function (ALP) on LBL-modified PDLLA film was found significantly higher (p < 0.01) than that of virgin PDLLA films. These data suggests that PSS/Chi pair was successfully employed to surface modify PDLLA film via a layer-by-layer technique, and enhanced its cell biocompatibility.     

太极图形符号在家具设计中的创新设计研究

目的实现太极图形符号在家具设计中的创新运用。方法从设计艺术学的角度,结合实例,详细论述了太极图形特有的形态特征和深刻的思想内涵在家具设计中的应用。结论分析了太极图形符号在家具设计中的运用,归纳了家具设计的基本原则,从而启发从中国传统图形中去思考家具设计,使传统符号进行创新,以期为家具设计提供理论指导。     

Surface modification of titanium thin film with chitosan via electrostatic self-assembly technique and its influence on osteoblast growth behavior

Chitosan (Chi) and poly (styrene sulfonate) (PSS) were employed to surface modify titanium thin film via electrostatic self-assembly (ESA) technique in order to improve its biocompatibility. The surface chemistry, wettability and surface topography of the coated films with different number of deposited layers were investigated by using X-ray photoelectron spectroscopy (XPS), water contact angle measurement and atomic force microscopy (AFM), respectively. The results indicated that a full surface coverage for the outmost layer was achieved at least after deposition of five layers, i.e., PEI/(PSS/Chi)₂ on the titanium films. The formed multi-layered structure of PEI(PSS/Chi) _x (x ≥ 2) on the titanium film was stable in air at room temperature and in phosphate buffered solution (PBS) for at least 3 weeks. Cell proliferation, cell viability, DNA synthesis as well as differentiation function (alkaline phosphatase) of osteoblasts on chitosan-modified titanium film (PEI/(PSS/Chi)₆) and control sample were investigated, respectively. Osteoblasts cultured on chitosan-modified titanium film displayed a higher proliferation tendency than that of control (p < 0.01). Cell viability, alkaline phosphatase as well as DNA synthesis measurements indicated that osteoblasts on chitosan-modified titanium films were greater (p < 0.01) than those for the control, respectively. These results suggest that surface modification of titanium film was successfully achieved via deposition of PEI/(PSS/Chi) _x layers, which is useful to enhance the biocompatibility of the titanium film.     

Guanidinium-induced differential kinetic denaturation of alkaline phosphatase isozymes

This paper examines the solution kinetics of bovine intestinal and liver alkaline phosphatase (ALP) isozymes. 4-Methylumbelliferyl phosphate is used as the substrate to study the differential kinetic behavior of ALP isozymes in the presence of guanidinium hydrochloride, a denaturant. The recursive Kalman filter algorithm for parameter estimation is used for analysis of the resulting kinetic data. A two-component first-order kinetic model with a zero-order component is used to successfully quantify intestinal and liver isozymes in synthetic mixtures. This work serves as a basis for the development of an electrophoresis separation method for ALP isozyme quantification with differential kinetic detection.     

Improved estimation of hydrologic data using the chi‐square goodness‐of‐fit test

Abstract

This study considers the complete Chi‐Square goodness‐of‐fit test procedures and numerical analysis to improve the intermediate value method for an approximate solution of the cumulative distribution function. The goodness‐of‐fit test is applied for evaluating the appropriate distribution of frequency analysis for estimating annual maximum flows of hydrologic data, from four measurement stations (Lounung, Yuemei, Santimem, and Kaoping stations) located in the Kaoping River Basin. Analytical results indicate that the error percentage of the Chi‐Square statistical values is improved by from 0.3 to 38.3%. Thus, the improved estimation of the Chi‐Square goodness‐of‐fit test procedure increases the accuracy of some frequency distributions.     

Biomimetic properties of an injectable chitosan/nano-hydroxyapatite/collagen composite

To meet the challenges of designing an injectable scaffold and regenerating bone with complex three-dimensional (3D) structures, a biomimetic and injectable hydrogel scaffold based on nano-hydroxyapatite (HA), collagen (Col) and chitosan (Chi) is synthesized. The chitosan/nano-hydroxyapatite/collagen (Chi/HA/Col) solution rapidly forms a stable gel at body temperature. It shows some features of natural bone both in main composition and microstructure. The Chi/HA/Col system can be expected as a candidate for workable systemic minimally invasive scaffolds with surface properties similar to physiological bone based on scanning electron microscopic (SEM), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR) results.     

Chitosan-Drug Conjugate Microspheres: Preparation and Drug Release Properties of Microspheres Composed of the Conjugate of 2'- or 3'-(4-Carboxy-butyryl)-5- Fluorouridine with Chitosan

The conjugate microspheres (Chi-glu-FUR-m) were prepared by the dry-in-oil method using chitosan-5-fuorouridine conjugate. Chi-glu-FUR-m were characterized by drug content, particle shape and size, swelling property, and drug release. Their characteristics were compared with those of the simple microspheres (Chi/ FUR-m), which were prepared under similar conditions using a mixture of chitosan and 5-fluorouridine. Both microspheres prepared showed a high retention of the drug after preparation and similar particle size and shape. Swelling ratios after incubation in aqueous buflers of pH 7.4 for 6 hr were similar for both microspheres. Chi-glu-FUR-m swelled quickly in aqueous buffers of pH 7.4 and the disintegration was observed to occur gradually from 24 hr afrer the incubation. Chi-glu-FUR-m showed a gradual drug release (50% release time = 61 hr), while Chi/FUR-m released the drug very rapidly, Such characteristics of Chi-glu-FURm as swelling, slow disintegration, and gradual drug release propose its usefulness for localization or chemoembolization therapy.