Production and isolation of azaspiracid-1 and -2 from Azadinium spinosum culture in pilot scale photobioreactors

Azaspiracid (AZA) poisoning has been reported following consumption of contaminated shellfish, and is of human health concern. Hence, it is important to have sustainable amounts of the causative toxins available for toxicological studies and for instrument calibration in monitoring programs, without having to rely on natural toxin events. Continuous pilot scale culturing was carried out to evaluate the feasibility of AZA production using Azadinium spinosum cultures. Algae were harvested using tangential flow filtration or continuous centrifugation. AZAs were extracted using solid phase extraction (SPE) procedures, and subsequently purified. When coupling two stirred photobioreactors in series, cell concentrations reached 190,000 and 210,000 cell · mL(-1) at steady state in bioreactors 1 and 2, respectively. The AZA cell quota decreased as the dilution rate increased from 0.15 to 0.3 day(-1), with optimum toxin production at 0.25 day(-1). After optimization, SPE procedures allowed for the recovery of 79 ± 9% of AZAs. The preparative isolation procedure previously developed for shellfish was optimized for algal extracts, such that only four steps were necessary to obtain purified AZA1 and -2. A purification efficiency of more than 70% was achieved, and isolation from 1200 L of culture yielded 9.3 mg of AZA1 and 2.2 mg of AZA2 of >95% purity. This work demonstrated the feasibility of sustainably producing AZA1 and -2 from A. spinosum cultures.     

Separation of fine organic particles by a low-pressure hydrocyclone (LPH)

The separation performance of a low-pressure hydrocyclone was tested using fine organic particles from 1 to 700 μm. The dimensions of the low-pressure hydrocyclone were an inflow diameter of 30 mm, a cylinder length of 575 mm, an overflow diameter of 60 mm, an underflow diameter of 50 mm, a cylinder diameter of 335 mm and a cone angle of 68°. The low-pressure hydrocyclone was operated with a lower inlet pressure (average 1.38–5.56 kPa) that could be maintained under water level differences that ranged from 17.5 to 53.5 cm between the water surface of the feeding mass cylinder and the middle of the inlet pipe of the low-pressure hydrocyclone. By varying the inflow rate, underflow ratio and feed concentration, the separation performance of the low-pressure hydrocyclone was affected. The separation performances were determined from total separation efficiency and grade efficiency. Separation performances were determined according to the different inflow rates of 400, 600, 800 and 1000 ml s⁻¹ and their respective underflow ratios that ranged from 5% to 30%. The maximum total separation efficiencies for each inflow rate were 41%, 46% and 46% at 400, 800 and 1000 ml s⁻¹ inflow rates, respectively, and at underflow rates of 30% of the inflow rates. In addition, a total separation efficiency of 46% was employed at 600 ml s⁻¹ of inflow rate and with an underflow rate of 25% its inflow rate. As the feed concentration increased from 25 to 150 mg l⁻¹, the separation performances were gradually decreased. For the fine particles ranging 1–200 μm, the grade efficiency was higher at the higher inflow rate (higher than 600 ml s⁻¹) and higher underflow rate. However, for the coarse particles ranging 400–700 μm, the grade efficiency was higher at the lower inflow rate (lower than 600 ml s⁻¹) and higher underflow rate. The cut-point (d₅₀) values ranged from 30 to 200 μm for a feed size range of 1–700 μm. The Response Surface Method (RSM) model predicted an optimum operating inflow rate and underflow ratio of 721 ml s⁻¹ of inflow rate and 30%, respectively, for the low-pressure hydrocyclone at a maximum total separation efficiency. Based on these findings, further design and operating adaptation of low-pressure hydrocyclones used for fine solids removal in recirculating aquaculture systems is expected.     

Effects of Cement Additives on Properties of CSA Cement from MSWI

Municipal solid waste incineration fly ash (MSWI) was successfully used as raw material in sintering and preparing a calcium sulphoaluminate (CSA) cement clinker in laboratory. The effects of different types and different addition percentages of cement additives on compressive strength and hydration properties were investigated. The results showed that lime (LI) powder / slag (SL) powder was compatible in CSA cement system while the activities of fly ash(FA)/ MSWI was low. Adding these four types of cement additives in CSA cement system respectively, there were negative effects on compressive strengths of hardened cement at early age while LI/SL improved the compressive strength at later age. The performance of combined additives was better, especially, the specimens with 10%LI+10%SL, 10%LI+10%MSWI, and 5%LI+15%SL.     

枯草芽孢杆菌固体发酵豆粕条件的优化

本试验采用枯草芽孢杆菌对豆粕进行固体发酵,通过正交试验探讨不同碳源组合、发酵时间、培养温度、水分比例、接种菌量等因素对发酵豆粕粗蛋白质增加率和蛋白质水解度的影响。结果表明,影响枯草芽孢杆菌固体发酵豆粕粗蛋白质增加率的条件因素的主次顺序为玉米粉比例>培养时间>温度>麸皮比例>水分比例>次粉比例>接种菌量,表观分析固体发酵条件最佳组合为A₃B₁C₃D₃E₂F₂G₁,粗蛋白质增加率达到28.05%;影响发酵豆粕蛋白质水解率的条件因素的主次顺序为温度>培养时间>玉米粉比例>接种菌量>麸皮比例>次粉比例>水分比例,表观分析固体发酵条件最佳组合为A₂B₃C₁D₃E₃F₃G₁,蛋白质水解度达到46.46%。枯草芽孢杆菌固体发酵豆粕的粗蛋白质增加率与蛋白质水解率存在明显的正相关(R²=0.556)。     

防治青枯病工程菌Hrp-菌株的固体剂型研究

本文对工程菌 Hrp-菌株的固体剂型进行了初步研究。经载体、保护剂和抑菌剂的筛选,最终得出固体菌剂的制作方法。具体为将发酵菌液以5000 r/min离心10 min后弃上清,收集菌泥;将菌泥加至石蜡油中,振荡15～20 min使其在石蜡油中充分分散,制成菌悬液;再将此菌液按1：1比例与灭菌后的草炭混合均匀,制成固体菌剂;再向这种固体菌剂中加入杂菌抑制剂乳酸链球菌素（0.75 g/kg）和纳他霉素（0.075 g/kg）,并混合均匀。制作的固体菌剂在pH值5.3左右、含水量35%、不抽真空及温度25℃的条件下保存6个月,存活率为44.4%,有效活菌量为1.12×1010 cfu/g。     

基于表面引发原子转移自由基聚合技术制备2,4-二氯苯氧乙酸分子印迹聚合物及其识别特性分析

首次以氯甲基化交联聚苯乙烯树脂(CMCPS)为载体和大分子引发剂、2,4-二氯苯氧乙酸(2,4-D)为模板、丙烯酰胺(AM)为单体、乙二醇二甲基丙烯酸酯(EDMA)为交联剂、溴化铜/2,2'-联吡啶(CuBr/Bpy)为催化剂,采用表面引发原子转移自由基聚合(SI-ATRP)技术,制备了2,4-二氯苯氧乙酸分子印迹聚合物(2,4-D MIPs),并研究了模板分子与功能单体比例对该印迹聚合物吸附量的影响。通过动态、静态及竞争试验考察了该印迹聚合物对2,4-D的吸附性能。结果表明:2,4-D MIPs对模板分子2,4-D具有良好的特异性识别作用;与2,4-二氯苯酚和2,4-二氯苯甲醛相比,2,4-D MIPs对2,4-D的选择性系数分别为2.84和3.75,相对选择性系数分别为2.31和2.29。采用Scatchard模型分析,可以得到两类结合位点,计算得到最大表观吸附量(Qmax)分别为76.92和142.91 mg/g,离解常数Kd分别为632.91和2 309.47 mg/L。将2,4-D MIPs作为固相萃取剂,对豆芽样品进行添加回收试验,回收率为86%~104%,相对标准偏差(RSD)为1.9%~10%,方法的检出限为20 ng/g。该印迹聚合物可以富集分离测定2,4-D,稳定性好,并且能重复使用。     

整体硬质合金螺旋木工铣刀的合理选用

从刀具旋向特点、加工方式、切削参数以及刀具磨损等方面介绍整体硬质合金螺旋木工铣刀的应用和选择。     

微生物固体发酵提取杜仲胶工艺参数的筛选

【目的】探讨菌株D-5在不同条件下对杜仲种皮固体发酵时羧甲基纤维素(CMC)酶、果胶酶活力及纤维类物质去除率的变化规律,筛选利用微生物固体发酵提取杜仲胶的发酵工艺参数。【方法】以曲霉D-5为发酵菌株,研究发酵基质不同的含水率、初始pH、起爆剂用量、发酵温度对CMC酶、果胶酶活力及纤维类物质去除率的影响。【结果】发酵基质的含水率、初始pH、起爆剂用量、发酵温度对菌株D-5的酶活力及纤维类物质的去除率有显著的影响。菌株D-5发酵的适宜条件为:基质含水率为60%、初始pH6.0、发酵温度30℃、起爆剂用量1.5%,在此条件下发酵6dCMC酶及果胶酶的活力最高,发酵18d纤维类物质去除率可达86.43%。【结论】利用微生物发酵去除纤维类物质是杜仲胶提取的重要途径之一,菌株D-5在杜仲胶提取中有一定的应用价值。     

生防放线菌153固态发酵条件的优化及其耐热力检测

【目的】探讨生防放线菌153的固态发酵条件,并考察其耐热力,为生防放线菌的大规模生产及活体菌剂的研制提供技术支持。【方法】将发酵物的含菌量作为筛选指标,以玉米粉、甘油、蔗糖、葡糖糖和可溶性淀粉为碳源,以小米粉、蛋白胨、酵母浸粉、黄豆粉和尿素为氮源,通过单因素试验获得适宜的碳、氮源,再采用L25(56)正交设计优化固体培养基配方,在此基础上确定固态发酵终点以及所需的接种量和种子液种龄。最后将壳聚糖、非耕作层黄土、高岭土、炉渣、碳酸钙、面粉和几丁质分别与生防放线菌153菌粉混合,55℃水浴至致死中时,测定生防放线菌153的耐热力,筛选适宜的载体。【结果】生防放线菌153固体培养基配方为:80mg/g麦麸、8mg/g玉米粉、10mg/g小米粉和2mg/g蛋白胨,初始含水量(含0.5g/L石灰的自来水)为60%,在此条件下培养,培养物的含菌量较对照增加了94.1%。生防放线菌153固态发酵培养终点为216h,接种量为20%(每100g固态培养基接种20mL种子液),种子液种龄为84h。生防菌153的适宜载体为壳聚糖和非耕作层黄土。【结论】确定了生防放线菌153的固态发酵条件,该条件可以促进生防菌153的生长产孢,并提高菌株的保存稳定性;确定了筛选生放菌153载体的方法,即55℃条件下水浴处理136min。     

固相萃取—高效液相色谱法测定兔血浆中山奈素的含量

兔的血浆样品经过β-葡萄糖醛酸酶和硫酸酯酶酶解、提取、C18固相萃取小柱富集净化后,在Agilent SB-C18柱(250mm×4.6 mm,5μm)上,以甲醇-0.4%磷酸(55∶45)为流动相,流速为1.0 mL.min-1,检测波长为360 nm,对山奈素进行测定的结果表明,血浆样品的最佳酶解条件为:β-葡萄糖醛酸酶和硫酸酯酶的终用量分别为400和20 U.mL-1,酶解时间为2h;血浆中山奈素含量的线性范围为0.019-0.608μg.mL-1,方法回收率为80.23%-84.27%,日内和日间精密度的RSD分别≤7.32%、10.17%.可见,本试验建立的高效液相色谱测定方法可以准确、灵敏地测定兔血浆中山奈素的含量,适用于山奈素血药含量检测和药代动力学研究.