洋葱假单胞菌乳糖酸发酵条件的优化

以选育的洋葱假单胞菌NTG-15-03为生产菌株,通过单因素和回归正交设计试验考察菌株种龄、接种量、发酵时间、发酵液初始pH值对乳糖酸产量的影响。结果表明：在pH7.0、种龄24h、接种量2%、发酵时间106h的条件下,该菌株的乳糖酸产量为10.08g/L。     

高效产脂肪酶菌株Burkholderia cepacia C1产酶条件优化

从油菜地土壤中分离到一株高效产脂肪酶菌株C1,经鉴定为Burkholderia cepacia。为了进一步提高C1 脂肪酶的产量,对其产酶的发酵条件进行优化。首先采用单因子试验筛选出最佳碳源为麸皮,最佳氮源为蛋白胨。通过Plackett-Burman 设计对发酵产酶的11 个相关因子进行试验,筛选出3 个主效因子,即蛋白胨质量浓度、橄榄油质量浓度及装液量。利用Box-Behnken 试验设计和响应曲面法分析确定主效因子的最优水平,得出产脂肪酶菌株C1 的最优产酶条件：1.50g/100mL 麸皮、1.05g/100mL 蛋白胨、1.63g/100mL 橄榄油、0.2g/100mL K2HPO4、0.05g/100mL MgSO4、初始pH 值为7.0、装液量为30mL。在优化条件下30℃,180r/min 培养72h,脂肪酶活力达到89.65U/mL,比未优化前的28.50U/mL 提高了2.15 倍。     

Speciation of Pb(II) sorbed by Burkholderia cepacia/goethite composites

Bacterial-mineral composites are important in the retention of heavy metals such as Pb due to their large sorption capacity under a wide range of environmental conditions. However, the partitioning of heavy metals between components in such composites is not probed directly. Using Burkholderia cepacia biofilms coated with goethite (alpha-FeOOH) particles, the partitioning of Pb(II) between the biological and iron-(oxyhydr)oxide surfaces has been measured using an X-ray spectroscopic approach. EXAFS spectra were fit to quantitatively determine the fraction of Pb(II) associated with each component as a function of pH and [Pb]. At pH < 5.5, at least 50% of the total sorbed Pb(II) is associated with the biofilm component, whereas the total uptake within the composite is dominated by goethite (> 70% Pb/goethite) above pH 6. Direct comparison can be made between the amount of Pb(II) bound to each component in the composite vs separate binary systems (i.e., Pb/biofilm or Pb/goethite). At high pH, Pb(II) uptake on the biofilm is dramatically decreased due to competition with the goethite surface. In contrast, Pb uptake on goethite is significantly enhanced at low pH (2-fold increase at pH 5) compared to systems with no complexing ligands. The mode of Pb(II)-binding to the goethite component changes from low to high [Pb]. Structural fitting of the EXAFS spectra collected from 10(-5.6) to 10(-3.6) M [Pb]eq at pH 6 shows that the Pb-goethite surface complexes at low [Pb] are dominated by inner-sphere bidentate, binuclear complexes bridging two adjacent singly coordinated surface oxygens, giving rise to Pb-Fe distances of approximately 3.9 A. At high [Pb], the dominant Pb(II) inner-sphere complexes on the goethite surface shift to bidentate edge-sharing complexes with Pb-Fe distances of approximately 3.3 A.     

A psychrotrophic Burkholderia cepacia strain isolated from refrigerated raw milk showing proteolytic activity and adhesion to stainless steel

The proteolytic activity of a psychrotrophic strain of Burkholderia cepacia isolated from refrigerated raw milk was characterized. Bur. cepacia produced proteolytic activity during growth at refrigeration temperature, with maximum activity at pH 6-7. The enzyme showed relative thermal stability in the range 40-50°C during 25 min, and maintained 80% its initial activity at 76°C/30 s. Milk coagulation assay showed that the crude protease from Bur. cepacia caused coagulation from day 2 for skimmed milk, whereas coagulation was observed from day 5 for whole milk. The adherence of this strain to stainless steel was evaluated, and the substrata had around 107 CFU/cm2 after 15 to 60 min incubation. Results on biofilm development suggest that this bacterium could adhere and to form biofilms even at refrigeration temperatures. These results indicate that Bur. cepacia may represent a potential hazardous to milk and dairy products.     

Burkholderia cepacia中具有解脂作用的胆固醇酯酶的酶学特性

为了更好地应用于工业领域,系统研究Burkholderia cepacia ZWS15胆固醇酯酶(EC 3. 1. 1. 13 cholesterol esterase,CHE)的酶学特性。通过DEAE离子交换从该菌发酵液中获得一种CHE,并探讨其有机溶剂与表面活性剂耐受性,以及底物水解特性。CHE在p H 5. 5～9. 0保持稳定,最适反应温度为40℃,在70℃温育2 h仍能保留70%以上的酶活,具有显著耐热性;在50%(体积分数)有机溶剂或5%(体积分数)表面活性剂存在下也具有较高酶活。飞行时间质谱鉴定该酶分子质量为37 k Da,其与胆固醇酯酶(源于洋葱伯尔霍尔德菌,Burkholderia cepacia)及脂肪酶(源于假单胞菌属,Pseudomonas sp.)具有较高匹配度。与商品化酶相比,CHE不仅具有胆固醇酯酶活性(对长链胆固醇酯类底物有明显偏好性),还具有解脂功能(可作用于三酰基甘油酯及对硝基苯酯)。该酶的优良性能可为其在食品、诊断、制浆造纸等领域的工业应用提供参考与借鉴。     

Enhancing activity and stability of Burkholderia cepacia lipase by immobilization on surface-functionalized mesoporous silicates

Burkholderia cepacia lipase was immobilized on various types of phenyl-functionalized mesoporous silicates (MPS). MPS, anchored with a phenyl group on the silica wall and with three dimensional (3D) mesoporosity, showed highest lipase adsorption capacity and best activities both in aqueous and organic reagents.     

Development of a time-temperature integrator system using Burkholderia cepacia lipase

A time-temperature integrator (TTI) is a device used to show a time-temperature dependent change that reflects the temperature history and quality status of the food to which it is attached. An enzymatic TTI system based on the reaction between Burkholderia cepacia lipase and tricaprylin, which causes a pH change, was developed. The temperature dependence of the response rate of this new lipase-type TTI was modeled using the Arrhenius equation, and the estimated activation energy was calculated as 70.61±11.10 kJ/mol (±95% confidence interval). The TTI response was validated under dynamic storage conditions with independent variable temperature experiments, and a good agreement was obtained between the predicted and measured values.     

Recovery of Burkholderia pseudomallei and B. cepacia from drinking water

Samples of drinking water were examined in order to evaluate the occurrence of two gram-negative bacteria: Burkholderia pseudomallei and B. cepacia. A total of 85 samples were collected from public and private buildings in the province of Bologna (Italy). Other bacteriological indicators (heterotrophic plate count at 22 and 36 degrees C) were also examined, together with physical and chemical parameters (temperature, pH, residual chlorine, total hardness and chemical oxygen demand (COD)). High levels of B. pseudomallei were recovered (mean value = 578 cfu/100 ml) in about 7% of samples, while B. cepacia was recovered in 3.5% (mean value = < 1) of the samples. The two microorganisms were found to correlate positively with heterotrophic plate counts at 22 and 36 degrees C, but not with the physical and chemical parameters taken into consideration.     

Sorption versus biomineralization of Pb(II) within Burkholderia cepacia biofilms

X-ray spectroscopy measurements have been combined with macroscopic uptake data and transmission electron microscopy (TEM) results to show that Pb(II) uptake by Burkholderia cepacia is due to simultaneous sorption and biomineralization processes. X-ray microprobe mapping of B. cepacia biofilms formed on alpha-Al2O3 surfaces shows that Pb(II) is distributed heterogeneously throughout the biofilms because of the formation of Pb "hot spots". EXAFS data and TEM observations show that the enhanced Pb accumulation is due to the formation of nanoscale crystals of pyromorphite (Pb5(PO4)3(OH)) adjacent to the outer-membrane of a fraction of the total population of B. cepacia cells. In contrast, B. cepacia cell suspensions or biofilms that were heat-killed or pretreated with X-rays do not form pyromorphite, which suggests that metabolic activity is required. Precipitation of pyromorphite occurs over several orders of magnitude in [H-] and [Pb] and accounts for approximately 90% of the total Pb uptake below pH 4.5 but only 45-60% at near-neutral pH because of the formation of additional Pb(II) adsorption complexes. Structural fits of Pb L(III) EXAFS data collected for heat-treated cells at near-neutral pH suggest that Pb(II) forms inner-sphere adsorption complexes with carboxyl functional groups in the biofilms.     

Oxidation of aromatic aldehyde to aromatic carboxylic acid by Burkholderia cepacia TM1 isolated from humus

Burkholderia cepacia TM1 isolated from humus was able to oxidize aromatic aldehydes to the corresponding aromatic carboxylic acids with an extreme by high yield in distilled water containing only the aromatic aldehyde as the substrate. The molar yields of vanillic acid from vanillin, p-hydroxybenzoic acid from p-hydroxybenzaldehyde, and syringic acid from syringaldehyde were approximately 94%, 92% and 72%, respectively. Until the added aromatic aldehyde was significantly consumed, consumption of the produced acid hardly occurred. The findings of this study indicate that an appropriate reactor residence time for continuous production can be set, and that aromatic carboxylic acid can be produced efficiently and continuously from the corresponding aldehyde.     

洋葱伯克霍尔德氏菌Lu10-1脂肪酶的异源表达

通过PCR扩增出洋葱伯克霍尔德氏菌Lu10-1脂肪酶基因,将基因片段分别克隆到大肠杆菌、毕赤酵母和枯草杆菌的表达载体,转入表达菌株中。结果表明,重组脂肪酶在大肠杆菌中主要以包涵体形式表达,在毕赤酵母中未有表达,而在枯草杆菌中实现了胞外分泌表达,测得发酵上清液酶活为13.8 U/m L。对重组枯草杆菌发酵条件进行了摇瓶初步优化和3 L的反应器分批培养。当以TB为出发培养基,初始p H 6.5,温度为37℃时,在3 L的发酵罐上最终酶活达到34.5 U/m L,是野生菌表达量的4.2倍。     

Continuous oxidation of aromatic aldehyde to aromatic carboxylic acid by Burkholderia cepacia TM1 in a cell-holding reactor

The continuous oxidation of aromatic aldehydes to the corresponding aromatic carboxylic acids was performed. In the continuous oxidation of aromatic aldehydes by Burkholderia cepacia TM1 in a cell-holding reactor, the concentration of the aromatic aldehyde in the reactor was kept extremely low by appropriately adjusting the initial turbidity of the cells in the reactor, the feeding rate of the aromatic aldehyde to the reactor and the average residence time. Thus, the feeding concentration at the inlet of vanillin, p-hydroxy-benzaldehyde, or syringaldehyde was 20.0, 20.0, or 4.0 g/l, respectively. Under the indicated reaction conditions, the steady state of the reaction continued for approximately 650 h, 450 h, and 160 h, respectively, for vanillin, p-hydroxybenzaldehyde and syringaldehyde as the substrate. The molar yield of vanillic acid, p-hydroxybenzoic acid and syringic acid in the steady state was, respectively, approximately 95, 80 and 96%, and the productivity was, respectively, 0.770, 0.350 and 0.160 (g/l.h). Moreover, since the recovered reaction solution consisted almost predominantly of only one type of aromatic carboxylic acid, separation and purification of the product was considered to be unnecessary. To prevent a decrease in the pH of the reaction solution and to maintain a high solubility of the substrate and the product, a phosphate buffer (pH 7.2) is better than distilled water as a reaction solution for feeding.     

洋葱伯克霍尔德氏菌L68邻苯二酚2,3-双加氧酶基因克隆、表达和纯化

采用自行设计的引物,从洋葱伯克霍尔德氏菌L68中PCR扩增得到phnE（邻苯二酚2,3-双加氧酶）基因,亚克隆到高表达载体pET 32a上,转入表达菌株中.经双向测序,证实构建过程中未出现突变.重组子经IPTG诱导后,可以表达有活性的重组双加氧酶.选择26 ℃进行重组蛋白诱导.薄层扫描显示,表达重组蛋白总量最高可占总蛋白的64.92%.利用金属螯合层析对重组蛋白进行了一步纯化,SDS-PAGE结果显示,重组蛋白纯度达到95%.     

基于BP神经网络的洋葱伯克霍尔德菌脂肪酶发酵软测量建模

为建立洋葱伯克霍尔德菌(Burkholderia cepacia)脂肪酶发酵过程的软测量模型,运用BP神经网络对洋葱伯克霍尔德菌脂肪酶的发酵过程进行软测量建模,并利用遗传算法对神经网络的初始权值和阈值进行优化,实现模型加快收敛速度,达到全局最优解效果.该模型能够比较精确地模拟菌体生长、底物消耗以及发酵产酶的过程动态,具有良好的泛化能力,说明BP神经网络结合遗传算法在洋葱伯克霍尔德茵脂肪酶发酵过程的模拟与预测中是一种高效快速的方法.     

Factors affecting patulin production by Penicillium expansum

Patulin, a mycotoxin produced by Penicillium spp. during fruit spoilage, is a major concern with regard to human health because exposure can result in severe acute and chronic toxicity, including carcinogenic, mutagenic, and teratogenic effects. In this study, we investigated the effects of Penicillium expansum isolate, apple cultivar, storage temperature and time, and pH on the production of patulin. Patulin was analyzed by a previously developed micellar electrokinetic capillary electrophoresis method. P. expansum isolates originating from across Ontario produced widely differing levels of patulin, ranging from 0 to >6 mg/g by dry mycelial weight. The highest patulin levels were those for isolates displaying aggressive growth (characterized by rapidly increasing acidity) accompanied by profuse mycelial development. Distinct patterns in fungal growth rates and patulin production were evident among isolates grown in McIntosh, Empire, and Mutsu ciders. Extensive fungal growth and higher patulin levels (538 to 1,822 microg/ml on day 14) in apple ciders were associated with incubation at room temperature (25 degrees C), although potentially toxic patulin levels (75 to 396 microg/ml on day 24) were also found in refrigerated ciders (4 degrees C) inoculated with P. expansum.     

脂肪酶位置选择性的影响因素

为开发高效、绿色、低成本的脂肪酶应用于结构脂质的制备，从内源性(脂肪酶的来源、基因序列、空间位阻)和外源性(底物空间结构、反应介质、固定化条件及反应pH)两方面讨论了脂肪酶位置选择性的影响因素。脂肪酶催化特异性由内源性因素和外源性因素共同决定。改变底物空间结构、反应介质、固定化条件和反应pH,常被用作提高脂肪酶催化性能的处理方式。只有充分考虑内外两方面因素的影响，才能充分提高脂肪酶的位置选择性和催化效率。该综述有望为高选择性脂肪酶的开发开辟新的研究方向，为后续结构脂质的高效合成提供启发性的研究思路。     

Factors limiting the biodegradation of Ulva sp cell-wall polysaccharides

The dietary fibres of the seaweed Ulva sp (sea-lettuce) consist of water-soluble ulvan, alkali-soluble β(1,4)-D -glucuronan and β(1,4)-D -glucoxylan, and an insoluble α-cellulose containing xylose residues. They are poorly degraded by human colonic bacteria particularly when associated within the intact plant cell wall. In order to better understand this resistance to microbial attack, their organisation in the cell-wall has been investigated by light and electron microscopy after sequential chemical extractions. Their susceptibility to enzymatic degradation and their accessibility to bacteria and enzyme were also studied. Microscopic localisation in native and sequentially extracted Ulva sp demonstrated that ulvan is in all the cell-walls of the algae and particularly between the two cell layers constituting the thallus. Glucuronan is close to the cytoplasmic membrane facing the outside of the seaweed and between adjacent cells. The xylose and glucose containing polysaccharides form packed layers surrounding the cells. A model of the spatial distribution of the different polysaccharides within the algae is proposed. Ulvan and glucuronan did not limit the xyloglucan and α-cellulose degradation by an endo-xylanase in the whole seaweed and its insoluble dietary fibre but the α-cellulose was not affected by a cellulase. The cell-wall of Ulva sp was accessible to enzymes but poorly to bacteria as assessed from porosity measurements. These results established that the poor fermentation of sea-lettuce by human colonic flora is primarily due to the ubiquitous presence of the degradation-resistant ulvan in the cell wall of Ulva sp. ©1997 SCI     

圆网制版感光工艺的影响因素

探讨圆网制版感光工艺中曝光过程的各种因素，如曝光时间与光源、光敏材料的特性、用量、暗反应，及其涂敷时胶层的厚度及湿度等对感光制版的影响；结合实际生产，阐述感光制版的应用原理。     

影响单酶法制备核桃多肽的因素研究

为寻求核桃多肽的最佳制备方法,以酶法水解核桃蛋白为基础,通过单因素和正交试验对核桃多肽制备工艺进行优化。结果表明,在酶解时间3 h、酶解温度38 ℃、pH 6.8、酶添加量4%、底物浓度为2.4%时,核桃多肽的抗氧化活性最佳,此时DPPH清除率可达92.35%。     

乳酸菌发酵代谢合成叶酸的影响因素

对嗜酸乳杆菌以及乳酸乳球菌发酵合成叶酸的影响因素进行了研究。结果表明,乳酸菌代谢合成叶酸的产率为17~100μg/L,菌种、培养时间、pH值、对氨基苯甲酸(PABA)质量浓度会影响乳酸菌合成叶酸的产量。与乳酸乳球菌乳酸亚种相比,嗜酸乳杆菌CH-2生成的叶酸产量要高。不同菌株生成叶酸的能力与pH值有关,嗜酸乳杆菌在pH值为4.2叶酸产率明显下降,乳酸乳球菌乳酸亚种产叶酸的能力则不受pH值影响。添加PABA可以显著提高乳酸菌的叶酸产率。选择适宜的乳酸菌菌株,优化发酵工艺参数可以提高乳及相关食品中叶酸的质量浓度,达到生物方式强化叶酸的效果。