59 resultados para Symbiotic fungus
Resumo:
Photosynthesis by phytoplankton cells in aquatic environments contributes to more than 40% of the global primary production (Behrenfeld et al., 2006). Within the euphotic zone (down to 1% of surface photosynthetically active radiation [PAR]), cells are exposed not only to PAR (400-700 nm) but also to UV radiation (UVR; 280-400 nm) that can penetrate to considerable depths (Hargreaves, 2003). In contrast to PAR, which is energizing to photosynthesis, UVR is usually regarded as a stressor (Hader, 2003) and suggested to affect CO2-concentrating mechanisms in phytoplankton (Beardall et al., 2002). Solar UVR is known to reduce photosynthetic rates (Steemann Nielsen, 1964; Helbling et al., 2003), and damage cellular components such as D1 proteins (Sass et al., 1997) and DNA molecules (Buma et al., 2003). It can also decrease the growth (Villafane et al., 2003) and alter the rate of nutrient uptake (Fauchot et al., 2000) and the fatty acid composition (Goes et al., 1994) of phytoplankton. Recently, it has been found that natural levels of UVR can alter the morphology of the cyanobacterium Arthrospira (Spirulina) platensis (Wu et al., 2005b). On the other hand, positive effects of UVR, especially of UV- A (315-400 nm), have also been reported. UV- A enhances carbon fixation of phytoplankton under reduced (Nilawati et al., 1997; Barbieri et al., 2002) or fast-fluctuating (Helbling et al., 2003) solar irradiance and allows photorepair of UV- B-induced DNA damage (Buma et al., 2003). Furthermore, the presence of UV-A resulted in higher biomass production of A. platensis as compared to that under PAR alone (Wu et al., 2005a). Energy of UVR absorbed by the diatom Pseudo-nitzschia multiseries was found to cause fluorescence (Orellana et al., 2004). In addition, fluorescent pigments in corals and their algal symbiont are known to absorb UVR and play positive roles for the symbiotic photosynthesis and photoprotection (Schlichter et al., 1986; Salih et al., 2000). However, despite the positive effects that solar UVR may have on aquatic photosynthetic organisms, there is no direct evidence to what extent and howUVR per se is utilized by phytoplankton. In addition, estimations of aquatic biological production have been carried out in incubations considering only PAR (i. e. using UV-opaque vials made of glass or polycarbonate; Donk et al., 2001) without UVR being considered (Hein and Sand-Jensen, 1997; Schippers and Lurling, 2004). Here, we have found that UVR can act as an additional source of energy for photosynthesis in tropical marine phytoplankton, though it occasionally causes photoinhibition at high PAR levels. While UVR is usually thought of as damaging, our results indicate that UVR can enhance primary production of phytoplankton. Therefore, oceanic carbon fixation estimates may be underestimated by a large percentage if UVR is not taken into account.
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The early life-history of Chinese rock carp Procypris rabaudi was investigated during a 56-day rearing period: 318 artificially propagated P. rabaudi larvae were reared throughout metamorphosis in a small-scale recirculation system (345 L water volume, 10 x 18 L rearing tanks, 150 L storage and filter compartment with bioballs, 20-30 larvae L-1) at the Institute of Hydrobiology, Wuhan, Hubei Province, China. The newly hatched larvae had an initial total length of 8.93 +/- 0.35 mm SD (n = 10) at 3 days post-hatch and reached an average total length of 33.29 mm (+/- 1.88 mm SD, n = 10) 56 days after hatching. Length increment averaged 0.45 mm day(-1), resulting in a mean growth of 24.4 mm within the 56-day period. High mortality rates of up to 92% derived from an introduced fungus infection and subsequent treatment stress with malachite green. Our results indicate that Chinese rock carp can be raised successfully from artificially fertilized eggs. We therefore assume this species to be a candidate for commercial aquaculture.
Resumo:
Interspecific symbiotic relationships involve a complex network of interactions, and understanding their outcome requires quantification of the costs and benefits to both partners. We experimentally investigated the costs and benefits in the relationship between European bitterling fish (Rhodeus sericeus) and freshwater mussels that are used by R. sericeus for oviposition. This relationship has hitherto been thought mutualistic, on the premise that R. sericeus use mussels as foster parents of their embryos while mussels use R. sericeus as hosts for their larvae. We demonstrate that R. sericeus is a parasite of European mussels, because it (i) avoids the cost of infection by mussel larvae and (ii) imposes a direct cost on mussels. Our experiments also indicate a potential coevolutionary arms race between bitterling fishes and their mussel hosts; the outcome of this relationship may differ between Asia, the centre of distribution of bitterling fishes, and Europe where they have recently invaded.
Resumo:
非豆科树木共生固氮体系抗逆性强,具有耐干旱、耐盐碱、耐低PH、耐水湿等特点,且分布广泛,资源丰富,因此开展Frankia-非豆科树木联合共生体系的理论与应用研究,不仅对丰富和发展联合共生理论具有重要意义,而且在加速树木生长、解决农村资源短缺、减少水土流失、改良土壤和改善生态环境等方面将产生重大的经济和生态环境效益.该文通过对非豆科固氮树木色赤杨的菌根、根瘤联合共生体人工构建技术以及赤杨联合共生增效作用的研究,选择出色赤杨的最适共生菌根菌、弗兰克氏菌,为联合共生体接种技术在造林生产实践尤其是在逆境条件下的应用提供理论依据.
Resumo:
本实验表明:外生菌根真菌彩色豆马勃、劣味乳菇、丝膜菌对PH的适应范围较广,最适生长BH呈酸性。模拟酸雨对马尾松幼苗菌根的外部形态和内部结构有明显影响。在温室栽培中,模拟酸雨(PH2.0)显著抑制菌根侵染率,在田间实验中,对菌根侵染率有一定的影响。菌根PH和土壤PH值随模拟酸雨PH下降而逐渐降低,接种菌根菌可略提高菌根PH和土壤PH值。菌根真菌过氧化氢酶对培养基中PH的变化不敏感,模拟酸雨对菌根过氧化氢酶活性影响也不明显。但沙培中,模拟酸雨(PH2.0)显著抑制菌根过氧化氢酶活性。模拟酸雨(PH2.0)显著刺激菌根过氧化物酶活性,接种菌根菌可以降低菌根过氧化物酶活性。不同PH的培养基对菌体硝酸还原酶活性有明显影响,而且菌体生长速度与硝酸还原酶活性呈正相关。模拟酸雨(PH2.0)显著抑制菌根硝酸还原酶活性,而接种菌根菌明显提高根系硝酸还原酶活性。菌体酸性磷酸酶活性对培养基中PH变化不敏感,同样菌根酸性磷酸酶活性对模拟酸雨的影响也不明显,但是接种菌根菌可明显提高根系酸性磷酸酶活性。模拟酸雨对马尾松幼苗茎的高生长影响不显著。但是对幼苗茎、根系的干重和侧根总长度有显著抑制作用。轻度酸雨(PH4.5-3.0)对马尾松幼苗生长有促进作用,接种菌可提高幼苗生长。从菌根形态结构和生理活性上看,接种菌根菌可减轻模拟酸雨对马尾松幼苗根系的危害,增强对模拟酸雨的抗性。4dThe result of experiment showed that ectomycorrhizal fungi Pisolithus tinctorins. Lactarius insulsus. Cortinarius russus can be growth in broad PH rang in pure culture, the optimum growth PH is acidity. The external morphology and internal structure of ectomycorrhiza of P. massoniana are affected with simulated acid rain. In greenhouse, simulated acid rain (PH2.0) treatment caused significant decrease in the percent infection, but it's not marked in field. The PH of mycorrhizal and soil are reduced with reducing rainfall PH. These PH are slight higher for inoculation with ectomycorrhizal fungi. Catalase activity of ectomycorrhizal fungus is not sensitive to medium with different PH. Mycorrhiza catalase activiyt is not affected significantly with simulated acid rain, but it's inhibited significantly with simulated acid rain (PH2.0) in the sand culture. Peroxidase atcivity of mycorrhiza is enhanced significantly with simulated acid rain (PH2.0), but it's universally lower for inoculation with ectomycorrhizal fungus. Ectomycorrhizal fungus nitrate reductase activity is affected significantly to medium with differdnt PH, the rates of these fungi growth and nitrate reductase activity is significant correlation. Nitrate reductase activity of mycorrhiza is inhibited significantly with simulated acid rain (PH 2.0), but it's increased significantly for inocnlation with mycorrhizal fungi. Ectomycorrhizal fungas acid phosphatase activity is not affected to medium with different PH, Mycorrhiza acid phosphatase activity is not affected with simulated acid rain too, the acid phosphatase activity of roots inoculated with mycorrhizal fungas is increased significantly. The highest acidity level simulated rain reduced signhficantly root system biomass and the dry weight of stem. Iower acidity level simulated rain can stimulated the growth of P. massoniana, the growth of seedling inocnlated with mycorrhizal fungus can be increased.
Resumo:
毛壳霉属(Chaetomium)和曲霉属(Aspergillus)真菌产生多种具有生物活性的化合物。为系统阐明两属微生物的次生代谢物,对三种毛壳霉、两种曲霉真菌分别进行固态发酵,以色谱和波谱技术研究发酵物中的成分,分离鉴定了51个化合物,其中23个为新化合物,测试了部分化合物对肿瘤细胞的活性。 1、从螺卷毛壳霉(C. cochloides)固态发酵物中分离鉴定了11个化合物,3个新化合物为螺卷毛壳霉素A~C(1~3)。化合物1、3及dethio-tetra (methylthio) chetomin(4)对Bre-04、Lu-04和N-04细胞株生长抑制的GI50值为0.05~7.0 μg/mL。 2、从印度毛壳霉(C. indicum)固态发酵物中鉴定的三个异喹啉生物碱印度毛壳霉素A~C(12~14)代表两类骨架新颖的异喹啉生物碱。 3、从巴西毛壳霉(C. brasiliense)固态发酵物中鉴定了11个化合物,其中Mollicellins I~J(15~16)、2-Hydroxymethyl-6-methylmethyleugenin(19)为新化合物。化合物16和Mollicellin H(18)对Bre-04、Lu-04、N-04细胞株生长抑制的GI50值在2.5~8.6 μg/mL。 4、从土曲霉(A. terreus)固态发酵物中鉴定了18个化合物。5个为新化合物为Terretonin A~D(24~27)和Asterrelenin(28),24~27为二倍半萜化合物,28为吲哚生物碱。 5、从杂色曲霉(A. versicolor)固态发酵物中鉴定了16个化合物。9个新的化合物Brevianamides K~N (40~43)、Averins A~C (44~46)和Glyanphenines A~B (47~48)代表三种类型的生物碱。 6、综述了1997-2007年间新的二倍半萜的研究进展。 The fungi of the genera Chaetomium and Aspergillus produce various secondary metabolites with biological activities. In order to systematically study the secondary metabolites, the solid-state fermented rice culture of three species of Chaetomium and two of Aspergillus were chemically studied. By the means of chromatograhy and spectroscopy, 55 compounds were isolated and identified, among of them 23 were new ones. The biological activities of some compounds were investigated. 1. From the fungus C. cochliodes, three new epipolythiodioxopiperazines, chaetocochins A-C (1-3) were isolated, together with 8 known ones (4-11). Compounds 1, 3 and 4 showed growth inhibitory effects against cancer cell lines Bre-04, Lu-04 and N-04 with GI50 values from 0.05 to 7.0 μg/mL. 2. Three novel isoquinolines Chaetoindicins A-C (12-14) were isolated and identified from the fungus C. indicum. Chaetoindicin A, Chaetoindicins B-C represented two classes of novel carbon skeletons. 3. Three new compounds, Mollicellins I-J (15-16), and 2-hydroxymethyl-6-methylmethyleugenin (19), were isolated from C. brasiliense. Compound 16 and Mollicellin H (18) showed growth inhibitory effects against cancer cell lines Bre-04, Lu-04 and N-04 with GI50 values from 2.5 to 8.6 μg/mL. 4. Eighteen compounds were isolated from the fungus A. terreus. Terretonin A-D(24 - 27)and Asterrelenin(28) are new compounds belonging to sesterterpoids and indole-ralated alkaloid, respectively. 5. From the fungus A. versicolor, sixteen secondary metabolites, including nine new ones, Brevianamides K-N (40-43), Averins A-C (44-46), and Glyanphenines A-B (47-48), were isolated and identified. Brevianamides K-N (40-43), Averins A-C (44-46), and Glyanphenines A-B (47-48) represented three classes of alkaloids. 6. New sesterterpenes and their bioactivities reported from 1997 to 2007 were summarized.
Resumo:
对15株白腐真菌进行了以玉米秸秆为基质的初步筛选,从中获得一株选择性系数较高的菌株Y10,并对其降解玉米秸秆的情况进行了研究。结果表明,在30天的培养过程中菌株Y10对玉米秸秆降解的选择性系数都大于1,第15天选择性系数最高为3.88。对未经降解和降解过的玉米秸秆分别作了紫外光谱和红外光谱分析,结果表明,经该菌降解后玉米秸秆的化学成分发生了很大变化,且木质素的降解程度要大于纤维素的降解程度。对菌株Y10进行了ITS-5.8S rDNA序列鉴定,初步判定其为Cerrena sp.。 为了考查不同的外源添加物对菌株Y10降解玉米秸秆的影响,在以玉米秸秆为基质的固态发酵培养基中分别添加了7种金属离子、8种碳源、6种氮源。结果显示,这7种金属离子均能促进木质素的降解,并且一定浓度的某些离子明显抑制纤维素的降解;其中添加0.036%的MnSO4·H2O和0.36%的MgSO4·7H2O对纤维素降解的抑制作用比较强,降解率分别为0.96%和1.31%,木质素的选择性系数分别达到了34.40和20.17。8种碳源中除麦芽糖外都能促进木质素的降解,除微晶纤维素外都明显促进纤维素的降解。6种氮源中酒石酸铵、硫酸铵、草酸铵和氯化铵的添加都会使该菌生长变慢,而且氮源浓度越高菌丝生长越慢。外加碳源和金属离子对半纤维素降解和选择性系数的影响不大。 同时对菌株Y10在液态培养下产木质素降解酶的条件和培养基做了优化。结果表明,在初始产酶培养基中,菌株Y10的漆酶酶活在第10d达到最高,锰过氧化物酶酶活在第11d达到最高,基本上检测不到木质素过氧化物酶。菌株Y10产漆酶的最适温度为32℃,最适PH为6.0;产锰过氧化物酶的最适温度为32℃,最适PH为6.5。菌株Y10产漆酶的最佳碳源为甘露糖,最佳氮源为酒石酸铵,最适诱导剂VA浓度为3 mmol/L,最适表面活性剂TW-80浓度为1%。 利用响应面法对其产漆酶的培养基进行优化,优化后的培养基配方为葡萄糖10.00 g/L,酒石酸铵0.50 g/L,大量元素296.50 ml/L,微量元素100.00 ml/L,NTA 1.40 g/L,VA 5.00 mmol/L,吐温-80加入量为0.10%。进行了菌株Y10产漆酶的验证实验,实测酶活为5282.56 U/L,与预测酶活5162.73 U/L接近。在优化后培养基中,菌株Y10在第14 d达到生长的最高峰,第20 d时,漆酶酶活最高,为11325.00 U/L;第16 d时,锰过氧化物酶酶活最高,为30.77 U/L。 对菌株Y10的漆酶酶学性质做了初步的研究,结果显示,酶反应的最适温度为40℃-65℃,最适PH为3.0。在40℃,PH=3.0时,漆酶催化ABTS反应的米氏方程为 。 Fifteen white-rot fungi based on corn stalk were screened. One white-rot fungus Y10 with high selectivity value was obtained. The degradation of corn stalk was initially studied. The results indicated that the selectivity value was above 1 during the 30 day-cultivation and the highest was 3.88 after 15 days. The composition of untreated and treated stalk was analyzed through ultraviolet spectroscopy and infrared spectroscopy. It was found that the composition of treated stalk was greatly altered and the degree of the degradation of lignin is greater than the cellulose. Y10 was identified as Cerrena sp. by ITS -5.8S rDNA sequence analysis. The influence of metal ions, carbon sources and nitrogen sources on corn stalk degradation by white-rot fungus was studied. While all seven metal ions could promote lignin degradation, the cellulose degradation was best inhibited at certain ion concentrations. Notably, when 0.036% MnSO4·H2O and 0.36% MgSO4·7H2O were added into the medium, the cellulose degradation was restrained to the extents that the coefficients of lignin selectivity rose to 34.40 and 20.17 respectively. It was also found that all carbon sources except maltose can promote lignin degradation. The addition of carbon sources other than microcrystalline cellulose significantly promoted cellulose degradation. The addition of the nitrogen sources, ammonium tartrate, ammonium sulfate, oxalate, ammonium chloride, resulted in remarkable inhibition to mycelium growth; the larger the concentrations of nitrogen sources are, the slower the mycelium grew. The addition of carbon sources and metal ions had less impact on the degradation of hemicellulose and selectivity value. Meanwhile, we optimized the conditions and culture medium of the lignin-degrading enzyme production of strain Y10. The results showed that in the initial culture medium, the Lac activity was highest at the 10th day, the MnP activity was highest at the 11th day and the LiP could not be detected. The optimum condition of Lac was at temperature 32 and PH =6.0 and the optimum condition of MnP was at temperature 32 and PH =6.5. The optimum carbon source for Lac was seminose, the optimum nitrogen source was ammonium tartrate, the optimum content of VA was 3 mmol/L, the optimum content of TW-80 was 1%. PB and RSM were used to optimize the culture medium of laccase by white-rot fungus Y10. The optimum culture medium was consist of glucose 10.00 g/L, ammonium tartrate 0.50 g/L, macro elements 296.50 ml/L, trace elements 100.00 ml/L, NTA 1.40 g/L, VA 5.00 mmol/L, TW-80 0.10%. Under the optimal conditions, the activity of laccase was 5282.56 U/L and the experimental value agreed with the predicted value 5162.73 U/L. The biomass was highest at the 14th day, the Lac activity was highest at the 20th day, the MnP activity was highest at the 16th day. The results of the studies on the characteristics of Lac showed that the optimum temperature for Lac activity is 40℃-65℃ ; the optimum PH for Lac activity is 3.0 and under 40℃,PH=3.0, the Michaelis-menten equation of Lac catalized ABTS oxidation was .
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组特殊自养氨氧化混合种群,表现:无机环境种群生长迅速、生物量高;在一个完全无机的自养生长环境中,不仅保持高氨氧化速率,并出现丰富的异养微生物种群;该种群置于异养、厌氧环境中,迅速表现出产氢特征。对于这样一个特殊的生态体系,研究其共生机理,以及联接这些种群之间的碳源和能源问题,将具有非常重要意义。我们拟从种群特征、细胞表面分泌产物、游离体系产物多糖、蛋白和脂肪酸方面开展研究。 第一部分,自养氨氧化混合种群的基本特征。采用氨氧化培养基,进行种群氨氧化特征研究;采用扫描电镜观察自养混合种群的微观特征;沉降、离心去除微生物种群,分析水相中的总有机碳、糖类等物质;利用LB培养基进行种群的分离、纯化,并采用DGGE手段对微生物种群结构进行分析。结果表明,接入菌种后(2/5000(V/V)),培养液中氨(200mg/L)在3-5天内快速降解;亚硝酸盐与氨氮变化呈负相关趋势,仅有少量硝酸盐含量(< 30mg/L)。氨氧化种群的生物量增长与氨氧化趋势一致,初始生物量7.75 mg/L(蛋白含量),3-5天后生物量快速增长,并达到最高63.06 mg/L(蛋白含量)。电镜图片显示,种群外包裹一层粘液。离心除去菌体后,检测培养液总有机碳和糖的含量,同样表现出与生物量增长相似的特征,分别由初始的3.73、2.35 mg/L,3-5内天迅速增加,并分别达到最大值35.19、27.45 mg/L。经初步分离、纯化并对纯化菌株进行测序,获得了10株异养微生物分别为布鲁氏菌科苍白杆菌属、纤维单孢菌、类芽孢菌属、黄杆菌属、无色杆菌、鞘脂单胞菌、嗜麦芽寡养单胞菌、噬氢菌属、硫红球菌、假单胞菌;DGGE显示,约有20分条离带,我们对其中的两条优势条带进行切割回收测序,鉴定为欧洲亚硝化单胞菌(Nitrosomonas eur)。 第二部分:混合种群自养-异养菌共生的可能机制。在对微生物种群特征初步分析基础上,针对胞外糖类组分可能被微生物代谢分解,我们重点对微生物细胞蛋白质与糖类进行分析。采用超声结合RIPA裂解液裂解,SDS-PAGE电泳分析混合种群总蛋白种类,并通过氨基酸分析仪及红外光谱法分析氨基酸组成及蛋白红外特征。采用超声破碎结合反复冻融对细胞样品进行处理,提取液采用醇沉、Sevage脱氮白,凝胶过滤方法脱盐和分级分离。对提取物的糖分析包括:紫外扫描,红外光谱,核磁共振,单糖组成分析;扫描电镜观察菌群破裂现象。SDS-PAGE分析结果表明:氨氧化种群不同生长阶段都显示出42kD蛋白表达量很高,d4时42kD蛋白表达已经很强,4-7d内一直持续这种过量表达,直到d8后表达开始减弱。说明42kD蛋白可能与氨氧化密切相关。红外光谱分析显示:细胞提取物的特征峰分布在3427.42cm-1、1718.18 cm-1和1681.72 cm-1、1160.07和1086.74 cm-1,分别对应为OH、 C=O、C-O-C基团,表明具有蛋白的典型特征;氨基酸分析显示蛋白中的Gly,Asp,Ala,Glu含量相对较高。 提取物中胞外多糖分离谱图得到不均一组分,共得到6个收集峰;紫外扫描在201-213 nm处有多糖吸收峰,同样表明多糖成分不均一性;多糖红外光谱特征峰主要分别在3400.49 cm-1、2920.28 cm-1、1154.54和1087.52 cm-1,对应OH、-CH2- or CH 、C-O-H or C-O-C等多糖特征基团;多糖提取物核磁共振1H d4.3~5.9之间出现强吸收峰,这是1H中,多糖存在的明显证据,1H NMR中,其中O-乙酰基的甲基上的氢信号为d1.1~1.3之间。糖肟全苯甲酸酯衍生物的HPLC测定中,得到单一的单糖峰,由于时间问题,还未进行更深入的试验;电镜图片显示,种群中的细胞有大量的破裂现象。 实验表明,自养氨氧化混合种群显示出快速的氨氧化速率,氨氧化过程生物量和有机质的增加明显。微生物种群包裹粘液层,并分离纯化出大量的异养菌;去除菌体后的游离培养液中存在有机质(包括多糖)说明无机自养生长体系中存在异养菌生长、繁殖的二次碳源;细胞提取物中蛋白条带数目多、种类丰富;细胞多糖提取物具有明显的多糖特征,以及单糖的存在。结合种群的显微特征和游离体系中的有机质的检测结果,我们认为,无机自养生长体系中,种群细胞生长过程中发生的破裂现象可能是导致大量的蛋白、多糖释放到游离胞外,并成为其他异养菌生长的碳源和氮源。这可能是自养体系中,大量异养菌共生的可能机制,至于是什么原因引起种群生长过程中产生的破裂现象,还有待下一步深入研究。 A group of mixed autotrophic ammonia oxidizing populations, having much biological characteristic tested by concerned personnel for pilot test: Performed rapid population growth and obtained high biomass in inorganic environment; Not only maintained a high rate of ammoxidation, promoted a wealth of heterotrophic microbial populations growth in a totally inorganic and autotrophic growth environment; Placed in heterotrophic and anaerobic environment,had the performance characteristics that could rapidly produce hydrogen.For such a special ecological system, Study its symbiotic mechanism and the connection between these populations of carbon and energy issues, will have a very important significance. We intended from the characteristics of the population, the secretion product of cell surface, free substance in the liquid medium like polysaccharide, protein and fatty acids carrying out research. Part I: The basic features of mixed autotrophic ammonia oxidizing populations . Use inorganic liquid medium, processed study for ammonia oxidation characteristics of the population; we used scanning electron microscopy to get micro-features of autotrophic ammonia oxidizing populations .The medium was carried out settlement and centrifugal then removed the microbial populations, after all of that we analysis the water phase for total organic carbon(TOC), carbohydrate and other substances; Solid ammonia oxidizing medium was adopted to separation and purification of population, DGGE means was for structure analysis of microbial population. The results showed that after the inoculum of bacteria (2 / 5000 (V / V)), ammonia in the culture medium (200 mg / L) was rapid degradation in 3-5 days; ammonia and nitrite have the negative correlation between changes in the trend, then only a small amount of nitrate content (<30mg / L). The biomass growth of ammoxidation population in line with the trend of ammonia oxidation, the initial volume of it was 7.75 mg / L (protein content), in 3-5 days upto 63.06 mg / L (protein content). Electron microscope image showed, the populations were wrapped in a layer of mucus, including the a large number ruptted micorbe , Centrifuge to remove bacteria, then detected the medium for total organic carbon and sugar content, result took on the same characteristics with biomass growth, that were from the initial 3.73、2.35 mg / L respectively, in 3-6 days achieved rapid increase in the maximum to 35.19、27.45 mg / L respectively. After initial separation、 purification ,then processed sequencing to strains purified and got the result that there were 10 heterotrophic microorganisms : Brucella Branch pale bacillus, Cellu lomonas, Bacillus species category, a Flavobacterium, colorless Bacteria, Aeromonas sheath fat, little support maltophilia Aeromonas, macrophages species hydrogen, sulphur-MI, Pseudomonas bacteria spores; DGGE display, there were 20 separation bands approximately. Part II: Mixed populations that autotrophic - heterotrophic bacteria symbiotic mechanism. On the basis of preliminary analysis of microbial population characteristics, aiming at extracellular carbohydrate components might be decomposition by microbial, we focused on microbial cell protein and carbohydrate analysis. Using ultrasound combined with RIPA lysis cracking the cells, SDS-PAGE electrophoresis analysis the total protein species of the population, and through the amino acid analyzer studied the compositions of amino acid and infrared spectroscopy analysis of a protein infrared characteristics. Using ultrasound combined with repeatedly freezing and thawing to treated the cell sample, then took the means that alcohol precipitation, deproteinization by Sevage, gel filtration aimed at desalination and grade separation to deal with the lysates . The extraction of sugar analysis included: UV scanning, IR, NMR, single-sugar composition analysis. SDS-PAGE analysis showed that: 42 kD protein expression was very high at different growth stages of mixed autotrophic ammonia oxidizing populations , on the fourth day, 42 kD protein expression had been very strong, 4-7d, it had continued this excessive expression, then started to weaken after 7 days. 42 kD protein that might be closely associated with ammonia oxidation. Infrared spectral analysis showed that: cell extracts with the characteristic that the peak distribution in 3427.42 cm-1、1718.18 cm-1 and 1681.72 cm-1、1160.07 cm-1 and 1086.74 cm-1 corresponding to OH、C = O、C-O-C Groups which had the typical characteristics of protein; and analysis showed that amino acids including Gly, Asp, Ala, Glu ,the content in the protein is relatively high. Exopolysaccharide in the extracts had the separation map that it was uneven, received a total of six collection peaks by the detection mode of phenol-sulphruic acid method ; ultraviolet scan in the 201-213 nm department had polysaccharide absorbing peak, the same ingredients that polysaccharide heterogeneity; infrared polysaccharide spectral characteristics of the main peak at 3400.49 cm-1, 2920.28 cm-1, 1154.54 and 1087.52 cm-1, corresponding OH,-CH2-or CH, C-O-H or C-O-C;and other characteristics of polysaccharide group; 1H NMR of polysaccharide extract appeared absorption peak between d4.3 ~5.9, which is the apparent evidence of polysaccharide, In 1H NMR, the hydrogen signal of one of O-acetyl was between 1.1 to 1.3. The determination of Sugar oxime whole benzoate derivatives by HPLC, there was a single-sugar peak, as a matter of time, yet more in-depth test. Summary: Mixed autotrophic ammonia oxidizing populations show us that it had the ability in ammonia oxidizing and it was great, organic matter and biomass increased significantly in the process of ammonia oxidation. Microbial populations was wrapped up slime layer, the phenomenon of cell breakdown obviously, and there were a lot of separation and purification of the heterotrophic bacteria; a lot of organic matter (including polysaccharides)remined in the medium that removal of cell indicated the inorganic system existed secondary carbon sources that could be used by the heterotrophic bacteria ; there were a large number proteins bands of cell extract, rich variety; cell extracts of polysaccharide had obvious characteristics of polysaccharide, and the existence evidence of single-sugar. Combined population of microscopic characteristics and free of organic matter in the test results, we believe that the health of inorganic system, population growth occurred in the course of the breakdown of the phenomenon is likely to lead to a lot of protein and polysaccharide released into the extracellular free, And other heterotrophic bacteria use them to the growth as carbon and nitrogen. This may be autotrophic system, the large number of heterotrophic bacteria symbiotic mechanism.
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本文主要研究了具有己酸乙酯酯化活性的真菌菌株的筛选和发酵条件优化。从大曲和糟醅样品中分离纯化获得79株产生透明圈的丝状真菌,菌落形态初步识别结果显示分离菌株包括红曲霉属、根霉属及曲霉属等菌株。其中菌株EM-56酯化酶活力最强,发酵获得的粗酶制剂酶活为172.36 u。根据显微形态、菌落形态及生理生化特征,初步鉴定该菌株为曲霉科红曲霉属紫色红曲霉(Monascus purpureus)。 在此基础上重点研究了菌株EM-56在不同培养基成分及不同培养条件下的产酶情况,确定了最佳培养基和培养条件。通过单因素实验确定在基础培养基中添加最佳碳源为葡萄糖,最佳氮源为蛋白胨。正交优化实验结果确定了最佳培养基组成:以麸皮为基础培养基,添加葡萄糖 2%,蛋白胨 0.3 %,KH2PO4.3H2O 0.05 %,MgSO4.7H2O 0.06 %。菌株EM-56在上述培养基中的最佳发酵条件为:初始pH 5.5,发酵温度为35°C,发酵时间7d,种龄48h,接种量8%,装瓶量50g / 瓶(500mL)。在最佳培养基和发酵条件下,菌株EM-56发酵获得的粗酶制剂酶活达到241.56 u,比优化前提高了40.15%。 In this paper, the research focuses on the selection of fungus with esterifying activity and optimization of fermentation conditions. We isolated 79 strains which had transparent zones from Daqu and fermented grains. The isolated strains contained Monascus、Rhizopus and Aspergillus through primary morphology analysis. The strain of EM-56 which produces strongest esterase was selected. The enzyme activity reached 172.36u. According to related literature, EM-56 was identified as Monascus purpureus through morphology analysis and biochemical determination. We also studied the effects of different medium and fermentation conditions on the esterase production of strain EM-56. The optimal medium and fermentation conditions were determined. Single factor experiment result shows that the optimal carbon source added is glucose and the optimal nitrogen source added is peptone. The optimal fermentation medium determined by orthogonal optimization test is as follows: wheat bran as substrate, glucose 2%, peptone 0.3%, KH2PO4.3H2O 0.05%,MgSO4.7H2O 0.06%. The optimal fermentation conditions are: initial pH 5.5, cultural temperature 35°C, cultural time 7d, seed age 48h, inoculation 8%, medium mass 50g / flask(500mL). The esterse activity of EM-56 cultivated in the optimal medium and fermentation conditions reached 241.56u and increased by 40.15% compared with the original activity.
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目前对PVA生物降解研究重点逐渐转移到对PVA降解菌和PVA降解酶的研究开发上,随着对PVA降解高效新菌株的不断发现和PVA降解酶作用机理和分泌机制的深入了解,利用高效微生物或酶法治理PVA这类高聚物的污染将具有较大的应用潜力。本论文研究工作正是基于这种客观条件下进行的,对本实验室前期分离的PVA降解菌株P1、共生菌B1+B2、Pa、Pb为研究对象,重点研究了菌株P1和共生菌B1+B2的产酶条件和产酶特性,验证找出了影响菌株P1产酶的生长因子,论证了菌株B1+B2的产酶特性,优化得出了菌株B1+B2的最佳产酶条件;然后对共生菌B1+B2的PVA降解酶的稳定性进行了研究;最后研究了最佳组合菌的产酶特性和最佳产酶条件。主要研究结果如下: 1 通过对菌株P1产酶因子的研究,找出了核黄素是菌株P1产酶的必须因子,在以淀粉为碳源时,核黄素只是产酶的必须因子,而不是菌体生长的必须因子;在以PVA为碳源时,核黄素既是生长的必须因子,也是产酶的必须因子,是菌株P1的生长因子。 2 对共生菌B1+B2的产酶条件和产酶特性进行了研究,并通过正交实验找出了影响菌株产酶的主要条件和菌株产酶的最佳条件。 3 对共生菌PVA降解酶的稳定性进行了研究,确定了影响酶稳定性的主要理化条件。 4 通过对菌株降解性能的比较,确定菌株Pa、Pb、共生菌、P1的作为组合菌的组成菌,然后通过复配实验确定出菌株的最佳组合为菌株Pa、P1、共生菌,最后通过正交实验确定最佳组合菌的最佳配比。 5对影响组合菌产酶的因素进行了研究,通过正交实验确定了影响组合菌产酶的主要因素和最佳产酶条件。 本文通过对PVA降解菌株产酶条件和特性的研究,旨在为PVA降解菌生产酶制剂及进一步优化PVA降解菌在PVA废水治理中的应用提供理论和应用依据。 Now the PVA-degrading bacteria and polyvinyl alcohol-degrading enzyme are the key studies on the PVA biological degradation. It has great application potential using special bacteria and enzyme to treat pollution of PVA, with some high efficient Strain and enzyme were found. The study of this paper was based on that objective condition. The stain P1, symbiotic bacteria B1+B2, stain Pa and strain Pb were studied .The conditions of enzyme production and enzyme production characteristic of stain P1, symbiotic bacteria B1+B2 were our key study, we tested and verified the growth factor which effected enzyme production of strain P1, demonstrated enzyme production characteristic of symbiotic bacteria B1+B2, optimized and obtained the optimum conditions of enzyme production; then we studied the stability of polyvinyl alcohol-degrading enzyme of strain B1+B2; last the enzyme characteristic and the optimum conditions of alcohol-degrading enzyme production of optimum combination stains were studied. The main study results are below: 1. Through the study of enzyme production factor we found that lactoflavin is the necessary factor in strain P1 enzyme production. When we used starch to be carbon energy, lactoflavin is only the necessary factor of enzyme production, but not growth factor. When we used PVA to be carbon energy, lactoflavin was not only the necessary growth factor ,but also the necessary enzyme production factor.So it was the growth factor of strain P1 2. The enzyme production conditions and enzyme production characteristic of symbiotic bacteria B1+B2 were studied. Through the orthogonal experimental design, the main conditions which effected the enzyme production and the optimum conditions of enzyme production were obtained 3. Through the study of the stability of polyvinyl alcohol-degrading enzyme, the main physical and chemical conditions which effected the enzyme stability were 4. The stain P1,symbiotic bacteria B1+B2, stain Pa and strain Pb were selected to form combination bacteria. The stain P1,symbiotic bacteria B1+B2,stain Pa were the optimum combination through duplication experiment. Then the optimum ratio was obtained through orthogonal experiment. 5. Studied the factors which effected the polyvinyl alcohol-degrading enzyme activity, then through orthogonal experiment, the main factors and condition of enzyme production which effected the combination bacteria were achieved. The result of the study was valuable for the ferment of the PVA-degrading enzyme and the optimization of the PVA-degrading performance in the PVA wastewater.
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齐墩果酸(OA)是一个分布广泛、含量丰富的天然三萜化合物,常以皂苷元的形式广泛存在于植物中,具有多种重要生物活性。但是OA许多活性较弱,且生物利用度低,限制了其在临床上的应用。一是OA水溶性差;二是抗癌活性仍与临床应用的抗癌药物相差比较大。 真菌在微生物转化中具有种类多、培养条件比较简单等特点,为了寻找到具有转化OA能力的菌株,采取一步发酵的方法,在18株实验室保藏真菌菌株中筛选到5株目的菌株,TLC分析显示有转化效果。 随后采用二步发酵的方法作为复筛,验证5株菌株转化能力,波谱分析结果表明5株菌株对OA确实有转化作用。 选择5株菌种中代号1F-2 2菌株作为放大实验菌株,分离转化产物,得到OA衍生物108(相对分子量414m/z)和1010(相对分子量340 m/z),分离出的产物用于活性检测。寻找到产物108的RP-HPLC分离条件,质谱得出二者相对分子质量。 为验证OA转化产物抗肿瘤活性,首次研究了OA对卵巢癌细胞株IGROV1和人乳腺癌细胞株MDA-MB-231作用,通过细胞增殖抑制实验、用MTT法检测细胞活性,结果表明齐墩果酸可降低卵巢癌细胞株IGROV1和乳腺癌细胞MDA-MB-231细胞增殖能力并呈剂量依赖性,对肿瘤细胞株的半数有效抑制浓度化IC50 分别为36.58μg/mL和38.8μg/mL (P<0.01)。OA能抑制肿瘤细胞活性,并且OA对卵巢癌细胞株IGROV1抑制活性高于乳腺癌细胞MDA-MB-231。 在此基础上,转化产物108和1010对卵巢癌细胞株IGROV1和人乳腺癌细胞株MDA-MB-231的抑制作用也进行研究,MTT实验结果表明,转化产物对两株癌细胞也有抑制活性(P<0.01)。 总之,本文工作为进一步开展齐墩果酸类化合物结构改造和抗肿瘤活性的研究奠定了基础。 Oleanolic acid (OA) is a triterpenoid widely distributed in the nature which possesses various important bioactivities. OA also serves as aglycon of many natural saponins. However, the relatively weak activities and poor bioavailability hinder its clinical use. Firstly, poor water-solubility results in worse bioavailability. Secondly, compared with clinical antitumor drug, the antitumor effect of OA has a great difference, it is worse. Many fungi have ability to transform nature products into a variety of derivatives, and transformation conditions of fungi are simple. Attempt to obtain fungi strains able to biotransform OA, we carried out the following experiments: To investigate the biotransformation 0f OA by strains supplied firstly, we used one-step fermentation method to screen the aimed strains from 18 fungus strains stored in our laboratory. On the basis of the initial screening experiments, we found 5 aimed strains. The TLC results showed that the 5 fungi strains could transform OA into other components derivatives. Then we used two-step fermentation method as secondly screening. We repeated the five strains to do the experiments, analytical data of the results proved the transformation indeed. In the followed experiments work, we chose 1F-2 2 strain as large-scale transformation fungus from the aimed fungi. We got two biotransformation products of OA by 1F-2 2, and named those derivatives 108 and 1010. We found RP-HPLC separation conditions of product 108. The two products were characterized by ESI-MS. To verify the anti-tumor activity of biotransformation products of OA, we studied the inhibition effect of oleanolic acid on the ovarian carcinomas IGROV1 and breast cancer cell line MDA-MB-231 firstly. With an assay based on a tetrazolium dye (MTT), the effects of various concentrations of oleanolic acid on ovarian carcinomas IGROV1 and breast cancer cell line MDA-MB-231 were studied. MTT method was used to measure the tumor cells viability. Compared with the control group, oleanolic acid can significantly inhibit the viability of the ovarian carcinoma cells IGROV1 and MDA-MB-231 breast cancer cell line (P<0.01), IC50 values were 36.58μg/mL or 38.8μg/mL. Oleanolic acid can inhibit the malignant tumor cells viability, and inhibitory activity of OA to ovarian carcinomas IGROV1 was higher than to breast cancer cell line MDA-MB-231. On this basis, we studied the anti-tumor activity of the two derivatives of OA [called 108 (414 m/z) and 1010(340 m/z)]. It came to the conclusion that the two derivatives also showed potent inhibitory effect on the growth of these tumor cells(P<0.01). Therefore, the results of studies will benefit the further investigating on the relationships of structures and antitumor activities of OA.
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本文筛选出一株能利用木糖产乙醇的丝状真菌Z7,对其利用木糖和半纤维素水解产物产乙醇的发酵条件进行了研究,并对Z7 利用玉米芯产木聚糖酶的条件进行了优化。全文分为三部分: 第一部分:目标微生物筛选、纯化及系统发育分析。以木糖为唯一碳源,采用梯度稀释和平板化线法从高温、中温酒曲中分离到16 株能利用木糖良好生长的丝状真菌;通过发酵试验复筛,获得一株能产乙醇的丝状真菌Z7;综合形态学和ITS 序列分析,初步鉴定为Aspergillus flavus。 第二部分:Z7 的乙醇发酵条件研究。以木糖为碳源,通过单因素试验确定最佳氮源和发酵温度;通过正交试验及SPSS 软件分析得到了不同N、P、K 成分对乙醇、残糖和菌体干重的影响。获得最佳的发酵条件为:(g/L)木糖50,尿素1, NH4NO3 1, K2HPO4 2 , KCl 0.5 , MgSO4.7H2O 0.5 , NaNO3 1 , pH 自然,培养温度33 ℃。以玉米芯半纤维素稀酸水解液为底物进行乙醇发酵,根据稀酸水解的单糖释放量和乙醇产量,确定115 ℃,1 h 为最佳玉米芯预处理条件;结合最佳发酵条件,添加1 g/L 的吐温20 能获得最大的乙醇浓度8.31 g/L。因此,Aspergillus flavus Z7 能利用半纤维素水解产物产乙醇,其中木糖的利用率80%以上。 第三部分:Z7 利用玉米芯产木聚糖酶条件优化。Aspergillus flavus Z7 在具有产乙醇能力的同时还具有产木聚糖酶的能力。本文通过单因素和正交试验得到最佳产酶培养基组分为:(g/L)玉米芯20,尿素2, 酵母膏2.5, K2HPO4 5,NaNO31, MgSO4.7H2O 1。单因素试验表明,用纱布代替塑料布密封摇瓶封口能显著提高产酶量;Z7 在碱性条件下具有更强的产酶性能。在最优条件下发酵,能产生最大木聚糖酶活122.23IU/mL。通过薄层分析,验证了Z7 产生的木聚糖酶具有水解木聚糖生成木糖及木寡糖的能力。 A strain of filamentous fungus which can produce ethanol by using the xylose was isolated in this research. The ethanol fermention conditions from xylose and dilute-acid hydrolyzate of the corn core were studied. The conditions of xylanase production by Z7 were also optimized. The paper involved three parts. Part1: Isolation, purification and phylogenetic analysis of the microbe. By using xylose as the single carbon source and the pla te streaking method, several filamentous fungi were isolated from the wine starter; through the fermentation test, a filamentous fungus Z7 which can produce ethanol was further recognized; furthermore, according to the morphologic observation and ITS seque nces analysis, Z7 was identified as Aspergillus flavus at the first step. Part2: Research on the condition of ethanol fermentation by Z7. By single factor experiment, the optional nitrogen resource and temperature of the fermentation were fixed; meanwhile, through the orthogonal array tests and the analysis of statistic software SPSS, the optional component of the culture medium and the fermentation condition were organized as follows: (g/L) xylose 50, urea 1, NH4NO3 1, K2HPO4 2, KCl 0.5 , MgSO4.7H2O 0.5, NaNO31, pH nature, temperature 33℃. Based on these optimal parameters, the fermentation of dilute-acid hydrolyzate of the corn core was carried on by Z7. According to the quantities of released sugar monomers and content of the ethanol, 115℃ in 1h is the best pretreatment condition; the maximal ethanol content can be obtained when 1g/L Tween 20 was added to. Therefore, the filamentous fungus Aspergillus flavus can use the hydrolysate of hemicellulose to produce ethanol, and the rate of xylose utilization was over 80%. Part3: Optimization of Z7’s xylanase producing condition from corn core. Aspergillus flavus Z7, which can utilize xylose or the hydrolysate of hemicellulose to produce ethanol, also had the ability of xylanase production. The optional component of the culture medium were fixed by the single factor experiment and the orthogonal array tests, and they were organized as follows: (g/L) corn core 20, Urea 2, Yeast extract 2.5, K2HPO4 5, NaNO31, MgSO4.7H2O 1; it was testified by the single factor experiment that sealing the shaking flasks with pledget other than plastic paper can obviously increase the xylanase activity; moreover, Z7 showed better xylanase production ability when in the alkali environment. Under the optional fermentation condition, the maximal xylanase activity 122.23IU/mL was proved. Through the analysis of thin- layer chromatography (TLC), the ability of xylanase from Z7, which can hydrolyze xylan to xylose monomer and oligomer, was vividly displayed.
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本文主要研究了泸州老窖古酿酒作坊内外环境空气真菌和空气细菌的群落结构和分布特征。结果如下: 作坊内外环境空气微生物浓度差别显著,并随季节变换而变化,春、夏季微生物浓度较高,秋、冬季较低,空气真菌在夏季达到最高,细菌在春季最高。 古作坊内外环境检测到的真菌均为16 属,但优势菌属不同,作坊外的优势菌属为青霉属(Penicillium)、曲霉属(Aspergillus)、无孢菌(non-sporing)、枝孢霉属(Cladosporium)和链格孢属(Alternaria);而作坊内优势菌属为曲霉属、青霉属、酵母菌(Yeast)、无孢菌,作坊内还含有较高浓度的根霉属(Rhizopus)、毛霉属(Mucor)、短梗霉属(Aureobasidiu),枝孢霉属和链格孢属等,曲霉属、酵母菌、根霉属、毛霉属为古酿酒作坊重要的酿酒真菌,青霉属、链格孢属为酿酒不利菌群。对古作坊内曲霉属进行了初步鉴定,主要是小冠曲霉(A.cristatellus)、米曲霉(A.oryzae)、黑曲霉(A.niger)和白曲霉(A.cadidus)。 空气细菌10 属21 种,作坊内外环境的优势菌属均为芽孢杆菌属(Bacillus)、微球菌属(Micrococcus)、葡萄球菌属(Staphylococcus)、假单胞菌属(Pseudomonad),其中芽孢杆菌属在作坊内占有绝对的优势,浓度比在40℅以上,是古酿酒作坊重要的酿酒细菌,另外还检测到较高浓度的乳酸杆菌(lactobucillus),这类菌容易使酒味发涩发苦,为酿酒不利菌。 作坊内外环境空气微生物表现出明显的交流现象。作坊内,青霉属、枝孢霉属、链格孢属、葡萄球菌属等杂菌占有一定比例;而在作坊外,芽孢杆菌属、曲霉属、根霉属(Rhizopus)、酵母菌等处于相对较高水平,绿化环境较好的营沟头作坊内的短梗霉属,枝孢霉属和链格孢属等杂菌含量低于什字头和新街子作坊。 The community structure and distribution characteristic of airborne microbes was investigated in ancient brewage workshops of luzhoulaojiao. The results are as follows: The concentration of airborne microbes was different in interior and exterior environment of ancient workshops, and also varied by seasons. microbial concentration was higher in spring and summer, and lower in fall and winner. The highest levels of airborne bacteria was in spring, but the fungal’s in summer. The identified genus of fungi were 16 in interior and exterior environment of the ancient workshops. But the dominant genus were different , The advantage genus in the interior were Aspergillus, Yeasts, Penicillum and Nonsporing and in the exterior were Penicillum, Nonsporing, Cladosporium, Aspergillus and Aureobasidiu. Rhizopus ,mucor, Aureobasidiu, Cladosporium, Alternaria and all also were at a higher level. Among these, Aspergillus, Yeasts, Rhizopus ,mucor are important vintage flora . Penicillum, Alternaria do harm to vintage. Aspergillus of ancient workshops was identified , the preponderant aspergillus species were A.cristatellus, A.oryzae, A.niger and A.cadidus in ancient brewage workshops. 10 genus 21 species bacteria were identified, the advantage genuses among the interior and exterior of the three workshops were bacillus, microccus, Staphylococcus Pseudomonas. Bacillus, which account for beyond 40℅ of the total bacteria concentration in all sampling pots, was the most dominant genus. Lactobacillus was identified at a high level in ancient workshops, it makes spirit taste bitter and astringent. So it is not a kind of good bacterium for vintage. The fungus in the interior and exterior atmosphere characterized intercommunion phenomenon. Obviously, the concentration of profitless fungus such as Penicillum, Cladosporium, Alternaria appeared in the interior, and the fungus such as Bacillus, Aspergillus, Rhizopus and Yeasts in the exterior were at a relatively high level. the harmfull fungus in yinggoutou workshops such as Aureobasidiu, Cladosporium, Alternaria and all were lower than shenzitou and xinjiezi workshops.
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Immobilized with PVA,sodium alginate and activated carbon,both Zoogloea sp. and Fusarium sp.strains could degrade phenanthrene and pyrene efficiently.The optimal carrier was made of 100ρ·g -1 L PVA,5 sodium alginateρ·g -1 L and 50 activated carbon ρ·g -1 L.The degradation rates of phenanthrene and pyrene in 10 days were 87.48% and 75.34% by the immobilized bacterium,37.04% and 20.85% higher than those by the free bacterium,and the rates in 15 days were 84.36% and 74.87% by the immobilized fungus,5.35% and 11.23% higher than those by the free fungus.
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研究了黄土塬区塬面不同套种作物的幼树苹果园土壤微生物的分布和酶活性。结果表明,果园土壤中微生物的分布在果树行与套种行中是不均匀的,细菌和真菌多为果树行多于套种行;土壤的过氧化氢酶,蔗糖酶,脲酶以及中性磷酸酶活性在套种小麦的果园中以套作行高于果树行,而套种烤烟则使脲酶和中性磷酸酶活性有所降低。套种豆类有利于土壤肥力的均衡发展。