Enhanced bio-decolorization of1-amino-4-bromoanthraquinone-2-sulfonic acid by Sphingomonas xenophaga with nutrient amendment

Bacterial decolorization of anthraquinone dye intermediates is a slow process under aerobic conditions. To speed up the process, in the present study, effects of various nutrients on 1-amino-4-bromoanthraquinone-2-sulfonic acid(ABAS) decolorization by Sphingomonas xenophaga QYY were investigated. The results showed that peptone, yeast extract and casamino acid amendments promoted ABAS bio-decolorization. In particular,the addition of peptone and casamino acids could improve the decolorization activity of strain QYY. Further experiments showed that L-proline had a more significant accelerating effect on ABAS decolorization compared with other amino acids. L-Proline not only supported cell growth, but also significantly increased the decolorization activity of strain QYY. Membrane proteins of strain QYY exhibited ABAS decolorization activities in the presence of L-proline or reduced nicotinamide adenine dinucleotide, while this behavior was not observed in the presence of other amino acids. Moreover, the positive correlation between L-proline concentration and the decolorization activity of membrane proteins was observed, indicating that L-proline plays an important role in ABAS decolorization. The above findings provide us not only a novel insight into bacterial ABAS decolorization, but also an L-proline-supplemented bioaugmentation strategy for enhancing ABAS bio-decolorization.     

Quinone-mediated decolorization of sulfonated azo dyes by cells and cell extracts from Sphingomonas xenophaga

The effects of various quinone compounds on the decolorization rates of sulfonated azo dyes by Sphingomonas xenophaga QYY were investigated. The results showed that anthraquinone-2-sulfonate (AQS) was the most effective redox mediator and AQS reduction was the rate-limited step of AQS-mediated decolorization of sulfonated azo dyes. Based on AQS biological toxicity tests, it was assumed that AQS might enter the cells to kill them. In the cytoplasmic extracts from strain QYY, AQS effectively increased decolorization rates of sulfonated azo dyes than other quinone compounds. In addition, we found a NADH/FMN-dependent AQS reductase using nondenaturing polyacrylamide gel electrophoresis (Native-PAGE).     

菌株HP3对溴胺酸的降解机理

通过正交试验,得到菌株HP3降解溴胺酸最佳条件为温度30℃,pH值7.0,摇床转速160r/min,培养基中不含NaCl有利于溴胺酸的降解.在此条件下,溴胺酸的降解遵从负指数模式.葡萄糖和硫酸铵的加入显著地提高菌体降解溴胺酸的速度.不同金属化合物对菌株HP3降解溴胺酸有不同程度的抑制作用,其中以HgCl₂和AgNO₃最显著.菌株HP3能降解化合物蒽醌、1,4,5,8-四羟基蒽醌和1-氨基蒽醌-2-磺酸钠,表明该菌株对底物的降解没有严格的专一性.溴胺酸经菌株HP3作用后吸收光谱发生明显变化.TOC分析表明,菌株HP3将溴胺酸的蒽醌环破坏后利用其中的一部分有机碳.液相色谱检测有中间产物邻苯二甲酸生成.溴胺酸降解终产物分子量为289和290.     

黄曲霉A5p1脱色类黑精及其机理初探:葡萄糖氧化酶的作用

类黑精是一种高分子难降解色素污染物质,在糖蜜酒精废水中大量存在.选取在前期研究中对糖蜜酒精废水具有较好脱色作用的黄曲霉(Aspergillus flavus)A5p1(保藏号CGMCC.4292),以合成的类黑精为对象研究脱色机理,试图为实际废水的生物脱色提供理论基础.结果显示,此菌株对合成类黑精具有生物吸附和生物降解的双重作用,以后者为主,最高脱色率可达65%.相较于漆酶、依赖/不依赖于锰的过氧化物酶等常规氧化脱色酶,此菌株中的产过氧化氢酶对脱色起主要作用,而且脱色过程中作为产过氧化氢酶之一的葡萄糖氧化酶与类黑精脱色率之间存在着一定的正相关性.添加葡萄糖氧化酶激活剂和抑制剂进行验证,发现葡萄糖氧化酶酶活力、过氧化氢产量和脱色率三者之间呈相关性变化.初步推测,葡萄糖氧化酶是黄曲霉A5p1生物降解脱色类黑精的关键酶之一.     

酵母Candida krusei对合成染料的脱色

通过筛选实验,从土壤中新分离到1株对活性艳红K-2BP具有明显脱色效果的酵母菌株Y-G-1,经鉴定为克鲁斯假丝酵母Candida krusei.该菌株对含活性艳红K-2BP起始浓度为200mg/L的培养基,最大脱色率为99%,达到最大脱色率的时间是12h.克鲁斯假丝酵母的最佳接种量应不低于5%(体积分数),培养基最适pH在4～9之间,氮源(NH₄)₂SO₄浓度不低于0.1%,相对应的碳源葡萄糖浓度不低于0.5%.对脱色机理的研究表明,该菌株对活性艳红K-2BP的去除属于降解脱色.此外,该菌株对另外9种染料(50mg/L)的脱色率在62%～96%之间.其中,对偶氮染料弱酸艳红B、活性黑KN-B和活性红M-3BE的脱色率都达到了90%以上,对三苯甲烷染料(酸性媒介漂蓝B)的脱色率达到了93%.表明克鲁斯假丝酵母在染料废水的处理上可能具有较好地应用潜能.     

Trametes sp.SQ01和Chaetomium sp.R01混合培养对四种染料的脱色

利用本实验室新构建的白腐菌Trametes sp.SQ01和毛壳菌Chaetomium sp.R01混合培养体系,对刚果红、酸性红、橙黄G和溴酚蓝4种染料进行了脱色研究.结果表明,SQ01与R01混合培养所产生的锰过氧化物酶(MnP)酶活比菌株SQ01单独培养时提高了约5.5倍.菌株R01的接种量、接种时间对混合培养中...     

固定化蒽醌对偶氮染料生物降解促进作用研究

采用非水溶性蒽醌固定化技术对偶氮染料生物降解促进作用进行了研究,对比了海藻酸钙,聚乙烯醇-硼酸,聚乙烯醇-海藻酸钙和琼脂4种固定化技术,探讨了溶解氧对脱色过程的影响和固定化蒽醌系统脱色广谱性.研究结果表明,固定化蒽醌可提高多种偶氮染料生物厌氧脱色速度1.5～2倍和降低偶氮染料脱色过程氧化还原电位-10～-15 mV;经4次循环使用后,其加速作用仍保持在90%以上;固定化蒽醌微生物系统具有很强抗氧冲击能力.     

Evaluation of Bacillus sp. MZS10 for decolorizing Azure B dye and its decolorization mechanism

To evaluate decolorization and detoxification of Azure B dye by a newly isolated Bacillus sp. MZS10 strain, the cultivation medium and decolorization mechanism of the isolate were investigated. The decolorization was discovered to be dependent on cell density of the isolate and reached 93.55%(0.04 g/L) after 14 hr of cultivation in a 5 L stirred-tank fermenter at 2.0 g/L yeast extract and 6.0 g/L soluble starch and a small amount of mineral salts. The decolorization metabolites were identified with ultra performance liquid chromatography-tandem mass spectroscopy(UPLC-MS). A mechanism for decolorization of Azure B was proposed as follows: the C=N in Azure B was initially reduced to –NH by nicotinamide adenine dinucleotide phosphate(NADPH)-dependent quinone dehydrogenase, and then the –NH further combined with –OH derived from glucose to form a stable and colorless compound through a dehydration reaction. The phytotoxicity was evaluated for both Azure B and its related derivatives produced by Bacillus sp. MZS10 decolorization, indicating that the decolorization metabolites were less toxic than original dye. The decolorization efficiency and mechanism shown by Bacillus sp. MZS10 provided insight on its potential application for the bioremediation of the dye Azure B.     

芘胁迫对5种羊茅属植物根系分泌的几类低分子量有机物的影响

根系分泌物与有机污染物的植物修复过程密切相关,研究胁迫条件下不同修复潜力植物根系分泌物的释放特征有助于揭示植物修复的内在机制.借助根际袋土培试验研究了芘胁迫(10~160 mg·kg~(-1))下5种羊茅属植物在不同胁迫期(30~70d)时根系分泌物中几种低分子量有机物的释放特征.结果表明:1芘胁迫促进了根系对可溶性糖的分泌:随着胁迫水平的升高、胁迫期的延长,其分泌量呈"先升后降"变化趋势,胁迫水平为C3(40.36 mg·kg~(-1))、胁迫期为40 d时达到最大值;修复潜力越大,趋势越明显.2芘胁迫增强了根系对低分子量有机酸的释放,修复潜力越大,释放高峰值出现时的胁迫浓度越高,且主要以草酸、乙酸、乳酸和苹果酸为主(98.15%),但修复潜力较强物种的根系分泌物中也检测到微量反丁烯二酸.3芘胁迫对氨基酸的种类影响不大,但对分泌量影响较大:苏氨酸、丝氨酸、甘氨酸、丙氨酸的分泌量随着胁迫水平的升高而剧增;脯氨酸、羟脯氨酸和天冬氨酸对芘胁迫的响应近乎以功能群的形式参与植物修复过程,参与的组分越多,修复潜力越强.可见,芘胁迫下根系分泌物中可溶性糖、低分子量有机酸以及氨基酸的释放特征与植物自身的修复潜力有关;修复潜力越强,释放量越多且成分越复杂,并表现出更强的环境适应性及生理可塑性.     

醌介导染料脱色菌株的分离鉴定及特性

实验分离获得1株能够利用醌化合物使磺酸化偶氮染料脱色的菌株JL,通过形态特征、16S rDNA与16S-23S区间序列分析表明,该菌株为蜡状芽孢杆菌(命名为Bacillus cereus JL).菌株JL使酸性大红3R脱色的最佳条件为葡萄糖浓度1g/L, pH值为5~7,温度30℃,接种量0.25g/L.蒽醌-2-磺酸(AQS)、蒽醌-2,6-二磺酸(AQDS)和2-羟基-1,4-萘醌(Lawsone)均能显著提高酸性大红3R的脱色速率,其中AQS的促进作用最为明显.研究发现, 0.1mmol/L AQS能够使菌株JL对2.0mmol/L酸性大红3R保持较高的脱色速率,而且能使多种偶氮染料脱色,表现出较好的底物广谱性.利用高效液相色谱-质谱鉴定了AQS介导的酸性大红3R脱色产物,表明酸性大红3R的偶氮键发生断裂, AQS在这一过程中仅起到电子传递的作用.     

壳聚糖铁固定疣孢漆斑菌球处理染料的研究

选择新疆本土产的疣孢漆斑菌作为固定化菌种,并与壳聚糖铁凝胶通过固定化手段有效结合起来,综合分析了壳聚糖铁固定化真菌脱色处理染料的过程机制,以及不同条件下的脱色率.结果表明:固定化真菌对染料的去除首先是染料积累的过程,其次是菌种降解的过程,壳聚糖铁的加入有效缩短了染料积累的时间,24 h的染料脱色率提升了1.5倍;壳聚糖铁固定化真菌对初始浓度为50~250 mg·L-1的酸性染料都有很好的去除效果.振荡培养使疣孢漆斑菌菌球的脱色效率最高提升14.4%.壳聚糖铁固定化菌球投加到染料后有较高的漆酶产量,48 h达到(81.40±3.18)U·mL-1.     

黄杆菌(Flavobacterium sp.)对溴氨酸脱色的研究

从受污土壤中筛选出对溴氨酸具有高效脱色能力的黄杆菌BX26．通过监测液体培养基中细胞浊度和溴氨酸吸光度的变化,考察了碳源浓度、溴氨酸浓度和以溴氨酸为唯一碳、氮、硫源时对菌体生长和溴氨酸脱色的影响．结果表明,BX26对溴氨酸的脱色在于酶的催化作用．该脱色酶属诱导性胞外酶．酶的诱导和产生需要液体培养基中有足量的氮源和硫源,但对碳源没有要求．溴氨酸对菌体的生长具有明显的促进作用,同时菌株也表现出很高的溴氨酸脱色活性,可使高达1000mg/L的溴氨酸降解脱色．菌体不能以溴氨酸为唯一碳、氮、硫源,但可以溴氨酸的脱色产物为唯一碳源．     

一株芽孢杆菌在低氮源浓度培养基中的生长与氨氮去除特性

对菌株YB3进行了16S rRNA基因序列进化分析,并分别以NH_4Cl、NaNO_2、NaNO_3、尿素和蛋白胨为单一氮源,配制了5种低氮源浓度培养基,研究YB3在与养殖水体相近营养水平条件下的生长与氨氮去除特性.结果显示,菌株YB3属于蜡样芽孢杆菌(Bacullis cereus),在5种培养基中均能够生长,菌悬液(吸光度OD600为1.0)接种量为1.0%(v/v)时,OD600由0.010增长到0.100~0.117.在NH_4Cl培养基中,YB3的氨氮去除速率为1.23 mg·L~(-1)·d~(-1),去除率为93.5%.在尿素、蛋白胨等有机氮源培养基中,YB3将首先导致氨氮的积累,累积倍数分别为51.69和3.38,之后开始去除,去除速率为1.56和0.29 mg·L~(-1)·d~(-1),去除率为93.7%和26.8%.结果也表明,提高YB3接种量至8.0%(v/v),可以使蛋白胨培养基氨氮累积倍数下降至2.02,去除速率提高至1.07 mg·L~(-1)·d~(-1),去除率最终达到98.4%.NaNO_2和NaNO_3培养基中均未检测到氨氮,而NH_4Cl、尿素和蛋白胨培养基中也未检测到NO_2~--N和NO_3~--N,表明YB3的硝化、亚硝化和反硝化作用均不强烈,去除氨氮的同时将不会造成NO_2~--N和NO_3~--N等的大量积累.本文为菌株YB3在养殖水体调控与净化中的应用研究提供了实验基础和理论支持.     

1株异养硝化菌胞外聚合物的研究

研究异养硝化菌(Acinetobacter sp.YY-5)在不同生长时期胞外聚合物(EPS)组成成分及其变化规律.利用热处理提取YY-5菌的EPS,分析该菌株在不同生长时期EPS组分,包括蛋白质、总糖、核酸和游离氨基酸等.并进一步把蛋白质水解成氨基酸,分析其氨基酸的变化规律及其对蛋白质理化性质的影响.结果表明,该菌株具有较高含量的EPS,且其在稳定期时EPS含量最高.在不同时期分泌的EPS组分均以蛋白质为主.EPS中各组分含量及其变化规律与该菌生长状态密切相关,在该菌株生长过程中,EPS中各组分的变化情况:蛋白质从14.599 mg.g-1增加到28.489 mg.g-1后减少为15.139 mg.g-1;总糖从6.757 mg.g-1增加到10.199 mg.g-1后减少为7.857 mg.g-1.核酸从1.56 mg.g-1一直增加到6.287 mg.g-1;游离氨基酸从3.713 mg.g-1增加到4.374 mg.g-1后减少为1.299 mg.g-1.蛋白质水解后,其中极性氨基酸呈先增加后减少的趋势,非极性氨基酸一直呈增加趋势;在生长过程中一直以带负电的氨基酸占优.     

变色栓菌对直接大红染料的脱色研究

从野外倒木上分离得到一株对直接染料具有高效脱色能力的白腐真菌CB1,采用形态学与分子生物学鉴定法对菌株CB1进行鉴定.结果表明,菌株CB1为变色栓菌(Trametes versicolor);在固体培养基上,利用T.versicolor CB1对5种直接染料进行脱色,结果显示,T.versicolor CB1对5种直接染料均可脱色,其中对直接大红的脱色效果最为明显;采用单因素分析方法对T.versicolor CB1脱色直接大红的培养条件进行优化,结果显示,以果糖为碳源、硝酸铵为氮源、p H=5、添加Cu SO_4和Ca Cl_2利于T.versicolor CB1对直接大红的脱色;通过正交试验优化T.versicolor CB1对直接大红的脱色条件,优化结果为:果糖10 g·L~(-1)、硝酸铵2 g·L~(-1)、Cu SO_41 mmol·L~(-1)、Ca Cl_23 mmol·L~(-1)、p H=5.优化条件下第7 d时的脱色率为78.23%,与优化前相比,脱色率提高了24.04%.     

Catalyzed degradation of disperse dyes by calcium alginate-pectin entrapped bitter gourd （Momordica charantia） peroxidase

Calcium-alginate pectin entrapped bitter gourd peroxidase (BGP) has been employed for the treatment of disperse dyes: Disperse Brown 1 (DB 1) and Disperse Red 17 (DR 17). Peroxidase alone was unable to decolorize DR 17 and DB 1. However, the investigated dyes were decolorized maximally by BGP in the presence of 0.2 mmol/L redox mediator, violuric acid (VA). A slow decrease in percent decolorization was observed when VA concentration was higher than 0.2 mmol/L which could likely be due to the high reactivity of its aminoxyl radical (> N–O.) intermediate, that might undergo chemical reactions with aromatic amino acid side chains of the enzyme thereby inactivating it. Maximum decolorization of the dyes was observed at pH 3.0 and 40°C within 2 hr of incubation. Immobilized peroxidase decolorized 98% DR 17 and 71% DB 1 using 35 U of BGP in batch process in 90 min. Immobilized enzyme decolorized 85% DR 17 and 51% DB 1 whereas soluble enzyme decolorized DR 17 to 48% and DB 1 to 30% at 60°C. UV-visible spectral analysis was used to evaluate the degradation of these dyes and their toxicity was tested by Allium cepa test. The generally observed higher stability of the bioaffinity bound enzymes against various forms of inactivation may be related to the specific and strong binding of enzyme with bioaffinity support which prevents the unfolding/denaturation of enzyme. Thus entrapped peroxidase was found to be effective in the decolorization of the investigated dyes.     

Biodegradation of Crystal Violet by Agrobacterium radiobacter

Agrobacterium radiobacter MTCC 8161 completely decolorized the Crystal Violet with 8 hr (10 mg/L) at static anoxic conditions. The decreased decolorization capability by A. radiobacter was observed, when the Crystal Violet concentration was increased from 10 to 100 mg/L. Semi-synthetic medium containing 1% yeast extract and 0.1% NH4Cl has shown 100% decolorization of Crystal Violet within 5 hr. A complete degradation of Crystal Violet by A. radiobacter was observed up to 7 cycles of repeated addition (10 mg/L). When the effect of increasing inoculum concentration on decolorization of Crystal Violet (100 mg/L) was studied, maximum decolorization was observed with 15% inoculum concentration. A significant increase in the activities of laccase (184%) and aminopyrine N-demethylase (300%) in cells obtained after decolorization indicated the involvement of these enzymes in decolorization process. The intermediates formed during the degradation of Crystal Violet were analyzed by gas chromatography and mass spectroscopy (GC/MS). It was detected the presence of N,N,N′,N′′-tetramethylpararosaniline, [N; N-dimethylaminophenyl] [N-methylaminophenyl] benzophenone, N; N-dimethylaminobenzaldehyde, 4-methyl amino phenol and phenol. We proposed the hypothetical metabolic pathway of Crystal Violet biodegradation by A. radiobacter. Phytotoxicity and microbial toxicity study showed that Crystal Violet biodegradation metabolites were less toxic to bacteria (A. radiobacter, P. aurugenosa and A. vinelandii) contributing to soil fertility and for four kinds of plants (Sorghum bicolor, Vigna radiata, Lens culinaris and Triticum aestivum) which are most sensitive, fast growing and commonly used in Indian agriculture.     

Biological treatment of chicken feather waste for improved biogas production

A two-stage system was developed which combines the biological degradation of keratin-rich waste with the production of biogas. Chicken feather waste was treated biologically with a recombinant Bacillus megaterium strain showing keratinase activity prior to biogas production. Chopped, autoclaved chicken feathers (4%, W/V) were completely degraded, resulting in a yellowish fermentation broth with a level of 0.51 mg/mL soluble proteins after 8 days of cultivation of the recombinant strain. During the subsequent anaerobic batch digestion experiments, methane production of 0.35 Nm³/kg dry feathers (i.e., 0.4 Nm³/kg volatile solids of feathers), corresponding to 80% of the theoretical value on proteins, was achieved from the feather hydrolyzates, independently of the pre-hydrolysis time period of 1, 2 or 8 days. Cultivation with a native keratinase producing strain, Bacillus licheniformis resulted in only 0.25 mg/mL soluble proteins in the feather hydrolyzate, which then was digested achieving a maximum accumulated methane production of 0.31 Nm³/kg dry feathers. Feather hydrolyzates treated with the wild type B. megaterium produced 0.21 Nm³ CH₄/kg dry feathers as maximum yield.     

堆肥腐殖酸形成及其电化学活性

为探究堆肥腐殖酸形成过程及腐殖酸的氧化还原性能,以城市生活垃圾不同堆肥阶段提取的腐殖酸样品为研究对象,运用现代色谱学技术、光谱学方法和电化学分析,研究了堆肥过程不同形态氮和碳官能团形成腐殖酸的特征,探究了堆肥腐殖酸的氧化还原性能及影响因素. 结果表明:堆肥腐殖酸中50%~75%的氮为氨基酸态氮,检出的15种氨基酸中酸性氨基酸(Asp和Glu)含量(高于100 mg/g)最高、丝氨酸(Ser)含量(3.78 mg/g)最低. 与堆肥腐殖酸中其他组分和官能团相比,氨基酸易被生物降解和利用,其含量随着发酵的进行呈下降趋势,后期氨基酸态氮占比逐渐降低. 腐殖酸中碳有多种形态,包括苯环、羧基、脂肪族等多种官能团,随着堆肥发酵的进行,木质纤维素降解形成腐殖酸,脂肪族官能团亦被氧化降解;与之相反的是,腐殖酸中羧酸类、醛、酮及苯环等官能团不断增加,导致腐殖酸氧化还原性能增强. 氧化和还原过程中,腐殖酸得失电子均会造成部分官能团结构破坏,大分子腐殖酸被降解和转化为小分子有机物. 研究显示,堆肥腐殖酸活性较高,可利用腐殖酸修复环境过程,但需要适当补充活性腐殖酸,防止腐殖酸因修复氧化还原过程导致降解后只能作为碳源.      

玉米根内和根际土壤溶磷细菌生物学特性研究

从玉米根内和根际土壤中分离筛选到22株溶磷细菌。根据16SrRNA基因序列分析,2株溶磷效果最好的菌株分别被鉴定为伯克霍尔德氏菌(Burkholderia sp.SY9)和泛菌(Pantoea sp.Gym7c)。同时对菌株SY9和Gym7c的溶磷能力和植物促生特性进行了进一步研究。利用磷酸钙和开阳磷矿矿粉研究了菌株释放P、Ca和Fe的效能。液体培养条件下,菌株在在磷酸钙培养基中的生长好于在开阳磷矿粉培养基中的生长,同时菌株能够合成铁载体。菌株SY9从开阳磷矿粉中释放P、Ca,和Fe的能力比菌株Gym7c强。然而菌株Gym7c从磷酸钙中释放P和Ca的能力比菌株SY9强。菌株Gym7c主要是通过合成的有机酸溶解磷酸钙并释放其中的P和Ca,而菌株SY9是通过合成的有机酸和铁载体溶解开阳磷矿粉并释放出其中的P、Ca,和Fe。溶解开阳磷矿粉的有机酸主要是葡萄糖酸而溶解磷酸钙的有机酸主要包括丁二酸、丙酸、苹果酸和酒石酸等。菌株Gym7c能够合成生长素、铁载体和1-氨基环丙烷-1-羧酸脱氨酶。菌株SY9和Gym7c对酸碱、盐和温度均表现出一定的抗性。