Influence of microbial biomass on the biodegradability of organic compounds

The influence of different types of inocula as well as the amount of inoculum (microbial biomass) on the biodegradation pattern of acetate, 4-nitrophenol, and the three methoxyaniline isomers was investigated in the Modified OECD test and a NPR guidline. Using sediment of the river Rhine as inoculum 4-nitrophenol could not be degraded, while an inoculum from garden soil gave only 60% degradation in the OECD test. Effluent of an activated sludge plant however was able to degrade 4-nitrophenol at a concentration of 19 mg/1 in the OECD test completely. At a concentration of 94 mg/1 no degradation was observed. Testing the methoxyanilines for biodegradability it was found that at a low inoculum level (OECD protocol) no degradation of the three compounds occurred. Using activated sludge (1.5 ml/1) as inoculum 3- and 4-methoxy aniline could be degraded for 60% respectively completely while 2-methoxyaniline was still refactory to degradation. Measuring the microbial biomass by means of ATP during biodegradation strongly suggested that the microbial flora which rapidly metabolize acetate is quite another microflora than the microflora responsible for the degradation of 4-nitrophenol.     

生物降解实验中的接种物标准化研究

针对降解实验中接种物采集困难、活性差异大、实验可重复性及结果可比性差的问题展开接种物标准化研究。对多来源的接种物在实验室可控条件下进行驯养,驯养后生长状态良好的活性污泥分别固定于灭菌麸皮和藻酸钙凝胶等2种微生物载体上,形成2种固定化接种物(分别简称为Bran及Ball)。对上述固定化接种物进行微生物活性、适应性、降解能力以及稳定性研究:通过2种固定化接种物与商品化接种物Polyseed对葡萄糖-谷氨酸(GGA)五日生化需氧量(BOD5)的结果与活菌细胞数的关系,确认3种接种物在降解实验中的用量可控制在相同数量级;比较不同批次及保存时间的2种固定化接种物、相同批次不同保存时间的Polyseed以及污水处理厂曝气池污泥为接种物对苯胺及二甘醇的快速生物降解实验(OECD301F)结果,发现Ball及Bran较Polyseed及新鲜采集活性污泥的微生物适应性及降解能力更佳。此外,在较长保存期内2种固定化接种物活性相对稳定。2种固定化接种物初具成为标准化接种物的必要条件。     

Degradation of inhibitory substances by heterotrophic microorganisms during bioleaching of heavy metals from anaerobically digested sewage sludge

The presence of organic acids was found to be inhibitory to the bioleaching of sewage sludge and the objective of the present study was to elucidate the roles of heterotrophic microorganisms in removing organic acids during the bioleaching of heavy metals from anaerobically digested sewage sludge. Microbiological analysis showed that acetic and propionic acids posed a severe inhibitory effect on iron-oxidizing bacteria as reflected by a sharp decrease in their viable counts in the first 4d and it only started to increase 2d after the depletion of both acids. Biodegradation of these inhibitory organic acids was revealed by sharp increases in total fungi and acidophiles between day 3 and day 5 which coincided with degradation of organic acids. This was further confirmed by the increases in total counts of both acetate and propionate degraders in the same period. Two yeast strains Y4 and Y5 with strong ability to degrade acetate and/or propionate were isolated and identified as Pichia sp. and Blastoschizomycetes capitatus, respectively. B. capitatus Y5 was an more important player in removing the inhibitory organic acids during the bioleaching process since it could utilize both acetate and propionate as sole carbon source while Pichia sp. Y4 was an strict acetate degrader. Results from the present study not only provided the evidence for biodegradation of organic acids by heterotrophs, but also disclosed a biological mechanism for the initiation of bioleaching of organic acid-laden sewage sludge.     

Use of acid preconditioning for enhanced dewatering of wastewater treatment sludges from the pulp and paper industry.

In municipal and industrial practices, wastewater treatment sludges are generally conditioned with organic polymers before dewatering. The dewatering polymers are expensive and contribute significantly to the overall sludge management cost. This paper discusses a preconditioning strategy that holds great promise for enhancing dewatering properties of wastewater treatment sludges, while reducing the cost. In this approach, the waste activated sludge (WAS) is briefly preconditioned with an acid before flocculating with an organic polymer. Experimental results showed that acid preconditioning significantly enhanced dewatering. Separately acidifying WAS and subsequently combining it with primary sludge produced higher presscake solids than acidifying the combined sludge to the same final pH. Acidification exhibited the added benefit of reducing Escherichia coli counts in sludge, thus improving its biological character. This may provide flexibility in choosing a beneficial use application.     

Assessing microbial communities for a metabolic profile similar to activated sludge.

To search for reliable testing inocula alternatives to activated sludge cultures, several model microbial consortia were compared with activated sludge populations for their functional diversity. The evaluation of the metabolic potential of these mixed inocula was performed using the Biolog EcoPlates and GN and GP MicroPlates (Biolog, Inc., Hayward, California). The community-level physiological profiles (CLPPs) obtained for model communities and activated sludge samples were analyzed by principal component analysis and hierarchic clustering methods, to evaluate the ability of Biolog plates to distinguish among the different microbial communities. The effect of different inocula preparation methodologies on the community structure was also studied. The CLPPs obtained with EcoPlates and GN MicroPlates showed that EcoPlates are suitable to screen communities with a metabolic profile similar to activated sludge. New, well-defined, standardized, and safe inocula presenting the same metabolic community profile as activated sludge were selected and can be tested as surrogate cultures in activated-sludge-based bioassays.     

Anaerobic inhibition and biodegradation of antibiotics in ISO test schemes

Municipal sewage is the main exposure route for antibiotics that are used in human medical care. Antibiotics that adsorb to the primary sludge and/or sur-plus activated sludge will enter the anaerobic digesters of municipal sewage treatment plants. Here anaerobic biodegradation or inhibition of anaerobic bacteria resulting in a disturbance of the process might occur. ISO standards 13641 (2003) and 11734 (1999) were used for assessing the anaerobic inhibition of 16 and the anaerobic biodegradability of 9 antibiotics respectively. Digestion sludge from a municipal sewage treatment plant (1g/l d.s.) was used as inoculum in both tests. In ISO 13641 (2003) most antibiotics showed only moderate inhibition effects after a 7 day incubation period, with EC50 values between 24 mg/l and more than 1000 mg/l (equal to mg/g d.s.). In contrast, metronidazol was decisively toxic to anaerobic bacteria with an EC50 of 0.7 mg/l. In the anaerobic degradation tests according to ISO standard 11734 (1995), only benzylpenicillin showed certain ultimate biodegradation after 60 days and most antibiotics inhibited the digesting sludge in the respective parallel tested inhibition controls. Thus the inhibition of anaerobic bacteria by antibiotics observed in the degradation tests was higher than expected from the results of the inhibition tests. The possible explanations are that distinct substrates are used (yeast extract versus sodium benzoate), that the digestion sludge loses activity during the washing steps performed for the degradation tests and that the exposure time in the degradation tests was 8 times longer than in the inhibition test.     

4种脂肪酸浮选捕收剂的生物降解性能研究

在好氧的条件下,以活性污泥为接种体,使用改良斯特姆法测试脂肪酸浮选捕收剂(油酸钠、9-羟基硬脂酸钠、硬脂酸钠、蓖麻油酸钠)的生物降解性能,并与利用BOD5/COD进行的可生化评价方法进行比较。结果表明:(1)油酸钠、蓖麻油酸钠的BOD5/COD分别为0.54、0.63,属易生物降解物质;9-羟基硬脂酸钠、硬脂酸钠的BOD5/COD分别为0.36、0.37,属可生物降解物质。(2)油酸钠、9-羟基硬脂酸钠、蓖麻油酸钠28d的生物降解率分别为88.6%、62.2%、100.0%,达到了经济合作与发展组织的OECD-301B生物降解性能实验标准,属于易生物降解物质;硬脂酸钠28d的生物降解率为49.8%,没有达到OECD-301B生物降解性能实验标准,属难生物降解物质。4种脂肪酸浮选捕收剂的生物降解性能依次为:蓖麻油酸钠油酸钠9-羟基硬脂酸钠硬脂酸钠。(3)当有机酸中存在碳碳双键和羟基时,有机酸的生物降解性能都会提高,并且碳碳双键的影响要大于羟基的影响;当二者同时存在时,生物降解性能会大幅度提高。     

A method for measuring the anoxic biodegradability under denitrifying conditions

A test for assessing the anoxic biodegradability of organic compounds under denitrifying conditions is proposed. The method is based on the recovery and quantification of the CO2 produced, which is evidence of complete biodegradation of the test compound (added as the sole carbon source). The tests were carried out in a mineral medium, with nitrate as electron acceptor. Whole lake sediments, sediment extracts and a commercial inoculum were assayed as a possible inoculum source by means of glucose biodegradability tests. It was found that the sediment extracts constitute a suitable and environmentally-relevant inoculum source, since they add non-significant amounts of carbon to the tests. Two xenobiotic compounds, namely, aniline and phenol, were tested in the aforementioned conditions as well as in a standard aerobic biodegradability test. Both aniline and phenol attained a biodegradation level higher than 60% in a short time period (<28 days) and thus can be considered as readily biodegradable in denitrifying environments. Nevertheless, the kinetics obtained in the anoxic test were slower than in aerobic conditions, and even suggested the accumulation of intermediate metabolites in the case of phenol. The results of this study indicate that the fate of xenobiotic compounds under anoxic conditions differs from that observed in an oxic environment, and therefore it should be considered by standard biodegradability testing procedures.     

Xenobiotic benzotriazoles—biodegradation under meso- and oligotrophic conditions as well as denitrifying,sulfate-reducing,and anaerobic conditions

The intensive use of benzotriazoles as corrosion inhibitors for various applications and their application in dishwasher detergents result in an almost omnipresence of benzotriazole (BTri), 4-methyl- and 5-methyl-benzotriazole (4-TTri and 5-TTri, respectively) in aquatic systems. This study aims on the evaluation of the biodegradation potential of activated sludge communities (ASCs) toward the three benzotriazoles regarding aerobic, anoxic, and anaerobic conditions and different nutrients. ASCs were taken from three wastewater treatment plants with different technologies, namely, a membrane bioreactor (MBR-MH), a conventional activated sludge plant CAS-E (intermittent nitrification/denitrification), and CAS-M (two-stage activated sludge treatment) and used for inoculation of biodegradation setups. All ASCs eliminated up to 30 mg L^?1 5-TTri and BTri under aerobic conditions within 2–7 and 21–49 days, respectively, but not under anoxic or anaerobic conditions. 4-TTri was refractory at all conditions tested. Significant differences were observed for BTri biodegradation with non-acclimated ASCs from MBR-MH with 21 days, CAS-E with 41 days, and CAS-M with 49 days. Acclimated ASCs removed BTri within 7 days. Furthermore, different carbon and nitrogen concentrations revealed that nitrogen was implicitly required for biodegradation while carbon showed no such effect. The fastest biodegradation occurred for 5-TTri with no need for acclimatization, followed by BTri. BTri showed sludge-specific biodegradation patterns, but, after sludge acclimation, was removed with the same pattern, regardless of the sludge used. Under anaerobic conditions in the presence of different electron acceptors, none of the three compounds showed biological removal. Thus, presumably, aerobic biodegradation is the major removal mechanism in aquatic systems.     

Exploring the potential of applying proteomics for tracking bisphenol A and nonylphenol degradation in activated sludge

A significant percentage of bisphenol A and nonylphenol removal in municipal wastewater treatment plants relies on biodegradation. Nonetheless, incomplete information is available concerning their degradation pathways performed by microbial communities in activated sludge systems. Hydroquinone dioxygenase (HQDO) is a specific degradation marker enzyme, involved in bisphenol A and nonylphenol biodegradation, and it can be produced by axenic cultures of the bacterium Sphingomonas sp. strain TTNP3. Proteomics, a technique based on the analysis of microbial community proteins, was applied to this strain. The bacterium proteome map was obtained and a HQDO subunit was successfully identified. Additionally, the reliability of the applied proteomics protocol was evaluated in activated sludge samples. Proteins belonging to Sphingomonas were searched at decreasing biomass ratios, i.e. serially diluting the bacterium in activated sludge. The protein patterns were compared and Sphingomonas proteins were discriminated against the ones from sludge itself on 2D-gels. The detection limit of the applied protocol was defined as 10^?3 g TTNP3 g^?1 total suspended solids (TSSs). The results proved that proteomics can be a promising methodology to assess the presence of specific enzymes in activated sludge samples, however improvements of its sensitivity are still needed.     

废水产氢产酸/同型产乙酸耦合产酸条件优化

为了更好地发挥产氢产酸/同型产乙酸耦合系统在废水厌氧发酵生产乙酸方面的优势,有必要寻找一种简单有效的方法以获得该系统产酸的优化条件.利用经过加热处理并活化的厌氧污泥作种泥,以模拟废水中的葡萄糖为底物,针对发酵时间、底物浓度、种泥浓度、初始pH进行4因素10水平均匀设计实验,得到了乙酸生产指标与产酸条件之间关系的回归方程;也得到了以高乙酸产量为主要目标导向同时兼顾高乙酸产率和高乙酸生产强度目标的优化条件;优化条件实验乙酸浓度比均匀设计中最高乙酸浓度提高20%左右.研究表明,将均匀设计应用于废水产氢产酸/同型产乙酸耦合产酸条件优化,可以避免盲目性,迅速获得满意结果.     

A practical method for quantification of phosphorus- and glycogen-accumulating organism populations in activated sludge systems.

Enhanced biological phosphorus removal (EBPR) from wastewater relies on the enrichment of activated sludge with phosphorus-accumulating organisms (PAOs). The presence and proliferation of glycogen-accumulating organisms (GAOs), which compete for substrate with PAOs, may be detrimental for EBPR systems, leading to deterioration and, in extreme cases, failure of the process. Therefore, from both process evaluation and modeling perspectives, the estimation of PAO and GAO populations in activated sludge systems is a relevant issue. A simple method for the quantification of PAO and GAO population fractions in activated sludge systems is presented in this paper. To develop such a method, the activity observed in anaerobic batch tests executed with different PAO/GAO ratios, by mixing highly enriched PAO and GAO cultures, was studied. Strong correlations between PAO/GAO population ratios and biomass activity were observed (R2 > 0.97). This served as a basis for the proposal of a simple and practical method to quantify the PAO and GAO populations in activated sludge systems, based on commonly measured and reliable analytical parameters (i.e., mixed liquor suspended solids, acetate, and orthophosphate) without requiring molecular techniques. This method relies on the estimation of the total active biomass population under anaerobic conditions (PAO plus GAO populations), by measuring the maximum acetate uptake rate in the presence of excess acetate. Later, the PAO and GAO populations present in the activated sludge system can be estimated, by taking into account the PAO/GAO ratio calculated on the basis of the anaerobic phosphorus release-to-acetate consumed ratio. The proposed method was evaluated using activated sludge from municipal wastewater treatment plants. The results from the quantification performed following the proposed method were compared with direct population estimations carried out with fluorescence in situ hybridization analysis (determining Candidatus Accumulibacter Phosphatis as PAO and Candidatus Competibacter Phosphatis as GAO). The method showed to be potentially suitable to estimate the PAO and GAO populations regarding the total PAO-GAO biomass. It could be used, not only to evaluate the performance of EBPR systems, but also in the calibration of potential activated sludge mathematical models, regarding the PAO-GAO coexistence.     

Comparison of mechanical pretreatment methods for the enhancement of anaerobic digestion of pulp and paper waste activated sludge

The conventional anaerobic digestion process, requiring long solids retention times (SRTs) to digest solids, is currently viewed as impractical for the pulp and paper industry because of high capital costs associated with the construction of new digesters. Recent developments in sludge solubilization technology could be promising in reducing digester size, which also allows for the potential use of decommissioned tanks, both of which can reduce the capital cost. Three pretreatment technologies for use with anaerobic digestion were tested on laboratory-scale to investigate their feasibility. The SRTs in all three digesters systematically decreased from 20 to 3 days. The reference digester was fed waste activated sludge (WAS) to serve as the control at the same SRTs. The other digesters were fed WAS that had been preconditioned using mechanical shearing, sonication, or high-pressure homogenization technology. Anaerobic digestion with high-pressure homogenization produced as much methane at 3-day mean SRT as that from the reference digester operated at 20-day SRT. Therefore, a new digester can theoretically be 85% smaller than a conventional digester. An added benefit of WAS to methane conversion is the recovery of nutrients nitrogen and phosphorus.     

Biodegradation of poly(propylene glycol)s under the conditions of the OECD screening test

Two poly(propylene glycol)s (PPGs): PPG 425 and PPG 725 were tested under the conditions of the OECD Screening Test with activated sludge as inoculum. Tested PPG were the sole source of organic carbon in the test. Quantitative determination of the biodegradation progress was performed by the HPLC with fluorescence detection after derivatisation of PPG with naphthyl isocyanate. The liquid chromatography-mass spectrometry technique was used for identification and semiquantitative determination of metabolites. Separation of PPG and metabolites from the water matrix was performed by liquid-liquid extraction with chloroform. It was found that the shorter PPG 425 is biodegraded significantly worse than the longer PPG 725 and that biodegradation occurs without shortening of the PPG chain for both PPG. PPG molecules are oxidised to ketones and/or aldehydes during the aerobic biodegradation process.     

Investigation of triclosan fate and toxicity in continuous-flow activated sludge systems

The purpose of this research was to study the fate and toxicity of triclosan (TCS) in activated sludge systems and to investigate the role of biodegradation and sorption on its removal. Two continuous-flow activated sludge systems were used; one system was used as a control, while the other received TCS concentrations equal to 0.5 and 2mgl(-1). At the end of the experiment, 1mgl(-1) TCS was added in the control system to investigate TCS behaviour and effects on non-acclimatized biomass. For all concentrations tested, more than 90% of the added TCS was removed during the activated sludge process. Determination of TCS in the dissolved and particulate phase and calculation of its mass flux revealed that TCS was mainly biodegraded. Activated sludge ability to biodegrade TCS depended on biomass acclimatization and resulted in a mean biodegradation of 97%. Experiments with batch and continuous-flow systems revealed that TCS is rapidly sorbed on the suspended solids and afterwards, direct biodegradation of sorbed TCS is performed. Regarding TCS effects on activated sludge process, addition of 0.5mgl(-1) TCS on non-acclimatized biomass initially deteriorated ammonia removal and nitrification capacity. After acclimatization of biomass, nitrification was fully recovered and further increase of TCS to 2mgl(-1) did not affect the performance of activated sludge system. The effect of TCS on organic substrate removal was minor for concentrations up to 2mgl(-1), indicating that heterotrophic microorganisms are less sensitive to TCS than nitrifiers.     

共代谢基质对苯酚降解菌XTT-3降酚作用的影响

采用驯化的方法从活性污泥中分离到一株苯酚降解菌XTT-3,经16SrDNA鉴定为Sphingobiumsp.。对该菌株进行碳饥饿处理,发现其降解苯酚的能力受到抑制。以只含苯酚的M9培养基为参照,添加0.2g/L酵母膏作为共代谢基质,对XTT-3菌株降解苯酚有较明显的促进作用,36h后苯酚降解率为68%。在含0.2g/L酵母膏的M9培养基中,同时添加20mg/L邻苯二酚,XTT-3降解苯酚作用显著增加,24h后苯酚降解率达75%,苯酚降解速度达0.261mg/min。     

Effect of activated sludge properties and membrane operation conditions on fouling characteristics in membrane bioreactors

Biofouling control is considered to be a major challenge in operating membrane bioreactors (MBRs) for the treatment of wastewater. This study examined the impact of biological, chemical, and physical properties of activated sludge on membrane filtration performance in laboratory-scale MBRs. Sludges with different microbial communities were produced using pseudo-continuous stirred-tank reactors and pseudo-plug flow reactors treating a synthetic paper mill wastewater. Various filtration resistances were used to investigate membrane fouling characteristics, and molecular biology tools targeting 16S ribosomal DNA gene sequences were used to identify predominant bacterial populations in the sludges or attached to the fouled membranes. Filtration experiments using axenic cultures of Escherichia coli, Acinetobacter calcoaceticus, and Gordonia amarae were also performed to better understand the initiation and development of biofouling. The results showed that the tendency of membranes to biofoul depended upon membrane operating conditions as well as the properties of the activated sludge in the MBR systems. Specific bacterial populations, which were not dominant in the activated sludges, were selectively accumulated on the membrane surface leading to the development of irreversible biofouling.     

Aerobic biodegradation of organotin compounds in activated sludge batch reactors

The biodegradation behavior of four organotin (OT) compounds, namely tributyltin (TBT), dibutyltin (DBT), monobutyltin (MBT) and triphenyltin (TPhT), was studied in lab-scale activated sludge batch reactors. The activated sludge was spiked with the OT compounds at a level of 100 microg l(-1) as Sn. Determination of the OT compounds by GC-FPD after ethylation in the dissolved and particulate phase revealed that 24 h after the start of the experiments, almost the total of OT compounds has been removed from the dissolved phase and is associated with the suspended solids. Calculation of mass balance in batch reactors showed that OT compounds biodegradation was performed via a sequential dealkylation process. Removals due to biodegradation were differentiated according to the parent compound. In experiments with non-acclimatized biomass, a percentage of 27.1, 8.3, 73.8 and 51.3 was still present as TBT, DBT, MBT and TPhT, respectively, at the end of the experiment (18th day). Half-lives (t1/2) of 10.2 and 5.1 days were calculated for TBT and DBT, respectively, whereas apparent t1/2 values could not be determined for MBT and TPhT (t1/2>18 days). The capacity of activated sludge to biodegrade OT compounds in the absence of supplemental substrate indicated that these compounds can be metabolized as single sources of carbon and energy in activated sludge systems. Excluding TBT, the presence of low concentrations of supplemental substrate did not affect the biodegradation potential of activated sludge. The acclimatization of biomass on OT compounds enhanced significantly biodegradation, resulting in significant decreases of half-lives of OT compounds. As a result in the presence of acclimatized biomass, half-lives of 1.4, 3.6, 9.8 and 5.0 days were calculated for TBT, DBT, MBT and TPhT, respectively.     

SBBR挂膜前活性污泥培养与驯化的研究

在序批式生物膜反应器（SBBR）挂膜前驯化活性污泥对养猪场废水进行生物降解实验。结果表明,在驯化期结束时,COD和NH₃-N的去除率分别达到85%和74%。对驯化过程中出现的污泥膨胀、上浮、泡沫等异常现象进行了分析,并提出了相应的解决措施。     

反硝化条件下有机碳低丰度河床沉积层中的苯胺降解

采集渭河河床沉积物,研究了反硝化条件下有机碳低丰度的河床沉积层中苯胺降解.结果表明,反硝化条件下,苯胺在有机碳低丰度的河床沉积层中可生物降解.使该环境中苯胺(约50 mg/L)降解近95%,当硝酸盐为30.69、184.16、245.54 mg/L时,降解时间分别约为20、45、70 d.在上述环境中另加35.98 mg/L乙酸盐后,苯胺降解速度在硝酸盐为184.16 mg/L时最大,硝酸盐为30.69 mg/L时最小.当硝酸盐为30.69 mg/L,不加乙酸盐,27 d苯胺降解约95%;添加35.98 mg/L乙酸盐后,实验进行了47 d还仍有近13 mg/L苯胺残留,说明外加碳源(乙酸盐)对苯胺降解具有抑制作用.但当硝酸盐为184.16、245.54 mg/L时,外加碳源(乙酸盐)则强化苯胺降解.水合金属氧化物对苯胺降解具有促进作用.