Biodegradation of oily sludge in Norwegian soils

Summary The degradation rate of oily sludge in soil under Norwegian conditions has been studied in field experiments during a 32 months' period. The experimental plots were added 0, 200, 400, and 600 kg N per ha. In uncultivated soil the oil content was reduced by respectively 4, 9, 22, and 26% during the first 9 months. In the same period the corresponding biodegradation in cultivated soil were 10, 15, 39, and 45%. At the end of the experiment only minor differences between the two different soil types were found. The mean degradation in percent was now 35, 50, 74, and 83, respectively. The optimum temperature for oil degradation in soil was found to be about 18°C. About 2/3 of the optimum activity was retained at 12°C. No leaching of oil or lead through soil columns infiltrated with oily sludge could be observed. A close relationship between oil content of the soil and the rate of water infiltration was found. Due to the extremely high content of lead in this oily sludge, a second application of sludge could not be recommended.     

Comparison of bio-augmentation and composting for remediation of oily sludge: A field-scale study in China

Two bioremediation technologies were performed in order to explore a better treatment process for an oily sludge restoration in China during 2004. The bioremediation by augmentation of biopreparation was compared with a conventional composting. The oily sludge and oil-polluted soil were received from an oil production plant. The total hydrocarbon content (THC) varied from 327.7 to 371.2 g kg⁻¹ of dry sludge and the THC in contaminated soil was 151.0 g kg⁻¹. Before application of preparation, straw, sawdust, top sand and pure soil were added in different proportions to the sludge and soil and mixed thoroughly. Such sludge and soil composites were used for negative controls and for activation of indigenous oil degrading microorganisms with addition of fertilizer (positive controls). For composting, crude manure and straw were added to the oily sludge and the THC was 101.4 g kg⁻¹. The biopreparation was applied every 2 weeks and experiment lasted 56 days under the ambient temperature. The sludge was mixed and watered every 3 days. After three times of biopreparation application, the THC decreased by 46–53% in the oily sludge and soil, while in the positive controls (activation of indigenous microorganisms) the THC decreased by 13–23%, and there was no oil degradation in negative controls After composting, the THC decreased by 31% in the oily sludge. The planting of Tall Fescue (Festuca arundinace) revealed a decrease of sludge toxicity after application of both bioremediation technologies and additionally decreased the THC by 5–7%.     

Biodegradation of Total Petroleum Hydrocarbons from Oily Sludge

The present study concerns the biotreatment of oily sludge of petroleum refineries. Experiments were performed to assess the degradation potential of the microbial species present in the oily sludge through augmentation, and using the augmented microbial inoculums to treat oily sludge in a slurry form containing mineral medium and water. The optimum pH of the slurry was found to be 8 for the biodegradation of oily sludge. The effect of oily sludge concentration, stirring rate, and treatment time on degradation of total petroleum hydrocarbons (TPH) was studied. It was found that the degradation process consists of two consecutive stages and that each stage follows a first-order kinetics. The first stage lasted 4 days followed by a second stage up to 7 days. The faster first stage had a rate constant of 0.1 day^?1, whereas the slower second stage had a rate constant of 0.056 day^?1. The kinetics was found to be time-dependent. The study showed that the mineral medium provided essential nutrients to the microbial species and that the degradation efficiency of the whole microbial species present in the oily sludge was quite high (～90%).     

Degradation of oily sludge from a flotation unit by free and immobilized microorganisms

Summary The degradation rate of hydrocarbons in oily sludge obtained from a flotation unit by free and immobilized cells in shaking flasks and in a stirred tank reactor was investigated. For the biodegration of 3.3% hydrocarbons free cells and cells immobilized on granular clay were used. Free cells needed 7–8 weeks to use 30% of the 3.3% hydrocarbons, whereas with immobilized cells the same result was obtained after 3–4 weeks only. In shaken flasks with high hydrocarbon concentrations (8%), immobilized Candida parapsilosis degraded 90% of the hydrocarbons in the oily sludge within 3 weeks, while free cells degraded only 27.5% in the same period. In degradation experiments with a bioreactor, free and immobilized cells of the isolate ISO-OS BÜ 20 showed better results compared to cultures in shaken flasks due to better aeration and mixing. Free cells degraded 50% of the 5% hydrocarbon-containing oily sludge in 7 weeks, whereas immobilized cells gave the same result after only 4 weeks.Offprint requests to: H.-J. Rehm     

Biodegradation potential of oily sludge by pure and mixed bacterial cultures

The biodegradation capacity of aliphatic and aromatic hydrocarbons of petrochemical oily sludge in liquid medium by a bacterial consortium and five pure bacterial cultures was analyzed. Three bacteria isolated from petrochemical oily sludge, identified as Stenotrophomonas acidaminiphila, Bacillus megaterium and Bacillus cibi, and two bacteria isolated from a soil contaminated by petrochemical waste, identified as Pseudomonas aeruginosa and Bacillus cereus demonstrated efficiency in oily sludge degradation when cultivated during 40 days. The bacterial consortium demonstrated an excellent oily sludge degradation capacity, reducing 90.7% of the aliphatic fraction and 51.8% of the aromatic fraction, as well as biosurfactant production capacity, achieving 39.4% reduction of surface tension of the culture medium and an emulsifying activity of 55.1%. The results indicated that the bacterial consortium has potential to be applied in bioremediation of petrochemical oily sludge contaminated environments, favoring the reduction of environmental passives and increasing industrial productivity.     

Biotreatment of Total Petroleum Hydrocarbons from an oily Sludge Using Co-Composting Approach

Discharging of non-treated oily sludge from oil refineries has undesirable impacts on the environment. In this research, the biotreatment of total petroleum hydrocarbons (TPHs) from Abadan Petroleum Refinery, Iran was done using co-composting method. A 5 kg mixture of oily sludge and compost (oily sludge: compost ratios 1:0, 1:0.1, 1:0.3, 1:0.5, and 1:0.7 w/w) and a bulking agent of wheat straw were used as treatments. All treatments were placed in a wooden box inside a laboratory and aerated every 2–3 days by mixing during two months of the experiments. The variation of TPHs concentration, bacterial density, C/N ratio, pH, and temperature were assessed during the 63 days- experiments. Results showed that the majority of TPHs of samples was removed at first 30 days. The maximum TPHs removal (65%) was obtained in the sludge: compost = 1:0.5 at the operation time of 63 day. The significant different between removal efficiency of oily sludge: compost ratios and the control sample (p value < 0.05) indicated the appropriate density of TPHs degrader-bacteria in treated samples. The highest variation in C/N ratio was observed 15± 0.58 in the oily sludge: compost = 1:0.1. At the beginning of experiments, the pH values of all treatments was alkaline, but became neutral at the end of the reaction time. The TPHs degradation kinetic for all oily sludge: compost ratios followed pseudo second-order model. In general, co-composting has high efficiency in oily sludge remediation and can be potentially applied as a simple and cost-effective approach for remediation of oily sludge.     

不同结构配比的鼠李糖脂乳化活性与油泥清洗效果

以大庆油田原油和含油污泥为对象,研究不同结构配比鼠李糖脂表面活性剂乳化活性及其对含油污泥清洗效果的影响,并优化清洗工艺参数。结果表明:单鼠李糖脂比例越高,其表面活性越好;双鼠李糖脂比例越高,其对原油的乳化能力越强;临界胶束浓度随着双鼠李糖脂比例的增加而逐渐加大;单、双鼠李糖脂配比不同的表活剂对油泥的清洗效果也不同,质量比为50∶50时清洗效率最高;鼠李糖脂浓度为1.0 g·L^-1、热洗时间为1.5h、热洗温度为65℃、洗脱强度为220 r·min^-1、固液质量比为1∶5条件下,油泥的清洗效率最高,可达81.3%;含油率为29.6%的落地油泥,经一级洗涤后油泥残油率降至5.5%,原油回收率达到87.3%,清洗出的原油无明显乳化,易于分离。由此可知,鼠李糖脂的单、双糖脂比例不同对其理化性质和清洗含油污泥的效果均有不同程度的影响。     

Effect of environmental parameters on the biodegradation of oil sludge.

A laboratory study was conducted with the aim of evaluating and optimizing the environmental parameters of "landfarming", i.e., the disposal by biodegradation in soil of oily sludges generated in the refining of crude oil and related operations. Oil sludge biodegradation was monitored by CO2 evolution and by periodic analysis of residual hydrocarbons. The parameters studied were soil moisture, pH, mineral nutrients, micronutrients, organic supplements, treatment rate, teratment frequency, and incubation temperature. Oil sludge biodegradation was optimal at a soil water-holding capacity of 30 to 90%, a pH of 7.5 to 7.8, C:N and C:P ratios of 60:1 and 800:1, respectively, and a temperature of 20 degrees C or above. Addition of micronutrients and organic supplements was not beneficial; sewage sludge interfered with hydrocarbon biodegradation. Breakdown of the saturated hydrocarbon (alkane and cycloalkane) fraction was the highest at low application rates, but higher application rates favored the biodegradation of the aromatic and asphaltic fractions. An application rate of 5% (wt/wt) oil sludge hydrocarbon to the soil (100,000 liters/hectare) achieved a good compromise between high biodegradation rates and efficient land use and resulted in the best overall biodegradation rate of all hydrocarbon classes. Frequent small applications resulted in higher biodegradation than single large applications. Two 100,000-liter/hectare (255 barrels per acre) or four 50,000-liter/hectare oil sludge hydrocarbon applications per growing season seem appropriate for most temperate zone disposal sites.     

Biodegradation Kinetics of Hydrocarbons in Soil during Land Treatment of Oily Sludge

This study focuses on the processes influencing hydrocarbon residue persistence in soil, following land treatment of refinery oily sludge. Treating sludge applied to soil resulted in 70% to 90% degradation of total petroleum hydrocarbon (TPH) during 2 months, regardless of their initial concentrations (9 to 60 g/kg soil). Kinetic analyses performed on TPH degradation, in laboratory and field systems, revealed a degradation pattern characterized by two consecutive first-order kinetics reactions in all experimental settings. The first stage lasted about 3 weeks and was characterized by a temperature dependent rate constant of 0.047 day^-1 at 24°C. That value was comparable to the rate constant obtained when combining the individual rate constants of the saturated, aromatic, asphaltene and polar fractions. The subsequent slower stage rate constant was 0.012 day^-1, insensitive to temperature and to hydrocarbon composition. The transition between the two stages (about 21 days) was independent of the experimental temperature and the biodegradation extent during the first stage. It was concluded that the extent of residual accumulation in the soil was determined by the biodegradation efficiency during the first three weeks of treatment when biological processes dominated. During the following period, abiotic processes leading to reduced bioavailability of the TPH were limiting the degradation rate. Practically, as the first few weeks of treatment determine its efficiency, efforts to enhance the biological activity should be directed to that period.     

Construction of a whole-cell catalyst displaying a fungal lipase for effective treatment of oily wastewaters

To combine the advantage of the oleaginous yeast Yarrowia lipolytica with the high activity of some fungal lipases for oily wastewater treatment, an effective lipase-displaying arming yeast was constructed using the flocculation functional domain of Saccharomyces cerevisiae as the protein anchor. To estimate the effect of the whole-cell oily wastewater treatment, the lipase-displaying arming yeast was added into an open activated sludge bioreactor. Within 72 h of whole-cell treatment, 96.9% of oil and 97.6% of chemical oxygen demand (COD) were removed, while only 87.1% of oil and 91.8% of COD were removed in control A (Y. lipolytica Polg was added), 45.1% of oil and 67.5% of COD were removed in control B (no cell was added) in 72 h. The lipase-displaying arming yeast exhibited remarkable oil removal and COD degradation effect compared with the control samples, exemplifying its application potential.     

Progressive Degradation of Crude Oil n-Alkanes Coupled to Methane Production under Mesophilic and Thermophilic Conditions

Although methanogenic degradation of hydrocarbons has become a well-known process, little is known about which crude oil tend to be degraded at different temperatures and how the microbial community is responded. In this study, we assessed the methanogenic crude oil degradation capacity of oily sludge microbes enriched from the Shengli oilfield under mesophilic and thermophilic conditions. The microbial communities were investigated by terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA genes combined with cloning and sequencing. Enrichment incubation demonstrated the microbial oxidation of crude oil coupled to methane production at 35 and 55°C, which generated 3.7±0.3 and 2.8±0.3 mmol of methane per gram oil, respectively. Gas chromatography-mass spectrometry (GC-MS) analysis revealed that crude oil n-alkanes were obviously degraded, and high molecular weight n-alkanes were preferentially removed over relatively shorter-chain n-alkanes. Phylogenetic analysis revealed the concurrence of acetoclastic Methanosaeta and hydrogenotrophic methanogens but different methanogenic community structures under the two temperature conditions. Candidate divisions of JS1 and WWE 1, Proteobacteria (mainly consisting of Syntrophaceae, Desulfobacteraceae and Syntrophorhabdus) and Firmicutes (mainly consisting of Desulfotomaculum) were supposed to be involved with n-alkane degradation in the mesophilic conditions. By contrast, the different bacterial phylotypes affiliated with Caldisericales, “Shengli Cluster” and Synergistetes dominated the thermophilic consortium, which was most likely to be associated with thermophilic crude oil degradation. This study revealed that the oily sludge in Shengli oilfield harbors diverse uncultured microbes with great potential in methanogenic crude oil degradation over a wide temperature range, which extend our previous understanding of methanogenic degradation of crude oil alkanes.     

Organic Bulking Agents for Enhancing Oil Bioremediation in Soil

Soil contaminated with oil is bioremediated by optimizing conditions for microbial activity. Often the question arises about the benefits of bulking with organic materials to improve soil conditions to enhance degradation of the less biodegradable or less bioavailable components. An investigation was undertaken in the laboratory with the objective of measuring the influence of bulking with dried plant material, bermudagrass, and alfalfa on the degradation of oily sludge added to soil. The oily sludge was diluted 50:50 on a weight basis with soil to achieve a final concentration of 100 g oil and grease kg^-1 of final soil mixture. Bulking agents were added 40 d after dilution of the sludge and optimization of environmental conditions to allow time for the readily decomposable fraction to be degraded before amendment with bulking agents. Populations of heterotrophic microorganisms increased approximately ten times by 40 and 80 d after addition of bulking agents, but the numbers of hydrocarbon-degrading microorganisms did not significantly increase above the number in the nonbulked control. Bulking agents increased the quantity of total petroleum hydrocarbons degraded by approximately 20% during the first 40 d after being added. Disappearance of hydrocarbons for bulked treatments was much slower during the next 40 d, such that the total petroleum hydrocarbon content for both bulked and nonbulked treatments generally was not significantly different at the end. It appears that adding bulking agents may enhance the rate of decomposition of total petroleum hydrocarbons by stimulating the general heterotrophic population of microorganisms, but the influence may not be sustained to influence the extent of decomposition.     

Methanogenic Microbial Community Composition of Oily Sludge and Its Enrichment Amended with Alkanes Incubated for Over 500 Days

Methanogenic microbial community is responsive to the availability of hydrocarbons and such information is critical for the assessment of hydrocarbon degradation in remediation and also in biologically enhanced recovery of energy from non-producing oil reserves. In this study, methanogenic enrichment cultures from oily sludge amended with n-alkanes (C₁₅-C₂₀) showed a development of active methanogenic alkanes-degrading consortium for over a total of 1000 days of incubation at 37°C. Total genomic DNAs were extracted from three types of samples, the original oily sludge (OS), the sludge after incubation for 500 days under methanogenic condition without any external carbon addition (EC), and the enrichment culture from the EC amended with n-alkanes (ET) incubated for another 500 days. The phylogenetic diversities of microbial communities of the three samples were analyzed by PCR amplification of partial 16S rRNA genes. The catabolic genes encoding benzylsuccinate synthase (bssA) and alkylsuccinate synthase (assA) were also examined by PCR amplification. These results provide important evidence in that microbial populations in an oily sludge shifted from methanogenic aromatic compounds degrading communities to potential methanogenic alkane-degrading communities when the enrichment was supplemented with n-alkanes and incubated under anaerobic conditions.     

Influence of 2,4-dinitrophenol on the characteristics of activated sludge in batch reactors

The response of activated sludge characteristics to the presence of 2,4-dinitrophenol (dNP) in batch cultures was investigated in this study. The sludge yield slightly decreased with an increase in dNP concentration. At 10 mg l(-1), or lower, dNP significantly reduced sludge yield and relative specific growth rates (mu/mu0), but didn't substantially affect its relative specific chemical oxygen demand removal rate (q/q0). Presence of dNP at 1-20 mg l(-1) increased the specific oxygen uptake rate of activated sludge, and slightly changed its hydrophobicity. An analysis on inhibition indicated that the reduction in sludge yield in the presence of dNP was mainly attributed to the significant decreased sludge growth, rather than the reduced substrate degradation.     

Biodegradation of diesel oil by an Arabian Sea sediment culture isolated from the vicinity of an oil field

Laboratory scale batch studies were performed to test the diesel oil biodegradation ability of ES1 cultures isolated from Arabian Sea sediments obtained from the vicinity of an oil field. This culture could utilize diesel as the sole source of carbon and energy. Under aerobic conditions, 39% loss of diesel oil was observed over 8 days where 80% of the loss was due to aliphatic constituents. Under anoxic nitrate reducing conditions the rate and extent of degradation was significantly lower, i.e., 18% over 50 days. Salt acclimatized cultures could tolerate salinities up to 3.5% and demonstrated optimal performance at a salinity of 0.5%. The optimum N/P ratio for these cultures was found to be in the range of 2:1-5:1. Addition of two trace elemental substance formulations exhibited a significant inhibitory effect on culture growth. This culture has good potential for decontamination of oil-contaminated marine and subsurface environments.     

Comparison of autochthonous bacteria and commercially available cultures with respect to their effectiveness in fuel oil degradation

Summary This study examined the microbial degradation of fuel oil by nine highly adapted different commercially available mixed bacterial cultures (DBC-plus, Flow Laboratories, Meckenheim, F.R.G.) and a bacterial community from a domestic sewage sludge sample. All mixed cultures were cultivated under aerobic batch conditions shaking (110 rpm) at 20°C in a mineral base medium containing 1 or 5% (v/v) fuel oil as the sole carbon source. Percent degradation of fuel oil and the n-alkane fraction was recorded for the nine DBC-plus cultures and the mixed population of the activated sludge sample. The increase in colony counts, protein, and optical density was studied during a 31-day incubation period for DBC-plus culture A, DBC-plus culture A2 and the activated sludge sample. The activated sludge mixed culture was most effective in degrading fuel oil, but various isolated bacterial strains from this bacterial community were not able to grow on fuel oil as the sole carbon source. In contrast, the n-alkane degradation rates of the DBC-cultures were lower, but single strains from the commercially available mixed cultures were able to mineralize fuel oil hydrocarbons. Strains ofPseudomonas aeruginosa were isolated most frequently and these organisms were able to grow very rapidly on fuel oil as a complex sole carbon source. The results indicate that fuel oil degradation in domestic sewage sludge is performed by mixed populations of naturally occurring bacteria and does not depend on the application of highly adapted commercially available cultures.     

Biodegradation of chrysene by the bacterial strains isolated from oily sludge

The biodegradation studies were conducted to test the ability of the bacterial strains (Chry2 and Chry3) isolated from the oily sludge obtained from Gujarat refinery, India, for utilization of chrysene in the liquid medium. Biodegradation of the compound was confirmed using gas chromatography and the percent degradation was calculated to be 15.0 and 17% by Chry2 and Chry3, respectively. The biodegradation results were supported by increase in viable cell count and dry biomass, in the presence of chrysene as the sole carbon source. Both the cultures produced biosurfactant which was indicated by the reduction in surface tension of the growth medium. Presence of catechol 2, 3-dioxygenase gene in Chry3 indicated its potential for degradation of PAHs through meta cleavage degradation pathway. Both the strains were found to possess catechol 1,2-dioxygenase and catechol 2,3-dioxygenase enzyme activities. Based on morphological and biochemical tests, the cultures were tentatively identified as Bacillus sp. (Chry2) and Pseudomonas sp. (Chry3).     

Anaerobic microbial associations degrading aromatic amino acids

Anaerobic microbial associations have been isolated that degrade aromatic amino acids to methane and carbon dioxide at high rates. Significant differences between the morphological, cytological, and physiological traits of cultures isolated from samples of adapted and unadapted sludge are shown. The effects of cultivation temperature, illumination, and presence of mineral nitrogen and bicarbonate in the medium upon adaptation of enrichment cultures to substrates and subsequent behavior of the anaerobic associations have been studied. Intermediate and final products of degradation of aminoaromatic compounds and the sequence of their formation in the cultures have been determined. We have also studied the effects of exogenous electron acceptors and additional carbon sources on the degradation of aminoaromatic compounds.     

Screening of herbaceous plants for peat-enhanced rehabilitation of contaminated soil with oily sludge

A batch pot experiment using nine herbaceous species were conducted for peat enhanced rehabilitation of contaminated soil with oily sludge in the initial contents of 0%, 1.3%, 7.4%, and 12.2%, respectively. The results showed that petroleum hydrocarbons removal, plant growth indices and enzyme activities varied depending on plant species and oil contents. Cotton, ryegrass and tall fescue were effective in the rehabilitation of oily sludge contaminated soils. The total petroleum hydrocarbon (TPH) removal ranged from 30.0% to 40.0% after 170 days of treatment. Plant biomass was shown to be the preferred indicator for screening phytoremediation plant because it was closely correlated with TPH removal and enzyme activities. Peat-enhanced plant rehabilitation could be a good strategy for the treatment of oily sludge contaminated saline soils.     

The response of maize (Zea mays L.) plant assisted with bacterial consortium and fertilizer under oily sludge

The objective of this study was to evaluate the role of PGPR consortium and fertilizer alone and in combination on the physiology of maize grown under oily sludge stress environment as well on the soil nutrient status. Consortium was prepared from Bacillus cereus (Acc KR232400), Bacillus altitudinis (Acc KF859970), Comamonas (Delftia) belonging to family Comamonadacea (Acc KF859971) and Stenotrophomonasmaltophilia (Acc KF859973). The experiment was conducted in pots with complete randomized design with four replicates and kept in field. Oily sludge was mixed in ml and Ammonium nitrate and Diammonium phosphate (DAP) were added at 70 ug/g and 7ug/g at sowing. The plant was harvested at 21 d for estimation of protein, proline and antioxidant enzymes superoxide dismutase (SOD) and peroxidase (POD). To study the degradation, total petroleum hydrocarbon was extracted by soxhelt extraction and extract was analyzed by GC-FID at different period after incubation. Combined application of consortium and fertilizer enhanced the germination %, protein and, proline content by 90,130 and 99% higher than untreated maize plants. Bioavailability of macro and micro nutrient was also enhanced with consortium and fertilizer in oily sludge. The consortium and fertilizer in combined treatment decreased the superoxide dismutase (SOD), peroxidase dismutase (POD) of the maize leaves grown in oily sludge. Degradation of total petroleum hydrocarbon (TPHs) was 59% higher in combined application of consortium and fertilizer than untreated maize at 3 d. The bacterial consortium can enhanced the maize tolerance to oily sludge and enhanced degradation of total petroleum hydrocarbon (TPHs). The maize can be considered as tolerant plant species to remediate oily sludge contaminated soils.