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Large quantities of oily wastewater are produced from the activities and processes in the petroleum industry, and draining of these effluents not only pollutes the environment but also reduces the yield of oil and water. This chapter gives a general introduction to the control and treatment of water emissions in the petroleum industry. It begins with an overview of wastewaters and management, followed by wastewater characterization, selection of oil–water separation and treatment technologies, and wastewater treatment (process wastewater pretreatment, primary treatment, secondary treatment, and tertiary treatment or polishing). Primary wastewater treatment includes separation of oil, water, and solids in two stages. In the first stage, oil–water separators are used to remove large quantities of free oil and heavy suspended solids from wastewater and are usually used when oil concentrations in the raw wastewater exceed approximately 500 mg/L. The second stage of primary treatment is designed to remove small oil droplets and suspended solids, oil emulsions, and oil-wetted solids that have not been separated in the first stage of primary treatment. In secondary treatment, dissolved oil and other organic pollutants are sometimes consumed by microorganisms. Biological-treatment processes can generally be classified as suspended growth processes such as activated sludge (AS) processes, sequencing batch reactors (SBRs), continuous stirred tank bioreactors (CSTBs), membrane bioreactors (MBRs), and aerated lagoons, and attached growth processes such as trickling filters (TFs), fluidized bed bioreactors (FBBs), and rotating biological contactors (RBCs). Tertiary treatment or polishing refers to any treatment that takes place downstream of the secondary treatment plant to obtain a high-quality effluent to meet discharge limits or possibly for reuse. Sand filtration, activated carbon adsorption, chemical oxidation, pressure-driven membrane-separation technologies (e.g., microfiltration (MF), ultrafiltration (UF), nanofiltration (NF), reverse osmosis (RO)), other advanced wastewater-treatment methods such as ion-exchange, electrodialysis (ED) and electrodialysis reversal (EDR), and advanced oxidation processes (AOPs) (e.g., hydrogen-peroxide/ultraviolet (H 2 O 2 /UV), ozonation process, Fenton and photo-Fenton processes, heterogeneous photocatalysis, electrochemical oxidation, wet-air oxidation (WAO), and supercritical water oxidation (SCWO)) are also discussed. Finally, wastewater-treatment plants (WWTPs) in the petroleum industry are summarized.

Rotating biological contactors (RBCs) for wastewater treatment began in the 1970s. Removal of organic matter has been targeted within organic loading rates of up to 120 g m−2 d−1 with an optimum at around 15 g m−2 d−1 for combined BOD and ammonia removal. Full nitrification is achievable under appropriate process conditions with oxidation rates of up to 6 g m−2 d−1 reported for municipal wastewater. The RBC process has been adapted for denitrification with reported removal rates of up to 14 g m−2 d−1 with nitrogen rich wastewaters. Different media types can be used to improve organic/nitrogen loading rates through selecting for different bacterial groups. The RBC has been applied with only limited success for enhanced biological phosphorus removal and attained up to 70% total phosphorus removal. Compared to other biofilm processes, RBCs had 35% lower energy costs than trickling filters but higher demand than wetland systems. However, the land footprint for the same treatment is lower than these alternatives. The RBC process has been used for removal of priority pollutants such as pharmaceuticals and personal care products. The RBC system has been shown to eliminate 99% of faecal coliforms and the majority of other wastewater pathogens. Novel RBC reactors include systems for energy generation such as algae, methane production and microbial fuel cells for direct current generation. Issues such as scale up remain challenging for the future application of RBC technology and topics such as phosphorus removal and denitrification still require further research. High volumetric removal rate, solids retention, low footprint, hydraulic residence times are characteristics of RBCs. The RBC is therefore an ideal candidate for hybrid processes for upgrading works maximising efficiency of existing infrastructure and minimising energy consumption for nutrient removal. This review will provide a link between disciplines and discuss recent developments in RBC research and comparison of recent process designs are provided (Section 2 ). The microbial features of the RBC biofilm are highlighted (Section 3 ) and topics such as biological nitrogen removal and priority pollutant remediation are discussed (Sections 4 Biological nutrient removal in RBCs , 5 Priority pollutant remediation in RBCs ). Developments in kinetics and modelling are highlighted (Section 6 ) and future research themes are mentioned.

Population growth in urban areas is leading to an expansion in industrial development, which in turn creates increased volumes of wastewater with ever more complex compositions. The wastewater that is generated is discharged into water sources that supply drinking water to other urban areas, causing higher levels of pollution. One option for the treatment of water and wastewater is the use of biological processes, which offer certain advantages over other common treatment technologies. The organic contaminants to be destroyed are used and transformed by bacteria or other organisms as a source of food. Thus the water/wastewater treatment process is used to dispose of waste in a non-toxic and sanitary manner. This chapter discusses the major biological options for water and wastewater treatment.

The removal of nitrate from a mature landfill leachate with high nitrate load in a lab-scale anoxic rotating biological contactor (RBC) was studied. Under a phosphorus-phosphate concentration of 10mg P-PO4 3− L−1 and nitrogen-nitrate concentrations above 530mgN-NO3 − L−1 the reactor achieved nitrogennitrate removal efficiencies close to 100%, without nitrite or nitrous oxide accumulation. Although the reactor presented a very good denitrification performance, the effluent carbon concentration was still above the legal discharge value. In order to increase the biodegradability of the leachate recalcitrant carbon load, a pre-ozonation was further investigated. The pre-ozonation led to a total organic carbon (TOC) removal of 28%. The sequence of treatments, leachate ozonation followed by RBC denitrification did not affect the denitrification efficiency. In fact, it was possible to attain a denitrification rate of 123mg N-NO3 − L−1 h−1. The moderate decrease in the carbon load of the final effluent indicated that some recalcitrant compounds were still present after ozonation. The anoxic RBC showed to be a promising technology for removing nitrate from landfill leachate., Fundação para a Ciência e a Tecnologia (FCT)

Current environmental legislation is focused in removal and/or reducing emissions of pollutants. Biological treatments of wastes are considered as an alternative opposite traditional physicochemical methods. In recent decades, the use of biofilters to removal of contaminants from wastewater and waste gases is being developed. Biofilters use microorganisms, which are capable of degrading many compounds, fixed to an inorganic/organic medium (carrier) to break down pollutants present in a fluid stream. In this article, basic aspects about biofilter configuration, filter media, microorganisms involved, and properties that affect the biofilter performance are presented. Once these aspects are clarified, work deals with the biofilter design. Efforts are focused to summarize the terminology, operational features, and design equations that are essential for this kind of bioreactors. In the design, treatments of liquid and gaseous waste streams, due to the singular differentiating characteristics of these equipments, are distinguished.

Water treatment has become a source of concern as new pollutants and higher volumes of waste water must be treated. Emerging biological approaches, namely the use of bioreactors, for cleaning processes have been introduced. The use of bioreactors requires the development of efficient monitoring tools, preferably with real-time measurements. In this work, a couple of flow injection systems were developed and optimized for the potentiometric determination of fluoride to monitor a rotating biological contactor (RBC) bioreactor and a sequencing batch reactor (SBR) with off-line and on-line sampling. Both the RBC and the SBR bioreactors were set up for the biodegradation of the halogenated organic compound 2-fluorophenol and, as fluoride was a degradation byproduct, the process was monitored by following up its concentration. The described flow injection potentiometric methods enabled the fluoride determination within the required quantification range 0.10-100 mM. The possible interferences from the growth medium were minimized in-line. The determination rate was 78 h(-1) for the off-line monitoring of RBC and 50(-1) h for the on-line monitoring of the SBR, with a sample consumption of 0.500 mL and 0.133 mL per determination, respectively. Furthermore, the overall reagent consumption was quite low. The accuracy of the system was evaluated by comparison with a batch procedure. The SBR efficiency was monitored both on-line by the flow system and off-line by HPLC, for comparison purposes.

This paper describes the mathematical modeling of a set of rotating biological contactors (RBC) used for ammonia removal by nitrification of the wastewater stream that comes from a petroleum refinery. The entire biological wastewater treatment unit is composed by an aerated facultative lagoon, four sets of RBCs, and a pair of clarification decanters. In this paper we are concerned with the nitrification step. It has two sequential biochemical pathways (nitritation and nitratation) for the nitrification process and the existence of two distinct biological agents, related to Nitrobacter and Nitrosomonas autotrophic bacteria. We discuss some characteristics about the kinetics and important points that should be concerned for such a study. We also propose an alternative way to model the spatial distribution on the rotating bioreactor, with necessary simplifications and the discussion on their limitations. Available data sets from on-line analyzers for ammonium, composed by 3-hours sampled input and output, were used for parameter estimation and model validation. The process modeling, dynamic simulation, and parameter estimation were completely accomplished in the software EMSO. The results show the good prediction capability of the proposed model. It is expected to use the developed model for monitoring the wastewater treatment and for operator training. Meanwhile, the acquired knowledge obtained during the model development has improved the efficiency and the operational procedures of the industrial unit.

Publisher Summary This chapter focuses on rotating biological contactors (RBC), which consist of a series of closely spaced circular disks of polystyrene or polyvinyl chloride. RBC are submerged in wastewater and rotated slowly through it. Approximately 70% of RBC systems are used for removing carbonaceous biochemical oxygen demand (BOD) only, 25% are used for combined carbonaceous BOD removal and nitrification with the remainder being used for nitrification of secondary effluents. Biomass (slime or film) grows on to the surface of the disks, and eventually covers the entire wetted surface. The rotation of the disks alternately contacts the biomass with organic matter in the wastewater and then with the atmosphere for the adsorption of oxygen. The disk rotation affects oxygen transfer and maintains the biomass in an aerobic condition. The rotation also removes excess solids from the disks by the shearing forces it creates as the disk passes through the wastewater. The sloughed or sheared biomass is then maintained in suspension, so that it can be removed from the unit to the sedimentation tanks. Some RBC are designed to allow the sloughed solids to settle to the bottom of the RBC tank and breakdown, with periodical removal of the sludge. The principle elements of a RBC are shaft, media, drive system, tankage, enclosure, and settling tank. Usually, RBC are covered with a fiberglass or reinforced plastic shell over each separate drive shaft assembly that gives protection to the biofilm and plant in adverse weather conditions.

Phanerochaete chrysosporium immobilized on different support materials, such as polyurethane foam (PUF) and scouring web (SW), in shake cultures, was able to decolourize efficiently the sugar refinery effluent in a long-term repeated-batch operation. The decolourization medium composition was optimized using PUF-immobilized fungus. Addition of glucose was obligatory and the minimum glucose concentration was found to be 5 g/l. A rotating biological contactor (RBC) containing P. chrysosporium immobilized on PUF disks was operated with optimized decolourization medium, in continuous mode with a retention time of 3 days. By simply reversing the feed inlet of the reactor, after 17 days of operation, it was possible to double the active fungal lifetime. During the course of operation the colour, total phenols and chemical oxygen demand were reduced by 55, 63 and 48%, respectively., PEDIP Program - Project EUREKA EUROAGRI 1974 RESINAS.

Wastewater treatment systems for individual houses are required to be cheap, robust, compact, hygienic at site, odourless, require little maintenance and be installed, and operated, by unskilled labour. Their designs are based on full-scale units but are highly modified. Selection depends on: (a) the loading to the unit (Table 22.1), (b) the treatment objectives, (c) site conditions and location within the site, (d) effluent discharge requirements, (e) unit requirements (e.g. power, maintenance, etc.) and finally (f) cost. The most commonly employed systems are cesspools, septic tanks, rotating biological contactors (RBCs), constructed wetlands and small complete treatment systems (National Water Council, 1980) . Some basic guidelines to the process selection of treatment systems for small communities up to a population equivalent of 50 are given in Fig. 22.1.

Publisher Summary This chapter discusses the type of reactors that are available for treating wastewater either aerobically or anaerobically. In the conventional aerobic system, the substrate is used as a source of carbon and energy. It serves as an electron donor, resulting in bacterial growth. The extent of degradation is correlated with the rate of oxygen consumption, as well as the previous acclimation of the organism in the same substrate. Since the overall rate of the anaerobic process is controlled by the methanogenic step, the rate of biomethanation can be accelerated only by enhancement of the rate conversion of volatile fatty acids to methane. In addition, proper attention must be paid to the safety aspects during the reactor design, because large quantities of potentially explosive gas are generated.. Anaerobic rotating biological contactors are similar to their aerobic counterparts except that they are maintained at anaerobic conditions. Similarly, anaerobic ponds are comparable to their aerobic counterparts. In the fixed film reactors, the biomass is made to grow on a fixed support material such as PVC, wood, carbon, and rock. These reactors have a simple construction, good for higher loading rates, and can withstand higher toxic and organic loadings.

Rotating biological contactor is being widely used for wastewater treatment but there is an apparent lack of knowledge about the rate at which oxygen transfer occurs, in physical and biological system. In this study the transfer of oxygen from air to water by a rotating disc air–liquid contactor in physical system is investigated. The oxygen transfer model suggested by Kim and Molof, Water Sci. Technol. 14 (1982) 569, was modified and the developed model is termed as modified Kim and Molof model. The model was calibrated by using available data in literature and validated by experiments conducted in this study. The effect of significant physical parameters was integrated into a single term and is termed as volume renewal number. The modified Kim and Molof model was compared with the other available models. The coefficient of determination (R2) for the modified Kim and Molof model obtained is 0.95 which is much higher than in the other available models. Thereby the model is expected to estimate oxygen transfer more accurately. Further, a simplified linear model between KLa and the volume renewal number is proposed. Both modified Kim and Molof and linear model estimate the overall oxygen transfer coefficient (KLa) accurately., © Elsevier

This chapter discusses the various fixed film technologies for wastewater treatment. The fixed film processes are based on the capacity of different microorganisms to grow on surfaces. There are certain conditions that are important for the sloughing off to occur and are used technologically to control the biofilm. The hydraulic erosion acts continually on the surface of the biofilm and leads to a steady sloughing off on the outer side. The degradation of bacteria in the endogenous phase at the bottom of the biofilm may cause a weakening of the adhesion. Gas formation within the biofilm may destroy the adhesion. Development of a bacterial biofilm for substrate degradation is a complex phenomenon that is influenced by factors such as wastewater characteristics (nature of substrate, nutrients ratio and environmental conditions), operational factors (inoculation, organic loading rate, hydrodynamics), and the support medium. The various fixed film techniques include the trickling filter (TF), the rotating biological contactors (RBC), the biological aerated filters (BAF), and the suspended growth processes such as the activated sludge.

Several studies have employed rotating biological contactors (RBCs) for nitrogen removal. However, partially submersed disks are used for nitrification, while completely submersed disks are used for denitrification. The objective of this study was to investigate the effect of disk submergence in the performance of rotating biological contactors, in terms of the denitrification process. Two RBCs, one with completely submersed disks (100% submergence) and the other with partially submersed disks (64.5%), were operated under the same conditions. Their performance was evaluated in terms of denitrification efficiency as well as biofilm characteristics, composition and activity. As far as the denitrification process is concerned, the RBC with a completely submersed biofilm was more efficient than the other but had a longer delay in start-up. The biofilm of both reactors was very thick (0.6 mm), but with different structures. Biofilm activity seems to be directly dependent on the biofilm structure, namely on the degree of hydration., Instituto de Biotecnologia e Química Fina (IBQF)., Fundação para a Ciência e a Tecnologia (FCT) – PRAXIS XXI - grant BD/9121/96.

The lack of quality water supplies in the Western Australian gold fields is a limiting factor in the use of biological oxidation to treat refractory gold ores prior to the extraction of gold by cyanidation. Clearly, the recycling of water from the CIP process back to a biological oxidation plant is desirable in regions where supplies of fresh water are a limiting resource. However, thiocyanate, a by-product of the cyanidation process and a major contaminant in tailings dams, must be removed if water is to be recycled because it is toxic to sulphide-oxidising bacteria. Two strains of bacteria that were capable of degrading thiocyanate were isolated from the Youanmi mine in Western Australia. Both strains demonstrated the ability to utilise thiocyanate as their sole energy and nitrogen source. Physiological characterization of the strains indicated that both species of bacteria had the potential to tolerate the variable conditions encountered in Youanmi tails water. Phylogenetic analysis showed that one strain was a member of the genus Thiobacillus and the other was a member of the genus Halomonas but neither could be accommodated within any described species. Phosphate was the only additional nutrient required for thiocyanate degradation. Both strains were inoculated into a laboratory-scale Rotating Biological Contactor (RBC). The success and extent of colonization was confirmed by recovery of signature lipids of both strains from the fixed-film biomass. The bacteria degraded thiocyanate to ammonium, sulphate and carbon dioxide. The biomass supported on the reactor surface area (20 m 2 ) was capable of degrading approximately 2800 mg L −1 to less than 1 mg L −1 thiocyanate at a flow rate of 30 mL min −1 . The process operated in saline, low nutrient water.

This chapter provides a brief overview of biological treatment of industrial wastes with mutant bacteria. Biological treatment processes include: activated sludge, trickling filter, land disposal, aerobic digestion, stabilization, aerated lagoons, and variations of these such as rotating biological contactors (RBC), deep shaft, pure oxygen, activated carbon, and mutant seeding. Activated sludge remains the predominant biological treatment system for industrial wastes. With proper detention, activated sludge systems perform well for biological oxygen demand (BOD) removal. Biological treatment processes are probably the most cost effective techniques for treating aqueous waste streams containing organic contaminants. Mutant bacteria additives are now available to augment the naturally developed bacterial population of activated sludge, trickling filter, or lagoon treatment plants. A regular schedule of bacterial additions can provide faster system response to problems such as start-ups, plant upsets to variable and shock loads, and cold weather operations.

Laboratory studies on bacterial desulfurization of Pittsburgh seam coal are being conducted to determine the mechanisms of biodepyritization and the resulting implications for design of microbial processes for depyritization of coal. We are currently comparing leach rates of Pittsburgh No. 8 (150 − 600 μm) coal by ferric ions in columns under sterile, anoxic conditions with leach rates of coal inoculated with Thiobacillus ferrooxidans under aerobic conditions. In addition, shake flask studies are in progress to determine if finely ground coal (75 − 150 μm) is depyritized by T. ferrooxidans and Leptospirillum ferrooxidans at similar rates. If the ferric ion mechanism dominates in coal biodepyritization, depyritization of coal fines in slurry ponds could be based on ferric ion leaching. In this system, soluble Fe(III) ion would percolate through the slurry to oxidize pyritic sulfur and, in turn, be reduced to Fe(II). Regeneration of the Fe(III) oxidant from Fe(II) could occur in a rotating biological contactor or other bioreactor as a loop in a recycle stream. This design would eliminate the need for costly aeration of the bulk coal slurry.

Malaysia is fast becoming an urbanised developing country. In 1980, it was estimated that 34.2 per cent of the population lived in urban areas. The urban population increased to 37.4 per cent in 1985. It was estimated that by the year 1990, there will be about 40.7 per cent urban population. Problems arise in keeping pace with this growth in the provision of adequate facilities for removing the community's wastewater. Especially for those in the urban or peri-urban fringes and run-down slums of large towns and cities without any proper sanitation facilities, a compact package on-site domestic wastewater treatment plant could be the answer to their sanitation woes. A full scale sewage treatment plant with primary and secondary clarifiers incorporating a Rotating Biological Contactor (RBC) as secondary treatment was employed to evaluate the process performance. The treatment plant was operated at different organic loading rates that ranged from 15.0 to 37.0 g C0D/m2/d. The overall carbonaceous and ammonia-nitrogen removal efficiencies in the RBC were from 82–92 per cent and 82–97 per cent respectively. The organic removal percentages in the primary and secondary clarifiers were between 22.2–28.0 percent and 19.7–24.1 percent respectively. The package treatment plant incorporating the RBC has demonstrated to be an efficient wastewater process in the removal of biologically degradable carbonaceous compounds.

Experiments were conducted to describe the kinetic performance of a pilot scale biological fluidized bed reactor for municipal wastewater denitrification. A half-order kinetic model (8 to 48 mg·1 –1 NO 3 +NO 2 -N), coupled with an Arrhenius temperature dependence relationship (4 to 27°C) was adequate to describe denitrification. Neither biofilm support media (granular activated carbon and sand) or hydraulic flux (0.25 to 1.7 m 3 ·m –2 ·min –1 ) were found to be significant factors in controlling the denitrification rate. Under similar influent wastewater conditions, the fluidized bed process provided equivalent NO 3 +NO 2 -N removal at volumetric denitrification rates approximately ten times as fast as suspended growth or submerged rotating biological contactor (RBC) processes. Temperature dependency of the removal rate was less than for suspended growth or submerged RBC processes, but similar to the dependency in a packed column.

In a 3 stage RBC with diameter 3m. discs the experiment of the flowing waterfilm which was entrained by biomass on the discs in the air phase showed that in 30–40 seconds waterfilm was adequately treated. The average COD of the first stage waterfilm was about 35% less than that of the same stage tankwater. For nitrification of the third stage the waterfilm also had a higher NH4-N removal and N0x-N/N% than the same stage tankwater. The NH4-N removal of the waterfilm was about 35% also. The removal of, the contaminants in the waste waterfilm depended on the biosurface form and structure and rotational speed of the discs. The shaggy biosurface with many macroscopic filaments was more effective than with genergal flat biosurface. While the rotational speed of the discs slowed down the water quality of waterfilm became better, but the waterfilm rate reduced. This experiment also showed that the thin flowing waterfilm not only quickly absorbed oxygen from the air, but also voltilized the C02 of the waterfilm so quickly into the air, that the pH value a little rose at a very short retention time in the air phase.

The relationship between the power economy and the BOD removal rate per unit floor area or unit volume of the aerobic wastewater treatment processes, such as activated sludge, trickling filter, rotating biological contactor, fluidized bed, oxidation ditch, deep shaft aerator were investigated under their conventional operating conditions for the domestic wastewater treatment. It was clarified that the higher BOD removal rate per unit volume or unit floor area of the plants brings about the lower power economy. Appropriate selections of wastewater treatment processes and their operating conditions are significant to maintain the high quality effluent and the high power economy.

Synthetic phenolic wastewater with sole carbon source were treated by RBC, and biomass, SCOD and phenol concentration were measured under the steady-state after step-change of influent phenol concentration, together with the observation of the microorganisms. Also, each factors; rotational speed, influent phenol concentration, reactor volume to surface area ratio(G value), affecting to the treatment of RBC were studied. As the results, effluent phenol concentration below 2.0mg/L were maintained to the phenol loading rate, influent phenol concentration were 24.6g of phenol/m/day and 314.5mg/L respectively by the 4stage RBC(at 25 C, 20rpm). At the phenol concentrations less than lOOmg/L, there is no evidence of substrate inhibition. More than 200mg/L, however, there was substrate inhibition and organic removal kinetics could be expressed as a first-order reaction between influents and effluents. Increase of disc area improved removal efficiency but at G value more than 8.2, the rate of increase was reduced. Rotational speed increased removal efficiency. However, it was reduced in the case of rotational speed 50rpm(1.05m/s) because of attached biomass sloughing.

The alcoholic distillery waste is characterized as a concentrated organic waste with high suspended solids content. It has been reported that this type of waste can be processed anaerobically to produce methane using the biofilm reactor. Solid-liquid separation prior to the anaerobic digestion using the biofilm reactor is required, because the negative effect of the adsorbed organic particles on the anaerobic digestion may be caused by producing the additional load due to the dissolution of the adsorbed organic particles. In this paper, it was demonstrated that anaerobic digestion of the alcoholic distillery waste using the submerged Rotating Biological Contactor(RBC)was quite effective to reduce the TOC concentration of the waste when the original waste was diluted to about 2500 mg TOC/1 and 3000 mg SS/1. It was also shown that dissolved- air flotation, where the pressurized dilution water is mixed with the original waste, produced the wastewater suitable for the anaerobic digestion using the RBC. Based on the experimental results obtained in this study, the authors proposed the combined system of the dissolved- air flotation and anaerobic digestion using the RBC to treat the alcoholic distillery waste.

A full-scale combined anaerobic random packed-cage and aerobic disc rotating biological contactor was operated under fluctuating organic loading rates treating a brewery wastewater. The robustness of the combined system in coping with greatly fluctuating influent COD's and pH's is demonstrated. Excessive turbulence along with the variable operating conditions are identified as the major reasons for the failure to establish a significant anaerobic biofilm attached to the media. The importance of suspended sludge and its retention within the anaerobic RBC is demonstrated by the changing biological population in the suspended solids.

Publisher Summary Several procedures have been described for growing Phanerochaete chrysosporiurn for ligninase production. These procedures differ somewhat in the medium formulation and types of growth vessels: (1) shallow stationary cultures, (2) agitated liquid cultures, and (3) rotating biological contactors (RBCs; disk fermenters). The recently developed use of agitated culture for production of ligninase permits easier “scale up.” Although ligninase can be produced in agitated flask cultures, the reliable use of stirred tank fermenters awaits further development, which is ongoing in several laboratories. This chapter describes the production of ligninase in shallow stationary cultures and in agitated cultures. Shallow stationary cultures (10 ml) are grown in rubber-stoppered, 125-ml Edenmyer flasks at 39 ° under 100% oxygen. The stationary cultures give somewhat more reliable and reproducible results than the agitated cultures.