Biocatalyst removal

Entezari MH, Mostafai M, Sarafraz-yazdi A (2006) A combination of ultrasound and a biocatalyst removal of 2-chlorophenol from aqueous solution. Ultrason Sonochem 13 37—41... 

The saturation factor 9 plays a significant role and tends to enlarge the region where kinetics controls the process. In the above conditions, the minimization of the external concentration gradients is required. By a proper choice of system fluid-dynamics, provided that the enzyme-membrane bonds are strong enough to avoid biocatalyst removal by shear. 

Biocatalysts in nature tend to be optimized to perform best in aqueous environments, at neutral pH, temperatures below 40 °C, and at low osmotic pressure. These conditions are sometimes in conflict with the need of the chemist or process engineer to optimize a reaction with respect to space-time yield or high product concentration in order to facilitate downstream processing. Furthermore, enzymes and whole cells are often inhibited by products or substrates. This might be overcome by the use of continuously operated stirred tank reactors, fed-batch reactors, or reactors with in situ product removal [14, 15]. The addition of organic solvents to increase the solubility of substrates and/or products is a common practice [16]. 

Several scouting experiments were performed to find the best pH conditions. Figure 3 reports the ratio between the PG specific activity measured after the purification procedure (ASf) and the initial PG specific activity (ASi). At pH 3.5, the microspheres are able to remove from the broth the major part of the protein without PG activity, thus providing a four time increase of the enzyme specific activity. The purified PG from Kluyveromyces marxianus was immobilised following the above procedure. Batch reactions in the packed bed reactor were done to evaluate the biocatalyst stability. After an initial loss, due to enzyme release, the residual PG activity reaches a plateau value corresponding to about 40% of the initial activity. Probably, some broth component interfered during the immobilisation reaction weakening the protein-carrier interactions. 

Another important argument for the use of the organic solvent is the reverse hydrolytic reactions that become feasible [61,75]. The inhibition of the biocatalyst can be reduced, since the substrate is initially concentrated in the organic phase and inhibitory products can be removed from the aqueous phase. This transfer can shift the apparent reaction equilibrium [28,62] and facilitates the product recovery from the organic phase [20,29,33]. A wide range of organic solvents can be used in bioreactors, such as alkanes, alkenes, esters, alcohols, ethers, perfluorocarbons, etc. (Table 1). 

Biodesulfurization (BDS) is the excision (liberation or removal) of sulfur from organosul-fur compounds, including sulfur-bearing heterocycles, as a result of the selective cleavage of carbon-sulfur bonds in those compounds by the action of a biocatalyst. Biocatalysts capable of selective sulfur removal, without significant conversion of other components in the fuel are desirable. BDS can either be an oxidative or a reductive process, resulting in conversion of sulfur to sulfate in an oxidative process and conversion to hydrogen sulfide in a reductive process. However, the reductive processes have been rare and mostly remained elusive to development due to lack of reproducibility of the results. Moderate reaction conditions are employed, in both processes, such as ambient temperature (about 30°C) and pressure. 

Thus, several improvements have been made in the Rhodococcus strains to make desulfurization application possible or attractive however, the sulfur removal rate still remains the biggest bottleneck and no biocatalysts capable of rates needed for commercialization exist as of yet. 

In addition to desulfurization activity, several other parameters are important in selecting the right biocatalyst for a commercial BDS application. These include solvent tolerance, substrate specificity, complete conversion to a desulfurized product (as opposed to initial consumption/removal of a sulfur substrate), catalyst stability, biosurfactant production, cell growth rate (for biocatalyst production), impact of final desulfurized oil product on separation, biocatalyst separation from oil phase (for recycle), and finally, ability to regenerate the biocatalyst. Very few studies have addressed these issues and their impact on a process in detail [155,160], even though these seem to be very important from a commercialization point of view. While parameters such as activity in solvent or oil phase and substrate specificity have been studied for biocatalysts, these have not been used as screening criteria for identifying better biocatalysts. 

A process innovation was introduced by Valentine [61], who added an SOx sorbent for mitigating the inhibiting effects of the formed oxysulfides. This process was developed for sulfur removal from extra heavy oils, bitumens, and its emulsions, such as the trade mark Orimulsion. Any active biocatalyst may be used in this process carried out at temperatures close to 50°C. The main features disclosed in patents protecting the use of R. rhodochrous-bas d biocatalysts, in desulfurization reactions are summarized in Table 12. 

Due to the water requirement of biocatalytic systems, BDS is typically carried out as a two-phase aqueous-oil process. However, increased sulfur removal rates could be accomplished by using an aqueous-alkane solvent catalytic system [46,203,220,255], The BDS catalytic activity depends on both, the biocatalysts and the nature of the feedstock. It can vary from low activity for crude oil to as high as 60% removal for light gas-oil type feedstocks [27,203,256], or 70% for middle distillates, 90% for diesel, 70% for hydrotreated diesel, and 90% for cracked feedstocks [203,256], The viscosity of the crude oil poses mixing issues in the two-phase oil-water systems however, such issues are minimal for distillate feedstocks, such as diesel or gasoline [257]. 

In a biodesulfurization process, there are actually three phases. For a liquid mixture containing the three phases - liquid fossil fuel, water, and the biocatalyst, more than one filter would be required. One filter will preferentially collect either the liquid fossil fuel or aqueous phase as the filtrate. The retentate will then flow to the second filter, which will collect the component not removed before. The remaining retentate, containing the biocatalyst, can then, preferably, be recycled. The process can be used to resolve an emulsion or microemulsion of the liquid fossil fuel and aqueous phase resulting from a... 

Desulfurization of other diesel feedstocks from Total Raffinage was also reported by EBC. In these studies, different engineered biocatalysts were used. Two different middle distillate fractions, one containing 1850 ppm sulfur and other containing 650 ppm sulfur, were tested. R. erythropolis sp. RA-18 was used in one experiment and was reported to desulfurize the diesel from 1850 to < 1200ppm sulfur within 24 hours. On the other hand, it removed sulfur from a middle distillate with 650ppm sulfur to below 200 ppm sulfur [222], Various Pseudomonas strains were also tested in this study and reported to remove less amounts of sulfur. A favorable characteristic of the Pseudomonas strains is their inability to form stable emulsions, which can be useful trait for product recovery. 

Pseudomonas was also tested as a biocatalyst host in another study [224], In this biocatalyst, the plasmid carrying the dsz genes from R. erythropolis KA2-5-1 was cloned into Pseudomonas host resulting in a biocatalyst, Pseudomonas strain PAR41. This strain was able to remove about 17.5% sulfur from a LGO containing 360 mg/L sulfur. Most... 

Another Pseudomonas strain P. delafieldii R-8 was reported to remove 90.5% sulfur from highly desulfurized diesel oil [259], The biocatalyst achieved desulfurization via a pathway similar to the 4S pathway. The rate of desulfurization was reported to be 11.25 mmol sulfur/kg dcw/h, with the sulfur being reduced from 591 to 56 mg/L. This was achieved via two biocatalyst treatments lasting 20 hours each, although the biocatalyst was active only for first 6h in each treatment. Up to C4-DBTs were reported to be removed. Almost 100% of Q and C2 DBTs were removed and about 94% C3 DBTs and 97% C4 DBTs were removed. This strain of Pseudomonas thus appears to have a mechanism to uptake up to C4 DBTs through its cell membrane. 

A microbial contacting process for the oxidation of H2S was disclosed [22], in which a chemoautotrophic bacterium T. thiooxidants or T. ferroxidans) is used to remove sulfides from gaseous streams, at aerobic conditions and low pH. The low pH is preferred since the optimum pH for growth of the bacteria is below 4.0. and for the elimination of undesired contaminant bacterial strains. A contactor is employed, the flow of the sulfur-containing stream is contacted counter-currently with the biocatalytic aqueous solution. The sulfate is recovered from the aqueous solution, which contains the biocatalyst, as well. 

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