Biodiesel synthesis in a solvent-free system by recombinant Rhizopus oryzae: comparative study between a stirred tank and a packed-bed batch reactor

A simultaneous synthesis of biodiesel, as fatty acid methyl esters, and monoacylglycerols catalysed by the recombinant Rhizopus oryzae lipase immobilized by adsorption on Relizyme OD/403M is presented. The use of this 1(3)-positional specific lipase prevents the formation of glycerol as a by-product, thus avoiding its drawbacks. The synthesis was carried out in a solvent-free system and it has been studied in two different reactor systems: stirred tank and packed-bed reactor. Stirred tank reactor presented a high-initial reaction rate and achieved a 33.6% yield, which corresponds to a value of 50.4% of the maximum yield that can be achieved with a 1(3)-positional specific lipase. In packed-bed reactor there was a smaller initial reaction rate, but it was achieved a 49.1% yield, which corresponds to a 73.6% of the maximum yield. When a second batch is performed, the yield decreased only 4% when packed-bed reactor is employed whereas a drastic decrease is observed in a stirred tank operation. Therefore, packed-bed reactor showed a best performance and minor damage to the biocatalyst.     

Enzymatic transesterification of olive oil and its precursors

The effect of different solvents and three different acyl acceptors on the transesterification of triolein (as a model compound) was investigated. The yield of biodiesel (methyl or ethyl ester) was monitored as a function of time. The yield of the product was also determined in a solvent-free system for two different modes of stirring. The results indicate that the highest yield is obtained in a solvent-free system with mechanical stirring. Methyl acetate is also effective as a solvent and acyl acceptor. Biodiesel was also produced by transesterification of triglycerides (triolein) present in olive oil with methanol and Novozym® 435. The effect of the molar ratio of methanol to triolein, mode of methanol addition, enzyme activity and reaction temperature on overall conversion and yield was determined. The final conversion and yield of biodiesel after a reaction time of 24 h were unaffected by changes in these parameters over the range studied. Preliminary findings indicate that the results obtained from small scale reactors and fresh oil can be extended to larger reactors and used oil.     

Enzymatic transesterification of olive oil and its precursors

The effect of different solvents and three different acyl acceptors on the transesterification of triolein (as a model compound) was investigated. The yield of biodiesel (methyl or ethyl ester) was monitored as a function of time. The yield of the product was also determined in a solvent-free system for two different modes of stirring. The results indicate that the highest yield is obtained in a solvent-free system with mechanical stirring. Methyl acetate is also effective as a solvent and acyl acceptor. Biodiesel was also produced by transesterification of triglycerides (triolein) present in olive oil with methanol and Novozym® 435. The effect of the molar ratio of methanol to triolein, mode of methanol addition, enzyme activity and reaction temperature on overall conversion and yield was determined. The final conversion and yield of biodiesel after a reaction time of 24 h were unaffected by changes in these parameters over the range studied. Preliminary findings indicate that the results obtained from small scale reactors and fresh oil can be extended to larger reactors and used oil.     

Enzymatic conversion of sunflower oil to biodiesel in a solvent-free system: Process optimization and the immobilized system stability

The feasibility of using the commercial immobilized lipase from Candida antarctica (Novozyme 435) to synthesize biodiesel from sunflower oil in a solvent-free system has been proved. Using methanol as an acyl acceptor and the response surface methodology as an optimization technique, the optimal conditions for the transesterification has been found to be: 45 ^oC, 3% of enzyme based on oil weight, 3:1 methanol to oil molar ratio and with no added water in the system. Under these conditions, >99% of oil conversion to fatty acid methyl ester (FAME) has been achieved after 50 h of reaction, but the activity of the immobilized lipase decreased markedly over the course of repeated runs. In order to improve the enzyme stability, several alternative acyl acceptors have been tested for biodiesel production under solvent-free conditions. The use of methyl acetate seems to be of great interest, resulting in high FAME yield (95.65%) and increasing the half-life of the immobilized lipase by about 20.1 times as compared to methanol. The reaction has also been verified in the industrially feasible reaction system including both a batch stirred tank reactor and a packed bed reactor. Although satisfactory performance in the batch stirred tank reactor has been achieved, the kinetics in a packed bed reactor system seems to have a slightly better profile (93.6 ± 3.75% FAME yield after 8–10 h), corresponding to the volumetric productivity of 48.5 g/(dm³ h). The packed bed reactor has operated for up to 72 h with almost no loss in productivity, implying that the proposed process and the immobilized system could provide a promising solution for the biodiesel synthesis at the industrial scale.     

Biodiesel production from algal oil using cassava (<Emphasis Type="Italic">Manihot esculenta</Emphasis> Crantz) as feedstock

As a potential source of biomass supplies, cassava (Manihot esculenta Crantz) has been studied for bioethanol production, but not for the production of biodiesel. In this study, we used cassava hydrolysate as an alternative carbon source for the growth of microalgae (Chlorella protothecoides) which accumulated oil in vivo, with high oil content up to 53% by dry mass under a 5-L scale fermentation condition. The oils were extracted and converted into biodiesel by transesterification. The biodiesel obtained consisted of mainly unsaturated fatty acids methyl ester (over 82%), cetane acid methyl ester, linoleic acid methyl ester, and oleic acid methyl ester. This work suggests the feasibility of an alternative choice for producing biodiesel from cassava by microalgae fermentation. We report herewith the optimized condition for the fermentation and for the hydrolysis of cassava as the carbon source.     

Continuous-flow biodiesel production using slit-channel reactors

Slit-channel reactors are reactors whose active surface areas are orders of magnitude higher than those of micro-reactors but have low fabrication costs relative to micro-reactors. We successfully produced biodiesel with different degrees of conversion using homogeneous catalyst in the slit-channel reactor. The reactor performance shows that percent conversion of soybean oil to biodiesel increases with channel depth, as expected, due to more efficient mixing. Shallow slit-channels require short average residence times for complete product conversion. Present results show that the slit-channel reactor provides an improved performance over traditional batch reactors using homogeneous sodium alkoxide catalyst. It is aimed to couple the reactors with solid catalysts in converting soybean oil to biodiesel and implementation method is suggested. The cost advantages resulting from the ease of fabrication of slit-channel reactors over micro-reactors and how these factors relate to the oil conversion efficiency to biodiesel are briefly noted and discussed.     

Integrated biodiesel production: a comparison of different homogeneous catalysts systems

The most common catalysts for biodiesel production are homogeneous basic catalysts. In the present paper, a comparison is made of different basic catalysts (sodium methoxide, potassium methoxide, sodium hydroxide and potassium hydroxide) for methanolysis of sunflower oil. All the reactions were carried out under the same experimental conditions in a batch stirred reactor and the subsequent separation and purification stages in a decanter. The analytical methods included gas chromatography and the determination of fat and oil conventional parameters. The biodiesel purity was near 100 wt.% for all catalysts. However, near 100 wt.% biodiesel yields were only obtained with the methoxide catalysts. According to the material balance of the process, yield losses were due to triglyceride saponification and methyl ester dissolution in glycerol. Obtained biodiesel met the measured specifications, except for the iodine value, according to the German and EU draft standards. Although all the transesterification reactions were quite rapid and the biodiesel layers achieved nearly 100% methyl ester concentrations, the reactions using sodium hydroxide turned out the fastest.     

Optimisation of integrated biodiesel production. Part II: a study of the material balance

A study was made of the material balance for the fatty acid methyl ester (biodiesel) synthesis from sunflower oil using potassium hydroxide as the catalyst. A factorial design of experiments and a central composite design have been used to evaluate the influence of operating conditions on the process material balance. The responses chosen were the biodiesel yield and the yield losses due to triglyceride saponification and methyl ester dissolution in glycerol, while the variables studied were temperature, initial catalyst concentration and the methanol:vegetable oil molar ratio. The biodiesel yield increased and therefore the yield losses decreased by decreasing catalyst concentration and temperature. However, the methanol:sunflower oil molar ratio did not affect the material balance variables significantly. Second-order models were obtained to predict the biodiesel yield and both yield losses. Within the experimental range studied, these models largely matched the results from the experiments.     

Lipase-catalysed synthesis of ester oils from biodiesel by-products

Ester oils obtained from natural long-chain fatty acids and alcohols are versatile substitutes for many petroleum-based products. Their efficient synthesis with the solvent-free esterification of free fatty acids (FFA) from by-products of biodiesel fabrication and 2-ethyl-1-hexanol with immobilised lipase from Thermomyces lanuginosa was investigated. The immobilisation of the biocatalyst in static emulsion yielded a specific esterification activity that was higher by a factor of 4.9–9.4 than the activity of the native enzyme. Favourable properties of the silicone-based immobilisation matrix in terms of stability and immobilisation yield were observed. In biodiesel by-products, the immobilised lipase catalysed the esterification of FFA as well as the transesterification of residual fatty acid methyl esters (FAME) to the desired ester oils. A conversion of 90% FFA and 35% FAME gave a total yield of 60%. The inactivation coefficients during repeated use in a stirred-tank reactor with intermittent pressure reduction were exceptionally low.     

Immobilization of steapsin lipase on macroporous immobead-350 for biodiesel production in solvent free system

Commercially available steapsin lipase was immobilized on macroporous polymer beads (IB-350) and further investigated for biodiesel production under solvent free conditions. The fatty acid methyl ester (biodiesel) synthesis was carried out by the methanolysis of fresh and used cooking sunflower oil. The enzymatic reaction for biodiesel synthesis was optimized with various reaction parameters and the obtained reaction conditions were 1: 6 molar ratio (oil: methanol), 50 mg biocatalyst and 20% water content at 45°C for 48 h under solvent free conditions. It was observed that 94% of biodiesel was produced under the optimized reaction conditions. The four step addition of methanol at the interval of 12 h was found to be more effective. Moreover the biocatalyst was effectively reused for four consecutive recycles and was appreciably stable for 90 days. The results obtained highlight potential of immobilized steapsin lipase for biodiesel production.     

Production of Cold-Flow Quality Biodiesel from High-Acidity On-Edible Oils—Esterification and Transesterification of Macauba (<Emphasis Type="Italic">Acrocomia aculeata</Emphasis>) Oil Using Various Alcohols

Biodiesel is an alternative fuel that has been used for partial or total substitution of diesel to reduce its environmental impacts. Prior studies on this topic have focused on the quest for better synthesis process, new catalysts and low-cost non-food and raw materials to improve the economic and sustainable production as well as product quality. In this study, acidic oil from macauba, a palm tree native to South America that has no food uses, was converted into biodiesel. The esterification and transesterification reactions were performed with methanol, ethanol and isobutanol with the goal of improving the cold properties of the biodiesel. The isobutyl ester exhibited the lowest freezing point temperature but underperformed outside of international specifications for kinematic viscosity; it also exhibited a low ester content. The methyl and ethyl esters were within the specifications of the international standards for ester content, density, kinematic viscosity and sulphur content. The ethyl ester produced from macauba oil displayed better properties in cold conditions than methyl and isobutyl esters studied here, with a cold filter plugging point of 0 °C. Its onset crystallisation temperature was reduced from ?5.96 to ?13.41 °C when subjected to fractional crystallisation. The ethyl ester exhibited the best lubricity value among the other esters studied.     

Biodiesel production in a magnetically-stabilized, fluidized bed reactor with an immobilized lipase in magnetic chitosan microspheres

Biodiesel production by immobilized Rhizopus oryzae lipase in magnetic chitosan microspheres (MCMs) was carried out using soybean oil and methanol in a magnetically-stabilized, fluidized bed reactor (MSFBR). The maximum content of methyl ester in the reaction mixture reached 91.3 (w/v) at a fluid flow rate of 25 ml/min and a magnetic field intensity of 150 Oe. In addition, the MCMs-immobilized lipase in the reactor showed excellent reusability, retaining 82 % productivity even after six batches, which was much better than that in a conventional fluidized bed reactor. These results suggested that a MSFRB using MCMs-immobilized lipase is a promising method for biodiesel production.     

Process engineering and optimization of glycerol separation in a packed-bed reactor for enzymatic biodiesel production

A process model for efficient glycerol separation during methanolysis in an enzymatic packed-bed reactor (PBR) was developed. A theoretical glycerol removal efficiency from the reaction mixture containing over 30% methyl esters was achieved at a high flow rate of 540 ml/h. To facilitate a stable operation of the PBR system, a batch reaction prior to continuous methanolysis was conducted using oils with different acid values and immobilized lipases pretreated with methyl esters. The reaction system successfully attained the methyl ester content of over 30% along with reduced viscosity and water content. Furthermore, to obtain a high methyl ester content above 96% continuously, long-term lipase stability was confirmed by operating a bench-scale PBR system for 550 h, in which the intermediates containing methyl esters and residual glycerides were fed into the enzyme-packed columns connected in series. Therefore, the developed process model is considered useful for industrial biodiesel production.     

Effect of water on lipase NS81006-catalyzed alcoholysis for biodiesel production

Biodiesel production catalyzed by free lipase has been drawing attention for its lower cost and faster reaction rate compared to immobilized lipase. It has been found that free lipase NS81006 could efficiently catalyze alkyl esters production and a certain amount of water is demonstrated to be necessary for the catalytic process. The effect of water content on liquid lipase NS81006-mediated methanolysis and ethanolysis for biodiesel production was first explored respectively in this paper. It was found that with water content ranging from 3% to 10% (based on oil weight), there was no significant difference in the final alkyl ester yield either in NS81006-mediated methanolysis or ethanolysis process, while the quality of biodiesel varied obviously. The acid value as well as the contents of monoglyceride and diglyceride were much lower in the lower water-containing system. With the water content decreasing from 10% to 3%, the acid value reduced from 8.24 to 4.89 mg KOH/g oil, and the content of MAG and DAG dropped to 0.31 and 0.22, from 0.62 and 0.74, respectively. Lipase could maintain rather good stability with proper alcohol adding strategy and the gradual reduction in biodiesel yield in the repeated uses resulted from the accumulation of by-product glycerol. The continuous running of lipase-mediated methanolysis of waste cooking oil was successfully realized at 30L reactor and a final methyl ester yield of over 90% could be obtained.     

Lipase-catalysed synthesis of ester oils from biodiesel by-products

Ester oils obtained from natural long-chain fatty acids and alcohols are versatile substitutes for many petroleum-based products. Their efficient synthesis with the solvent-free esterification of free fatty acids (FFA) from by-products of biodiesel fabrication and 2-ethyl-1-hexanol with immobilised lipase from Thermomyces lanuginosa was investigated. The immobilisation of the biocatalyst in static emulsion yielded a specific esterification activity that was higher by a factor of 4.9-9.4 than the activity of the native enzyme. Favourable properties of the silicone-based immobilisation matrix in terms of stability and immobilisation yield were observed. In biodiesel by-products, the immobilised lipase catalysed the esterification of FFA as well as the transesterification of residual fatty acid methyl esters (FAME) to the desired ester oils. A conversion of 90% FFA and 35% FAME gave a total yield of 60%. The inactivation coefficients during repeated use in a stirred-tank reactor with intermittent pressure reduction were exceptionally low.     

A monolithic lipase reactor for biodiesel production by transesterification of triacylglycerides into fatty acid methyl esters

An enzymatic reactor with lipase immobilized on a monolithic polymer support has been prepared and used to catalyze the transesterification of triacylglycerides into the fatty acid methyl esters commonly used for biodiesel. A design of experiments procedure was used to optimize the monolithic reactor with variables including control of the surface polarity of the monolith via variations in the length of the hydrocarbon chain in alkyl methacrylate monomer, time of grafting of 1-vinyl-4,4-dimethylazlactone used to activate the monolith, and time used for the immobilization of porcine lipase. Optimal conditions involved the use of a poly(stearyl methacrylate-co-ethylene dimethacrylate) monolith, grafted first with vinylazlactone, then treated with lipase for 2 h to carry out the immobilization of the enzyme. Best conditions for the transesterification of glyceryl tributyrate included a temperature of 37°C and a 10 min residence time of the substrate in the bioreactor. The reactor did not lose its activity even after pumping through it a solution of substrate equaling 1,000 reactor volumes. This enzymatic reactor was also used for the transesterification of triacylglycerides from soybean oil to fatty acid methyl esters thus demonstrating the ability of the reactor to produce biodiesel.     

Influence of transitional states on the microbial ecology of anaerobic digesters treating solid wastes

A better understanding of the microbial ecology of anaerobic processes during transitional states is important to achieve a long-term efficient reactor operation. Five wastes (pig manure, biodiesel residues, ethanol stillage, molasses residues, and fish canning waste) were treated in five anaerobic reactors under the same operational conditions. The influence of the type of substrate and the effect of modifying feeding composition on the microbial community structure was evaluated. The highest biomethanation efficiency was observed in reactors fed with fish canning waste, which also presented the highest active archaeal population and the most diverse microbial communities. Only two Bacteria populations could be directly related to a particular substrate: Ilyobacter with biodiesel residues and Trichococcus with molasses residues. Results showed that the time to achieve steady-state performance after these transitional states was not dependent on the substrate treated. But reactors needed more time to handle the stress conditions derived from the start-up compared to the adaptation to a new feeding. Cluster analyses showed that the type of substrate had a clear influence on the microbiology of the reactors, and that segregation was related to the reactors performance. Finally, we conclude that the previous inoculum history treating solid waste and higher values of active Archaea population are important factors to face a successful change in substrate not entailing stability failure.     

Developing the reaction kinetics for a biodiesel reactor

The aim of this paper was to investigate the kinetics of the biodiesel reaction in order to find out how best to reach 96.5% methyl ester. The purity of the biodiesel product was examined using gas chromatography to the EN14214 FAME standard and real-time optical microscopy was used to observe the reaction. The problem was the reaction does not reach completion and the mechanism is not understood. It was observed that droplet size had a major influence on reaction end point and that the reaction was mass-transfer limited. This observation was confirmed by developing a mass-transfer based reaction model using the data from the batch reactor which agreed with results from other researchers. The model predicted better conversion with more mixing intensity. The results show that significant improvements could be made to the conventional FAME process.     

Dimethyl carbonate as potential reactant in non-catalytic biodiesel production by supercritical method

In this study, the non-catalytic supercritical method has been studied in utilizing dimethyl carbonate. It was demonstrated that, the supercritical dimethyl carbonate process without any catalysts applied, converted triglycerides to fatty acid methyl esters with glycerol carbonate and citramalic acid as by-products, while free fatty acids were converted to fatty acid methyl esters with glyoxal. After 12 min of reaction at 350 degrees C/20 MPa, rapeseed oil treated with supercritical dimethyl carbonate reached 94% (w/w) yield of fatty acid methyl ester. The by-products from this process which are glycerol carbonate and citramalic acid are much higher in value than glycerol produced by the conventional process. In addition, the yield of the fatty acid methyl esters as biodiesel was almost at par with supercritical methanol method. Therefore, supercritical dimethyl carbonate process can be a good candidate as an alternative biodiesel production process.     

Glycerol extracting dealcoholization for the biodiesel separation process

By means of utilizing sunflower oil and Jatropha oil as raw oil respectively, the biodiesel transesterification production and the multi-stage extracting separation were carried out experimentally. Results indicate that dealcoholized crude glycerol can be utilized as the extracting agent to achieve effective separation of methanol from the methyl ester phase, and the glycerol content in the dealcoholized methyl esters is as low as 0.02 wt.%. For the biodiesel separation process utilizing glycerol extracting dealcoholization, its technical and equipment information were acquired through the rigorous process simulation in contrast to the traditional biodiesel distillation separation process, and results show that its energy consumption decrease about 35% in contrast to that of the distillation separation process. The glycerol extracting dealcoholization has sufficient feasibility and superiority for the biodiesel separation process.