Enzymatic Synthesis Of Biodiesel Research Articles

Enzymatic In Situ Interesterification of Rapeseed Oil with Methyl Formate in Diesel Fuel Medium

The purpose of this research was to evaluate the process of enzymatic biodiesel synthesis by directly using rapeseed as a raw material, extracting the oil contained within and interesterifying with a mixture of methyl formate and mineral diesel, choosing the amount of mineral diesel so that the ratio between it and the rapeseed oil in the seeds was 9:1. As the final product of the interesterification process, a mixture of mineral diesel and biodiesel was obtained directly, which is conventionally produced by mixing the mineral diesel and biodiesel. The tests were performed using enzymatic catalysis using the lipase Lipozyme TL TIM. Process optimization was performed using the response surface methodology. A model describing the interaction of three independent variables and their influence on the yield of rapeseed oil methyl esters was developed. The physical and chemical indicators of the product obtained under optimal interesterification conditions were evaluated.

Ion Exchange Resin and Entrapped Candida rugosa Lipase for Biodiesel Synthesis in the Recirculating Packed-Bed Reactor: A Performance Comparison of Heterogeneous Catalysts

Ion exchange resins and immobilized lipase as heterogeneous catalysts are used to synthesize biodiesel for alternative fossil fuels. The use of ion exchange resins in the solid and activated phase can ease the separation process. Furthermore, resins can be reactivated and used repeatedly, reducing the need for catalysts. On the other hand, an immobilized enzyme is biodegradable and can catalyze the transesterification process to produce biodiesel with a lower alcohol-to-oil ratio, minimizing side reactions and impurities. Therefore, the catalysts used in this study are ion exchange resins, such as Lewatit MP-64, Amberlite IRA410Cl, and Diaion PK208LH, as well as immobilized Candida rugosa lipase. By using vegetable oil as a feedstock and methanol for the transesterification, biodiesel production was carried out in a packed bed reactor. The present study aims to investigate the optimum process parameters, including the concentration of resin and enzyme, resin activation time, resin types, flowrate, and stability of resin and enzyme on the biodiesel yield. The results showed that the optimum conditions for biodiesel production with ion exchange resin were 4 g of resin, activated for 3 h, and synthesized for 3 h; Lewatit obtained a biodiesel yield of 94.06%, Amberlite obtained 90.00%, and Diaion obtained 73.88%. Additionally, the stability test of the reactivated Lewatit resin showed that it still has the capability of producing biodiesel with a yield of more than 80% after three regeneration cycles. In contrast, Candida rugosa lipase as was immobilized by entrapment in sodium alginate before being used in the biodiesel production for 12 h. The results showed that lower flowrate in enzymatic biodiesel synthesis produced a higher amount of biodiesel, of up to 71.1%. Nonetheless, immobilized lipases can be used up to three times without a significant loss in biodiesel yield.

Enzymatic biodiesel synthesis from Trichosporon shinodae yeast through circular economy: A greener approach

In the present study, crude glycerol, a by-product of biodiesel production, was supplemented to Trichosporon shinodae for higher lipid synthesis and further extracted (lipid) using ultrasonic-assisted extraction (UAE). Optimization of enzymatic transesterification (ET) using Response Surface Methodology (RSM) coupled with Central Composite Design (CCD) quadratic model of single-cell oil resulted in 90.03%(v/v) of maximum FAME's yield with methanol to oil ratio of 14.98, solvent to oil ratio of 0.270 in 12.52 h at 25.99 °C with 1.26%(v/v) initial water content. The developed model was significant, with an R2 value of 95.20% and an adj.R2 value of 91.97%. The developed RSM model was further utilized for optimization through the Modified Chaotic Invasive Weed Optimization (MC-IWO). The optimization results showed a maximum FAME's yield of 90.03% utilizing the MC-IWO parameters of 5 seeds with a 6% initial standard deviation using an exponent value of 2 with a population size of 15 in 56 generations. FAME's characterization of T. shinodae has revealed 58.8% (v/v) of saturated FAME's and 41.2% (v/v) of FAME's predominantly with oleic acid methyl ester content (38%,v/v). Fuel properties of T. shinodae showed kinematic viscosity of 3.9–4.4 mm2/s, iodine value (113–115 g I2/100 g), cetane index (46–48), saponification value (203–206 mg/KOH), acid value (1.2–1.4 mg KOH/g) and higher heating value (39–40 MJ/kg). Blending experiments of T. shinodae biodiesel with conventional petroleum showed calorific values of B5, B20 and B100 were 48.63 MJ/kg, 43.45 MJ/kg and 38.35 MJ/kg, respectively. This study showed the potential of T. shinodae biodiesel, which can be used either as an additive or as a blend of conventional fuels.

Obtention of biodiesel through an enzymatic two-step process. Study of its performance and characteristic emissions.

We describe the enzymatic synthesis of biodiesel from waste cooking oil (WCO) in a two-step production process: hydrolysis of WCO, followed by acid-catalyzed esterification of free fatty acids (FFAs). Among the three commercial enzymes evaluated, the inexpensive lipase Lipex® 100L supported on Lewatit® VP OC 1600 produced the best overall biodiesel yield (96.3%). Finally, we assessed the combustion efficiency of the obtained biodiesel and its blends. All blends tested presented lower emissions of CO and HC compared to diesel. The NOx emissions were higher due to biodiesel's high volatility and viscosity. The cost of biodiesel production was calculated using the process described.

Mitigation of methanol inactivation of lipases by reaction medium engineering with glycine betaine for enzymatic biodiesel synthesis

Lipases are used for enzymatic production of fatty acid methyl esters (FAME/ biodiesel), an alternative fuel. However, lipases are intolerant to methanol, which is one of the substrates in the transesterification reaction to produce biodiesel. Methanol denatures lipases and thereby significantly reduces FAME yield. We have used molecular dynamics simulations to investigate methanol induced denaturation of Candida rugosa lipase and the effects of adding glycine betaine (GB), (an osmolyte), to the reaction medium, to keep lipase catalytically active in the presence of methanol. Simulation studies indicate a significant reduction in lipase denaturation, in the presence of 3 M and above concentrations of GB. The simulation results were experimentally validated by assessing FAME yield in the presence and absence of GB. Higher yields of FAME in the presence of GB confirm catalytically active lipase, indicating a stable enzyme and stabilizing action of GB on lipase in the presence of methanol. This study can therefore be regarded as a proof-of-concept to ascertain the stabilizing effect of glycine betaine, on lipase, in the presence of methanol. Furthermore, the fuel properties of the FAME synthesized were assessed by ASTM methods.

Model-to-model: Comparison of mathematical process models of lipase catalysed biodiesel production in a microreactor

Based on the experimental results and model simulation it was shown that Bi-Bi Ping Pong mathematical process model is not suitable for the description of the lipase catalysed biodiesel production in a microreactor. Therefore, three additional mathematical process models were proposed. Prior to transport model development, the reaction rates were described with double substrate Michaelis-Menten kinetics, Bi-Bi Ping-Pong kinetics, and Hill kinetics. The Hill kinetic model was proposed as the best kinetic model based on the model selection criterion. In order to validate the proposed mathematical process models, biodiesel synthesis in a microreactor was performed at four different initial process conditions. In all validation experiments, free fatty acid concentration and enzyme concentration were kept constant and the oil to methanol ratio in the inlet streams was altered. An increase of biodiesel yield was observed for the higher methanol concentration in the system. If a large excess of methanol was used (oil to methanol ratio 1:90) the yield was higher than 90% for the residence time of only 40 min. In comparison to the batch process, where the yield of 96% was achieved for 48 h, this was a significant improvement. Two out of three proposed mathematical process models described experimental data very well for all analysed residence times. Considering the level of complexity and accuracy, a mathematical process model of steady-state two parallel plug flow reactors was proposed as the optimum solution for the mathematical description of enzymatic biodiesel synthesis performed in a microreactor.

Lipid Extraction Maximization and Enzymatic Synthesis of Biodiesel from Microalgae

Microalgae has received overwhelming attention worldwide as a sustainable source for energy generation. However, the production of biofuel from microalgae biomass consists of several steps, of which lipid extraction is the most important one. Because of the nature of feedstock, extraction needs special attention. Three different methods were studied to extract algal oil from two different algae variant, Chlorella sp. and Spirulina sp. The highest percentage oil yield was obtained by ultrasonication (9.4% for Chlorella sp., 6.6% for Spirulina sp.) followed by the Soxhlet and solvent extraction processes. Ultrasonication and Soxhlet extraction processes were further optimized to maximize oil extraction as solvent extraction was not effective in extracting lipid. For ultrasonication, an amplitude of 90% recorded the highest percentage yield of oil for Spirulina sp. and a 70% amplitude recorded the highest percentage yield of oil for Chlorella sp. On the other hand, for Soxhlet extraction, a combination of chloroform, hexane, and methanol at a 1:1:1 ratio resulted in the highest yield of algal oil. Afterward, the crude algae oil from the ultrasonication process was transesterified for 5 h using an immobilized lipase (Novozyme 435) at 40 °C to convert triglycerides into fatty acid methyl ester and glycerol. Thus, ultrasonic-assisted lipid extraction was successful in producing biodiesel from both the species.

Specific Anion Effects on Lipase Adsorption and Enzymatic Synthesis of Biodiesel in Nonaqueous Media.

Pseudomonas fluorescens lipase (Pfl) was adsorbed on macroporous polypropylene to obtain a heterogeneous biocatalyst. The effect of NaCl concentration and of different 100 mm sodium salts on the Pfl loading and catalytic performance toward biodiesel synthesis via the solvent-free methanolysis of triglycerides was studied. Although lipase adsorption onto polypropylene is governed by hydrophobic interactions, both salt concentration and anion type affect lipase loading. Protein loading decreased along the series: Cl- > SO42- ≈ no salt > Br- > I- > SCN- > F- > AcO-. This nonmonotonic ion-specific trend can be the result of opposite mechanisms occurring during the adsorption step. A similar trend is observed also for triglyceride conversion and biodiesel yield. It is likely that ions affect the microenvironment of the adsorbed lipase by interacting specifically with the hydration water and polypeptide chains, thus affecting enzyme catalysis.

Enzymatic Synthesis of Biodiesel by Direct Transesterification of Rapeseed Cake

Abstract Increase in energy demand and limited sources of fossil fuels force the world’s population to seek new energy sources Biodiesel obtained by transesterification of oils with an alcohol in the presence of catalysts is an example of renewable fuel. The aim of the study was to assess the best conditions for enzymatic biodiesel synthesis by direct transesterification of rapeseed cake using Taguchi method. The influence of alcohol (methanol, ethanol, propanol, and butanol), temperature (30–60 °C), C. antarctica lipase B concentration (0.5–2 %), time (6–48 h) and rapeseed cake to alcohol ratio (1:2–1:5) was examined in the synthesis of fatty acid alkyl esters. Optimum conditions for direct enzymatic transesterification of rapeseed cake are: 30 °C, 12 h, 0.5 % of lipase and ethanol in 4:1 ratio to rapeseed cake.

A new reactor for enzymatic synthesis of biodiesel from waste cooking oil: A static-mixed reactor pilot study

Nowadays, biodiesel has become a globally accepted renewable liquid fuel. The environmentally friendly enzymatic synthesis process is being widely researched. To solve the problem that production efficiency is low in enzymatic process, we optimized the structure of the static mixer by computational fluid dynamics (CFD) modeling. The optimal static mixer consists of 6 mixer units, and the length-diameter ratio of every mixer unit is 1.5. The enzymatic synthesis of biodiesel was conducted between waste cooking oil (WCO) and methanol by transesterification and esterification. The reaction conditions were also optimized in this designed static-mixed reactor. Under optimal conditions, the reaction time can save half time when compared with the traditional stirred reactor, which indicates that the static mixer has higher production efficiency.

Improving the Yields and Reaction Rate in the Ethanolysis of Soybean Oil by Using Mixtures of Lipase CLEAs.

Due to the heterogeneity of oils, the use of mixtures of lipases with different activity for a large number of glycerol-linked carboxylic acids that compose the substrate has been proposed as a better alternative than the use of one specific lipase preparation in the enzymatic synthesis of biodiesel. In this work, mixtures of lipases from different sources were evaluated in their soluble form in the ethanolysis of soybean oil. A mixture of lipases (50% of each lipase, in activity basis) from porcine pancreas (PPL) and Thermomyces lanuginosus lipase (TLL) gave the highest fatty acid ethyl ester (FAEE) yield (around 20 wt.%), while the individual lipases gave FAEE yields 100 and 5 times lower, respectively. These lipases were immobilized individually by the cross-linked enzyme aggregates (CLEAs) technique, yielding biocatalysts with 89 and 119% of expressed activity, respectively. A mixture of these CLEAs (also 50% of each lipase, in activity basis) gave 90.4 wt.% FAEE yield, while using separately CLEAs of PPL and TLL, the FAEE yields were 84.7 and 75.6 wt.%, respectively, under the same reaction conditions. The mixture of CLEAs could be reused (five cycles of 6 h) in the ethanolysis of soybean oil in a vortex flow-type reactor yielding an FAEE yield higher than 80% of that of the first batch.

Lipases in liquid formulation for biodiesel production: Current status and challenges

Enzymatic synthesis of biodiesel showed advantageous characteristics in relation to other technologies once it works under bland conditions, no generation of wastewater, no occurrence of saponifications reactions and production of a biodiesel with high quality. Although many researches still apply immobilized lipases, the high costs associated with this biocatalyst hamper the economic viability of the process. Lipases in free/soluble/liquid formulation employed to biodiesel production via hydroesterification reaction have attracted interest from researchers because they are more cost effective than the immobilized form, making the enzymatic route more competitive. In addition, soluble lipases present higher reaction rates, reducing the time required to obtain a satisfactory biodiesel yield. Despite the fact that already exist industrial plants producing biodiesel with the assistance of lipases in liquid formulation, results of researches show that the process still needs to overcome some drawbacks. This paper is a comprehensive and critical discussion on the publications where soluble lipases were applied on biodiesel synthesis, as well as the challenges that the technology faces and its current status in pilot and industrial applications.

Improving the soluble lipase-catalyzed biodiesel production through a two-step hydroesterification reaction system.

The application of lipases in liquid formulation instead of immobilized forms in the enzymatic biodiesel synthesis can make the process cost-efficient, more competitive, and sustainable. However, despite the benefits, the long reaction times required to achieve satisfactory yields is still a drawback of this biotechnological process. In this sense, employing the novel low-cost soluble NS40116 lipase, this paper proposes an innovative two-step hydroesterification reaction (TSHR) system as a technique of improving the reaction rate of an enzymatic biodiesel production. With the employment of two central composite statistical design to optimize the parameters of each of the reactions involved, the influence of the parameters "water concentration added to the reaction," "methanol-to-oil molar ratio," and "lipase load" on the process yield, besides the acid value of the samples, was investigated. After only 8 h of reaction, the highest fatty acid methyl ester yield reached was 97.1% with an acid value of 4.62 mg KOH g-1 utilizing a total of 8 wt% water, methanol-to-oil molar ratio of 6.3:1, and 0.70 wt% of lipase. Furthermore, the statistical models for both reactions indicated to be significant with 95% of reliability. Considering that the papers published using soluble lipases in a one-step batch process normally reach similar yields to those obtained in this research after 16 h to 24 h of reaction, the proposed system demonstrated to be a promising option of process configuration for the enzymatic production of biodiesel.

Enzymatic Biodiesel Synthesis by the Biphasic Esterification of Oleic Acid and 1-Butanol in Microreactors

The enzymatic esterification of oleic acid and 1-butanol to butyl oleate was performed in an aqueous–organic system in capillary microreactors with various inner diameters operated under slug flow....

Lipase NS40116 as catalyst for enzymatic transesterification of abdominal chicken fat as substrate

Enzymatic biodiesel, as an alternative biodiesel obtained by chemical route and alternative to petroleum fuel, becomes attractive by using low-cost raw materials such as abdominal chicken fat and low-cost free enzyme as catalyst. In this sense, the main goal of this work was to optimize enzymatic biodiesel synthesis to obtain high methyl esters content from abdominal chicken fat by using free NS 40116 lipase as a catalyst in a solvent-free system. Kinetics studies were carried out to investigate the influence of enzyme in relation to water content (0.0–2.0 wt%), NS 40116 concentration (0.1–0.7%), temperature (30–45 °C), molar ratio oil to ethanol (1:3–1:6) and different additions of methanol (1–6 shots). Under optimized conditions of reaction, 77% of conversion was obtained using 0.3 wt% NS 40116, 2 wt% water, 1:4.5 molar ratio oil to methanol (3 shots), 30 °C and 250 rpm in 24 h.

Characterization and compositional analysis of highly acidic karanja oil and its potential feedstock for enzymatic synthesis of biodiesel

In the present study, we focused on the synthesis of biodiesel fromPseudomonas cepacia, crude karanja oil by the process of enzymatic transesterification using bio-support materials, such as lipase immobilized on polyvinyl alcohol/AlgNa.

Effect of acyl‐acceptor stepwise addition strategy using alperujo oil as a substrate in enzymatic biodiesel synthesis

AbstractBACKGROUNDUsing renewable feedstock sources for biodiesel production seems to be a promising strategy and even more so when enzymatic catalysis with lipases are used. However, it is well known that these enzymes could be inactivated by reaction conditions such as temperature or alcohol concentration. In this work, the effect of temperature and initial water activity (aw) value on immobilised recombinant Rhizopus oryzae lipase (rROL) were studied. Methanolysis and ethanolysis reactions using alperujo oil with three different stepwise addition strategies were employed.RESULTSRecombinant 1,3‐positional selective rROL covalently immobilised on polymethacrylate amino‐epoxy activated support showed maximum initial reaction rate at low aw value (0.093). It was found that 30 °C was the optimal temperature in terms of biocatalyst stability during transesterification reactions. Adding alcohol at once, ethanol was clearly a better acyl‐acceptor in terms of stability than methanol. Productivity was found to be 2‐fold higher when five pulses of ethanol were used instead of methanol.CONCLUSIONSAlperujo oil has great potential as a low cost feedstock for biodiesel production through enzymatic catalysis using a nearly semi‐continuous alcohol addition strategy. © 2017 Society of Chemical Industry

Optimization studies to develop a low-cost medium for production of the lipases of Rhizopus microsporus by solid-state fermentation and scale-up of the process to a pilot packed-bed bioreactor

Abstract A low-cost lipase preparation is required for enzymatic biodiesel synthesis. One possibility is to produce the lipase in solid-state fermentation (SSF) and then add the fermented solids (FS) directly to the reaction medium for biodiesel synthesis. In the current work, we scaled up the production of FS containing the lipases of Rhizopus microsporus. Initial experiments in flasks led to a low-cost medium containing wheat bran and sugarcane bagasse (50:50 w/w, dry basis), supplemented only with urea. We used this medium to scale-up production of FS, from 10 g in a laboratory column bioreactor to 15 kg in a pilot packed-bed bioreactor. This is the largest scale yet reported for lipase production in SSF. During scale-up, the hydrolytic activity of the FS decreased 57%: from 265 U g−1 at 18 h in the laboratory bioreactor to 113 U g−1 at 20 h in the pilot bioreactor. However, the esterification activity decreased by only 14%: from 12.1 U g−1 to 10.4 U g−1. When the FS produced in the laboratory and pilot bioreactors were dried and added directly to a solvent-free reaction medium to catalyze the esterification of oleic acid with ethanol, both gave the same ester content, 69% in 48 h.

Sugar-Improved Enzymatic Synthesis of Biodiesel with Yarrowia lipolytica Lipase 2

In this research, it was found that in Yarrowia lipolytica Lipase 2 (YLLIP2)-catalyzed biodiesel production, the content of fatty acid methyl esters (FAMEs) was increased by about 10% with the addition of d(+)-glucose. The result indicated that the d(+)-glucose could be used as an effective additive in YLLIP2-catalyzed biodiesel production. In accordance with the results above, the single factor experiments of key parameters in the process were first carried out. On the basis of the single factor experiment results, a five-factor, three-level response surface method was adopted to obtain the optimal reaction conditions: lipase dosage 40 IU/g oil, d(+)-glucose to lipase 1:1.05 (w/w), water content 1.95%, and reaction temperature 39.4 °C. A stoichiometric amount of methanol (11.5%, methanol/waste oil) was added in 6 times (every 4 h for each addition). A subsequent pilot scale production in a 5-ton reactor was carried out to check the performance of this method, and a good biodiesel content of 91.4% was obt...

Optimization of the Enzymatic Synthesis of Biodiesel from Terminalia cattapa L. Kernel Oil Using Response Surface Methodology.

The synthesis of biodiesel using Terminalia cattapa L. (local language: Ketapang) kernel oil and ethanol has been conducted. The effect of variable parameters of synthesis includes reaction time (3-7 h), temperature (35-60°C), amount of enzyme (0.1-0.3 g) and substrate molar ratio (ketapang kernel oil to ethanol, 1:1-1:3) were studied. Response surface methodology based on a five-level and four-variable central composite rotatable design were used to evaluate the interactive effects of the sinthesis parameters on the percentage yield of biodiedsel. The analysis of variance shows that the optimum conditions of the synthesis reaction were at 4.03 h of reaction time, 0.25 g of weight of enzyme, 45.04°C of temperature and 1.50 of substrate molar ratio. The actual experimental yield was 83.9% under optimum condition, which compared well to the maximum predicted value of 83.71%.