Whole-cell Lipase Research Articles

Evaluation of biodiesel produced from some oleaginous fungi oil using whole-cell lipase

Evaluation of biodiesel produced from some oleaginous fungi oil using whole-cell lipase

Whole-cell Lipase Catalytic Synthesis of Short-chain Fragrance Esters using Aspergillus flavus

Background: Fragrances are the collection of unlike functional assemblies, most likely alcohols, esters, aldehydes, ketones, and acids in organic products/hydrocarbons. Short-chain aliphatic fragrance esters have immense applications as flavors in the food, pharmaceutical and cosmetic industries and also have remarkable commercial significance in cosmetics and personal care products like perfumes, face creams, shampoos, soaps, lotions, jams, jellies, etc. Objective: This study aimed to synthesize short-chain fragrance esters using a whole-cell lipase catalyst from Aspergillus flavus (RBD-01). Method: The present study emphasizes the synthesis of artificial flavoring compounds by using a wholecell biocatalytic process, which can have wide significance. Herein, the preparation of ethyl alkanoates (ethyl propanoate to ethyl decanoate) was performed to investigate the flavors and fragrance excellence. The biomass from Aspergillus flavus (RBD-01) was used as a catalyst to facilitate the remarkable esterification activities towards the synthesis of important aroma esters with the help of a series of short-chain acids and alcohols. Result: The ethyl hexanoate (4) among all synthesized alkanoates was found to have a fruity fragrance with a good conversion rate. Further synthesized alkyl hexanoates (4A-4I) were found to have good fruity/pineapple/berry flavors and significant aroma quality. Conclusion: These results implied that whole-cell lipase of Aspergillus flavus (RBD-01) is a promising biocatalyst in the production of flavor aroma esters and can boost production in the food/cosmetic manufacturing industries.

Production of Methanol Tolerant Cell-bound Lipase of Magnusiomyces spicifer SPB2 and Application as Whole-cell Biocatalyst in Transesterification Reaction

Cell-bound lipase (CBL) is a potential biocatalyst for production of fatty acid alkyl ester (FAAE), which is feasible replacement for diesel fuel. This study aimed to improve both hydrolysis and transesterification activities of CBL from the yeast Magnusiomyces spicifer SPB2, which could be employed as whole-cell biocatalyst for transesterification reactions through optimizing the growth medium using Taguchi orthogonal array experimental method (L9). The optimized medium, resulting the highest CBL transesterification activity, consisted of soybean oil, peptone, Gum Arabic and an initial pH of 5. Interestingly, the fatty acid methyl ester (FAME) yield increased up to 33.28 times when compared with non-optimized medium. Moreover, cell morphology of M. spicifer SPB2 varied depending on the culture medium component. The single cell was more active to be used as the whole-cell biocatalyst than the pseudomycelial form for transesterification reaction. The highest FAME yield was 93.86 % when refined palm oil to methanol molar ratio of 1:8 was used as substrates. Whole-cell lipase from M. spicifer showed favorable activity to iso-butanol as an acyl accepter, resulting in fatty acid butyl ester (FABE) yield of 86.80 % at 1:3 molar ratio of palm oil to iso-butanol. The FAME and FABE yields dramatically decreased from 93.86 and 86.80 % to 57.02 and 4.48 %, respectively after the reuse of the enzyme to the second batch of reaction. These results indicated that the whole-cell biocatalyst developed from M. spicifer was an efficient and economical approach to improve enzyme activity and stability in the transesterification reaction. HIGHLIGHTS Methanol tolerant cell-bound lipase and extracellular lipase of spicifer SPB2 were produced The whole-cell biocatalyst of spicifer SPB2 was developed for FAME production The optimized medium was developed to prepare whole-cell lipases from spicifer SPB2 Gum Arabic was the most influencing factor for synthetic activity of whole-cell CBL from yeast Single cell morphology exhibited higher transesterification activity than pseudomycelial form GRAPHICAL ABSTRACT

Palm Oil Decanter Cake Wastes as Alternative Nutrient Sources and Biomass Support Particles for Production of Fungal Whole-Cell Lipase and Application as Low-Cost Biocatalyst for Biodiesel Production

This is the first report on the possible use of decanter cake waste (DCW) from palm oil industry as alternative nutrient sources and biomass support particles for whole-cell lipase production under solid-state fermentation (SSF) by newly isolated fungal Aspergillus sp. MS15 and their application as a low-cost and environment-friendly biocatalyst for biodiesel production. The results found that DCW supplemented with 0.1% K2HPO4, 0.05% MgSO4·7H2O, 1% peptone and 2% urea and pH adjusted to 6.0 was optimal for whole-cell lipase production. The optimal moisture content and fermentation temperature was 60% and 37.5 °C, respectively. Environmentally friendly biodiesel production, through either esterification or transesterification using whole-cell lipase immobilized on DCW as a biocatalyst, was optimized. The optimal reaction temperature for both reactions was 37 °C. The whole-cell lipase effectively esterified oleic acid into >95% biodiesel yield through esterification under optimal water activity at 0.71 and an optimal methanol to oleic acid molar ratio of 2:1, and also effectively transesterified palm oil under optimal water activity at 0.81 and an optimal methanol to oil molar ratio of 3:1. The fuel properties of produced biodiesel are close to the international biodiesel standards. These results have shown the circular utilization of palm oil mill waste for the low-cost production of an effective biocatalyst, and may contribute greatly to the sustainability of renewable bioenergy production.

Response surface methodology for optimization of Rhizopus stolonifer 1aNRC11 mutant F whole-cell lipase production as a biocatalyst for methanolysis of waste frying oil

Biodiesel, fatty acid methyl ester (FAME), is an environmentally friendly alternative fuel produced by transesterification of renewable lipid feedstocks for diesel engines. Methanolysis of waste frying oil (WFO) was catalyzed by Rhizopus stolonifer 1aNRC11 mutant F whole-cell lipase that produced under solid state fermentation (SSF) using wheat bran (WB) and black cumin meal as the main substrate. Cultivation SSF parameters of R. stolonifer mutant were optimized using Plackett-Burman (PB) experimental design and Face-centred Central Composite Design (FCCD), taking FAME yield as a response for whole-cell lipase production ability. PB results showed that cultivation temperature, medium pH and moisture content were the most significant parameters, which were selected for optimization by FCCD. The optimal determined conditions were as follows: WB (10 g), black cumin meal (6%, w/w of WB), fodder yeast (1%, w/w of WB), moisture content (1.5 g water/g solid), inoculum size (3 discs), initial pH (8), and cultivation time (4 day) at 28 °C. With R. stolonifer mutant cultivated in the optimized conditions, the FAME yield reached 82.6% that was close to the predicted yield of 85.7%. The analysis of variance showed that the model was statistically significant (p value of .0001) and could explain 89% of the variability in the response. These results indicated that the optimization conditions increased biodiesel production by about 43% compared to the basal medium (57.60%). The optimization of SSF conditions using low cost components makes the whole-cell lipase production process from the R. stolonifer 1aNRC11 mutant F economically effective for application in biodiesel production from WFO.

Biotransesterification capabilities of Mucorales whole-cell lipase isolates and mutants

Abstract In this study, enzymatic production of biodiesel (fatty acid methyl esters, FAME) from waste frying oil (WFO) was investigated using fungal whole-cell lipase as a sustainable technique. Seven Mucorales whole-cell isolates and 6 mutants were tested for WFO transesterification to produce biodiesel. All tested isolates showed positive respond to qualitative lipase and esterase activity tests. A relatively strong positive correlation was detected between quantitative lipase activity that ranged between 15.91 and 25.42 U/g and transesterification capabilities in terms of FAME that reached up to 46.35%. Two isolates that showed potent transesterification capability were identified molecularly using rRNA ITS sequences as Rhizopus americanus 2aNRC11 and Rhizopus stolonifer 1aNRC11. Mutagenic effect of ethyl methanesulfonate (EMS) improved transesterification capabilities of R. americanus 2aNRC11 mutant B and R. stolonifer 1aNRC11 mutant G by 7.89 and 11.54%, respectively, compared to their corresponding wild types. Random amplified polymorphic DNA (RAPD) analysis supported the probability of genetic mutations as the total polymorphism reached to 70.59 and 91.3% among R. stolonifer 1aNRC11 and R. americanus 2aNRC11 mutants and their corresponding wild types, respectively.

Novel Bacterial Surface Display System Based on the Escherichia coli Protein MipA.

Bacterial surface display systems have been developed for various applications in biotechnology and industry. Particularly, the discovery and design of anchoring motifs is highly important for the successful display of a target protein or peptide on the surface of bacteria. In this study, an efficient display system on Escherichia coli was developed using novel anchoring motifs designed from the E. coli mipA gene. Using the C-terminal fusion system of an industrial enzyme, Pseudomonas fluorescens lipase, six possible fusion sites, V140, V176, K179, V226, V232, and K234, which were truncated from the C-terminal end of the mipA gene (MV140, MV176, MV179, MV226, MV232, and MV234) were examined. The whole-cell lipase activities showed that MV140 was the best among the six anchoring motifs. Furthermore, the lipase activity obtained using MV140 as the anchoring motif was approximately 20-fold higher than that of the previous anchoring motifs FadL and OprF but slightly higher than that of YiaTR232. Western blotting and confocal microscopy further confirmed the localization of the fusion lipase displayed on the E. coli surface using the truncated MV140. Additionally the MV140 motif could be used for successfully displaying another industrial enzyme, α-amylase from Bacillus subtilis. These results showed that the fusion proteins using the MV140 motif had notably high enzyme activities and did not exert any adverse effects on either cell growth or outer membrane integrity. Thus, this study shows that MipA can be used as a novel anchoring motif for more efficient bacterial surface display in the biotechnological and industrial fields.

Assessment of waste frying oil transesterification capacities of local isolated Aspergilli species and mutants

Abstract Biodiesel (fatty acids methyl esters, FAME) has attracted considerable attention as an environmentally and eco-friendly alternative for diesel engines. This research manipulates the use of Aspergillus whole-cell lipase as a biocatalyst for biodiesel production from waste frying oil (WFO). A total of 17 isolates of Aspergillus species screened for lipase and esterase production abilities. Qualitatively, 11 Aspergillus isolates showed lipase and/or esterase activities and only 4 isolates were able to perform WFO transesterification under the tested conditions. Two Aspergillus isolates showed relatively high FAME yields, thus were selected as good enzyme producers. These two isolates were molecularly identified using rRNA gene sequence ITS1 and ITS2 as A. tamarii NDA03a (Genbank Accession Number MK849615) and A. flavus NDA04a (Genbank Accession Number MK811208), respectively. These identified isolates were exposed to ethyl methanesulfonate (EMS) for producing hyper lipolysis mutants. Mutagenesis led to 13.15 and 14.45% improvement of the WFO transesterification by A. tamarii NDA03a and A. flavus NDA04a, respectively. Random amplified polymorphic DNA (RAPD) analysis of the produced mutants confirmed the genetic basis of the activity variation. Genetic polymorphism reached to 79.31% and 80.65% between A. flavus NDA04a and A. tamarii NDA03a mutants and their corresponding wild types, respectively.

Valorization of Palm Biomass Waste for Sustainable Production of Lipase-Carbon Matrices And Biodiesel

The conversion of arable land to produce plant-based biofuels can produce more greenhouse gas (GHG) emissions than the displacing effect that is expected from the use of biofuels instead of fossil fuels. Utilizing agricultural waste offers an alternative pathway to reducing GHG emissions. Typical palm oil mills, for instance, produce palm kernel shells (PKS) and palm oil mill effluents (POME) as wastes in enormous amounts which accounts for > 60% of solid and liquid waste from the mill. Here, we explore the feasibility of a circular palm bioeconomy where waste PKS was directly converted to activated carbons (AC) in a technique (550 °C, under N 2 ) that departs from the conventional two-step (carbonization and activation) AC synthesis route. Using FT-IR, FE-SEM, and EDX as surface analysis tools, the synthesized carbons, PKAC, showed excellent morphological and surface functionalities similar to bituminous coal-derived activated carbons. The 1 – 10 μm pore diameters of PKAC were relevant in the immobilization of Aspergillus oryzae whole-cells expressing recombinant Fusarium heterosporum lipase. For biodiesel synthesis, the higher specific activity of the PKAC-immobilized whole-cell lipase (81.6 IU/mg BSP) ensured the conversion of POME to ≥97.5 wt. % fatty acid methyl ester (FAME). Whilst offering the benefits of cheap immobilization carriers and greener synthesis routes, the produced FAME showed properties (48.8 cetane number) that conform to ASTM and EN specifications. This study thus serves as a framework for the design of hydrocarbon synthesis platforms for a future palm bioeconomy.

Biodiesel-mediated biodiesel production: A recombinant Fusarium heterosporum lipase-catalyzed transesterification of crude plant oils

Production of biodiesel from unrefined plant-derived oils has been considerably investigated but, to date, there appears to be a limit by phospholine gums and other contaminants in its conversion to biodiesel. The presence of such contaminants increases the melting point of the oils, thereby, making it unfavorable for lipase-catalyzed reactions at mild temperatures. Here, to circumvent this limiting effect, we explore the potential of biodiesel as a replacement for conventional organic solvents in solubilizing gum-formation contaminants in crude palm oil (CPO). A strategy of 1:1 CPO/biodiesel molar amount consolidates degumming and trans/esterification into one-step using immobilized Aspergillus oryzae expressing Fusarium heterosporum lipase. The innocuous biodiesel solvent not only ensured a 98.8 wt% biodiesel yield but also improved the catalytic activity of the whole-cell lipase. This allowed repeated use of the recombinant lipase in nine consecutive batches. At room temperature, biodiesel as a solvent saves on post-separation and reduces environmental footprints of the biodiesel production process.

ArgR directly inhibits lipA transcription in Pseudomonas protegens Pf-5

ArgR, a transcriptional regulator belonging to the AraC/XylS family, plays a key role in arginine metabolism regulation. ArgR has also been found to repress the transcription of a lipase gene, but its molecular mechanism is still unknown. In this study, we investigated the molecular mechanism acting on the expression of intracellular lipase gene lipA regulated by ArgR in Pseudomonas protegens Pf-5 through knockout and overexpression of argR, detection of DNA-protein interaction in vivo, determining whole-cell lipase activities of various strains derived from Pf-5, and examining β-galactosidase activities of various lacZ fusions. The results demonstrated that ArgR inhibits lipA expression at the transcriptional level. Further results showed that the inhibition of lipA transcription by ArgR is mediated by binding to the ArgR binding site of lipA promoter to produce steric hindrance, in which the common sequence, TGTCGC is crucial for the ArgR binding. Besides, arginine inhibits lipA expression in both wild-type and argR mutant, and shows a synergistic inhibition on lipA expression when combined with ArgR. To the best of our knowledge, this is the first report on ArgR directly repressing the transcription of a lipase gene.

Chemical and Biological Investigation of Organic Wastes of Frying Oils and Beef Fats: Valorization for Biodiesel Production

The present study investigates the different approaches of biodiesel production by exploiting low cost feedstocks such as organic wastes of frying oils (WFO) and wastes of beef fats (WBF). The aim was to compare not only two different sources of waste raw materials but also different approaches of biodiesel production. Biodiesel which refers to fatty acid methyl esters (FAME) was produced by both chemical and enzymatic transesterification. The characterization of the biodiesel produced by both approaches was performed according to the European standard EN 14214. The results showed that the biological method gave a richer FAME biodiesel through the catalysis of whole-cell lipase. However, for the chemical method, better biodiesel physicochemical properties were observed for the two raw materials. Therefore, it would be interesting to compromise by optimizing the biological biodiesel production approach in order to obtain a better quality in coherence with EN 14214 requirements.

Effects of organic solvent, water activity, and salt hydrate pair on the sn-1,3 selectivity and activity of whole-cell lipase from Aspergillus niger GZUF36.

We previously screened a whole-cell lipase EC 3.1.1.3 from the novel strain Aspergillus niger GZUF36, which exhibited 1,3-selectivity in the synthesis of 1,3-diacylglycerol via glycerolysis. However, the mechanism of lipase selectively in catalyzing the sn-1,3 position remains ambiguous. This work was performed to investigate the 1,3-selective mechanism of lipase using glycerolysis to synthesize 1,3-diacylglycerol (1,3-DG) as a model reaction by changing solvent(s) and water activity (aw), and addition of salt hydrate pair. The measured diacylglycerol yield was also used to examine lipase activity. Results indicated that not only organic solvent and aw have strong effect on the sn-1,3 selectivity, but also ions of salt hydrate pair also affected selectivity. Lipase conformation was altered by hydrophobic interactions of the solvent, aw, or ions of salt hydrate, resulting in distinct sn-1,3 selectivity of the lipase. The salt hydrate pair changed the lipase conformation and selectivity not only by aw but also by static interactions, which was rarely reported. These parameters also affected lipase activity. The lipase displayed the highest selectivity (about 88%) and activity in solvents of t-butanol and n-hexane (1:29, v/v) at aw 0.43. The results demonstrated that the sn-1,3 selectivity and activity of the lipase from A. niger GZUF36 may be improved by control of some crucial factors. This work laid a foundation for the application of lipase in the synthesis of 1,3-DG and other structural and functional lipids.

Production of Whole-Cell Lipase from Streptomyces clavuligerus in a Bench-Scale Bioreactor and Its First Evaluation as Biocatalyst for Synthesis in Organic Medium.

This work evaluated a wild-type Streptomyces clavuligerus strain as a whole-cell lipase (Sc-WCL) producer by submerged fermentation. In an orbital shaker, lipase hydrolytic activity of 3000UL-1, measured at pH9.0 and 37°C by using p-nitrophenyl palmitate as substrate, was achieved after 36h fermentation using glycerol-free production medium in a baffled Erlenmeyer flask at 28°C and pH6.8. Maximum productivity of 52.5UL-1h-1 was achieved after 24h in bioreactor using glycerol-free production medium at pH6.8 and 28°C, with agitation at 400rpm and aeration at 1vvm. Sc-WCL was shown to be more active at 60°C and pH10.7, while higher activity retention was observed at 30-40°C after 1h incubation at pH10. Sc-WCL showed to have potential to be used as biocatalyst in hydrolysis and esterification reactions. In the hydrolysis of p-nitrophenyl palmitate, lyophilized Sc-WCL expressed a hydrolytic activity (330unitsg-1 solid, measured at 37°C and pH9.0) around 100-fold higher than the ones declared by a supplier of lyophilized powders of mixtures of intracellular lipases from Thermus thermophiles and Thermus flavus (≥3.0unitsg-1 solid, measured at 65°C and pH8.0). In the synthesis of butyl butyrate in anhydrous medium, 85% ester conversion was achieved at 37°C after 8h reaction. Thus, Sc-WCL showed to be a promising biocatalyst because it is cheaper than the isolated and purified lipases.

The production of immobilized whole-cell lipase from Aspergillus nomius ST57 and the enhancement of the synthesis of fatty acid methyl esters using a two-step reaction

The production of immobilized whole-cell lipase from Aspergillus nomius ST57 was optimized by cultivation in 50mL of growth medium with the addition of 100, 0.5×0.5×0.5cm3 pieces of biomass support particles at 150rpm, 30°C for 84h. The hydrolytic activity of the immobilized whole-cell lipase was 30.77% higher than the activity obtained from free biomass (based on 1mg dry cell weight). Response surface methodology with a central composite design, which consisted of three parameters: the amount of the immobilized whole-cell lipase, the reaction temperature and the water content, was applied to optimize the synthesis of fatty acid methyl esters (FAME) from palm oil in a solvent-free system by the transesterification reaction using the immobilized whole-cell lipase from A. nomius ST57 as a biocatalyst. The highest percentage of FAME (78.23%) was obtained from a reaction mixture containing 1g palm oil, 1:4 molar ratio of oil to methanol, 23% (w/w of palm oil) of 50mM sodium acetate buffer pH 5 and 16 pieces of immobilized whole-cell lipase at 40°C and 1200rpm for 4h. The water-free transesterified palm oil mixture was then esterified using the immobilized whole-cell lipase. The highest percentage of FAME (94.77%) was obtained at 1g transesterified palm oil mixture, 16 pieces of immobilized whole-cell lipase and 0.06g of methanol for 2h. The immobilized whole-cell lipase retained 90.95% transesterification activity and 100% esterification activity after use for 10 batches.

Cell-bound lipases from Burkholderia sp. ZYB002: gene sequence analysis, expression, enzymatic characterization, and 3D structural model.

BackgroundThe whole-cell lipase from Burkholderia cepacia has been used as a biocatalyst in organic synthesis. However, there is no report in the literature on the component or the gene sequence of the cell-bound lipase from this species. Qualitative analysis of the cell-bound lipase would help to illuminate the regulation mechanism of gene expression and further improve the yield of the cell-bound lipase by gene engineering.ResultsThree predictive cell-bound lipases, lipA, lipC21 and lipC24, from Burkholderia sp. ZYB002 were cloned and expressed in E. coli. Both LipA and LipC24 displayed the lipase activity. LipC24 was a novel mesophilic enzyme and displayed preference for medium-chain-length acyl groups (C10-C14). The 3D structural model of LipC24 revealed the open Y-type active site. LipA displayed 96 % amino acid sequence identity with the known extracellular lipase. lipA-inactivation and lipC24-inactivation decreased the total cell-bound lipase activity of Burkholderia sp. ZYB002 by 42 % and 14 %, respectively.ConclusionsThe cell-bound lipase activity from Burkholderia sp. ZYB002 originated from a multi-enzyme mixture with LipA as the main component. LipC24 was a novel lipase and displayed different enzymatic characteristics and structural model with LipA. Besides LipA and LipC24, other type of the cell-bound lipases (or esterases) should exist.Electronic supplementary materialThe online version of this article (doi:10.1186/s12896-016-0269-6) contains supplementary material, which is available to authorized users.

Kinetic Study of Immobilized Whole-Cell Lipase Rhizopus Oryzae as Biocatalyst in Biodiesel Synthesis through Non-Alcohol Route

Kinetic Study of Immobilized Whole-Cell Lipase Rhizopus Oryzae as Biocatalyst in Biodiesel Synthesis through Non-Alcohol Route

The two-component GacS-GacA system activates lipA translation by RsmE but not RsmA in Pseudomonas protegens Pf-5.

In Pseudomonas spp., the Gac-Rsm signal transduction system is required for the production of lipases. The current model assumes that the system induces lipase gene transcription mediated through the quorum-sensing (QS) system. However, there are no reports of a QS system based upon N-acyl homoserine lactones or the regulation of lipase gene expression in Pseudomonas protegens. In this study, we investigated the regulatory mechanism acting on lipA expression activated by the Gac-Rsm system in P. protegens Pf-5 through deletion and overexpression of gacA, overexpression of rsmA or rsmE, expression of various lacZ fusions, reverse transcription-PCR analysis, and determination of whole-cell lipase activity. The results demonstrated that the GacS-GacA (GacS/A) system activates lipA expression at both the transcriptional and the translational levels but that the translational level is the key regulatory pathway. Further results showed that the activation of lipA translation by the GacS/A system is mediated through RsmE, which inhibits lipA translation by binding to the ACAAGGAUGU sequence overlapping the Shine-Dalgarno (SD) sequence of lipA mRNA to hinder the access of the 30S ribosomal subunit to the SD sequence. Moreover, the GacS/A system promotes lipA transcription through the mediation of RsmA inhibiting lipA transcription via an unknown pathway. Besides the transcriptional repression, RsmA mainly activates lipA translation by negatively regulating rsmE translation. In summary, in P. protegens Pf-5, the Gac-RsmE system mainly and directly activates lipA translation and the Gac-RsmA system indirectly enhances lipA transcription.

Molecular Identification of Lipase LipA from Pseudomonas protegens Pf-5 and Characterization of Two Whole-Cell Biocatalysts Pf-5 and Top10lipA

To identify lipase LipA (PFL_0617) from Pseudomonas protegens Pf-5, a lipA deletion mutant (Pf0617) and a complementary strain (Pf0617lipA) were constructed, and their effects on the lipase production were examined. Pf0617 remarkably decreased its whole-cell lipase activity, whereas Pf0617lipA made its whole-cell lipase activity not only restore to wild-type level but also get a further increment. However, the deletion and overexpression of lipA did not affect the extracellular lipase activity. In addition, the unbroken whole cells of these strains were able to catalyze the hydrolysis of membrane-permeable p-nitrophenyl esters, but could not hydrolyze the membrane-impermeable olive oil. These results confirmed that LipA was an intracellular lipase and Pf-5 could also be used as a natural whole-cell biocatalyst. To evaluate the potential of Pf-5 as a whole-cell biocatalyst and separately characterize the whole-cell LipA, the properties of the whole-cell lipases from Pf-5 and Top10lipA were characterized. The results demonstrated that both Pf-5 and Top10lipA exhibited high tolerance to alkaline condition, high temperature, heavy metal ions, surfactants, and organic solvents. Taken together, lipA can realize functional expression in E. coli Top10, and Pf-5 and Top10lipA as whole-cell biocatalysts may have enormous potential in applications.

Improving Yield of 1,3-Diglyceride by Whole-Cell Lipase fromA. NigerGZUF36 Catalyzed Glycerolysis via Medium Optimization

1,3-Diglyceride (1,3-DG) has attracted considerable attention as a healthful food oil component. In this study, we reported a novel method to synthesis of 1,3-DG via glycerolysis of triglyceride (TG) catalyzed by whole-cell lipase from a new isolated strain, Aspergillus niger GZUF36. The glycerolysis products include major diglyceride and minor monoglyceride. To improve yield of 1,3-DG, the optimized medium for the strain was obtained by combination of single-factor experiments, Plackett-Burman (PB), the steepest ascent and Box-Behnken design (BBD). Using the optimized medium, yield of 1,3-DG synthesized by the whole-cell lipase was 26.90 mol% and 1.42-fold yield was improved. The optimized medium lays the foundation for further studies of the reaction system for synthesis of 1,3-DG. The whole-cell catalyzed-glycerolysis process represents a significant advance toward achieving economical production of 1,3-DG at industrial scale.