Progress and perspectives of enzymatic preparation of human milk fat substitutes.

Human milk fat substitutes (HMFS) with triacylglycerol profiles highly similar to those of human milk fat (HMF) play a crucial role in ensuring the supply in infant nutrition. The synthesis of HMFS as the source of lipids in infant formula has been drawing increasing interest in recent years, since the rate of breastfeeding is getting lower. Due to the mild reaction conditions and the exceptionally high selectivity of enzymes, lipase-mediated HMFS preparation is preferred over chemical catalysis especially for the production of lipids with desired nutritional and functional properties. In this article, recent researches regarding enzymatic production of HMFS are reviewed and specific attention is paid to different enzymatic synthetic route, such as one-step strategy, two-step catalysis and multi-step processes. The key factors influencing enzymatic preparation of HMFS including the specificities of lipase, acyl migration as well as solvent and water activity are presented. This review also highlights the challenges and opportunities for further development of HMFS through enzyme-mediated acylation reactions.

Immobilization of Lipase from Thermomyces lanuginosus in Magnetic Macroporous ZIF-8 Improves Lipase Reusability in Biodiesel Preparation.

In recent years, metal-organic frameworks (MOFs) have emerged as a promising support for immobilizing enzymes due to their high designability and structural diversity. Previous studies show that MOFs with single-crystal-ordered macroporous structures can effectively improve the accessibility of large-size enzyme and reduce the mass transfer resistance compared to conventional MOFs. In order to further enhance the reusability of lipase immobilized on macroporous MOFs, modification of MOFs through some magnetic particles could be an efficient approach. In this work, magnetic macroporous zeolitic imidazolate framework-8 (ZIF-8), referred to as m-M-ZIF-8 (with an average macropore size of about 140 nm), was synthesized and used for the immobilization of Thermomyces lanuginosus lipase (TLL). It was found that enzyme loading and the specific enzyme activity of the immobilized lipase were greatly enhanced through this magnetic modification. The enzyme loading of TLL@C-ZIF-8, T LL@M-ZIF-8, and TLL@m-M-ZIF-8 was 0.060, 0.074, and 0.076 mg/mg respectively. Besides, the activity of 93.5% was maintained after the immobilized lipase being repeatedly used for five batches, which was much higher than that of the immobilized lipase without magnetic modification, which was only 73.4%.

An overview to process design, simulation and sustainability evaluation of biodiesel production.

The overwhelming concerns due to over exploitation of fossil resources necessitate the utilization of alternative energy resources. Biodiesel has been considered as one of the most adaptable alternative to fossil-derived diesel with similar properties and numerous environmental benefits. Although there are various approaches for biodiesel production, development of cost-effective and robust catalyst with efficient production methods and utilization of a variety of feedstock could be the optimum solution to bring down the production cost. Considering the complexity of biodiesel production processes, process design, quantitative evaluation and optimization of the biodiesel from whole systems perspectives is essential for unlocking the complexity and enhancing the system performances. Process systems engineering offers an efficient approach to design and optimize biodiesel manufacturing systems by using a variety of tools. This review reflects state-of-the-art biodiesel research in the field of process systems engineering with a particular focus on biodiesel production including process design and simulation, sustainability evaluation, optimization and supply chain management. This review also highlights the challenges and opportunities for the development of potentially sustainable and eco-friendly enzymatic biodiesel technology.

Lipase Immobilization on Macroporous ZIF-8 for Enhanced Enzymatic Biodiesel Production.

Immobilization of enzyme on metal-organic frameworks (MOFs) has drawn increasing interest owing to their many well-recognized characteristics. However, the pore sizes of MOFs (mostly micropores and mesopores) limit their application for enzyme immobilization to a great extent owing to the large size of enzyme molecules. Synthesis of MOFs with macropores would therefore solve this problem, typically encountered with conventional MOFs. In this work, macroporous zeolitic imidazolate frameworks (ZIF-8), referred to as M-ZIF-8, were synthesized and used for immobilization of Aspergillus niger lipase (ANL). Immobilization efficiency using M-ZIF-8 and enzymatic catalytic performance for biodiesel preparation were investigated. The immobilized ANL on M-ZIF-8 (ANL@M-ZIF-8) showed higher enzymatic activity (6.5-fold), activity recovery (3.8-fold), thermal stability (1.4- and 3.4-fold at 80 and 100 °C, respectively), reusability (after five cycles, 68% of initial activity was maintained), and porosity than ANL on conventional ZIF-8 (ANL/ZIF-8). In addition, by using ANL@M-ZIF-8 for catalyzing a biodiesel production reaction, a higher fatty acid methyl ester yield was achieved.

Kinetics and Mechanism of Solvent Influence on the Lipase-Catalyzed 1,3-Diolein Synthesis.

1,3-Diacylglycerol preparation has roused increasing attention in recent years as the 1,3-diacylglycerol-rich oils can suppress the deposition of visceral fat and prevent the body weight increasing. Lipozyme TL IM-mediated esterification of oleic acid with monoolein was effective for 1,3-diacylglycerol production. During the esterification process, the solvent shows obvious influence on the diolein synthesis as well as the 1,3-diolein production. This work investigated the related kinetics and mechanism of the solvent effect on the esterification and Lipozyme TL IM performance. The results indicated that both the esterification rate constant and the acyl migration rate constant positively correlated with the logP of the solvent, while the site specificity of lipase has negative correlation with solvent logP. The acylation toward the 2-position of 1-monoolein was more sensitive to the solvent logP compared to the 1-position of glycerides. Molecular dynamics simulation revealed that solvents with different logP influenced the structure of Lipozyme TL IM including RMSD, hydrogen bond, and radial distribution function to a large extent, which subsequently led to the catalytic activity and selectivity variation of the lipase.

Hydrophobic pore space constituted in macroporous ZIF-8 for lipase immobilization greatly improving lipase catalytic performance in biodiesel preparation.

BACKGROUND: During lipase-mediated biodiesel production, by-product glycerol adsorbing on immobilized lipase is a common trouble that hinders enzymatic catalytic activity in biodiesel production process. In this work, we built a hydrophobic pore space in macroporous ZIF-8 (named as M-ZIF-8) to accommodate lipase so that the generated glycerol would be hard to be adsorbed in such hydrophobic environment. The performance of the immobilized lipase in biodiesel production as well as its characteristics for glycerol adsorption were systematically studied. The PDMS (polydimethylsiloxane) CVD (chemical vapor deposition) method was utilized to get hydrophobic M-ZIF-8-PDMS with hydrophobic macropore space and then ANL (Aspergillus niger lipase) was immobilized on M-ZIF-8 and M-ZIF-8-PDMS by diffusion into the macropores. RESULTS: ANL@M-ZIF-8-PDMS presented higher enzymatic activity recovery and better biodiesel production catalytic performance compared to ANL@M-ZIF-8. Further study revealed that less glycerol adsorption was observed through the hydrophobic modification, which may attribute to the improved immobilized lipase performance during biodiesel production and ANL@M-ZIF-8-PDMS remained more than 96% activity after five cycles' reuse. Through secondary structure and kinetic parameters' analysis, we found that ANL@M-ZIF-8-PDMS had lower extent of protein aggregation and twice catalytic efficiency (V max/K m) than ANL@M-ZIF-8. CONCLUSIONS: Hydrophobic pore space constituted in macroporous ZIF-8 for lipase immobilization greatly improved lipase catalytic performance in biodiesel preparation. The hydrophobic modification time showed negligible influence on the reusability of the immobilized lipase. This work broadened the prospect of immobilization of enzyme on MOFs with some inspiration.

Integrated Production of Biodiesel and Concentration of Polyunsaturated Fatty Acid in Glycerides Through Effective Enzymatic Catalysis.

DHA (docosahexaenoic acid) and EPA (eicosapentaenoic acid) contained in glycerides have been reported to be more advantageous for their intake than their counterpart in the form of free fatty acid or fatty acid esters. This work attempts to achieve the flexible concentration of DHA and EPA in glycerides as well as biodiesel production via a two-step process catalyzed by lipases. In the first step, several commercial lipases were investigated and Novozym ET2.0 demonstrated the highest potential in selective concentration of DHA and EPA. Over 85% of EPA and other fatty acids were converted to its corresponding FAEEs (fatty acid ethyl esters), while over 80% of DHA remained in glycerides under the optimized conditions. After the first step ethanolysis, the oil phase was subject to molecular distillation and a 97.5% biodiesel (FAEE) content could be obtained. Further flexible enrichment of DHA and EPA in glycerides was realized by immobilized lipase Novozym 435-mediated transesterification of glycerides (remaining in the heavy phase after molecular distillation) with DHA- or EPA-rich EE, and glycerides with 67.1% DHA and 13.1% EPA, or glycerides with 41.1% EPA and 38.0% DHA could be obtained flexibly. This work demonstrated an effective approach for DHA and EPA enrichment combined with biodiesel production through enzymatic catalysis.

[Pretreatment of oil palm residues by dilute alkali for cellulosic ethanol production].

In the study, we used oil palm residues (empty fruit bunch, EFB) as raw material to produce cellulosic ethanol by pretreatment, enzymatic hydrolysis and fermentation. Firstly, the pretreatment of EFB with alkali, alkali/hydrogen peroxide and the effects on the components and enzymatic hydrolysis of cellulose were studied. The results show that dilute alkali was the suitable pretreatment method and the conditions were first to soak the substrate with 1% sodium hydroxide with a solid-liquid ratio of 1:10 at 40 degrees C for 24 h, and then subjected to 121 degrees C for 30 min. Under the conditions, EFB solid recovery was 74.09%, and glucan, xylan and lignin content were 44.08%, 25.74% and 13.89%, respectively. After separated with alkali solution, the pretreated EFB was washed and hydrolyzed for 72 h with 5% substrate concentration and 30 FPU/g dry mass (DM) enzyme loading, and the conversion of glucan and xylan reached 84.44% and 89.28%, respectively. We further investigated the effects of substrate concentration and enzyme loading on enzymatic hydrolysis and ethanol batch simultaneous saccharification and fermentation (SSF). The results show that when enzyme loading was 30 FPU/g DM and substrate concentration was increased from 5% to 25%, ethanol concentration were 9.76 g/L and 35.25 g/L after 72 h fermentation with Saccharomyces cerevisiae (inoculum size 5%, V/V), which was 79.09% and 56.96% of ethanol theory yield.

[Stability of whole cell biocatalyst for biodiesel production from renewable oils].

Lipase-mediated biodiesel production becomes increasingly important because of mild reaction conditions, pollution free during the process and easy product separation. Compared with traditional immobilized lipase, whole cell biocatalyst is promising for biodiesel production because it is easy to prepare and has higher enzyme activity recovery. Rhizopus oryzae IFO4697 can be used as the catalyst for biodiesel production. To further study the stability of the whole cell biocatalyst is extremely important for its further application on large scale. This paper focuses on the stability study of Rhizopus oryzae IFO4697 when used for the methanolysis of renewable oils for biodiesel production. The results showed that water content was important for achieving high catalytic activity and good stability of the biocatalyst. The optimum water content was found to be 5%-15%. Both particle size and desiccation methods showed no obvious effect on the stability of the biocatalyst. With GA cross-linking pretreatment, the stability of the biocatalyst could be improved significantly. When Rhizopus oryzae IFO4697 repeatedly used for next batch reaction, direct vacuum filtration was found to be a good way for the maintenance of good stability of the biocatalyst. Under the optimum reaction conditions, the methyl ester yield could keep over 80% during 20 repeated reaction batches.

Mechanism exploration during lipase-mediated methanolysis of renewable oils for biodiesel production in a tert-butanol system.

tert-Butanol has been developed as a novel reaction medium for lipase-mediated methanolysis of renewable oils for biodiesel production, in which lipase could maintain high catalytic activity, although the log P value of tert-butanol was just about 0.35. The related mechanism exploration has been carried out, and it has been proposed first in this manuscript that in the biodiesel production system, log P(environment) (log P(environment) = x(methanol) log P(methanol) + x(oils) log P(oils) + x(solvent) log P(solvent)) including reactants and organic solvent should be taken into account to consider the effect of the whole environment on lipase activity instead of just considering the effect of the organic solvent itself. Further study showed that the operational stability of the lipase could be improved significantly in this system and there was no loss in lipase activity even after its being continuously used for 200 batches. The phase diagrams of the ternary-components tert-butanol/methanol/rapeseed oils were plotted further, and it was found that the methanol tolerance was the saturated methanol concentration in the system. It was demonstrated first here that the improved stability of the lipase was due to the elimination of negative effects caused by methanol and byproduct glycerol in the tert-butanol system.