Novozym® 435 and Lipozyme® RM IM as Biocatalysts for Benzyl Benzoate Synthesis

With the ever-increasing demand for clean technology in the industrial sector, natural methods, such as enzyme-catalyzed, represent a sustainable alternative to industrial chemical processes. In this context, the synthesis of benzyl benzoate ester using commercial immobilized lipases was evaluated. For this, a kinetic study was carried out to determine the reaction time (24 h) and enzyme concentration (10 wt%). Then, a 22 full factorial design was proposed to evaluate the effect of molar ratio (benzyl alcohol to benzoic anhydride) and temperature on conversion of benzyl benzoate in the presence of tert-butanol as solvent. For the Novozym® 435, maximum conversion (32%) was achieved at 60 ºC, using a molar ratio of 1:5 (alcohol to anhydride). A maximum conversion of 51% was obtained for Lipozyme® RM IM at 40 ºC and the molar ratio of 1:5. The benzyl benzoate showed moderate antimicrobial action against S. aureus (MIC = 0.05 mg μL-1). With the results, the conclusion was that the methodology of design of experiments was adequate for the proposed system and allowed the optimization of the production of benzyl benzoate.

Co-Immobilization of Rhizopus oryzae and Candida rugosa Lipases onto mMWCNTs@4-arm-PEG-NH2—A Novel Magnetic Nanotube–Polyethylene Glycol Amine Composite—And Its Applications for Biodiesel Production

This article describes the successful synthesis of a novel nanocomposite of superparamagnetic multi-walled nanotubes with a four-arm polyethylene glycol amine polymer (mMWCNTs@4-arm-PEG-NH2). This composite was then employed as a support for the covalent co-immobilization of Rhizopus oryzae and Candida rugosa lipases under appropriate conditions. The co-immobilized lipases (CIL-mMWCNTs@4-arm-PEG-NH2) exhibited maximum specific activity of 99.626U/mg protein, which was 34.5-fold superior to that of free ROL, and its thermal stability was greatly improved. Most significantly, CIL-mMWCNTs@4-arm-PEG-NH2 was used to prepare biodiesel from waste cooking oil under ultrasound conditions, and within 120 min, the biodiesel conversion rate reached 97.64%. This was due to the synergy effect between ROL and CRL and the ultrasound-assisted enzymatic process, resulting in an increased biodiesel yield in a short reaction time. Moreover, after ten reuse cycles, the co-immobilized lipases still retained a biodiesel yield of over 78.55%, exhibiting excellent operational stability that is attractive for practical applications. Consequently, the combined use of a novel designed carrier, the co-immobilized lipases with synergy effect, and the ultrasound-assisted enzymatic reaction exhibited potential prospects for future applications in biodiesel production and various industrial applications.

A review between whole-cells and immobilized lipases technologies for biodiesel and flavor esters production

As lipases são enzimas versáteis que catalisam diversas reações e podem ser aplicadas em várias áreas da indústria. No entanto, a biocatálise enfrenta um obstáculo econômico para poder competir com as rotas convencionais. Diferentes estratégias têm sido estudadas com o objetivo de reduzir o custo dos biocatalisadores e melhorar a atividade e estabilidade catalítica. A imobilização enzimática é uma das estratégias mais eficientes para tornar a aplicação de enzimas competitiva em larga escala industrial, proporcionando reutilização contínua, facilidade de separação do meio reacional e maior eficiência do processo. No entanto, apesar dos avanços tecnológicos alcançados, os biocatalisadores ainda são caros para usos industriais devido às etapas de recuperação, purificação e imobilização de enzimas. Nesse contexto, surge a tecnologia de células integras (CI) como uma forma de imobilização, na qual as próprias células, geralmente cultivadas em um suporte, são aplicadas no processo de biotransformação, contendo as proteínas de interesse aderidas à sua superfície, permitindo fácil separação, reutilização e dispensando a etapa de purificação. Embora a tecnologia de células inteiras tenha se tornado uma ferramenta valiosa para muitos processos de biotransformação, existem alguns inconvenientes inerentes, que impediram seu avanço em escala industrial, e muitos estudos foram realizados com o objetivo de otimizar seu desempenho. Este artigo apresenta uma revisão entre as tecnologias de células íntegras e de lipases imobilizadas, principalmente relacionadas às sínteses de biodiesel e de ésteres de sabor, uma vez que estas são amplamente relatadas na literatura.

Comparative Study on Lipase Immobilized onto Organo-Cation Exchanged Kaolin and Metakaolin: Surface Properties and Catalytic Activity

Clay mineral has received much attention to be used as biocatalysts as it is cheaper, easily available and environmentally friendly. However, the use of unmodified clay, in particular kaolin for enzyme immobilization showed unsuitability of this support due to its negative charge. In this study, the hydrophobic properties of kaolin and metakaolin (kaolin heated to 650 °C) were adjusted by the intercalation with benzyltriethylammonium chloride (BTEA-Cl), at concentrations 2.0 times the cation exchange capacities (CEC) of the clays. The supports were then used for immobilization of lipase from Candida rugosa (CRL). From the study, the highest percentage of lipase immobilization was achieved (70.14%), when organo-modified metakaolin (2.0 MK) was used. The supports as well as the immobilized biocatalysts were then characterized by X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy, and nitrogen adsorption techniques. Comparisons of the efficiencies of immobilized with free CRL in the synthesis of nonyl hexanoate showed that immobilized CRL achieved enzymatic activities of between 5.24×10−3 to 3.63×10−3 mmol/min/mg, while free CRL achieved enzymatic activity of 3.27×10−3 mmol/min/mg after 5 h of reaction at 30 ℃. The immobilized CRLs also maintained 70.81% – 80.59% thermostabilities at 70 ℃ as compared to the free CRL (28.13%). CRL immobilized on 2.0 NK and 2.0 MK also maintained 38.54% and 62.56%, respectively, of the initial activities after 10 continuous cycles, showing the excellent stability and reusability of the immobilized lipases, suitable as substitute for expensive, hazardous catalysts used in industries. Copyright © 2021 by Authors, Published by BCREC Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).

Design of a New Gemini Lipoaminoacid with Immobilized Lipases Based on an Eco-Friendly Biosynthetic Process

Lipoaminoacids (LAA) are an important group of biosurfactants, formed by a polar hydrophilic part (amino acid) and a hydrophobic tail (lipid). The gemini LAA structures allow the formation of a supramolecular complex with bioactive molecules, like DNA, which provides them with good transfection efficiency. Since lipases are naturally involved in lipid and protein metabolism, they are an alternative to the chemical production of LAA, offering an eco-friendly biosynthetic process option. This work aimed to design the production of novel cystine derived gemini through a bioconversion system using immobilized lipases. Three lipases were used: porcine pancreatic lipase (PPL); lipase from Thermomyces lanuginosus (TLL); and lipase from Rizhomucor miehei (RML). PPL was immobilized in sol-gel lenses. L-cystine dihydrochloride and dodecylamine were used as substrates for the bioreaction. The production of LAA was evaluated by thin layer chromatography (TLC), and colorimetric reaction with eosin. The identification and quantification was carried out by High Performance Liquid Chromatographer-Mass Spectrometry (HPLC-MS/MS). The optimization of media design included co-solvent (methanol, dimethylsulfoxide), biphasic (n-hexane and 2-propanol) or solvent-free media, in order to improve the biocatalytic reaction rates and yields. Moreover, a new medium was tested where dodecylamine was melted and added to the cystine and to the biocatalyst, building a system of mainly undissolved substrates, leading to 5 mg/mL of LAA. Most of the volume turned into foam, which indicated the production of the biosurfactant. For the first time, the gemini derived cystine lipoaminoacid was produced, identified, and quantified in both co-solvent and solvent-free media, with the lipases PPL, RML, and TLL.

Solvent-free esterifications mediated by immobilized lipases: a review from thermodynamic and kinetics perspective

Esters are a highly relevant class of compounds in the industrial context, and biocatalysis applied to esters syntheses is already a reality for some chemical companies. Their syntheses in solvent-free...