A novel hybrid biocatalyst from immobilized Eversa^® Transform 2.0 lipase and its application in biolubricant synthesis

Abstract

In this study, a Taguchi experimental design was used for optimzing the immobilization of the lipase Eversa^® Transform 2.0 (EVS) onto a hybrid support consisting of chitosan (CHI) and agarose (AGA), with glutaraldehyde (GLU) used as the support activator. The biocatalyst obtained was characterized by X-Ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FTIR), Thermogravimetry (TGA), Energy Dispersive Spectroscopy (EDS), and Scanning Electron Microscopy (SEM). The optimized reaction conditions (60 min, 5 mM ionic strength, 1% GLU concentration, and 5 mg protein load per g of support) resulted in a highly active biocatalyst (74.39 ± 0.48 U/g) and delivered an immobilization yield of 74.20 ± 0.28%. The biocatalyst produced was observed to lose only 15.3% of its activity after 61 days of storage. The activity was also observed to increase by 96.70% ± 0.76, 27.34% ± 2.34, and 84.35% ± 1.68 in the presence of the organic solvents hexane, cyclohexane, and methanol, respectively. Additionally, the byocatalist showed more pronounced activity at temperatures above 50 °C and was still able to retain approximately 30% of it at 70 °C. These values were found to be higher at alkaline pHs, as the activity of Eversa^® 2.0 Transform saw an increase of up to 140% at pH 9. The desorption tests performed did not reveal any enzymatic detachment from the support. The novel biocatalyst also showed promising ester-lubricating properties. Furthermore, the in silico study revealed a binding affinity of −5.1 kcal/mol between oleic acid and the enzyme, suggesting that the combination of the substrate and the lipase was more stable and therefore, suitable for esterification.

Graphical Abstract

Keywords:

• Eversa^® Transform 2.0
• enzymatic immobilization
• chitosan
• agarose
• biolubricants
• docking studies

Disclosure statement

The authors and funding agencies have no conflicts of interest to declare. No conflicts, informed consents, or human or animal rights apply to this paper.

Additional information

Funding

The authors acknowledge and are grateful for the financial support of the following Brazilian Scientific and Technological Development Agencies: Conselho Nacional de Desenvolvimento Científico e Tecnológico [311062/2019-9], and Coordenação de Aperfeiçoamento de Ensino Superior. Centro Nordestino de Aplicação e Uso da Ressonância Magnética Nuclear (CENAUREMN) for NMR spectra.