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Production and Characterization of Whole-Cell Rhizopus oryzae CCT3759 to be Applied as Biocatalyst in Vegetable Oils Hydrolysis

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Abstract

The present study aims to produce a mycelium-bound lipase of the fungus Rhizopus oryzae CCT3759 by submerged fermentation in order to be applied as biocatalyst in the hydrolysis of different vegetable oils. Optimal cultivation conditions have been achieved in a medium containing olive oil as inducer for 72 h of fermentation, thus obtaining 30.5 g/L of dry biomass concentration and hydrolytic activity of 389.1 U/g, which corresponds to a total lipase activity around of 12,000 U/L. Maximum hydrolytic activity was observed at pH 6.0 and 40 °C. Kinetic parameters concerning apparent Michaelis–Menten constant (Km = 50.5 mM) and maximum reaction rate (Vmax = 815.4 µmol/g min) have been determined in olive oil emulsion hydrolysis. Thermal stability tests revealed that the enzyme retained 75% of its initial activity after 4 h at 50 °C, whose thermal inactivation constant (Kd) and half-life (t1/2) was 0.073 h−1 and 9.4 h, respectively. The effect of biocatalyst concentration, expressed as activity units—U (200 and 400 U), on the hydrolysis of vegetable oils was investigated under fixed conditions: oil/buffer mass ratio of 25% (m/m), 100 mM buffer sodium phosphate pH 6.0, 40 °C and the mechanical stirring frequency of 600 rpm. As expected, increasing the initial activity from 200 to 400 U leads to higher values of initial reaction rates and hydrolysis percentage. However, initial reaction rate values were similar for six different vegetable oils due to the high accessibility of the lipase to the substrate under such experimental conditions. A complete hydrolysis of olive, cottonseed, sunflower and canola oils has been achieved after 26–30 h of reaction using 400 U of activity. These results suggest a promising application of the produced biocatalyst in the production of free fatty acids, an important class of compounds for oleochemical industries.

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Acknowledgements

This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) – Finance Code 001. W.S.M.R. thanks CAPES for the student fellowship. A.A.M. thanks Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq – Brazil) for the research fellowship (Process 310633/2020-6).

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Authors and Affiliations

  1. Federal University of Alfenas (UNIFAL-MG), Alfenas, MG, 37130-001, Brazil

    Willian S. M. Reis, Adriano A. Mendes & Ernandes B. Pereira

  2. Laboratory of Bioprocesses, Federal University of Alfenas (UNIFAL-MG), Alfenas, MG, 37130-001, Brazil

    Alexandre B. Matias

  3. Institute of Chemistry, Federal University of Alfenas (UNIFAL-MG), Alfenas, MG, 37130-001, Brazil

    Adriano A. Mendes

  4. Department of Chemical Engineering, Engineering School of Lorena (EEL), University of São Paulo (USP), São Paulo, Brazil

    Heizir F. de Castro

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Contributions

Willian S. M. Reis and Alexandre B. Matias carried out the experimental work. Willian S. M. Reis, Adriano A. Mendes and Ernandes B. Pereira carried out the final editing of the manuscript and the writing of the article. Adriano A. Mendes, Ernandes B. Pereira and Heizir F. de Castro were responsible for conceptualization, supervision and funding acquisition. All authors have read and agreed to the published version of the manuscript.

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Correspondence to Willian S. M. Reis or Ernandes B. Pereira.

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Reis, W.S.M., Matias, A.B., Mendes, A.A. et al. Production and Characterization of Whole-Cell Rhizopus oryzae CCT3759 to be Applied as Biocatalyst in Vegetable Oils Hydrolysis. Catal Lett 152, 1–11 (2022). https://doi.org/10.1007/s10562-021-03622-8

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