Production of low-calorie structured lipids from spent coffee grounds or olive pomace crude oils catalyzed by immobilized lipase in magnetic nanoparticles
Graphical abstract
Introduction
Lipids are important compounds of the daily diet providing energy, essential fatty acids, liposoluble vitamins and antioxidants. The nutritional value of triacylglycerols (TAG) is determined by the fatty acid (FA) composition and the positional distribution of FA. TAG structure can be modified or restructured to obtain novel TAG (structured lipids, SL) with improved nutritional or functional properties. Low-calorie TAG are examples of structured lipids with improved benefits compared to the natural oils and fats. These TAG are usually of the type MLM since they contain a long-chain fatty acid (L) at the position sn-2 and medium-chain fatty acids (M), at the positions sn-1,3 (Kim and Akoh, 2015). They present a caloric value of 5–7 kcal/g against 9 kcal/g of natural fats (Smith et al., 1994), can be synthesized either (i) by acidolysis of a TAG or an oil containing long-chain fatty acids, with a medium-chain fatty acid (Costa et al., 2018), or (ii) by interesterification of a TAG or oil with methyl or ethyl esters of medium-chain fatty acids (Ferreira-Dias et al., 2019, Zhang et al., 2020). Ethyl esters are preferred over methyl esters as acyl donors for the production of food products due to the toxicity of methanol.
Structured lipids can be synthesized either enzymatically or chemically. Chemical methods are inexpensive compared to the enzymatic methods but lack specificity. Thus, the production of SL such as MLM can only be efficient if sn-1,3 regioselective lipases (EC 3.1.1.3.; triacylglycerol acyl-hydrolase) are used as catalysts. They can be used non-immobilized or immobilized in solid supports in order to be reused in batch or used in continuous bioreactors, decreasing process costs. Currently, the high prices of commercial immobilized lipases, together with their frequently low operational stability, have been recognized as the major constraints for industrial implementation of lipase-catalyzed processes. Thus, the search for novel supports for immobilization promoting high activity and operational stability is needed to develop sustainable industrial enzymatic processes. Magnetic nanoparticles (MNP) have been recently used for enzyme immobilization (Alex et al., 2014, Xu et al., 2014) due to their unique properties such as superparamagnetism, high surface area, large surface-to-volume ratio, lower mass transfer resistance and lesser fouling, easy separation under external magnetic fields, long term storage and re-usability (Xie and Ma, 2009). Moreover, functionalized MNP may facilitate covalent bonding of enzymes increasing their stability, activity and reusability. Magnetic nanoparticles appear to be very promising for their applications in oil biotransformation (Alex et al., 2014).
In lipase-catalyzed synthesis of SL, the use of ethyl esters as acyl donors instead of free fatty acids (FFA) may be a way to avoid the presence of high amounts of FFA in the reaction medium with inactivation effects on some enzymes (Tecelão et al., 2019). Also, ethyl esters, being more volatile than free fatty acids, are more easily recovered from the reaction media by distillation, which leads to more cost-effective downstream processing. In addition, the use of cheap oils extracted from agro-industry residues, as raw materials for SL synthesis, will contribute for the implementation of low-cost sustainable enzymatic processes. Spent coffee grounds (SCG) and olive pomace (OP) are examples of agro-food residues with significant environmental impacts (Cho et al., 2020). Both OP and SCG are produced in large amounts around the world, approximately 2.9 million and 8,5 million tons per year, respectively (Nasopoulou and Zabetakis, 2013, Nguyen et al., 2020). Therefore, it is important to valorize the by-products that result from olive and coffee processing. The OP is obtained after olive oil extraction by mechanical processes. This residue still contains about 3–4.5% (wet basis) of residual oil, known as olive pomace oil, with a composition similar to that of the olive oil. This oil may be extracted by solvent (usually n-hexane) and is currently used for edible purposes after refining (refined olive pomace oil or refined olive residue oil). Also, olive pomace and spent coffee grounds oils have been used for biodiesel production (Lama-Muñoz et al., 2014, Tarigan et al., 2019). Both OP and SCG are important sources of edible oils rich in long chain fatty acids (FA). SCG oil consists predominantly of linoleic acid (C18:2, 42.1–44.1%) and palmitic acid (C16:0, 32.9–33.6%) (Ramos-Andrés et al., 2019, Tarigan et al., 2019), while the OP oil consists predominantly of oleic acid (C18:1, 55–85%) (EEC, 1991). The use of SCG and OP oils to produce SL will be a revenue for the coffee and olive oil extraction sectors, respectively.
In the present study, the crude oils extracted from SCG and OP were used as raw-material for the production of low-calorie SL, either by acidolysis with capric acid, or by interesterification with ethyl caprate, in solvent-free media, catalyzed by immobilized lipases. The Rhizopus oryzae lipase (ROL) was immobilized in glutaraldehyde functionalized magnetite nanoparticles (MNP–ROL) and tested as a novel biocatalyst. The results obtained with MNP-ROL were compared with the performance of the commercial immobilized preparation of Thermomyces lanuginosus lipase (Lipozyme TL IM) in the same reactions.
Section snippets
Materials
The olive pomace crude oil was provided by the olive pomace oil extraction plant of UCASUL – União de Cooperativas UCRL (Alvito, Portugal). The spent coffee grounds (SCG) were kindly supplied by coffee shops in Brazil. Iron (II) chloride (FeCl2), Iron (III) chloride (FeCl3), 3-Triethoxysilylpropylamine (APTES) and the lyophilized Rhizopus oryzae lipase (ROL) were from Sigma Chemical Co. (St. Louis, MO, USA). The commercial sn-1,3 regioselective Thermomyces lanuginosus lipase immobilized on a
Characterization of the magnetic nanoparticles
The MNP presented density of 1.1 ± 0.1. APTES-modified MNP showed activity because of the higher density of enzymes immobilized on the pre-treated nanoparticles. These results demonstrated that surface modification of MNP with APTES enables ROL to be covalently immobilized on the surface of MNP, thereby maintaining the catalytic activity of enzyme (Park et al., 2009).
The DLS analysis of the magnetic nanoparticles indicated an average particle size of 122 nm which has been theoretically
Conclusion
This study shows that crude oils from SCG and OP can be valorized to produce low-calorie structured lipids, by acidolysis or interesterification, catalyzed by sn-1,3 regioselective lipases. Rhizopus oryzae lipase immobilized in magnetic nanoparticles showed to be a promising biocatalyst for SL production presenting (i) higher activity than Lipozyme TL IM, with both oils, with a preference towards acidolysis, and (ii) high stability when reused in acidolysis and interesterification of OP oil.
CRediT authorship contribution statement
Danyelle A. Mota: Conceptualization, Writing - original draft, Writing - review & editing. Devi Rajan: . Giuditta C. Heinzl: Methodology, Writing. Natália M. Osório: Conceptualization, Methodology, Writing - review & editing, Supervision. Jorge Gominho: Conceptualization, Methodology, Writing - review & editing, Supervision. Laiza C. Krause: Conceptualization, Supervision. Cleide M.F. Soares: Conceptualization, Methodology, Writing - review & editing, Resources, Funding acquisition,
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
The authors thank Mr. Ramiro Martínez, from Novozymes, Madrid, Spain, for providing the commercial immobilized lipase “Lipozyme TL IM”, and UCASUL- União de Coooperativas UCRL, Alvito, Portugal, for providing the crude olive pomace oil used in this study. The authors also thank Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Apoio à Pesquisa e à Inovação Tecnológica do Estado de Sergipe (FAPITEC) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
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