Natália Melo Osório

In this chapter, some examples of laboratory protocols to produce nutritional structured lipids, namely human milk fat substitutes, dietetic triacylglycerols, and interesterified fat blends with improved functional and rheological properties, catalyzed by either immobilized commercial or noncommercial lipase preparations, are presented. The use of crude oils instead of refined counterparts to reduce oil purification costs is also addressed. In addition to batch synthesis, the continuous production in packed- or fluidized-bed bioreactors is addressed, as well as the evaluation of operational stability of the biocatalysts used (either in batch reuses or in continuous mode).

The sustainable production of healthy structured lipids (SLs) using oils extracted from agro-industry by-products or non-conventional lipid sources is of utmost importance in the framework of a circular bioeconomy, toward a zero-waste goal. In this study, low-calorie triacylglycerols (TAGs) containing a long-chain (L) fatty acid (FA) at position sn-2 and medium-chain (M) FAs at positions sn-1,3 (MLM type SL) were obtained from virgin cold-pressed milk thistle (51.55% linoleic acid; C18:2), grapeseed (66.62% C18:2), and apricot kernel (68.61% oleic acid; C18:1) oils. Lipase-catalyzed acidolysis with capric acid (C10:0) or interesterification with ethyl caprate (C10 Ethyl) in solvent-free media were performed. In batch reactions, immobilized Rhizomucor miehei lipase (Lipozyme RM) was used as a biocatalyst. For all tested oils, new TAG (SL) yields, varying from 61 to 63%, were obtained after 6 h of interesterification. Maximum new TAG yields were reached after 6, 24, and 30 h of acidolysis with grapeseed (64.7%), milk thistle (56.1%), or apricot kernel (69.7%) oils, respectively. Continuous acidolysis and interesterification of grapeseed oil were implemented in a packed-bed bioreactor, catalyzed by immobilized Thermomyces lanuginosus lipase (Lipozyme TL IM). Throughout 150 h of continuous operation, no lipase deactivation was observed, with average SL yields of 79.2% ± 4.1 by interesterification and 61.5% ± 5.91 by acidolysis.

Crude olive pomace oil (OPO) is a by-product of olive oil extraction. In this study, low-calorie structured triacylglycerols (TAGs) were produced by acidolysis of crude OPO with medium-chain fatty acids (caprylic, C8:0; capric, C10:0) or interesterification with their ethyl ester forms (C8EE, C10EE). These new TAGs present long-chain fatty acids (L) at position sn-2 and medium-chain fatty acids (M) at positions sn-1,3 (MLM). Crude OPO exhibited a high acidity (12.05–28.75% free fatty acids), and high contents of chlorophylls and oxidation products. Reactions were carried out continuously in a packed-bed bioreactor for 70 h, using sn-1,3 regioselective commercial immobilized lipases (Thermomyces lanuginosus lipase, Lipozyme TL IM; and Rhizomucor miehei lipase, Lipozyme RM IM), in solvent-free media at 40 °C. Lipozyme RM IM presented a higher affinity for C10:0 and C10EE. Lipozyme TL IM preferred C10:0 over C8:0 but C8EE over C10EE. Both biocatalysts showed a high activity and operational stability and were not affected by OPO acidity. The New TAG yields ranged 30–60 and the specific productivity ranged 0.96–1.87 g NewTAG/h.g biocatalyst. Lipozyme RM IM cost is more than seven-fold the Lipozyme TL IM cost. Therefore, using Lipozyme TL IM and crude acidic OPO in a continuous bioreactor will contribute to process sustainability for structured lipid production by lowering the cost of the biocatalyst and avoiding oil refining.

In this chapter, some examples of laboratory protocols to produce functional structured lipids, namely, human milk fat substitutes, dietetic triacylglycerols, and interesterified fat blends with improved nutritional and rheological properties, catalyzed either by immobilized commercial or noncommercial lipase preparations, are presented. In addition to batch synthesis, the continuous production in packed- or fluidized-bed bioreactors is addressed, as well as the evaluation of operational stability of the biocatalysts used (either in batch reuses or in continuous mode).

Low calorie triacylglycerols (TAG) presenting medium‐chain fatty acids (M) at positions sn‐1,3 and long‐chain fatty acids (L) at position sn‐2 are known as MLM. This study aims at the production of MLM by acidolysis of grapeseed oil with medium‐chain caprylic (C8:0) or capric (C10:0) acids. Grapeseed oil is used as source of long‐chain polyunsaturated fatty acids (FAs), especially linoleic acid, at sn‐2 position in TAG. Reactions are performed in batch, in solvent‐free systems, during 48 h. Novel non‐commercial sn‐1,3 regioselective lipases are used as alternative to high‐cost commercial biocatalysts, namely, the heterologous lipase from Rhizopus oryzae (rROL) immobilized in Amberlite™ IRA 96 and Carica papaya lipase (CPL) self‐immobilized in papaya latex. The highest productions of new TAG are achieved at 40 °C, molar ratio TAG:M of 1:2, after 48 h, with both biocatalysts, with yields varying between 38 and 69%. rROL immobilized in Amberlite IRA 96 shows a preference toward caprylic acid while CPL shows no preference toward caprylic or capric acid. First‐order deactivation kinetics is observed for both biocatalysts. Half‐lives at 40 °C are 166 and 118 h for rROL and 96 and 81 h for CPL, in the acidolysis of grapeseed oil with C8:0 or C10:0, respectively.Practical Applications: Grapeseeds of Vitis vinifera L. are a by‐product of the wine industry. Using grapeseed oil to produce added‐value functional oils rich in linoleic acid (essential fatty acid) may be a way of improving the revenues of the enological and oil sectors. Both lipases, and mainly Carica papaya lipase self‐immobilized in papaya latex, due to its low‐cost production and easy preparation, are promising non‐commercial biocatalysts for the synthesis of MLM in solvent‐free media. The use of a solvent‐free system, in addition of being a green option, is also preferred for economic reasons, avoiding the costs with solvent and solvent recovery. Also, the use of the stoichiometric molar ratio TAG:M of 1:2 will 1) decrease costs related with the recovery and reutilization of medium chain fatty acids in excess, product recovery and purification, and 2) avoid biocatalyst deactivation by high amounts of free fatty acids in reaction media.Both Carica papaya lipase (CPL) self‐immobilized in papaya latex and rROL in Amberlite IRA 96 catalyze the synthesis of MLM from grapeseed oil in solvent‐free medium. The highest productions of new TAG are achieved at 40 °C, molar ratio TAG:M of 1:2, after 48 h, with both biocatalysts, with yields varying between 38 and 69%. rROL immobilized in Amberlite IRA96 shows a preference toward caprylic acid while CPL shows no preference toward caprylic or capric acid.