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Enzymatic Modification of Antioxidants Towards Omega-3 Oil Protection

Publikation: Bog/antologi/afhandling/rapportPh.d.-afhandlingForskning


  • Zhiyong Yang, Danmark
  • Molekylærbiologisk Institut
This PhD dissertation entitled “Enzymatic Modification of Antioxidants Towards Omega-3 Oil Protection” is primarily focused on synthesize of novel antioxidant from natural sources for better protection of oxidation-prone omega 3 oil. Selected phenolic acids were conjugated with fatty alcohols in different chain length and triacylglycerol (TAG). Several synthesis strategies were evaluated.

Synthesis of lipophilic phenolic fatty alcohols esters were initially conducted in a binary organic solvent system, which was composed of hexane and butanone. Preliminary studies were conducted to identify the main parameters affecting yield of these novel lipophilized phenolic fatty alcohol esters in a binary solvent system. The parameters studied in these preliminary studies included volume ratio of hexane to butanone, chain lengths of fatty alcohols and types of phenolic acids (dihydrocaffeic acid (DHCA), ferulic acid, and caffeic acid). The optimal mixed volume ratio was found to be 65/35 (hexane/butanone, v/v), in which both substrates have reasonably good solubility without compromising the enzyme activity. Medium chain fatty alcohols were easier to be esterified with phenolic acids, and highest conversion was achieved when hexanol (C6) was used. It was possible to dissolve ferulic and caffeic acids in this reaction system, but the conversions of these two phenolic acids were much lower than DHCA. Based on the optimal mixed volume ratio from the preliminary experiments and using esterification of octanol and DHCA as model, effects of different reaction parameters (temperature, reaction time, enzyme load, substrate molar ratio) were evaluated according to the experimental designs generated by response surface methodology (RSM). Optimal reaction conditions were predicted by the model and conversion of DHCA was increased significantly to above 90%.

To further increase the productivity of lipophilic phenolic esters, another system using ionic liquid (IL) Methyltrioctylammonium Trifluoroacetate (tOMA•TFA) as an assisting solvent was developed. Several parameters including the effect of mixed volume ratio between tOMA•TFA and octanol, temperature, water removal techniques (molecular sieves or vacuum), and concentration of DHCA were studied. Conversion of DHCA increased slightly with increasing volume ratio of IL/octanol. There was a 10 to 20% higher conversion when the reaction was conducted at 80 oC and 70 oC respectively comparing to at 60 oC. Significant increase in conversion was observed when water was removed from the reaction. The conversion of DHCA was increased to 90% when molecular sieves were added, which is 30% higher than the reaction without molecular sieves. Moreover, the initial reaction rate increased as the concentration of DHCA was increased. Compared to the reactions conducted in parallel experiments where tOMA•TFA was not added, system composed of IL/octanol increased both solubility of DHCA and productivity of lipophilic phenolic esters significantly. This binary system was also proved to be very efficient in the conversion of ferulic and caffeic acids. Conversions were very low in the system without tOMA•TFA.

Besides synthesis of amphiphilic phenolic esters with fatty alcohol, other novel antioxidants can also be produced by conjugating phenolic acids with TAG. Two individual studies were conducted to achieve this goal. In the first study, enzymatic transesterification was conducted between octyl dihydrocaffeate and tricaprylin. Octyl dihydrocaffeate was initially synthesized from esterification of octanol with DHCA and then used for subsequent incorporation into tricaprylin. Due to the good compatibility between octyl dihydrocaffeate and TAG, an improved volumetric productivity and high enzymatic activity were obtained within 72 hour. In the second study, ferulate was incorporated directly into the highly unsaturated fish oil through enzymatic transesterification of ethyl ferulate (EF) with fish oil. In both of these studies, significant increases in production of phenolic TAG were achieved after optimizing the reaction conditions with RSM.

Micro-emulsion systems have the potential to be efficient synthesis reaction systems for the present study. Three different systems, phenolic acids (DHCA and ferulic) in IL/octanol, rutin in IL/linoleic, and ferulic in IL/triolein, were developed and investigated. These systems were characterized using dynamic light scattering (DLS) and small angle X-ray scattering (SAXS). It was suspected that the first system was deduced to be a homogenous system because no particles were found in it. Particles with a size of 45 nm and above 1000 nm were found in the second and third system respectively. Nevertheless, the third system composed of phenolic and IL/triolein was not as stable emulsion system as the second system composed of rutin and IL/linoleic.

Overall, the study in this dissertation explored several different routes for synthesis of lipophilic antioxidants. Several novel antioxidants were synthesized though esterification of selected phenolic acids with different fatty alcohols and TAG. These antioxidants are amphiphilic molecules and can be used to prevent oxidation in lipid-based food products.
ForlagAarhus Universitet
Antal sider147
StatusUdgivet - jul. 2011

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