Optimization of extraction methods for the analysis of antibiotics in poultry and mammalian meal

"In Portugal, the canning industries can generate up to 40 % of fish by-products that are mainly intended for animal feed. The main product resulting from the processing of fish products is fish oil due to own source of long chain polyunsaturated fatty acids, especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are part of the omega-3 series. Due to the high amount of free fatty acids (FFA) present in the fish oil it is necessary to reduce the acid value to make it apt for human or animal consumption. The high amount of FFA affects the quality of the fish oil, its processing and storage. This work aims to study the reduction of acidity of fish oil by enzymatic esterification with ethanol, converting FFA in ethyl esters, and thus contribute to improving their quality and consequently increase their commercial value. The sample of fish oil was collected in the Savinor UTS company and it was characterized for the following parameters: acidity value (12-14 mgKOH/gamostra), iodine value (158.2 giodo/100gamostra), density (939.4 kg/m3), kinematic viscosity (35.83 mm2/s) and moisture content (1.437 %). To catalyze the enzymatic esterification, four enzymes were tested, the Novozyme 435, the Lipozyme TL 100L, the Lipozyme CALB L and Lecitase Ultra by selecting the one that caused the greater reduction of the acid value (Lipozyme TL 100L). The reaction operating conditions were also studied, such as temperature (32.5; 35; 37.5; 40; 42.5; 45 and 47.5 °C), weight ratio enzyme: oil (0.0025; 0.005; 0.0075; 0.01 and 0.02 w/w) and weight ratio alcohol: FFA (1.62; 2.43; 3.24; 4.04 and 4.85 w/w). The operating conditions for which it was obtained a greater reduction of the acid value are: temperature of 40 °C, mass ratio of enzyme: oil 0.01 m/m, mass ratio alcohol: FFA of 3.24 w/w, in the presence of ethanol at 96 % v/v for 180 minutes. The esterification carried out with these operating conditions led to the removal of 75 % of acidity, reaching a final acid value of 3.13 mgKOH/gamostra corresponding to the FFA content of 1.57 %. With this percentage of FFA reduction, it is concluded that the quality of fish oil improved and ensured an increase of its commercial value. Also, it was concluded that the enzymatic esterification is the appropriate catalysis for the FFA reduction, due to the fact that it has shown good results, it is a clean reaction and is conducted at low temperatures."