Enzyme-aided production of lipid emulsifiers from side-streams of the food industry: rapeseed press cake and oat oil

Abstract: The goal of this thesis project was to utilize side-streams of the food industry, namely rapeseed cake and oat oil, by enzyme-aided production of lipid emulsifiers. The first objective was to extract lipids from the rapeseed press cake and increase protein yield, by adding an enzymatic step to the already existing protein extraction procedure developed by the food department of Lund University. A pre-treatment of the rapeseed press cake with Pectinex Ultra SP-L aimed to degrade the cell-wall matrix showed an increase in protein yield of approximately 10%. Lipids were not extracted as no separate oil layer was seen during any of the treatments. It is hypothesised that the lipids were hydrolysed during the procedure leading to the bitter taste of the extract and solubilisation of the lipids. No starting material for modification of lipids was therefore obtained from rapeseed press cake. The second objective was to determine the effect of enzymatic modification on the composition and emulsification properties of lipids. Two different oils were used: crude oat oil and polar lipid enriched oat oil (PL40). Firstly the unmodified oils were characterized. Column separation was performed, which successfully produced a neutral lipid, phospholipid and glycolipid fraction. Thin layer chromatography (TLC) showed that the neutral lipid fraction mainly contained di- and triglycerides and the phospholipid fraction contained phosphatidyl choline (PC), phosphatidyl ethanolamine (PE), phosphatidyl inositol (PI) and lyso-PC. Fatty acid profiles were determined by fatty acid methylation and gas chromatography. Main fatty acids in both oils were palmitic acid (16:0), oleic acid (18:1) and linoleic acid (18:2). Minor fatty acids were palmitoleic acid (16:1), stearic acid (18:0) linolenic acid (18:3), arachidic acid (20:0), eicosenoic acid (20:1) and avenoleic acid (18:2 15OH), of which the latter was only found in the glycolipid fraction. Differences in fatty acid profiles of the two oils could be explained by the profiles of the different fractions. Emulsion stability was assessed by a fine-tuned spectrophotometric method supplemented by visual observations and light microscopy. Stability was significantly increased by the presence of unmodified oils, especially by PL40 oil. Surprisingly, contact time between oat oil and water before mixing also significantly affected stability, which was ascribed to the formation of liquid crystals. Modification of the lipids was performed by a lipase from Rhizopus arrhizus and hydrolysed lipids were initially extracted by adding chloroform and methanol. However, this method was inconsistent and three other extraction methods were tested. Addition of water, chloroform and methanol proofed most appropriate and was used for further experiments. Emulsion stabilizing ability increased as expected with incubation time for both crude oat oil and PL40 oil. Unexpectedly, the PL40 oil incubated without enzyme showed a higher stability than the hydrolysed oil. It is hypothesised that due to the hydrolysis more elaborate lipid structures are formed, which prevent coalescence but can cover less area than unmodified lipids, leading to bigger droplets and an increased creaming rate. From the results it is clear that the polar lipids in oat oil can be utilized as emulsifiers of which the properties can be changed by enzymatic hydrolysis. In conclusion it can be said that both rapeseed press cake and oat oil can be utilized to a greater extent by aid of enzymes. The fine-tuned methods reported in this thesis for enzymatic hydrolysis and assessing change in molecular structure and emulsification properties can be used as a starting point for future research. This could eventually lead to implementation in industry and an increased utilisation of the side-streams of the food industry.