Master of Science / Food Science Institute / Jayendra K. Amamcharla / In recent years, the interest in functional components such as phospholipids (PLs) is increasing as a result of growing awareness of their health benefits. PLs affect several cell functions, such as growth, molecular transport system, memory processing, stress responses, and central nervous system myelination. Many studies have shown that the neutral lipids can be successfully extracted using supercritical carbon dioxide (SCO₂) from different types of foods such as egg, canola, pumpkin seed, fish and dairy powders. It is an alternative method to avoid the use of large quantities organic solvents. The SCO₂ is a safe, environmentally friendly and economical process to extract edible lipids from a variety of matrices. However, a modifier such as ethanol is needed to fractionate PLs due to limited solubility of PLs in SCO₂. The objectives of this study were to optimize the SFE process parameters and to determine the effect of pressure, temperature, and ethanol concentration on the extraction efficiency of PLs from whey protein phospholipid concentrate (WPPC) and buttermilk powder (BMP). Three different batches of WPPC and BMP were obtained from a commercial manufacturer and followed a unique two-step extraction process to isolate PLs from WPPC and BMP. In Step-1, neat supercritical CO₂ was used to remove all the neutral lipids at 414 bar pressure, 60 °C sample temperature, and 5 L/min CO₂ flow rate. The spent solids, the powder left after the first step extraction, were used to extract PLs in the second step. The Step-2 (SCO₂-Ethanol) process was optimized in terms of pressure (350, 414 and 550 bar), temperature (40 °C and 60 °C) and concentration of ethanol (10%, 15% and 20%) as independent factors. All the lipid fractions were analyzed by high performance lipid chromatography (HPLC) and thin layer chromatography (TLC). For WPPC, only ethanol concentration had significant effect (P < 0.05) on the amount of PLs extracted after the Step-2. On the other hand, temperature and ethanol concentration were significantly (P < 0.05) affected the efficiency of SFE for BMP. The optimal processing conditions for WPPC and BMP were 350 bar pressure, 60 °C sample temperature and 15% concentration of ethanol, and 550 bar of pressure, 60 °C sample temperature and 15% concentration of ethanol, respectively. This study allowed obtaining PLs from dairy co-products such as WPPC and BMP as a separate ingredient and this could be useful in nutraceutical and infant formulations as well as different food products formulations.