Study On Ultrasound Enhanced Sub-critical CO₂ Extraction Of Lutein From Chlorella

Many drawbacks in traditional extraction techniques had hindered the further application of natural active ingredient, such as low efficiency, pollution and organic solvent residual. The supercritical fluid extraction attracted a lot of attention, but the high demand of pressure, energy and investment had impeded its large-scale application. As a result, the sub-critical CO2 extraction (SCCE) technique emerged. As a modified method of the supercritical CO2 extraction (SCE) technique, SCCE has lower temperature and pressure, but with higher density and extraction power. These advantages enable SCCE a better technique to extract the ingredient that was heat sensitive, easily oxidized and decomposed. Furthermore, SCCE was able to obtain higher extraction yield with the enhancement of ultrasound. In this dissertation, ultrasound enhanced sub-critical CO2 extraction (USCCE) technique was introduce to extract lutein from chlorella. The effects of factors involved in the USCCE process were systematically investigated and the optimal extraction condition was determined. The performance of USCCE was compared with soxhlet extraction (SE), sub-critical water extraction (SWE), SCE, SCCE with and without pretreatment in terms of process conditions and extraction yield. Moreover, experimental results were confirmed by the kinetic model built by sonochemistry technology team and mechanism of ultrasound enhancement in sub-critical CO2 extraction was discussed by presenting the ultrasound Blake threshold theory, cavitation experiment and scanning electron microscope (SEM) experiment results.Many factors involved could affect the process of USCCE, such as pretreatment, extraction temperature, extraction pressure, CO2 flow rate, cosolvent amount, extraction time and ultrasound power. Extraction pressure, which had a significant influence on the lutein extraction yield, was the most effective factor. The significance of the four major factors affecting lutein extraction yield was in the following order: extraction pressure>ultrasound power>cosolvent amount>extraction temperature. The optimal extraction condition was extraction temperature of 27℃, extraction pressure of 21MPa, cosolvent amount of 1.5mL/g ethanol, ultrasound power of 1000W. The lutein extraction yield obtained under this condition was 124.01mg/100g chlorella.The results of comparison of six methods indicated that the USCCE was able to increase the lutein extraction yield significantly at lower extraction temperature and pressure with the introduction of ultrasound and cell wall breaking pretreatment. Besides, USCCE was more favorable for its lower cost compared with SCE and advantage of no use of toxic reagent. SWE was unavailable on extraction of lutein for its overheated condition. Lutein and the other two unknown carotenoids were detected in the extration by all of the methods except for SWE.Based on the extraction kinetic model built by sonochemistry technology team, the kinetic models of USCCE and SCCE of lutein from chlorella were set as E = 130 .64×(1?e?0. 6599t) and E = 101 .82×(1?e?0. 5683t) respectively. The simulated results of the two models fit the experimental results well. Ultrasound was able to enhance the extraction process of SCCE according to the differences of parameters in the kinetic models, such as E∞and k.The ultrasound cavitation in sub-critical CO2 was studied by calculating Blake threshold values and tin foil material experiment. SEM was combined with the cavitation results to discuss the ultrasound enhancement mechanism in SCCE. The Blake threshold intensities for sub-critical CO2 in various temperatures and pressures were listed. Moreover, the results showed that the phenomena of tin foil material after ultrasound treatment under the certain condition displayed similar trend with the calculation. Relatively lower pressure and higher temperature facilitated the occurrence of cavitation. However, cavity was detected under the high-pressure condition that cavitation was suppose to be unavailable according to the calculation. The reason accounted for this was still unknown and further research was required. The surface of chlorella without any pretreatment was destructed to some extent with the effect of ultrasound, which facilitated the extraction of inner components. The surface structure of chlorella of which cell wall was broken by cellulose enzyme turned loose, exposing the inner part toward the ultrasound, decreasing the internal mass transfer resistance, which resulted in the highest extraction yield. The ultrasound enhancement mechanism of USCCE differed with the one of regular solvent. When temperature was 27℃, pressure was 21MPa, ultrasound intensity was 0.63×106 W/m2 in sub-critical CO2, besides the effect of shock wave and micro jet brought by transient cavitation, both the intense and high speed vibration of particles induced by the mechanical effect and the tremendous stress imposed by stable cavitation when resonance occurred had played important roles in ultrasound enhancement mechanism.