Preparation And Bioactivity Of Protein Hydrolysates And Polysaccharides From Camellia Oleifera Seed Cake

Camellia oleifera Abel is an unique oilseed tree crop which is widely distributed in central and south China. Camellia oil extracted from its seeds, with a reputation "orential olive oil", is rich of unsaturated fatty acid and various bioactive components. During the oil productive process, many byproducts, such as seed cake and soapstock, were generated and unually used as fertilizer and fuel, even as waste which brought some enviromental stress without any economic benefit. Thus, it had practical and exploiting significance to futher utilize camellia seed cake. Protein and polysaccharide, as biomacromolecules with various bioactivities and high nutrition value, are becoming more and more attractive. Camellia seed cake included abundant protein and polysaccharide. Thus, in this work, functionial property and antioxidant activity of camellia seed cake protein and its enzymatic hydrolysates have been investigated, and anticancer and antioxidant activity of camellia seed cake polysaccharide also have been evaluated. This work provided a theotical basis and method reference for futher utilization of protein and polysaccharide from camellia seed cake, and for the improvement of their economic benefit.1. The combined extraction of camellia seed cake protein (SCP) and its polysaccharide (COP-c) was optimazed by orthogonal design. The optimazed extraction conditions were as follow:pH of alkali solution was 10, ratio of material to liquid was 1:30, extraction time was 3 h, and extraction temperature was 60℃. Under these conditions, the yield of SCP and COP-c were 4.93±0.13% and 4.91±0.09%, respectively.2. Camellia seed cake protein (SCP) was separated into two faractions (≥10 kDa and < 10 kDa) by ultrafiltration. Fraction< 10 kDa had an excellent oil-holding capacity but less water-holding capacity, while fraction≥10 kDa had an opposite behavior. In addition, fraction< 10 kDa possessed a prominent protein solubility, but its foaming stability was poor. SCP had the same behavior as fraction≥10 kDa. Fraction≥10 kDa exhibited excellent emulsifying capability and foaming stability. The influence of pH and temperature on functional property was varied with protein molecular weight. Fraction< 10 kDa was strongly tolerant to temperature, and could maintain good emulsifying stability and protein solubility at 80℃. But its emulsifying and foaming stability was poor at an acidic environment. At 60℃, SCP and its ultrafiltration fractions had the best protein solubility and emulsifying capability.3. SCP was hydrolyzed into protein hydrolysates (SCPH) by five commercial proteases (Flavorzyme, Trypsin, Neutrase, Papain, Alcalase). Amino acid composition, molecular weight distribution, antioxidant activity and functional property of SCPH were investigated. Enzymatic hydrolysis improved protein solubility significantly but impaired the foaming and emulsifying property. Hydrolysate generated by alcalase had the highest hydrolysis degree (DH) and antioxidant activity, and displayed excellent protein solubility over wide range of pH, while hydrolysate prepared by flavorzyme showed better copper chelating capacity and emulsifying stability with low molecular weight distribution. Trypsin-treated SCPH showed better foaming property than original protein. The results indicated that enzyme type greatly influenced the molecular weight, functional property and antioxidant activity of SCPH.4. Antioxidant activities and functional properties of SCPH prepared using alcalase with different DH values (10%,20%,30% and 40%) were investigated. As the hydrolysis time was extended, the DPPH radical scavenging activity increased and finally reached a plateau, the copper chelating capacity decreased, and the superoxide radical scavenging and iron chelating activities increased initially, then subsequently slowed. The solubility, foaming properties, and emulsification properties of SCPH were affected by pH and DH. SCPH with DH≤30% showed a good functional properaty, but further hysrolysis (DH 40%) impaired protein functional property extremely. SCPH at DH 20% and 30% values exhibited good functional property and antioxidant activity. So, it was also found that electing appropriate protease type and controlling the DH could be enhanced or reduced functional property according to actual applications.5. Four membrane ultrafiltration fractions (COP-1, COP-2, COP-3 and COP-4) were obtained from crude camellia seed cake polysaccharide (COP-c) and homogenized through DEAE cellulose-52 and sephedex G-100 columns. The molecular weights of COP-1, COP-2, COP-3 and COP-4 were around 7.9,36,83, and 225 kDa, respectively. All samples showed similar FT-ER. spectrum. COP-2 and COP-3 showed highest radical scavenging activity. COP-1 exhibited the strongest metal chelating capability. While COP-4 showed the lowest free radical scavenging activity and metal chelating capacity. The results indicated that the bioactivity of polysaccharide was influced by molecular weight. Medium molecular weight camellia seed cake polysaccharide possessed the highest antioxidant activity. Low molecular weight polysaccharide had the best metal chelating capability. High molecular weight polysaccharide had the weast antioxidant and metal chelating activity. All samples showed no significant growth inhibitory effect of two cancer cells both HeLa and HepG2 cells.