Judul: Formation of Amyloid Fibrils from Bogor Nut [Vigna subterranea (L.) Verdc.] Protein Isolates, 7S Globulin, and 11S Globulin
Abstrak: One of the alternatives to enhance the benefits of Bogor nut which had examined in this dissertation is modifying nut protein to be functioned as a food ingredient. The globular structure of nut protein is modified into fibrillar structure by fibrillation. The fibrillation process of nut protein is carried out by giving special treatment in vitro. These treatments include giving temperature above denaturation temperature, low pH, and low ionic strength. This condition causes the peptide fragments resulting from protein hydrolysis will aggregate into length and forming semi flexible fibrils, hereinafter is referred to as amyloid fibrils. The changes of proteins conformation from globular to fibrillar will cause physical and chemical properties changes on the fibrils surface, thus providing different functions than its globular proteins. These structural changes resulted in an increase of the fibril solution viscosity, changes of flow behavior (rheology), and also surface properties of the fibrils. Thus, fibrillation can increase the protein functional characteristics to be developed as food ingredient according to the needs for processed food. The first stage of the research was preparing the beans and flour of Bogor nut. After 120 days of planting, fresh Bogor nut pods are harvested and then to be processed into Bogor nut flour. From 100 kg of fresh pods, it was obtained about 18.65 kg of Bogor nut flour, or from 100 kg of dry beans, 91.7 kg of Bogor nut flour was obtained. The protein content of nut flour is 21.66% (w/w) and the fat content is around 8.76%. The low-fat content is very beneficial to the fat extraction process, making it more economical. Thus, Bogor bean flour becomes a potential protein source to be isolated and fractionated. The second stage, through gradual pH adjustment starting from pH 6.4 to precipitate the 11S globulin, then followed by pH adjustment on the supernatant to pH 4.8 to precipitate the 7S globulin, this research succeeded in obtaining the protein isolate of Bogor nut, as well as the globulin fraction 11S and 7S which are purer as an ingredient for the formation of amyloid fibrils. In relation to the characteristics of protein sedimentation (the term S = Svedberg Unit), this research also studied the effect of globulin deposition at different centrifuge speeds, on 10,000 ×g and 19,000 ×g. The results showed that 10,000 ×g centrifuge speed was more economical and capable of harvesting isolates and globulin (PI-10, 7SG-10, 11SG-10) with higher protein yield and protein content compared to 19,000 ×g centrifuge speed (PI-19, 7SG -19, 11SG-19). The third stage, the six types of isolates and globulins that were extracted from the protein of Bogor Nut (PI-10, 7SG-10, 11SG-10, PI-19, 7SG-19, 11SG-19) has succeeded in forming amyloid fibril. Through various treatments such as the temperature of 65, 75, 85 oC, and heating duration of 6, 12, 18, 24, 30 hours, has obtained process conditions that produce a significant number of fibrils, which are on temperature 85 oC, 24 hours of heating duration for 1% concentration, 21 hours heating duration for 2% concentration, medium with pH 2, and medium agitation of 150 stokes per minute.
Keyword: amyloid fibrils, Bogor nut, isolation of 7S and 11S globulin, rheometerSDS PAGE-TEM-FTIR-CR spectroscopy assay, Gel filtration chromatography