IMMOBILIZATION OF ?-AMYLASE ON POLYPIROLLE MEMBRANE USING ELECTROPOLIMERIZATION

• Main Author: SUKMAWARDANI NIM:30512016, YULIA
• Format: Dissertations
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/31817
• Institution: Institut Teknologi Bandung

Starch is the largest component in human foodstock, especially in Indonesia, and is a raw material for various industrial processes, especially food and beverage manufacturing industries. Currently the process of hydrolyzing starch polymers into simpler molecules has become one of the important stages in the industrial world. Hydrolysis of starch is done in two ways by using acid or starch breaking enzyme for example from the amylase group. The use of amylases is more desirable because it is environmentally friendly, more specific and not causing deviation at the end product. Starch hydrolysis process using amylase to become oligosacharide can reach degree of starch hydrolysis to 42-97% depending on the type of substrate and incubation time. Along with the rapidly growing science, analytical techniques in chemistry are also growing, including the technique of starch hydrolysis by using enzymes. An enzyme's weakness, such as its low probability, expected to be overcome by new immobilizing technique on a polymer membrane. <br /> <br /> Development and novelty in this research are combination of two important aspect, which are optimum condition of PPy film preparation electrochemically as media of ?-amylase captured and optimum condition of amobilization ?-amyase in PPy film through physicohemistry antaraction. Furthermore, in the study of electrometry, we studied the use of steel gauze as the electrodes for PPy electrodeposition and amobilization medium on PPy matrix. The phenomenon of ?-amylase immobilization on PPy matrix is confirmed by electrometry study, i.e by studying cyclic voltammogram profiles PPy, ?-amylase, and PPy /?-amylase (ie, ?-amylase immobilized in PPy film). These two aspects then used as the basis for preparation PPy /?-amylase membrane, which in this study was applied in the process of enzymatic starch hydrolysis. <br /> <br /> To streamline the performance of ?- amylase, this enzyme needs to be immobilized on the surface of PPy film which is conductive by using electropolimerization method. In this study, amobilization of ?- amylase was performed in four stages of work. First, preparation of PPy film by electrodeposition of pyrrole on steel gauze electrode. Second, the growth of polymer in PPy membrane and PPy /?- amylase membrane analyzed by cyclic voltammogram. Furthermore, in the third stage, the manufacturing of PPy/?-amylase membrane on the surface of the steel gauze is <br /> <br /> carried out by electrolysis method. The last step is testing the performance of membrane bioreactor PPy ?- amylase. <br /> <br /> In this research, synthesis of five types of PPy/?-amylase membrane with steel gauze support material had been done. Five types of PPy/?-amylase electrodeposed on the surface of the steel gauze using 0.1 M Py solution and five variations of ?-amylase concentration (0.1535, 0.307, 0.614, 0.921, 1.228 ?g/mL) in phosphate buffer 50 mM pH 6.5 at room temperature with optimum electrolysis conditions (potential 5V, electrolysis time 5 minutes and distance 1.5 cm between two electrodes). <br /> <br /> The experimental results confirm two performance roles of PPy/?-amylase membrane, ie as selective barrier membrane to starch. The rejection range of PPy /?-amylase membranes 1 to 5 on successive starch is 12,45,70-71,30%. It was also found that the membrane PPy/? amylase 5 was the best performing membrane. The application of the membrane PPy/?-amylase 5 to the sample after 10 successive recurrent uses showed that the more usage, the less membrane rejection of starch, the rejection at first use was 71,05%, while in the tenth usage of 21,70%. <br /> <br /> In the second part of the present study, each of the above optimum conditions is utilized for the synthesis and application of the PPy/?--amylase membrane on the steel gauze surface for the determination of starch. In this case, the performance of the immobilized enzyme on the PPy matrix is performed by comparing the activity of the immobilized enzyme and the activity of the free enzyme. Determination of ?-amylase activity was done by spectrophotometric UV-visible. Activated ?- amylase activity in PPy/?-amylase 5 is 1426 U/L, while free enzyme activity is 1299 U/L. <br /> <br /> The amyloidization of ?-amylase in PPy matrix is a physical-chemistry between the carboxylate groups in ?-amylase and NH groups in PPy, as confirmed by shifting poultry and the formation of new peaks in the FTIR spectrum. SEM image results with 600 to 3000x magnification indicates that the entire surface of the steel gauze is covered by a PPy layer. The result of SEM-EDS analysis to polymer layer obtained C 24,51% and N 14,08% as PPy constituent elements. The white layer attached to and bound to the polymer film is identified as ?- amylase and comprises C 44.60% elements; O 19.78%. <br /> <br /> The membrane synthesis method of PPy / ?- amylase by electropolimerization provides the ease of immobilization of ?- amylase in PPy matrix with steel gauze support material as electrode prepared by cyclic voltammetric method. Thus, the overall research data is expected to contribute to the development of specific analytical chemistry and related fields.