Angelia, Cindy
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Sanjaya, Astia
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Aida, Aida
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Tanudjaja, Ellen
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Victor, Hans
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Cahyani, Antari Daru
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Tan, Tjie Jan
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
,
Pinontoan, Reinhard
(Biology Department, Faculty of Science and Technology, Universitas Pelita Harapan)
The use of GRAS microorganisms isolated from fermented foods during amylase production using an economical food-waste medium provides more opportunities to produce amylase with a wider range of applications. Hence, this study aimed to isolate a good amylase-producing fungi from the traditional Indon...
The use of GRAS microorganisms isolated from fermented foods during amylase production using an economical food-waste medium provides more opportunities to produce amylase with a wider range of applications. Hence, this study aimed to isolate a good amylase-producing fungi from the traditional Indonesian fermented cassava, gatot, and to identify the amylase-producing capability of the isolate in a potato peel waste (PPW) medium. Black-colored fungi isolated from gatot was morphologically identified and the amylase produced was characterized using SDS-PAGE and Native PAGE. The isolate was then grown on PPW medium, and the amylase produced was further characterized. Morphological identification and enzyme characterization revealed that the Aspergillus niger aggregate F isolated from gatot secreted an active extracellular ${\alpha}$-amylase with an optimum pH of 5-6. In conclusion, Aspergillus niger aggregate F isolated from gatot can be used to produce ${\alpha}$-amylase using PPW as a medium.
The use of GRAS microorganisms isolated from fermented foods during amylase production using an economical food-waste medium provides more opportunities to produce amylase with a wider range of applications. Hence, this study aimed to isolate a good amylase-producing fungi from the traditional Indonesian fermented cassava, gatot, and to identify the amylase-producing capability of the isolate in a potato peel waste (PPW) medium. Black-colored fungi isolated from gatot was morphologically identified and the amylase produced was characterized using SDS-PAGE and Native PAGE. The isolate was then grown on PPW medium, and the amylase produced was further characterized. Morphological identification and enzyme characterization revealed that the Aspergillus niger aggregate F isolated from gatot secreted an active extracellular ${\alpha}$-amylase with an optimum pH of 5-6. In conclusion, Aspergillus niger aggregate F isolated from gatot can be used to produce ${\alpha}$-amylase using PPW as a medium.
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문제 정의
Isolation of microbes from fermented food like gatot offers a higher chance of finding generally recognized as safe (GRAS) microbes which potentially produce amylase with a wider range of application. Therefore, this study aimed to determine and characterize the ability of Aspergillus niger aggregate isolated from gatot to produce amylase using PPW as its culture medium.
Isolation of gatot’s microorganisms has successfully selected a number of different multicellular fungi, including the one whose criteria matched the target candidate, Aspergillus niger. This fungus was denoted as isolate F and further used for morphological identification and amylase characterization in this study.
제안 방법
2. Identification of amylase produced by Aspergillus niger aggregate F grown in PDB medium using (A) SDS-PAGE and (B) Native-PAGE and activity staining analysis. (A) Lane 1: HyperPage Prestained Protein Marker (Bioline); Lane 2: crude extract of Aspergillus niger aggregate F; Lane 3: partially purified and concentrated crude extract of Aspergillus niger aggregate F; (B) Lane 1: crude extract of Aspergillus niger aggregate F; Lane 2: a single band with an active amylase activity.
Characterization of amylase activity produced by Aspergillus niger aggregate F was done by determining its optimum pH and substrate concentration. In this study, DNS method was used to measure the level of glucose released when the crude extract of Aspergillus niger aggregate F was mixed with starch solution and incubated for 15 min. Based on prior experiment (data not shown), amylase produced by Aspergillus niger aggregate F had an optimum temperature of 60℃, thus substrate incubation for DNS analysis was done in that temperature.
대상 데이터
In this study, the candidate of amylase-producing fungi were isolated from Indonesian traditional fermented food, gatot (Fig. 1A). Gatot is a product of spontaneous cassava fermentation by indigenous microorganisms, primarily fungi and lactic acid bacteria.
이론/모형
As the main goal of this study was to isolate a potential amylase-producing fungi from gatot that can be applied in SSF with PPW as its sole medium, the ability of the isolate to produce extracellular amylase was first needed to be confirmed. Both crude and partially purified extracts of Aspergillus niger aggregate F grown in PDB were collected and their protein content were measured using Biuret assay. The crude extract contained 0.
As much as 500 μl of that solution was then mixed with 20 μl of crude extract of Aspergillus niger aggregate F and incubated at 60℃ for 15 min. The amount of glucose resulted from the hydrolysis of starch by amylase contained in the extract were measured using DNS method, as decribed by Miller [17].
The crude and partially purified extracts of Aspergillus niger aggregate F were analyzed by SDS-PAGE using 4% stacking gel and 10% running gel according to the method of Laemmli [17]. Electrophoresis was run at 50 V for 60 min, followed by 100 V for another 60 min.
Optimum substrate concentration of the amylase was determined by mixing 500 μl of starch in various concentration (4, 6, 8, 10, 12, 14, 18, 22, and 30 mg/ml) with 20 μl of crude extract of Aspergillus niger aggregate F and incubated them at 60℃ for 15 min. The glucose level resulted from the hydrolysis of starch by amylase contained in the extract were measured using DNS method, as decribed by Miller [17].
성능/효과
As the main goal of this study was to isolate a potential amylase-producing fungi from gatot that can be applied in SSF with PPW as its sole medium, the ability of the isolate to produce extracellular amylase was first needed to be confirmed. Both crude and partially purified extracts of Aspergillus niger aggregate F grown in PDB were collected and their protein content were measured using Biuret assay.
Based on the results of this study, it was known that Aspergillus niger aggregate F isolated from gatot secreted extracellular amylase when grown both in commercial potato-based medium and PPW. In the spontaneous fermentation of gatot, the high starch content of the cassava (64−84% of dry weight) supports the growth of Aspergillus niger, a fungus most commonly found in the environment including food and feed ingredients [32, 33].
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