SKRINING BAKTERI TERMOFILIK POTENSIAL AMILOLITIK DARI SUMBER AIR PANAS WAY BELERANG KALIANDA LAMPUNG SELATAN

Thermophilic bacteria that produced amylase and protease have been isolated from Way Belerang hot spring, Kalianda, South Lampung. This research aims to screen and identify thermophilic bacteria that have the potential to produce thermostable amylase and protease enzymes.The research procedures included sampling, isolation of enzyme-producing thermophilic bacteria, a series of phenotypic and biochemical tests, and molecular identification by 16s rRNA. This study used 2 treatments, namely incubation temperature 37 and 50 ºC with 3 repetitions. The results showed that the optimum temperature for growth of thermophilic bacterial isolates and thermophilic bacterial isolates producing amylase enzymes was 50ºC. The bacteria isolate that had the best amylolytic enzyme activity was Isolate A.WB.50.1 with a diameter of the inhibitory zone was 15.44 mm. Isolate A.WB.50.1 has been identified by the species Pseudomonas stutzeri.

Characterization of Thermostable Amylase From Halophilic Lactobacillus Plantarum TS1

Amylase is an important enzyme use extensively in various industrial processes. It is mainly involved in the catalysis of starch that requires harsh conditions; therefore, it is required to isolate amylases with unique properties that makes it more applicable. Extremophiles are the major resource of such enzymes; therefore, amylase positive strains were isolated from the saline soil where the temperature is also exceptionally high. Five amylase positive strains were isolated from the Karak salt range, Kohat Pakistan and were identified by phylogenetic analysis. DNS based assay was employed to compare the activities of different amylases obtained from five strains while using the starch as a substrate. The amylase obtained from Lactobacillus plantarum TS1 was found to be more efficient, which was purified and characterized. The SDS-PAGE of purified amylase showed a single band with an estimated size of 10 kDa. The kinetic parameters were measured at two temperatures i.e. at 37 °C and 50 °C. The Kcat as well as the Kcat/KM were found to be high when temperature increased from 37 °C to 50 °C. Amylase was active at wide range of temperature as well as pH and work efficiently in the presence of salts and various organic solvents.

Screening and Characterization of Thermostable Amylase-Producing Bacteria Isolated from Soil Samples of Afdera, Afar Region, and Molecular Detection of Amylase-Coding Gene

Studying thermostable amylase-producing bacteria in extreme environments has a crucial role to overcome different industrial challenges. Afar Region is one of the hottest and salty areas, making it the home of extremophiles. This study aimed at screening and characterizing amylase-producing bacteria isolated from soil samples of Afdera, Afar Region, and detection of their amylase-coding genes. Thus, a total of 49 bacterial isolates were obtained from the collected soil samples. Out of these, three isolates (M2, M8, and M13) were selected on the basis of diameter of the average clear zone formation and time taken to decolorize iodine solution. Based on their morphological and biochemical characteristics, the isolates were identified as genus Bacillus. PCR amplification and detection of the amylase-coding gene confirmed the presence of the amylase gene in the three bacterial isolates. Optimum amylase production time for these isolates was 48 hrs (M13 and M8) and 72 hrs (M2) corresponding to the amylase activity of 0.67 U/mL for M13, 0.74 U/mL for M8, and 0.73 U/mL for M2 with an optimum temperature of 55°C. Studies on the effect of temperature revealed that the crude enzyme had a maximum activity and stability at 75°C, 70°C, and 65°C for isolates M13, M8, and M2, respectively. Additionally, amylase produced from all isolates retained more than 66.41% of their original activity after incubating them at a temperature range from 55 to 80°C for 50 min. Optimum pH for the activity of all crude amylases was in the range from 5 to 9 with a peak activity at pH 8. Their activity decreased significantly by the presence of Zn+2 and Mg2+; however, their activity increased by the presence of Ca+2. Moreover, the three crude amylases were stable (0–3 M) with NaCl concentration. Amylases of this finding with thermophilic and halophilic characteristics offer a wide range of applications in food, brewing, textile, starch, paper, and deterrent industries. Thus, identification of these Bacillus isolates at a molecular level and purification as well as detailed characterization of the types of amylases are recommended for effective utilization in different industries.

Aktivitas amilase bakteri amilolitik asal larva black soldier fly (Hermetia illucens)

Amylase is an enzyme that has been widely used as a biocatalyst in foodand bioethanol industries. The availability of thermostable amylase will further expand the market and extend the shelf life of this enzyme. Amylase is produced by amylolytic bacteria using media with high-costnitrogen sources, such as pepton. Black soldier fly (BSF) is a potential source of amylolitic bacteria since its ability to degrade organic matters rapidly. This research aimedtoexploreamylolitic bacteria from the larvae of BSF with highest amylase activity that can be produced using low-cost media. The screening ofamylase activity was conducted by culturing the bacteria on starch containing media.Bacteria with the highest amylase activity were cultured in liquid media with twodifferentnitrogensources (urea and nitrate). Determinations of the optimum pH and temperature for this enzyme activity were carried out in the pH range 4to 7 and temperature 35to 65 ºC. Three amylase-producing isolates were obtained in this study. M1 isolate which has the highest activity was characterized based oncatalase activity and Gram staining. The results showed that the M1 isolate mightbelong togenus Proteussp. At the optimum condition (45ºC and pH 7), amylase activityin nitrate mediawas0.791U/mL, which was about 18-folds higher than that in ureamedia (0,041U/mL). Thus, amylase isolated from BSF larvae can be classified as a mesophilic enzyme and has the potential to be developed commercially at lower production costs.[Keywords:crude extract enzyme,Proteus sp.,thermostable] AbstrakAmilase merupakan salah satu enzim yangtelah digunakan secara luas sebagai biokatalis dalam industri pangan dan bioetanol.Ketersediaan amilase termostabil akan semakin memperluas pasar dan memperpanjang daya simpan enzim ini. Selama ini, produksi amilase dilakukan dengan memanfaatkan bakteri amilolitik menggunakan media dengan sumber nitrogen yang mahal, misalnya pepton. Black soldier fly (BSF) merupakan sumber bakteri amilolitik yang potential karena BSF memiliki kemampuan mendegradasi bahan organik dengan cepat. Penelitian ini bertujuan untuk mengeksplorasi bakteri amilolitik dengan kemampuan amilase tinggi yang dapat diproduksi menggunakan media yanglebihmurah.Skrining bakteri penghasil amilase dilakukan dengan menumbuhkan bakteri pada media yang mengandung pati. Bakteri dengan aktivitas amilase tertinggi dikulturkan dalam media cair dengan dua sumber nitrogenyang berbeda, yaitu urea dan nitrat. PenentuanpH dan suhu optimum aktivitas enzim ini dilakukan pada rentang pH 4sampai 7 dan suhu 35sampai 65 ºC.Tiga isolat penghasil amilase diperoleh dalam penelitian ini. Isolat M1 yang memiliki aktivitas tertinggi dikarakterisasi berdasarkan uji katalasedan uji pewarnaan Gram. Hasilnya menunjukkan bahwa isolat M1 termasukgenus Proteus sp. Pada kondisi optimum (suhu 45oC dan pH 7), aktivitas amilase pada media nitrat adalah 0,791 U/mL, lebih kurang 18 kali lebih tinggi dibanding aktivitas pada media urea (0,041 U/mL). Dengan demikian, amilase yang dihasilkan oleh bakteri asallarva BSF merupakan enzim mesofilik dan berpotensi untuk dikembangkan secara komersial dengan biaya produksi yang lebih murah.[Kata kunci: enzim ekstrak kasar, Proteussp.,termostabil]