Özet
Dayanak
Amilazlar, nişastayı parçalayan ve meyve sularının arıtılmasına yardımcı olan enzimlerdir.
Amaçlar
Bu çalışma, toprak örneklerinden izole edilen amilolitik Streptomyces türlerinin amilaz üretimi ve meyve suyu berraklaştırma potansiyelinin taranmasına odaklanmıştır. Seçilen organizmalar, hücrelerin indirgen şeker içeriği ölçülerek test edilen amilaz üretmiştir.
Yöntemler
Amilaz sentezini optimize etmek için pH, karbon ve azot kaynağı ile çalkalama ve inkübasyon sürelerinin etkileri değerlendirilmiştir.
Bulgular
Toplam 22 tür izole edildi ve bunlardan beş tanesi (FE4, ELI1, FL2, MS2 ve MS5) hafif asidik ila hafif alkali pH aralığında yüksek amilaz üretim kabiliyeti gösterdi. Manyok kabukları, Streptomyces spp. A4 (0,834), ELI1 (0,910) ve FE4 (0,814 U/mL) türlerinde optimal amilaz üretimini destekledi. Üre azot kaynağı olarak kullanıldığında, 100-150 rpm çalkalama hızında ve fermantasyonun dördüncü gününde zirveye ulaştığında, ELI1 ile 0,930 U/mL’lik maksimum verim gözlemlenmiştir. 16S rRNA gen dizilemesi kullanılarak S. griseoflavus ELI_1 olarak tanımlanmış ve GenBank’a OQ930232 erişim numarasıyla kaydedilmiştir. Bu suş tarafından üretilen amilaz kısmen saflaştırılmış ve spesifik aktivitesi 1,50’den 4,56 U/mL’ye önemli ölçüde artmıştır. Portakal suyunu berraklaştırma kabiliyeti test edilmiş ve amilaz tedavisi sonrasında bulanıklık %16 oranında önemli ölçüde azalmıştır (p <0,05).
Sonuç
Böylece, toprak örneğinden izole edilen amilolitik tür S. griseoflavus ELI_1, portakal suyunu en iyi şekilde berraklaştırabildi.
Anahtar Kelimeler:
Kaynaklar
1
Abd-Elhalem, B. T., El-Sawy, M., Gamal, R. F., & Abou-Taleb, K. A. (2015). Production of amylases from Bacillus amyloliquefaciens under submerged fermentation using some agro-industrial by-products. Annals of Agricultural Sciences, 60 (2), 193–202. https://doi.org/10.1016/j.aoas.2015.06.001
2
Adebare, B. S., Johnson, A. A., Olanrewaju, T. M., Titilope, A. O., Femi, B. P., Bankole, S. A., Joseph, I. O., & Joshua, A. A. (2021). Production and optimization of alpha amylase from Aspergillus niger using TME 419 cassava peel as substrate. African Journal of Biological Sciences, 3 (4), 50–59. https://doi.org/10.33472/AFJBS.3.4.2021.50-59
3
Adrio, J. L., & Demain, A. L. (2014). Microbial enzymes: Tools for biotechnological processes. Biomolecules, 4 (1), 117–139. https://doi.org/10.3390/biom4010117
4
Ahmed, K., Munawar, S., & Khan, M. A. (2015). Cultural conditions for maximum alpha-amylase production by Penicillium notatum IBGE 03 using shaken flask technique of submerged fermentation. Pure Applied Biology, 4 (3), 306–312. http://dx.doi.org/10.19045/bspab.2015.43005
5
Al-Dhabi, N. A., Esmail, G. A., Ghilan, A. M., Arasu, M. V., Duraipandiyan, V., & Ponmurugan, K. (2020). Isolation and purification of starch hydrolysing amylase from Streptomyces sp. Al-Dhabi-46 obtained from the Jazan region of Saudi Arabi with industrial applications. Journal of King Saud University – Science, 32 , 1226–1232. https://doi.org/10.1016/j.jksus.2019.11.018
6
Ali, I., Sultan, S., Mahmood, R. T., Tariq, M., Shamim, Z., Mushtaq, A., & Asiri, M. (2023). Production and characterisation of α-amylase from indigenously isolated Streptomyces sp. Bioresources, 18 (1), 6–18. https://doi.org/10.15376/biores.18.1.6-18
7
Arifeen, S., Jamil, J., Sarwar, A., Ullah, N., Nelofer, R., Aziz, T., Alharbi, M., Alasmari, A. F., Alshammari, A., & Albekairi, T. H. (2024). Biosynthesis and optimization of amylase from Bacillus sp. isolated from soil samples using agro-industrial waste as substrate. Applied Ecology and Environmental Research, 22 (4), 2927–2940. http://dx.doi.org/10.15666/aeer/2204_29272940
8
Bamigboye, C. O., Okonji, R. E., Oluremi, I. O., & James, V. (2022). Stain removing, juice-clarifying, and starch liquefying potentials of amylase from Pleurotus tuberregium in submerged fermentation system. Journal of Genetic Engineering and Biotechnology, 20 , Article 23. https://doi.org/10.1186/s43141-022-00298-4
9
Barman, D., & Dkhar, M. S. (2024). Purification and characterization of moderately thermostable raw-starch digesting α-amylase from endophytic Streptomyces mobaraensis DB13 associated with Costus speciosus. The Journal of General and Applied Microbiology, 69 (6), 293–300. https://doi.org/10.2323/jgam.2023.08.001
10
Baysal, Z., Uyar, F., & Aytekin, C. (2003). Solid state fermentation for production of α-amylase by a thermotolerant Bacillus subtilis from hot spring water. Process Biochemistry, 38 , 1665–1668. https://doi.org/10.1016/S0032-9592(02)00150-4
11
Casas, S. D., Orozco, J. L., Cardenas, A. J., Perez, H. L. R., & Brizuelas, L. G. (2025). Influence of the use of alpha amylase for the splitting of starches on color, ash and reducing sugars. Afinidad. Journal of Chemical Engineering Theoretical and Applied Chemistry, 82 , 359–366. https://doi.org/10.55815/431129
12
Chowdary, A. R., Prakash, P. O., & Kishor, W. A. (2018). Optimization of amylase production by Bacillus cereus using solid state fermentation. Journal of Biotechnology Resource, 4 , 56–65.
13
El-Sayed, M. H., Gomaa, A. E., Atta, O. M., & Hassane, A. M. (2024). Characteristics and kinetics of thermophilic actinomycetes’ amylase production on agro-wastes and its application for ethanol fermentation. World Journal of Microbiology and Biotechnology, 40 , Article 255. https://doi.org/10.1007/s11274-024-04009-8
14
Fahmy, N. M. (2022). Characterization of amylase-producing Streptomyces sp. NAA-28 isolated from mangrove sediment, Red Sea, Egypt. Egyptian Journal of Aquatic Biology and Fisheries, 26 (4), 1053–1065.
15
Fasiku, S. A., Bello, M. A., & Odeniyi, O. A. (2023). Production of xylanase by Aspergillus niger GIO and Bacillus megaterium through solid-state fermentation. Access Microbiology, 5 , 000506.v5. https://doi.org/10.1099/acmi.0.000506.v5
16
Fasiku, S. A., & Wakil, S. M. (2022). Screening of factors responsible for conversion of maize straw into bioethanol. Journal of Microbiology, Biotechnology and Food Sciences, 12 (2), e5901. https://doi.org/10.55251/jmbfs.5901
17
Fasiku, S. A., Ogunsola, O. F., Fakunle, A., & Olanbiwoninu, A. A. (2020). Isolation of bacteria with potential of producing extracellular enzymes (amylase, cellulase and protease) from soil samples. Journal of Advances in Microbiology, 20 (3), 21–26. https://doi.org/10.9734/jamb/2020/v20i330224
18
Gómez-Villegas, P., Viagra, J., Rumero, L., Gotor, C., Raposo, S., Gonçalves, B., & Léon, R. (2021). Biochemical characterization of the amylase activity from the new haloarchaeal strain Haloarcula sp. HS isolated in the Odiel Marshlands. Biology, 10 (4), 337. https://doi.org/10.3390/biology10040337
19
Gusakov, A. V., Kondratyeva, E. G., & Sinitsyn, A. P. (2011). Comparison of two methods for assaying reducing sugars in the determination of carbohydrase activities. International Journal of Analytical Chemistry, 2011 , 283658. https://doi.org/10.1155/2011/283658
20
Haque, A., Marwaha, S., Deb, S. K., Nigam, S., Arora, A., Hooda, K. S., Soujanya, P. L., Aggarwal, S. K., Lall, B., Kumar, M., Islam, S., Panwar, M., Kumar, P., & Agrawal, R. C. (2022). Deep learning based approach for the identification of diseases in maize crop. Scientific Reports, 12 (1), 1–14. https://doi.org/10.1038/s41598-022-10140-z
21
Hossain, K. M., Khan, U., Rahman, S. M. M., & Khan, M. S. (2024). Potential antimicrobial and fruit juice clarification activity of amylase enzyme from Bacillus strains. Biotechnology Reports, 44 , e00861. https://doi.org/10.1016/j.btre.2024.e00861
22
Ibrahim, D., Weloosamy, H., & Lim, S. (2015). Effect of agitation speed on the morphology of Aspergillus niger HFD5A-1 hyphae and its pectinase production in submerged fermentation. World Journal of Biological Chemistry, 6 (3), 265. https://doi.org/10.4331/wjbc.v6.i3.265
23
Illanes, A., Cauerhff, A., Wilson, L., & Castro, G. R. (2012). Recent trends in biocatalysis engineering. Bioresource Technology, 115 , 48–57. https://doi.org/10.1016/j.biortech.2011.12.050
24
Iram, N., Shakir, H. A., Irfan, M., Khan, M., Ali, S., Answer, A., Saeed, S., & Qazi, J. I. (2020). Statistical optimization of amylase production using grape fruit peels in submerged fermentation. Acta Scientiarum Technology, 43 , e50538. https://doi.org/10.4025/actascitechnol.v43i1.50538
25
Kahraman, O., Lee, H., Zhang, W., & Feng, H. (2017). Manothermosonication (MTS) treatment of apple–carrot juice blend for inactivation of Escherichia coli O157:H7. Ultrasonic Sonochemistry, 38 , 820–828. https://doi.org/10.1016/j.ultsonch.2016.11.024
26
Kareem, S. O., & Adebowale, A. A. (2007). Clarification of orange juice by crude fugal pectinase from citrus peel. Nigeria Food Journal, 24 , 130–137. https://doi.org/10.4314/nifoj.v25i1.33661
27
Kavitha, A., & Vijayalkshmi, M. (2010). Production of amylases by Streptomyces tendea TK-VL-333. International Journal of Current Resources, 10 , 110–114.
28
Khan, F. K. (2025). Recent advances in microbial enzymes applications for sustainable textile processing and waste management. Sci, 7 , Article 46. https://doi.org/10.3390/sci7020046
29
Kharel, M., Shepherd, M. D., Nybo, S. E., Smith, M. L., & Bosserman, M. A. (2010). Isolation of Streptomyces species from soil. Current Protocols in Microbiology, 19 (1), 4–5. https://doi.org/10.1002/9780471729259.mc10e04s19
30
Kizhakedathil, M. P. J., & Subathra, D. C. (2021). Acid stable α-amylase from Pseudomonas balearica VITPS19 – Production, purification and characterization. Biotechnology Reports, 30 , e00603.
31
Krishna, N., Muvva, V., Munaganti, R. K., & Bindu, H. (2015). Studies on optimization of amylase production by Streptomyces cheonanensis VUK-A isolated from mangroove habitats. Journal of Advances in Biology and Biotechnology, 3 (4), 165–172. https://doi.org/10.9734/JABB/2015/18025
32
Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura, K. (2018). MEGA X: Molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 35 , 1547–1549. https://doi.org/10.1093/molbev/msy096
33
Li, S., Yang, X., Yang, S., Zhu, M., & Wang, X. (2012). Technology prospecting on enzymes: Application, marketing and engineering. Computational and Structural Biotechnology Journal, 2 (3), 10–12. https://doi.org/10.5936/csbj.201209017
34
Liu, L., Yang, H., & Shin, H. D. (2013). How to achieve high-level expression of microbial enzymes: Strategies and perspectives. Bioengineering, 4 (4), 212–223. https://doi.org/10.4161/bioe.24761
35
Liviu, G., Olivian, C., & Cristian, H. (2022). Fruit juices clarification with α-amylases. Annals of the University of Craiova, Biology, Horticulture, Food Products Processing Technology, Environmental Engineering, 27 (63), 173–176.
36
Lowry, O. H., Rosebrough, N. J., Farr, A. L., & Randall, R. J. (1951). Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry, 193 , 265–275.
37
Mokrani, S., & Nabti, E. H. (2024). Recent status in production, biotechnological applications, commercial aspects, and future prospects of microbial enzymes: A comprehensive review. International Journal of Agricultural Science and Food Technology, 10 (1), 006–020. https://dx.doi.org/10.17352/2455-815X.000202
38
Niyomukiza, S., Owino, W., Mathara, J. M., & Maina, N. (2023). Isolation, purification and biochemical characterization of alkaline amylase from Bacillus subtilis strain W3SFR5 isolated from kitchen wastes. Applied Food Biotechnology, 20 (1), 9–19. http://dx.doi.org/10.22037/afb.v10i1.39495
39
Olakusehin, V. O., & Oyedeji, O. (2021). Production of α-amylase from Aspergillus flavus S2-OY using solid substrate fermentation of potato ( Solanum tuberosum L.) peel. International Journal of Biological and Chemical Sciences, 15 (5), 1950–1967. https://dx.doi.org/10.4314/ijbcs.v15i5.21
40
Oliveira, R. L., Dias, J. L., da Silva, O. S., & Porto, T. S. (2018). Immobilization of pectinase from Aspergillus aculeatus in alginate beads and clarification of apple and umbu juices in a packed bed reactor. Food Bioproduction Process, 109 , 9–18. https://doi.org/10.1016/j.fbp.2018.02.005
41
Oussadi, M. I., & Kitouni, M. (2015). Statistical optimization of cultural conditions of an halophilic alpha-amylase production by halophilic Streptomyces sp. grown on orange waste powder. Biocatalysis and Agricultural Biotechnology, 4 (4), 685–693. http://dx.doi.org/10.1016/j.bcab.2015.08.011
42
Oyenado, O. A., & Omoruyi, I. M. (2024). Review of amylase production by microorganisms and their industrial application. Ife Journal of Science, 26 (2), 537–553. https://dx.doi.org/10.4314/ijs.v26i2.23
43
Patel, A. K., Dong, C. D., Chen, C. W., Pandey, A., & Singhania, R. R. (2023). Production, purification, and application of microbial enzymes. In Biotechnology of microbial enzymes (pp. 25–57). https://doi.org/10.1016/B978-0-443-19059-9.00019-0
44
Prabakaran, G., & Pugalvendhan, R. (2009). Production and immobilization of alpha-amylase by using Bacillus subtilis. Recents Research in Science and Technology, 1 (4), 189–194. https://doi.org/10.3923/pjbs.2007.2039.2047
45
Rathore, D. S., & Singh, S. P. (2021). Kinetics of growths and co-production of amylase and protease in novel marine actinomycete, Streptomyces lopnurensis KaM5. Folia Microbiologia, 66 , 303–316. https://doi.org/10.1007/s12223-020-00843-z
46
Roheen, T., Ramzan, R., Nadeem, M., Atif, F. A., Munir, M., & Qureshi, T. M. (2024). Synthesis and characterization of CMC/PAM-amy hydrogel and its efficacy in apple juice clarification. Processes, 12 , 2264. https://doi.org/10.3390/pr12102264
47
Saad, W. F., Othman, A. M., Abdel-Fatah, M., & Ahmad, M. S. (2021). Response surface methodology as an approach for optimization of α-amylase production by the new isolated thermotolerant Bacillus licheniformis WF67 strain in submerged fermentation. Biocatalysis and Agricultural Biotechnology, 32 , 101944. https://doi.org/10.1016/j.bcab.2021.101944
48
Saadoun, I., Alawawdeh, M., Jaradat, Z., Ababneh, Q., Al-joubori, B. M., & Elsheikh, E. A. E. (2023). Characterization of diesel-degrading, hydrolytic enzymes-producing Streptomyces spp. isolated from fuel-oil polluted soils. Arab Journal of Basic and Applied Sciences, 30 (1), 248–255. https://doi.org/10.1080/25765299.2023.2196110
49
Sahu, P. K., Singh, R., Shrivastava, M., Darjee, S., Mageshwaran, V., Phurailtpan, L., & Rohatgi, B. (2024). Microbial production of α-amylase from agro-waste: An approach towards biorefinery and bio-economy. Energy Nexus, 14 , 100293. https://doi.org/10.1016/j.nexus.2024.100293
50
Saravanan, A., Kumar, P. S., Dai-Viet, N. V., Jeevanantham, S., Karishma, S., & Yaashikaa, R. (2021). A review on catalytic-enzyme degradation of toxic environmental pollutants: Microbial enzymes. Journal of Hazardous Materials, 419 , 126451. https://doi.org/10.1016/j.jhazmat.2021.126451
51
Sharma, H. P., Patel, H., & Sugandha. (2017). Enzymatic added extraction and clarification of fruit juices – A review. Critical Reviews in Food Science and Nutrition, 57 (6), 1215–1227. http://dx.doi.org/10.1080/10408398.2014.977434
52
Shu, W., Ruckert-Reed, C., Gromyko, O., Tistechok, S., Kalinowski, J., Luzhetskyy, A., & Wittmann, C. (2025). Description of Streptomyces explomaris sp. nov., isolated from the coastal soil rhizosphere of Juniperus excelsa and reclassification of Streptomyces libani as a later heterotypic synonym of Streptomyces nigrescens. International Journal of Systematic and Evolutionary Microbiology, 75 , 006711. https://doi.org/10.1099/ijsem.0.006711
53
Shwe, K. K. W., & Win, S. S. (2019). Effect of amylolytic enzyme treatment on banana juice preparation, yield and clarification: A preliminary approach. International Journal of Science and Engineering Applications, 8 (2), 75–78.
54
Singh, R., Kumar, M., Mittal, A., & Mehta, P. K. (2016). Microbial enzymes: Industrial progress in 21st century. 3 Biotechnology, 6 , 174. https://doi.org/10.1007/s13205-016-0485-8
55
Singh, T. A., Jajoo, A., & Bhasin, S. (2020). Production and application of glucose isomerase from Streptomyces enissocaesilis and amylase from Streptomyces sp. for the synthesis of high fructose corn syrup. SN Applied Sciences, 2 (12), 1968. https://doi.org/10.1007/s42452-020-03757-0
56
Sondhi, S., Kaur, P. S., Sura, H., Juglani, M., & Sharma, D. (2021). Amylase based clarification of apple, orange and grape juice. International Journal of Contemporary Technology and Research, 3 (3), 187–190. https://doi.org/10.46860/cgcijctr.2021.06.31.187
57
Suthar, S., Joshi, D., Patel, H., Patel, D., & Kikani, B. A. (2024). Optimization and purification of a novel calcium-independent thermostable, α-amylase produced by Bacillus licheniformis UDS-5. World Journal of Microbiology and Biotechnology, 40 (12), Article 385. https://doi.org/10.1007/s11274-024-04188-4
58
Tamura, K., & Nei, M. (1993). Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Molecular Biology and Evolution, 10 , 512–526. https://doi.org/10.1093/oxfordjournals.molbev.a040023
59
Thulasisingh, A., Ananthakrishnan, K., Raja, A., & Kannaiyan, S. (2024). Bioprospecting of novel and industrially appropriate enzymes: A review. Water, Air and Soil Pollution, 235 , 12. https://doi.org/10.1007/s11270-023-06831-6
60
Vojnovic, S., Aleksic, I., Ilic-Tomic, T., Stevanovic, M., & Nikodinovic-Runic, J. (2024). Bacillus and Streptomyces spp. as hosts for production of industrially relevant enzymes. Applied Microbiology and Biotechnology, 108 , Article 185. https://doi.org/10.1007/s00253-023-12900-x
61
Wang, H., Yuan, J., Chen, L., Ban, Z., Zheng, Y., Jiang, Y., Jiang, Y., & Li, X. (2022). Effects of fruit storage temperature and time on cloud stability of not from concentrated apple juice. Foods, 11 , 2568. https://doi.org/10.3390/foods11172568
