جداسازی وغربال گری قارچ های هالوفیل اختیاری تولید کننده آنزیم های صنعتی از پارک های جنگلی تهران
محورهای موضوعی : میکروبیولوژی
محدثه لاری پور
1
*
,
انسیه کارگر فراغه
2
,
Monireh Movahedi
3
1 - دانشگاه آزاد اسلامی واحد تهران شمال
2 - کارشناس ارشد بیوتکنولوژی میکروبی، دانشکده علوم زیستی، دانشگاه آزاد اسلامی واحد تهران شمال
3 - Assistant Professor of Biochemistry , Faculty of Biological Science, Department of biochemistry, Islamic Azad University, Tehran North Branch, Iran.
کلید واژه: پکتیناز, قارچ, پارک جنگلی, هالوفیل,
چکیده مقاله :
چکیده سابقه وهدف: قارچ های ساپروفیت هالوفیل اختیاری به دلیل سهولت در کشت و تحمل شرایط سخت می توانند مقادیر زیادی آنزیم خارج سلولی را با مصرف صنعتی تولید کنند. در این مطالعه سعی شده است فعالیت آنزیمی قارچ های هالوفیل اختیاری از پارک های جنگلی تهران مورد سنجش قرار گیرد. مواد و روش‌ها: بنابراین نمونه‌برداری از خاک و هوای پارک جنگلی تهران، ساپروفیت‌ها با استفاده از تکنیک میکروسکوپی و کشت اسلاید شناسایی شدند. سپس قارچ های جدا شده از نظر فعالیت آنزیمی مورد بررسی کیفی قرار گرفتند و اکثر سویه ها آنزیم های پکتیناز، لیپاز، پروتئاز و آمیلاز تولید کردند. پنی سیلیوم هالوتولرانت و سویه های E01، E03، E014 اینورتاز تولید کردند. و تنها سویه E13 سلولاز تولید کرد. بنابراین فعالیت آنزیمی پکتیناز (از طریق بررسی آنزیم تولید شده در محیط با پکتین از پاگ) در سویه حاوی آنزیم. بهترین سویه آنزیمی تولید شده با روش PCR شناسایی و درخت فیلوژنی ترسیم می شود. يافته ها: بيشترين قارچ هاي جدا شده آسپرژيلوس، پني سيليوم، فوزاريوم، موكور و ريزوپوس بود. از بین قارچ‌های جدا شده و شناسایی شده، 19 جدایه برای تولید آنزیم‌های خارج سلولی در نظر گرفته شد و با استفاده از روش‌های مولکولی مانند Fusarium sublunatum، Aspergillus subramanianii، Penicillium hallotolerans، Mucor circinelloides و Purpureocillium lilacinum شناسایی شدند. سویه های Cladosporium sp.، Penicillium halotolerans بیشترین فعالیت را نشان دادند (با فعالیت های 0.71 و 0.79 U/mL). نتیجه گیری: Cladosporium sp و Penicillium halotolerans به دلیل کشت روی بسترهای ارزان قیمت و استخراج آسان آنزیمی از محیط کشت، می توانند برای تولید آنزیم صنعتی مورد استفاده قرار گیرند.
Abstract Aim and Background: Facultative halophilic saprophytic fungi, can produce high amounts of extracellular enzymes with industrial consumption, due to their ease of cultivation, and endurance of severe conditions. In this study, it has been attempted to Assay of enzymatic activity of facultative halophilic fungi from Tehran Forest Parks. Materials and methods: So sampling of soil and air of forest park of Tehran, saprophytes are been identified by the use of microscopic technique and slide culture. Then, the fungi separated were examined qualitatively regarding enzyme activity, and most strains were produced pectinase, lipase, protease, and amylase enzymes. Penicillium halotolerant and E01, E03, E014 strains produced invertase. And only the E13strain produced cellulase.So the enzyme activity of pectinase (via examining the enzyme produced in the media with pectin from pug) in a strain containing the enzyme. In the end stage, the best of enzyme produced strain are been identification to PCR technique and drawn phylogeny tree. Results: Most separated fungi were Aspergillus sp., Penicillium sp., Fusarium sp., Mucor sp., and Rhizopus sp. from among the fungi separated and identified, 19 isolates were considered for producing extracellular enzymes and were identified using molecular methods, such as Fusarium sublunatum, Aspergillus subramanianii, Penicillium hallotolerans, Mucor circinelloides, and Purpureocillium lilacinum. Cladosporium sp., Penicillium halotolerans strains showed the most activity (with activities of 0.71, and 0.79 U/mL). Conclusion: Cladosporium sp, and Penicillium halotolerans due to being cultivated on inexpensive substrates and easy enzyme extraction from the culture media, can be used for industrial enzyme production.
Haki GDaR, S.K. Developments in industrially important thermostable enzymes; areview. Bioresource Technology. 2003;89:17-34.
Kirk O, Borchert, T.V and Fuglsang, C.C. . Industrial enzyme applications. Current Opinion in Biotechnology 2002;13:345-51.
3. Oyeleke SBaO, A.A. Production of amylase by bacteria isolated from a cassava waste dumpsite in Minna, Niger state, Nigeria. African Journal of Microbiology Research 2009;3(4):143-6.
1. Haki GDaR, S.K. Developments in industrially important thermostable enzymes; areview. Bioresource Technology. 2003;89:17-34. 2. Kirk O, Borchert, T.V and Fuglsang, C.C. . Industrial enzyme applications. Current Opinion in Biotechnology 2002;13:345-51. 3. Oyeleke SBaO, A.A. Production of amylase by bacteria isolated from a cassava waste dumpsite in Minna, Niger state, Nigeria. African Journal of Microbiology Research 2009;3(4):143-6. 4. Gropinath SCBaea. Extracellular enzymatic activity profiles Fungi isolated from oil-rich enviroments. Mycorience. 2005;46:119-26.
5. Mishra BKaD, S.K. . Production of amylase and xylanase enzymes from soil fungi of Rajasthan. Journal of Advances in developmental Research 2010;1(1):21-3.
6. Walsh G. Proteins biochemistry and biotechnology: John Wiley and Sons. ; 2002. Chapter 6.
7. Ronivaldo Rodrigues da Silva. Bacterial and Fungal Proteolytic Enzymes: Production, Catalysis and Potential Applications. Applied Biochemistry and Biotechnology.2017;183(1):1-19.
8. Gostincar C, LENASSI M.Gunde- Cimerman N.,plemenitas A. Fungal adaption to extremely high salt concentration. Adv ApplMicrobial. 2011;77:71-96.
9. Mishra RV, D . Pandey ,BK . Pathak,N and Zeeshan ,M. Direct Colony Nested-PCR for the Detection of Fusarium oxysporum f. sp. Psidii Causing Wilt Disease in Psidium guajaval. Jof Horticulture 2014;1(2).
10. Imran A, Akbar.A, Yanwisetpakdee.B, Prasongsuk.S, Lotrakul.P ,and Punnapayak.H. Purification, Characterization, and Potential of Saline Waste Water Remediation of a Polyextremophilic α-Amylase from an Obligate Halophilic Aspergillus gracilis. BioMed Research International 2014; Article ID 106937, 7.
11. Espinel-Ingroff A. KT. Spectrophotometric method of inoculum preparation for the in vitro susceptibility testing of filamentous fungi. J Clin Microbial. 1991;29(2):393-4.
12. Brizzio Saea. Extracellular enzymatic activities of basidiomycetous yeasts isolated from glacial and subglacial waters of Northwest Patagonia (Argentina). Can J Microbiol. 2007;53(519-525).
13. Martmez - Trujillo Aaea. Constitutive and mducible pectinolytic enzymes from AspergUlustlavipes FP - 500 and their modulation by PH and carbon source. Braz J Microbiol. 2009;1(40).
14. Rajan A, Kumar, D.R.S and Nair, A.J. Isolation of a novel alkaline lipase producing fungus Aspergillus fumigatus MTCC 9657 from aged and crude rice bran oiland quantification by HPTLC. Isolation of a novel alkaline lipase producing fungus Aspergillus fumigatus MTCC 9657 from aged and crude rice bran oiland quantification by HPTLC. 2011;5(2):116-26.
15. Poonam S. N. Microbial Enzymes with Special Characteristics for Biotechnological Applications. Biomolecules. 2013 ; 3(3): 597–611.
16. Oluwayemisi OA. Effect of Blanching, Ripening and Other Treatments on the Production Characteristics of Pectinolytic Enzymes from Banana Peels by Aspergillus Niger. Global Journal of Science Frontier Research Chemistry. 2012;12(2).
17. Nouri goushki A, Kargar,M. ,Amini,J. . Identification and optimization of fungal pectinase isolated in screening stage for fruit juice industry. Journal of Microbial World. 2015;8(1).
18. Miller G. Use of Dinitrosalicylic acid reagent for determination of reducing sugars. Analytical Chemistry. 1959;31:426-9.
19. White TJ, Bruns, T., Lee, S., Taylor, J. W. . Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Academic Press, Inc. 1990:315-22.
20. Mueller GMaS, j.p. fungeal biodiversity : what do we know? what can we predict? Biodiversconserv. 2007;16:1-5.
21. Sandhya Caea. Comparative evaluation of neutral protease production by Aspergillus oiyzae in submerged and solid - state fermentation. Process Biochemistry. 2005;40:2689 - 94.
22. Gopinath SCBaea. Extracellular enzymatic activity profiles in fungi isolated from oil - rich environments. Mycoscience. 2005;46:119-26.
23. Kashyap DR, Soni SK, Tiwari R. Enhanced production of pectinase by Bacillus sp. DT7 using solid sate fermentation. Bioresource Technology. 2003;88(3):251-4.
24. Jayani R, Saxena, S. and Gupta, R. Microbial pectinolytic enzymes: A review. Process Biochemistry. 2005;40:2931-44.
25. Souza J, Silva, ES., Maia MLS and Teixeira MFS. Screening of fungal strains for pectinolytic activity: endopolygalacturonase production by Peacilomyces clavisporus 2A.UMIDA.1. Process Biochemistry. 2003;39:455-8.
26. De Souza PMaEM, P. O. . Application of microbial a - amylase in industry - a review. Brazilian Journal of Microbiology. 2010;41:850-61.
27. Padayachee T. Application of thermostable α - amylase from thermomyces lannginosus ATCC 58157 to nutritionally enhance starch based food.: Durban University of Technology, South Africa.; 2006.
28. Djamal C. Acid protease production by isolated species of penidllium. European Journal of Science Research. 2009;25(3):469-77.
29. Lakshmi P, Suresh Babu.B, Radhaiah.A and Sreeramulu.A. . Screening, Identification and Isolation of Cellulolytic fungi from soils of Chittoor District, India. IntJCurrMicrobiolAppSci 2014;3(7):661-71.
30. Anisa S K,Ashwini S, Girish K .Isolation and screening of Aspergillus spp. for pectinolytic activity. Electronic Journal of Biology 2013;9(2):37-41 ·
31. Mukunda S, Onkarappa.R, Prashith Kekuda.TR . Isolation and Screening of Industrially Important Fungi from the Soils of Western Ghats of Agumbe and Koppa, Karnataka. Technology and Arts Research Journal. 2012;1(4):27-32.
32. Maciel M, Herculano PN, Porto TS, eixeira MFST , Moreira KA and De Souza-Motta CM. . Production and partial characterization of pectinases from forage palm by Aspergillus niger URM4645. African Journal of Biotechnology. 2011;10(13):2469-79.