تولید و تعیین ویژگی های بیوسورفکتانت سویه بومی میکروباکتریوم تحمل کننده شوری جداشده از خاک های دریاچه نمک قم
محورهای موضوعی : میکروب شناسی محیطی
سید سهیل آقایى
1
,
فرزانه فخاریان
2
,
محمد رضا ذوالفقاری
3
,
محمد سلیمانی
4
1 - گروه میکروبیولوژی، دانشکده علوم پایه، دانشگاه آزاد اسلامی ، واحد قم، ایران.
2 - گروه میکروبیولوژی،دانشکده علوم پایه،دانشگاه آزاد اسلامی،قم،ایران
3 - دانشگاه آزاد اسلامی، واحد قم، گروه میکروبیولوژی
4 - گروه میکروبیولوژی،دانشکده علوم پایه،دانشگاه آزاد اسلامی،قم،ایران
کلید واژه: تحمل کننده نمک, بیوسورفکتانت, اکتینوباکتریا, میکروباکتریوم, خاک نمکی,
چکیده مقاله :
سابقه و هدف: سورفکتانت ها با منشا زیستی، ترکیبات آلی تولیدی توسط میکروارگانیسم ها از جمله کپک ها، مخمرها و باکتری ها هستند که با قرار گرفتن در بین سطوح، باعث کاهش کشش سطحی و بین سحطی می شوند. در این تحقیق به جداسازى سویه های اکتینوباکتریا تولید کننده بیوسورفکتانت از خاک دریاچه نمک قم پرداخته شد.مواد و روش ها: 110سویه اکتینوباکتریا از خاک جداسازی و برای تولید بیوسورفکتانت مورد سنجش قرار گرفتند. آزمون هاى رایج تولید بیوسورفکتانت(همولیز گلبول قرمز، آزمون پخش نفت، سنجش کشش سطح و غیره) صورت گرفت و در نهایت آنالیز 16S rRNA روى جدایه برتر مولد بیوسورفکتانت انجام شد. آزمون هاى کروماتوگرافى لایه نازک، طیف سنجى مادون قرمز و آنالیز ساختاری روى بیوسورفکتانت صورت گرفت. بهینه سازى تولید بیوسورفکتانت در حضور منابع کربن و نیتروژن مختلف و عوامل دما، pH و دور همزن انجام شد.نتایج: از بین ١١٠ سویه اکتینوباکتریا، ١٥ سویه قادر به تحمل نمک تا ١٠% بودند. با توجه به آزمون هاى سنجش تولید بیوسورفکتانت، ٨ سویه قادر به تولید بیوسورفکتانت بودند که از این میان سویه شماره 9 به عنوان بهترین سویه انتخاب شد و با آنالیز 16S rRNA در جنس میکروباکتریوم قرار گرفت. آنالیز هاى ساختارى گلیکولیپیدى بودن بیوسورفکتانت را مشخص نمودند. ساکارز و عصاره مخمر به عنوان بهترین منع کربن و نیتروژن و دمای 27 درجه سلسیوس، pH ، ١١ و دور همزن ١٧٠ rpm به عنوان شرایط بهینه انتخاب شدند.نتیجه گیری: نتایج این پژوهش پتانسیل سویه میکروباکتریوم تولیدکننده بیوسورفکتانت به منظور استفاده کاربردی در پاکسازی زیستی آب و اکوسیستم خاک را نشان می دهد.
Background & Objectives: Biosurfactants are biological surface active compounds produced by fungi, yeast and bacteria. These amphiphilic compounds can reduce surface tension and interfacial tension between individual molecules. The aims of this investigation was screening of biosurfactant producing halotolerant Actinobacteria species from the unexplored regions of Qom saline lake. Materials & methods: About 110 soil actinobacteria isolated strains were initially screened and then teste for their ability to BS production. Conventional screening methods of BS carried out using blood hemolysis, drop collapse method,oil spreading and surface tension measurements. 16S rRNA sequencing was done for the best biosurfactant producer strain. Further the partially purified BS was characterized by TLC, FTIR and compositional analysis. BS production was optimized using different carbon & nitrogen sources and optimized by different culture conditions such as temperatures, pH and stirring rate. Results: 15 out of 110 isolates were able to tolerate high salt concentrations up to 10% . 8 isolated strains were BS producer. Isolate No.9 showed 99% similarity to Microbacterium paraoxidans by 16S rRNA gene sequencing method. Compositional analysis methods proved a glycolipid structure of BS. Sucrose and yeast extract were identified as the most appropriate carbon and nitrogen source, respectively. Maximum production of BS was obtained in pH 7,at temperature 27 oC and stirring rate 170 rpm.Conclusion: These findings emphasize that such bacterial strains with superior BS production may find their potential application in bioremediation marine and soil ecosystem.
1. Desai JD, Banat IM. Microbial production of surfactants and their commercial potential.
Microbiol Mol Biol Rev. 1997;61(1):47-64.
2. Lin S-C. Review Biosurfactants: Recent Advances. J Chem Technol Biot. 1996;66(2):109-20.
3. Banat IM, Makkar RS, Cameotra SS. Potential commercial applications of microbial
surfactants. Applied microbiology and biotechnology. 2000;53(5):495-508.
4. Cameotra SS, Makkar RS. Recent applications of biosurfactants as biological and
immunological molecules. Current opinion in microbiology. 2004;7(3):262-6.
5. Cooper D . Biosurfactants. Microbiological sciences. 1986;3(5):145-9.
6. Hajfarajollah H, Mokhtarani B, Noghabi KA. Newly antibacterial and antiadhesive lipopeptide
biosurfactant secreted by a probiotic strain, Propionibacterium freudenreichii. Applied
biochemistry and biotechnology. 2014;174(8):2725-40.
7. Větrovský T, Baldrian P. An in-depth analysis of actinobacterial communities shows their high
diversity in grassland soils along a gradient of mixed heavy metal contamination. Biol Fert
Soils. 2015;51(7):827-37.
8. Augustine SK, Bhavsar SP, Kapadnis BP. A non-polyene antifungal antibiotic from
Streptomyces albidoflavus PU 23. J BiosciJ Biosci. 2005;30(2):201-11.
9. Richter M, Willey JM, Süßmuthßmuth R, Jung n, Fiedler H-P. Streptofactin, a novel
biosurfactant with aerial mycelium inducing activity fromStreptomyces tendaeTü 901/.8c.
FEMS Microbiology Letters. 1998;163(2):165-71.
10. Deepika L, Krishnan K. Biosurfactant and Heavy Metal Resistance Activity of Streptomyces
spp. Isolated from Saltpan Soil. Br J Pharmacol Toxicol. 2010;1.
11. Pacheco J, Ciapina EM, omes Ede B, Junior NP. Biosurfactant production by rhodococcus
erythropolis and its application to oil removal. Braz J Microbiol. 2010;41(3):685-93.
12. Vyas TK, Dave BP. Production of biosurfactant by Nocardia otitidiscaviarum and its role in
biodegradation of crude oil. International journal of systematic and evolutionary microbiology.
2011;8(2):425-32.
13. Manivasagan P, Sivasankar P, Venkatesan J, Sivakumar K, Kim SK. Optimization, production
and characterization of glycolipid biosurfactant from the marine actinobacterium, Streptomyces
sp. MAB36. Bioprocess and biosystems engineering. 2014;37(5):783-97.
Acknowledgment
We would gratefully acknowledge the
support from the University of
Tehran research center of new technologies
in life science engineering.
Conflict of nterest
The authors declare that there is no conflict of
interests regarding the publication of this
paper.
437
Journal of Microbial World, Volume 12, No. 4, January 2020. Production and characterization of biosurfactant by indigenous halotolerant… Farzaneh Fakharian et al.
14. Thavasi R. Biosurfactants from marine hydrocarbonoclastic bacteria and their application in
marine oil pollution abatement Annamalai University, Chidambaram.
15. Khopade A, Ren B, Liu , Mahadik K, Zhang L, Kokare C. Production and characterization
of biosurfactant from marine Streptomyces species B3. Journal of colloid and interface science.
2012;367(1):311-8.
16. Carrillo P , Mardaraz C, Pitta-Alvarez SI, iulietti AM. Isolation and selection of
biosurfactant-producing bacteria. World journal of microbiology & biotechnology. 1996;12
(1):82-4.
17. Morikawa M, Hirata , Imanaka T. A study on the structure-function relationship of
lipopeptide biosurfactants. Biochimica et biophysica acta. 2000;1488(3):211-8.
18. oussef NH, Duncan KE, Nagle DP, Savage KN, Knapp RM, McInerney MJ. Comparison of
methods to detect biosurfactant production by diverse microorganisms. Journal of
microbiological methods. 2004;56(3):339-47.
19. andhimathi R, Seghal Kiran , Hema TA, Selvin J, Rajeetha Raviji T, Shanmughapriya S.
Production and characterization of lipopeptide biosurfactant by a sponge-associated marine
actinomycetes Nocardiopsis alba MSA10. Bioprocess and biosystems engineering. 2009;32
(6):825-35.
20. Cooper D , oldenberg B . Surface-active agents from two bacillus species. Appl Environ
Microbiol. 1987;53(2):224-9.
21. Bergey DH, Buchanan RE, ibbons NE, American Society for M. Bergey s manual of
determinative bacteriology. Baltimore: Williams & Wilkins Co.; 1974.
22. Cook AE, Meyers PR. Rapid identification of filamentous actinomycetes to the genus level
using genus-specific 16S rRNA gene restriction fragment patterns. International journal of
systematic and evolutionary microbiology. 2003;53(Pt 6):1907-15.
23. Altschul SF, ish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool.
Journal of molecular biology. 1990;215(3):403-10.
24. Sharafi H, Abdoli M, Hajfarajollah H, Samie N, Alidoust L, Abbasi H, et al. First report of a
lipopeptide biosurfactant from thermophilic bacterium Aneurinibacillus thermoaerophilus
MK01 newly isolated from municipal landfill site. Applied biochemistry and biotechnology.
2014;173(5):1236-49.
25. Saikia RR, Deka S, Deka M, Banat IM. Isolation of biosurfactant-producing Pseudomonas
aeruginosa RS29 from oil-contaminated soil and evaluation of different nitrogen sources in
biosurfactant production. Annals of Microbiology. 2011;62(2):753-63.
26. Cury JA, Rebello MA, Del Bel Cury AA. In situ relationship between sucrose exposure and
the composition of dental plaque. Caries research. 1997;31(5):356-60.
27. Folch J, Lees M, Sloane Stanley H. A simple method for the isolation and purification of
total lipides from animal tissues. The Journal of biological chemistry. 1957;226(1):497-509.
28. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of
protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72(1):248-54.
1. Desai JD, Banat IM. Microbial production of surfactants and their commercial potential.
Microbiol Mol Biol Rev. 1997;61(1):47-64.
2. Lin S-C. Review Biosurfactants: Recent Advances. J Chem Technol Biot. 1996;66(2):109-20.
3. Banat IM, Makkar RS, Cameotra SS. Potential commercial applications of microbial
surfactants. Applied microbiology and biotechnology. 2000;53(5):495-508.
4. Cameotra SS, Makkar RS. Recent applications of biosurfactants as biological and
immunological molecules. Current opinion in microbiology. 2004;7(3):262-6.
5. Cooper D . Biosurfactants. Microbiological sciences. 1986;3(5):145-9.
6. Hajfarajollah H, Mokhtarani B, Noghabi KA. Newly antibacterial and antiadhesive lipopeptide
biosurfactant secreted by a probiotic strain, Propionibacterium freudenreichii. Applied
biochemistry and biotechnology. 2014;174(8):2725-40.
7. Větrovský T, Baldrian P. An in-depth analysis of actinobacterial communities shows their high
diversity in grassland soils along a gradient of mixed heavy metal contamination. Biol Fert
Soils. 2015;51(7):827-37.
8. Augustine SK, Bhavsar SP, Kapadnis BP. A non-polyene antifungal antibiotic from
Streptomyces albidoflavus PU 23. J BiosciJ Biosci. 2005;30(2):201-11.
9. Richter M, Willey JM, Süßmuthßmuth R, Jung n, Fiedler H-P. Streptofactin, a novel
biosurfactant with aerial mycelium inducing activity fromStreptomyces tendaeTü 901/.8c.
FEMS Microbiology Letters. 1998;163(2):165-71.
10. Deepika L, Krishnan K. Biosurfactant and Heavy Metal Resistance Activity of Streptomyces
spp. Isolated from Saltpan Soil. Br J Pharmacol Toxicol. 2010;1.
11. Pacheco J, Ciapina EM, omes Ede B, Junior NP. Biosurfactant production by rhodococcus
erythropolis and its application to oil removal. Braz J Microbiol. 2010;41(3):685-93.
12. Vyas TK, Dave BP. Production of biosurfactant by Nocardia otitidiscaviarum and its role in
biodegradation of crude oil. International journal of systematic and evolutionary microbiology.
2011;8(2):425-32.
13. Manivasagan P, Sivasankar P, Venkatesan J, Sivakumar K, Kim SK. Optimization, production
and characterization of glycolipid biosurfactant from the marine actinobacterium, Streptomyces
sp. MAB36. Bioprocess and biosystems engineering. 2014;37(5):783-97.
Acknowledgment
We would gratefully acknowledge the
support from the University of
Tehran research center of new technologies
in life science engineering.
Conflict of nterest
The authors declare that there is no conflict of
interests regarding the publication of this
paper.
437
Journal of Microbial World, Volume 12, No. 4, January 2020. Production and characterization of biosurfactant by indigenous halotolerant… Farzaneh Fakharian et al.
14. Thavasi R. Biosurfactants from marine hydrocarbonoclastic bacteria and their application in
marine oil pollution abatement Annamalai University, Chidambaram.
15. Khopade A, Ren B, Liu , Mahadik K, Zhang L, Kokare C. Production and characterization
of biosurfactant from marine Streptomyces species B3. Journal of colloid and interface science.
2012;367(1):311-8.
16. Carrillo P , Mardaraz C, Pitta-Alvarez SI, iulietti AM. Isolation and selection of
biosurfactant-producing bacteria. World journal of microbiology & biotechnology. 1996;12
(1):82-4.
17. Morikawa M, Hirata , Imanaka T. A study on the structure-function relationship of
lipopeptide biosurfactants. Biochimica et biophysica acta. 2000;1488(3):211-8.
18. oussef NH, Duncan KE, Nagle DP, Savage KN, Knapp RM, McInerney MJ. Comparison of
methods to detect biosurfactant production by diverse microorganisms. Journal of
microbiological methods. 2004;56(3):339-47.
19. andhimathi R, Seghal Kiran , Hema TA, Selvin J, Rajeetha Raviji T, Shanmughapriya S.
Production and characterization of lipopeptide biosurfactant by a sponge-associated marine
actinomycetes Nocardiopsis alba MSA10. Bioprocess and biosystems engineering. 2009;32
(6):825-35.
20. Cooper D , oldenberg B . Surface-active agents from two bacillus species. Appl Environ
Microbiol. 1987;53(2):224-9.
21. Bergey DH, Buchanan RE, ibbons NE, American Society for M. Bergey s manual of
determinative bacteriology. Baltimore: Williams & Wilkins Co.; 1974.
22. Cook AE, Meyers PR. Rapid identification of filamentous actinomycetes to the genus level
using genus-specific 16S rRNA gene restriction fragment patterns. International journal of
systematic and evolutionary microbiology. 2003;53(Pt 6):1907-15.
23. Altschul SF, ish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool.
Journal of molecular biology. 1990;215(3):403-10.
24. Sharafi H, Abdoli M, Hajfarajollah H, Samie N, Alidoust L, Abbasi H, et al. First report of a
lipopeptide biosurfactant from thermophilic bacterium Aneurinibacillus thermoaerophilus
MK01 newly isolated from municipal landfill site. Applied biochemistry and biotechnology.
2014;173(5):1236-49.
25. Saikia RR, Deka S, Deka M, Banat IM. Isolation of biosurfactant-producing Pseudomonas
aeruginosa RS29 from oil-contaminated soil and evaluation of different nitrogen sources in
biosurfactant production. Annals of Microbiology. 2011;62(2):753-63.
26. Cury JA, Rebello MA, Del Bel Cury AA. In situ relationship between sucrose exposure and
the composition of dental plaque. Caries research. 1997;31(5):356-60.
27. Folch J, Lees M, Sloane Stanley H. A simple method for the isolation and purification of
total lipides from animal tissues. The Journal of biological chemistry. 1957;226(1):497-509.
28. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of
protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72(1):248-54.