Advertisement

Annals of Microbiology

, Volume 69, Issue 9, pp 895–907 | Cite as

Surface properties and exopolysaccharide production of surface-associated microorganisms isolated from a dairy plant

  • Dilay Kütük AyhanEmail author
  • Ayhan Temiz
  • Farzin Asghari Sana
  • Menemşe Gümüşderelioğlu
Original Article
  • 42 Downloads

Abstract

Purpose

The purpose of this study was to isolate the surface-associated microorganisms from the dairy plant surfaces with a high probability of biofilm formation and determine the most adhesive strains in terms of surface properties and exopolysaccharide production.

Methods

Four hundred and ninety-five surface-associated microorganisms were isolated from potential biofilm-forming surfaces of a dairy plant. One hundred and seventy of these were isolated after cleaning/disinfection of the pasteurized milk, white cheese and butter tank, yogurt and ice cream filling unit, ice cream air pressing, and condensed milk pipe. It is noteworthy that some isolates might cause post-production contamination, food infection, and intoxication. Selected 42 isolates were identified by Gram staining, physiological and biochemical tests, and 16S rRNA gene sequencing. Then, surface properties and exopolysaccharide production of 10 selected isolates were determined. To evaluate the surface properties, microbial adhesion to hydrocarbons, static water contact angle, salt aggregation, and surface zeta potential tests were performed.

Result

The microbial adhesion to hydrocarbons (MATH) test exhibited the lowest standard deviations, and the most consistent results between the replicates. The highest hydrophilic characteristics and exopolysaccharide production were exhibited by Gram-negative Pseudomonas aeruginosa, followed by Gram-positive Bacillus toyonensis. Also, a significant diversity of neutral sugar was determined in their alditol acetate forms by using gas chromatography–mass spectrometry. In this context, it is believed that the determination of the EPS content of the isolates would contribute to establishing an effective cleaning/disinfection procedure for dairy plants.

Conclusion

This study indicated that microbial adhesion is still a common problem in the dairy industry. Because of this situation, dairy plants should be organized and constructed to be suitable for hygiene and sanitary applications.

Keywords

Dairy plant Adhesion Identification Surface property Exopolysaccharide production Neutral sugar content 

Notes

Funding information

This work was supported by the Hacettepe University Scientific Research Projects Coordination Unit (Project Codes: 014 D01 602 003 and FDK-2016-13096).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Research involving human participants and/or animals

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed consent

Not applicable.

References

  1. Abasolo-Pacheco F, Saucedo PE, Mazon-Suastegui JM et al (2015) Isolation and use of beneficial microbiota from the digestive tract of lions-paw scallop Nodipecten subnodosus and winged pearl oyster Pteria sterna in oyster aquaculture. Aquac Res:1–10.  https://doi.org/10.1111/are.12754
  2. Absolom DR, Lamberti FV, Policova Z et al (1983) Surface thermodynamics of bacterial adhesion. Appl Environ Microbiol 46:90–97Google Scholar
  3. Ahimou F, Paquot M, Jacques P et al (2001) Influence of electrical properties on the evaluation of the surface hydrophobicity of Bacillus subtilis. J Microbiol Methods 45:119–126.  https://doi.org/10.1016/S0167-7012(01)00240-8 CrossRefGoogle Scholar
  4. Arana I, Santorum P, Muela A, Barcina I (1999) Chlorination and ozonation of waste-water: comparative analysis of efficacy through the effect on Escherichia coli membranes. J Appl Microbiol 86:883–888CrossRefGoogle Scholar
  5. Bağcı U (2012) Determination of some important properties of lactic acid bacteria isolated from human milk based on food technology. Dissertation, Hacettepe University, Graduate School of Science and EngineeringGoogle Scholar
  6. Bales PM, Renke EM, May SL et al (2013) Purification and characterization of biofilm-associated EPS exopolysaccharides from ESKAPE organisms and other pathogens. PLoS One 8:1–8.  https://doi.org/10.1371/journal.pone.0067950 CrossRefGoogle Scholar
  7. Barnes RL, Caskey DK (2002) Using ozone in the prevention of bacterial biofilm formation and scaling. In: Water cond purification, technical report. October, vol 44, pp 1–3Google Scholar
  8. Bernardes PC, de Andrade NJ, Ferreira SO et al (2010) Assessment of hydrophobicity and roughness of stainless steel adhered by an isolate of Bacillus cereus from a dairy plant. Brazilian J Microbiol 41:984–992.  https://doi.org/10.1590/S1517-83822010000400017 CrossRefGoogle Scholar
  9. Bjerkan G, Witsø E, Bergh K (2009) Sonication is superior to scraping for retrieval of bacteria in biofilm on titanium and steel surfaces in vitro. Acta Orthop 80:245–250.  https://doi.org/10.3109/17453670902947457 CrossRefGoogle Scholar
  10. Blackman IC, Frank JF (1996) Growth of Listeria monocytogenesas a biofilm on various food-processing surfaces. J Food Prot 59:827–831CrossRefGoogle Scholar
  11. Blumenkrantz N, Asboe-Hansen G (1973) New method for quantitative determination of uronic acids. Anal Biochem 54:484–489.  https://doi.org/10.1016/0003-2697(73)90377-1 CrossRefGoogle Scholar
  12. Boonaert CJP, Dufrêne YF, Derclaye SR, Rouxhet PG (2001) Adhesion of Lactococcus lactis to model substrata: direct study of the interface. Colloids Surf B Biointerfaces 22:171–182.  https://doi.org/10.1016/S0927-7765(01)00196-5 CrossRefGoogle Scholar
  13. Boulange-Petermann L, Baroux B, Bellon-Fontaine M-N (1993) The influence of metallic surface wettability on bacterial adhesion. J Adhes Sci Technol 7:221–230.  https://doi.org/10.1163/156856193X00673 CrossRefGoogle Scholar
  14. Brolazo EM, Leite DS, Tiba MR et al (2011) Correlation between api 50 ch and multiplex polymerase chain reaction for the identification of vaginal lactobacilli in isolates. Brazilian J Microbiol 42:225–232.  https://doi.org/10.1590/S1517-83822011000100028 CrossRefGoogle Scholar
  15. Bryers JD (1987) Biologically active surfaces: processes governing the formation and persistence of biofilms. Biotechnol Prog 3:57–68.  https://doi.org/10.1002/btpr.5420030202 CrossRefGoogle Scholar
  16. Chen X, Stewart PS (2002) Role of electrostatic interactions in cohesion of bacterial biofilms. Appl Microbiol Biotechnol 59:718–720.  https://doi.org/10.1007/s00253-002-1044-2 CrossRefGoogle Scholar
  17. Cherif-Antar A, Moussa–Boudjemâa B, Didouh N et al (2016) Diversity and biofilm-forming capability of bacteria recovered from stainless steel pipes of a milk-processing dairy plant. Dairy Sci Technol 96:27–38.  https://doi.org/10.1007/s13594-015-0235-4 CrossRefGoogle Scholar
  18. Donlan RM, Costerton JW (2002) Biofilms: survival mechanisms of clinically relevant microorganisms. Clin Microbiol 15:167–193.  https://doi.org/10.1128/CMR.15.2.167 CrossRefGoogle Scholar
  19. Drenkard E (2003) Antimicrobial resistance of Pseudomonas aeruginosa biofilms. Microbes Infect 5:1213–1219.  https://doi.org/10.1016/j.micinf.2003.08.009 CrossRefGoogle Scholar
  20. DuBois M, Gilles KA, Hamilton JK et al (1956) Colorimetric method for determination of sugars and related substances. Anal Chem 28:350–356.  https://doi.org/10.1021/ac60111a017 CrossRefGoogle Scholar
  21. Feng L, Li X, Du G, Chen J (2009) Adsorption and fouling characterization of Klebsiella oxytoca to microfiltration membranes. Process Biochem 44:1289–1292.  https://doi.org/10.1016/j.procbio.2009.07.017 CrossRefGoogle Scholar
  22. Fox A (1999) Carbohydrate profiling of bacteria by gas chromatography – mass spectrometry and their trace detection in complex matrices by gas chromatography – tandem mass spectrometry. J Chromatogr A 843:287–300CrossRefGoogle Scholar
  23. Frank JF (2001) Microbial attachment to food and food contact surfaces. Adv Food Nutr Res 43:319–370.  https://doi.org/10.1016/S1043-4526(01)43008-7 CrossRefGoogle Scholar
  24. Frank JF, Koffi RA (1990) Surface adherence growth of Listeria monocytogenes is associated with increased resistance to surfactant sanitizers and heat. J Food Prot 53:550–554CrossRefGoogle Scholar
  25. Gallardo-Moreno AM, Pacha-Olivenza MA, Saldana L et al (2009) In vitro biocompatibility and bacterial adhesion of physico-chemically modified Ti6Al4V surface by means of UV irradiation. Acta Biomater 5:181–192.  https://doi.org/10.1016/j.actbio.2008.07.028 CrossRefGoogle Scholar
  26. Gómez-Suárez C, Pasma J, van der Borden AJ et al (2002) Influence of extracellular polymeric substances on deposition and redeposition of Pseudomonas aeruginosa to surfaces. Microbiology 148:1161–1169.  https://doi.org/10.1099/00221287-148-4-1161 CrossRefGoogle Scholar
  27. Hamadi F, Latrache H (2008) Comparison of contact angle measurement and microbial adhesion to solvents for assaying electron donor – electron acceptor (acid – base) properties of bacterial surface. Colloids Surf B Biointerfaces 65:134–139.  https://doi.org/10.1016/j.colsurfb.2008.03.010 CrossRefGoogle Scholar
  28. Hanlon GW, Olliff CJ, Brant JA, Denyer SP (1999) A novel image-analysis technique for measurement of bacterial cell surface tension. J Pharm Pharmacol 51:207–214CrossRefGoogle Scholar
  29. Harrigan WF (1998) Laboratory methods in food microbiology. Academic Press, San DiegoGoogle Scholar
  30. Hyde FW, Alberg M, Smith K (1997) Comparison of fluorinated polymers against stainless steel, glass and polypropylene in microbial biofilm adherence and removal. J Ind Microbiol Biotechnol 19:142–149.  https://doi.org/10.1038/sj.jim.2900448 CrossRefGoogle Scholar
  31. Jang A, Szabo J, Hosni AA et al (2006) Measurement of chlorine dioxide penetration in dairy process pipe biofilms during disinfection. Appl Microbiol Biotechnol 72:368–376.  https://doi.org/10.1007/s00253-005-0274-5 CrossRefGoogle Scholar
  32. Jones DS, Adair CG, Mawhinney WM, Gorman SP (1996) Standardisation and comparison of methods employed for microbial cell surface hydrophobicity and charge determination. Int J Pharm 131:83–89.  https://doi.org/10.1016/0378-5173(95)04368-3 CrossRefGoogle Scholar
  33. Kajiyama S, Tsurumoto T, Osaki M et al (2009) Quantitative analysis of Staphylococcus epidermidis biofilm on the surface of biomaterial. J Orthop Sci 14:769–775.  https://doi.org/10.1007/s00776-009-1405-0 CrossRefGoogle Scholar
  34. Kiran GS, Priyadharshini S, Anitha K et al (2015) Characterization of an exopolysaccharide from probiont Enterobacter faecalis MSI12 and its effect on the disruption of Candida albicans biofilm. RSC Adv 5:71573–71585.  https://doi.org/10.1039/C5RA10302A CrossRefGoogle Scholar
  35. Knight GC, Craven HM (2010) A model system for evaluating surface disinfection in dairy factory environments. Int J Food Microbiol 137:161–167.  https://doi.org/10.1016/j.ijfoodmicro.2009.11.028 CrossRefGoogle Scholar
  36. Kwaszewska AK, Brewczyńska A, Szewczyk EM (2006) Hydrophobicity and biofilm formation of lipophilic skin corynebacteria. Polish J Microbiol 55:189–193Google Scholar
  37. Li B, Logan BE (2004) Bacterial adhesion to glass and metal-oxide surfaces. Colloids Surf B: Biointerfaces 36:81–90.  https://doi.org/10.1016/j.colsurfb.2004.05.006 CrossRefGoogle Scholar
  38. Li J, McLandsborough LA (1999) The effects of the surface charge and hydrophobicity of Escherichia coli on its adhesion to beef muscle. Int J Food Microbiol 53:185–193.  https://doi.org/10.1016/S0168-1605(99)00159-2 CrossRefGoogle Scholar
  39. Li Z, Zhong S, Lei H et al (2009) Production of a novel bioflocculant by Bacillus licheniformis X14 and its application to low temperature drinking water treatment. Bioresour Technol 100:3650–3656.  https://doi.org/10.1016/j.biortech.2009.02.029 CrossRefGoogle Scholar
  40. Ljungh A, Wadström T (1995) Growth conditions influence expression of cell surface hydrophobicity of staphylococci and other wound infection pathogens. Microbiol Immunol 39:753–757CrossRefGoogle Scholar
  41. Ljungh A, Hjerten S, Wadstrom T (1985) High surface hydrophobicity of autoaggregating Staphylococcus aureus strains isolated from human infections studied with the salt aggregation test. Infect Immun 47:522–526Google Scholar
  42. Lortal S, Di Blasi A, Madec MN et al (2009) Tina wooden vat biofilm: a safe and highly efficient lactic acid bacteria delivering system in PDO Ragusano cheese making. Int J Food Microbiol 132:1–8.  https://doi.org/10.1016/j.ijfoodmicro.2009.02.026 CrossRefGoogle Scholar
  43. Mafu AA, Roy D, Goulet J, Hagny P (1990) Attachment of Listeria monocytogenes to stainless steel, glass, polypropylene and rubber surfaces after short contact times. J Food Prot 53:742–746CrossRefGoogle Scholar
  44. Marín ML, Benito Y, Pin C et al (1997) Lactic acid bacteria: hydrophobicity and strength of attachment to meat surfaces. Lett Appl Microbiol 24:14–18CrossRefGoogle Scholar
  45. Marques SC, Rezende JDGOS, Alves LADF et al (2007) Formation of biofilms by Staphylococcus aureus on stainless steel and glass surfaces and its resistance to some selected chemical sanitizers. Brazilian J Microbiol 38:538–543.  https://doi.org/10.1590/S1517-83822007000300029 CrossRefGoogle Scholar
  46. Marshall KC (1992) Biofilms: an overview of bacterial adhesion activity and control at surfaces. Am Soc Microbiol 58:202–207Google Scholar
  47. Martín R, Olivares M, Pérez M et al (2010) Identification and evaluation of the probiotic potential of lactobacilli isolated from canine milk. Vet J 185:193–198.  https://doi.org/10.1016/j.tvjl.2009.04.014 CrossRefGoogle Scholar
  48. Meyer B (2003) Approaches to prevention, removal and killing of biofilms. Int Biodeterior Biodegrad 51:249–253.  https://doi.org/10.1016/S0964-8305(03)00047-7 CrossRefGoogle Scholar
  49. Minagi S, Miyake Y, Yumi F et al (1986) Cell-surface hydrophobicity of Candida species as determined by the contact-angle and hydrocarbon-adherence methods. J Gen Microbiol 132:1111–1115Google Scholar
  50. Mozzi F, Vaningelgem F, Hebert ME et al (2006) Diversity of heteropolysaccharide-producing lactic acid bacterium strains and their biopolymers. Appl Environ Microbiol 72:4431–4435.  https://doi.org/10.1128/AEM.02780-05 CrossRefGoogle Scholar
  51. Mustapha A, Liewen MB (1989) Destruction of Listeria monocytogenes by sodium hypochlorite and quaternary ammonium sanitizers. J Food Prot 52:306–311CrossRefGoogle Scholar
  52. Nel HA, Bauer R, Wolfaardt GM, Dicks LMT (2002) Effect of bacteriocins pediocin PD-1, plantaricin 423, and nisin on biofilms of Oenococcus oeni on a stainless steel surface. Am J Enol Vitic 53:191–196Google Scholar
  53. Obuekwe CO, Al-Jadi ZK, Al-Saleh ES (2009) Hydrocarbon degradation in relation to cell-surface hydrophobicity among bacterial hydrocarbon degraders from petroleum-contaminated Kuwait desert environment. Int Biodeterior Biodegrad 63:273–279.  https://doi.org/10.1016/j.ibiod.2008.10.004 CrossRefGoogle Scholar
  54. Ophir T, Gutnick DL (1994) A role for exopolysaccharides in the protection of microorganisms from desiccation. Appl Environ Microbiol 60:740–745Google Scholar
  55. Palmer JS, Flint SH, Schmid J, Brooks JD (2010) The role of surface charge and hydrophobicity in the attachment of Anoxybacillus flavithermus isolated from milk powder. J Ind Microbiol Biotechnol 37:1111–1119.  https://doi.org/10.1007/s10295-010-0758-x CrossRefGoogle Scholar
  56. Pasmore M, Todd P, Smith S et al (2001) Effects of ultrafiltration membrane surface properties on Pseudomonas aeruginosa biofilm initiation for the purpose of reducing biofouling. J Memb Sci 194:15–32.  https://doi.org/10.1016/S0376-7388(01)00468-9 CrossRefGoogle Scholar
  57. Rijnaarts HHM, Norde W, Bouwer EJ et al (1995) Reversibility and mechanism of bacterial adhesion. Colloids Surf B Biointerfaces 4:5–22.  https://doi.org/10.1016/0927-7765(94)01146-V CrossRefGoogle Scholar
  58. Rinker KD, Kelly RM (1996) Growth physiology of the hyperthermophilic archaeon Thermococcus litoralis: development of a sulfur-free defined medium, characterization of an exopolysaccharide, and evidence of biofilm formation. Appl Environ Microbiol 62:4478–4485Google Scholar
  59. Roberson EB, Firestone MK (1992) Relationship between desiccation and exopolysaccharide production in a soil Pseudomonas sp. Appl Environ Microbiol 58:1284–1291Google Scholar
  60. Rosenberg M, Gutnick D, Rosenberg E (1980) Adherence of bacteria to hydrocarbons: a simple method for measuring cell-surface hydrophobicity. FEMS Microbiol Lett 9:29–33.  https://doi.org/10.1017/CBO9781107415324.004 CrossRefGoogle Scholar
  61. Saini G (2010) Bacterial hydrophobicity: assessment techniques, applications and extension to colloids. Dissertation, Oregon State University, Chemical EngineeringGoogle Scholar
  62. Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Nati Acad Sci USA 74:5463–5467CrossRefGoogle Scholar
  63. Sassaki GL, Souza LM, Serrato RV et al (2008) Application of acetate derivatives for gas chromatography-mass spectrometry: novel approaches on carbohydrates, lipids and amino acids analysis. J Chromatogr A 1208:215–222.  https://doi.org/10.1016/j.chroma.2008.08.083 CrossRefGoogle Scholar
  64. Sinde E, Carballo J (2000) Attachment of Salmonella spp. and Listeria monocytogenes to stainless steel, rubber and polytetrafluorethylene: the influence of free energy and the effect of commercial sanitizers. Food Microbiol 17:439–447.  https://doi.org/10.1006/fmic.2000.0339 CrossRefGoogle Scholar
  65. Smoot LM, Pierson MD (1998) Effect of environmental stress on the ability of Listeria monocytogenes Scott a to food contact surfaces. J Food Prot 61:1293–1298CrossRefGoogle Scholar
  66. Soares JC, Marques MR, Tavaria FK et al (2011) Biodiversity and characterization of Staphylococcus species isolated from a small manufacturing dairy plant in Portugal. Int J Food Microbiol 146:123–129.  https://doi.org/10.1016/j.ijfoodmicro.2011.02.008 CrossRefGoogle Scholar
  67. Stewart PS, Roe F, Rayner J et al (2000) Effect of catalase on hydrogen peroxide penetration into Pseudomonas aeruginosa biofilms. Appl Environ Microbiol 66:836–838.  https://doi.org/10.1128/AEM.66.2.836-838.2000 CrossRefGoogle Scholar
  68. Strathmann M, Wingender J, Flemming HC (2002) Application of fluorescently labelled lectins for the visualization and biochemical characterization of polysaccharides in biofilms of Pseudomonas aeruginosa. J Microbiol Methods 50:237–248.  https://doi.org/10.1016/S0167-7012(02)00032-5 CrossRefGoogle Scholar
  69. Styriak I, Laukova A, Fallgren C, Wadström T (1999) Binding of selected extracellular matrix proteins to enterococci and Streptococcus bovis of animal origin. Curr Microbiol 39:327–335.  https://doi.org/10.1007/s002849900467 CrossRefGoogle Scholar
  70. Sutherland IW (1982) Microbial exopolysaccharides - their role in microbial adhesion in aqueous systems. Crit Rev Microbiol 10:173–201.  https://doi.org/10.3109/10408418209113562 CrossRefGoogle Scholar
  71. Suzuki C, Kobayashi M, Kimoto-nira H (2013) Novel exopolysaccharides produced by Lactococcus lactis subsp. lactis, and the diversity of epsE genes in the exopolysaccharide biosynthesis gene clusters. Biosci Biotechnol Biochem 77:2013–2018.  https://doi.org/10.1271/bbb.130322 CrossRefGoogle Scholar
  72. Tang L, Pillai S, Revsbech NP et al (2011) Biofilm retention on surfaces with variable roughness and hydrophobicity. Biofouling 27:111–121.  https://doi.org/10.1080/08927014.2010.544848 CrossRefGoogle Scholar
  73. Temiz A (2010) Genel Mikrobiyoloji Uygulama Teknikleri. Hatipoğlu Yayınları, AnkaraGoogle Scholar
  74. Triandafillu K, Balazs DJ, Aronsson BO et al (2003) Adhesion of Pseudomonas aeruginosa strains to untreated and oxygen-plasma treated poly(vinyl chloride) (PVC) from endotracheal intubation devices. Biomaterials 24:1507–1518.  https://doi.org/10.1016/S0142-9612(02)00515-X CrossRefGoogle Scholar
  75. Ukuku DO, Fett WF (2002) Relationship of cell surface charge and hydrophobicity to strength of attachment of bacteria to cantaloupe rind. J Food Prot 65:1093–1099CrossRefGoogle Scholar
  76. Valeriano C, de Oliveira TLC, de Carvalho SM et al (2012) The sanitizing action of essential oil-based solutions against Salmonella enterica serotype Enteritidis S64 biofilm formation on AISI 304 stainless steel. Food Control 25:673–677.  https://doi.org/10.1016/j.foodcont.2011.12.015 CrossRefGoogle Scholar
  77. van Merode AEJ, Duval JFL, van der Mei HC et al (2008) Increased adhesion of Enterococcus faecalis strains with bimodal electrophoretic mobility distributions. Colloids Surf B Biointerfaces 64:302–306.  https://doi.org/10.1016/j.colsurfb.2008.02.004 CrossRefGoogle Scholar
  78. Vanhaecke E, Remon J, Moors M et al (1990) Kinetics of Pseudomonas aeruginosa adhesion to 304 and 316-L stainless steel: role of cell surface hydrophobicity. Appl Environ Microbiol 56:788–795Google Scholar
  79. Waines PL, Moate R, Moody AJ et al (2011) The effect of material choice on biofilm formation in a model warm water distribution system. Biofouling 27:1161–1174.  https://doi.org/10.1080/08927014.2011.636807 CrossRefGoogle Scholar
  80. Wang SY, Chen KN, Lo YM et al (2012) Investigation of microorganisms involved in biosynthesis of the kefir grain. Food Microbiol 32:274–285.  https://doi.org/10.1016/j.fm.2012.07.001 CrossRefGoogle Scholar
  81. Watnick P, Kolter R (2000) Biofilm, city of microbes. J Bacteriol 182:2675–2679.  https://doi.org/10.1128/JB.182.10.2675-2679.2000 CrossRefGoogle Scholar
  82. Wingender J, Strathmann M, Rode A et al (2001) Isolation and biochemical characterization of extracellular polymeric substances from Pseudomonas aeruginosa. Methods Enzymol 336:302–314CrossRefGoogle Scholar
  83. Yang Z (2000) Antimicrobial compounds and extracellular polysaccharides produced by lactic acid bacteria: structures and properties. Dissertation, University of Helsinki, Department of Food TechnologyGoogle Scholar
  84. Yokota S, Kaya S, Sawada S et al (1987) Characterization of a polysaccharide component of lipopolysaccharide from Pseudomonas aeruginosa IID 1008 (ATCC 27584) as D-rhamnan. Eur J Biochem 209:203–209CrossRefGoogle Scholar

Copyright information

© Università degli studi di Milano 2019

Authors and Affiliations

  1. 1.Department of Food EngineeringHacettepe UniversityAnkaraTurkey
  2. 2.Department of Nanotechnology & NanomedicineHacettepe UniversityAnkaraTurkey
  3. 3.Department of Chemical EngineeringHacettepe UniversityAnkaraTurkey

Personalised recommendations