Formal- and high-structured kinetic process modelling and footprint area analysis of binary imaged cells: Tools to understand and optimize multistage-continuous PHA biosynthesis

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Abstract

Competitive polyhydroxyalkanoate (PHAs) production requires progress in microbial strain performance, feedstock selection, downstream processing, and more importantly according to the process design with process kinetics of the microbial growth phase and the phase of product formation. The multistage continuous production in a bioreactor cascade was described for the first time in a continuously operated, flexible five-stage bioreactor cascade that mimics the characteristics involved in the engineering process of tubular plug flow reactors. This process was developed and used for Cupriavidus necator-mediated PHA production at high volumetric and specific PHA productivity (up to 2.31 g/(Lh) and 0.105 g/(gh), respectively). Based on the experimental data, formal kinetic and high structured kinetic models were established, accompanied by footprint area analysis of binary imaged cells. As a result of the study, there has been an enhanced understanding of the long-term continuous PHA production under balanced, transient, and nutrient-deficient conditions that was achieved on both the micro and the macro kinetic level. It can also be concluded that there were novel insights into the complex metabolic occurrences that developed during the multistage- continuous production of PHA as a secondary metabolite. This development was essential in paving the way for further process improvement. At the same time, a new method of specific growth rate and specific production rate based on footprint area analysis was established by using the electron microscope.

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  • 1. Khosravi-Darani K. Bucci DZ. Application of poly(hydroxyalkanoate) in food packaging: Improvements by nanotechnology. Chem Biochem Eng Q 2015; 29(2): 275-285.

  • 2. Nigmatullin R Thomas P Lukasiewicz B Puthussery H Roy I. Polyhydroxyalkanoates a family of natural polymers and their applications in drug delivery. J Chem Technol Biotechnol 2015; 90(7): 1209-1221.

  • 3. Koller M. Poly(hydroxyalkanoates) for food packaging: Application and attempts towards implementation. Appl Food Biotechnol 2014; 1(1): 3-15.

  • 4. Ong SY Sudesh K. Effects of polyhydroxyalkanoate degradation on soil microbial community. Polym Degrad Stab 2016; 131: 9-19.

  • 5. Berezina N Yada B Lefebvre R. From organic pollutants to bioplastics: insights into the bioremediation of aromatic compounds by Cupriavidus necator. New Biotechnol 2015; 32(1): 47-53.

  • 6. Jendrossek D Pfeiffer D. New insights in the formation of polyhydroxyalkanoate granules (carbonosomes) and novel functions of poly(3‐hydroxybutyrate). Environ Microbiol 2014; 16(8): 2357-2373.

  • 7. Masood F Yasin T Hameed A. Polyhydroxyalkanoates-what are the uses? Current challenges and perspectives. Crit Rev Biotechnol 2015; 35(4): 514-521.

  • 8. Obruca S Sedlacek P Mravec F Samek O Marova I. Evaluation of 3-hydroxybutyrate as an enzyme-protective agent against heating and oxidative damage and its potential role in stress response of poly(3-hydroxybutyrate) accumulating cells. Appl Microbiol Biotechnol 2016; 100(3): 1365-1376.

  • 9. Reddy CSK Ghai R Kalia V. Polyhydroxyalkanoates: an overview. Biores Technol 2003; 87(2): 137-146.

  • 10. Steinbuchel A. Perspectives for biotechnological production and utilization of biopolymers: metabolic engineering of polyhydroxyalkanoate biosynthesis pathways as a successful example. Macromol Biosci 2001; 1(1): 1-24.

  • 11. Keshavarz T Roy I. Polyhydroxyalkanoates: bioplastics with a green agenda. Curr Opin Microbiol 2010; 13(3): 321-326.

  • 12. Chen GQ Hajnal I. The ‘PHAome’. Trends Biotechnol 2015; 33(10): 559-564.

  • 13. Koller M Maršalek L Miranda de Sousa Dias M Braunegg G. Producing microbial polyhydroxyalkanoate (PHA) biopolyesters in a sustainable manner. New Biotechnol 2017; 37(A): 24-38.

  • 14. Narodoslawsky M Shazad K Kollmann R Schnitzer H. LCA of PHA Production-Identifying the Ecological Potential of Bio-plastic. Chem Biochem Eng Q 2015; 29(2): 299-305.

  • 15. Novak M Koller M Braunegg M Horvat P. Mathematical modelling as a tool for optimized PHA production. Chem Biochem Eng Q 2015; 29(2): 183-220.

  • 16. Kaur G Roy I. Strategies for large-scale production of polyhydroxyalkanoates. Chem Biochem Eng Q 2015; 29(2): 157-172.

  • 17. Haas C El-Najjar T Virgolini N Smerilli M Neureiter M. High cell-density production of poly (3-hydroxybutyrate) in a membrane bioreactor. New Biotechnol 2017; 37(A): 117-122.

  • 18. Luo HP Kemoun A Al-Dahhan MH Sevilla JF Sanchez JG Camacho FG Grima EM. Analysis of photobioreactors for culturing high-value microalgae and cyanobacteria via an advanced diagnostic technique: CARPT. Chem Eng Sci 2003; 58(12): 2519-2527.

  • 19. Dionisi D Majone M Vallini G Di Gregorio S Beccari M. Effect of the applied organic load rate on biodegradable polymer production by mixed microbial cultures in a sequencing batch reactor. Biotechnol Bioeng 2006; 93(1): 76-88.

  • 20. Koller M Muhr A. Continuous production mode as a viable process- engineering tool for efficient poly(hydroxyalkanoate) (PHA) bio-production. Chem Biochem Eng Q 2014; 28(1): 65-77.

  • 21. Koller M Braunegg G. Potential and prospects of continuous polyhydroxyalkanoate (PHA) production. Bioengineering 2015; 2(2): 94-121.

  • 22. Braunegg G Lefebvre G Renner G Zeiser A Haage G Loidl-Lanthaler K. Kinetics as a tool for polyhydroxyalkanoate production optimization. Can J Microbiol 1995: 41(13): 239-248.

  • 23. Moser A (1988) Bioprocess technology: kinetics and reactors. Springer New York

  • 24. Atlić A Koller M Scherzer D Kutschera C Grillo-Fernandes E Horvat P Chiellini E Braunegg G. Continuous production of poly([R]-3-hydroxybutyrate) by Cupriavidus necator in a multistage bioreactor cascade. Appl Microbiol Biotechnol 2001; 91(2): 295-304.

  • 25. Patnaik PR. Perspectives in the Modeling and Optimization of PHB Production by Pure and Mixed Cultures. Cit Rev Biotechnol 2005: 25(3); 153-171.

  • 26. Koller M Horvat P Hesse P Bona R Kutschera C Atlić A. Braunegg G. Assessment of formal and low structured kinetic modeling of polyhydroxyalkanoate synthesis from complex substrates. Bioproc Biosyst Eng 2006; 29(5-6): 367-377.

  • 27. Špoljarić IV Lopar M Koller M Muhr A Salerno A Reiterer A Malli K Angerer H Strohmeier K Schober S Mittelbach M. Mathematical modeling of poly[(R)-3-hydroxyalkanoate] synthesis by Cupriavidus necator DSM 545 on substrates stemming from biodiesel production. Biores Technol 2013; 133: 482-494.

  • 28. Vadlja D Koller M Novak M Braunegg G Horvat P. Footprint area analysis of binary imaged Cupriavidus necator cells to study PHB production at balanced transient and limited growth conditions in a cascade process. Appl Microbiol Biotechnol 2016; 100(23): 10065-10080.

  • 29. Horvat P Špoljarić IV Lopar M Atlić A Koller M Braunegg G. Mathematical modelling and process optimization of a continuous 5-stage bioreactor cascade for production of poly[-(R)-3-hydroxybutyrate] by Cupriavidus necator. Bioproc. Biosyst. Eng. 2013; 36(9): 1235-1250.

  • 30. Luedeking R Piret EL. A kinetic study of the lactic acid fermentation. Batch process at controlled pH. J Biochem Microbiol Technol Eng 1959; 1(4): 393-412.

  • 31. Megee III RD Drake JF Fredrickson AG Tsuchiya HM. Studies in intermicrobial symbiosis. Saccharomyces cerevisiae and Lactobacillus casei. Can J Microbiol 1972; 18(11): 1733-1742.

  • 32. Mankad T Nauman EB. Modeling of microbial growth under dual limitations. The Chem Eng J 1992; 48(2): B9-B11.

  • 33. Špoljarić IV Lopar M Koller M Muhr A Salerno A Reiterer A Horvat P. In silico optimization and low structured kinetic model of poly[(R)-3-hydroxybutyrate] synthesis by Cupriavidus necator DSM 545 by fed-batch cultivation on glycerol. J Biotechnol 2013; 168(4): 625-635.

  • 34. Lopar M Špoljarić IV Atlić A Koller M Braunegg G Horvat P. Fivestep continuous production of PHB analyzed by elementary flux modes yield space analysis and high structured metabolic model. Biochem Eng J 2013; 79 57-70.

  • 35. Lopar M Špoljarić IV Cepanec N Koller M Braunegg G Horvat P. Study of metabolic network of Cupriavidus necator DSM 545 growing on glycerol by applying elementary flux modes and yield space analysis. J Ind Microbiol Biotechnol 2014; 41(6): 913-930.

  • 36. Krzyzanek V Hrubanova K Samek O Obruca S Marova I Bernatova S Siler M Zemanek P. Cryo-SEM and Raman Spectroscopy study of the involvement of polyhydroxyalkanoates in stress response of bacteria. Microsc Microan 2015; 21(S3): 183-184.

  • 37. Mravec F Obruca S Krzyzanek V Sedlacek P Hrubanova K Samek O Kucera D Benesova P Nebesarova J. Accumulation of PHA granules in Cupriavidus necator as seen by confocal fluorescence microscopy. FEMS Microbiol Lett 2016; 363(10) fnw094.

  • 38. Wang Y Yin J Chen GQ. Polyhydroxyalkanoates challenges and opportunities. Curr Opin Biotechnol 2014; 30: 59-65.

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