Evaluation of clavulanic acid recovery techniques from Streptomyces clavuligerus cultivation broths

Yeison Agudelo-Arenas Rigoberto Ríos-Estepa David Gómez-Ríos   

Open Access   

Published:  May 23, 2024

DOI: 10.7324/JAPS.2024.178446

Clavulanic acid (CA) is a soft β-lactam antibiotic with a strong inhibitory effect on β-lactamase enzymes and it is produced by the filamentous Gram-positive bacterium Streptomyces clavuligerus (S. clavuligerus) as a secondary metabolite; the complete pathway (known as the clavam pathway) derives from the condensation of arginine and glyceraldehyde-3-phosphate, as anabolic precursors. In this work, the potential of two distinct operations for CA recovery was re-evaluated: first, the adsorption of CA through the utilization of the anion exchange resin Amberlite IRA 400, and second, the application of liquid–liquid extraction systems. For the liquid–liquid extraction, two experimental designs were developed aimed at finding the best conditions of the process. For the case of adsorption, the same experimental design, used in the previous strategy, was carried out, for comparison purposes. CA loss was minimized at 10°C and pH 2.0. The adsorption was favored by increasing the adsorbent-to-liquid ratio. Thus, the highest separation was attained in the range of 40%–45% solid/liquid ratio, adsorbing a mean value of 47.7% of the CA present in the broth. Results showed that there is still room for further improvements in CA recovery using both, adsorption and extraction, as the most advantageous techniques.

Keyword:     Streptomyces clavuligerus clavulanic acid separation processes liquid–liquid extraction adsorption bioprocess


Agudelo-Arenas Y, Ríos-Estepa R, Gómez-Ríos D. Evaluation of clavulanic acid recovery techniques from Streptomyces clavuligerus cultivation broths. J Appl Pharm Sci. 2024. Online First. http://doi.org/10.7324/JAPS.2024.178446

Copyright: © The Author(s). This is an open-access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

HTML Full Text


1. Gómez-Ríos D, Ramírez-Malule H. Bibliometric analysis of recent research on multidrug and antibiotics resistance (2017-2018). J Appl Pharm Sci. 2019;9(05):112-6. https://doi.org/10.7324/JAPS.2019.90515

2. López-Agudelo VA, Gómez-Ríos D, Ramirez-Malule H. Clavulanic acid production by Streptomyces clavuligerus: insights from systems biology, strain engineering, and downstream processing. Antibiotics. 2021;10(1):84. https://doi.org/10.3390/antibiotics10010084

3. Ramirez-Malule H, López-Agudelo VA, Gómez-Ríos D, Ochoa S, Ríos-Estepa R, Junne S, et al. TCA cycle and its relationship with Clavulanic acid production: a further interpretation by using a reduced genome-scale metabolic model of Streptomyces clavuligerus. Bioengineering. 2021;8(8):103. https://doi.org/10.3390/bioengineering8080103

4. Gómez-Ríos D, Ramírez-Malule H, Neubauer P, Junne S, Ríos-Estepa R. Data of clavulanic acid and clavulanate-imidazole stability at low temperatures. Data Brief. 2019;23:103775. https://doi.org/10.1016/j.dib.2019.103775

5. Gómez-Ríos D, Ramírez-Malule H, Neubauer P, Junne S, Ríos-Estepa R. Degradation kinetics of clavulanic acid in fermentation broths at low temperatures. Antibiotics. 2019;8(1):6. https://doi.org/10.3390/antibiotics8010006

6. Yepes J. Mejoramiento de la producción de ácido clavulánico mediante el cultivo de Streptomyces clavuligerus en fermentación extractiva, usando biorreactores operados en lote alimentado. Tesis de magister. Medellín, Colombia: Universidad de Antioquia; 2020.

7. Ser HL, Law JW, Chaiyakunapruk N, Jacob SA, Palanisamy UD, Chan KG, et al. Fermentation conditions that affect clavulanic acid production in Streptomyces clavuligerus: a systematic review. Front Microbiol. 2016;7:522. https://doi.org/10.3389/fmicb.2016.00522

8. Pinilla L, Toro LF, Avignone-Rossa C, Peñuela M, Rios-Estepa R. Streptomyces clavuligerus strain selection for clavulanic acid biosynthesis: a study based on culture composition effects and statistical analysis. Dyna. 2018;85(205):111-8. https://doi.org/10.15446/dyna.v85n205.69560

9. Gómez-Rios D, Ramírez-Malule H, Ochoa S, Ríos-Estepa R. Rational selection of culture medium for clavulanic acid production by Streptomyces Clavuligerus based on a metabolic modeling approach. Agric Nat Resour. 2022;56(2):267-76. https://doi.org/10.34044/j.anres.2022.56.2.05

10. Costa CL, Badino AC. Overproduction of clavulanic acid by extractive fermentation. Electron J Biotechnol. 2015;18:154-60. https://doi.org/10.1016/j.ejbt.2015.03.001

11. Barboza M, Almeida M, Hokka C. Influence of temperature on the kinetics of absorption and desorption of clavulanic acid by ionic exchange. Biochem Eng J. 2003;14:19-26. https://doi.org/10.1016/S1369-703X(02)00103-1

12. Abdo T, Kumar V. An overview of membrane science and technology. Nanomater Polym Membranes. 2016;1-23. https://doi.org/10.1016/B978-0-12-804703-3.00001-2

13. Abreu DC, Figueiredo KC. Bromelain separation and purification processes from pineapple extract. Braz J Chem Eng. 2019;36(2):1029-39. https://doi.org/10.1590/0104-6632.20190362s20180417

14. Mancilha M, Guimaraes G, Nardi J, Oliveira J, Hirata D. Optimization of liquid-liquid extraction step for clavulanic acid from fermentation broth using solvent mixtures. Quim Nova. 2014;37(8):1335-41. https://doi.org/10.5935/0100-4042.20140213

15. Brites L, Oliveira J, Barboza M, Hokka C. Effect of physicochemical properties of solvents on clavulanic acid extraction from fermentation broth. Lat Am Appl Res. 2012;42(1):65-70.

16. Carvalho V, Brandão JF, Brandão R, Rangel-yagui CO, Couto JA, Converti A, et al. Stability of clavulanic acid under variable pH, ionic strength and temperature conditions. A new kinetic approach. Biochem Eng J. 2009;45:89-93. https://doi.org/10.1016/j.bej.2009.02.013

17. Raven PH, Johnson GB, Mason KA, Losos JB, Singer SR. The nature of molecules and properties of water. In Biology. 10th ed. New York, NY: McGraw-Hill; 2014. pp 17-30.

18. Reece JB, Urry LA, Cain ML, Wasserman SA, Minorsky PV, Jackson RB. Water and life. In Campbell biology. 10th edition. San Francisco, CA: Pearson; 2011. pp 44-54.

19. Bersanetti PA, Almeida RM, Barboza M, Araújo ML, Hokka CO. Kinetic studies on clavulanic acid degradation. Biochem Eng J. 2005;23(1):31-6. https://doi.org/10.1016/j.bej.2004.10.007

20. Mayer AF, Deckwer WD. Simultaneous production and decomposition of clavulanic acid during Streptomyces clavuligerus cultivations. Appl Microbiol Biotechnol. 1996;45(2):41-6. https://doi.org/10.1007/s002530050646

21. Forte MB, Luna EC, Pastore HO, Rodrigues MI, Filho FM. Evaluation of clavulanic acid adsorption in MgAl-layered double hydroxide: kinetic, equilibrium and thermodynamic studies. Adsorp Sci Technol. 2012; 30(1):65-80. https://doi.org/10.1260/0263-6174.30.1.65

22. Hirata DB, Oliveira JH, Leão KV, Rodrigues MI, Ferreira AG, Giulietti M, et al. Precipitation of clavulanic acid from fermentation broth with potassium 2-ethyl hexanoate salt. Sep Purif Technol. 2009;66(3):598-605. https://doi.org/10.1016/j.seppur.2009.01.010

23. Martino M. Tensoactivos. CMC a temperaturas bajas o moderadas. Influencia sobre la CMC de la presencia de compuestos neutros o polares: alanina. Trabajo de grado, España: Universidad de la Laguna; 2020.

24. Luz DA, Rodrigues AK, Silva FR, Torres AE, Cavalcante CL, Brito ES, et al. Adsorptive separation of fructose and glucose from an agroindustrial waste of cashew industry. Bioresour Technol. 2008;99:2455-65. https://doi.org/10.1016/j.biortech.2007.04.063

25. Barboza M, Almeida R, Hokka C. Kinetic studies of clavulanic acid recovery by ion exchange chromatography. Bioseparation. 2002;10(4-5):221-7. https://doi.org/10.1023/A:1016365827265

26. Seader JD, Henley J, Roper DK. Separation process principles, chemical and biochemical operations. New York, NY: John Wiley & Sons; 2011.

27. Barboza M, Silva C, Cuel M, Barreto V, Hokka CO. Separation of clavulanic acid from fermented broth of amino acids by an aqueous two-phase system and ion exchange adsorption. N Biotechnol. 2012;29(3):428-31. https://doi.org/10.1016/j.nbt.2011.05.012

28. López VA, Gómez-Ríos D, Ramirez-Malule H. Clavulanic acid production by Streptomyces clavuligerus. Antibiotics. 2021;10(1):84. https://doi.org/10.3390/antibiotics10010084

29. da Silva KC, Souza AT, Badino AC, Pedrolli DB, Cerri MO. Screening of medium constituents for clavulanic acid production by Streptomyces clavuligerus. Braz J Microbiol. 2018;49(4):832-9. https://doi.org/10.1016/j.bjm.2018.01.006

30. Saudagar PS, Survase SA, Singhal RS. Clavulanic acid: a review. Biotechnol Adv. 2008;26:335-51. https://doi.org/10.1016/j.biotechadv.2008.03.002

31. Almeida R, Barboza M, Hokka C. Continuous clavulanic acid adsorption process. Appl Biochem Biotechnol. 2003; 108(1-3):867-80. https://doi.org/10.1385/ABAB:108:1-3:867

32. da Silva CS, Cuel MF, Barreto VO, Kwong WH, Hokka CO, Barboza M. Separation of clavulanic acid from fermented broth of amino acids by an aqueous two-phase system and ion-exchange adsorption. N Biotechnol. 2012;29(3):428-31. https://doi.org/10.1016/j.nbt.2011.05.012

33. Scott MJ. Principles of ion exchange technology. Amsterdam, Netherlands: Elsevier Science; 2003.

34. Chen G, Liu Y, Cao Y. Recovery of chromate ions from industrial wastewater using ion exchange resins: a review. J Ind Eng Chem. 2018; 66:1-13.

35. Gutiérrez H, Salazar R. Elementos de inferencia estadística: experimentos con uno y dos tratamientos. In Análisis y Diseño de Experimentos. 2th ed. New York, NY: McGraw-Hill; 2008.

Article Metrics
129 Views 16 Downloads 145 Total



Related Search

By author names