Development and characterization of a novell-asparaginase/MWCNT nanobioconjugate


The enzymel-asparaginase (ASNase) presents effective antineoplastic properties used for acute lymphoblastic leukemia treatment besides their potential use in the food sector to decrease the acrylamide formation. Considering their applications, the improvement of this enzyme's properties by efficient immobilization techniques is in high demand. Carbon nanotubes are promising enzyme immobilization supports, since these materials have increased surface area and effective capacity for enzyme loading. Accordingly, in this study, multi-walled carbon nanotubes (MWCNTs) were explored as novel supports for ASNase immobilization by a simple adsorption method. The effect of pH and contact time of immobilization, as well as the ASNase to nanoparticles mass ratio, were optimized according to the enzyme immobilization yield and relative recovered activity. The enzyme-MWCNTs bioconjugation was confirmed by thermogravimetric analysis (TGA), Fourier transform infrared spectroscopy (FTIR), Raman and transmission electron microscopy (TEM) studies. MWCNTs have a high ASNase loading capacity, with a maximum immobilization yield of 90%. The adsorbed ASNase retains 90% of the initial enzyme activity at the optimized conditions (pH 8.0, 60 min, and 1.5 x 10(-3)g mL(-1)of ASNase). According to these results, ASNase immobilized onto MWCNTs can find improved applications in several areas, namely biosensors, medicine and food industry.



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Cristovao, RO; Almeida, MR; Barros, MA; Nunes, JCF; Boaventura, RAR; Loureiro, JM; Faria, JL; Neves, MC; Freire, MG; Ebinuma-Santos, VC; Tavares, APM; Silva, CG

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This work was financially supported by Base Funding - UIDB/EQU/50020/2020 of the Associate Laboratory LSRE-LCM - funded by national funds through FCT/MCTES (PIDDAC), and POCI-01-0145-FEDER-031268 - funded by FEDER, through COMPETE2020 - Programa Operacional Competitividade e InternacionalizacAo (POCI), and by national funds (OE), through FCT/MCTES. This work was developed within the scope of the project CICECO-Aveiro Institute of Materials, UIDB/50011/2020 & UIDP/50011/2020, financed by national funds through the Portuguese Foundation for Science and Technology/MCTES. Ana P. M. Tavares and Claudia G. Silva acknowledge the FCT Investigator Programme and Exploratory Project (IF/01634/2015 and IF/00514/2014, respectively) with financing from the European Social Fund and the Human Potential Operational Programme. Valeria C. Santos-Ebinuma acknowledges FAPESP (2018/06908-8). Raquel O. CristovAo acknowledges FCT funding under DL57/2016 Transitory Norm Programme. JoAo C. F. Nunes acknowledges FCT for the PhD fellowship (SFRH/BD/150671/2020).

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