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Entrapment of Aspergillus niger MYA 135 mycelium for recycles in biocatalysis Erika L. Regner1, Hebe N. Salvatierra1, Mario D. Baigorí1, 2 and Licia M. Pera1 1 Laboratorio de Morfogénesis y Fermentaciones (PROIMI - CONICET)- 2 Universidad Nacional de Tucumán. E-mail: regnererika@gmail.com .Keywords: Alginate Beads; Mycelium-bound Lipase; Plackett-Burman design INTRODUCTION Entrapment in alginate beads is a technique that is widely used to improve operational stability and easy manipulation of biocatalyst applied in industrial processes [1, 2]. The aim of this work was to analyze the feasibility of reuse of the entrapped Aspergillus niger MYA 135 mycelium-bound lipase (Mb) in alginate beads. The beads components were evaluated through the Plackett?Burman (PB) statistical experimental design as screening experiments tool. RESULTS AND DISCUSSION Plackett-Burman statistical method was performed to identify the factors contributing to the feasibility of biocatalyst reutilization. The effect of ten factors such as: Alginate (0.5-1%), polyethylene glycol (0-0.25%), glycerol (0-0.25%), glutaraldehyde (0-1%), biomass concentration (0.01-0.02 g/ml), cross linking time (30-60 minutes), MgCl2 (0-0.01 M), CaCl2 (0.05-0.1 M), BaSO4 (0-0.01 M) and SrCl2 (0-0.01 M) in two levels (-1 and +1) was examined. In these experiments, all analyzed variables had a significant effect on the hydrolytic activity using p-nitrophenyl palmitate as substrate after five reactions cycles. The effect of each variable on the biocatalyst recycles was represented in Figure 1. It can be clearly seen that the high level of alginate showed a strong influence on the biocatalyst durability. Additionally, the presence of divalent cations such as magnesium and barium had an important effect on the response studied. It has been reported an increase in the stability and a swelling reduction of the alginate beads in presence of barium ions. Although it has been described the efficient generation of stable beads using strontium as cross linking agent [3], in this study, the strontium ion has a negative influence on the biocatalyst reuse. Furthermore, the high level of calcium had a positive effect on the biocatalyst recycle. In this sense, it is well known that a decrease in the swelling rate constant is associated with an elevated calcium concentration, thereby increasing the stability of the beads [4]. Finally, it was found that a cross linking time of 30 minutes (low level) was adequate for this approach. Figure 1. Main effects of the studied factors on the Mb hydrolytic activity from A. niger MYA 135 after five cycles based on PB experimental results. CONCLUSION The statistical approach employed in this study allows for rapid identification of important variables affecting the biocatalyst reutilization of a Mb hydrolytic activity from A. niger 135. Finally, further statistical optimization using response surface methodology needs to be conducted. ACKNOWLEDGEMENTS This work was supported by grant PICT 2011?2158 (FONCyT) y PIP 297 (CONICET) REFERENCES [1] Smidsrød O.; Skjåk-Bræk, G. Alginate as immobilization matrix for cells. Tibtech 1990, 8, 71-78. http://www.sciencedirect.com/science/article/pii/016777999090139O. [2] Peart, P. C.; Chen, A. R. M.; Reynolds, W. F.; Reese, P. B. Entrapment of mycelial fragments in calcium alginate: A general technique for the use of immobilized filamentous fungi in biocatalysis. Steroids 2012, 77, 85-90. http://www.sciencedirect.com/science/article/pii/S0039128X11003126. [3] Mørch,Ý. A.; Donati, I.; Strand, B. L.; Skjåk-Bræk, G. Effect of Ca2+, Ba2+, and Sr2+ on Alginate Microbeads. Biomacromolecules, 2006, 7, 1471?1480. http://pubs.acs.org/doi/abs/10.1021/bm060010d. [4] Davidovich-Pinhas, M.; Bianco-Peled, H. A quantitative analysis of alginate swelling. Carbohydrate Polymers 2010, 79, 1020?1027. http://www.sciencedirect.com/science/article/pii/S0144861709005906