BIOPROCESSING AND ENGINEERING New Biotechnology · Volume 31S · July 2014

ylation of androst-4-ene-3,17-dione (AD). AD in turn is producedprimarily with microbial biotransformation of natural sterols bysome actinobacteria.

The aim of this work was to develop a bioprocess for obtainingof 11�-hydroxyandrost-4-ene-3,17-dione from phytosterol.

Specific biochemical activities of two microbial strains wereused as a basis for the two-stage bioprocess. On the first stage phy-tosterol was converted to AD by Mycobacterium sp. NRRL 3805B.The conditions of biotransformation were optimized to get approx.70% molar yield of AD from 12 g/l of the substrate. On the secondstage AD accumulated in the biotransformation broth was regio-and stereo-specifically hydroxylated into C-11-alpha position bythe mycelial fungus Aspergillus ochraceus VKM F-830Y. Cultivationconditions for preparation of the fungal biocatalyst with higherspecific activity and mode of the biocatalyst application were opti-mized.

Both biotransformations were carried out in a single laboratory-scale bioreactor thus allowing exclude AD isolation andpurification procedures. The two-stage bioprocess provided 65-68% molar yield of 11�HAD from phytosterol for 65-72 h. Theproduct was separated and purified to 95% by step-wise crystalliza-tion and re-crystallizations from a system of polar organic solvents.

For our knowledge, microbial production of 11�-HAD fromphytosterol was not so far reported.

http://dx.doi.org/10.1016/j.nbt.2014.05.1903

PF-16

Plasmid DNA purification by integrating membranetechnology with arginine affinity chromatography

João Queiroz ∗ , Catherine Nunes, Ângela Sousa, José Nunes,António Morão, Fani Sousa

University of Beira Interior

The implementation of clarification and purification processesto isolate the supercoiled (sc) plasmid isoform at industrial scalebecomes crucial. In the present study, membrane filtration tech-nology was performed to isolate and clarify the sc plasmid DNA(pDNA) from lysates. Microfiltration process was implemented toeliminate the suspended solids and to perform a diafiltration of thesolution, followed by an ultrafiltration technique to concentratethe plasmid and to remove the different types of RNA [1]. Finally,a suitable chromatographic strategy is essential to remove residualimpurities and to obtain the sc pDNA as a highly pure prod-uct. Affinity chromatography with amino acids as ligands, suchas arginine, has been employed for this objective due to its highselectivity for the sc isoform and also because of the mild elutionconditions required to achieve its purification [2]. Thereby, thesample resultant from the ultrafiltration process was applied in thearginine chromatographic matrix, to attain an adequate strategyfor sc pDNA purification. The nature of the arginine support andthe use of moderate salt concentrations render this operation moreeconomically sustainable and viable to be used in large scale sys-tems. The separation of sc isoform was proved by electrophoreticand HPLC analysis. Overall, the integration of membrane technol-

ogy with affinity chromatography to efficiently purify the pDNAresults in a powerful tool for industrial manufacturing.

References

[1].Nunes JC, et al. Journal of Membrane Science 2012;415-416:24–35.[2].Sousa A, et al. Journal of Separation Science 2009;32:1665–72.

http://dx.doi.org/10.1016/j.nbt.2014.05.1904

PF-17

Establishment of Efficient Microalgal Harvesting Tech-nique by the Concomitant Application of Red or BlueLight Wavelength with Chitosan

Dae Geun Kim1,∗ , Yoon-E. Choi2

1 Department of Bioprocess Engineering, Chonbuk National University2 LED Agri-bio Fusion Technology Research Center, Chonbuk National Univer-sity

Microalgae are considered to be one of the most promisingfeedstocks for biodiesel, due to their rapid growth and high lipidcontent. However, microalgal harvesting is one of indispensablestep claiming almost 20-30% of total biomass production cost. Chi-tosan is a natural, non-toxic, polycationic polymer with multipleapplications in pharmaceuticals, food, agricultural, and chemicalindustries. There are multiple lines of reports that chitosan can alsobe a promising alternative flocculants for microalgal harvesting. Inour previous study, light-emitting diodes(LEDs) especially red andblue color were demonstrated to govern the specific microalgalcell biology. Blue light illumination led to significantly increasedcell size, whereas red light resulted in small-sized cell with activedivisions. Based on that, in this study, we attempted to establish anovel harvesting strategy of microalgal biomass by the concomi-tant applications of both blue light illumination and chitosan. Wesuccessfully proved that microalgal cells cultivated under blue lightsettled more rapidly than those of biomass cultivated under redlight illumination. Next, the combinationary effects of differentlight wavelengths and chitosan concentration were thoroughlytested. The data suggested that the optimal harvest conditionunder the blue light with chitosan were significantly deviated fromthose under the red light with chitosan. Our strategy, based onthe illumination of blue light in conjunction with chitosan, willcontribute to setting up the future biomass harvest process usingmicroalgae.

http://dx.doi.org/10.1016/j.nbt.2014.05.1905

PF-18

Optimization of an industrial primary protein recoveryprocess by Design of Experiment

Tanja Buch ∗ , Ian Marison

Dublin City University

The primary recovery of proteins at the end of a fermentationprocess can be considered as a keystone for the overall protein lossduring purification. Recombinant proteins are widely produced in

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