PubMed · 13133788
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B BOGDANIKOWA, T BOGDANIK. 1953-11-09. [Certain so-called flocculation reaction].. https://pubmed.ncbi.nlm.nih.gov/13133788/
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Biomass separation represents a critical bottleneck in Komagataella phaffii-based biopharmaceutical processes, as typically high cell densities of 40 - 50 % create significant operational, technical and economic challenges for harvest operations. Yeast cell aggregation (flocculation) provides a solution to accelerate cell sedimentation by increasing particle size, thus allowing to improve biomass-supernatant separation efficiency during both natural gravity settling and (continuous) centrifugation operations. This study demonstrates successful engineering of K. phaffii strains with an inducible flocculation phenotype using CRISPR/Cas9-based genome editing to integrate the Saccharomyces cerevisiae FLO1 (ScFLO1) gene under control of various regulatory elements, including methanol-inducible and derepressible promoters. Flocculation strength could be enhanced by implementing transcriptional positive feedback circuits based on the methanol-inducible AOX1 promoter. To address methanol-free production requirements, we developed alternative systems to retrofit PAOX1-based ScFLO1 expression and exploited the derepressible PDF promoter, offering broader compatibility with biopharmaceutical manufacturing facilities. Flocculating cells cultivated in a bioreactor demonstrated significantly improved sedimentation behavior, with considerably lower supernatant turbidity after short low-speed centrifugation or gravity sedimentation compared to non-flocculating controls. Crucially, cell flocculation had no negative impact on product amount and quality when expressing a multivalent NANOBODY® VHH molecule with pharmaceutical relevance. Thus, this work establishes the first genetically engineered flocculation system in K. phaffii compatible with recombinant protein production, providing the basis for an innovative approach to streamline harvest operations in biopharmaceutical processes.
Coagulation-flocculation is a proven technique for the treatment of high suspended solids wastewater. In this study, the central composite face-centered design (CCFD) and response surface methodology (RSM) have been applied to optimize two most important operating variables: coagulant dosage and pH, in the coagulation-flocculation process of pulp and paper mill wastewater treatment. The treated wastewater with high total suspended solids (TSS) removal, low SVI (sludge volume index) and high water recovery are the main objectives to be achieved through the coagulation-flocculation process. The effect of interactions between coagulant dosage and pH on the TSS removal and SVI are significant, whereas there is no interaction between coagulant dosage and water recovery. Quadratic models have been developed for the response variables, i.e. TSS removal, SVI and water recovery based on the high coefficient of determination (R(2)) value of >0.99 obtained from the analysis of variances (ANOVA). The optimum conditions for coagulant dosage and pH are 1045mgL(-1) and 6.75, respectively, where 99% of TSS removal, SVI of 37mLg(-1) and 82% of water recovery can be obtained.
Moringa oleifera seeds, an environmental friendly and natural coagulant are reported for the pretreatment of palm oil mill effluent (POME). In coagulation-flocculation process, the M. oleifera seeds after oil extraction (MOAE) are an effective coagulant with the removal of 95% suspended solids and 52.2% reduction in the chemical oxygen demand (COD). The combination of MOAE with flocculant (NALCO 7751), the suspended solids removal increased to 99.3% and COD reduction was 52.5%. The coagulation-flocculation process at the temperature of 30 degrees C resulted in better suspended solids removal and COD reduction compared to the temperature of 40, 55 and 70 degrees C. The MOAE combined with flocculant (NALCO 7751) reduced the sludge volume index (SVI) to 210mL/g with higher recovery of dry mass of sludge (87.25%) and water (50.3%).