Process cost and facility considerations in the selection of primary cell culture clarification technology

The bioreactor volume delineating the selection of primary clarification technology is not always easily defined. Development of a commercial scale process for the manufacture of therapeutic proteins requires scale‐up from a few liters to thousands of liters. While the separation techniques used for protein purification are largely conserved across scales, the separation techniques for primary cell culture clarification vary with scale. Process models were developed to compare monoclonal antibody production costs using two cell culture clarification technologies. One process model was created for cell culture clarification by disc stack centrifugation with depth filtration. A second process model was created for clarification by multi‐stage depth filtration. Analyses were performed to examine the influence of bioreactor volume, product titer, depth filter capacity, and facility utilization on overall operating costs. At bioreactor volumes <1,000 L, clarification using multi‐stage depth filtration offers cost savings compared to clarification using centrifugation. For bioreactor volumes >5,000 L, clarification using centrifugation followed by depth filtration offers significant cost savings. For bioreactor volumes of ～2,000 L, clarification costs are similar between depth filtration and centrifugation. At this scale, factors including facility utilization, available capital, ease of process development, implementation timelines, and process performance characterization play an important role in clarification technology selection. In the case study presented, a multi‐product facility selected multi‐stage depth filtration for cell culture clarification at the 500 and 2,000 L scales of operation. Facility implementation timelines, process development activities, equipment commissioning and validation, scale‐up effects, and process robustness are examined. © 2013 The Authors. American Institute of Chemical Engineers Biotechnol. Prog., 29:1239–1245, 2013     

Economic impact of combined torrefaction and pelletization processes on forestry biomass supply

The cost of supplying wood biomass from forestry operations in remote areas has been an obstacle to expansion of forest‐based bioenergy in much of the western United States. Economies of scale in the production of liquid fuels from lignocellulosic biomass feedstocks favor large centralized biorefineries. Increasing transportation efficiency through torrefaction and pelletization at distributed satellite facilities may serve as a means to expand the utilization of forestry residuals in biofuel production. To investigate this potential, a mixed‐integer linear program was developed to optimize the feedstock supply chain design with and without distributed pretreatment. The model uses techno‐economic assessment of scale‐dependent biomass pretreatment processes from existing literature and multimodal biomass transportation cost evaluations derived from a spatially explicit network analysis as input. In addition, the sensitivity of the optimal system configuration was determined for variations of key input parameters including the production scale of pretreatment facilities, road and rail transportation costs, and feedstock procurement costs. Torrefaction and densification were found to reduce transportation costs by $0.84 per GJ and overall delivered costs by $0.24 per GJ, representing 14.5% and 5.2% cost reductions compared to feedstock collection without pretreatment. Significant uncertainties remain in terms of the costs associated with deploying torrefaction equipment at the scales modeled, but the level of potential cost savings suggests further analysis and development of these alternatives.     

Explaining the reductions in Brazilian sugarcane ethanol production costs: importance of technological change

Over the period 1975–2010, unit production costs of sugarcane ethanol in Brazil declined by 67%, while the per‐unit processing costs decreased by more than 70%. This article examines the role of various factors that lead to these cost reductions, including learning‐by‐doing (LBD), economies of scale, rising input prices, market competitiveness, and exogenous technological changes. Using the aggregate industry‐level data, we show that the traditional experience curve approach will lead to biased estimate of the learning effect when economies of scale, rising input prices, market competitiveness, and exogenous technological changes are excluded as explanatory variables in explaining these cost reductions. With the inclusion of these variables and LBD, we find that the reductions in production/processing costs of Brazilian sugarcane ethanol were primarily driven by autonomous technological changes and unrelated to LBD. Economies of scale, market competitiveness, and rising input prices had insignificant impacts on the reductions in production/processing costs of sugarcane ethanol over the sample period.     

A framework for the identification of promising bio‐based chemicals

Recent progress in metabolic engineering and synthetic biology enables the use of microorganisms for the production of chemicals—“bio‐based chemicals.” However, it is still unclear which chemicals have the highest economic prospect. To this end, we develop a framework for the identification of such promising ones. Specifically, we first develop a genome‐scale constraint‐based metabolic modeling approach, which is used to identify a candidate pool of 209 chemicals (together with the estimated yield, productivity, and residence time for each) from the intersection of the high‐production‐volume chemicals and the KEGG and MetaCyc databases. Second, we design three screening criteria based on a chemical’s profit margin, market volume, and market size. The total process cost, including the downstream separation cost, is systematically incorporated into the evaluation. Third, given the three aforementioned criteria, we identify 32 products as economically promising if the maximum yields can be achieved, and 22 products if the maximum productivities can be achieved. The breakeven titer that renders zero profit margin for each product is also presented. Comparisons between extracellular and intracellular production, as well as Escherichia coli and Saccharomyces cerevisiae systems are also discussed. The proposed framework provides important guidance for future studies in the production of bio‐based chemicals. It is also flexible in that the databases, yield estimations, and criteria can be modified to customize the screening.     

Utilization of agricultural residues for poly(3-hydroxybutyrate) production by Halomonas boliviensis LC1

Aims: Utilization of cheap and readily available agricultural residues as cheap carbon sources for poly(3‐hydroxybutyrate) (PHB) production by Halomonas boliviensis. Methods and Results: Wheat bran was hydrolysed by a crude enzyme preparation from Aspergillus oryzae NM1 to provide a mixture of reducing sugars composed mainly of glucose, mannose, xylose and arabinose. Growth of H. boliviensis using a mixture of glucose (0·75% w/v) and xylose (0·25% w/v) in the medium led to a PHB content and concentration of 45 wt% and 1 g l^?1, respectively, after 30 h. A similar PHB concentration was attained when H. boliviensis was grown on wheat bran hydrolysate but with a lower PHB content, 34 wt%. In a batch cultivation mode in a fermentor, using 1·8% (w/v) reducing sugars, the maximum PHB accumulation by H. boliviensis was attained in 20 h, but was reduced to about 30 wt%. By adding butyric acid (0·8% v/v), sodium acetate (0·8% w/v) and decreasing the reducing sugars concentration to 1·0% w/v in the medium, PHB accumulation and concentration were increased to 50 wt% and 4 g l^?1, respectively, after 20 h. Butyric acid and sodium acetate for PHB production could also be provided by anaerobic digestion of solid potato waste. Conclusions: Cheap and readily available agricultural residues can be used as substrates to produce PHB. The production of PHB by H. boliviensis using wheat bran hydrolysate as source of carbon is expected to reduce the production cost and motivates further studies. Significance and Impact of the Study: Large‐scale commercial utilization of PHB is mainly hampered by its high production cost. Carbon source for PHB production accounts up to 50% of the total production costs. Thus, the use of waste agricultural residues can substantially reduce the substrate cost (and in turn even provide value to the waste), and can downsize the production costs. This improves the market competitiveness. Studies on PHB production by moderate halophiles were recently initiated with H. boliviensis and findings show that it has potential for commercial exploitation. PHB production by H. boliviensis using wheat bran and potato waste is hence interesting.     

Economic analysis of pilot‐scale production of B‐phycoerythrin

β‐Phycoerythrin is a color protein with several applications, from food coloring to molecular labeling. Depending on the application, different purity is required, affecting production cost and price. Different production and purification strategies for B‐phycoerythrin have been developed, the most studied are based on the production using Porphyridium cruentum and purified using chromatographic techniques or aqueous two‐phase systems. The use of the latter can result in a less expensive and intensive recovery of the protein, but there is lack of a proper economic analysis to study the effect of using aqueous two‐phase systems in a scaled‐up process. This study analyzed the production of B‐Phycoerythrin using real data obtained during the scale‐up of a bioprocess using specialized software (BioSolve, Biopharm Services, UK). First, a sensitivity analysis was performed to identify critical parameters for the production cost, then a Monte Carlo analysis to emulate real processes by adding uncertainty to the identified parameters. Next, the bioprocess was analyzed to determine its financial attractiveness and possible optimization strategies were tested and discussed. Results show that aqueous two‐phase systems retain their advantages of low cost and intensive recovery (54.56%); the costs of production per gram calculated (before titer optimization: US$15,709 and after optimization: US$2,374) allowed to obtain profit (in the range of US$millions in a 10‐year period) for a potential company taking this production method by comparing the production cost against commercial prices. The bioprocess analyzed is a promising and profitable method for the generation of a highly purified B‐phycoerythrin. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1472–1479, 2016     

The Production of Pyrethrins by Plant Cell and Tissue Cultures of Chrysanthemum cinerariaefolium and Tagetes Species

Pyrethrins, the most economically important natural insecticide, comprise a group of six closely related monoterpene esters. The industrial production is based on their extraction from Chrysanthemum cinerariaefolium (Pyrethrum) capitula. The world production of natural pyrethrins still falls short of global market demand stimulating the research in in vitro production as an alternative to conventional cultivation methods. The different biotechnological alternatives such as callus cultures, shoot and root cultures, plant cell suspension cultures, and bioconversion of precursors by means of enzymatic synthesis or genetically engineered microorganisms, as well as the progress achieved in methods for the identification and quantitation of insecticidal compounds have been reviewed. Although technology for plant cell culture exists, industrial applications have, to date, been limited due to both the low economical viability and technological feasibility at large scale. Bioconversion of readily available precursors looks more attractive, but more research is needed before this technology is used for the industrial production of pyrethrins.     

The Production of Pyrethrins by Plant Cell and Tissue Cultures of Chrysanthemum cinerariaefolium and Tagetes Species

Pyrethrins, the most economically important natural insecticide, comprise a group of six closely related monoterpene esters. The industrial production is based on their extraction from Chrysanthemum cinerariaefolium (Pyrethrum) capitula. The world production of natural pyrethrins still falls short of global market demand stimulating the research in in vitro production as an alternative to conventional cultivation methods. The different biotechnological alternatives such as callus cultures, shoot and root cultures, plant cell suspension cultures, and bioconversion of precursors by means of enzymatic synthesis or genetically engineered microorganisms, as well as the progress achieved in methods for the identification and quantitation of insecticidal compounds have been reviewed. Although technology for plant cell culture exists, industrial applications have, to date, been limited due to both the low economical viability and technological feasibility at large scale. Bioconversion of readily available precursors looks more attractive, but more research is needed before this technology is used for the industrial production of pyrethrins.     

Molecular characterization of podoviral bacteriophages virulent for Clostridium perfringens and their comparison with members of the Picovirinae

Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium responsible for human food-borne disease as well as non-food-borne human, animal and poultry diseases. Because bacteriophages or their gene products could be applied to control bacterial diseases in a species-specific manner, they are potential important alternatives to antibiotics. Consequently, poultry intestinal material, soil, sewage and poultry processing drainage water were screened for virulent bacteriophages that lysed C. perfringens. Two bacteriophages, designated ΦCPV4 and ΦZP2, were isolated in the Moscow Region of the Russian Federation while another closely related virus, named ΦCP7R, was isolated in the southeastern USA. The viruses were identified as members of the order Caudovirales in the family Podoviridae with short, non-contractile tails of the C1 morphotype. The genomes of the three bacteriophages were 17.972, 18.078 and 18.397 kbp respectively; encoding twenty-six to twenty-eight ORF's with inverted terminal repeats and an average GC content of 34.6%. Structural proteins identified by mass spectrometry in the purified ΦCP7R virion included a pre-neck/appendage with putative lyase activity, major head, tail, connector/upper collar, lower collar and a structural protein with putative lysozyme-peptidase activity. All three podoviral bacteriophage genomes encoded a predicted N-acetylmuramoyl-L-alanine amidase and a putative stage V sporulation protein. Each putative amidase contained a predicted bacterial SH3 domain at the C-terminal end of the protein, presumably involved with binding the C. perfringens cell wall. The predicted DNA polymerase type B protein sequences were closely related to other members of the Podoviridae including Bacillus phage Φ29. Whole-genome comparisons supported this relationship, but also indicated that the Russian and USA viruses may be unique members of the sub-family Picovirinae.     

Life Cycle Energy Assessment of Alternative Water Supply Systems (9 pp)

Goal, Scope and Background This paper discusses the merging of methodological aspects of two known methods into a hybrid on an application basis. Water shortages are imminent due to scarce supply and increasing demand in many parts of the world. In California, this is caused primarily by population growth. As readily available water is depleted, alternatives that may have larger energy and resource requirements and, therefore, environmental impacts must be considered. In order to develop a more environmentally responsible and sustainable water supply system, these environmental implications should be incorporated into planning decisions. Methods Comprehensive accounting for environmental effects requires life cycle assessment (LCA), a systematic account of resource use and environmental emissions caused by extracting raw materials, manufacturing, constructing, operating, maintaining, and decommissioning the water infrastructure. In this study, a hybrid LCA approach, combining elements of process-based and economic input-output-based LCA was used to compare three supply alternatives: importing, recycling, and desalinating water. For all three options, energy use and air emissions associated with energy generation, vehicle and equipment operation, and material production were quantified for life-cycle phases and water supply functions (supply, treatment, and distribution). The Water-Energy Sustainability Tool was developed to inform water planning decisions. It was used to evaluate the systems of a Northern and a Southern California water utility. Results and Discussion The results showed that for the two case study utilities desalination had 2–5 times larger energy demand and caused 2–18 times more emissions than importation or recycling, due primarily to the energy-intensity of the treatment process. The operation life-cycle phase created the most energy consumption with 56% to 90% for all sources and case studies. For each water source, a different life-cycle phase dominated energy consumption. For imported water, supply contributed 56% and 86% of the results for each case study; for desalination, treatment accounted for approximately 85%; for recycled water, distribution dominated with 61% and 74% of energy use. The study calculated external costs of air pollution from all three water supply systems. These costs are borne by society, but not paid by producers. The external costs were found to be 6% of desalinated water production costs for both case studies, 8% of imported water production costs in Southern California, and 1–2% for the recycled water systems and for the Northern California utility's imported water system. Conclusion Recycling water was found to be more energy intensive in Northern than in Southern California, but the results for imported water were similar. While the energy demand of water recycling was found to be larger than importation in Northern California, the two alternatives were competitive in Southern California. For all alternatives in both case studies, the energy consumed by system operation dominated the results, but maintenance was also found to be significant. Energy production was found to be the largest contributor in all water provision systems, followed by materials production. The assessment of external costs revealed that the environmental effects of energy and air emissions caused by infrastructure is measurable, and in some cases, significant relative to the economic cost of water. Recommendation and Perspective This paper advocates the necessity of LCA in water planning, and discusses the applicability of the described model to water utilities.     

Resource supply and the evolution of public-goods cooperation in bacteria

Background  

Explaining public-goods cooperation is a challenge for evolutionary biology. However, cooperation is expected to more readily evolve if it imposes a smaller cost. Such costs of cooperation are expected to decline with increasing resource supply, an ecological parameter that varies widely in nature. We experimentally tested the effect of resource supply on the evolution of cooperation using two well-studied bacterial public-good traits: biofilm formation by Pseudomonas fluorescens and siderophore production by Pseudomonas aeruginosa.     

Metabolic engineering of Clostridium acetobutylicum for the enhanced production of isopropanol‐butanol‐ethanol fuel mixture

Butanol is considered as a superior biofuel, which is conventionally produced by clostridial acetone‐butanol‐ethanol (ABE) fermentation. Among ABE, only butanol and ethanol can be used as fuel alternatives. Coproduction of acetone thus causes lower yield of fuel alcohols. Thus, this study aimed at developing an improved Clostridium acetobutylicum strain possessing enhanced fuel alcohol production capability. For this, we previously developed a hyper ABE producing BKM19 strain was further engineered to convert acetone into isopropanol. The BKM19 strain was transformed with the plasmid pIPA100 containing the sadh (primary/secondary alcohol dehydrogenase) and hydG (putative electron transfer protein) genes from the Clostridium beijerinckii NRRL B593 cloned under the control of the thiolase promoter. The resulting BKM19 (pIPA100) strain produced 27.9 g/l isopropanol‐butanol‐ethanol (IBE) as a fuel alcohols with negligible amount of acetone (0.4 g/l) from 97.8 g/l glucose in lab‐scale (2 l) batch fermentation. Thus, this metabolically engineered strain was able to produce 99% of total solvent produced as fuel alcohols. The scalability and stability of BKM19 (pIPA100) were evaluated at 200 l pilot‐scale fermentation, which showed that the fuel alcohol yield could be improved to 0.37 g/g as compared to 0.29 g/g obtained at lab‐scale fermentation, while attaining a similar titer. To the best of our knowledge, this is the highest titer of IBE achieved and the first report on the large scale fermentation of C. acetobutylicum for IBE production. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1083–1088, 2013     

Considerations for the process development of insect‐derived antimicrobial peptide production

Antimicrobial peptides (AMPs) could evolve into new therapeutic lead molecules against multi‐resistant bacteria. As insects are a rich source of AMP, the identification and characterization of insect‐derived AMPs is particularly emphasized. One challenge of bringing these molecules into market, e.g., as a drug, is to develop a cost‐efficient large‐scale production process. Due to the fact that a direct AMP isolation from insects is not economical and that chemical synthesis is recommended for peptide sizes below 40 amino acids, a viable option is heterologous AMP production. Therefore, previous knowledge concerning the expression of larger proteins can be adapted, but due to the AMP nature (e.g., small size, bactericide) additional challenges have to be faced during up and downstream processing. Nonetheless the bottleneck for large‐scale AMP production is the same as for proteins; mainly the downstream process. This review introduces opportunities for insect‐derived AMP production, like the choice of the expression system (based on previously derived data), depending on the AMP nature, as well as new purification strategies like elastin‐like peptide/intein based purification strategies. All of these aspects are discussed with regard to large‐scale processes and costs. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:1–11, 2015     

Xanthophyllomyces dendrorhous for the industrial production of astaxanthin

Astaxanthin is a red xanthophyll (oxygenated carotenoid) with large importance in the aquaculture, pharmaceutical, and food industries. The green alga Haematococcus pluvialis and the heterobasidiomycetous yeast Xanthophyllomyces dendrorhous are currently known as the main microorganisms useful for astaxanthin production at the industrial scale. The improvement of astaxanthin titer by microbial fermentation is a requirement to be competitive with the synthetic manufacture by chemical procedures, which at present is the major source in the market. In this review, we show how the isolation of new strains of X. dendrorhous from the environment, the selection of mutants by the classical methods of random mutation and screening, and the rational metabolic engineering, have provided improved strains with higher astaxanthin productivity. To reduce production costs and enhance competitiveness from an industrial point of view, low-cost raw materials from industrial and agricultural origin have been adopted to get the maximal astaxanthin productivity. Finally, fermentation parameters have been studied in depth, both at flask and fermenter scales, to get maximal astaxanthin titers of 4.7 mg/g dry cell matter (420 mg/l) when X. dendrorhous was fermented under continuous white light. The industrial scale-up of this biotechnological process will provide a cost-effective method, alternative to synthetic astaxanthin, for the commercial exploitation of the expensive astaxanthin (about $2,500 per kilogram of pure astaxanthin).     

Bioprocess design and economic analysis for the commercial production of environmentally friendly bioinsecticides from Bacillus thuringiensis HD-1 kurstaki

A production process for B. thuringiensis (Bt) bioinsecticides was designed in detail, including alternative batch, low-density fed-batch (LDFB), and high-density fed-batch (HDFB) fermentation configurations. Capital and operating costs, as well as profitability based on simple rate of return, were performed using a purpose-written FORTRAN program, explicitly analyzing production of a water-based flowable product used in forestry applications.The total capital cost was 18 million dollars (Canadian dollars) for a stand-alone plant with base-scale capacity of 3 x 10(7) billion international units (BIU)/year. Raw material costs amounted to 1.5 million dollars yearly, of which approximately half was for formulation ingredients. Per-unit production cost rose sharply for scales of less than 1 x 10(7) BIU/year, but was little affected by scale above 3 x 10(7) BIU/year. Product cost was much lower at all scales for a LDFB as opposed to batch fermentation process, but HDFB gave relatively little additional cost benefit. Profitability analysis performed by co-varying scale and selling price showed that break-even occurred at a price of 0.45 dollars/BIU for a batch process at base scale, while with LDFB fermentation the same production volume sold at 0.35 dollars/BIU gave a 12% rate of return. Since the assumed base scale would represent 8-15% of current world Bt bioinsecticide production, based on value or volume, it was concluded that profitability would require some or all of the following elements: targeting higher-value markets such as disease vector control, in addition to forestry; a potentially lower plant capacity (although at least 1 x 10(7) BIU/year;) and coproduction of other large-volume microbial products to absorb capacity and match bioinsecticide output to market demand.     

Isolation and characterization of Campylobacter bacteriophages from retail poultry

The ability of phages to survive processing is an important aspect of their potential use in the biocontrol of Campylobacter in poultry production. To this end, we have developed a procedure to recover Campylobacter bacteriophages from chilled and frozen retail poultry and have validated the sensitivity of the method by using a characterized Campylobacter phage (i.e., NCTC 12674). By using this method, we have shown that Campylobacter phages can survive on retail chicken under commercial storage conditions. Retail chicken portions purchased in the United Kingdom were screened for the presence of endogenous Campylobacter phages. Thirty-four Campylobacter bacteriophages were isolated from 300 chilled retail chicken portions, but none could be recovered from 150 frozen chicken portions. The phage isolates were characterized according to their lytic profiles, morphology, and genome size. The free-range products were significantly more likely to harbor phages (P < 0.001 by single-factor analysis of variance) than were standard or economy products. This study demonstrates that Campylobacter bacteriophages, along with their hosts, can survive commercial poultry processing procedures and that the phages exhibited a wide range of recovery rates from chicken skin stored at 4 degrees C.     

正辛酸沉淀在单克隆抗体纯化中的工艺优化与应用

为应对治疗性抗体快速增长的市场需求,抗体上游细胞培养规模和表达量水平已显著提高,而下游纯化工艺的生产效率则相对落后,下游处理能力已成为限制抗体产能的瓶颈。本研究以单克隆抗体mab-X为实验材料,优化了细胞培养液、低pH病毒灭活收集液2种模式的正辛酸(caprylic acid,CA)沉淀工艺条件,并研究了CA处理去除聚体、CA处理灭活病毒等2种应用,在小试的基础上,采用低pH病毒灭活收集液CA沉淀的模式进行了500 L细胞培养规模生产放大研究,对沉淀前后的产品质量和收率进行了检测和对比。结果显示,两种模式的CA沉淀均可显著降低宿主细胞蛋白(host cell protein,HCP)残留和聚体含量,在聚体去除实验中CA沉淀可去除约15%的聚体,病毒灭活研究显示CA对逆转录模型病毒具有完全的病毒灭活能力。在放大生产规模中,下游依次进行了深层过滤收获、亲和层析、低pH病毒灭活、CA沉淀及深层过滤、阳离子交换层析,CA沉淀过程中混合时间和搅拌速度显著影响CA沉淀效果,CA沉淀处理后低pH病毒灭活液中的HCP残留量降低了895倍,沉淀后产品纯度和HCP残留均已控制在单克隆抗体质量要求范围内,CA沉淀可以减少传统纯化工艺中的一个精纯步骤。总之,下游工艺中采用CA沉淀,能够精简传统纯化工艺,并完全满足mab-X的纯化质量要求,而且能提高生产效率、降低生产成本。本研究结果将推动CA沉淀在单克隆抗体下游纯化生产中的应用,为解决目前传统纯化工艺的问题提供参考。     

Softening the agricultural matrix: a novel agri‐environment scheme that balances habitat restoration and livestock grazing

The loss and degradation of woody vegetation in the agricultural matrix represents a key threat to biodiversity. Strategies for habitat restoration in these landscapes should maximize the biodiversity benefit for each dollar spent in order to achieve the greatest conservation outcomes with scarce funding. To be effective at scale, such strategies also need to account for the opportunity cost of restoration to the farmer. Here, we critique the Whole‐of‐Paddock Rehabilitation program, a novel agri‐environment scheme which seeks to provide a cost‐effective strategy for balancing habitat restoration and livestock grazing. The scheme involves the revegetation of large (minimum 10 ha) fields, designed to maximize biodiversity benefits and minimize costs while allowing for continued agricultural production. The objectives and design of the scheme are outlined, biodiversity and production benefits are discussed, and we contrast its cost‐effectiveness with alternative habitat restoration strategies. Our analysis indicates that this scheme achieves greater restoration outcomes at approximately half the cost of windbreak‐style plantings, the prevailing planting configuration in southeastern Australia, largely due to a focus on larger fields, and the avoidance of fencing costs through the use of existing farm configuration and infrastructure. This emphasis on cost‐effectiveness, the offsetting of opportunity costs through incentive payments, and the use of a planting design that seeks to maximize biodiversity benefits while achieving production benefits to the farmer, has the potential to achieve conservation in productive parts of the agricultural landscape that have traditionally been “off limits” to conservation.     

Economic analysis of royalactin production under uncertainty: Evaluating the effect of parameter optimization

Royalactin is a protein with several different potential uses in humans. Research, in insects and in mammalian cells, has shown that it can accelerate cell division and prevent apoptosis. The method of action is through the use of the epidermal growth factor receptor, which is present in humans. Potential use in humans could be to lower cholesterolemic levels in blood, and to elicit similar effects to those seen in bees, e.g., increased lifespan. Mass production of Royalactin has not been accomplished, though a recent article presented a Pichia pastoris fermentation and recovery by aqueous two‐phase systems at laboratory scale as a possible basis for production. Economic modelling is a useful tool with which compare possible outcomes for the production of such a molecule and in particular, to locate areas where additional research is needed and optimization may be required. This study uses the BioSolve software to perform an economic analysis on the scale‐up of the putative process for Royalactin. The key parameters affecting the cost of production were located via a sensitivity analysis and then evaluated by Monte Carlo analysis. Results show that if titer is not optimized the strategy to maintain a low cost of goods is process oriented. After optimization of this parameter the strategy changes to a product‐oriented and the target output becomes the critical parameter determining the cost of goods. This study serves to provide a framework for the evaluation of strategies for future production of Royalactin, by analyzing the factors that influence its cost of manufacture. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:744–749, 2015     

Pretreatments for enhancing clarification efficiency of depth filtration during production of monoclonal antibody therapeutics

High cell density, high product titer mammalian cell culture is the new paradigm for production of recombinant proteins. While the typical motivation is to get a high product titer, additional undesirable outcomes often include an increase in percentage solids in the cell culture fluid (cellular debris and sub-micron colloids), thereby offering new challenges to downstream processing. This article focuses on scouting and comparison of different approaches used for clarification of cell culture fluid. The approaches include centrifugation followed by depth filtration, direct depth filtration without centrifugation and feed pretreatment with use of specially designed density gradient filtration to improve efficiency of clarification and removal of process contaminants from feed stream. The work also evaluates impact of three different pretreatment approaches, namely pH adjustment to acidic condition, metal cation (calcium phosphate) flocculation, and polycationic polymer flocculation (using polymer-I and polymer-II). The results obtained indicate that the use of pretreatment significantly improves the clarification efficiency of depth filtration. Pretreatment options like polycationic polymer-I based flocculation resulted in a >5 fold reduction in filter area requirement as well as >6 fold reduction in HCDNA while retaining acceptable recovery of the IgG (>98%). Thus, pretreatment offers a significant reduction in the depth filtration footprint (~5–6 fold decrease in filter area requirement). However, one must take into consideration the process development time required, capital cost, consumable cost, cost of the pretreatment chemical, cost of testing to demonstrate clearance of treatment agent, ease of scale-ability, and process robustness when finalizing the optimal clarification approach.