Browsing by Author "Carsanba, Erdem"
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- Carnosic acid production from sugarcane syrup by engineered yeast in fed-batch fermentationPublication . Carsanba, Erdem; Fernandes, Sara; Beato, Felipe; Carvalho, Luís Carlos; Pintado, Ana; Lopes, Ana; Ribeiro, Mónica; Leal, Tânia; Pintado, Manuela; Oliveira, CarlaPhenolic diterpene carnosic acid (CA) is widely used in the food, nutritional health, and cosmetic industries due to its antioxidative and antimicrobial properties. This work aimed to overproduce CA in Saccharomyces cerevisiae from sugarcane syrup in fed-batch 2 L bioreactor fermentation. A geranylgeranyl diphosphate (GGPP)-producing strain modified with genes encoding the enzymes copalyl diphosphate synthase (Pv.CPS), miltiradiene synthase (Ro.KSL2), hydroxy ferruginol synthase (Ro.HFS), CA synthase (Ro.CYP76AK8), CYP reductase (At.ATR1), and transketolase (TKL1) was used. Lowering the feed rate from 12–26 g/L/h to 7–8 g/L/h, and the use of a dynamic dissolved oxygen (DO) trigger (min. 10%, max. 40%, threshold 70%) instead of a DO trigger of 30%, enhanced CA production by 27%. As a result, the highest CA titer ever reported to date, 191.4 mg/L, was obtained in 4-day fermentation. This study shows the feasibility of engineered yeast to produce CA from the sustainable feedstock sugarcane syrup.
- Fermentation strategies for production of pharmaceutical terpenoids in engineered yeastPublication . Carsanba, Erdem; Pintado, Manuela; Oliveira, CarlaTerpenoids, also known as isoprenoids, are a broad and diverse class of plant natural products with significant industrial and pharmaceutical importance. Many of these natural products have antitumor, anti-inflammatory, antibacterial, antiviral, and antimalarial effects, support transdermal absorption, prevent and treat cardiovascular diseases, and have hypoglycemic activities. Production of these compounds are generally carried out through extraction from their natural sources or chemical synthesis. However, these processes are generally unsustainable, produce low yield, and result in wasting of substantial resources, most of them limited. Microbial production of terpenoids provides a sustainable and environment-friendly alternative. In recent years, the yeast Saccharomyces cerevisiae has become a suitable cell factory for industrial terpenoid biosynthesis due to developments in omics studies (genomics, transcriptomics, metabolomics, proteomics), and mathematical modeling. Besides that, fermentation development has a significant importance on achieving high titer, yield, and productivity (TYP) of these compounds. Up to now, there have been many studies and reviews reporting metabolic strategies for terpene biosynthesis. However, fermentation strategies have not been yet comprehensively discussed in the literature. This review summarizes recent studies of recombinant production of pharmaceutically important terpenoids by engineered yeast, S. cerevisiae, with special focus on fermentation strategies to increase TYP in order to meet industrial demands to feed the pharmaceutical market. Factors affecting recombinant terpenoids production are reviewed (strain design and fermentation parameters) and types of fermentation process (batch, fed-batch, and continuous) are discussed.
- Microbial production of terpenesPublication . Pimentel, Lígia; Carsanba, Erdem; Teixeira, Francisca; Vidigal, Susana; Pintado, Manuela; Oliveira, Carla; Rodríguez-Alcalá, Luis M.Sustainability is one of the main challenges facing humankind in the twenty-first century due to the continuous increase in the demand for energy and resources that characterizes our current industrial activity, driven in part by exponential population growth. This problem can be tackled from different strategies and among them the search for biomolecules, either recovered through processes based on the circular economy or through synthetic biology, are highly promising. In this sense, terpenes, the largest family of secondary metabolites in the plant kingdom, have attracted much of the research in recent decades in pursuit of alternative, more ecological, and sustainable ways for their obtention. The reasons for this interest are due to their extensive structural diversity and the possibility of gaining new functionalities, simply by chemical modification, which also makes them excellent candidates in areas such as biomaterials and pharmaceuticals. But also, the fact that the natural biosynthetic pathways of terpenes are well known from the point of view of metabolites and enzymes facilitates their industrial production using genetically modified microorganisms. This chapter aims to give the reader a broad but at the same time comprehensive view of the production of microbial terpenes in a sustainability context. Starting by the circumstances that lead to the need to look for renewable sources of biomolecules and following by why terpenes represent a very promising opportunity even if only their characteristics from a chemical and bioactivity point of view were considered. Finally, it will be discussed which microorganisms can produce these unique lipids and how, the main option followed nowadays is using synthetic biology strategies, involving modified organisms that are already being used on an industrial scale for applications ranging from biofuels to pharmaceuticals.
- Modulation of polyphenols in beta-farnesene fed-batch fermentationPublication . Carvalho, Luís Carlos; Oliveira, Ana L. S.; Carsanba, Erdem; Pintado, Maria Manuela; Oliveira, Carla
- Peptide extract from spent yeast improves resistance of Saccharomyces cerevisiae to oxidative stressPublication . Lopes, Ana; Azevedo-Silva, João; Carsanba, Erdem; Pintado, Manuela; Oliveira, Ana Sofia; Ferreira, Carlos; Pereira, Joana Odila; Carvalho, Ana P.; Oliveira, CarlaYeast cells face various stress factors during industrial fermentations, since they are exposed to harsh environmental conditions, which may impair biomolecules productivity and yield. In this work, the use of an antioxidant peptide extract obtained from industrial spent yeast was explored as supplement for Saccharomyces cerevisiae fermentation to prevent a common bottleneck: oxidative stress. For that, a recombinant yeast strain, producer of β-farnesene, was firstly incubated with 0.5 and 0.7 g/L peptide extract, in the presence and absence of hydrogen peroxide (an oxidative stress inducer), for 1–5 h, and then assayed for intracellular reactive oxygen species, and growth ability in agar spot assays. Results showed that under 2 mM H2O2, the peptide extract could improve cells growth and reduce reactive oxygen species production. Therefore, this antioxidant effect was further evaluated in shake-flasks and 2-L bioreactor batch fermentations. Peptide extract (0.7 g/L) was able to increase yeast resistance to the oxidative stress promoted by 2 mM H2O2, by reducing reactive oxygen species levels between 1.2- and 1.7-fold in bioreactor and between 1.2- and 3-fold in shake-flask fermentations. Moreover, improvements on yeast cell density of up to 1.5-fold and 2-fold, and on biomolecule concentration of up to 1.6-fold and 2.8-fold, in bioreactor and shake-flasks, respectively, were obtained. Thus, culture medium supplementation with antioxidant peptide extracted from industrial spent yeast is a promising strategy to improve fermentation performance while valuing biomass waste. This valorization can promote a sustainable and eco-friendly solution for the biotechnology industry by the implementation of a circular economy model.
- Phenolic compounds modulation in β-farnesene fed-batch fermentation using sugarcane syrup as feedstockPublication . Carvalho, Luís Carlos; Oliveira, Ana L. S.; Carsanba, Erdem; Pintado, Manuela; Oliveira, CarlaSugarcane syrup is the feedstock used in the industrial production of β-farnesene through Saccharomyces cerevisiae fermentation. However, it contains phenolic compounds, which may interfere with yeast performance. The aim of this work was to modulate the transfer of phenolic compounds from sugarcane syrup to the broth, throughout 13 days of β-farnesene fed-batch fermentation in 2-L bioreactors. This was accomplished by applying two mathematical calculations: a mass balance and the Weibull kinetic model. The concentrations of most phenolic compounds increased until day 3 of fermentation and then remained constant until the end of the process. However, quantities of hydroxybenzaldehyde, protocatechuic, caffeic, ferulic and p-coumaric acids, decreased after day 2. The Weibull model exhibited better fit to the data, with an R2 of 0.85 or higher. This work provides for the first time a model describing phenolic accumulation in β-farnesene fermentation, which can be a valuable tool to apply in similar processes.
- Valorisation of waste bread for the production of yeast biomass by yarrowia lipolytica bioreactor fermentationPublication . Carsanba, Erdem; Agirman, Bilal; Papanikolaou, Seraphim; Fickers, Patrick; Erten, HuseyinThe increase in the wastage of bread, representing 12.5 million tons per year, causes ecological problems, such as the production of methane and CO2, when that waste bread (WB) is improperly managed. To reduce this ecological footprint, a more sustainable system of WB management must be set up. Based on its chemical composition, WB has a high potential to be used as feedstock for microbial growth and conversion into value-added bio products. The microbial valorisation of WB is a novel biotechnological approach to upgrading a waste into a renewable feedstock for bio-based industry, thus favouring the circular economy concept. Based on this, the aim of this study was to test WB as a feedstock for biomass production by Yarrowia lipolytica, which can be considered as a promising supplement for animal and human dietary products. The enzymatic hydrolysis of WB was primarily optimized for large-scale production in a bioreactor. The biomass production of Y. lipolytica strain K57 on WB hydrolysate-based media in batch bioreactor culture was then investigated. As a result, a very high starch to glucose conversion yield of 97% was obtained throughout optimised hydrolysis. At the end of 47 h of batch culture, a biomass higher than 62 g/L, specific growth rate of 0.37 h−1 and biomass yield of 0.45 g/g were achieved from a WB hydrolysate. Therefore, this study demonstrates that WB hydrolysate has a promising potential to be used as a feedstock for biomass production by Y. lipolytica strain K57 for food and animal diet applications.