Cultivation of Phaffia rhodozyma (Xanthophyllomyces dendrorhous) yeast in discontinuous system to obtain astaxanthin

A levedura Phaffia rhodozyma, produtora de astaxantina, pigmento carotenóide largamente empregado na aqüicultura de peixes e crustáceos, pode ser eficientemente cultivada num meio de cultura de baixo custo, à base de caldo de cana diluído 1:10 e uréia a 1 g/L. No entanto, a produção de biomassa e a formação do carotenóide sofrem a inibição pelo substrato (efeito "Crabtree"), limitando desta forma a utilização do caldo de cana com concentrações da fonte de carbono superiores a 20 g/L, importante consideração na produção industrial de astaxantina. No presente trabalho, o cultivo da levedura P. rhodozyma foi realizado em processo descontínuo alimentado, no qual se obteve produtividade volumétrica de 0,024 mg astaxantina/L.h. em relação aos 0,013 mg astaxantina/L.h. obtidos no cultivo controle, que não sofreu alimentação da fonte de carbono.The yeast Phaffia rhodozyma produces astaxanthin, a carotenoid pigment widely applied in fish and crustaceous cultivation. This yeast can be efficiently cultured in a low cost medium, sugar cane broth diluted 1:10 and supplemented with 1 g/L urea. However, the biomass and astaxanthin production undergo inhibition by the substrate (Crabtree effect), limiting the utilization of sugar cane broth up to 20 g/L total sugar concentration. Therefore, this effect must be considered during the industrial production of astaxanthin. In the present work, using fed batch system to cultivate P. rhodozyma we were able to obtain 0.024 mg astaxanthin/l.h compared to 0.013 mg astaxanthin/l.h obtained by the discontinuous cultivation system

Depolimerizacija škroba pomoću razrijeđene fosforne kiseline i primjena hidrolizata u fermentaciji astaksantina

An innovative alternative for cassava starch hydrolysis has been developed using diluted (about 0.1 %) phosphoric acid at 160 °C. This technology is advantageous for developing countries where enzyme costs are prohibitive and hydrochloric acid is currently the only catalyst used for starch depolymerization. Lower concentrations of the byproduct hydroxymethyl furfural (HMF) were generated during starch hydrolysis when using phosphoric acid as compared to hydrochloric acid at any given acidic pH. Glucose was the major product from phosphorolysed starch under most reaction conditions, although maltosaccharides with degrees of polymerization from 2 to 7 were also produced, with their relative amounts depending on hydrolysis conditions. Neutralization of the acid with aqueous ammonia produced a hydrolysate with sources of C (free sugars), P (phosphate), and N (ammonium) that could find several applications. We demonstrated one of these, namely the potential for the use of the hydrolysate as a fermentation feedstock, by cultivating the astaxanthin-producing red yeast Xanthophyllomyces dendrorhous on it. Cassava wastewater, a polluting byproduct of starch processing, was found to be a convenient source of nitrogen for this fermentation process.Razvijen je inovativni postupak hidrolize škroba manioke pomoću razrijeđene 0,1 %-tne fosforne kiseline pri 160 °C. Taj je postupak prikladan za zemlje u razvoju gdje je cijena enzima ograničavajući faktor, a klorovodična kiselina jedini katalizator koji se primjenjuje pri hidrolizi škroba. Primjenom fosforne kiseline nastaje manje nusprodukta hidroksimetilfurfurala nego s klorovodičnom kiselinom. Pri skoro svim uvjetima hidrolize glavni je produkt glukoza, iako nastaju i maltosaharidi stupnja polimerizacije od 2 do 7, ovisno o uvjetima hidrolize. Neutralizacijom kiseline vodenom otopinom amonijaka dobiven je hidrolizat koji može služiti kao izvor ugljika (slobodni šećeri), fosfora (fosfati) i dušika (amonijak) za razne primjene. U radu je navedena mogućnost uporabe hidrolizata kao podloge za uzgoj crvenoga kvasca Xanthophyllomyces dendrorhous pri proizvodnji astaksantina. Otpadne vode od proizvodnje škroba manioke, koje inače zagađuju okoliš, prikladan su izvor dušika u tom postupku

Otimização da produção de biomassa e astaxantina pela levedura Phaffia rhodozyma

A combinação de processos descontínuo alimentado e matérias-primas de baixo custo (caldo de cana-de-açúcar e uréia) foi estudada a fim de otimizar a produção de biomassa e astaxantina pela levedura Phaffia rhodozyma ATCC 24202. No processo otimizado, produtividades em biomassa e astaxantina de 0,327 g/L/h e 0,124 mg/L/h foram obtidas, respectivamente. Comparadas com o processo descontínuo estudado, verificou-se aumento de 4,55 vezes na produtividade em biomassa e 4,73 vezes na produtividade em astaxantina.The combination of fed-batch processes and low cost substrates (sugar cane juice and urea) was studied in view of the optimization of biomass and astaxanthin production by the yeast Phaffia rhodozyma ATCC 24202. In the optimized process, a biomass and astaxanthin productivity of 0.327 g/l/h and 0.124 mg/l/h was achieved, respectively. Compared to the batch process studied, an increase of approximately 4.55-fold in the biomass productivity and 4.73-fold in the astaxanthin productivity was found

Starch Depolymerization with Diluted Phosphoric Acid and Application of the Hydrolysate in Astaxanthin Fermentation

An innovative alternative for cassava starch hydrolysis has been developed using diluted (about 0.1 %) phosphoric acid at 160 °C. This technology is advantageous for developing countries where enzyme costs are prohibitive and hydrochloric acid is currently the only catalyst used for starch depolymerization. Lower concentrations of the byproduct hydroxymethyl furfural (HMF) were generated during starch hydrolysis when using phosphoric acid as compared to hydrochloric acid at any given acidic pH. Glucose was the major product from phosphorolysed starch under most reaction conditions, although maltosaccharides with degrees of polymerization from 2 to 7 were also produced, with their relative amounts depending on hydrolysis conditions. Neutralization of the acid with aqueous ammonia produced a hydrolysate with sources of C (free sugars), P (phosphate), and N (ammonium) that could find several applications. We demonstrated one of these, namely the potential for the use of the hydrolysate as a fermentation feedstock, by cultivating the astaxanthin-producing red yeast Xanthophyllomyces dendrorhous on it. Cassava wastewater, a polluting byproduct of starch processing, was found to be a convenient source of nitrogen for this fermentation process

Thermopressurized diluted phosphoric acid pretreatment of ligno(hemi)cellulose to make free sugars and nutraceutical oligosaccharides

Ligno(hemi)cellulosics (L(h)Cs) as sugarcane bagasse and loblolly pine sawdust are currently being used to produce biofuels such as bioethanol and biobutanol through fermentation of free sugars that are often obtained enzymatically. However, this bioconversion requires a pretreatment to solubilize the hemicellulose fractions, thus facilitating the action of the cellulolytic enzymes. Instead of the main free monosaccharides used in these current models, the modulation of thermopressurized orthophosphoric acid as a pretreatment, in the ranges of 3–12 atm and pH 1.5–2.5, can produce nondigestible oligosaccharides (NDOS) such as xylo-oligosaccharides (XOS) because heteroxylan is present in both types of hardwood and softwood hemicelluloses. A comparative thin-layer chromatographic analysis of the hydrolytic products showed the best conditions for NDOS production to be 7 atm/water, pH 2.25 and 2.50, and 8.5 atm/water for both sources. Particular hydrolysates from 7 atm (171 °C) at pHs 2.25 and 2.50 both for cane bagasse and pine sawdust, with respective oligosaccharide contents of 57 and 59 %, once mixed in a proportion of 1:1 for each plant source, were used in vitro as carbon sources for Bifidobacterium or Lactobacillus. Once both bacteria attained the stationary phase of growth, an unforeseen feature emerged: the preference of B. animalis for bagasse hydrolysates and, conversely, the preference of L. casei for pine hydrolysates. Considering the fact that nutraceutical oligosaccharides from both hemicelluloses correspond to higher value-added byproducts, the technology using a much diluted thermopressurized orthophosphoric acid pretreatment becomes an attractive choice for L(h)Cs

Accelerating the Morphogenetic Cycle of the Viral Vector Aedes aegypti Larvae for Faster Larvicidal Bioassays

Any bioassay to test new chemically synthesized larvicides or phytolarvicides against Culicidae and more harmful mosquito species, such as Aedes aegypti and Aedes albopictus, which specifically transmit dengue, yellow fever, chikungunya viral fevers as well as Zika virus, or Anopheles gambiae, a vector for malaria and philariasis, requires thousands of well-developed larvae, preferably at the fourth instar stage. The natural morphogenetic cycle of Aedes spp., in the field or in the laboratory, may extend to 19 days at room temperature (e.g., 25°C) from the first permanent contact between viable eggs and water and the last stage of larval growth or metamorphosis into flying adults. Thus, accelerated sequential molting is desirable for swifter bioassays of larvicides. We achieved this goal in Aedes aegypti with very limited strategic and low-cost additions to food, such as coconut water, milk or its casein, yeast extract, and to a lesser extent, glycerol. The naturally rich coconut water was excellent for quickly attaining the population of instar IV larvae, the most advanced one before pupation, saving about a week, for subsequent larvicidal bioassays. Diluted milk, as another food source, allowed an even faster final ecdysis and adults are useful for mosquito taxonomical purpose

Effect of feeding methods on the astaxanthin production by Phaffia rhodozyma in fed-batch process

The effect of feeding methods on the production of astaxanthin by the yeast Phaffia rhodozyma ATCC 24202 was studied, using continuous and pulsed fed-batch processes and low cost materials as substrates (sugar cane juice and urea). In continuous fed-batch processes, a cellular astaxanthin concentration of 383.73 µg/g biomass was obtained. But in pulsed fed-batch processes a reduction in the cellular astaxanthin concentration (303.34 µg/g biomass) was observed. Thus the continuous fed-batch processes could be an alternative to industrial production of astaxanthin, allowing an increase in the biomass productivity without losses on astaxanthin production by the yeast.<br>O efeito da alimentação na produção de astaxantina pela levedura Phaffia rhodozyma ATCC 24202 foi estudado, utilizando processos descontínuo alimentado com alimentação contínua e intermitente, e matérias-primas de baixo custo como substratos (caldo de cana de açúcar e uréia). Em processos descontínuo alimentado com alimentação contínua, uma concentração celular de astaxantina de 383,73 µg/g biomassa foi obtida. Entretanto, em processos descontínuo alimentado com alimentação intermitente, uma redução na concentração celular de astaxantina (303,34 µg/g biomassa) foi observada. Desta forma, processos descontínuo alimentado com alimentação contínua poderiam ser uma alternativa na produção industrial de astaxantina, permitindo um aumento na produtividade de biomassa sem perdas na produção de astaxantina pela levedura