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1.
Methods Mol Biol ; 2704: 231-243, 2023.
Article in English | MEDLINE | ID: mdl-37642848

ABSTRACT

Phytosterols, coming as a by-product of vegetable oils or wood pulp, contain the cyclopentanoperhydrophenanthrene nucleus and can be bioconverted into steroid intermediates by removing the C17 side chain. This chapter shows the scale-up, from flask to bioreactor, of phytosterols bioconversion into 4-androstene-3,17-dione (androstenedione; AD) using Mycolicibacterium neoaurum B-3805. Due to the fact that phytosterols and AD are nearly insoluble in water, two-phase systems and the use of chemically modified cyclodextrins have been described as methods to solve it. Here, we use a water-oil two-phase system that allows the bioconversion of up to 20 g/L of phytosterols into AD in 5 L and 20 L bioreactors.


Subject(s)
Androstenedione , Phytosterols , Androstenes , Bioreactors , Water
2.
Methods Mol Biol ; 2296: 351-363, 2021.
Article in English | MEDLINE | ID: mdl-33977458

ABSTRACT

Daptomycin is a cyclic lipopeptide antibiotic with potent activity against gram-positive bacteria. It has a calcium-dependent mechanism of action that disrupts multiple features of the bacterial membrane function. This antibiotic is highly demanded due to its effectiveness against to microorganisms resistant to other antibiotics, including vancomycin-resistant Staphylococcus aureus (VRSA) and methicillin-resistant S. aureus (MRSA). Daptomycin is produced by fermentation of Streptomyces roseosporus, currently identified as Streptomyces filamentosus. However, low fermentation yields and high production costs are reported. This chapter describes a method of strain improvement involving random mutagenesis, rational screening by bioassay, and flask fermentation. The ultimate objective is to select mutants of S. roseosporus overproducing daptomycin in order to design a more cost-effective daptomycin production.


Subject(s)
Daptomycin/biosynthesis , Streptomyces/metabolism , Anti-Bacterial Agents/biosynthesis , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Daptomycin/pharmacology , Fermentation/physiology , Genetic Engineering/methods , Methicillin-Resistant Staphylococcus aureus/drug effects , Methicillin-Resistant Staphylococcus aureus/genetics , Mutagenesis/genetics , Streptomyces/drug effects , Streptomyces/genetics
3.
Methods Mol Biol ; 1852: 257-268, 2018.
Article in English | MEDLINE | ID: mdl-30109636

ABSTRACT

Lycopene is a carotenoid mainly present in red-colored fruits and vegetables. Its value in the pharmaceutical and food industry is linked to its benefits for the human health, including properties against cancer and cardiovascular diseases, and its use as a food colorant. Lycopene can be produced either by synthetic or natural means, but there is a preference for the second, since it is considered a more eco-friendly and less harmful process. Among natural methods for obtaining lycopene, microbial fermentation is a good alternative to extraction from plants that naturally contain lycopene, since it implies obtaining higher and more specific amounts of this carotenoid. This chapter describes lycopene production by fermentation of the fungus Blakeslea trispora, a naturally carotenoid producer, at 30 L scale. This procedure involves separated growth of the two sexual mating types of B. trispora during the vegetative stages and the use of a lycopene cyclase inhibitor to achieve lycopene accumulation during the production stage.


Subject(s)
Fermentation , Lycopene/metabolism , Mucorales/metabolism , Biomass , Bioreactors , Biosynthetic Pathways , Carotenoids/analysis , Carotenoids/biosynthesis , Carotenoids/chemistry , Glucose/metabolism , Lycopene/analysis , Lycopene/chemistry , Molecular Structure , Phosphates/metabolism , Viscosity
4.
Methods Mol Biol ; 1645: 199-210, 2017.
Article in English | MEDLINE | ID: mdl-28710630

ABSTRACT

Phytosterols, generated as a by-product of vegetable oils or wood pulp, contain the cyclopentane-perhydro-phenanthrene nucleus, and can be converted into steroid intermediates by removing the C17 side chain. This chapter shows the scale-up, from flask to fermentor, of the phytosterols bioconversion into 4-androstene-3,17-dione (androstenedione; AD) with Mycobacterium neoaurum B-3805. Due to the fact that phytosterols and AD are nearly insoluble in water, two-phase systems and the use of chemically modified cyclodextrins have been described as methods to solve it. Here we use a water-oil two-phase system that allows for the bioconversion of up to 20 g/L of phytosterols into AD in 20 L fermentor.


Subject(s)
Androstenedione/biosynthesis , Biotransformation , Mycobacterium/genetics , Phytosterols/chemistry , Androstenedione/chemistry , Mycobacterium/metabolism , Plant Oils/chemistry , Glycine max/chemistry , Water/chemistry
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