title: Production of Limonoids with Insect Antifeedant Activity in a Two-Stage Bioreactor Process with Cell Suspension Culture of Azadirachta indica.
abstract: Neem tree (Azadirachta indica) cell suspension culture is an alternative for the production of limonoids for insect control that overcomes limitations related to the supply of neem seeds. To establish conditions for cell growth and azadiracthin-related limonoid production, the effect of different sucrose concentrations, nitrate and phosphate in Murashige and Skoog (MS) medium, and the addition of one precursor and three elicitors was evaluated in shake flasks. The process was scaled up to a 3-l stirred tank bioreactor in one- and two-stage batch cultivation. In shake flasks, more than fivefold increase in the production of limonoids with the modified MS medium was observed (increase from 0.77 to 4.52 mg limonoids/g dry cell weight, DCW), while an increase of more than fourfold was achieved by adding the elicitors chitosan, salicylic acid, and jasmonic acid together (increase from 1.03 to 4.32 mg limonoids/g DCW). In the bioreactor, the volumetric production of limonoids was increased more than threefold with a two-stage culture in day 18 (13.82 mg limonoids/l in control single-stage process and 41.44 mg/l in two-stage process). The cultivation and operating mode of the bioreactor reported in this study may be adapted and used in optimization and process plant development for production of insect antifeedant limonoids with A. indica cell suspension cultures.
authors.affiliation: Grupos de Biotecnología Industrial y Biotecnología Vegetal, Facultad de Ciencias, Universidad Nacional de Colombia Sede Medellín, Calle 59A No 63- 20, Medellín, Colombia, andresvasquezrivera@gmail.com.
authors.forename: Andrés
authors.initials: A
authors.lastname: Vásquez-Rivera
authors.forename: Diego
authors.initials: D
authors.lastname: Chicaiza-Finley
authors.forename: Rodrigo A
authors.initials: RA
authors.lastname: Hoyos
authors.forename: Fernando
authors.initials: F
authors.lastname: Orozco-Sánchez
mesh_terms: D000818:Animals; D031601:Azadirachta / Q000737:chemistry*; D001681:Biological Assay; D018533:Biomass; D019149:Bioreactors; D018929:Cell Culture Techniques / Q000379:methods*; D048271:Chitosan / Q000494:pharmacology; D003470:Culture Media; D003517:Cyclopentanes / Q000494:pharmacology; D005247:Feeding Behavior / Q000187:drug effects; D007306:Insecticides / Q000633:toxicity*; D036701:Limonins / Q000633:toxicity*; D009566:Nitrates / Q000494:pharmacology; D054883:Oxylipins / Q000494:pharmacology; D010710:Phosphates / Q000494:pharmacology; D020156:Salicylic Acid / Q000494:pharmacology; D019346:Sodium Acetate / Q000494:pharmacology; D018411:Spodoptera / Q000187:drug effects* / Q000502:physiology; D013395:Sucrose / Q000494:pharmacology; D013535:Suspensions
chemical_list: D003470:Culture Media; D003517:Cyclopentanes; D007306:Insecticides; D036701:Limonins; D009566:Nitrates; D054883:Oxylipins; D010710:Phosphates; D013535:Suspensions; D019346:Sodium Acetate; D013395:Sucrose; C011006:jasmonic acid; D048271:Chitosan; D020156:Salicylic Acid