title: Ultrasound-assisted extraction of azadirachtin from dried entire fruits of Azadirachta indica A. Juss. (Meliaceae) and its determination by a validated HPLC-PDA method.
abstract: Azadirachta indica A. Juss., also known as neem, is a Meliaceae family tree from India. It is globally known for the insecticidal properties of its limonoid tetranortriterpenoid derivatives, such as azadirachtin. This work aimed to optimize the azadirachtin ultrasound-assisted extraction (UAE) and validate the HPLC-PDA analytical method for the measurement of this marker in neem dried fruit extracts. Box-Behnken design and response surface methodology (RSM) were used to investigate the effect of process variables on the UAE. Three independent variables, including ethanol concentration (%, w/w), temperature (°C), and material-to-solvent ratio (gmL(-1)), were studied. The azadirachtin content (µgmL(-1)), i.e., dependent variable, was quantified by the HPLC-PDA analytical method. Isocratic reversed-phase chromatography was performed using acetonitrile/water (40:60), a flow of 1.0mLmin(-1), detection at 214nm, and C18 column (250×4.6mm(2), 5µm). The primary validation parameters were determined according to ICH guidelines and Brazilian legislation. The results demonstrated that the optimal UAE condition was obtained with ethanol concentration range of 75-80% (w/w), temperature of 30°C, and material-to-solvent ratio of 0.55gmL(-1). The HPLC-PDA analytical method proved to be simple, selective, linear, precise, accurate and robust. The experimental values of azadirachtin content under optimal UAE conditions were in good agreement with the RSM predicted values and were superior to the azadirachtin content of percolated extract. Such findings suggest that UAE is a more efficient extractive process in addition to being simple, fast, and inexpensive.
authors.affiliation: Unidade Universitária de Ciências Exatas e Tecnológicas, Universidade Estadual de Goiás, Anápolis, Goiás, Brazil. Electronic address: joelma.paula@ueg.br.
authors.forename: Joelma Abadia Marciano
authors.initials: JAM
authors.lastname: de Paula
authors.affiliation: Curso de Farmácia, Instituto de Ciências da Saúde, Universidade Paulista, Goiânia, Goiás, Brazil.
authors.forename: Lucas Ferreira
authors.initials: LF
authors.lastname: Brito
authors.affiliation: Laboratório de Pesquisa, Desenvolvimento and Inovação de Bioprodutos, Universidade Federal de Goiás, Goiânia, Goiás, Brazil.
authors.forename: Karen Lorena Ferreira Neves
authors.initials: KLFN
authors.lastname: Caetano
authors.affiliation: Laboratório de Pesquisa, Desenvolvimento and Inovação de Bioprodutos, Universidade Federal de Goiás, Goiânia, Goiás, Brazil.
authors.forename: Mariana Cristina
authors.initials: MC
authors.lastname: de Morais Rodrigues
authors.affiliation: Curso de Farmácia, Pontifícia Universidade Católica de Goiás, Goiânia, Goiás, Brazil.
authors.forename: Leonardo Luiz
authors.initials: LL
authors.lastname: Borges
authors.affiliation: Laboratório de Pesquisa, Desenvolvimento and Inovação de Bioprodutos, Universidade Federal de Goiás, Goiânia, Goiás, Brazil.
authors.forename: Edemilson Cardoso
authors.initials: EC
authors.lastname: da Conceição
mesh_terms: D031601:Azadirachta; D002851:Chromatography, High Pressure Liquid; D000431:Ethanol / Q000737:chemistry; D005638:Fruit / Q000737:chemistry; D057230:Limit of Detection; D036701:Limonins / Q000032:analysis* / Q000737:chemistry*; D015203:Reproducibility of Results; D013696:Temperature; D000069453:Ultrasonic Waves
chemical_list: D036701:Limonins; D000431:Ethanol; C010329:azadirachtin
keywords: Azadirachta indica; Azadirachtin; Box–Behnken design; HPLC-PDA; Response surface methodology; Ultrasound-assisted extraction