Screening of Metarhizium spp. strains for anticancer indolizidine alkaloid production and its rapid detection by MS analysis

Abstract

Six fungi strains (M. anisopliae 3935, 4516, 4819, PL57, PL43 and M. flavoviride CG291) were studied regarding their ability to produce an anticancer indolizidine alkaloid. The culture process was carried out in Shaken flask at 26ºC and 200 rpm using three different culture medium containing oat meal extract supplemented with glucose and DL-lysine or Czapek culture medium. The mycelial extracts produced by Metarhizium spp. cultures were directly submitted to electrospray ionization mass spectrometry (ESI-MS) analysis and the highest alkaloid concentration (approximately, 6 mg.L -1 ) was reached when M. anisopliae 3935 was tested.

Optimization of different process variables for the production of an indolizidine alkaloid, swainsonine from Metarhizium anisopliae

“…Swainsonine being a widely studied asparagine-linked mannosidase inhibitor is a potentially important drug against deadly diseases like cancer, AIDS or any kind of metastasis [15]. Production of this indolizidine alkaloid from M. anisopliae is possible at the fermenter level [10,16], but no studies have, as yet, been conducted using the stepwise classical, statistical or machine learning techniques (MLT) optimization procedures. In the present study, ten isolates of M. anisopliae were screened and isolate UM 8 (ARSEF 1724) showed high swainsonine production at 72 h of incubation.…”

“…Isolates of M. anisopliae were initially screened for their growth and sporulation characteristics on oatmeal agar media and then ten selected isolates were subsequently screened for swainsonine production. Spore suspension (4 × 10 8 spores/ml) were used to inoculate 250 ml duplicate Erlenmeyer flasks containing complex oatmeal media [10] at a fixed concentration of 2% (w/v) [17] with incubation conditions being 180 rpm and 28 °C for 168 h. Samples were removed after every 24 h and assayed for swainsonine production [12].…”

“…There has been only one fungal species reported to date which produces this indolizidine alkaloid, the insect pathogenic fungus Metarhizium anisopliae [9]. Substantial improvement in swainsonine levels from fed batch fermentations has been achieved in a complex medium based on oatmeal extract fed intermittently with Dglucose and lysine [10]. However, there is no optimization process at the shake flask level reported so far by using any of the classical, statistical or the advanced optimization methods.…”

Efeito da adição de precursores na produção de alcaloide anticancerígeno usando a técnica de planejamento experimental

“…16 Para a quantificação do alcaloide uma curva padrão foi preparada adicionando-se 0,5 μL de ácido fórmico e uma solução de metanol (1:1 v/v) à solução de alcaloide padrão (500 μL). As amostras de fermentação também foram preparadas como descrito acima, cujas análises foram feitas por infusão direta com um fluxo de 10 μL min -1 usando uma bomba-seringa (Harvard apparatus).…”

“…16 e uma linhagem de M. anisopliae 3935 foi a que alcançou, comparativamente entre as cepas avaliadas, maior rendimento de alcalóide produzido (da ordem de 6 mg L -1 ). Neste contexto, este trabalho teve como foco avaliar a potencialidade desta cepa de produzir quantidades maiores de alcalóide em função da adição de precursores, como lisina e glicose, usando a metodologia de planejamento experimental com base nos estudos previamente reportados, 10 com o propósito de estabelecer uma correlação empírica para estimar a quantidade de alcalóide formado em função dos precursores adicionados em escala de laboratório.…”