Optimization of conditions for increasing of saffron cell biomass and crocin production in stirred bioreactor

Abstract

Bioreactors provide suitable conditions for the growth of cells and production of secondary metabolites by regulating physical and chemical factors. In this study, saffron cell growth and the production of crocin metabolite were evaluated in Erlenmeyer flask and in a stirred bioreactor. To optimize the conditions, the effect of different concentrations of 2-(N-morpholino) ethanesulfonic acid (MES) (0, 2.5 and 5 mM) as a buffering agent and sucrose (3%, 6% and gradually 6%) were addressed on saffron cell growth and crocin production in the Erlenmeyer flask. This aim was then pursued in a stirred bioreactor through investigation on the effect of aeration and pH medium adjustment. The results of the first step experiment showed that Schenk and Hildebrandt basal medium with naphthalene acetic acid (2 mg l−1) and 6-benzylaminopurine (1 mg l−1) supplemented with 2.5 mM of MES as well as gradual increment of sucrose from 3 to 6% caused the highest cell biomass and crocin production. The spectrophotometry measurement showed that the highest crocin content of the cells was 0.8 mg g−1 after 6 weeks. The results of the second part revealed that in the stirred bioreactor, constant pH (5.8) during the growth period is a limiting factor for the cell growth and crocin production. Aeration was also found to be an inhibiting factor for the production of crocin. The results showed that, if the evaporated volume of water caused by aeration is constituted, it can be an effective factor to increase cell growth around 2 folds. In addition, total crocin content of the proliferated cells in stirred bioreactor system under the non-constant pH and without aeration conditions, could be raised up to 2 mg g−1 of cell dry weight, based on the HPLC determination. According to the results of this study, it can be concluded that MES and gradual increment of sucrose could increase the cell growth and crocin production. In the tested stirred bioreactor, it was found that the natural pH fluctuation is a suitable condition for saffron biomass cell growth and crocin production. The proper aeration could increase the cell biomass, if the evaporated water is replaced. However based on the obtained results it can be concluded that, aeration at tested value is not capable of increasing the crocin production.