Abstract
Shiraia mycelial culture is a promising biotechnological alternative for the production of hypocrellin A (HA), a new photosensitizer for anticancer photodynamic therapy (PDT). The extractive fermentation of intracellular HA in the nonionic surfactant Triton X-100 (TX100) aqueous solution was studied in the present work. The addition of 25 g/L TX100 at 36 h of the fermentation not only enhanced HA exudation to the broth by 15.6-fold, but stimulated HA content in mycelia by 5.1-fold, leading to the higher production 206.2 mg/L, a 5.4-fold of the control on day 9. After the induced cell membrane permeabilization by TX100 addition, a rapid generation of nitric oxide (NO) and hydrogen peroxide (H2O2) was observed. The increase of NO level was suppressed by the scavenger vitamin C (VC) of reactive oxygen species (ROS), whereas the induced H2O2 production could not be prevented by the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO), suggesting that NO production may occur downstream of ROS in the extractive fermentation. Both NO and H2O2 were proved to be involved in the expressions of HA biosynthetic genes (Mono, PKS and Omef) and HA production. NO was found to be able to up-regulate the expression of transporter genes (MFS and ABC) for HA exudation. Our results indicated the integrated role of NO and ROS in the extractive fermentation and provided a practical biotechnological process for HA production.
Highlights
Hypocrellin A (HA) is a perylenequinone pigment extracted from the fruiting bodies of a pathogenic fungus Shiraia bambusicola in bamboos
Our results demonstrated that increase of HA production in the extractive fermentation was from HA in the cultural broth, and the stimulated biosynthesis of HA in mycelial cells (Figure 1)
We have successfully demonstrated that the extractive fermentation by Triton X-100 (TX100) could enhance greatly HA production in Shiraia cultures, suggesting an effective process strategy to mycelium cultures
Summary
Hypocrellin A (HA) is a perylenequinone pigment extracted from the fruiting bodies of a pathogenic fungus Shiraia bambusicola in bamboos. Liu et al (2016) managed to mutagenize Shiraia spores using cobalt-60 gamma irradiation to obtain mutated strains for higher HA production [10] Apart from these conventional optimization methods, extractive fermentation in water-organic solvent two-phase system, known as perstractive fermentation or milking process, is becoming an efficient strategy to enhance fungal products [11]. Organic surfactant is added to permeabilize cells for intracellular products across the cell membrane and extract the fungal products consecutively in the surfactant micelle aqueous solution. Another two-phase system is formed when a nonionic surfactant micelle aqueous solution is at above a certain temperature (cloud point). The application of the concept of nonionic surfactant micelle aqueous solution or cloud point system to submerged Shiraia fermentation has not been studied
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