Bioprocess development and optimization of melanin from Pseudomonas stutzeri

Abstract

Melanins are water insoluble polyphenol compounds that are responsible for black, brown and grey pigmentation in eukaryotes and prokaryotes. The presence of indole ring and many functional groups, such as, =O, -OH, -NH, and –COOH makes melanin an ideal choice for many medical, cosmetic and environmental applications. The current market price of melanin is about USD 500 because of unavailability of sustainable manufacturing process. Microorganisms are reported to produce melanin; however, sustainability of a microbial bioprocess depends on raw material requirements and water footprint. A non-pathogenic microorganism capable of growing in seawater and require raw materials, which do not compete for human consumption is ideal for any bioprocess development. Therefore, a marine bacterium known as Pseudomonas stutzeri, which readily grows in seawater and does not require any tyrosine addition was selected for this work on melanin production. The best operating conditions were identified for melanin production. Inoculum age of 12 hours and 10% inoculum gave the best melanin production. Different carbon, nitrogen sources and trace elements were screened and glucose and coconut cake meal were selected as the best carbon and nitrogen sources, respectively. Sulphates of copper, magnesium and iron enhanced melanin production. All these media components were optimized using central composite design and melanin production was increased to 3.4 folds to about 520 mg/L. However, melanin demonstrated biomass growth inhibition as well as feedback inhibition on to the enzyme tyrosinase. An alumina adsorbent based novel adsorptive bioprocess was developed to reduce the feedback inhibition by melanin, which resulted in 8.8 fold enhancement in the production of melanin by the marine bacteria Pseudomonas stutzeri (153 mg/L to 1349.03 mg/L). Further, an adsorption based downstream process was developed using alumina, at acidic pH (< 3) which resulted in 98% recovery of melanin from the fermentation broth. Particle size analysis of the biosynthesized melanin indicated that they are nanoparticles with a size of 32 ± 0.98 nm and preliminary investigation indicated that melanin nanoparticles adsorbed different heavy metals at very low metal concentrations.