Studies on nutritional requirement for the culture of lichen Ramalina nervulosa and Ramalina pacifica to enhance the production of antioxidant metabolites Neeraj Verma & B. C. Behera & Ankita Joshi Received: 14 October 2011 / Accepted: 12 January 2012 / Published online: 22 February 2012 # Institute of Microbiology, Academy of Sciences of the Czech Republic, v.v.i. 2012 Abstract In the present report, nutritional requirement for the culture of two lichen species Ramalina nervulosa and Ramalina pacifica were studied in order to enhance their growth rate and antioxidant metabolite production. Extract of R. nervulosa cultured in Bold’ s basal medium (BBM) showed higher antioxidant activity than R. pacifica cultured in Murashige and Skoog (MS) medium. The lichen species were sub-cultured in standardized nutrient media. R. nervu- losa in BBM (1% glucose, 50 ppb asparagines, pH 6.5) yielded 2.76 g biomass with 26.18 mg sekikaic acid, 24.32 mg usnic acid/g dry biomass in a period of 60 days. R. pacifica in MS media (3% sucrose, 100 ppb thiamine, pH 5.9) yielded 3.54 g biomass and 58.92 mg salazinic acid, 40.16 mg usnic acid in the same time period. The standard- ized culture conditions implemented on bioreactor, R. ner- vulosa yielded 17.7 g biomass with the production of sekikaic acid 122.8 mg, usnic acid 75.4 mg in 4.5 days. R. pacifica produced 10.3 g biomass along with salazinic acid 200 mg and usnic acid 136.8 mg in the same duration. Lichen secondary metabolites produced in bioreactor showed moderate antioxidant activity; sekikaic acid 42% to 56.4%; salazinic acid 33.6% to 41.9% and usnic acid 19.9% to 29.5%. Introduction Lichen is a symbiotic association between a mycobiont and a photobiont, which may be an alga or cyanobacteria, result- ing in a stable thallus of specific structure (Ahmadjian 1993). Lichens produces unique variety of secondary metabolites belongs to depsides, depsidones, dibenzofurans and pulvinic acid derivatives family (Huneck and Yoshimura 1996). Till date, 1,050 lichen secondary metab- olites including those from cultures are reported (Molnár and Farkas 2010). Out of which less than 10% lichen metabolites have been investigated for various biological activities. Antioxidant potential of several lichens and their metabolites have been reported by Hidalgo et al. (1994), Behera et al. (2005) and Verma et al. (2008a) and are proved to be a good source of natural antioxidants. However, lichen growth in nature is very slow and not available in sufficient quantity for commercial exploitation. In recent past, Japanese scientists (Yamamoto et al. 1985) have developed methodology for lichen culture. Subsequent to this several workers could be able to obtain cell aggre- gates comprising of both symbionts, which have been used for the screening of many biological activities of lichens and their metabolites for possible pharmaceutical and therapeut- ical applications (Yamamoto et al. 1998; Higuchi et al. 1993; Behera et al. 2005; 2006; 2009; Verma et al. 2008a, b). Although in the laboratory conditions, lichen symbionts as cell aggregates grow much faster than in nature, but they N. Verma (*) : B. C. Behera Mycology and Plant Pathology Group, Plant Science Division, Agharkar Research Institute, G.G. Agarkar Road, Pune 411004, India e-mail: neerajverma2310@gmail.com A. Joshi School of Biochemistry, Devi Ahilya Vishwavidyalaya, Indore 452017, India Folia Microbiol (2012) 57:107–114 DOI 10.1007/s12223-012-0100-2