Investigation of antioxidant, antimicrobial and toxicity activities of lichens from high altitude regions of Nepal

Abstract

Background: Several lichen species are reported to be used tradiationally in many theraupatic practices. Many lichen species are reported as sources of several bioactive natural compounds. Several lichen species of Nepal are so far chemically unexplored.

Methods: The morphological, anatomical and phytochemical characteristics of lichens were compared for the taxonomic identification of the species. Methanol- water extract of lichens were sub fractionated into hexane, dichloromethane and methanol fractions for bioactivity assays. Antimicrobial activities of extracts were evaluated agaisnt pathogenic bacteria and fungal species. DPPH test was used for antioxidant potential evaluation. Brineshrimp test was perfermed to evaluate toxicity of the extracts.

Results: A total of 84 lichen specimens were collected and identified from Annapurna Conservation Area (ACA) Nepal. The specimens were identified as belonging to 19 genera and 47 species. Methanol fractions of 16 specimens and dichloromethane (DCM) fractions of 21 lichens specimens showed antioxidant activities comparable with commercial standards (BHA, Butylated hydroxyanisole, IC50=4.9±0.9 μg/mL) even at crude extract level. Similarly, the DCM fraction of 17 lichens showed potential antimicrobial activity against a Gram-positive bacterium (Staphylococcus aureus KCTC3881) and DCM fractions of 45 lichens showed antimicrobial activity against a Gram-negative bacterium (Klebsiella pneumoniae KCTC2242). DCM fractions of three lichens showed antifungal activity against the yeast, Candida albicans KCTC 7965. Likewise, methanol fractions of 39 lichens and DCM fractions of 74 lichens showed strong toxicity against brine shrimp nauplii with more than 80% mortality.

Conclusion: Such biological activity-rich lichen specimens warrant further research on exploration of natural products with antioxidant, antimicrobial and anti cancer (toxic) potential.

Keywords: Antimicrobial; Antioxidant; DPPH; Lichen; Thin layer chromatography.

Fig. 1

TLC based chemical screening of lichen extracts. Plate viewed under UV (254 nm). Mobile phase for TLC development was 10% methanol in DCM. W-methanol soluble water fraction, D-DCM fractions. The sample in the TLC plate is as follows: 1, SAR1W; 2, SAR9W; 3, SAR11W; 4, SAR13W; 5, SAR15W; 6, SAR18W; 7, SAR27W; 8, GHAN5W; 9, GHAN8W; 10, GHAN9W; 11, GHAN13W; 12, JOM6W; 13, DAN1W; 14, DAN6W; 15, DAN11W; 16, CHA8W; 17, SAR1D; 18, SAR5D; 19, SAR6D; 20, SAR12D; 21, SAR23D; 22, SAR24D; 23, SAR25D; 24, SAR26D; 25, GHAN5D; 26, GHAN8D; 27, GHAN15D; 28, JOM14D; 29, JOM15D; 23, JOM16D; 31, DAN1D; 32, DAN3D; 33, DAN6D; 34, DAN8D; 35, DAN11D; 36, CHA1D; 37 CHA8D

Fig. 2

TLC based antioxidant screening of the active extracts. Mobile phase for TLC development was 10% methanol in DCM. The purple color was from DPPH and antioxidant active fractions showed reduced in purple color to yellow. The darkness of color spots indicated the potential of antioxidant activity, the size of color spot indicated the content of antioxidant active compounds in the extracts. W- methanol soluble water fraction, D- DCM fractions of lichen extracts. The sample in the TLC plate is as follows:1, SAR1W; 2, SAR9W; 3, SAR11W; 4, SAR13W; 5, SAR15W; 6, SAR18W; 7, SAR27W; 8, GHAN5W; 9, GHAN8W; 10, GHAN9W; 11, GHAN13W; 12, JOM6W; 13, DAN1W; 14, DAN6W; 15, DAN11W; 16, CHA8W; 17, SAR1D; 18, SAR5D; 19, SAR6D; 20, SAR12D; 21, SAR23D; 22, SAR24D; 23, SAR25D; 24, SAR26D; 25, GHAN5D; 26, GHAN8D; 27, GHAN15D; 28, JOM14D; 29, JOM15D; 23, JOM16D; 31, DAN1D; 32, DAN3D; 33, DAN6D; 34, DAN8D; 35, DAN11D; 36, CHA1D; 37, CHA8D