Extraction and identification of secondary metabolites from mangrove rare actinomycete actinophytocola sp. K4-08 with bioactivity potential

Abstract

Actinomycetes are aerobic filamentous Gram-positive bacteria that produce various secondary metabolites, notably antibiotics. Unfortunately, the effectiveness of this bacteria has been jeopardized in recent years due to the rise of multidrug-resistance bacteria. Hence, researchers have switched to ‘non-streptomycetes’ to gain novel metabolic compounds. The current study was designed to extract and identify the microbial compounds from mangrove rare actinomycete, Actinophytocola sp. K4-08 (KR902625) was previously isolated from Kuantan mangrove sediments. To date, this is the first study that demonstrates the properties of the genus Actinophytocola sp. concerning their biological potential. Colonies of Actinophytocola sp. K4-08 was subjected to morphological characterization using gram staining and scanning electron microscope (SEM). Actinophytocola sp. K4-08 is a Gram-positive bacterium with branched substrate mycelium fragmented into a rod-like shape and regular round chain spore formation. Moreover, this strain utilised more than 10 carbon sources and tolerated up to 10 % sodium chloride (NaCl), demonstrating its adaptation to the marine environment. Crude extracts from both supernatant and cells of Actinophytocola sp. K4-08 were prepared using different solvent namely, ethyl acetate, methanol, and acetone with XAD-2 resins. Extraction with ethyl acetate produced dark yellow liquid residue and brownish solid residues with the highest crude at 1.35 g. Solid acetone (AE) and liquid methanol (ME) crudes showed significant antibacterial activities against Bacillus subtilis with inhibition of 7.9 ± 0.1 mm and 12.0 ± 0.0 mm respectively through disc diffusion susceptibility test. Overall, liquid crude extracts exhibited higher antagonistic activity against B. subtilis than solid crude extracts. The antioxidant activities of crude extracts were further assessed using total phenolic content (TPC), total flavonoid content (TFC), free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) assay. Liquid ethyl acetate (EA) crude demonstrated higher TPC and TFC, while solid EA crude showed higher DPPH scavenging and FRAP assays. Both solid and liquid EA crude extracts have moderately good antioxidant potential. Liquid EA crude was further analysed for cytotoxicity assays against human non-small lung cancer cells. A549 cell was the most sensitive toward the liquid EA crude than the H1299 cell line, with a higher reduction in cell viability at 62.52 ± 0.76 % and 79.13 ± 0.90 %, respectively. The presence of O-H, C-H, and C=C bonding was identified using Fourier-transform infrared spectroscopy (FT-IR) and 2,4-bis (1,1-dimethyl ethyl)-phenol (2,4-DTP), an anticancer drug was detected in solid ME crude using gas chromatography-mass spectrometry (GC-MS). Chemical profiling using reversed-phase thin-layer chromatography (RP-TLC) and high-performance liquid chromatography (HP-LC) showed good separation of microbial compounds in liquid EA crude with acetonitrile and water (1: 1.5, v/v) solvent ratio. Ningpeisinoside, lamiophlomiol A and pseudolaric acid AO-β-D-glucopyranoside were detected using liquid chromatography-mass spectrometry (LC-MS). The present findings suggested that rare mangrove actinomycete, Actinophytocola sp. K4-08 is a high-potential candidate with interesting biosynthetic capabilities for the drug discovery program.