Isolation and identification of cyanobacterial samples
Cyanobacterium was isolated from the jhum land soil (25°10/22.9//N and 93°07/10.9//E at an elevation of 600.9 m) of Dima Hasao district of Assam, North-East India in BG-110 media (Rippka 1979) without supplementation of combined nitrogen. The isolated cyanobacterium was identified as Nostoc muscorum Ag. ex Born. et Flah. using the key mentioned in Desikachary (1959).
Biomass production using organic manure
Media was optimized for cyanobacteria biomass production using organic manures – cow dung (CD) and vermicompost (VC). The nutrient extract from thoroughly dried cow dung and vermicompost were filtered, sterilized and diluted ranging from 100-900 mg L-1. Nutrients were analyzed following …show more content…
Eq. 2
YEPS - Change in RPS yield, dS – time (number of days)
Extraction of released polysaccharide (RPS)
Cyanobacterial cultures were centrifuged for 5000g for 10 minutes. The pellet was discarded and the supernatant was concentrated 10 times in oven at 40°C. To the solution three volumes of chilled absolute ethanol was added and kept overnight at -20°C (Jittawuttipoka et al. 2013). The white gel-like precipitate was separated by centrifuging at 10,000rpm for 10 minutes and lyophilized.
Biochemical characterization of crude released polysaccharide (RPS)
Crude RPS separated by centrifugation was washed twice with distilled water and was dissolved in 1mL of distilled water. The sample thus obtained was estimated for total carbohydrate by Anthrone method (Spiro 1966).
Monosaccharide analysis
Sample preparation for the monosaccharide analysis was followed as mentioned in El-Rassi (2002).In brief, lyophilized EPS samples (1.5 mg) was taken in 0.5 mL of methanolic HCl solution(prepared by adding 0.4 mL acetyl chloride) to 15mL of methanol. Methanolysis was performed at 80°C for 24h. It was followed by N-acetylation using acetic anhydride and trimethylsylilation. 1μl of sample was injected into the GC-MS (Perkin Elmer ClarusSQ 8 GCMS with autosampler) using autosampler. GC-MS conditions used was- Column : RTX-5MS 30M, 0.32mm ID, Gradient conditions: (Total run time: 30 min), Carrier gas: Helium at a flow rate of 1ml min-1. Initial temperature was 0°C min-1 to 100°C with hold time 1min. Ramp 1 was 8°C min-1 to 260°C min-1 with hold time 5min and Ramp 2 was 10°C min-1 to 300°C min-1 with no hold