2.2. Chemical analysis
All samples were filtered through 0.2 μm pore size membranes (Minisart, Sartorius stedim biotech, Germany) before analysis. The concentrations of TN and NH3–N were determined using second-derivative method. The concentration of TP was measured according to the ascorbic PR171 method prescribed in standard methods (AWWA, 1998). Culture pH and water-temperature were measured using a pH electrode (pH 3110 SET 2/SenTix® 41, WTW, Germany). The COD was measured according to standard methods (AWWA, 1998).
2.3. Isolation and identification of microalgae
2.4. Estimation of biovolume of microalgal strains
Biovolume of each strain isolated and identified as mentioned above was calculated every week by volumetric method (Hillebrand et al., 1999). Observations were made using a growth rings light microscope (Nikon F) with magnification of 200, 400 and 1000×, directly connected to the camera and computer. ImageJ software was used to measure the cell number, length, width and height of microalgae. Consequently, volume of each microalga was calculated using Microsoft Excel 2010 (Park et al., 2013 and Kim et al., 2014b).