*3.2.4 Scanning electron microscopy (SEM) analysis*

The surface morphology and characteristics of the synthesized nanoparticle were observed using Scanning Electron Microscopy (SEM) according to the protocol mentioned by [21]. Images were taken by the model ZEISS SEM and performed at a beam accelerating voltage of 20 kV.

**Figure 2.**

*Sequential step for RPS extraction from P. tenue.*

*Green Synthesis of Silver Nano-Particle from Cyanobacteria and Effect on Microalgal Growth… DOI: http://dx.doi.org/10.5772/intechopen.106039*

#### **3.3 Studies with the effect of silver nanoparticles**

Silver nanoparticles synthesized from *P. tenue* were first filtered through a 0.22 μm membrane filter and then integrated into the control culture medium (ASN-III medium) of *Phormidium tenue* culture at different concentrations. The effects on the growth rate, biomass production, and EPS production were then monitored according to the protocol mentioned above. The total biomass yield and EPS yield were analyzed at regular intervals.

### **3.4 Extraction of EPS**

#### *3.4.1 Extracellular or released polysaccharides (RPS)*

A known volume of culture was centrifuged at 10,000 rpm for 15 min. The supernatant so obtained was used for the extraction of released polysaccharides by addition of a measured volume of extraction solvent (acetone) followed by incubation at 4°C for 48 h. The released polysaccharide was then precipitated and collected by centrifugation at 10,000 rpm for 10 minutes. The pellet thus obtained was freeze drier (**Figure 2**).

### *3.4.2 Cell-bound or capsular polysaccharides (CPS)*

A known amount of culture was centrifuged at 10,000 rpm for 10 min. The supernatant was discarded, and the pellet obtained was used to estimate capsular polysaccharides. It was carried out by the addition of 36.5% formaldehyde (0.06 mL) to the pellet, followed by incubation for 1 hour at 4°C [8], after which 60 mL of 1 N NaOH was introduced and further kept for incubation at 4°C for 3 hours. The treated sample was then centrifuged, and capsular polysaccharides

**Figure 3.**

*Sequential step for CPS extraction from Phormidium tenue.*

in that supernatant were extracted by adding acetone (10 mL) and incubated for 48 hours at 4°C. The capsular polysaccharides were precipitated by centrifugation and freeze drier (**Figure 3**).
