This research was concerned with the fiber separation and non-woven preparation form sugar canes leaves. The separation process was carried out by chemical pulping method at 100 degree Celsius. In order to study the suitable condition for separation process the concentration of sodium hydroxide ranging from 0 -20 grams per liter and boiling time 15 30 45 60 75 90 105 and 120 minutes were performed. Web formation process was performed by wet laid method and polyvinyl alcohol was selected as binder in web bonding process. in order to study the suitable amount of binder in bonding process concentration of polyvinyl alcohol ranging from 1 2 4 6 8 and 10 % w/v coated on web surface and cured at 120 degree Celsius. Bursting strength as well as physical properties evaluation of non-woven fabrics was performed. For antimicrobial finishing process was performed by spray method and pad method. The finishing result was evaluated by MTCC test method 147-1998 (Qualitative) and 100 -1999 (Quantitative) The result was found that appropriate condition for fiber separation was recommended as follows: the amount of sodium hydroxide of 10 grams per liter and boiling time 60 minutes. The suitable amount of polyvinyl alcohol for web bonding process was 2 % w/v. Bursting strength of non-woven fabric was 53 kN/m 3 and padding method was a suitable method for antimicrobial finishing (antimicrobial area 0.3 millimeters )