OBJECTIVE:To set up a method for determination of molecular weight and content of polysaccharide in Bletilla striata. METHODS:HPSEC-ELSD method was adopted. The determination was performed on TSK-GEL G4000 PWXL column with mobile phase consisted of pure water at the flow rate of 0.6 mL/min.The column temperature was 30 ℃,and sample size was 20 μL. Evaporative light scattering detector was used.The molecular weight and content of polysaccharide in 3 batches of B. striata were established. RESULTS:The linear range of weight average molecular weight of B. striata polysaccharide were 24.17-178.00 kD(R2=0.985 5). The linear range of BT07 content determination were 0.508 0-5.080 mg/mL(R2=0.998 4). The limits of detection and quantitation were 0.116 5 and 0.274 0 mg/mL. RSDs of precision,stability and reproducibility tests were all lower than 2%(n=6 or n=7).Average recoveries rate were 99.16%-100.20%(RSD=0.39%-0.64%,n=9).Average retention time of 3 batches of samples was 14.28 min(RSD=4.35%,n=3),and weight average molecular weight was 23.54 kD(RSD=3.78%, n=3). Polydispersity coefficient D,MW/Mn)ranged 1.463-1.578. Average content of sample was 93.4%(RSD=4.22%,n=3). CONCLUSIONS:HPSEC-ELSD method is simple,rapid,accurate and reliable,which can be used for simultaneous determination of B.striata polysaccharide.