Objective:To establish a method for the separation and purification of haemocoagulase from agkistrodon blomhoffii in Changbai Mountain. Methods: An enzyme component with clotting activity was isolated from the venin of agkistrodon blomhoffii in Changbai Mountain by gel filtration, anion-exchange chromatography and heparin-Sepharose affinity chromatography. SDS-Page and RP-HPLC were used to determine its purity, SDS-Page was applied to study its action mode with bovine fibrinogen, HPSEC was used to determine the molecular weight, an IEF method was employed to detect its isoelectric point, and Lowry method was used for the deter-mination of protein concentration. Results:One haemocoagulase was purified from agkistrodon in Changbai Mountain. SDS-Page dis-played one band, and HPLC showed one single chromatographic peak. The haemocoagulase acted only on α chain of fibrinogen. Its molecular weight was 32. 2kD with isoelectric point of 5. 21. The enzyme had clotting activity in vitro. Conclusion:The method can be used for the separation and purification of haemocoagulase from agkistrodon blomhoffii in Changbai Mountain.

Objective:To establish a method for the determination of content and related substances of troxipite tablets by HPLC. Methods:A Waters Symmetry-C18 (150 mm × 4. 6 mm,5 μm) column was used. The mobile phase was methanol -0. 4% phosphoric acid solution (50∶50). The flow rate was 1. 0 ml·min-1. The detection wavelength was 260nm. The column temperature was 35℃and the injection volume was 20 μl. Results:The linear range of troxipite was 3-75 μg·ml-1(r=0.999 7). The average recovery was 99. 7%,RSD=0. 95%(n=9). Conclusion:The method is simple, accurate and specific, and can be used in the quality control of troxipite tablets.

OBJECTIVE:To establish a method for the determination of related substances in Methylphenidate hydrochloride for injection. METHODS:HPLC method was adopted. The determination was performed on Waters symmetry C18 column with mo-bile phase consisted of methanol-0.01 mol/L potassium dihydrogen phosphate solution(60:40,V/V)at the flow rate of 1.0 mL/min. The detection wavelength was set at 210 nm,the column temperature was 35 ℃ and sample size was 10 μL. RESULTS:The linear range of impurity A and B were 0.02-3.0 μg/mL(r=0.9998). The limits of quantitation were 0.2,0.6 ng,and the limits of detec-tion were 0.06,0.2 ng,respectively. RSDs of precision,stability and reproducibility were all lower than 2.0%;recoveries were 98.2%-100.0%(RSD=0.56%,n=9),98.0%-100.3%(RSD=0.70%,n=9),respectively. CONCLUSIONS:The method is sim-ple,accurate and suitable for the determination of related substance in Methylphenidate hydrochloride for injection.