[Human thrombin: enzymatic properties, stability and standardization of preparation].
The work deals with estimation of thrombin preparation having such features as: sedimentation activity 3000-3200 NIH un. per 1 mg of protein and 97% of active centres. The enzyme isolated has been estimated according to the amidolytic activity on synthetic substrates S-2160 and BAPNA being equal 5200 and 185 milli un/mg of protein, respectively. According to the electrophoresis in PAAG in the presence of Ds-Na the preparation is homogenous, its molecular mass is 36000. The fibrinogen sedimentation time dependence on the isolated thrombin concentration has been estimated as well as the comparative analysis with the thrombin of the firm "Sigma" with the previously calibrated activity using the international standartion (coded P4) has been conducted. The absence of proportionality between the substrate sedimentation time and the preparation concentration has been determined. It has been revealed, that if the experimental findings are presented in the units 1/t against the thrombin units NIH the right lines are received within the limits used. The defreezing and secondary freezing of the preparation preserved under -20 degrees C have been showed as rendering an essential effect on thrombin activity. In order of the enzyme stabilizing at preserving the thrombin isolated has been concentrated applying the amycon membranes (MWCo: 30,000). While applying the thrombin water-saline solution in the conditions selected the preparation has showed itself practically stable during a year without utilizing any admixtures. The essential effect on thrombin has been found from the side of 1% glycin, 0.5% PEG, 1% saccharose and so on. The thrombin isolated high functional homogeneity, its stability permit to recommend the preparation as an operative standard.
Kolodzeĭskaia
M V
MV
Palladin Institute of Biochemirtsy, NAS of Ukraine, Kyiv.
Chernyshenko
T M
TM
D001665:Binding Sites; D004356:Drug Storage; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005340:Fibrinogen / Q000378:metabolism; D005612:Freeze Drying; D006801:Humans; D008970:Molecular Weight; D012015:Reference Standards; D012964:Sodium / Q000378:metabolism; D012996:Solutions; D013379:Substrate Specificity; D013917:Thrombin / Q000302:isolation & purification / Q000378:metabolism* / Q000592:standards*; D013997:Time Factors; D014867:Water
D012996:Solutions; D014867:Water; D005340:Fibrinogen; D012964:Sodium; D013917:Thrombin