| 摘要 |
FIELD: medicinal industry. ^ SUBSTANCE: invention relates to a method for isolating biologically active substance from mammalian bladder and preparing a medicinal formulation for parenteral administration that can be used in medicine as an agent normalizing bladder tonus. Agent normalizing bladder tonus is made as a medicinal formulation for parenteral administration and represents peptide complex with the content of low-molecular fraction from 70% to 90%, molecular mass of its peptide components in the range from 74 to 394 Da and concentration of polypeptides 2.5-2.9 mg/ml. Agent is prepared from calf bladders (age is 12 months, not above) or pigs by extraction of tissue with acetic acid in the presence of zinc chloride. The proposed method for preparing agent normalizing bladder tonus from calves (age is 12 months, not above) or pigs involves defrosting tissue at temperature -40°C (not less), keeping at temperature -20-22°C for two months (not less) followed by milling and adding 3% acetic acid solution in the volume ratio = 1:5 at temperature 20 ± 5°C. Extraction is carried out at constant stirring and 1% zinc chloride solution is added to the prepared homogenous suspension in the volume ratio = 50:1 followed by cooling at constant stirring up to temperature 7-16°C and the following stirring for 1 h in each 4 h in settling for 48 h. Extract is separated from inert substances by separating, acetone is added to extract in the volume ratio = 1:5 and kept at temperature 3-5°C for 4 h. Formed homogenized deposit is precipitated repeatedly with acetone twice (not less) and deposit containing active substance is washed out on Nutch filter with two-fold volumes of acetone cooled to temperature 7-16°C up to preparing light-gray deposit. Deposit is rubbed through metallic sieve, dried, dissolved in distilled water at room temperature and constant stirring up to the concentration of polypeptides 2.5-2.9 mg/ml. Solution is centrifuged, filtered and subjected for ultrafiltration treatment in device under anti-pressure 1.0 kgf/cm2 (not above) through materials with retaining capacity 15000 Da. Glycocol is added to ultrafiltrate up to its final concentration 10-20 mg/ml at pH = 5.6-6.6, solution is subjected for sterilizing filtration under pressure 2.0 kgf/cm2 (not above), poured into ampoules in volume 2 ml and subjected autoclaving at temperature 120°C for 8 min and under atmosphere pressure 1.1 kgf/cm2. Invention provides optimal technology in isolating peptide complex with the content of low-molecular fraction from 70% to 90%, molecular mass of its peptide components in the range from 74 to 394 Da, and preparing aqueous solution of extract with the concentration of polypeptides 2.5-2.9 mg/ml/ Invention proves both purification of prepared product from impurities and to enhance its yield. The isolated substance differs from the known substances prepared early from mammalian raw by molecular mass of its peptide components, absence of toxicity and apyretic properties based on the complete removing impurities. ^ EFFECT: improved preparing method, valuable medicinal properties of agent. ^ 3 cl, 2 tbl, 2 dwg, 5 ex |
