A method for preparing urokinase

Abstract

The invention relates to a urinary protein enriching method which comprises the steps of directly adsorbing urinary proteins in urine in urinals or urinating buckets with filter cloth bags loaded with modified silica gel, and transporting the urinary protein adsorbed filter cloth bags to working points for subsequent treatment. According to the method, the isoelectric point property of specific urinary proteins is utilized, and urinary proteins such as urinary trypsin inhibitors, human urinary kininogenase, urokinase and the like are effectively absorbed directly by using the modified silica gel, macroporous resin, chitin, ionic resin and the like, so that a urine collecting step is avoided. The method is free of obvious influence on sanitary conditions of toilets, and the cost for urine transportation and a series of environmental problems resulting from urine transportation are greatly reduced.

Description

technical field

[0001] The invention belongs to the field of separation and purification, in particular to a method for preparing urokinase. Background technique

[0002] Urokinase is an alkaline protease that is produced by the kidneys and mainly exists in the urine of humans and mammals. It is a normal function of the human body when blood coagulates. However, under certain pathological conditions, blood acid is formed in the blood vessels to block blood circulation. Urokinase is a thrombolytic drug, but it is not directly used on the blood clot itself, but first activates plasmin in the body to become active plasmin, and plasmin acts on plasmin clot, causing it to break down into soluble polycarbonates.

[0003] Urokinase has various molecular weight forms, and the clinical effect of urokinase with high molecular weight on dissolving thrombus is about twice that of low molecular weight one. Therefore, the molecular weight of urokinase is regarded as one of the quality st...

Examples

Embodiment 1

[0034] Phenylamidated functional silica gels are prepared by the following method:

[0035] 1) After soaking the silica gel in hydrochloric acid solution, wash until there is no chloride ion, and dry;

[0036] 2) After activating the dried silica gel in step (1) at 200-300°C, add 2-chloro-6-methylaniline and phenyltrichlorosilane for reflux reaction, filter, precipitate, wash, and dry to obtain the precursor;

[0037] 3) Dissolve the precursor and 8.0mL pyridine in 70.6mL THF, place in an ice-water bath, control the temperature between 0-5°C, and slowly add 10.000g, 74.4mmol 3-ethoxyacryloyl chloride to the solution , about 15 minutes to finish dripping. The mixture was warmed to 20°C and stirred at this temperature for 2 hours, then the three-necked flask was placed in an ice-water bath (0-10°C), 13.5mL of 1mol / L hydrochloric acid was added, 35mL of water was added, and the solution was distilled under reduced pressure to Concentrate to a thick slurry, cool to room temperat...

Embodiment 2

[0040] A method for preparing urokinase, comprising:

[0041] 1) Place the filter cloth bag directly in the urinal or urinal of the toilet. The filter cloth bag includes a cotton gauze layer, a modified cellulose layer and a modified silica gel layer arranged sequentially from outside to inside. Modified silica gel is phenyl amidated functional group silica gel;

[0042] 2) Urine flows through the filter bag, and urine protein is absorbed;

[0043] 3) Collect the filter cloth bags that absorb urine protein and transport them to the processing point for subsequent processing.

Embodiment 3

[0044] Example 3 Collect the filter cloth bags that absorb urine protein and transport them to the processing point for subsequent treatment

[0045]Transport to the processing point for subsequent processing, including the following steps:

[0046] 1) Collect the adsorbent for adsorbing urinary protein, dissolve it with 400ml of pH6.0, 0.1mol / L phosphate buffer, stir for 30 minutes, let stand and pour out the supernatant, and then dissolve it with 400ml of 16% (w / v) ammonium sulfate , stirred for 30 minutes, and took the supernatant.

[0047] 2) Add 4.6g of ammonium sulfate, stir to dissolve, adjust the pH to 8.4 with 1N NaOH solution, and let stand for 2 hours;

[0048] 3) Filter paper to collect the precipitate, wash once with normal saline, and then wash with a certain concentration of EDTA solution to chelate Cu2+,

[0049] 4) Then add a certain amount of ammonia water to the precipitate to fully dissociate the urokinase in the precipitate to obtain a urokinase solution...

technical field

[0001] The invention belongs to the field of separation and purification, in particular to a method for preparing urokinase. Background technique

[0002] Urokinase is an alkaline protease that is produced by the kidneys and mainly exists in the urine of humans and mammals. It is a normal function of the human body when blood coagulates. However, under certain pathological conditions, blood acid is formed in the blood vessels to block blood circulation. Urokinase is a thrombolytic drug, but it is not directly used on the blood clot itself, but first activates plasmin in the body to become active plasmin, and plasmin acts on plasmin clot, causing it to break down into soluble polycarbonates.

[0003] Urokinase has various molecular weight forms, and the clinical effect of urokinase with high molecular weight on dissolving thrombus is about twice that of low molecular weight one. Therefore, the molecular weight of urokinase is regarded as one of the quality st...

Examples

Embodiment 1

[0034] Phenylamidated functional silica gels are prepared by the following method:

[0035] 1) After soaking the silica gel in hydrochloric acid solution, wash until there is no chloride ion, and dry;

[0036] 2) After activating the dried silica gel in step (1) at 200-300°C, add 2-chloro-6-methylaniline and phenyltrichlorosilane for reflux reaction, filter, precipitate, wash, and dry to obtain the precursor;

[0037] 3) Dissolve the precursor and 8.0mL pyridine in 70.6mL THF, place in an ice-water bath, control the temperature between 0-5°C, and slowly add 10.000g, 74.4mmol 3-ethoxyacryloyl chloride to the solution , about 15 minutes to finish dripping. The mixture was warmed to 20°C and stirred at this temperature for 2 hours, then the three-necked flask was placed in an ice-water bath (0-10°C), 13.5mL of 1mol / L hydrochloric acid was added, 35mL of water was added, and the solution was distilled under reduced pressure to Concentrate to a thick slurry, cool to room temperat...

Embodiment 2

[0040] A method for preparing urokinase, comprising:

[0041] 1) Place the filter cloth bag directly in the urinal or urinal of the toilet. The filter cloth bag includes a cotton gauze layer, a modified cellulose layer and a modified silica gel layer arranged sequentially from outside to inside. Modified silica gel is phenyl amidated functional group silica gel;

[0042] 2) Urine flows through the filter bag, and urine protein is absorbed;

[0043] 3) Collect the filter cloth bags that absorb urine protein and transport them to the processing point for subsequent processing.

Embodiment 3

[0044] Example 3 Collect the filter cloth bags that absorb urine protein and transport them to the processing point for subsequent treatment

[0045]Transport to the processing point for subsequent processing, including the following steps:

[0046] 1) Collect the adsorbent for adsorbing urinary protein, dissolve it with 400ml of pH6.0, 0.1mol / L phosphate buffer, stir for 30 minutes, let stand and pour out the supernatant, and then dissolve it with 400ml of 16% (w / v) ammonium sulfate , stirred for 30 minutes, and took the supernatant.

[0047] 2) Add 4.6g of ammonium sulfate, stir to dissolve, adjust the pH to 8.4 with 1N NaOH solution, and let stand for 2 hours;

[0048] 3) Filter paper to collect the precipitate, wash once with normal saline, and then wash with a certain concentration of EDTA solution to chelate Cu2+,

[0049] 4) Then add a certain amount of ammonia water to the precipitate to fully dissociate the urokinase in the precipitate to obtain a urokinase solution...