CN103333240B - Method for reclaiming human albumin from component IV precipitate - Google Patents
Method for reclaiming human albumin from component IV precipitate Download PDFInfo
- Publication number
- CN103333240B CN103333240B CN201310307858.8A CN201310307858A CN103333240B CN 103333240 B CN103333240 B CN 103333240B CN 201310307858 A CN201310307858 A CN 201310307858A CN 103333240 B CN103333240 B CN 103333240B
- Authority
- CN
- China
- Prior art keywords
- component
- precipitate
- liquid
- fraction
- pressure filter
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Abstract
The invention discloses a method for reclaiming human albumin from a component IV precipitate. Methods for reclaiming the human albumin from the component IV precipitate are heat treatment methods in the prior art, the heat treatment methods are low in recovery rate, all proteins except the albumin are denatured in the recovery process and other substances cannot be extracted from other proteins, so that the extraction cost is increased. According to the method, a component V is effectively separated from the component IV precipitate and is prepared into a human albumin finished product by steps of dissolving and purifying the component IV precipitate, preparing a component V precipitate, refining and purifying the component V, performing ultrafiltration, preparing a semi-finished product, performing Pasteur inactivation and the like. Impure proteins can be removed by a low temperature ethanol precipitation method, the coprecipitation of the human albumin is reduced, the yield of finished product is improved, the denaturation of the proteins is decreased, and the stability of the finished product is improved. After the human albumin is extracted, other components can be extracted, the comprehensive utilization of blood plasma is improved, and the method is a novel method for extracting the human albumin.
Description
Technical field
What the present invention relates to is the method reclaiming human serum albumin from fraction IV precipitate.Belong to the preparing technical field of blood products.
Background technology
Human serum albumin is a kind of blood products; extract from human plasma; the Q volume of blood of patient can be increased and maintain plasma colloid osmotic pressure; there is very important effect in loss in treatment in a large number because the reasons such as burn cause body fluid; in addition, human serum albumin also plays a part nutrition supply, transport and removing toxic substances and to the colloid of other plasma proteins protection and stable.
Each blood products manufacturer all suitably improves production human serum albumin with reference to Cohn6 method or Kistler-Nitchmann cold ethanol method in the world at present, this type of technology is cold ethanol fractionation precipitation technology, by effectively controlling five factors of protein precipitation: pH value, temperature, protein concn, ionic strength, alcohol concn precipitate the different proteins in blood plasma step by step, the step of general separation human serum albumin for first removing components I from blood plasma, remove compositionⅱ+III again, then remove component IV, finally precipitate extraction components V i.e. human serum albumin.The component IV produced in human serum albumin production process generally goes out of use as impurity, but because the influence factor in fractionation precipitation process is a lot, in traditional production technique, a certain amount of separated human serum albumin of failing is mingled with in fraction IV precipitate, also discarded simultaneously, cause the waste of material, also cause in short supply for manufacturer's seralbumin raw material, therefore, human serum albumin is reclaimed from discarded precipitation, the situation alleviating internal sources blood plasma in short supply seems very important, for this reason, notification number is CN1305903C, title is: a kind of from deposited components I, II, III, albuminous method is reclaimed in IV, and notification number is CN102311497A, title is: one cold ethanol method reclaims albuminous method from component IV, all provide from precipitation fraction I, II, III, albuminous method is reclaimed in IV, there is certain positive effect, but, the method provided is all heat treating process, by precipitation fraction I, II, III, IV dilution post-heating to 68 DEG C this stagnation point, make the equal sex change of all proteins except albumin, thus separate with albumin, extract albumin, but the rate of recovery is low on the one hand for heat treating process, the on the other hand equal sex change of all albumen in addition to albumin in removal process, can not therefrom extract other materials, extraction cost is increased.Therefore, seek a kind of more preferably do not need heat from precipitation fraction IV, reclaim albuminous method, be just provided with very important realistic meaning.
Summary of the invention
Object of the present invention is just to provide the method reclaiming human serum albumin from fraction IV precipitate, overcome the deficiency existing for prior art heat treating process, effectively solve and extract human serum albumin from component IV, to meet the needs of problems of people to human blood albumin products.
For achieving the above object, this invention takes following technical proposals, from fraction IV precipitate, reclaim the method for human serum albumin, comprise following concrete technological step:
(1) fraction IV precipitate dissolves
1 ~ 2 DEG C that fraction IV precipitate is put into 2 times of weight, in the sodium-chlor of 8 ~ 10g/L, stirring and dissolving, keep solution temperature-1 ~-2 DEG C, stirring and dissolving 5 ~ 6 hours, is fraction IV precipitate lysate.
(2) purifying
1. fraction IV precipitate lysate is squeezed in diluent and dilute, the amount of diluent is 3 ~ 4 times of fraction IV precipitate lysate, after dilution, goods are stirred and cool to-2 ~-3 DEG C, 1mol/L sodium bicarbonate is slowly added by the speed of 0.8 ~ 1.2L/ minute, adjustment goods pH value to 6.1 ~ 6.4, stir 6 ~ 8 hours under-2 ~-3 DEG C of conditions, obtain component IV dissolved solution for later use;
Wherein the preparation of diluent is: sodium chloride-containing 0.15mol/L, alcohol concn 27 ~ 30%(V/V), stir and cool to-2 ~-3 DEG C;
2. in component IV lysate, add the ethanol of less than-20 DEG C 95%, alcohol concn in component IV lysate is made to reach 40 ~ 42%(V/V), ethanol adds speed and remains to 70 ~ 80L/h, solution temperature-2 ~-3 DEG C is kept in adition process, ethanol finishes, with sodium bicarbonate adjustment solution ph to 6.6 ~ 6.7 of 1mol/L, cooling makes the temperature of component IV lysate to-5.5 ~-6 DEG C, and it is for subsequent use that constant temperature stirring obtains component IV diluent for 6 ~ 8 hours;
3. in component IV diluent, diatomite is added, add-on is every 100Kg fraction IV precipitate 3 ~ 5Kg diatomite, stir and use pressure filter press filtration after 30-40 minute, press filtration terminates rear cold reduction air and blows pressure filter intracavity liquid, to going out bubble, collect filtrate, obtain component IV pressing filtering liquid for subsequent use, precipitation is abandoned and is lost;
(3) preparation of component V precipitation
1. measure component IV pressing filtering liquid volume, open and stir, cooling, goods cool to-6 ~-7 DEG C, slowly add 2mol/L HAC, and adding speed is 50-60ml/min, goods are cooled to-8 ~-9 DEG C after having adjusted pH and stir 2 hours, quietly putting 6 ~ 8 hours by adjustment pH value of solution to 5.2 ~ 5.5;
2. open stirring, in component IV pressing filtering liquid, add diatomite, add-on adds 1 ~ 2Kg diatomite with every 100Kg fraction IV precipitate, stirs and starts press filtration after 30-40 minute, and filtrate abandoning is lost or carried out ethanol recycling after collecting; Collecting component V precipitates for subsequent use;
(4) component V polishing purification
1. the component V precipitation 8%(V/V of 5 times of volumes) ethanolic soln stirring and dissolving, after precipitation is dissolved completely, measuring pH value of solution should in pH to 4.50 ~ 4.70, and goods outlet temperature controls, for-3.0 DEG C ± 0.5 DEG C, to continue stirring press filtration in 2 ~ 3 hours;
2. press filtration: stir in pressure-filtering process and do not stop, after press filtration is complete, with liquid in low temperature cold Air blowing filter, stops after going out bubble, and rear FV refines balance liquid and rinses filter plate, and washing fluid is incorporated in filtered liquid that to obtain refined solution for subsequent use;
Wherein FV refines the preparation of balance liquid: containing ethanol 12%(V/V), solution ph 4.50 ~ 4. 70, is cooled to-3.0 DEG C ± 0.5 DEG C.
(5) ultrafiltration
1. stirring is opened, with 1M sodium bicarbonate adjustment refined solution pH value to 7.0 ~ 7.2;
2. by refined solution first ultrafiltration and concentration to protein concentration 100 ~ 150g/L, dialyse with the sodium chloride solution equal-volume of the 9g/L of 5 times, albumin stoste is obtained again with the water for injection equal-volume dialysis of 3 times, by final for albumin stoste ultrafiltration and concentration to more than protein content 210g/L, with appropriate water for injection washing ultra-filtration membrane, collect film washing liquid, be incorporated in above-mentioned protein liquid and be human serum albumin stoste, sampling is done and is carried out stoste detection;
(6) work in-process preparation
Dilution preparation is carried out: prepare according to stock protein matter content according to stoste calibrating, Sodium octoate is taken by every gram of protein 160 mmole, sodium-chlor is taken by every gram of protein 120 ~ 130 mmole, dissolve with a small amount of water for injection after taking, add human serum albumin stoste, again human serum albumin stoste is diluted to protein concn 195 ~ 200g/L, adjusted to ph is 6.8 ~ 7.0;
(7) pasteurization: goods are carried out 60 DEG C of 10 hours inactivation of viruses, degerming packing after deactivation.
Described diatomite refers to through 180 DEG C dry roasting 2.5 hours, is cooled to the diatomite of less than 0 DEG C.
Take the present invention of above-mentioned measure, adopt multistep ethanol precipitation and remove foreign protein, purification human serum albumin by parameters such as adjustment ionic strength, pH value, protein content, temperature.Chilled alcohol precipitation method effectively can remove foreign protein, reduces the coprecipitated of human serum albumin, improves product yield, reduces the sex change of protein, increases the stability of finished product.The present invention is compared with existing recovery method heat treating process, and it has clear superiority, and (1) cold ethanol method of the present invention, to both at home and abroad to produce production unit that human serum albumin adopts now similar, does not therefore need newly added equipment when large-scale recovery; (2) rate of recovery is high, traditional heat treating process, and its rate of recovery is 0.4 ~ 0.8Kg albumin/ton blood plasma, and the rate of recovery of the present invention is 1.0 ~ 1.2Kg albumin/ton blood plasma, and the rate of recovery improves 100%.(3) in conventional recovery method, all after FIV heating method, carry out cold ethanol method extraction again, its FIV sex change after heating, can not therefrom extract other materials, and the present invention utilizes classical cold ethanol method, after FIV is extracted human serum albumin, still can extract other components, this is the place of the present invention than traditional method advanced person.The present invention can effectively reduce the cost of the raw blood plasma in blood products production, improves the comprehensive utilization of blood plasma, is extract the innovation on human serum albumin, has huge economic and social benefit.
The present invention can effectively isolate component V and then be mixed with human serum albumin finished product from fraction IV precipitate, the situation alleviating internal sources blood plasma in short supply is of great practical significance, both adequately achieve making full use of raw material, prevent again the pollution to environment.
Human serum albumin stoste prepared by the present invention, can be packed as human serum albumin finished product after work in-process preparation and inactivation of virus, as the most important middle product producing human serum albumin, its quality meets following requirement:
1. purity: detect by cellulose acetate membrane electrophoresis method in " Chinese Pharmacopoeia " version in 2010 three annex, its purity should be not less than 96%;
2. protein content: detect by contracting method urea method two in " Chinese Pharmacopoeia " version in 2010 three annex, should 210g/L be greater than;
3. pH value: with physiological sodium chloride solution, trial-product protein content is diluted to 10g/L, pH value should be 6.4 ~ 7.4.
4. residual ethanol content: measure by Kang Wei diffusion boat method in " Chinese Pharmacopoeia " version in 2010 three annex, should not higher than 0.025%.
5. pyrogen test: after component V resolution of precipitate, undertaken by " Chinese Pharmacopoeia " version in 2010 three annex Ⅹ II E " bacterial endotoxins test ", its Bacterial endotoxin limit should be less than 1.67EU/ml.
Four, embodiment
Embodiment 1
The technological step of the embodiment of the present invention 1 is as follows:
(1) 1 ~ 2 DEG C that fraction IV precipitate is put into 2 times of weight, in the sodium chloride solution of 9g/L, stirring and dissolving, keep solution temperature-1 ~-2 DEG C, stirring and dissolving 6 hours, is fraction IV precipitate lysate.
(2) fraction IV precipitate lysate is added in diluent and dilute, the amount of diluent is 3.2 times of fraction IV precipitate lysate, after dilution, goods are stirred and cool to-2 ~-3 DEG C, 1mol/L sodium bicarbonate is slowly added by the speed of 1L/ minute, adjustment goods pH value to 6.25, stirs under-2 ~-3 DEG C of conditions and obtains fraction IV precipitate diluent in 7 hours;
The preparation of diluent: sodium chloride-containing 0.15mol/L, alcohol concn 28%(V/V), stir and cool to-2 ~-3 DEG C.
(3) in fraction IV precipitate diluent, add the ethanol of less than-20 DEG C 95%, make alcohol concn in fraction IV precipitate diluent reach 40.5%(V/V), ethanol adds speed and remains to 70 ~ 80L/h, keeps solution temperature-2 ~-3 DEG C in adition process.Ethanol finishes, adjustment solution ph to 6.65, and cooling makes fraction IV precipitate diluent temperature to-5.5 ~-6 DEG C, stirs and within 6 hours, obtains fraction IV precipitate alcohol dilution liquid.
(4) in fraction IV precipitate alcohol dilution liquid, add diatomite, add-on is every 100Kg fraction IV precipitate 4Kg diatomite, stirs and uses pressure filter press filtration after 30 minutes.Press filtration terminates rear cold reduction air and blows pressure filter intracavity liquid, stops after going out bubble.Pump into-6 ~-7 DEG C of FIV balance liquids and rinse pressure filter, every platform rinses 150L, finally blows goods in pressure filter chamber with cold reduction air.Collect filtrate, obtain component IV pressing filtering liquid, precipitation is abandoned.
Diatomaceous preparation: 180 DEG C are dry roasting 2.5 hours, be cooled to less than 0 DEG C for subsequent use.
The preparation of FIV balance liquid: amount of preparation is every platform pressure filter 600L.Compound method: containing 40%(V/V) ethanol, the sodium-chlor of 2g/L, the anhydrous sodium acetate of 4.8g/L, by acetic acid adjusted to ph to 6.0, solution temperature-6 ~-7 DEG C.
The preparation of pressure filter: every 100Kg fraction IV precipitate fills 25 filter plates, after installing filter plate, it is negative for rinsing pressure filter to bacterial endotoxin with water for injection.In FIV balance liquid, add diatomite with the amount of every platform pressure filter 5Kg, pumped into pressure filter and carry out circulating cooling after stirring, when pressure filter temperature out is to less than-3 DEG C, stop circulation, freezing air blowout pressure filter chamber inner equilibrium liquid is for subsequent use.
(5) measure component IV pressing filtering liquid volume, open and stir, cooling, goods cool to-6 ~-7 DEG C, slowly add 2mol/L HAC, and adding speed is 50ml/min, adjustment pH value of solution to 5.35, after having adjusted pH, component IV pressing filtering liquid is cooled to-8 ~-9 DEG C, stirs 2 hours, quiet putting obtains frozen composition IV pressing filtering liquid for 6 hours;
(6) open stirring, in frozen composition IV pressing filtering liquid, add diatomite, add-on adds 1Kg diatomite with every 100Kg fraction IV precipitate, stirs and starts press filtration after 30 minutes.Filtered liquid carries out ethanol recycling after abandoning or collecting; After press filtration, unclamp pressure filter, scraping component V precipitates.
Diatomaceous preparation: 180 DEG C are dry roasting 2.5 hours, be cooled to less than 0 DEG C for subsequent use.
The preparation of pressure filter: every 100Kg fraction IV precipitate installs filter plate 6, it is negative for rinsing to bacterial endotoxin with water for injection after installing filter plate.In FV balance liquid, add diatomite, add-on is every platform pressure filter 3Kg, adds the preparation circulation pressure filter of rear FV balance liquid, and ensure that pressure filter temperature out should reach less than-5 DEG C, cold reduction air blows pressure filter inner equilibrium liquid, stops after going out bubble, for subsequent use.
The preparation of FV balance liquid: amount of preparation is every platform pressure filter 400L.Compound method: water for injection 230L, adds 95% ethanol 170L, cools to-8 ~-9 DEG C.
(7) component V precipitation 8% ethanolic soln stirring and dissolving of 5 times of volumes, after precipitation is dissolved completely, adjustment pH to 4.50, goods outlet temperature controls as-3.0 DEG C ± 0.5 DEG C, continues stirring and within 2 hours, obtains component V resolution of precipitate liquid.
(8) press filtration: get the filter of component V resolution of precipitate hydraulic pressure, stir in process and do not stop, after press filtration is complete, with liquid in low temperature cold Air blowing filter, stop after going out bubble, rear F V refines balance liquid and rinses filter plate, and washing fluid is incorporated in filtered liquid and obtains component V sedimentation and filtration liquid.
The preparation of pressure filter: every 100Kg component V precipitates installs filter plate 15, it is negative for rinsing to bacterial endotoxin with water for injection after installing filter plate.With F V polishing purification balance liquid circulation pressure filter before using, make pressure filter temperature out should reach less than-2 DEG C for subsequent use.
F V refines balance liquid preparation: amount of preparation: every platform pressure filter 250L.Compound method is: containing ethanol 12%, solution ph 4.50, is cooled to-3.0 DEG C ± 0.5 DEG C.
(9) open stirring, obtain component V deposition and purification liquid by 1M sodium bicarbonate adjustment component V sedimentation and filtration liquid pH value to 7.05.
(10) component V deposition and purification liquid is concentrated into protein concentration 100g/L first, first dialyse with the sodium chloride solution equal-volume of the 9g/L of 5 times, obtain albumin stoste with the water for injection equal-volume dialysis of 3 times again, albumin stoste is finally concentrated into more than protein content 210g/L.With appropriate water for injection washing ultra-filtration membrane, collect film washing liquid, be incorporated in above-mentioned albumin concentrated liquor and be human serum albumin stoste, stoste detection is carried out in sampling.
(11) work in-process preparation: carry out dilution preparation according to the calibrating of human serum albumin stoste: prepare according to human serum albumin stock protein matter content, Sodium octoate is taken by every gram of protein 160 mmole, sodium-chlor is taken by every gram of protein 120 mmole, dissolve with a small amount of water for injection after taking, add in human serum albumin stoste, again human serum albumin stoste is diluted to protein concn 196g/L, adjusted to ph obtains human serum albumin solution 6.85.
(12) pasteurization: human serum albumin solution is carried out 60 DEG C of 10 hours inactivation of viruses, degerming packing after deactivation.
Embodiment 2
The technological step of the embodiment of the present invention 2 is as follows:
(1) 1 ~ 2 DEG C that fraction IV precipitate is put into 2 times of weight, in the sodium chloride solution of 10g/L, stirring and dissolving, keep solution temperature-1 ~-2 DEG C, stirring and dissolving 5 hours, obtains fraction IV precipitate lysate.
(2) fraction IV precipitate lysate is added in diluent and dilute, the amount of diluent is 3 times of fraction IV precipitate lysate, after dilution, goods are stirred and cool to-2 ~-3 DEG C, 1mol/L sodium bicarbonate is slowly added by the speed of 1L/ minute, adjustment fraction IV precipitate lysate pH value to 6.4, stirs 6 hours component IV and precipitates diluent under-2 ~-3 DEG C of conditions;
The preparation of diluent: sodium chloride-containing 0.15mol/L, alcohol concn 29%(V/V), stir and cool to-2 ~-3 DEG C.
(3) in fraction IV precipitate diluent, add the ethanol of less than-20 DEG C 95%, make the alcohol concn of fraction IV precipitate diluent reach 41.5%(V/V), ethanol adds speed and remains to 70 ~ 80L/h, keeps solution temperature-2 ~-3 DEG C in adition process.Ethanol finishes, adjustment solution ph to 6.7, and cooling makes products temperature to-5.5 ~-6 DEG C, stirs and within 8 hours, obtains fraction IV precipitate alcohol dilution liquid.
(4) in fraction IV precipitate alcohol dilution liquid, add diatomite, add-on is every 100Kg fraction IV precipitate 5Kg diatomite, stirs and uses pressure filter press filtration after 30 minutes.Press filtration terminates rear cold reduction air and blows pressure filter intracavity liquid, stops after going out bubble.Pump into-6 ~-7 DEG C of FIV balance liquids and rinse pressure filter, every platform rinses 200L, finally blows goods in pressure filter chamber with cold reduction air.Collect filtrate, obtain component IV pressing filtering liquid, precipitation is abandoned.
Diatomaceous preparation: 180 DEG C are dry roasting 2.5 hours, be cooled to less than 0 DEG C for subsequent use.
The preparation of FIV balance liquid: amount of preparation is every platform pressure filter 600L.Compound method: containing 40%(V/V) ethanol, the sodium-chlor of 2.5g/L, the anhydrous sodium acetate of 4.5g/L, by acetic acid adjusted to ph to 6.1, solution temperature-6 ~-7 DEG C.
The preparation of pressure filter: every 100Kg fraction IV precipitate fills 25 filter plates, after installing filter plate, it is negative for rinsing pressure filter to bacterial endotoxin with water for injection.In FIV balance liquid, add diatomite with the amount of every platform pressure filter 5Kg, pumped into pressure filter and carry out circulating cooling after stirring, when pressure filter temperature out is to less than-3 DEG C, stop circulation, freezing air blowout pressure filter chamber inner equilibrium liquid is for subsequent use.
(5) measure component IV pressing filtering liquid volume, open and stir, cooling, component IV pressing filtering liquid cools to-6 ~-7 DEG C, slowly adds 2mol/L HAC, and adding speed is 50ml/min, adjustment pH value of solution to 5.45, after having adjusted pH, component IV pressing filtering liquid is cooled to-8 ~-9 DEG C, stirs 2 hours, quiet putting obtains frozen composition IV pressing filtering liquid for 7 hours;
(6) open stirring, in frozen composition IV pressing filtering liquid, add diatomite, add-on adds 2Kg diatomite with every 100Kg fraction IV precipitate, stirs and starts press filtration after 30 minutes.Filtered liquid carries out ethanol recycling after abandoning or collecting; After press filtration, unclamp pressure filter, scraping component V precipitates.
Diatomaceous preparation: 180 DEG C are dry roasting 2.5 hours, be cooled to less than 0 DEG C for subsequent use.
The preparation of pressure filter: every 100Kg fraction IV precipitate installs filter plate 8, it is negative for rinsing to bacterial endotoxin with water for injection after installing filter plate.In FV balance liquid, add diatomite, add-on is every platform pressure filter 3Kg, adds the preparation circulation pressure filter of rear FV balance liquid, and ensure that pressure filter temperature out should reach less than-5 DEG C, cold reduction air blows pressure filter inner equilibrium liquid, stops after going out bubble, for subsequent use.
The preparation of FV balance liquid: amount of preparation is every platform pressure filter 400L.Compound method: water for injection 230L, adds 95% ethanol 170L, cools to-8 ~-9 DEG C.
(7) component V precipitation 8% ethanolic soln stirring and dissolving of 5 times of volumes, after precipitation is dissolved completely, adjusts pH to 4.70, and outlet temperature controls as-3.0 DEG C ± 0.5 DEG C, continues stirring press filtration in 3 hours and obtains component V resolution of precipitate liquid.
(8) press filtration: get the filter of component V resolution of precipitate hydraulic pressure, stir in process and do not stop, after press filtration is complete, with liquid in low temperature cold Air blowing filter, stop after going out bubble, rear F V refines balance liquid and rinses filter plate, washing fluid is incorporated in filtered liquid, obtains component V sedimentation and filtration liquid.
The preparation of pressure filter: every 100Kg component V precipitates and installs filter plate 20, it is negative for rinsing to bacterial endotoxin with water for injection after installing filter plate.Refine balance liquid circulation pressure filter with F V before using, make pressure filter temperature out should reach less than-2 DEG C for subsequent use.
FV refines balance liquid preparation: amount of preparation: every platform pressure filter 250L.Compound method is: containing ethanol 12%, solution ph 4. 70, is cooled to-3.0 DEG C ± 0.5 DEG C.
(9) open stirring, obtain component V deposition and purification liquid by 1M sodium bicarbonate adjustment component V sedimentation and filtration liquid pH value to 7.15.
(10) component V deposition and purification liquid is concentrated into protein concentration 150g/L first, first dialyse with the sodium chloride solution equal-volume of the 9g/L of 5 times, obtain albumin stoste with the water for injection equal-volume dialysis of 3 times again, albumin stoste is finally concentrated into more than protein content 210g/L.With appropriate water for injection washing ultra-filtration membrane, collect film washing liquid, be incorporated in above-mentioned albumin concentrated solution and be human serum albumin stoste, stoste detection is carried out in sampling.
(11) work in-process preparation: carry out dilution preparation according to the calibrating of human serum albumin stoste: prepare according to human serum albumin stock protein matter content, Sodium octoate is taken by every gram of protein 160 mmole, sodium-chlor is taken by every gram of protein 130 mmole, dissolve with a small amount of water for injection after taking, add in human serum albumin stoste, again human serum albumin stoste is diluted to protein concn 200g/L, adjusted to ph obtains human serum albumin solution 6.95.
(12) pasteurization: human serum albumin stoste is carried out 60 DEG C of 10 hours inactivation of viruses, degerming packing after deactivation.
Claims (1)
1. from fraction IV precipitate, reclaim the method for human serum albumin, it is characterized in that described method comprises following concrete technological step:
(1) fraction IV precipitate dissolves
1 ~ 2 DEG C that fraction IV precipitate is put into 2 times of weight, in the sodium-chlor of 8 ~ 10g/L, stirring and dissolving, keep solution temperature-1 ~-2 DEG C, stirring and dissolving 5 ~ 6 hours, is fraction IV precipitate lysate;
(2) purifying
1. fraction IV precipitate lysate is squeezed in diluent and dilute, the amount of diluent is 3 ~ 4 times of fraction IV precipitate lysate, after dilution, goods are stirred and cool to-2 ~-3 DEG C, 1mol/L sodium bicarbonate is slowly added by the speed of 0.8 ~ 1.2L/ minute, adjustment goods pH value to 6.1 ~ 6.4, stir 6 ~ 8 hours under-2 ~-3 DEG C of conditions, obtain component IV dissolved solution for later use;
Wherein the preparation of diluent is: sodium chloride-containing 0.15mol/L, alcohol concn 27 ~ 30%(V/V), stir cooling
To-2 ~-3 DEG C;
2. in component IV lysate, add the ethanol of less than-20 DEG C 95%, alcohol concn in component IV lysate is made to reach 40 ~ 42%(V/V), ethanol adds speed and remains to 70 ~ 80L/h, solution temperature-2 ~-3 DEG C is kept in adition process, ethanol finishes, with sodium bicarbonate adjustment solution pH value to 6.6 ~ 6.7 of 1mol/L, cooling makes the temperature of component IV lysate to-5.5 ~-6 DEG C, and it is for subsequent use that constant temperature stirring obtains component IV diluent for 6 ~ 8 hours;
3. in component IV diluent, diatomite is added, add-on is every 100Kg fraction IV precipitate 3 ~ 5Kg diatomite, stir and use pressure filter press filtration after 30-40 minute, press filtration terminates rear cold reduction air and blows pressure filter intracavity liquid, to going out bubble, collect filtrate, obtain component IV pressing filtering liquid for subsequent use, precipitation is abandoned and is lost; Wherein, described diatomite refers to through 180 DEG C dry roasting 2.5 hours, is cooled to the diatomite of less than 0 DEG C;
(3) preparation of component V precipitation
1. component IV pressing filtering liquid volume is measured, open and stir, cooling, goods cool to-6 ~-7 DEG C, slowly add 2mol/L HAC, adding speed is 50-60ml/min, adjustment pH value of solution to 5.2 ~ 5.5, after having adjusted pH, goods be cooled to-8 ~-9 DEG C and stir 2 hours, quietly putting 6 ~ 8 hours;
2. open stirring, in component IV pressing filtering liquid, add diatomite, add-on adds 1 ~ 2Kg diatomite with every 100Kg fraction IV precipitate, stirs and starts press filtration after 30-40 minute, and filtrate abandoning is lost or carried out ethanol recycling after collecting; Collecting component V precipitates for subsequent use; Wherein, described diatomite refers to through 180 DEG C dry roasting 2.5 hours, is cooled to the diatomite of less than 0 DEG C;
(4) component V polishing purification
1. the component V precipitation 8%(V/V of 5 times of volumes) ethanolic soln stirring and dissolving, after precipitation is dissolved completely, measuring pH value of solution should in pH to 4.50 ~ 4.70, and goods outlet temperature controls, for-3.0 DEG C ± 0.5 DEG C, to continue stirring press filtration in 2 ~ 3 hours;
2. press filtration: stir in pressure-filtering process and do not stop, after press filtration is complete, with liquid in low temperature cold Air blowing filter, stops after going out bubble, and rear FV refines balance liquid and rinses filter plate, and washing fluid is incorporated in filtered liquid that to obtain refined solution for subsequent use;
Wherein FV refines the preparation of balance liquid: containing ethanol 12%(V/V), solution pH value 4.50 ~ 4. 70, is cooled to-3.0 DEG C ± 0.5 DEG C;
(5) ultrafiltration
1. stirring is opened, with 1M sodium bicarbonate adjustment refined solution pH value to 7.0 ~ 7.2;
2. by refined solution first ultrafiltration and concentration to protein concentration 100 ~ 150g/L, dialyse with the sodium chloride solution equal-volume of the 9g/L of 5 times, albumin stoste is obtained again with the water for injection equal-volume dialysis of 3 times, by final for albumin stoste ultrafiltration and concentration to more than protein content 210g/L, with appropriate water for injection washing ultra-filtration membrane, collect film washing liquid, be incorporated in above-mentioned protein liquid and be human serum albumin stoste, stoste detection is carried out in sampling;
(6) work in-process preparation
Dilution preparation is carried out: prepare according to stock protein matter content according to stoste calibrating, Sodium octoate is taken by every gram of protein 160 mmole, sodium-chlor is taken by every gram of protein 120 ~ 130 mmole, dissolve with a small amount of water for injection after taking, add human serum albumin stoste, again human serum albumin stoste is diluted to protein concn 195 ~ 200g/L, adjustment pH value is 6.8 ~ 7.0;
(7) pasteurization: goods are carried out 60 DEG C of 10 hours inactivation of viruses, degerming packing after deactivation.
2. according to the method reclaiming human serum albumin from fraction IV precipitate described in claim 1, when it is characterized in that press filtration in preparation process, the preparation of pressure filter is: every 100Kg fraction IV precipitate dress 6-25 opens filter plate, after filter plate is installed, it is negative for rinsing pressure filter to washing fluid bacterial endotoxin with water for injection, in FIV balance liquid or FV balance liquid, diatomite is added with the amount of every platform pressure filter 3-5Kg, pumped into pressure filter after stirring and carried out circulating cooling, when pressure filter temperature out is to less than-3 DEG C, stop circulation, freezing air blowout pressure filter chamber inner equilibrium liquid, the compound method of described FIV balance liquid is: containing 40%(V/V) ethanol, the sodium-chlor of 1.5 ~ 2.5g/L, the anhydrous sodium acetate of 4.5 ~ 5.1g/L, with acetic acid adjustment pH value to 6.0 ~ 6.1, solution temperature-6 ~-7 DEG C, amount of preparation is every platform pressure filter 600L, the compound method of described FV balance liquid is: water for injection 230L, adds 95% ethanol 170L, cools to-8 ~-9 DEG C, and amount of preparation is every platform pressure filter 400L.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310307858.8A CN103333240B (en) | 2013-07-22 | 2013-07-22 | Method for reclaiming human albumin from component IV precipitate |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310307858.8A CN103333240B (en) | 2013-07-22 | 2013-07-22 | Method for reclaiming human albumin from component IV precipitate |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103333240A CN103333240A (en) | 2013-10-02 |
| CN103333240B true CN103333240B (en) | 2015-01-28 |
Family
ID=49241459
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310307858.8A Active CN103333240B (en) | 2013-07-22 | 2013-07-22 | Method for reclaiming human albumin from component IV precipitate |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103333240B (en) |
Families Citing this family (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103833844A (en) * | 2014-03-05 | 2014-06-04 | 贵州中泰生物科技有限公司 | Production method of human serum albumin |
| CN104086646B (en) * | 2014-07-03 | 2018-10-09 | 成都蓉生药业有限责任公司 | The preparation method that FV is precipitated in blood product |
| CN104086645A (en) * | 2014-07-03 | 2014-10-08 | 成都蓉生药业有限责任公司 | Method for preparing human serum albumin and globulin |
| CN104558156A (en) * | 2015-01-23 | 2015-04-29 | 郑州莱士血液制品有限公司 | Method for extracting human serum albumin from plasma and increasing yield |
| CN105367651B (en) * | 2015-12-22 | 2018-11-27 | 郑州莱士血液制品有限公司 | A kind of method of cold ethanol method removal human immunoglobulin(HIg) pyrogen |
| CN106008698B (en) * | 2015-12-25 | 2019-05-07 | 北海开元生物科技有限公司 | A method of human serum albumin pyrogen is removed using cold ethanol method |
| CN111589199B (en) * | 2020-06-02 | 2023-10-27 | 国药集团贵州血液制品有限公司 | Separation device and method for ultralow aluminum ion residual quantity of human serum albumin |
| CN112062833A (en) * | 2020-10-09 | 2020-12-11 | 国药集团武汉血液制品有限公司 | Method for extracting human serum albumin from plasma component IV precipitate |
| CN116589559B (en) * | 2023-06-05 | 2024-02-27 | 广东丹霞生物制药有限公司 | Process for preparing human serum albumin |
| CN117185300B (en) * | 2023-09-26 | 2024-04-05 | 中南大学 | Method for preparing fluorosilicate crystals based on black talc |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101792490A (en) * | 2010-01-19 | 2010-08-04 | 广东卫伦生物制药有限公司 | Method for recycling albumin in Cohn's fraction IV precipitate |
-
2013
- 2013-07-22 CN CN201310307858.8A patent/CN103333240B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101792490A (en) * | 2010-01-19 | 2010-08-04 | 广东卫伦生物制药有限公司 | Method for recycling albumin in Cohn's fraction IV precipitate |
Non-Patent Citations (1)
| Title |
|---|
| 国内外FIV沉淀回收白蛋白工艺调查与分析;刘力;《中国输血杂志》;20101231;第1091页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103333240A (en) | 2013-10-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103333240B (en) | Method for reclaiming human albumin from component IV precipitate | |
| CN102488713B (en) | Method for preparing sheep placenta extract and sheep placenta hydrolyzed collagen concentrated solution | |
| CN101974070B (en) | Preparation process of human prothrombin compound | |
| CN104342040A (en) | Preparation method of high-melt-temperature fish scale gelatin | |
| CN104672328A (en) | Production method of human antithrombin III | |
| CN101972479A (en) | Preparation process of intravenous injection human immunoglobulin | |
| CN108065413A (en) | The method that oyster oligopeptide is prepared using oyster fresh meat | |
| CN109875009A (en) | A kind of preparation method of privet-like premna leaf jelly | |
| CN102807511A (en) | Method for extracting taurine from mussel | |
| CN104558156A (en) | Method for extracting human serum albumin from plasma and increasing yield | |
| CN104004084B (en) | A kind of production method of human albumin | |
| CN112480243B (en) | Process and equipment for large-scale hirudin separation and purification production | |
| CN106676089A (en) | Method for preparing human prothrombin complex from plasma | |
| CN104341539B (en) | A kind of enzyme process combines the method that membrane technology one step prepares refined heparin sodium | |
| CN106928332A (en) | A kind of preparation method of different deliquescent mussel proteins and polypeptide | |
| CN106119328A (en) | A kind of high-quality shrimp oligopeptide powder, preparation method thereof of applicable industrialized production | |
| CN112521487A (en) | Improved production process of human serum albumin | |
| CN103103170B (en) | Production process for cow or sheep hyaluronidase | |
| CN108101980B (en) | Preparation method of high-purity phycocyanin | |
| CN103462166B (en) | Pueraria flavones functional drink and preparation method thereof | |
| CN112442136A (en) | Method for extracting functional components from tremella | |
| CN106318995A (en) | Method for preparing fructan and gluconic acid by using inulin | |
| CN205258363U (en) | Preparation facilities of high -quality gardenia uranidin | |
| CN108042569A (en) | A kind of method of impurity in separation gel class Chinese medicine | |
| CN107383185A (en) | A kind of extracting method of high-purity bovine serum albumin |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20170605 Address after: 450001 No. 22 Yulan street, Zhengzhou hi tech Development Zone, Henan, Zhengzhou Patentee after: ZHENGZHOU RAAS BLOOD PRODUCTS CO., LTD. Address before: 536000 No. 126 Beihai Avenue, Haicheng District, the Guangxi Zhuang Autonomous Region, Beihai 804 Patentee before: Beihai Kaiyuan Biological Technology Co., Ltd. |
|
| TR01 | Transfer of patent right |