NO137427B - APPARATUS FOR MANUFACTURE OF STEPED BUILDING ELEMENTS - Google Patents
APPARATUS FOR MANUFACTURE OF STEPED BUILDING ELEMENTS Download PDFInfo
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- NO137427B NO137427B NO1870/73A NO187073A NO137427B NO 137427 B NO137427 B NO 137427B NO 1870/73 A NO1870/73 A NO 1870/73A NO 187073 A NO187073 A NO 187073A NO 137427 B NO137427 B NO 137427B
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- Prior art keywords
- streptokinase
- column
- phosphate buffer
- concentration
- solution
- Prior art date
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- 238000004519 manufacturing process Methods 0.000 title 1
- 108010023197 Streptokinase Proteins 0.000 claims description 45
- 229960005202 streptokinase Drugs 0.000 claims description 45
- 239000008363 phosphate buffer Substances 0.000 claims description 18
- 238000000034 method Methods 0.000 claims description 12
- 239000001913 cellulose Substances 0.000 claims description 5
- 229920002678 cellulose Polymers 0.000 claims description 5
- 239000012535 impurity Substances 0.000 claims description 3
- JVIPLYCGEZUBIO-UHFFFAOYSA-N 2-(4-fluorophenyl)-1,3-dioxoisoindole-5-carboxylic acid Chemical compound O=C1C2=CC(C(=O)O)=CC=C2C(=O)N1C1=CC=C(F)C=C1 JVIPLYCGEZUBIO-UHFFFAOYSA-N 0.000 claims description 2
- 229920001425 Diethylaminoethyl cellulose Polymers 0.000 claims description 2
- 239000000243 solution Substances 0.000 description 16
- GUBGYTABKSRVRQ-WFVLMXAXSA-N DEAE-cellulose Chemical compound OC1C(O)C(O)C(CO)O[C@H]1O[C@@H]1C(CO)OC(O)C(O)C1O GUBGYTABKSRVRQ-WFVLMXAXSA-N 0.000 description 10
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 10
- 239000011347 resin Substances 0.000 description 6
- 229920005989 resin Polymers 0.000 description 6
- 239000000725 suspension Substances 0.000 description 6
- LWIHDJKSTIGBAC-UHFFFAOYSA-K tripotassium phosphate Chemical compound [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 239000000872 buffer Substances 0.000 description 5
- 239000011780 sodium chloride Substances 0.000 description 5
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- 229910019142 PO4 Inorganic materials 0.000 description 4
- 239000007853 buffer solution Substances 0.000 description 4
- 238000010828 elution Methods 0.000 description 4
- 235000021317 phosphate Nutrition 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 102000013566 Plasminogen Human genes 0.000 description 3
- 108010051456 Plasminogen Proteins 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000000356 contaminant Substances 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 3
- 239000010452 phosphate Substances 0.000 description 3
- 229910000160 potassium phosphate Inorganic materials 0.000 description 3
- 239000008057 potassium phosphate buffer Substances 0.000 description 3
- 235000011009 potassium phosphates Nutrition 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 239000001488 sodium phosphate Substances 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- 238000000855 fermentation Methods 0.000 description 2
- 230000004151 fermentation Effects 0.000 description 2
- 238000012856 packing Methods 0.000 description 2
- 229940012957 plasmin Drugs 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 229910000162 sodium phosphate Inorganic materials 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 238000011282 treatment Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 206010002660 Anoxia Diseases 0.000 description 1
- 241000976983 Anoxia Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 206010011091 Coronary artery thrombosis Diseases 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 206010011703 Cyanosis Diseases 0.000 description 1
- 102000009123 Fibrin Human genes 0.000 description 1
- 108010073385 Fibrin Proteins 0.000 description 1
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 1
- 101000605403 Homo sapiens Plasminogen Proteins 0.000 description 1
- 206010021143 Hypoxia Diseases 0.000 description 1
- 208000010378 Pulmonary Embolism Diseases 0.000 description 1
- 241000193996 Streptococcus pyogenes Species 0.000 description 1
- 208000007536 Thrombosis Diseases 0.000 description 1
- 206010044565 Tremor Diseases 0.000 description 1
- 229910052783 alkali metal Inorganic materials 0.000 description 1
- 150000001340 alkali metals Chemical class 0.000 description 1
- 229910052784 alkaline earth metal Inorganic materials 0.000 description 1
- 150000001342 alkaline earth metals Chemical class 0.000 description 1
- 230000007953 anoxia Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 230000036760 body temperature Effects 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000012461 cellulose resin Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 208000002528 coronary thrombosis Diseases 0.000 description 1
- 239000012531 culture fluid Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 229910000397 disodium phosphate Inorganic materials 0.000 description 1
- 235000019800 disodium phosphate Nutrition 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000012149 elution buffer Substances 0.000 description 1
- 239000003344 environmental pollutant Substances 0.000 description 1
- 210000000416 exudates and transudate Anatomy 0.000 description 1
- 229950003499 fibrin Drugs 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 230000002949 hemolytic effect Effects 0.000 description 1
- 230000002706 hydrostatic effect Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 1
- 235000019799 monosodium phosphate Nutrition 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 1
- 231100000719 pollutant Toxicity 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 1
- 239000012064 sodium phosphate buffer Substances 0.000 description 1
- 235000011008 sodium phosphates Nutrition 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 201000005060 thrombophlebitis Diseases 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000001226 triphosphate Substances 0.000 description 1
- 235000011178 triphosphate Nutrition 0.000 description 1
- 125000002264 triphosphate group Chemical class [H]OP(=O)(O[H])OP(=O)(O[H])OP(=O)(O[H])O* 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B28—WORKING CEMENT, CLAY, OR STONE
- B28B—SHAPING CLAY OR OTHER CERAMIC COMPOSITIONS; SHAPING SLAG; SHAPING MIXTURES CONTAINING CEMENTITIOUS MATERIAL, e.g. PLASTER
- B28B15/00—General arrangement or layout of plant ; Industrial outlines or plant installations
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B28—WORKING CEMENT, CLAY, OR STONE
- B28B—SHAPING CLAY OR OTHER CERAMIC COMPOSITIONS; SHAPING SLAG; SHAPING MIXTURES CONTAINING CEMENTITIOUS MATERIAL, e.g. PLASTER
- B28B23/00—Arrangements specially adapted for the production of shaped articles with elements wholly or partly embedded in the moulding material; Production of reinforced objects
- B28B23/02—Arrangements specially adapted for the production of shaped articles with elements wholly or partly embedded in the moulding material; Production of reinforced objects wherein the elements are reinforcing members
- B28B23/04—Arrangements specially adapted for the production of shaped articles with elements wholly or partly embedded in the moulding material; Production of reinforced objects wherein the elements are reinforcing members the elements being stressed
- B28B23/06—Arrangements specially adapted for the production of shaped articles with elements wholly or partly embedded in the moulding material; Production of reinforced objects wherein the elements are reinforcing members the elements being stressed for the production of elongated articles
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B28—WORKING CEMENT, CLAY, OR STONE
- B28B—SHAPING CLAY OR OTHER CERAMIC COMPOSITIONS; SHAPING SLAG; SHAPING MIXTURES CONTAINING CEMENTITIOUS MATERIAL, e.g. PLASTER
- B28B7/00—Moulds; Cores; Mandrels
- B28B7/0029—Moulds or moulding surfaces not covered by B28B7/0058 - B28B7/36 and B28B7/40 - B28B7/465, e.g. moulds assembled from several parts
- B28B7/0035—Moulds characterised by the way in which the sidewalls of the mould and the moulded article move with respect to each other during demoulding
- B28B7/0041—Moulds characterised by the way in which the sidewalls of the mould and the moulded article move with respect to each other during demoulding the sidewalls of the mould being moved only parallelly away from the sidewalls of the moulded article
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B28—WORKING CEMENT, CLAY, OR STONE
- B28B—SHAPING CLAY OR OTHER CERAMIC COMPOSITIONS; SHAPING SLAG; SHAPING MIXTURES CONTAINING CEMENTITIOUS MATERIAL, e.g. PLASTER
- B28B7/00—Moulds; Cores; Mandrels
- B28B7/10—Moulds with means incorporated therein, or carried thereby, for ejecting or detaching the moulded article
Landscapes
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Ceramic Engineering (AREA)
- Mechanical Engineering (AREA)
- Manufacturing & Machinery (AREA)
- Devices For Post-Treatments, Processing, Supply, Discharge, And Other Processes (AREA)
- Manufacturing Of Tubular Articles Or Embedded Moulded Articles (AREA)
- Moulds, Cores, Or Mandrels (AREA)
- Coating Apparatus (AREA)
- Casting Devices For Molds (AREA)
- Panels For Use In Building Construction (AREA)
- Road Signs Or Road Markings (AREA)
- Apparatuses And Processes For Manufacturing Resistors (AREA)
Description
Fremgangsmåte til rensing av uren streptokinase. Process for purification of impure streptokinase.
Foreliggende oppfinnelse angår en fremgangsmåte til rensing av uren streptokinase. The present invention relates to a method for purifying impure streptokinase.
Streptokinase er et fermentas]' onspro-dukt fra hemolytiske streptokokker. Det ble for noen tid siden fastslått at streptokinasen spiller en rolle ved oppløsning av blodpropper og særlig av eksudater som er rike på fibrin-bestanddeler. I avhengighet av blodproppenes natur og størrelse frembringer streptokinase, eller streptokinase i nærvær av plasminogen fra mennesker, oppløsning av disse. Imidlertid har hittil anvendelsen av. streptokinase (og av plasmin fremstillet fra plasminogen og streptokinase) vært begrenset på grunn av den kjennsgjerning at streptokinasen har vært forurenset med streptokokk-protein-bipro-dukter som frembringer giftvirkninger hos mennesker. Slike virkninger viser seg ved økning av legemstemperaturen, senkning av blodtrykket, rystelser, kuldegysninger, anoksi, cyanose og lignende. Streptokinase is a fermentation product from hemolytic streptococci. It was established some time ago that streptokinase plays a role in the dissolution of blood clots and especially of exudates that are rich in fibrin components. Depending on the nature and size of the clots, streptokinase, or streptokinase in the presence of human plasminogen, dissolves them. However, to date the application of streptokinase (and of plasmin produced from plasminogen and streptokinase) has been limited by the fact that the streptokinase has been contaminated with streptococcal protein by-products that produce toxic effects in humans. Such effects are manifested by an increase in body temperature, lowering of blood pressure, tremors, chills, anoxia, cyanosis and the like.
Ved hjelp av foreliggende oppfinnelse By means of the present invention
muliggjøres det å fjerne de forurensende proteiner fra streptokinase-preparater så at man får i høy grad ren streptokinase. Herved blir altså en i høy grad ren streptokinase tilgjengelig, hva der gjør det mulig makes it possible to remove the contaminating proteins from streptokinase preparations so that you get highly pure streptokinase. This means that a highly pure streptokinase is available, what makes this possible
å behandle pasienter etter ønske med streptokinase eller med plasminogen som er aktivert med streptokinase. Slike be-handlinger er funnet å være av enestående to treat patients as desired with streptokinase or with streptokinase-activated plasminogen. Such treatments have been found to be exceptional
viktighet ved behandling av forskjellige sykdommer som tromboflebit, flebothrom- importance in the treatment of various diseases such as thrombophlebitis, phlebothrom-
bose, lunge-emboli, coronær trombose og lignende. bose, pulmonary embolism, coronary thrombosis and the like.
Den urene streptokinase som renses ved fremgangsmåten ifølge oppfinnelsen, kan erholdes ved hjelp av kjente fremgangsmå-ter. I alminnelighet fremstilles den ved å anvende Streptococcus hemolyticus som podningsmiddel i et næringsmedium og la The impure streptokinase which is purified by the method according to the invention can be obtained using known methods. In general, it is prepared by using Streptococcus hemolyticus as an inoculum in a nutrient medium and la
veksten av bakteriene foregå i 8—10 timer. Etter forløpet av dette tidsrom avbrytes the growth of the bacteria takes place for 8-10 hours. After the expiry of this period, it is cancelled
fermentasjonen ved å lede kulturvæsken gjennom kjøleslanger hvorpå bakteriecel-lene fjernes ved sentrifugering eller filtre-ring. Streptokinasen kan isoleres fra kulturvæsken ved hjelp av hvilken som helst kjent metode. I alminnelighet har materialet som er isolert på denne måte, en renhet fra 15 til 60 %, bestemt ved fysikalske homogenitetsmetoder. Foreliggende oppfinnelse angår altså en fremgangsmåte til rensning av slike urene produkter. the fermentation by passing the culture liquid through cooling hoses, whereupon the bacterial cells are removed by centrifugation or filtration. The streptokinase can be isolated from the culture fluid by any known method. In general, the material isolated in this way has a purity of 15 to 60%, as determined by physical homogeneity methods. The present invention therefore relates to a method for cleaning such impure products.
Fremgangsmåten ifølge oppfinnelsen er karakterisert ved at en oppløsning av The method according to the invention is characterized in that a solution of
uren streptokinase i en fosfatpuffer med molkonsen tras jon fra 0,005 til 0,015, fortrinnsvis 0,01, bringes i kontakt med DEAE-cellulose, hvorefter cellulosen vaskes med en fosfatpuffer hvis molkonsentrasjon ligger mellom 0,02 og 0,05, fortrinsvis 0,06, hvorved forurensninger fjernes, hvorpå streptokinasen elueres fra cellulosen med en fosfatpuffer hvis molkonsentrasjon ligger mellom 0,06 og 0,15, fortrinsvis 0,08. impure streptokinase in a phosphate buffer with a molar concentration of from 0.005 to 0.015, preferably 0.01, is brought into contact with DEAE cellulose, after which the cellulose is washed with a phosphate buffer whose molar concentration is between 0.02 and 0.05, preferably 0.06, whereby impurities are removed, after which the streptokinase is eluted from the cellulose with a phosphate buffer whose molar concentration is between 0.06 and 0.15, preferably 0.08.
Kromatograferingen kan foregå i kolonne eller chargevis. DEAE-cellulose (dietylaminoetylcellulose) vil si cellulose av den art som er beskrevet av Peterson og Sober (JACS 78:751, 1956). Chromatography can take place in a column or batchwise. DEAE cellulose (diethylaminoethyl cellulose) means cellulose of the kind described by Peterson and Sober (JACS 78:751, 1956).
Mange av forurensningene adsorberes ved den lave ionestyrke og bindes fast til harpiksen så at det erholdte streptokinase-eluat er fritt for disse stoffer. Andre forurensninger adsorberes ikke i nogen stor utstrekning og er tilstede i gjennomløpet. Many of the contaminants are adsorbed by the low ionic strength and are firmly bound to the resin so that the streptokinase eluate obtained is free of these substances. Other pollutants are not adsorbed to any great extent and are present in the flow-through.
Ved på riktig måte å velge fosfatpuf-ferens konsentrasjoner og den passende fraksjon, er det mulig med meget godt ut-bytte og på enkel måte å oppnå en meget ren streptokinase med høy terapeutisk ef-fektivitet og uten forurensninger som frembringer bivirkninger. By correctly choosing the concentrations of the phosphate buffer and the appropriate fraction, it is possible with very good yield and in a simple way to obtain a very pure streptokinase with high therapeutic effectiveness and without contaminants that produce side effects.
Der kan i fremgangsmåten ifølge oppfinnelsen anvendes en hvilkensomhelst vanlig fosfatpuffer, men det foretrekkes å bruke natriumfosfat eller kaliumfosfat. Imidlertid kan der også med godt resultat anvendes mono-di- eller trifosfater av jordalkalimetallene, foruten sådanne fos-fater av alkalimetallene. In the method according to the invention, any common phosphate buffer can be used, but it is preferred to use sodium phosphate or potassium phosphate. However, mono-di- or triphosphates of the alkaline earth metals can also be used with good results, in addition to such phosphates of the alkali metals.
Ved elueringen av streptokinasen med fosfatpuffere som har en molar konsentrasjon innen det foran angitte område, er fraksjoner med en molar konsentrasjon fra 0,06 til 0,08 funnet å inneholde de største mengder streptokinase. In the elution of the streptokinase with phosphate buffers having a molar concentration within the range indicated above, fractions with a molar concentration from 0.06 to 0.08 have been found to contain the largest amounts of streptokinase.
En meget egnet utførelsesform for fremgangsmåten ifølge oppfinnelsen under anvendelse av økende konsentrasjon av fos-fatpufferen ved elueringen, er å la denne strømme gjennom en kolonne med opprin-nelig molar konsentrasjon på 0,01 og grad-vis tilføre kolonnen en fosfatpuffer med molar konsentrasjon på 0,15. På denne måte økes kontinuerlig den molare konsentrasjon av pufferen som anvendes til elueringen. Den fraksjon som går gjennom kolonnen opp til en molar konsentrasjon på 0,06, kastes vekk, mens den fraksjon som har en molar konsentrasjon over 0,06 og opp til 0,15 oppsamles. I stedet for kontinuerlig å øke den molare konsentrasjon på denne måte, er det mulig å vaske harpiksen suksessivt med adskilte pufferoppløs-ninger med økende konsentrasjon. Således kan f.eks. en første vaskning utføres med en 0,02 M oppløsning, den annen med en 0,04 M oppløsning og den tredje med en 0,06 M oppløsning, idet alle de herved erholdte vaskevæsker kastes vekk. Vaske-væskene med de derpå følgende høyere molare konsentrasjoner, nemlig 0,07 M, 0,09 og 0,15 oppsamles da de inneholder de største mengder streptokinase. A very suitable embodiment of the method according to the invention, using increasing concentration of the phosphate buffer during the elution, is to let this flow through a column with an initial molar concentration of 0.01 and gradually add to the column a phosphate buffer with a molar concentration of 0.15. In this way, the molar concentration of the buffer used for the elution is continuously increased. The fraction that passes through the column up to a molar concentration of 0.06 is thrown away, while the fraction that has a molar concentration above 0.06 and up to 0.15 is collected. Instead of continuously increasing the molar concentration in this way, it is possible to wash the resin successively with separate buffer solutions of increasing concentration. Thus, e.g. a first washing is carried out with a 0.02 M solution, the second with a 0.04 M solution and the third with a 0.06 M solution, with all the wash liquids obtained thereby being thrown away. The washings with the subsequent higher molar concentrations, namely 0.07 M, 0.09 and 0.15 are collected as they contain the largest amounts of streptokinase.
En del streptokinase vil finnes i den første eluerings-pufferoppløsning med konsentrasjon 0,06 M, følgelig kan en annen vaskning med samme molare konsentrasjon utføres og vaskevæsken oppsamles da den inneholder en betydelig mengde streptokinase. Alternativt er det mulig å fjerne forurensningene med en enkel pufferopp-løsning hvis konsentrasjon er 0,06 M og å oppsamle streptokinasen med en enkel puf-feroppløsning med konsentrasjon f.eks. 0,08 M. Herved vil imidlertid utbyttet av streptokinase bli relativt lavt. Some streptokinase will be present in the first elution buffer solution with a concentration of 0.06 M, therefore another wash with the same molar concentration can be performed and the wash liquid collected as it contains a significant amount of streptokinase. Alternatively, it is possible to remove the contaminants with a simple buffer solution whose concentration is 0.06 M and to collect the streptokinase with a simple buffer solution with a concentration of e.g. 0.08 M. In this way, however, the yield of streptokinase will be relatively low.
I det følgende beskrives som eksempler noen utførelsesformer for oppfinnelsen. In the following, some embodiments of the invention are described as examples.
Eksempel I Example I
Dette eksempel viser rensing av urent streptokinase ved kromatografi i DEAE-celluloseharpiks fremstillet i overensstem-melse med det ovenfor anførte litteratur-sted. This example shows the purification of impure streptokinase by chromatography in DEAE cellulose resin prepared in accordance with the literature reference cited above.
Det foretrekkes å kondisjonere harpiksen før anvendelsen på følgende måte: 400 g DEAE-cellulose suspenderes i 8 liter vann og suspensjonens pH-verdi innstilles på 2 med IN saltsyre. Der-tilsettes derpå natriumklorid så at konsentrasjonen bringes på 5 %. Blandingen filtreres gjennom osteklede og vaskes med 8 liter 5 %'s natriumkloridoppløsning. Materialet suspenderes i vann og suspensjonens pH-verdi innstilles på 10,5 med IN natriumhydroxyd-oppløsning. Natriumkloridkonsentrasj onen bringes på 5 %, hvorpå blandingen filtreres og det tilbakeværende på filteret vaskes med 8 liter 5 %'s natriumkloridoppløs-ning. DEAE-cellulosen suspenderes påny og suspensjonens pH-verdi innstilles på 3 med IN saltsyre. Konsentrasjonen bringes påny til 5 % og harpiksen vaskes med 8 liter 5 %' s natriumkloridoppløsning. Derpå vaskes DEAE-cellulosen med 20 liter pyrogenfritt vann og filtreres. It is preferred to condition the resin before use in the following way: 400 g of DEAE cellulose is suspended in 8 liters of water and the pH value of the suspension is adjusted to 2 with IN hydrochloric acid. Sodium chloride is then added so that the concentration is brought to 5%. The mixture is filtered through cheesecloth and washed with 8 liters of 5% sodium chloride solution. The material is suspended in water and the pH value of the suspension is adjusted to 10.5 with IN sodium hydroxide solution. The sodium chloride concentration is brought to 5%, after which the mixture is filtered and what remains on the filter is washed with 8 liters of 5% sodium chloride solution. The DEAE cellulose is resuspended and the pH value of the suspension is adjusted to 3 with 1N hydrochloric acid. The concentration is again brought to 5% and the resin is washed with 8 liters of 5% sodium chloride solution. The DEAE cellulose is then washed with 20 liters of pyrogen-free water and filtered.
DEAE-cellulosen suspenderes i 0,01 M fosfatpuffer, suspensjonen omrøres og man lar de faste stoffer bunnfelles flere ganger for å fjerne findelte bestanddeler. Det er av vesentlig betydning at findelte bestanddeler fjernes for å kunne få kolonner med en passende strømningshastighet. Den kolonne som ble anvendt i dette eksempel, har en strømningshastighet på ca. 500 ml pr. time. Det er meget fordelaktig å om-røre kolonnens innhold umiddelbart før anvendelsen for å forhindre adskillelse av de faste stoffer fra glass-overflaten. The DEAE cellulose is suspended in 0.01 M phosphate buffer, the suspension is stirred and the solids are allowed to settle several times to remove finely divided components. It is of essential importance that finely divided constituents are removed in order to obtain columns with a suitable flow rate. The column used in this example has a flow rate of approx. 500 ml per hour. It is very advantageous to stir the contents of the column immediately before use to prevent separation of the solids from the glass surface.
I en kolonne med 30 cm lengde og inn-vendig diameter 52 mm anbringes DEAE-cellulose suspendert i 0,01 M fosfatpuffer. Der tilsettes DEAE-cellulose i slik mengde at der etter bunnfelning dannes en kolonne med høyde 6—7 cm. DEAE cellulose suspended in 0.01 M phosphate buffer is placed in a column with a length of 30 cm and an internal diameter of 52 mm. DEAE cellulose is added in such a quantity that, after sedimentation, a column with a height of 6-7 cm is formed.
Oppløsningen av den urene streptokinase dialyceres mot en fosfatpuffer med følgende sammensetning (konsentrasjon 0,005—0,015 M, fortrinnsvis 0,01 M): Mononatriumfosfat (NaH2P04) H20 — 27,6 g/liter Oppi. A Dinatriumfosfat (Na2HPOi) — 28,4 g pr. liter Oppi. B 16 ml av A fortynnes til 200 ml med destil-84 ml av B lert vann så at man får en fosfatpuffer med konsentrasjon 0,1 M og pH-verdi 7,5. Denne fortynnes til en fosfatkonsen-trasjon på 0,01 M. The solution of the impure streptokinase is dialyzed against a phosphate buffer with the following composition (concentration 0.005-0.015 M, preferably 0.01 M): Monosodium phosphate (NaH2P04) H20 — 27.6 g/liter Oppi. A Disodium phosphate (Na2HPOi) — 28.4 g per liters Uppi. B 16 ml of A is diluted to 200 ml with 84 ml of B distilled water so that a phosphate buffer with a concentration of 0.1 M and a pH value of 7.5 is obtained. This is diluted to a phosphate concentration of 0.01 M.
Etter at likevekt har innstillet seg, er streptokinase-materialet ferdig til rensing. Innstillingen av likevekt kan bedømmes ved å måle oppløsningens elektriske mot-stand. After equilibrium has been established, the streptokinase material is ready for purification. The setting of equilibrium can be judged by measuring the electrical resistance of the solution.
Den streptokinase som anvendes som utgangsmateriale kan fåes i handelen. I en konsentrasjon på 16° enheter/ml, eller i det hele 100 X 16e enheter oppløst i 100 ml vann, dialyseres i 16—18 timer ved 3° C mot 2 liter 0,01 M fosfatpuffer, pH-verdi 7,5. Streptokinaseoppløsningen innføres i kolonnen og elueres med fosfatpuffer. Ved en konsentrasjon av avløpet på 0,06 og 0,07 M elueres hovedparten av streptokinasen som et produkt med høy renhet, og viser seg i avløpet. Fraksjonene undersøkes på innhold av streptokinase og protein, og de en-kelte fraksjoners renhet bestemmes ved å beregne den spesifikke aktivitet (enheter streptokinase pr. gamma nitrogen). De fraksjoner som er rike på streptokinase kan lyofiliseres eller kan anvendes til over-føring av plasminogen til plasmin. The streptokinase used as starting material is commercially available. In a concentration of 16° units/ml, or a total of 100 X 16e units dissolved in 100 ml of water, dialyze for 16-18 hours at 3° C against 2 liters of 0.01 M phosphate buffer, pH value 7.5. The streptokinase solution is introduced into the column and eluted with phosphate buffer. At an effluent concentration of 0.06 and 0.07 M, most of the streptokinase is eluted as a product of high purity, and appears in the effluent. The fractions are examined for content of streptokinase and protein, and the purity of the individual fractions is determined by calculating the specific activity (units of streptokinase per gamma nitrogen). The fractions that are rich in streptokinase can be lyophilized or can be used to transfer plasminogen to plasmin.
Eksempel II Example II
Fremstilling av kolonnen: Der fremstilles en suspensjon av 1,5 g DEAE-cellulose i 30 ml 0,005 M kaliumfosfatpuffer med pH 7,0. Denne suspensjon helles i en krom-atografisk kolonne (med høyde 30 cm og indre diameter 1 cm) forsynt med en tet-ningsskive og en kapillarhevert. Man lar harpiksen bunnfelles ved sin tyngde, hvorpå man pakker den til en høyde på 15 cm ved påsetning av trykkluft (filtrert luft med et trykk ganske lite høyere enn atmosfæretrykk). Et tynt lag bomull anbringes over harpiksen i kolonnen for å beskytte denne mot mekaniske forstyrrel-ser. Denne pakning av kolonnen nedsetter strømningshastigheten gjennom denne til 10—20 ml pr. time. Anbringelse av prøven på kolonnen: Uren streptokinase med ca. 250 mg protein, oppløses i ca. 15 ml vann og den erholdte oppløsning dialyseres ved lavere tempera-tur (2—5° C) mot en 0,005 M fosfatpuffer. Den tilføres derpå kolonnen. Man lar den urene streptokinase-oppløsning perkolere gjennom kolonnen (under det samme lave trykk som ble anvendt ved pakningen av kolonnen) før tilsetning av puffer med økende ionestyrke. Preparation of the column: A suspension of 1.5 g of DEAE cellulose in 30 ml of 0.005 M potassium phosphate buffer with pH 7.0 is prepared. This suspension is poured into a chrome-atographic column (with a height of 30 cm and an internal diameter of 1 cm) provided with a sealing disk and a capillary sieve. The resin is allowed to settle to the bottom by its own weight, after which it is packed to a height of 15 cm by applying compressed air (filtered air with a pressure slightly higher than atmospheric pressure). A thin layer of cotton is placed over the resin in the column to protect it from mechanical disturbances. This packing of the column reduces the flow rate through it to 10-20 ml per hour. Placing the sample on the column: Impure streptokinase with approx. 250 mg protein, dissolves in approx. 15 ml of water and the resulting solution are dialysed at a lower temperature (2-5° C) against a 0.005 M phosphate buffer. It is then fed to the column. The impure streptokinase solution is allowed to percolate through the column (under the same low pressure used when packing the column) before adding buffers of increasing ionic strength.
Eluering fra kolonnen: Kaliumfosfatpuffer med pH-verdi 7,0 og med kontinuerlig økende konsentrasjon tilføres kolonnen. Konsentrasjonsgradienten ble erholdt ved å la 0,3 M kaliumfosfat (pH-verdi 7,0) strømme inn i en 250 ml beholder med 0,005 M puffer. Puffer-oppløsningene blan-des ved hjelp av et magnetisk røreverk. Oppløsning fra blandingsbeholderen strøm-mer gjennom kolonnen, idet strømnings-hastigheten gjennom denne bestemmer strømningshastigheten av 0,3 M kaliumfos-fatoppløsningen til 250 ml beholderen. Den hydrostatiske søyle som driver pufferen gjennom kolonnen, kan varieres ved å heve eller senke pufferbeholderen. Elution from the column: Potassium phosphate buffer with a pH value of 7.0 and with continuously increasing concentration is added to the column. The concentration gradient was obtained by allowing 0.3 M potassium phosphate (pH value 7.0) to flow into a 250 ml container with 0.005 M buffer. The buffer solutions are mixed using a magnetic stirrer. Solution from the mixing container flows through the column, the flow rate through which determines the flow rate of the 0.3 M potassium phosphate solution into the 250 ml container. The hydrostatic column that drives the puffer through the column can be varied by raising or lowering the puffer container.
Hvert påfølgende 10 ml eluat ble opp-samlet separat. Undersøkelser på streptokinase-innhold i de således oppsamlede fraksjoner viser at under de angitte betin-gelser er hovedmengden av ren streptokinase i det 100. til 150. ml eluat, i hvilke fosfatkonsentrasjonen varierer fra 0,07 til 0,12 M. Ved lavere fosfatkonsentrasjoner ble forurensninger i den urene streptokinase eluert. Each subsequent 10 ml eluate was collected separately. Investigations into streptokinase content in the thus collected fractions show that under the specified conditions the main amount of pure streptokinase is in the 100 to 150 ml eluate, in which the phosphate concentration varies from 0.07 to 0.12 M. At lower phosphate concentrations impurities in the impure streptokinase were eluted.
Eksempel III Example III
Man går frem således som angitt i eksempel II, men under anvendelse av na-triumfosfatpuffer i stedet for kaliumfosfat-puffer. Hovedmengden av ren streptokinase er da tilstede i det 100. til 150. ml eluat, således som i eksempel II. The procedure is as indicated in example II, but using sodium phosphate buffer instead of potassium phosphate buffer. The main amount of pure streptokinase is then present in the 100 to 150 ml eluate, as in example II.
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US00251358A US3832118A (en) | 1972-05-08 | 1972-05-08 | Apparatus for production of cast concrete members |
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NO137427B true NO137427B (en) | 1977-11-21 |
NO137427C NO137427C (en) | 1978-03-01 |
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NO1870/73A NO137427C (en) | 1972-05-08 | 1973-05-07 | APPARATUS FOR MANUFACTURE OF CAST BUILDING ELEMENTS |
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US (1) | US3832118A (en) |
JP (1) | JPS5516045B2 (en) |
AR (1) | AR198311A1 (en) |
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AU (1) | AU464377B2 (en) |
BE (1) | BE799248A (en) |
BG (1) | BG27060A3 (en) |
BR (1) | BR7303329D0 (en) |
CA (1) | CA1000936A (en) |
CH (1) | CH579973A5 (en) |
CS (1) | CS168652B2 (en) |
DD (1) | DD105152A5 (en) |
DE (1) | DE2322542A1 (en) |
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FI (1) | FI50943C (en) |
FR (1) | FR2184316A5 (en) |
GB (1) | GB1434797A (en) |
IE (1) | IE37587B1 (en) |
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IN (1) | IN139764B (en) |
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US4004874A (en) * | 1974-06-19 | 1977-01-25 | Span-Deck, Inc. | Apparatus for production of cast concrete members |
FI61281C (en) * | 1978-10-02 | 1982-07-12 | Span Deck Inc | BETONGGJUTESUNDERLAGSANLAEGGNING |
US4289293A (en) * | 1978-10-02 | 1981-09-15 | Span-Deck, Inc. | Combination bed concrete casting apparatus |
US4252293A (en) * | 1979-10-31 | 1981-02-24 | Norwalk Concrete Industries, Inc. | Hydraulic mold release mechanism for precast concrete products |
EP0030247A1 (en) * | 1979-12-10 | 1981-06-17 | T.R.-Plast Kunststoffverarbeitung Inh. Theo Rosbach | Moulding apparatus for toy construction-elements |
DE3336655C2 (en) * | 1983-10-08 | 1995-07-27 | Karl Munte Betonwerke Gmbh | Casting mold and method for producing a quiver foundation |
ATE133601T1 (en) * | 1990-06-21 | 1996-02-15 | Nord Immobiliare Centro | METHOD AND APPARATUS FOR PRODUCING REINFORCED CONCRETE ITEMS |
BE1004584A3 (en) * | 1990-08-29 | 1992-12-15 | Coussee Marc | Method and device for producing t-shaped beams from pre-stressed concrete |
NL9002531A (en) * | 1990-11-20 | 1992-06-16 | Schokbeton Bv | Apparatus for the production of prestressed concrete sleepers for railway tracks, as well as sleepers manufactured using this apparatus. |
US5393033A (en) * | 1993-07-21 | 1995-02-28 | Wilson Concrete Company | Adjustable side form concrete mold |
ES2155293B1 (en) * | 1997-02-04 | 2001-12-16 | Extremadura 2000 De Estructura | MANUFACTURING PROCEDURE OF PRETENSED PLATES TYPE PI BY EXTRUSION. |
US20050183357A1 (en) * | 2004-02-10 | 2005-08-25 | The Cretex Companies, Inc. | Pre-formed concrete section |
FR2892740A1 (en) * | 2006-02-10 | 2007-05-04 | Conseil Service Investissement | Method of producing a concrete beam with a base plate with a top surface carrying a rib involves provision of an enclosure in the form of the beam, concrete pouring into the enclosure, and beam extraction |
US20160312466A1 (en) * | 2014-10-10 | 2016-10-27 | Norfa Enterprises Pty Ltd. | Components for masonry construction |
US11701795B1 (en) * | 2019-02-27 | 2023-07-18 | Hamilton Form Company, Ltd. | Concrete mold form |
CN112895116B (en) * | 2021-01-19 | 2022-03-11 | 陕西中天建筑工业有限公司 | Auxiliary equipment for prefabricated plate material distribution and processing technology thereof |
CN113246285B (en) * | 2021-05-25 | 2022-07-08 | 中交(福州)建设有限公司 | Automatic assembly line prefabrication construction method for concrete revetment hooking block |
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US1068613A (en) * | 1912-05-07 | 1913-07-29 | William J Volz | Mold for forming irregular bricks and tiles. |
US1642947A (en) * | 1927-01-31 | 1927-09-20 | Goliath Rubber Company | Mold |
US1770368A (en) * | 1929-04-18 | 1930-07-08 | Permold Co | Mold |
US2916795A (en) * | 1957-05-03 | 1959-12-15 | Henderson Albert | Apparatus for molding reinforced concrete building slabs, columns and girders |
US3132403A (en) * | 1961-06-22 | 1964-05-12 | Fmc Corp | Concrete beam molding apparatus |
CA763876A (en) * | 1962-07-06 | 1967-07-25 | Span-Deck | Apparatus and method for forming concrete planks or slabs |
US3168771A (en) * | 1963-01-28 | 1965-02-09 | Alfred T Nelson | Adjustable wing t form |
US3523343A (en) * | 1967-12-05 | 1970-08-11 | Span Deck Inc | System for the production of cast concrete members |
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1972
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1973
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- 1973-04-27 IL IL42129A patent/IL42129A/en unknown
- 1973-04-30 AU AU54976/73A patent/AU464377B2/en not_active Expired
- 1973-04-30 CH CH611873A patent/CH579973A5/xx not_active IP Right Cessation
- 1973-05-01 CA CA170,182A patent/CA1000936A/en not_active Expired
- 1973-05-01 IE IE683/73A patent/IE37587B1/en unknown
- 1973-05-01 GB GB2059173A patent/GB1434797A/en not_active Expired
- 1973-05-02 FI FI731396A patent/FI50943C/en active
- 1973-05-04 DE DE2322542A patent/DE2322542A1/en not_active Withdrawn
- 1973-05-07 NO NO1870/73A patent/NO137427C/en unknown
- 1973-05-07 AR AR247875A patent/AR198311A1/en active
- 1973-05-07 DD DD170656A patent/DD105152A5/xx unknown
- 1973-05-07 FR FR7316318A patent/FR2184316A5/fr not_active Expired
- 1973-05-07 OA OA54901A patent/OA04382A/en unknown
- 1973-05-07 SU SU731914244A patent/SU615844A3/en active
- 1973-05-07 SE SE7306381A patent/SE382008B/en unknown
- 1973-05-08 PL PL1973162408A patent/PL96612B1/en unknown
- 1973-05-08 RO RO7374718A patent/RO65677A/en unknown
- 1973-05-08 ES ES414526A patent/ES414526A1/en not_active Expired
- 1973-05-08 BE BE130875A patent/BE799248A/en unknown
- 1973-05-08 AT AT403973A patent/AT330638B/en not_active IP Right Cessation
- 1973-05-08 BG BG023540A patent/BG27060A3/en unknown
- 1973-05-08 JP JP5151073A patent/JPS5516045B2/ja not_active Expired
- 1973-05-08 NL NL7306389A patent/NL7306389A/xx not_active Application Discontinuation
- 1973-05-08 ZA ZA733117A patent/ZA733117B/en unknown
- 1973-05-08 CS CS3280A patent/CS168652B2/cs unknown
- 1973-05-08 IT IT23855/73A patent/IT994859B/en active
- 1973-05-08 BR BR3329/73A patent/BR7303329D0/en unknown
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1978
- 1978-09-15 KE KE2884A patent/KE2884A/en unknown
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