DD213947A1 - METHOD OF CULTURING BACTERIA OF SPECIES PASTEURELLA MULTOCIDA - Google Patents
METHOD OF CULTURING BACTERIA OF SPECIES PASTEURELLA MULTOCIDA Download PDFInfo
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- DD213947A1 DD213947A1 DD24857583A DD24857583A DD213947A1 DD 213947 A1 DD213947 A1 DD 213947A1 DD 24857583 A DD24857583 A DD 24857583A DD 24857583 A DD24857583 A DD 24857583A DD 213947 A1 DD213947 A1 DD 213947A1
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- pasteurella multocida
- production
- multocida
- cultivation
- species
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- 241000606856 Pasteurella multocida Species 0.000 title claims abstract description 12
- 238000000034 method Methods 0.000 title claims abstract description 10
- 238000012258 culturing Methods 0.000 title claims abstract description 6
- 241000894006 Bacteria Species 0.000 title claims abstract description 5
- 238000004519 manufacturing process Methods 0.000 claims abstract description 9
- 239000002028 Biomass Substances 0.000 claims abstract description 8
- 229940051027 pasteurella multocida Drugs 0.000 claims abstract description 3
- 241000894007 species Species 0.000 claims 1
- 229960005486 vaccine Drugs 0.000 abstract description 6
- 239000000463 material Substances 0.000 abstract description 4
- 238000011031 large-scale manufacturing process Methods 0.000 abstract 1
- 239000000203 mixture Substances 0.000 description 4
- 235000015097 nutrients Nutrition 0.000 description 4
- PXQPEWDEAKTCGB-UHFFFAOYSA-N orotic acid Chemical compound OC(=O)C1=CC(=O)NC(=O)N1 PXQPEWDEAKTCGB-UHFFFAOYSA-N 0.000 description 4
- CKLJMWTZIZZHCS-UHFFFAOYSA-N D-OH-Asp Natural products OC(=O)C(N)CC(O)=O CKLJMWTZIZZHCS-UHFFFAOYSA-N 0.000 description 3
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 3
- CKLJMWTZIZZHCS-UWTATZPHSA-N L-Aspartic acid Natural products OC(=O)[C@H](N)CC(O)=O CKLJMWTZIZZHCS-UWTATZPHSA-N 0.000 description 3
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 3
- -1 L-serine o Chemical class 0.000 description 3
- 229960005261 aspartic acid Drugs 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 244000052616 bacterial pathogen Species 0.000 description 3
- 229940041514 candida albicans extract Drugs 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 210000002966 serum Anatomy 0.000 description 3
- 239000012138 yeast extract Substances 0.000 description 3
- 241000337692 Bunia Species 0.000 description 2
- 235000013049 Bunias Nutrition 0.000 description 2
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 2
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 description 2
- ODKSFYDXXFIFQN-BYPYZUCNSA-N L-arginine Chemical compound OC(=O)[C@@H](N)CCCN=C(N)N ODKSFYDXXFIFQN-BYPYZUCNSA-N 0.000 description 2
- 229930064664 L-arginine Natural products 0.000 description 2
- 235000014852 L-arginine Nutrition 0.000 description 2
- LEVWYRKDKASIDU-IMJSIDKUSA-N L-cystine Chemical compound [O-]C(=O)[C@@H]([NH3+])CSSC[C@H]([NH3+])C([O-])=O LEVWYRKDKASIDU-IMJSIDKUSA-N 0.000 description 2
- 235000019393 L-cystine Nutrition 0.000 description 2
- 239000004158 L-cystine Substances 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 description 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 2
- DFPAKSUCGFBDDF-UHFFFAOYSA-N Nicotinamide Chemical compound NC(=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-UHFFFAOYSA-N 0.000 description 2
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 2
- JZRWCGZRTZMZEH-UHFFFAOYSA-N Thiamine Natural products CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 description 2
- 229960003067 cystine Drugs 0.000 description 2
- 229960002989 glutamic acid Drugs 0.000 description 2
- 229960005010 orotic acid Drugs 0.000 description 2
- 229940014662 pantothenate Drugs 0.000 description 2
- 239000011713 pantothenic acid Substances 0.000 description 2
- 229960001153 serine Drugs 0.000 description 2
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 description 2
- 229960003495 thiamine Drugs 0.000 description 2
- 235000019157 thiamine Nutrition 0.000 description 2
- 239000011721 thiamine Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 description 1
- 229930182844 L-isoleucine Natural products 0.000 description 1
- 239000004395 L-leucine Substances 0.000 description 1
- 235000019454 L-leucine Nutrition 0.000 description 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 244000309466 calf Species 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 229960002413 ferric citrate Drugs 0.000 description 1
- 239000001530 fumaric acid Substances 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- NPFOYSMITVOQOS-UHFFFAOYSA-K iron(III) citrate Chemical compound [Fe+3].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NPFOYSMITVOQOS-UHFFFAOYSA-K 0.000 description 1
- 229960000310 isoleucine Drugs 0.000 description 1
- 229960003136 leucine Drugs 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 229960003966 nicotinamide Drugs 0.000 description 1
- 235000005152 nicotinamide Nutrition 0.000 description 1
- 239000011570 nicotinamide Substances 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 229960005190 phenylalanine Drugs 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 229960004441 tyrosine Drugs 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Das Verfahren zur Kultivierung von Bakterien der Species Pasteurella multocida" bezieht sich auf das Anwendungsgebiet der Herstellung von Impfstoffen. Der Erfindung liegt die Aufgabe zugrunde, ein Kultivierungsverfahren zu beschreiben, das ohne erhoehten Material- und Energieeinsatz eine verbesserte Impfstoffproduktion erlaubt. Das Wesen der Erfindung besteht darin, dass die Kultivierung zur Biomasseproduktion von P. multocida bei erniedrigten Kultivierungstemperaturen, zwischen 25 und 30 Grad C, am besten bei 27 Grad C, erfogt. Die Erfindung kann in der Grossproduktion von Pasteurella-multocida-Vakzine Anwendung finden.The method for cultivating bacteria of the species Pasteurella multocida refers to the field of application of the production of vaccines The object of the invention is to describe a culturing method which allows improved vaccine production without increased use of material and energy in that the cultivation for biomass production of P. multocida is performed at reduced culturing temperatures, between 25 and 30 degrees C, most preferably at 27 degrees C. The invention can find application in the large scale production of Pasteurella multocida vaccine.
Description
Titel der ErfindungTitle of the invention
Verfahren zur Kultivierung von Bakterien der Species Pasteurella multocidaMethod of culturing bacteria of the species Pasteurella multocida
Anwendungsgebiet der ErfindungField of application of the invention
Die Erfindung betrifft ein Verfahren zur Kultivierung von Bakterien der Species Pasteurella multocida. Die Vermehrung dieser Mikroorganismen, die als Erreger von Krankheiten eine Holle spielen, ist für die Herstellung von Impfstoffen bedeutsam.The invention relates to a method for the cultivation of bacteria of the species Pasteurella multocida. The multiplication of these microorganisms, which play a major role as pathogens for diseases, is important for the production of vaccines.
Charakteristik der bekannten technischen LösungenCharacteristic of the known technical solutions
Die Massenkultivierung für diese Zwecke geschieht in bekannter Weise bei 370C in flüssigen Nährmedien unterschiedlicher Zusammensetzung - in chemisch definierten, teilweise definierten bzw. komplexen Medien mit mehr oder weniger hohen Anteilen Undefiniert zusammengesetzter natürlicher Bestandteile, z* B. Pepton, Trypton, Hefe-Extrakt, Serum, Pleischwasser u.a.The mass cultivation for these purposes is done in a known manner at 37 0 C in liquid nutrient media of different composition - in chemically defined, partially defined or complex media with more or less high levels of undefined composite natural ingredients, z * B. peptone, tryptone, yeast Extract, serum, pleural water, etc.
Die Biomasseproduktion ist je nach der Art der eingesetzten Nährlösungen ,und Kultivierungsbedingungen unterschiedlich hoch. Zur Massenkultivierung für die ImpfStoffproduktion eingesetzte Medien sind aufgrund ökonomischer Betrachtungen oft nicht geeignet, so daß die in Laboransätzen erreichten Ausbeuten dort kaum erreicht werden. Die Erfindung bezweckt deshalb, bei gleichem Materialeinsatz durch Veränderung der Kultivierungsbedingungen die Biomasseproduktion zu erhöhen.The biomass production varies depending on the type of nutrient solutions used, and cultivation conditions. Due to economic considerations, media used for mass cultivation for vaccine production are often unsuitable, so that the yields achieved in laboratory mixtures are barely reached there. The invention therefore aims to increase the biomass production with the same material use by changing the cultivation conditions.
Darlegung des Wesens der ErfindungExplanation of the essence of the invention
Der Erfindung liegt die Aufgabe zugrunde, ein Kultivierungsverfahren zu beschreiben, das ohne erhöhten Material- und Energieeinsatz eine verbesserte Impfstoffproduktion erlaubt.The invention has for its object to describe a cultivation process that allows improved material and energy use improved vaccine production.
Erfindungsgemäß wird diese Aufgabe dadurch gelöst, daß Stämme von Pasteurella multocida durch Erniedrigung der Bebrütungstemperatur kultiviert werden, wobei die Ausbeuten an Biomasse ohne zusätzlichen Materialeinsatz bei der Herstellung der Hährlösungen stark erhöht v/erden. Statt der bisher üblichen 37°C werden Temperaturen zwischen 25 und 3o C eingesetzt, wobei sich als praktikable Größe 270C erweist. Damit wird bei gleichem Nährstoffangebot unter Temperaturerniedrigung um 1o C (also gleichzeitiger Einsparung von Energie) eine Ausbeutesteigerung von 1o - oo % erreicht.According to the invention, this object is achieved in that strains of Pasteurella multocida are cultured by lowering the incubation temperature, wherein the yields of biomass strongly increased without additional use of material in the preparation of Hährlösungen v / earth. Instead of the usual 37 ° C temperatures between 25 and 3o C are used, with 27 0 C proves to be a practical size. Thus, with the same nutrient supply, with temperature reduction by 1o C (ie simultaneous saving of energy), a yield increase of 1o - oo% is achieved.
Ausführungsbeispieleembodiments
1.Am Beispiel von P. multocida 383 aus der Stammsammlung des Instituts für bakterielle Tierseuchenforschung Jena soll die Abhängigkeit der Biomasse-Ausbeuten von der Kultivierung stemperatur gezeigt werden (als Maß für die produziete Biomasse dient die bei 650 lim gemessen=Extinktion am Photometer, E^1- )j1.The example of P. multocida 383 from the strain collection of the Institute for Bacterial Epidemiology Jena shows the dependence of the biomass yields on the cultivation temperature (the measured biomass used is the measured at 650 lim = extinction on the photometer, E ^ 1 -) j
Die Zahl der Lebendkeime ist bei erniedrigter Temperatur ebenfalls erhöht (P. multocida 383; 370C: 5 χ 1o9/ml, 270C:The number of live germs is also increased at low temperature (P. multocida 383; 37 0 C: 5 χ 1o 9 / ml, 27 0 C:
7 x 1o /ml). Bei Kultivierung im Bakterienferment er werden gleichfalls analoge Ausbeuteerhöhungen beobachtet: 370C;7 x 10 / ml). When cultivated in the bacterial ferment he also analogous increases in yield are observed: 37 0 C;
Err- = o,6; 6 χ Ιο"1 Lebendkeime/ml; 270C: E/-r-_ = o,75; coo -ι 030Err- = o, 6; 6 χ Ιο " 1 live germs / ml; 27 0 C: E / -r-_ = o, 75; coo -ι 030
2 χ 1o Lebendkeime/ml.2 χ 1o live germs / ml.
Die vorgenannten Ergebnisse wurden mit einem chemisch definiertem Uährmedium (Rezept n9o6") folgender Zusammensetzung bei der Kultivierung erreicht (Angaben in g/l Aqua dest., pH 7,S - S):The abovementioned results were achieved with a chemically defined culture medium (recipe n 9o6 ") of the following composition during cultivation (data in g / l of distilled water, pH 7, S - S):
12 H2O 32,312H 2 O 32.3
1'36 1 '36
HaCl ^ 1,19HaCl ^ 1,19
MgSo1 . 7 H2O ο,25MgSo 1 . 7 H 2 O o, 25
Glucose 6Glucose 6
L-Arginin » HCl ο,2L-arginine »HCl o, 2
L-Asparaginsäure 1,6L-aspartic acid 1.6
L-Cystin o,12L-cystine o, 12
L-Glutaminsäure ο, 15L-glutamic acid ο, 15
DL-Serin ο,2DL serine ο, 2
(oder L-Serin o,1)(or L-serine o, 1)
L-Isoleucin 0,065L-isoleucine 0.065
L-Leucin 0,065L-leucine 0.065
L-Phenylalanin 0,085L-phenylalanine 0.085
L-Tyrosin o,o9L-Tyrosine o, o9
Ca-Pantothenat o,oo2Ca-pantothenate o, oo2
Mcotinsäureamid 0,00 5Mcotinic acid amide 0.005
Thiamin . HCl o5oo1Thiamine. HCl o 5 oo1
Orotsäure ' o,o15Orotic acid, o, o15
Eisen-III-citrat o5oo1Ferric citrate o 5 oo1
Die starke Ausbeuteerhöhung bei 270C ist jedoch nicht auf ein bestimmtes !Tährmedium bei der Durchführung des Kultivierungsverfahrens beschränkt, wie die Versuche mit verschiedenen Medien beweisen. Auf 6 unterschiedlichen ITährmedien, die sowohl chemisch definierten, teildefinierten als auch komplexen Charakter hatten, wurden stets bei 270C höhere Biomasseausbeuten erreicht (P. multocida 383):However, the strong increase in yield at 27 0 C is not limited to a particular! Tährmedium in carrying out the culturing process, as shown by the experiments with different media. 6 different ITährmedien chemically defined both in part defined as a complex character had been, always at 27 0 C higher biomass yields reached (P. multocida 383):
(270C)(27 0 C)
0,60 o,77 o,9o 0,64 0,60 0,600.60 o, 77 o, 9o 0.64 0.60 0.60
wsiehe Seite 4 w see page 4
die Nährmedienzusammensetzung ist v/ie folgt (g/l): 8o3: Na2HPO. . 12 H2O 32,3the nutrient composition is as follows (g / l): 8o3: Na 2 HPO. , 12H 2 O 32.3
24 1,36 24 1.36
MgSO4 . 7 H2O o,25MgSO 4. 7H 2 O o, 25
Glucose 6Glucose 6
L-Arginin . HCl . o,1L-arginine. HCl. o, 1
L-Asparaginsäure 1,6L-aspartic acid 1.6
L-Cystin ο,οβL-cystine o, o
!-Glutaminsäure o,o75! -Glutamic acid o, o75
DL-Serin o,1DL serine o, 1
(oder L-Serin o,o5)(or L-serine o, o5)
Ca-Pantothenat o,oo2Ca-pantothenate o, oo2
Nicotinsäureamid ο,005Nicotinamide ο, 005
Thiamin . HCl o,oo1Thiamine. HCl o, oo1
Orotsäure o,o15Orotic acid o, o15
Hefe-Extrakt 5Yeast extract 5
8o3S: 8o3 mit 1o % Pferdeserum oder Kälberserum·8o3S: 8o3 with 1o% horse serum or calf serum ·
808: Na2HPO4 . 12 H2O 32,3808: Na 2 HPO 4 . 12H 2 O 32.3
KH2PO4 1,36KH 2 PO 4 1.36
MgSO4 .7H2O o,25MgSO 4 .7H 2 O o, 25
Glucose 6Glucose 6
L-Asparaginsäure 1,6L-aspartic acid 1.6
Fumarsäure 5,8Fumaric acid 5,8
Hefe-Extrakt 5Yeast extract 5
9o2: Rezept 9o6 (siehe im Text weiter vorn) ohne Pe-Ill-citrat9o2: Recipe 9o6 (see above) without Pe-Ill citrate
9o9: Rezept 9o6 (siehe im Text weiter vorn) ohne Pe-Ill-citrat, ohne Tyrosin9o9: Recipe 9o6 (see above) without Pe-Ill citrate, without tyrosine
2. Mit 11 repräsentativen P.-multocida-Stämmen unterschiedlicher Typen aus der Stammsammlung des Instituts für bakterielle Tierseuchenforschung Jena (383, 383 ts, 6417, 35), dem Institut "Pasteur" 3ukarest (B 85o), dem Central Public Health Laboratory, London (ICTC 1o322, 1o324, 1o326, 1o2o4, 1o3o2) und dein National Institute of Animal Health, Tokyo (VA 3j Bunia II) wurde das Verfahren in der beschriebenen Weise bei 270C durchgeführt und ebenfalls in allen Fällen eine Ausbeutesteigerung erreicht:2. With 11 representative P. multocida strains of different types from the strain collection of the Institute for Bacterial Epidemiology Jena (383, 383 ts, 6417, 35), the Institute "Pasteur" 3ukarest (B 85o), the Central Public Health Laboratory, London (ICTC 1o322, 1o324, 1o326, 1o2o4, 1o3o2) and your National Institute of Animal Health, Tokyo (VA 3j Bunia II), the method was carried out in the manner described at 27 0 C and also achieved in all cases an increase in yield:
Stamm S^r- (3-7 C; Ξ,-.- (^ 7 C)Strain S ^ r- (3-7 C; Ξ, -.- (^ 7 C)
383 o,46 o,59383 o, 46 o, 59
383 ts · o,39 o,47383 ts · o, 39 o, 47
VA 3 o,45 o,5oVA 3 o, 45 o, 5o
6417 o,34 0,466417 o, 34 0.46
B 35o 0,46 o,5oB 35o 0.46 o, 5o
35 o,54 o,5835 o, 54 o, 58
Bunia II 0,48 o,52Bunia II 0.48, 52
I0324 o,5o 0,60I0324 o, 5o 0.60
1o2o4 o,57 o,621o2o4 o, 57 o, 62
I0326 o,5o o,54I0326 o, 5o o, 54
1o3o2 o,47 0,611o3o2 o, 47 0.61
Claims (2)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DD24857583A DD213947A1 (en) | 1983-03-08 | 1983-03-08 | METHOD OF CULTURING BACTERIA OF SPECIES PASTEURELLA MULTOCIDA |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DD24857583A DD213947A1 (en) | 1983-03-08 | 1983-03-08 | METHOD OF CULTURING BACTERIA OF SPECIES PASTEURELLA MULTOCIDA |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| DD213947A1 true DD213947A1 (en) | 1984-09-26 |
Family
ID=5545449
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| DD24857583A DD213947A1 (en) | 1983-03-08 | 1983-03-08 | METHOD OF CULTURING BACTERIA OF SPECIES PASTEURELLA MULTOCIDA |
Country Status (1)
| Country | Link |
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| DD (1) | DD213947A1 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5587166A (en) * | 1987-03-24 | 1996-12-24 | British Technology Group Limited | Vaccine against Pasteurella |
-
1983
- 1983-03-08 DD DD24857583A patent/DD213947A1/en not_active IP Right Cessation
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5587166A (en) * | 1987-03-24 | 1996-12-24 | British Technology Group Limited | Vaccine against Pasteurella |
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| A1 | Published as prov. economic patent | ||
| ENJ | Ceased due to non-payment of renewal fee |