ES2228281B1 - PRODUCTION METHOD OF GIBERELINAS GA3, THROUGH FERMENTATIONS WITH WILD STRAP IMI58289 OF HONGO GIBBERELLA FUJIKUROI. - Google Patents
PRODUCTION METHOD OF GIBERELINAS GA3, THROUGH FERMENTATIONS WITH WILD STRAP IMI58289 OF HONGO GIBBERELLA FUJIKUROI.Info
- Publication number
- ES2228281B1 ES2228281B1 ES200400724A ES200400724A ES2228281B1 ES 2228281 B1 ES2228281 B1 ES 2228281B1 ES 200400724 A ES200400724 A ES 200400724A ES 200400724 A ES200400724 A ES 200400724A ES 2228281 B1 ES2228281 B1 ES 2228281B1
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- Prior art keywords
- gibberellins
- imi58289
- wild
- gibberella fujikuroi
- production
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Método de producción de giberelinas GA3, mediante fermentaciones con la estirpe silvestre IMI58289 del hongo Gibberella Fujikuroi. Es objeto de esta invención un método para la producción de giberelinas con la estirpe silvestre de Gibberella fujikuroi IMI58289, sin necesidad de manipulación genética previa del hongo. En primer lugar, la estirpe IMI58289 se inocula durante 6 días en medio rico en nitrógeno, condiciones en las que se inhibe la producción de giberelinas. A los seis días, los cultivos se filtran y el micelio se lava con agua destilada y se resuspende en una solución de glucosa con microelementos. La incubación posterior en estas condiciones da lugar a una mezcla de giberelinas con una proporción inusualmente elevada de GA3.Method of production of gibberellins GA3, by fermentation with the wild line IMI58289 of the fungus Gibberella Fujikuroi. The object of this invention is a method for the production of gibberellins with the wild strain of Gibberella fujikuroi IMI58289, without the need for prior genetic manipulation of the fungus. First, the IMI58289 line is inoculated for 6 days in a medium rich in nitrogen, conditions in which the production of gibberellins is inhibited. At six days, the cultures are filtered and the mycelium is washed with distilled water and resuspended in a glucose solution with microelements. Subsequent incubation under these conditions results in a mixture of gibberellins with an unusually high proportion of GA3.
Description
Método de producción de giberelinas GA_{3} mediante fermentaciones con la estirpe silvestre IMI58289 del hongo Gibberella fujikuroi.Method of production of gibberellins GA3 by fermentation with the wild strain IMI58289 of the fungus Gibberella fujikuroi .
Es objeto de esta invención un método de producción de giberelinas GA_{3}, usando la estirpe silvestre IMI58289 de Gibberella fujikuroi, sin necesidad de manipulación genética previa del hongo.The object of this invention is a method of producing gibberellins GA3, using the wild strain IMI58289 of Gibberella fujikuroi , without the need for prior genetic manipulation of the fungus.
Las giberelinas (GAs) son fitohormonas naturales que regulan el crecimiento y la floración de las plantas entre otras actividades fisiológicas. Estas fitohormonas se encuentran a nivel de trazas en las plantas pero su aplicación exógena produce resultados espectaculares por lo que tienen un gran interés en agricultura intensiva y la industria de la cerveza. Se han descrito más de 100 giberelinas entre las cuales GA_{3} (1), también conocida como ácido giberélico, se ha considerado tradicionalmente la giberelina más activa en plantas (MacMillan, J, Natural Product Report, 1997, 221). Sin embargo estudios recientes han mostrado que GA_{1} (2) tiene una importante actividad biológica, incluso superior a GA_{3} en plantas como el arroz (Kobayashi M, MacMillan J, Phinney B, Gaskin P, Spray CR, Hedden P, Phytochemistry, 2000, 55, 317), el tomate (Grünzwig JM, Rabinowitch HD, Katan J, Wodner M, Ben-Tal Y, Phytochemistry 1997, 46, 811), el cerezo (Huanpu M, Blake PS, Browning G; Taylor JM, Phytochemistry, 2001, 56, 67) o el guisante (Yaxley JR, Ross JJ, Sherriff LJ, Reid JB, Plant Physiol, 2001, 125, 627).Gibberellins (GAs) are natural phytohormones that regulate the growth and flowering of plants among other physiological activities. These phytohormones are at the level of traces in plants but their exogenous application produces spectacular results so they have a great interest in intensive agriculture and the beer industry. More than 100 gibberellins have been described, among which GA3 (1), also known as gibberellic acid, has traditionally been considered the most active gibberellin in plants (MacMillan, J, Natural Product Report , 1997 , 221). However, recent studies have shown that GA 1 (2) has an important biological activity, even higher than GA 3 in plants such as rice (Kobayashi M, MacMillan J, Phinney B, Gaskin P, Spray CR, Hedden P , Phytochemistry , 2000 , 55 , 317), tomato (Grünzwig JM, Rabinowitch HD, Katan J, Wodner M, Ben-Tal Y, Phytochemistry 1997 , 46 , 811), cherry tree (Huanpu M, Blake PS, Browning G; Taylor JM, Phytochemistry, 2001 , 56 , 67) or the pea (Yaxley JR, Ross JJ, Sherriff LJ, Reid JB, Plant Physiol, 2001 , 125 , 627).
El hongo filamentoso Gibberella fujikuroi es un organismo capaz de producir giberelinas en cantidades suficientes para su explotación industrial.The filamentous fungus Gibberella fujikuroi is an organism capable of producing gibberellins in sufficient quantities for its industrial exploitation.
La producción de giberelinas por distintas estirpes silvestres o mutantes de Gibberella fujikuroi en diversos medios de cultivo ha sido descrita en varias ocasiones. Concretamente, la producción de GA_{3} (1) ha sido cubierta bajo diferentes patentes (GB844341, GB850018, GB886551, US2918413) así como la producción de la mezcla activa de giberelinas GA_{4} + GA_{7} (US6287800, CN1222575). Sin embargo, la producción de GA_{3} (2) o mezclas de GA_{1} + GA_{3} a niveles que puedan ser útiles en la industria agroquimica no ha sido descrita previamente, y precisamente la invención que se presenta en esta solicitud de patente consiste en un método de producción de giberelinas GA_{3}.The production of gibberellins by different wild or mutant strains of Gibberella fujikuroi in various culture media has been described several times. Specifically, the production of GA_ {3} (1) has been covered under different patents (GB844341, GB850018, GB886551, US2918413) as well as the production of the active mixture of gibberellins GA_ {4} + GA_ {(US6287800, CN1222575 ). However, the production of GA 3 (2) or mixtures of GA 1 + GA 3 at levels that may be useful in the agrochemical industry has not been previously described, and precisely the invention presented in this Patent application consists of a method of producing gibberellins GA3.
En esta solicitud de patente se describe un método para la producción de giberelinas con la estirpe silvestre de Gibberella fujikuroi IMI58289 sin necesidad de manipulación genética previa del hongo.This patent application describes a method for the production of gibberellins with the wild strain of Gibberella fujikuroi IMI58289 without the need for prior genetic manipulation of the fungus.
En primer lugar, la estirpe IMI58289 se incuba durante 6 días en medio de rico en nitrógeno, condiciones en las que se inhibe la producción de giberelinas. A los seis días, los cultivos se filtran y el micelio se lava con agua destilada y se resuspende en una solución de glucosa con microelementos. La incubación posterior en estas condiciones da lugar a una mezcla de giberelinas aproximada de 0.1 g/l de medio cultivado, con una proporción inusualmente elevada de GA_{3} .First, the IMI58289 line is incubated for 6 days in the middle of rich in nitrogen, conditions in which gibberellin production is inhibited. At six days, the cultures are filtered and the mycelium is washed with distilled water and resuspended in a glucose solution with microelements. The subsequent incubation under these conditions results in a mixture of gibberellins approximately 0.1 g / l of cultivated medium, with a unusually high proportion of GA3.
Los extractos de cada fermentación fueron analizados mediante espectroscopia ^{1}H RMN y la técnica combinada Cromatografía de Gases-Espectrometría de Masas (CG-EM) [Columna: HP-1 (Crosslinked Methyl Siloxane) 25 m x 0.2 mm x 0.33 \mum; gas portador: helio con un flujo de 0.6 ml/min; temperatura del inyector: 260ºC; temperatura del detector: 280ºC; temperatura inicial: 120ºC, incrementada a 5ºC/min hasta 220ºC, incrementada posteriormente a 3ºC/min hasta 280ºC, donde se mantiene durante 10 minutos].The extracts of each fermentation were analyzed by 1 H NMR spectroscopy and the technique Combined Gas Chromatography-Spectrometry of Masses (CG-EM) [Column: HP-1 (Crosslinked Methyl Siloxane) 25 m x 0.2 mm x 0.33 µm; gas carrier: helium with a flow of 0.6 ml / min; temperature of injector: 260 ° C; detector temperature: 280 ° C; temperature Initial: 120ºC, increased to 5ºC / min to 220ºC, increased subsequently at 3 ° C / min up to 280 ° C, where it is maintained for 10 minutes].
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La fermentación se llevó a cabo en dos etapas. En una primera etapa se distribuyó el medio de cultivo rico en nitrógeno en matraces Erlenmeyer, que se inocularon con una suspensión de esporas de la estirpe silvestre IMI58289 de Gibberella fujikuroi. Los matraces, tapados con algodón graso para facilitar la aireación, se incubaron a 30ºC con una agitación orbital de 200 rpm aproximadamente. A los 6 días se detuvo la fermentación y se separó el micelio del medio de cultivo por filtración. A continuación se resuspendió en una solución de glucosa con microelementos y se incubó durante 6 días en las condiciones arriba mencionadas. Finalmente se filtró el caldo de cultivo, se acidificó a pH 1.5-3 con HCl, y se extrajo con acetato de etilo. La evaporación a vacío del disolvente y posterior desengrasado del extracto generó un residuo constituido mayoritariamente por giberelinas. La giberelina mayoritaria de esta mezcla fue GA_{3} (en una proporción superior al 50%).The fermentation was carried out in two stages. In a first stage, the nitrogen-rich culture medium was distributed in Erlenmeyer flasks, which were inoculated with a spore suspension of the wild strain IMI58289 of Gibberella fujikuroi . The flasks, covered with fatty cotton to facilitate aeration, were incubated at 30 ° C with an orbital shaking of approximately 200 rpm. After 6 days the fermentation was stopped and the mycelium was removed from the culture medium by filtration. It was then resuspended in a glucose solution with microelements and incubated for 6 days under the conditions mentioned above. Finally, the culture broth was filtered, acidified to pH 1.5-3 with HCl, and extracted with ethyl acetate. Evaporation of the solvent under vacuum and subsequent degreasing of the extract generated a residue consisting mostly of gibberellins. The majority gibberellin in this mixture was GA3 (in a proportion greater than 50%).
Claims (2)
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
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ES200203050A ES2221786B1 (en) | 2002-12-20 | 2002-12-20 | PRODUCTION OF GIBERELINAS, GA1 AND GA3 THROUGH FERMENTATIONS WITH WILD STRAPS OF HONGO GIBBERELLA FUJIKUROI. |
Publications (2)
Publication Number | Publication Date |
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ES2228281A1 ES2228281A1 (en) | 2005-04-01 |
ES2228281B1 true ES2228281B1 (en) | 2006-05-16 |
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ES200400724A Expired - Fee Related ES2228281B1 (en) | 2002-12-20 | 2002-12-20 | PRODUCTION METHOD OF GIBERELINAS GA3, THROUGH FERMENTATIONS WITH WILD STRAP IMI58289 OF HONGO GIBBERELLA FUJIKUROI. |
ES200203050A Expired - Fee Related ES2221786B1 (en) | 2002-12-20 | 2002-12-20 | PRODUCTION OF GIBERELINAS, GA1 AND GA3 THROUGH FERMENTATIONS WITH WILD STRAPS OF HONGO GIBBERELLA FUJIKUROI. |
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ES200203050A Expired - Fee Related ES2221786B1 (en) | 2002-12-20 | 2002-12-20 | PRODUCTION OF GIBERELINAS, GA1 AND GA3 THROUGH FERMENTATIONS WITH WILD STRAPS OF HONGO GIBBERELLA FUJIKUROI. |
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Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
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CN108004290B (en) * | 2018-01-18 | 2021-06-29 | 四川龙蟒福生科技有限责任公司 | Fermentation medium for producing gibberellic acid |
CN108285915B (en) * | 2018-01-18 | 2021-07-23 | 四川龙蟒福生科技有限责任公司 | Fermentation method of gibberellic acid |
CN114231421A (en) * | 2021-12-01 | 2022-03-25 | 四川龙蟒福生科技有限责任公司 | Gibberella fujikuroi and fermentation method for producing GA3 |
Family Cites Families (6)
Publication number | Priority date | Publication date | Assignee | Title |
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BE567954A (en) * | 1957-02-05 | |||
GB838033A (en) * | 1957-02-13 | 1960-06-22 | Ici Ltd | Process for the production of gibberellic acid |
GB819110A (en) * | 1957-07-02 | 1959-08-26 | Pfizer & Co C | Recovery process for gibberellins |
GB839652A (en) * | 1957-09-09 | 1960-06-29 | Ici Ltd | A process for the manufacture of gibberellic acid |
ES2046129B1 (en) * | 1992-06-15 | 1994-09-01 | Consejo Superior Investigacion | A PROCEDURE FOR THE PRODUCTION OF GIBERELIC ACID, IN SUBMERGED CROP, FROM EFFLUENTS FROM THE INDUSTRIAL PROCESSING OF MUSSEL. |
JP2004517625A (en) * | 2000-11-14 | 2004-06-17 | フィブロ−テック,インコーポレイテッド,ドゥーイング ビジネス アズ アグトロール インターナショナル | Novel nucleic acids and methods for modulating gibberellin production and morphologically altered cells |
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2002
- 2002-12-20 ES ES200400724A patent/ES2228281B1/en not_active Expired - Fee Related
- 2002-12-20 ES ES200203050A patent/ES2221786B1/en not_active Expired - Fee Related
Non-Patent Citations (2)
Title |
---|
CANDAU, R. et al. Regulation of gibberellin biosynthesis in Gibberella fujikuroi. Plant Physiol., 1992, Vol. 100, páginas 1184-1188. * |
FERNÁNDEZ-MARTÍN, R. et al. ent-Kaurene and squalene synthesis in Fusarium fujikuroi cell-free extracts. Phytochemistry, 2000, Vol. 54, páginas 723-728. * |
Also Published As
Publication number | Publication date |
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ES2221786A1 (en) | 2005-01-01 |
ES2221786B1 (en) | 2006-04-01 |
ES2228281A1 (en) | 2005-04-01 |
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