CS248612B1 - Industrial mutant of sponge claviceps fusiformis w1 cc-1-85 - Google Patents
Industrial mutant of sponge claviceps fusiformis w1 cc-1-85 Download PDFInfo
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- CS248612B1 CS248612B1 CS883284A CS883284A CS248612B1 CS 248612 B1 CS248612 B1 CS 248612B1 CS 883284 A CS883284 A CS 883284A CS 883284 A CS883284 A CS 883284A CS 248612 B1 CS248612 B1 CS 248612B1
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- claviceps fusiformis
- sponge
- fusiformis
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- 241000124232 Claviceps fusiformis Species 0.000 title claims abstract description 10
- DAVNRFCJMIONPO-UHFFFAOYSA-N (7-methyl-6,6a,8,10a-tetrahydro-4h-indolo[4,3-fg]quinoline-9-yl)methanol Chemical compound C1=CC(C2C=C(CO)CN(C2C2)C)=C3C2=CNC3=C1 DAVNRFCJMIONPO-UHFFFAOYSA-N 0.000 claims abstract description 9
- 229930013930 alkaloid Natural products 0.000 abstract description 9
- 238000000855 fermentation Methods 0.000 abstract description 8
- 230000004151 fermentation Effects 0.000 abstract description 8
- 238000004519 manufacturing process Methods 0.000 abstract description 7
- 150000003797 alkaloid derivatives Chemical class 0.000 abstract description 3
- 244000005700 microbiome Species 0.000 abstract description 3
- 230000002906 microbiologic effect Effects 0.000 abstract 1
- 229920001817 Agar Polymers 0.000 description 8
- 239000008272 agar Substances 0.000 description 8
- 235000010633 broth Nutrition 0.000 description 6
- 229920001503 Glucan Polymers 0.000 description 2
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 2
- 230000001143 conditioned effect Effects 0.000 description 2
- 239000000049 pigment Substances 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- XJOOMMHNYOJWCZ-UHFFFAOYSA-N Agroclavine Natural products C1=CC(C2C=C(C)CN(C2C2)C)=C3C2=CNC3=C1 XJOOMMHNYOJWCZ-UHFFFAOYSA-N 0.000 description 1
- SAHHMCVYMGARBT-UHFFFAOYSA-N Isochanoclavine I Natural products C1=CC(C(C(NC)C2)C=C(C)CO)=C3C2=CNC3=C1 SAHHMCVYMGARBT-UHFFFAOYSA-N 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 229960001230 asparagine Drugs 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000007707 calorimetry Methods 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 230000001427 coherent effect Effects 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Řažení se týká nového průmyslového mutanta houby Claviceps fusiformis W1 produ- .kujicí alkaloid elymoklavin, uložený ve sbírce mikroorganismů Mikrobiologického ústavu ČSAV v Praze 4 pod 6. CC 1/85. Submersní fermentaci tohoto kmene lze několikanásobně zvýžit produkcí klavinových alkaloidů, zejména farmakologicky významného alkaloidu elyaoklavinu.The ripping involves a new industrial mutant fungi Claviceps fusiformis W1 producing alkaloid elymoclavin, deposited in collection of microbiological microorganisms Institute of the Czechoslovak Academy of Sciences in Prague 4 under the 6th CC 1/85. Submerged fermentation of this strain is possible increase the production by keyboard alkaloids, particularly pharmacologically significant alkaloid elyaoclavine.
Description
Vynález se týká nového průmyslového mutanta kmene Claviceps fusiformis Wi produkující farmaceuticky významný alkaloid elyaoklavin (M.Abe et el., US Pat. 2835675) strukturního vzorceThe invention relates to a novel industrial mutant of the strain Claviceps fusiformis Wi producing a pharmaceutically important alkaloid elyaoclavin (M.Abe et al., US Pat. 2835675) of the structural formula
Vzorky mutanta jsou uloženy ve sbírce mikroorganismů Mikrobiologického ústavu ČSAV v Praze 4 pod označením Claviceps fusiformis pod č. CC 1/85.Mutant samples are deposited in the collection of microorganisms of the Institute of Microbiology of the Czechoslovak Academy of Sciences in Prague 4 under the designation Claviceps fusiformis under No. CC 1/85.
Mutant byl získán ze 70 isolátů připravených níže popsaným mutačním působením ÚV zářením na laboratorní kmen Claviceps fusiformis MI525 (Mikrobiologický ústav ČSAV v Praze 4).The mutant was obtained from 70 isolates prepared by the UV radiation mutation described below on laboratory strain Claviceps fusiformis MI525 (Institute of Microbiology of the Czechoslovak Academy of Sciences in Prague 4).
Příprava vysokoprodukčního mutanta. Veákeré statické kultivace kmene probíhaly v temnu při 24+1 °C na agarová půdě T2 (C.Spalla, in Genetice of Industrial Microorganisms. Eds.Preparation of high-production mutant. All static cultures of the strain were performed in the dark at 24 + 1 ° C on T2 agar broth (C.Spalla, in Genetics of Industrial Microorganisms. Eds.
Z.Vaněk, Z.HoStálek, U.Cudlin. Elsevier, Amsterdam 1973, p.393). Půda T2 obsahovala tyto složky (g.L-15: sacharosa 00, L-asparagin i0, Ca(N0^)2<4H20 i, kvasničný extrakt 0.1,Z.Vanek, Z.HoStalek, U.Cudlin. Elsevier, Amsterdam 1973, p.393). Soil T2 contained the following components (gL -1 5: sucrose 00, L-asparagine 10, Ca (NO 2) 2 < 4H 2 0, yeast extract 0.1,
KH2PO4 0.25, MgS04.7H20 0.25, KC1 0.12, FeSO4.7H2O 0.02, ZnSO4.7H2O 0.015, agar 20, dest.KH 2 PO 4 0.25, MgSO 4 .7H 2 0 0.25, KCl 0.12, FeSO 4 .7H 2 O 0.02, ZnSO 4 .7H 2 O 0.015, agar 20, dest.
HjO, pH 5,2 po sterilizaci v autoklávu 15· min. při 105 °C.H 2 O, pH 5.2 after autoclaving 15 · min. at 105 ° C.
Spory 21 dsnní rodičovské kultury Claviceps fusiformis HI525, vyrostlé na seěikmená agarové půdě T2 v Endo zkumavkách, byly smyty s povrchu kultury destilovanou vodou. 5 ml takto získané suspenze obsahující přibližně 1,5 .-10’ spor v 1 ml bylo přeneseno na Petriho misku (0 10 cm) a ozářeno zdrojem UV nastaveným na 10 uJ.m“2.s“’ dávkou 0,1 mUvm“2 (1 OOO erg).The spores of the 21 day parental culture of Claviceps fusiformis HI525, grown on sloping T2 agar broth in Endo tubes, were washed off with the surface of the culture with distilled water. 5 ml of the resulting slurry containing about 1.5.-10 'spores in 1 ml was transferred to a petri dish (0 10 cm) and irradiated with UV source set at 10 uJ.m "2 .s'' dose of 0.1 mUvm" 2 (1000 erg).
Ozářená suspenze spor byla vhodně naředžna dest. vodou a spory obsažené v 5 ml výsledné suspenze byly vysety na povrch agarová půdy T2 v Petriho misce (0 10 cm) a inkubovány 21 dní.The irradiated spore suspension was suitably diluted with dist. water and spores contained in 5 ml of the resulting suspension were sown on the surface of T2 agar broth in a Petri dish (0 10 cm) and incubated for 21 days.
Kolonie vyrostlé z autogenisovaných spor, byly pak jednotlivě přeneseny na sešikmenou agarovou půdu T2 a inkubovány 21 až 28 dní.Colonies grown from autogenized spores were then individually transferred to slanted T2 agar broth and incubated for 21-28 days.
Narostlé kultury byly testovány na schopnost tvořit elymoklavin a další klavinové alkaloidy v podmínkách submersní fermentace (Seháček.et, al., čs. AO 199986).Growth cultures were tested for their ability to form elymoclavin and other clavin alkaloids under submersive fermentation conditions (Seháček.et, al., Cz. AO 199986).
Alkaloidy byly stanoveny ve fermentační půdá 21 denních submersnlch kultur kalorimetricky (G.J.Banks et sl., J.gen. Mikrobiol 82,345,1974) a metodou vysokoúčinné kapalinové chromatografie (M.Wurst et, al., J.Chromát. 150,477, 1978).The alkaloids were determined in fermentation broths of 21-day submersible cultures by calorimetry (G. J. Banks et al., J.gen. Microbiol.
Charakteristika vysokoprodukčního mutanta. Srovnání základních znaků vysokoprodukčního kmene W 1 se znaky rodičovského kmene MI 525 je uvedeno v tab. 1.Characteristics of high-production mutant. A comparison of the basic traits of the high-production strain W 1 with the traits of the parental strain MI 525 is given in Tab. 1.
a) Kultivace na agarovém mediu T 2.(a) Cultivation on T 2 agar medium.
Mutant W 1 roste na agarovém mediu T 2 při teplotě 24 +Ί °C v souvislé bohatě zvrásněné vrstvě tvořené šedým myceliem a oválnými konidiemi. Do media vylučuje fialový pigment.Mutant W 1 grows on T 2 agar medium at 24 + Ί ° C in a continuous richly creased layer consisting of gray mycelium and oval conidia. It exudes a violet pigment.
b) Submersní kultivaceb) Submerged cultivation
Submersní kultivace nového kmene W 1 i rodičovského kmene MI 525 byla provedena při teplotě 24+1 °C na rotačním třepacím stroji i v laboratorním fermentoru na upraveném fermentač ním médiu CS2 tis. A0 199986). Submersní kultury kmene W i se vyznačuji vláknitým, větveným mnohobuněčným myceliem, které vytváří četné mikro- a makrokor.idie a na konci fermentace seSubmerged cultivation of both the new strain W 1 and the parent strain MI 525 was carried out at 24 + 1 ° C on a rotary shaker and in a laboratory fermenter on a conditioned fermentation medium CS2 thous. A0 199986). Submerged cultures of strain W 1 are characterized by filamentous, branched, multicellular mycelia which produce numerous micro- and macrocorridia and at the end of fermentation
240612 se rozpadá na blastoarthrospóry. Submersní vývoj kmene W 1 je provázen tvorbou fialového exocelulárního pigaentu.240612 disintegrates into blastoarthrospores. Submerged development of strain W 1 is accompanied by the formation of a violet exocellular pigaent.
Ve srovnáni s rodičovským kmenem Claviceps fusiformis WI 525 se mutant W t vyznačuje téměř trojnásobně vyěší produkci olymoklavinu a polovičním obsahem nežádoucích exogenních glukanů. V podmínkách submersní fermentace produkuje v 1 1 media 4,2 g klavinových alkaloidů, * tohe 2,5 g elymoklavinu. Průvodní alkaloidy jsou agroklavin a chanoklavin. Srovnání základních znaků vysokoprodukčního mutanta W 1 se znaky rodičovského kmene C.fusiformis W1525 je uvedeno v tab. 1. Mutant podle vynálezu je uchován v temnu při 4 °C, zabezpečen proti vysychání media, a to ve formě 21 denních kultur vyrostlých na seěikmenám mediu T2 v Endo zkumavkách při teplotě 24 + 1 °C. Uchovávaná kultury jsou pravidelně přeočkovávány ve tříměsíčních intervalech.Compared to the parental strain Claviceps fusiformis WI 525, the mutant W t is characterized by almost three times higher olymoclavin production and half the content of unwanted exogenous glucans. Under submersive fermentation conditions, 4.2 g of clavin alkaloids, this 2.5 g of elymoclavin, are produced in 1 L of medium. The accompanying alkaloids are agroclavin and chanoclavin. A comparison of the basic traits of the high-production mutant W 1 with those of the parental strain C.fusiformis W1525 is shown in Tab. 1. The mutant according to the invention is stored in the dark at 4 ° C, protected against drying out of the medium in the form of 21 day cultures grown on seedlings of medium T2 in Endo tubes at 24 + 1 ° C. Preserved cultures are regularly revaccinated at three-month intervals.
Vynálezem chráněný vysokoprodukční mutant Claviceps fusiformis W 1, zabezpečuje klíčový úsek velkokapacitní přípravy elymoklavinu submersní fermentací.The high-production mutant Claviceps fusiformis W 1 protected by the invention provides a key section of the large-scale preparation of elymoclavin by submerged fermentation.
Tabulka 1Table 1
Kultura na agarové půdě T2 Submersní kultura^Culture on agar medium T2 Submerged culture ^
ZnakCharacter
tg.L-'ltg.L-1
PŘEDMĚT V Y H i. L SZUSUBJECT V Y H i L SZU
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CS883284A CS248612B1 (en) | 1984-11-19 | 1984-11-19 | Industrial mutant of sponge claviceps fusiformis w1 cc-1-85 |
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CS883284A CS248612B1 (en) | 1984-11-19 | 1984-11-19 | Industrial mutant of sponge claviceps fusiformis w1 cc-1-85 |
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CS248612B1 true CS248612B1 (en) | 1987-02-12 |
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1984
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