NO133271B

NO133271B -

Info

Publication number: NO133271B
Application number: NO1464/69A
Authority: NO
Inventors: F Arcamone; G Franceschi; S Penco
Original assignee: Farmaceutici Italia
Priority date: 1968-04-12
Filing date: 1969-04-10
Publication date: 1975-12-29
Also published as: FR2007443A1; DK126987B; DE1917874B2; IL31966A; YU85869A; YU34389B; SE359293B; CH518250A; ES365907A1; BE731398A; NL6904844A; DE1917874A1; NO133271C; DE1917874C3; GB1217133A; AT283597B; NL145536B; IL31966A0

Description

Foreliggende oppfinnelse angår en ny fremgangsmåte ved The present invention relates to a new method by

fremstilling av adriamycin og dets aglyconadriamycinon. preparation of adriamycin and its aglyconadriamycinone.

Adriamycin er et kjent antibiotikum med bakterostatisk og antitumoral virkning, som sammen med sine derivater er beskrevet i belgisk patentskrift nr. 713.773. Adriamycin og dets aglycon har folgende strukturformel: hvor R er hydrogen eller Adriamycin is a known antibiotic with bacteriostatic and antitumoral action, which, together with its derivatives, is described in Belgian patent document no. 713,773. Adriamycin and its aglycon have the following structural formula: where R is hydrogen or

Det ovennevnte patentskrift beskriver dessuten en mikro-biologisk fremgangsmåte for fremstilling av adriamycin, hvor mik-roorganismen Streptomyces peucetius var. caesius benyttes. Der er nu overraskende funnet en ny syntese for fremstilling av adriamycin og dets aglycon, ved hvilken der startes med henholdsvis an-tibiotikumet daunomycin og dets aglycon daunomycinon. Daunomycin og dets aglycon daunomycinon er beskrevet i britisk patentskrift nr. 1.003.383. The above-mentioned patent also describes a microbiological method for the production of adriamycin, in which the microorganism Streptomyces peucetius var. caesius is used. A new synthesis has now surprisingly been found for the production of adriamycin and its aglycone, which starts with the antibiotic daunomycin and its aglycone daunomycinon, respectively. Daunomycin and its aglycon daunomycinone are described in British Patent No. 1,003,383.

Fremgangsmåten ifolge oppfinnelsen er kjennetegnet ved at daunomycin, daunomycinon eller deres derivater av strukturformelen: The method according to the invention is characterized by daunomycin, daunomycinone or their derivatives of the structural formula:

hvor R' er hydrogen eller radikalet og X er omsettes i et inert organisk opplosningsmiddel som ikke fremkaller hydrolyse av glucosidbindingen, med brom eller jod, i det siste tilfelle fortrinnsvis i nærvær av en base slik som kalsiumoxyd, og at det erholdte 14-brom eller 14-jod-derivat enten a) omsettes med et alkalimetallacetat i nærvær av et polart opplosningsmiddel hvorefter det tilsvarende 14-acetoxyderivat overfores ved alkalisk hydrolyse til 14-hydroxyderivåtet, eller b) overfores ved alkalisk hydrolyse til 14-hydroxyderivåtet, hvorefter produktet erholdt i a) eller b) isoleres og renses, where R' is hydrogen or the radical and X is reacted in an inert organic solvent which does not induce hydrolysis of the glucosidic bond, with bromine or iodine, in the latter case preferably in the presence of a base such as calcium oxide, and that 14-bromo or 14-iodo derivative is either a) reacted with an alkali metal acetate in the presence of a polar solvent, after which the corresponding 14-acetoxy derivative is transferred by alkaline hydrolysis to the 14-hydroxy derivative, or b) transferred by alkaline hydrolysis to the 14-hydroxy derivative, after which the product obtained in a) or b) isolated and cleaned,

som, når utgangsma terialet er N-salicyliden-daunomycin, utfores ved eliminering av salicyliden-gruppen ved syrehydrolyse under dannelse av adriamycin, eller som når utgangsmaterialet er N-trifluoracetyl-daunomycin, utfores ved eliminering which, when the starting material is N-salicylidene-daunomycin, is carried out by elimination of the salicylidene group by acid hydrolysis to form adriamycin, or which, when the starting material is N-trifluoroacetyl-daunomycin, is carried out by elimination

av trifluoracetylgruppen ved at det erholdte N-trifluoracetyladriamycin behandles med ethylorthoformiat og p-toluensulfonsyre, og det erholdte N-trifluoracety ladriamycin-9,14-ethylorthoformiat underkastes alkalisk hydrolyse, hvorefter det erholdte adriamycin-9,14-ethylorthoformiat underkastes syrehydrolyse under dannelse av adriamycin. of the trifluoroacetyl group by treating the obtained N-trifluoroacetyladriamycin with ethylorthoformate and p-toluenesulfonic acid, and the obtained N-trifluoroacetyladriamycin-9,14-ethylorthoformate is subjected to alkaline hydrolysis, after which the obtained adriamycin-9,14-ethylorthoformate is subjected to acid hydrolysis to form adriamycin .

Når det gjelder antibiotika av den cycliske type har der tidligere ikke vært utfort en substitusjon av et hydrogen-atom med et hydroxylradika1 ved en fremgangsmåte analog med fo-religgende. Ennvidere må der taes i betraktning at daunomycin er en forbindelse med flere forskjellige reaktive kjemiske funk-sjoner og som er karakterisert ved en bemerkelsesverdig ustabilitet overfor oxydasjonsmidler, reduksjonsmidler, varme, baser, og forst og fremst syrer. In the case of antibiotics of the cyclic type, a substitution of a hydrogen atom with a hydroxyl radical1 by a method analogous to the present has not previously been carried out. Furthermore, it must be taken into account that daunomycin is a compound with several different reactive chemical functions and which is characterized by a remarkable instability towards oxidizing agents, reducing agents, heat, bases and above all acids.

Spesielt ustabiliteten overfor syrer, hvilken forårsa-ker splitting av glucosidbindingen, er meget viktig ved betraktning av bromeringsreaksjonen. Especially the instability towards acids, which causes splitting of the glucosidic bond, is very important when considering the bromination reaction.

Når en forbindelse bromeres for å erstatte et hydrogen-atom, skjer der alltid en dannelse av hydrogenbromid ifolge re-aksjon s skj ema et : When a compound is brominated to replace a hydrogen atom, hydrogen bromide is always formed according to the reaction scheme:

Da daunomycin er karakterisert ved ustabilitet overfor syrer, ville hydrogenbromidet (eller rettere hydrobromsyren) kunne hydrolysere glucosidbindingen i molekylet. As daunomycin is characterized by instability towards acids, the hydrogen bromide (or rather the hydrobromic acid) could hydrolyse the glucosidic bond in the molecule.

Det var derfor nddvendig å finne frem til en forbindelse som var istand til å bibeholde hydrobromsyren og samtidig væ-re et godt opplosningsmiddel. Dette ble oppnådd ved anvendelse av blandingen av methanolrdioxan. It was therefore necessary to find a compound that was able to retain the hydrobromic acid and at the same time be a good solvent. This was achieved by using the mixture of methanolrdioxane.

Dioxanet i blandingen bibeholder syren og forhindrer hydrolyse av glucosidbindingen. The dioxane in the mixture retains the acid and prevents hydrolysis of the glucosidic bond.

Vedrorende daunomycinon, inneholder denne forbindelse ingen glucosidbinding og krever derfor ikke denne spesielle opp-losningsmiddelblanding. Det efterfolgende eksempel 3 er derfor illustrert ved anvendelse av kloroform som opplosningsmiddel, fordi hydrolyse i dette tilfelle ikke kan finne sted. Regarding daunomycinone, this compound contains no glucosidic bond and therefore does not require this particular solvent mixture. The following example 3 is therefore illustrated using chloroform as solvent, because hydrolysis cannot take place in this case.

Når det gjelder daunomycin's ustabilitet overfor oxydasjonsmidler, er det å bemerke at daunomycin kan betraktes som et hydrokinonderivat, og hydrokinonets ustabilitet overfor oxydasjonsmidler er velkjent innen faget. Hvis således reaksjonen under alkalihydrolysen av 14-bromderivåtet av daunomycin ikke utfores som beskrevet i de efterfolgende eksempler, vil der finne sted en dannelse av nafthacenkinon. Regarding daunomycin's instability to oxidizing agents, it should be noted that daunomycin can be considered a hydroquinone derivative, and hydroquinone's instability to oxidizing agents is well known in the art. Thus, if the reaction during the alkali hydrolysis of the 14-bromo derivative of daunomycin is not carried out as described in the following examples, a formation of naphthacene quinone will take place.

Adriamycin og adriamycinon som fåes ved fremgangsmåten ifolge oppfinnelsen, isoleres og renses efter de kjente ekstrak-sjons- og rensemetoder. Adriamycin and adriamycinone obtained by the method according to the invention are isolated and purified according to the known extraction and purification methods.

De folgende eksempler illustrerer oppfinnelsen: The following examples illustrate the invention:

Eksempel 1 ADRIAMYCIN Example 1 ADRIAMYCIN

1,30 g daunomycin-hydroklorid i 30 ml methylalkohol og lOO ml vannfritt dioxan behandles med 3,3 ml av en bromopplosning i kloroform (10 g brom i lOO ml kloroform). Reaksjonsblan-dirigen tillates å stå i 4 timer ved romtemperatur, hvorefter den inndampes til tdrrhet under redusert trykk. Residuet tas opp 1.30 g of daunomycin hydrochloride in 30 ml of methyl alcohol and 100 ml of anhydrous dioxane are treated with 3.3 ml of a bromine solution in chloroform (10 g of bromine in 100 ml of chloroform). The reaction mixture is allowed to stand for 4 hours at room temperature, after which it is evaporated to dryness under reduced pressure. The residue is taken up

med 5 - 10 ml kloroform, og 3 - 5 volumdeler ethylether tilsettes. Der utfelles 1,10 g brom-daunomycin-hydroklorid, som ora-krystalliseres fra kloroform-methanol. Smeltepunkt 177 - 178°C. with 5 - 10 ml of chloroform, and 3 - 5 parts by volume of ethyl ether are added. There, 1.10 g of bromo-daunomycin hydrochloride is precipitated, which is ora-crystallised from chloroform-methanol. Melting point 177 - 178°C.

En opplosning av 1,0 g 14-brcm daunomycin-hydroklorid i 150 ml methylalkohol behandles med 70 ml destillert vann, hvorefter der tilsettes 0,1 N natriumhydroxyd inntil pH 10,3, hvilken pH-verdi opprettholdes-20 minutter. Det hele utfores under nitrogenatmosfære. Efter fortynning med destillert vann ekstraheres oppldsningen flere ganger med kloroform inntil alt det farvede materiale er gått over i opplosningsmidlet. De sammenslåtte kloroformekstrakter torres over vannfritt natriumsulfat, konsentreres under redusert trykk til et lite volum, tilsettes 1,8 ml 0,6 N saltsyre i vannfri methanol og behandles med 10 volumdeler ethylether. Der erholdes O,72 g av en amorf utfellning som utkrystallisert fra methanol-propanol gir 0,45 g adriamycin-hydroklorid som smelter ved 205 - 210°C (under spaltning). A solution of 1.0 g of 14-brcm daunomycin hydrochloride in 150 ml of methyl alcohol is treated with 70 ml of distilled water, after which 0.1 N sodium hydroxide is added until pH 10.3, which pH value is maintained for 20 minutes. It is all carried out under a nitrogen atmosphere. After dilution with distilled water, the solution is extracted several times with chloroform until all the colored material has passed into the solvent. The combined chloroform extracts are dried over anhydrous sodium sulfate, concentrated under reduced pressure to a small volume, 1.8 ml of 0.6 N hydrochloric acid in anhydrous methanol are added and treated with 10 parts by volume of ethyl ether. 0.72 g of an amorphous precipitate is obtained which, crystallized from methanol-propanol, gives 0.45 g of adriamycin hydrochloride which melts at 205 - 210°C (under decomposition).

Eksempel 2 ADRIAMYCIN Example 2 ADRIAMYCIN

En suspensjon av 6,60 g 14-bromdaunomycin-hydroklorid fremstillet i henhold til eksempel 1 i 200 ml vannfritt aceton tilsettes 1,8 g smeltet og pulverisert kaliumacetat. Suspensjo-nen filtreres efter å være oppvarmet med tilbakelopskjoling i 45 minutter, og den erholdte opplosning inndampes til torrhet under redusert trykk. Residuet tas opp med kloroform og behandles med ethylether. Utfellningen som fåes i en mengde av 0,45 g, renses ved filtrering gjennom en kiselsyresdyle under anvendelse av methylenklorid-methanol-vann i mengdeforholdet 100:20:2 som opplosningsmiddel-system. Der erholdes 150 mg 14-acetoxydaunomycin som smelter ved 196 - 198°C. To a suspension of 6.60 g of 14-bromodaunomycin hydrochloride prepared according to Example 1 in 200 ml of anhydrous acetone is added 1.8 g of melted and powdered potassium acetate. The suspension is filtered after being heated with reflux cooling for 45 minutes, and the resulting solution is evaporated to dryness under reduced pressure. The residue is taken up with chloroform and treated with ethyl ether. The precipitate, which is obtained in an amount of 0.45 g, is purified by filtration through a silicic acid filter using methylene chloride-methanol-water in the ratio of 100:20:2 as solvent system. 150 mg of 14-acetoxydaunomycin is obtained which melts at 196 - 198°C.

En opplosning av 0,lO g 14-acetoxydaunomycin i 30 ml aceton-methanol (2:1) tilsettes 10 ml av en 5 %'s natriumbicar« bonatopplosning. Blandingen tillates å stå 3 timer ved romtemperatur, hvorefter den ekstraheres grundig med kloroform efter fortynning med vann. Kloroformekstraktene slåes sammen og torres over vannfritt natriumsulfat, hvorefter de konsentreres til et lite volum under redusert trykk. Den konsentrerte opplosning tilsettes O,15 ml 0,6 N saltsyre i vannfri methanol og derefter 3 volumdeler ethylether. Den amorfe utfellning fores gjennom en kolonne fyllt med 40.g cellulosepulver. Det elueres med opplos-ningssystemet propanoltethylacetat:vann i volumforholdet 7:1:2. Fraksjonene som inneholder adriamycin, slåes sammen og inndampes til torrhet, hvorved der erholdes 43 mg adriamycin-hydroklorid som smelter ved 204 - 210°C (under spaltning). A solution of 0.10 g of 14-acetoxydaunomycin in 30 ml of acetone-methanol (2:1) is added to 10 ml of a 5% sodium bicarbonate solution. The mixture is allowed to stand for 3 hours at room temperature, after which it is thoroughly extracted with chloroform after dilution with water. The chloroform extracts are combined and dried over anhydrous sodium sulphate, after which they are concentrated to a small volume under reduced pressure. 0.15 ml of 0.6 N hydrochloric acid in anhydrous methanol and then 3 parts by volume of ethyl ether are added to the concentrated solution. The amorphous precipitate is passed through a column filled with 40 g of cellulose powder. It is eluted with the solvent system propanol/ethyl acetate:water in the volume ratio 7:1:2. The fractions containing adriamycin are combined and evaporated to dryness, whereby 43 mg of adriamycin hydrochloride is obtained which melts at 204 - 210°C (under decomposition).

Eksempel 3 ADRIAMYCINON Example 3 ADRIAMYCINONE

1 g daunomycinon opplbses i lOO ml kloroform og tilsettes 6,75 ml av en bromopplosning i kloroform (2 ml brom opplost i ICO ml kloroform). Efter natten over ved romtemperatur fra-filtreres det dannede krystallinske produkt, som derefter om-krysta lliseres fra ethylacetat. Der erholdes 1 g 14-brcm-daunomycinon med smeltepunkt 220 - 225°C (under spaltning). 1 g of daunomycinone is dissolved in 100 ml of chloroform and 6.75 ml of a bromine solution in chloroform is added (2 ml of bromine dissolved in 10 ml of chloroform). After overnight at room temperature, the formed crystalline product is filtered off, which is then recrystallized from ethyl acetate. 1 g of 14-brcm-daunomycinone with a melting point of 220 - 225°C (under decomposition) is obtained.

[a]p = + 165° (c = 0,1 i dioxan). [a]p = + 165° (c = 0.1 in dioxane).

0,6 g 14-brom-daunomycinon oppslemmes i 300 ml vannfritt aceton og tilsettes 1,3 g smeltet kaliumacetat. Blandin- 0.6 g of 14-bromo-daunomycinone is suspended in 300 ml of anhydrous acetone and 1.3 g of molten potassium acetate is added. Blendin-

gen oppvarmes med tilbakelbpskjoling i 30 minutter, hvorefter oppløsningen filtreres og inndampes til torrhet under redusert trykk. Residuet krystalliseres fra ethylacetat, hvorved der erholdes 0,5 g 14-acetoxy-daunomycinon som smelter ved 242 -245°C. is heated with reflux for 30 minutes, after which the solution is filtered and evaporated to dryness under reduced pressure. The residue is crystallized from ethyl acetate, whereby 0.5 g of 14-acetoxy-daunomycinone is obtained which melts at 242-245°C.

[a]p = + 192° (c = 0,1 i dioxan). [a]p = + 192° (c = 0.1 in dioxane).

O,1 g 14-acetoxydaunomycinon opploses i aceton og tilsettes IO ml av en IO %'s opplosning av natriumbicarbonat. Efter 3 timer ved romtemperatur ekstraheres reaksjonsblandingen med kloroform. Kloroformekstrakten torres og konsentreres til et lite volum under redusert trykk. Den kromatograferes derefter gjennom en kiselsyresoyle for den elueres med kloroform inneholdende 2 % ethanol for å fjerne utgangsmateriale som fortsatt måtte være tilstede. Derefter elueres adriamycinonet med kloroform som inneholder 5 % ethanol. Der erholdes 50 mg produkt som ved krystallisering fra ethylacetat smelter ved 228 - 230°C. 0.1 g of 14-acetoxydaunomycinone is dissolved in acetone and 10 ml of a 10% solution of sodium bicarbonate is added. After 3 hours at room temperature, the reaction mixture is extracted with chloroform. The chloroform extract is dried and concentrated to a small volume under reduced pressure. It is then chromatographed through a silica gel eluting with chloroform containing 2% ethanol to remove any starting material that may still be present. The adriamycinone is then eluted with chloroform containing 5% ethanol. 50 mg of product is obtained which, upon crystallization from ethyl acetate, melts at 228 - 230°C.

[a]^ = + 156° (c = 0,1 i dioxan). [a]^ = + 156° (c = 0.1 in dioxane).

Eksempel 4 ADRIAMYCINON Example 4 ADRIAMYCINONE

0,1 g 14-bromdaunomycinon erholdt i henhold til eksempel 3 ved behandling med IO ml 0,1 N natronlut ved 0°C i 15 minutter, overfores til adriamycinon, som isoleres ved kromatogra-fering gjennom en kiselsyresoyle. 0.1 g of 14-bromodaunomycinone obtained according to example 3 by treatment with 10 ml of 0.1 N caustic soda at 0° C. for 15 minutes is transferred to adriamycinone, which is isolated by chromatography through a silica gel.

Eksempel 5 ADRIAMYCIN Example 5 ADRIAMYCIN

4,6 g daunomycin, i form av fri base i 390 ml kloroform og 155 ml vannfri ether, behandles med 9,3 ml trifluoreddiksyre-anhydrid. Reaksjonsblandingen tillates å stå ved romtemperatur i 30 minutter, hvorefter den ekstraheres med vann, og den organiske fase inndampes til torrhet under vakuum. Residuet tas opp med 100 ml vannfri methanol og oppvarmes méd tilbakelop i IO minutter. Methanolopplosningen destilleres til torrhet, og residuet krystalliseres fra tetrahydrofuran-petroleum, hvorved der erholdes 4,5 g N-fluoracetyldaunomycin som smelter ved 169-171°C. 4.6 g of daunomycin, in the form of free base in 390 ml of chloroform and 155 ml of anhydrous ether, is treated with 9.3 ml of trifluoroacetic anhydride. The reaction mixture is allowed to stand at room temperature for 30 minutes, after which it is extracted with water, and the organic phase is evaporated to dryness under vacuum. The residue is taken up with 100 ml of anhydrous methanol and heated at reflux for 10 minutes. The methanol solution is distilled to dryness, and the residue is crystallized from tetrahydrofuran-petroleum, whereby 4.5 g of N-fluoroacetyldaunomycin is obtained which melts at 169-171°C.

0,6 g N-trifluoracetyldaunomycin i 30 ml tetrahydrofuran og 20 ml vannfri methanol tilsettes 0,50 g jod og 0,50 g pulverisert kalsiumoxyd. Blandingen rystes under nitrogenatmosfære i 5 timer, hvorefter den filtreres, og oppldsningen efter 0.6 g of N-trifluoroacetyl daunomycin in 30 ml of tetrahydrofuran and 20 ml of anhydrous methanol is added to 0.50 g of iodine and 0.50 g of powdered calcium oxide. The mixture is shaken under a nitrogen atmosphere for 5 hours, after which it is filtered, and the solution is left

fortynning med kloroform rystes med vann under surgjbring med saltsyre til omslag med lakmus. Den organiske fase inndampes til torrhet under vakuum, og residuet tas opp med litt kloroform og kromatograferes gjennom en sbyle ved eluering med kloroform inneholdende okende mengder ethylacetat. Det bnskede produkt elueres når konsentrasjonen av ethylacetat er ca. 10 %. Der erholdes 0,25 g 14-jod-N-trifluoracetyldaunomycin som smelter ved 172 - 175°C. dilution with chloroform is shaken with water while acidifying with hydrochloric acid to cover with litmus. The organic phase is evaporated to dryness under vacuum, and the residue is taken up with a little chloroform and chromatographed through a funnel by elution with chloroform containing increasing amounts of ethyl acetate. The desired product is eluted when the concentration of ethyl acetate is approx. 10%. 0.25 g of 14-iodo-N-trifluoroacetyldaunomycin is obtained which melts at 172 - 175°C.

0,8 g av denne forbindelse oppvarmes med tilbakeldp i 15 minutter med 60 ml vannfritt aceton i nærvær av 1,50 g smeltet og pulverisert kaliumacetat. Efter filtrering tilsettes opp-løsningen kloroform og rystes med vann under langsom surgjdring med saltsyre til omslag med lakmus. Den organiske fase inndampes nesten til torrhet under vakuum, og residuet tilsettes petrolether. Der utfelles 0,62 g 14-acetoxy-N-trifluoracetyldaunomycin som smelter ved 150 - 153°C. 0.8 g of this compound is refluxed for 15 minutes with 60 ml of anhydrous acetone in the presence of 1.50 g of molten and powdered potassium acetate. After filtration, chloroform is added to the solution and shaken with water while slowly acidifying with hydrochloric acid to cover with litmus. The organic phase is evaporated almost to dryness under vacuum, and petroleum ether is added to the residue. There, 0.62 g of 14-acetoxy-N-trifluoroacetyldaunomycin is precipitated, which melts at 150 - 153°C.

1,4 g av dette produkt opploses i en blanding av 50 ml methanol og 100 ml aceton og tilsettes 100 ml av en 5 %'s natri-umbicarbonatoppldsning. Blandingen holdes ved romtemperatur og under nitrogenatmosfære i 3 timer; Reaksjonsblandingen helles derefter over i vann, surgjores med vandig vinsyre til en pH-verdi mellom 5 og 6, og ekstraheres.flere ganger med kloroform, idet der totalt anvendes 350 ml av opplosningsmidlet. De organiske faser slåes sammen og inndampes under redusert trykk, hvorved der erholdes 0,95 g av et fast residuum som tas opp med kloroform og kromatograferes. Det således erholdte N-trifluoracetyladriamycin elueres med kloroform som inneholder 15 % ethylacetat. De sammenslåtte og inndampede eluater gir 0,56 g av det onskede produkt som smelter ved 174 - 176°C. 1.4 g of this product is dissolved in a mixture of 50 ml of methanol and 100 ml of acetone and 100 ml of a 5% sodium bicarbonate solution is added. The mixture is kept at room temperature and under a nitrogen atmosphere for 3 hours; The reaction mixture is then poured into water, acidified with aqueous tartaric acid to a pH value between 5 and 6, and extracted several times with chloroform, using a total of 350 ml of the solvent. The organic phases are combined and evaporated under reduced pressure, whereby 0.95 g of a solid residue is obtained which is taken up with chloroform and chromatographed. The N-trifluoroacetyladriamycin thus obtained is eluted with chloroform containing 15% ethyl acetate. The combined and evaporated eluates give 0.56 g of the desired product which melts at 174-176°C.

0,45 g N-trifluoracetyladriamycin i 80 ml dioxan tilsettes IO ml ethylorthoformiat og 3 mg p-toluensulfonsyre. Efter 48 timer ved romtemperatur fortynnes reaksjonsblandingen med kloroform og vaskes med vann under kraftig rysting i en skilletrakt. Den organiske fase inndampes til et lite volum, hvorefter den ved tilsetning av petrolether gir 35 g utfellning. Dette produkt kromatograferes gjennom en sdyle av 7 g kiselsyre ved eluering med kloroform som inneholder 10 % ethylacetat. Det 0.45 g of N-trifluoroacetyladriamycin in 80 ml of dioxane is added to 10 ml of ethyl orthoformate and 3 mg of p-toluenesulfonic acid. After 48 hours at room temperature, the reaction mixture is diluted with chloroform and washed with water under vigorous shaking in a separatory funnel. The organic phase is evaporated to a small volume, after which it gives 35 g of precipitation when petroleum ether is added. This product is chromatographed through a column of 7 g silicic acid, eluting with chloroform containing 10% ethyl acetate. The

inndampede eluat gir 80 mg 9,14-N-trifluoracetyladriamycin-ethylklorf ormiat som smelter ved 162 - 164°C. evaporated eluates give 80 mg of 9,14-N-trifluoroacetyladriamycin ethyl chloroformate which melts at 162-164°C.

0,20 g av dette produkt opploses i en blanding av 10 ml aceton og 40 ml O,1 N natriumhydroxyd under en atmosfære av rent nitrogen. Efter 20 minutter ved romtemperatur innstilles pH-ver-dien på 8,6 med 1 N saltsyre, og oppldsningen ekstraheres grundig mange ganger med 50 ml kloroform, inntil ekstrakten ikke len-ger farves. De sammenslåtte kloroformekstrakter inndampes til et lite volum og fores gjennom en soyle inneholdende 4 g kiselsyre. Elueringen utfores med oppldsningsmiddelblandingen methylenklorid-methanol-vann i volumforholdet 50:10:1. Ved inndampning av eluatet fåes O,11 g adriamycin-9,14-ethylorthoformiat som smelter ved 203 - 206°C. 0.20 g of this product is dissolved in a mixture of 10 ml of acetone and 40 ml of 0.1 N sodium hydroxide under an atmosphere of pure nitrogen. After 20 minutes at room temperature, the pH value is adjusted to 8.6 with 1 N hydrochloric acid, and the solution is thoroughly extracted many times with 50 ml of chloroform, until the extract is no longer coloured. The combined chloroform extracts are evaporated to a small volume and passed through a sieve containing 4 g of silicic acid. The elution is carried out with the solvent mixture methylene chloride-methanol-water in the volume ratio 50:10:1. Evaporation of the eluate yields 0.11 g of adriamycin-9,14-ethylorthoformate which melts at 203 - 206°C.

0,10 g av dette produkt i 9 ml aceton tilsettes 1 ml 0.10 g of this product in 9 ml of acetone is added to 1 ml

1 N saltsyre. Blandingen tillates å stå ved romtemperatur i 20 timer. Utfellingen som dannes, krystalliseres fra methanol-propanol,. hvorved der fåes 40 mg adriamycin-hydroklorid med smeltepunkt 204 - 210°C (spaltes). 1 N hydrochloric acid. The mixture is allowed to stand at room temperature for 20 hours. The precipitate that forms is crystallized from methanol-propanol. thereby obtaining 40 mg of adriamycin hydrochloride with a melting point of 204 - 210°C (decomposed).

Eksempel 6 ADRIAMYCIN Example 6 ADRIAMYCIN

l,O0 g daunomycin-hydroklorid i 30 ul destillert vann tilsettes 0,1 N natriumhydroxyd til pH 8,6 og behandles derefter med IO ml av en IO %'s opplosning av salicylaldehyd i methanol. Utfellingen tas opp i og vaskes med vann, hvorefter den krystalliseres fra ethanol. Der erholdes 0,97 g salicylidendaunomycin som smelter ved 168 - 170°C. 1.00 g of daunomycin hydrochloride in 30 ul of distilled water is added to 0.1 N sodium hydroxide to pH 8.6 and then treated with 10 ml of a 10% solution of salicylaldehyde in methanol. The precipitate is taken up in and washed with water, after which it is crystallized from ethanol. 0.97 g of salicylidene daunomycin is obtained which melts at 168 - 170°C.

0,50 g salicylidendaunomycin i 20 ml tetrahydrofuran tilsettes IO ml vannfri methanol, 1,0 g jod og 1,0 g kalsiumoxyd. Blandingen kokes forsiktig med tilbakeldpskjdling i 40 minutter, hvorefter den filtreres og inndampes til torrhet under redusert trykk. Residuet tas opp med 22 ml kloroform. Kloroformoppldsningen vaskes med vann, rystes og tdrres, hvorefter den konsentreres til et lite volum og tilsettes pétrol-ether for fullstendig utfelling. Der erholdes 0,41 g urent 14-jod-salicylidendaunomycin. Produktet tas opp med 20 ml vannfritt aceton, behandles med 0,80 g smeltet og pulverisert kaliumacetat og kokes med tilbakelopskjoling i 20 minutter. Suspen- 0.50 g of salicylidene daunomycin in 20 ml of tetrahydrofuran is added to 10 ml of anhydrous methanol, 1.0 g of iodine and 1.0 g of calcium oxide. The mixture is carefully refluxed for 40 minutes, after which it is filtered and evaporated to dryness under reduced pressure. The residue is taken up with 22 ml of chloroform. The chloroform solution is washed with water, shaken and dried, after which it is concentrated to a small volume and petroleum ether is added for complete precipitation. 0.41 g of impure 14-iodo-salicylidene daunomycin is obtained. The product is taken up with 20 ml of anhydrous acetone, treated with 0.80 g of melted and powdered potassium acetate and refluxed for 20 minutes. Suspense-

sjonen avkjbles derefter og filtreres, og filtratet inndampes til torrhet under vakuum. Residuet som utgjores av 14-acetoxy-salicylidendaunomycin, opploses i en blanding av like deler methanol og aceton og tilsettes 18 ml av en 5 %'s vandig opplosning av natriumbicarbonat. Reaksjonsblandingen ekstraheres med kloroform efter 3 timer ved romtemperatur. Kloroformekstrakten konsentreres til ca. 10 ml og kromatograferes gjennom en sdyle av 3 g kiselsyre. Den elueres med oppldsningsmiddelblandingen methylenklorid-petrolethermethanol i volumforholdet 100:50:5. Fraksjonene som inneholder utgangsmaterialet og andre forurensninger, kastes, mens de derpå fdlgende fraksjoner oppsamles.. Ved inndampning gir disse 65 mg salicylidenadriamycin som smelter ved 194°C (spaltes) . the reaction is then cooled and filtered, and the filtrate is evaporated to dryness under vacuum. The residue consisting of 14-acetoxy-salicylidene daunomycin is dissolved in a mixture of equal parts methanol and acetone and 18 ml of a 5% aqueous solution of sodium bicarbonate is added. The reaction mixture is extracted with chloroform after 3 hours at room temperature. The chloroform extract is concentrated to approx. 10 ml and chromatographed through a sieve of 3 g silicic acid. It is eluted with the solvent mixture methylene chloride-petroleum methanol in the volume ratio 100:50:5. The fractions containing the starting material and other impurities are discarded, while the following fractions are collected. When evaporated, these give 65 mg of salicylidene adriamycin which melts at 194°C (decomposes).

0,20 g salicylidenadriamycin i 20 ml kloroform ekstraheres tre ganger med porsjoner å IO ml O,1 N saltsyre. Alt det farvede produkt overfores til den vandige fase, som, under ni-trogena tmosf ære, gjdres alkalisk til pH 8,6 med O,1 N natriumhydroxyd og ekstraheres med kloroform inntil ekstrakten er farve-lds. Ekstraktene slåes sammen og tbrres over vannfritt natriumsulfat, hvorefter de konsentreres under redusert trykk til ca. 5 0.20 g of salicylidene adriamycin in 20 ml of chloroform is extracted three times with portions of 10 ml of 0.1 N hydrochloric acid. All the colored product is transferred to the aqueous phase, which, under a nitrogen atmosphere, is made alkaline to pH 8.6 with 0.1 N sodium hydroxide and extracted with chloroform until the extract is colored. The extracts are combined and filtered over anhydrous sodium sulphate, after which they are concentrated under reduced pressure to approx. 5

ml og tilsettes 0,5 ml 0,6 N saltsyre og 10 volumdeler vannfri ethylether. Der erholdes 0,12 g adriamycin-hydroklorid som smelter ved 205 - 210°C (spaltes). ml and add 0.5 ml of 0.6 N hydrochloric acid and 10 parts by volume of anhydrous ethyl ether. 0.12 g of adriamycin hydrochloride is obtained which melts at 205 - 210°C (decomposes).

Claims

Procedure for the production of adriamycin and adriamycinone of the structural formula

where R is hydrogen or characterized by that daunomycin, daunomycinone or their derivatives of the structural formula: where R<1> is hydrogen or the radical is reacted in an inert organic solvent which does not induce hydrolysis of the glucosidic bond, with bromine or iodine, in the latter case preferably in the presence of a base such as calcium oxide, and that the 14-bromo or 14-iodo derivative obtained is either a) reacted with an alkali metal acetate in the presence of a polar solvent after which the corresponding 14-acetoxy derivative is transferred by alkaline hydrolysis to the 14-hydroxy derivative, or b) transferred by alkaline hydrolysis to the 14-hydroxy derivative, after which the product obtained in a) or b) is isolated and purified, which, when the starting material is N-salicylidene-daunomycin, is carried out by elimination of the salicylidene group by acid hydrolysis to form adriamycin, or as when the starting material is N-trifluoroacetyl-daunomycin, is carried out by elimination of the trifluoroacetyl group by treating the obtained N-trifluoroacetyladriamycin with ethyl orthoformate and p -toluenesulfonic acid, and the obtained N-trifluoroacetyladriamycin-9, 14-ethylorthoformate submit es alkaline hydrolysis, after which the obtained adriamycin-9,14-ethylorthoformate is subjected to acid hydrolysis to form adriamycin.