DE1445689B - Process for the production of the alkaloids Leurosidin and Leuro cnstin, from their salts with acids and their quaternary ammonium salts - Google Patents

Description

Achieve ailments that accompany various cancerous diseases. Among the cancerous diseases which can be treated favorably are Hodgkin's disease, acute lymphocytosis (Leukemia), myelomatosis (formation of multiple myelomata), lymphosarcomatosis, astrocytomatosis and the cervical and prostate cancer. In Hodgkin's disease and leukemia In many cases the symptoms of the disease subside and in carcinomas and sarcomas a decrease in tumor size or a cessation of tumor growth can be determined.

example

a) Obtaining Leurocristin and Leurosidin

from one to 90% from leaves

existing plant material

800 kg of Vinca rosea plant material, which is about 90% leaves and 10% stems was moistened with 540 liters of 2% aqueous tartaric acid. The moistened leaves were extracted by stirring with five 4000 liter portions of benzene. The benzene extracts were combined and concentrated in vacuo to a volume of approximately 500 liters. To the benzene concentrate 1070 liters of 2% aqueous tartaric acid were added and the remainder of the Benzene removed by vacuum steam distillation. The acidic, aqueous concentrate was used for removal Insoluble material filtered and then filtered by the addition of 28% aqueous ammonia adjusted to a pH of about 3. The acidic solution was extracted with two 540 liter portions of benzene. The benzene extracts were separated and discarded. Then the pH of the aqueous solution - again by adding 28% aqueous ammonia - to a pH of about 8.5 to 9.0 set. The basic solution was extracted twice with 540 liters of benzene each time, and the benzene extracts were separated and combined. The combined extracts were dried with anhydrous sodium sulfate and concentrated in vacuo to a volume of about 15 liters. The benzene concentrate was in divided into three equal parts, which were separately chromatographed on 40 kg of deactivated aluminum oxide each. Each 40 kg batch of aluminum oxide was deactivated with 7.5 liters of water and 347 ecm of glacial acetic acid been.

Table I.

Table I above shows the results of the first, crude chromatographic separation. In the The first column shows the numbers of the fractions, and column 2 shows the eluents or mixtures used and in column 3 the alkaloids mainly obtained from the individual fractions are given. Vincaleukoblastin was obtained as crystalline sulfate from fractions 43 to 45. The received Mother liquors were mixed with fractions 47 to 52

ίο united. The combined fractions were under Stir slowly poured into a solution of 336 g of citric acid in 16 liters of water. The pH the solution was adjusted to about 4.4 with 28% aqueous ammonia and the solution twice with each 16 liters of benzene extracted. The benzene extracts were combined, dried over anhydrous sodium sulfate and evaporated to dryness in vacuo. The pH of the aqueous phase was then with 28% aqueous ammonia adjusted to about 7.0 and the resulting solution again twice with each 16 liters of benzene extracted. These benzene extracts were mixed with the residue obtained as above the benzene extraction at pH 4.4, the whole mixture was dried over anhydrous sodium sulfate dried and the solvent removed by evaporation in vacuo, leaving an alkaloid mixture remained as a residue.

40 g of the alkaloid residue was dissolved in 300 ecm of benzene, and the resulting solution was an 1335 g of aluminum oxide, which is partially deactivated by treatment with 80 ecm of water and 4 ecm of acetic acid was chromatographed.

The chromatogram was carried out using the same solvents or solvent mixtures, as indicated in Table I, developed. The results of the chromatography are as follows Table II indicated:

Table II

fraction Eluent alkaloid 1 benzene Catharanthin 2 benzene Vindolinine 3 to 19 benzene Ajmalicin 20 to 21 benzene Vindoline 34 to 42 Benzene- Leurosin chloroform (1: 1) 43 to 45 Benzene- ¹ vincaleukoblastine chloroform (1: 1) 46 chloroform Virosin 47 to 52 Chloroform- Amorphous substance, Methanol the Leurocristin and Leurosidin contains

fraction Eluent 1 benzene 2.3 Benzene-chloroform (3: 1) 4.5 Benzene-chloroform (3: 1) 6 to 8 Benzene-chloroform (3: 1) 9 to 14 Benzene-chloroform (3: 1) 15th Benzene-chloroform (3: 1) 16 Benzene-chloroform (3: 1) 17 to 29 Benzene-chloroform (3: 1) 30 to 32 Benzene-chloroform (3: 1) 33 Benzene chloroform (3: 1) 34 Benzene-chloroform (3: 1) 35 Benzene-chloroform (3: 1) 36 Benzene-chloroform (3: 1) 37 Benzene-chloroform (3: 1) 38. Benzene-Chloroform- ^: 1) 39 Benzene-chloroform (3: 1) 40 Benzene-chloroform (3: 1) 41 Benzene-chloroform (3: 1) 42 Benzene-chloroform (3: 1) ■ 43 Benzene-chloroform (3: 1) 44 chloroform 45 to 57 chloroform 58 Chloroform-methanol (19: 1)

Fractions 16 to 43 were found to contain leurosidin and leurocristin.

Each chromatography fraction containing leuosidin and / or leuurocristin was converted to in vacuo Evaporated to dryness and the remaining alkaloid material recrystallized from methanol. The leurosidiri - if present - was the first to go crystalline fraction from; and the Leurocristin was made by concentrating and cooling the mother liquors Leurösidin crystallization obtained;

Leurosidine melts with decomposition at about 208 to 211 ⁰ C and Leurbcristin with decomposition at about 218-220 ⁰ G;

Analysis:

Leurocristin *)
Leürösidiri

66; 73
67.08

7.06
7 ₅ 34

6.75
6.24

O molecular weight

19j44
18.81

824
826

*) Dried for minutes at 18Ö ° C.

Leurosidine has an optical rotation of [a)% = + 55.8 ° (fc = 1 in chloroform) - while the Leuro _r eristin an optical rotation of [a] f = J-17.0 ° (c = 1 in Ethylene dichloride). Both alkaloids possess two titratable basic groups if they are / titrated in 33 ° _o aqueous dimethylformamide. In the case of leucosidine, the ρΚ _α values of the basic groups are 5.0 and 8.8, while in the case of leuurocristin they are between 5.0 and 7.4. The USA.-Spectrum of Leurosidine shows absorption maxima at 214 and 265 ΐημ with shoulders at 286; 295 and 310 πιμ, while the spectrum of the Leurocristins maxima at 22Oj 255 and 296 πιμ with shoulders at 262 and 290 πιμ;

Table III Leurosidine (μ) Leurocristin (μ) 2.78 2.89 2.90 3.41; .. «« 3.41 5.74 5.74 5.93 6.17 6.24 6.64 6.67 6.85 6.86 6.97 6.97 7.28 7.35 7.50 7.53 7.74 7.74 8.15 8.15 8.56 8.74 8.75 8.84 8.87 9.04 9.07 9.15 9.28 9.60 9.70 9.89 9.91 10.44 10.45 10.73 10.86 10.86 11.13 11.23 11.99 ' 11.74 12.21 12.02

Table III above shows the IR absorption maxima of the alkaloids when measured in chloroform solution specified.

Depending on the geographical origin of the plant material, the harvest time and other unknowns Factors. be between 20 and 70 mg Leurocristin / kg. Sheet material received.

· '*>.'. ";:> ■! ■ V'Mb) ', production of Leurosidin-
and Leurocristin Sulphate

1 g of leurosidine was dissolved in 30 ecm acetone. A mixture was added to the acetone solution of the alkaloid from 5 ecm l% aqueous sulfuric acid and 42 ecm

Given water. The resulting mixture was evaporated to dryness in vacuo. The Leüros'iäinsulfat containing residue would be heated with heating dissolved in anhydrous ethanol. When the ethanol solution was cooled, practically quantitative

ao _ tative yield crystalline jLeurosidin-sülfat having a melting point of about 237 to receive up to 2 "45 ⁰ G. At 201 to 212 ° G leakage was observed from the solvent.

Analysis6: G = 59.84%, H = 6.75%, N - 5.23%, Ö = 24.08% _; Formula weight: 924.

The Leurocristinsuifat was completely analogous Made from the free base Leurocristin. The compound was produced with practically quantitative yield obtained and melted after recrystallization from ethanol at about 273 to 281 ° C. At 210 to • At 232 ° G there was an escape of solvent.

Analysis: C = 59.81%; H = 6.67%, N = 6.02%, O = 23.66% molecular weight: 922.

c) Production of Leurösidinuhd Leurocristin methojodid;

0.001 moles of Leurocristin became approximately 10 cc of benzene solved. 0.002 equivalents were added to the benzene solution Methyijodid added, and the resulting mixture was about 5 hours at room temperature.

proven, whereby crystalline Leurocristin methojodid parted. The mixture was cooled for about 2 hours to allow additional leurocristin methoiodide to crystallize. The precipitate was filtered off and recrystallized from ethanol. The thus prepared with a practically quantitative yield leurocristine-methoiodide melting at about 226-232 ⁰ C.

Analysis: J 13.08%.

The Leurosidine Methoiodide was made; by using leuosidine in place of Leurocristin was used. The Leurosidinmethojodid also obtained with practically quantitative yield melted out after recrystallization

Ethanol with decomposition at about 198-216 ⁰ C. At about 173 ° to 178 ° C leakage was observed from the solvent.

Analysis: J = 13.05%.

Claims (1)

1 2 Patent claim · together - of vinca plants, in which by preliminary investigations either one of the alkaloids Leuro- Process for the production of the alkaloids Leurocristin and Leurosidin or both have been determined, Sidin and Leurocristin, from their salts with acids, are dried and ground. The dried, and their quaternary ammonium salts, the ground material is mixed with dilute, watery wine characterized in that ceric acid is moistened and extracted with a water-miscible organic solvent with a portion of vinca plants that is not reduced in size with water. Darigen The more basic alkaloids in the will be moistened with an organic acid solution and then retained with a water-immiscible solid phase, while the weak organic solvent extracted, the extract io basic alkaloids, including the Leurocristin and the mixed with aqueous acid and the organic leukoblastin as well as the leurosin and vincaleukoblastin, Solvent removed, then the acidic aqueous extracted into the organic solvent. Phase after extraction with a not water The organic extract is separated and concentrated, The weakly basic alkaloid fraction is made from the weakly basic alkaloids extracted with an organic concentrate with water in aqueous acid, The immiscible solvent extracted, the The acidic, aqueous extract is separated and mixed with organic phase after drying and a water-immiscible organic solution extracted on deactivated aluminum oxide chromosome. The acidic, aqueous phase is graphed, the aluminum oxide first made basic with benzene and the leuurosidine-leucristin- or a weakly polar, then with a strong alkaloid fraction, then in a non-mixer with water back-extracted polar solvent or solvent-compatible organic solvent, eluted mixture containing the Leurosidin and / or Leuro- This extract is a chromatographic Trencristin containing fraction, optionally subjected to deactivated aluminum oxide after exposure, further purification by shaking out their acidic. In this process, Leurosin and and neutral solution with benzene, evaporation of the 25 the vincaleukoblastin with benzene or a weak Benzene solution to dryness and repeated chromopolar solvents, such as ether, eluted. Frakmatography on aluminum oxide, for drying, which contains the Leurosidin and the Leurocristin evaporates, the residue is fractionally crystallized and more strongly bound, is then with a and optionally then so obtained more polar solvent, such as chloroform or Leurosidin and / or Leurocristin. with acids in 30% methanol, eluted from the chromatography column. the salts or with alkyl or aralkyl halides This fraction is chromatographed again to or sulfates converted into the quaternary ammonium salts a further purification and enrichment of Leuro. sidin and Leurocristin. The raw alkali loids are produced by evaporation of the individual chromium 35 matography fractions obtained in vacuo to dryness. The amorphous residue obtained is in The invention relates to a process for a solvent such as methanol or ethanol, geGewinnung the alkaloids Leurosidin and Leuro- dissolves. Under these conditions it crystallizes cristin, of their salts with acids and quaternary leucosidine - if present - and can be filtered off Ammonium salts, characterized in that one turns 40. When concentrating and cooling the filtrate is Chopped up parts of Vinca plants' with a water- the Leurocristin - if present - as crystalline moistened solution of an organic acid and obtained substance, which is also filtered off. Election then With an organically immiscible with water, the alcoholic solutions that are caused by See solvent extracted, the extract with aqueous dissolving of the above-mentioned amorphous residue mixed acid and the organic solvent 45 can be obtained, removed directly with aqueous or ethanol, then the acidic aqueous phase can be treated according to Extrak- scher sulfuric acid, whereby tion with a water-immiscible solution Leurocristinsulfat. You can also use the Makes medium alkaline and the weakly basic filtrates, of which the Leurosidin and the Leuro-Alkaloids separated with a water-immiscible cristine as crystalline alkaloid bases Solvent extracted, the organic phase after 50, treat in a similar way to still Leurodem Drying and concentration to obtain deactivated alumi- cristine sulfate. In the above process, instead of next with benzene or a weakly polar, then tartaric acid, other organic acids such as vinegar with a more polar solvent or solvent acid or citric acid can also be used. Eluted in the medium mixture, the leucosidin and / or 55 different stages of the above process use-Leurocristin-containing fraction, optionally after bare, water-immiscible organic solutions further purification by shaking out their acidic medium are z. B. benzene, ether, ethyl acetate, chlorine and neutral solution with benzene, Eiridampfen the form, ethylene dichloride or methyl isobutyl ketone.
Benzene solution to dryness and repeated chromato- Leurocristin and Leurosidin are closely related digraphy on aluminum oxide, evaporated to dryness, 60 mere alkaloids, which have two indole ring systems per molecule of the residue fractionally crystallized and given. The free alkaloid bases are high-melting leucosidine and / or crystalline solid substances, if so obtained,
or Leurocristin with acids in the salts or with both bases form salts, alkyl or aralkyl halides or sulfates with suitable acids in the z. B. with sulfuric acid sulfates. Furthermore, both quaternary ammonium salts form converted. 65 bases by reaction with alkyl iodides, such as methyl The inventive method is z. B. following iodide, quaternary ammonium derivatives,
Carried out as follows: With Leurocristin and Leurosidin, leaves, stems or roots can be formed - separately or aside from the symptoms and a relief of the