DE1445619B - Process for the preparation of 7 amino cephalosporan acid esters - Google Patents

Description

1 2

The invention relates to a process for 19.3 mg extract no. 2 (chloroform-soluble lower ;:

Production of 7-amino-cephalosporanic acid esters, · Schlag), which according to thin-layer chromatogram (Ta-?

which is characterized in that one diester des Belle 1) and IR spectrum (bands at 5.61, 5.75 and

Cephalosporin C with low molecular weight alkyl alcohol 5.93 ηιμ) in addition to little starting material side pro-

get, p-nitrophenol, 2,4-dinitrophenol, 2,3,6-tri-5 contains.

nitrophenol, benzyl alcohol, p-nitrobenzyl alcohol, 4.3 mg extract No. 3 (from the proportion of buffer pH 3.3),

p-Methoxybenzyl alcohol, benzhydrol or p, p-di- according to thin-layer chromatogram (Table 1) little

containing methoxybenzhydrol with free amino group in Ben- starting material,

zol, nitromethane, dioxane or chlorinated carbon 26.8 mg (44% yield, based on cephalosporin

hydrogen in a concentration of about 0.2 ion overall yield: 25.4%) 7-amino-cephalospo-

up to 1% at room temperature, if necessary in rans acid benzyl ester in extract no. 4 (from 2% Phos-j!

Presence of acetic acid or aqueous acetic acid, 'phosphoric acid component). IR spectrum in methylene-!

if desired together with pyridine, rid for a few days: bands at 5.62 and 5.75 ηιμ. After thin-film!

lets stand and the 7-amino-cephalosporanic acid chromatogram (table) uniformly,

ester isolated. . 15 The product can be dissolved in glacial acetic acid in the presence of j

The esters can be converted to 7-amino-cephalospo-j |

genolysis can be hydrogenated into the free 7-amino-cephalosporanic acid. ||

be transferred. In this regard, especially the cephalosporin used as a starting material!

The benzhydryl ester is favorable, which can be prepared by means of trifluoro-C-dibenzyl ester as follows: t

acetic acid and anisole can be split, even the 20 9.43 g cephalosporin C (20 mmol) are in 250 ml!

p-Methoxybenzyl ester can be hydrolyzed in this way; 1 n-sodium bicarbonate dissolved, with 3.62 ml tertt-Bu-

p-nitrobenzene and tyloxycarbonylazide (26 mmol), dissolved in 150 ml of di-

Benzyl ester, which is added by means of catalytically excited water oxane, and the mixture is stirred at 40 ° for 5 hours. The other

be split into the fabric. closing in a vacuum at 0.5 mm Hg and 30 °

The diester 25 used as starting material, about 150 ml of concentrated solution, is diluted with of the cephalosporin C are produced, for example, 200 ml of water and extracted several times, by removing the free amino group of the cephalo ethyl acetate. The aqueous, saturated with table salt; sporins C by an amino protecting group, e.g. B. a phase is then generated at pH 2.0 in the cold; Acyl group such as trifluoroacetyl, tosyl, carbobenzoxy scooped out with ethyl acetate. Drying the! | or especially tert-butyloxycarbonyl, trityl or extract washed with saturated sodium chloride solution o-Nitrophenylsulfenyl, blocked, then the carboxylates over sodium sulphate and evaporation in vacuo !! groups esterified and the amino protective group gives off 10.7 g of amorphous, colorless N-tert-butyloxy-l splits. carbonyl-cephalosporin C (91.6% yield). ρ

As a by-product of the aminolysis, the ε-Ami-Rf value in the paper chromatogram in the system IjI

noadipic acid lactam monoester formed. This leaves 35 [n-butanol-acetic acid (10: 1) saturated with water] :! ¹

prove to be a neutral substance of the desired ba- 0.80; in system 2 (water-saturated n-butanol + 1% E

Easily separate sischen reaction product, z. B. by glacial acetic acid): 0.51. Bioautographic evidence with; ¹

Extraction with acidic solvents, chromato- staph. aureus. ■

graphy or countercurrent distribution. To a solution of 3.2 g (6.15 mmol) of N-tert.-!

In the following examples the temperatures are 40 butyloxycarbonyl-cephalosporin C in 64 ml abs .;

Doors indicated in degrees Celsius. Ethylene glycol dimethyl ether is left in the course of

. . 5 minutes 185 ml of essential phenyldiazomethane solution,

Example 1 _solution) containing 16 mmol of reagent, add dropwise and !;

100 mg of cephalospirin C-dibenzyl ester are stirred for 40 minutes at 25 ° in the dark. Then ji

Dissolve 20 ml of the purest methylene chloride and remove excess reagent by adding 2 ml of ice

angle left at 22 °. The intravinegar which runs off is destroyed, taking 30 minutes in the dark

Molecular reaction can easily be stirred further in the IR spectrum. The reaction solution is in vacuo;

Track trum (1 mm cell): evaporated, the residue in 120 ml of methylene-ij

The amide band of the starting material at 5.93 ηιμ rid added, twice with 60 ml of ice-cold n-Na- '

becomes steadily weaker over the course of several days, with 50 trium bicarbonate solution and then twice with 50 ml!

A new band of waxy 10% sodium chloride solution was washed next to it at 6.00 ΐημ. The water!

sender intensity develops (lactam bands of the split phase are sequentially with twice

Product o-carbobenzoxy-o-amino-valerolactam). 40 ml of methylene chloride extracted. The one with Na-;

After 20 days is concentrated by evaporation in a vacuum, taken triumsulfat and dried in vacuo eingedampf- ¹

in chloroform-ether (1: 3) and shaken after 55 th methylene chloride solutions give 6.17 g of amorphous,

each other with 0.1 molar phosphate buffer pH 3.3 and partly oily residue, which at the twenty-

2% aqueous phosphoric acid. In the process, times the amount by weight of silica gel separates out (deactivated with

a precipitate forms which is dissolved in chloroform. 5 weight percent water) is chromatographed.

The two aqueous extracts are extracted with a methylene chloride-acetone mixture (95: 5)

pH 8.5 separately with ethyl acetate. The over magnesium 60 2.73 g (yield 63%) of N-tert-butyloxycarbonyl-

Sulphate-dried organic phases give the cephalosporin C-dibenzyl ester elute, which from

Evaporation of the following residues: acetone — ether — petroleum ether (b.p. 50 to 70 °).

25.9 mg extract no. 1 [chloroform-ether (1: 3) - is installed. F. 88.5 to 91 °.

Proportion], according to the IR spectrum (bands at 5.75 and IR absorption spectrum in Nujol: bands below

6.00 ιτιμ in methylene chloride) from o-carbobenzoxy 65 others at 2.98 μ, 5.61 μ, 5.78 μ (with shoulder at

(Consists of 5-amino-valerolactam. Im thin-layer chromium 5.70 μ), 5.91 μ, 6.05 μ, 6.57 μ, 6.83 μ, 7.24 μ, 7.69 μ _{; 1}

matogram no spots with iodine starch and nin- 8.02 μ, 8.17 μ, 8.55 μ, 8.99 μ, 9.32 μ, 9.53 μ, 9.75 μ ,!

hydrin-collidine. 10.37 µ, 11.0 µ, 11.53 µ, 13.37 µ and 14.40 µ. UV.-Ab-i

Sorption spectrum in fine spirits: X _max 265 ηιμ (ε = 8200).

Rf value in the thin-layer chromatogram on silica gel (benzene-acetone system 8: 2): 0.46.

Reaction according to R e i η d e 1 and Hoppe (Ber., 87, P. 1103 [1954]): yellow with a purple halo.

Reaction with iodine-starch-acetic acid (R. Thomas, Nature, 191, p. 1161 [1961]; P. H. A. S η e a t h and Z. F. Collins, Biochem. J., 79, p. 512 [1961]): positive. ίο

140 mg N - tert. - Butyloxycarbonyl - cephalosporin C-dibenzyl ester (0.2 mmol) are left with 2 ml of trifluoroacetic acid at 25 ° for 5 minutes. The trifluoroacetic acid is then removed in vacuo. The oily residue is taken up in 20 ml of methylene chloride, washed twice with 10 ml of ice-cold n-sodium carbonate solution, then washed twice with 10 ml of 10 ° / _o sodium chloride solution. The aqueous phases are extracted twice more with 5 ml of methylene chloride. The methylene chloride solutions dried with sodium sulfate give 120 mg (quantitative yield) of cephalosporin C-dibenzyl ester on evaporation in vacuo, which has the following Rf values in the thin-layer chromatogram:

Systen Rf

Dioxane-water 0.77

Benzene-acetone (1: 1) 0.29

Chloroform-methanol (9: 1) 0.56

tert-benzyl alcohol — i-propanol—

Water (100: 40: 55) 0.62

n-butanol — acetic acid — water

(100: 100: saturated) 0.57;

Reaction with iodine-starch-acetic acid ... positive reaction according to R e i η d e 1 and

Hop yellow

Reaction with ninhydrin collidine, cherry red

Rf values in the thin-layer chromatogram on silica gel

Starting material extract no.2

Extract No. 3

Extract No. 4

system

n-butanol-acetic acid 10: 1 saturated with water

system
Benzene-acetone 6: 4

indicator

Ninhydrin collidine
indicator

Iodine starch

0.60

0.16

red-violet

positive (0.61)
0.80 to 0.95

0.0 to 0.22

flesh colored

positive

0.61

(0.47)

0.36

0.15
0.0

red-violet
positive

0.67

0.62

dark yellow positive

The reagent is prepared according to R. T hο m a s, Nature, 191, p. 1161 (1961). Weak spots are indicated by brackets.

Example 2

2.61 g of cephalosporin C-dibenzyl ester are dissolved in 520 ml of methylene chloride, with 5.2 ml of one Equimolecular mixture of pyridine and glacial acetic acid and left to stand for 4 days in the dark at 22 °. The work-up carried out as in Example 1 gives the following extracts:

945 mg extract No. 1 [chloroform-ether (1: 3) portion], consisting of o-carbobenzoxy-o-amino-valerolactam.

680 mg extract No. 2 (chloroform-soluble precipitate).

j 136 mg extract no. 3 (from the portion of buffer pH 3.3), [consisting of cephalosporin C-dibenzyl ester.
j 604 mg (38 ° / ₀ of theory) of 7-amino uniform; cephalosporanic acid benzyl ester in extract # 4 (2% IPhosphorsäureanteil) related to cephalosporin C. Yield:.! 21.9%.

All products are characterized as in Example 1 by means of an IR absorption spectrum and a thin-layer chromatogram (see table).

Example 3

100 mg of cephalosporin C-dibenzyl ester are dissolved in 20 ml of methylene chloride with 0.02 ml of an equimolar mixture of pyridine and acetic acid and treated with 0.02 ml of water and allowed to stand for I ¹ I ₂ days in the dark at 22 °. Working up as in Example 1 gives the following extracts:

46.2 mg extract No. 1 15.5 mg extract No. 2 9.5 mg extract No. 3

32.1 mg extract No. 4

Consists of a uniform 7-amino-cephalosporanic acid benzyl ester. Yield: 53%, based on cephalosporins: 30.5%.

Example 4

100 mg of cephalosporin C-dibenzyl ester, dissolved in 20 ml of methylene chloride, are mixed with 0.02 ml of 2η-acetic acid and left to _{stand for 7 V for 2} days at 22 °. Working up as in Example 1 delivers

42.8 mg extract no.1
20.0 mg extract no.2
15.2 mg extract No. 3 and

30.9 mg (yield: 51%, based on cephalosporin C

based on: 29.4%) 7-Amino-cephalosporanic acid benzyl ester in extract No. 4

60

Example 5

In the same way as described in Example 1 to 4, cephalosporin C-dimethyl ester, cephalo-

Claims (1)

5 6 sporin C-diethyl ester and cephalosporin C-di-n-butyl ester, which is yellowish from chloroform-ether (1: 4) ester treated. 7-Aminocephalospo crystals are obtained in this way. After recrystallization, the melts rans acid methyl ester, ethyl ester and n-butyl ester. Ester at 104 to 106 °. According to thin-layer chromatography These have the following Rf values in the paper chromatogram on silica gel is the compound uniform, grams, system I (n-butanol-acetic acid 10: 1, ge 5 The Rf value in the chloroform-methanol system saturated with water): methyl ester: Rf I = 0.13; Ethyl- (95: 5) is 0.79, in cyclohexane-ethyl acetate (1: 1) 0.19. ester: Rf I = 0.15; n-butyl ester: Rf I = 0.16; yellow - yellow spots are obtained with sodium hydroxide solution, colorless brown coloration with ninhydrin collidine or bioauto- stains with iodine-starch reagent. The tert-butyloxy graphic proof with staph. aureus after spraying carbonyl group is, as described in Example 1, hen with 1 molar pyridine in acetone-water (1: 1), split off with trifluoroacetic acid,
and L ° / _o sodium phenylacetyl chloride in acetone. The The UV spectrum shows a maximum at 263 πιμ B eis ρ ie 1 7
(ε = 8000). The diesters used as starting materials will be 12.6 g of cephalosporin C-di-benzhydryl ester prepared as follows: 15 dissolved in 2 liters of absolute methylene chloride, with 2 ml 1 g of N-tert-butyloxycarbonyl-cephalosporin C is mixed with 2N aqueous acetic acid and dissolved in 20 ml of methanol for 8 days, cooled to 0 ° and left to stand below 22 ° in the dark. It is evaporated with 15 ml of an ethereal, 4 ° / ₀ igen kuum in va swirling and the residue is taken in a Ge-diazomethane solution (or diazoethane or diazo mix of 5 parts of toluene, 2 parts of ethyl acetate, 3 tibutane solution) . After about 5 seconds, 20 liters of alcohol and 3 parts of 2 normal aqueous saltman stop the reaction by adding 3 ml of glacial acetic acid. acid on. After complete dissolution with vigorous The concentrated in vacuo, then shaking in 200 ml of vinegar, the phases are separated and the lower ester taken up mixture is washed with 1 η sodium phase with two other upper phases (5 parts of toluene bicarbonate and saturated saline solution, and 2 parts of ethyl acetate) shaken out. The three over sodium sulfate dried and phase in vacuum are evaporated four more times with alcohol-2N salt. This gives the N-tert-butyloxycarbo- acid (1: 1) after-extracted. The product-containing Unternylcephalosporin C-dimethyl (or -diäthyl- or phases are combined, washed with 50 ° / ₀ aqueous Trika- dibutyl) as an amorphous colorless residue. Liumphosphatlösung adjusted to pH 6 and in vacuo The Rf values of the compounds in System I are freed from alcohol. Then one places with the trika- [n-butanol-acetic acid (10: 1), saturated with water] 30 calcium phosphate solution to pH 8.0 and extracted three times or system III (n-butanol, saturated with water + 1% with ethyl acetate. Ex-glacial acetic acid dried over sodium sulfate) are: dimethyl ester: Rf I = 0.89; RfIII trakt releases the 7-amino-cephalo- = 0.84; Diethyl ester: Rf I = 0.91; Dibutyl ester: sporic acid benzhydryl ester, which from ether in to RfIII = 0.70 (bioautographic evidence with needles combined with drusen crystallized; F. 122 bis Staph. aureus). 35 124 °; it shows in the thin-layer chromatogram The tert-butyloxycarbonyl group is hydrolyzed using tri-silica gel in the system n-butanol-glacial acetic acid (10: 1), as described in example 1, saturated with water an Rf value of 0.64 (dirty cracks, yellow spot with ninhydrin collidine). The ester can be converted into the free 7-amino- Example 6 40 cephalosporanic acid are converted: one dissolves 6.8 g = 1 part ester in 1 part anisole and mixed with When using Cephalosporin C-di-p-nitrophenylesier, 5 parts of trifluoroacetic acid. You then steam as described in Example 1 to 4, hydrolyzed, so immediately at 0.2 mm of mercury within 20 Mi- 7-amino-cephalosporanic acid-p-nitrophenous is obtained, taken up in 30 ml of ethyl acetate and the nyl ester is poured. The compound migrating at the paper 45 solution simultaneously with about 13 ml of 50 ° / _o aqueous electrophoresis (pH 4.5; 2000VoIt, IV ₂ hours) tripotassium phosphate solution with stirring to 20 ml 8.2 cm towards the cathode. 3 ° / _o aqueous dipotassium hydrogen phosphate solution. The cephalosporin used as the starting material. The two inflows are to be dimensioned so that C-di-p-nitrophenyl ester can be prepared as follows, the pH fluctuates between 6 and 8 and in the end to 7 become: 50 stops. The separated aqueous phase is 6.47 g tert. - Butyloxycarbonyl - cephalosporin C, twice more with 10 ml of ethyl acetate each time and the three vinegar-4.24 g of p-nitrophenol and 8.56 g of dicyclohexylcarbo ester phases are dissolved twice with 5 ml of 3% aqueous diimide each time in 300 ml of acetonitrile and washed in riger dipotassium hydrogen phosphate solution. In the dark under a nitrogen atmosphere for 17 hours at Man discard the organic phases, combine the 22 ° left to stand. Then the precipitated aqueous solution is filtered off and dicyclohexylurea (4.38 g) is removed with about 4 ml of concentrated salts and evaporated to pH 3.5 in the acid. The vacuum after 15 hours of standing at 0 ". By triturating the deposited product three times, the product is filtered off, the water is washed with a little ice vapor residue with 100 ml of petroleum ether each time and dried. 3.90 g of excess dicyclohexylcarbodiimide is obtained (yield 92%) ) pure 7-amino-cephalosporanic acid (2.94 g) and filtered off from the insoluble product
The material was then dissolved in acetone, which deposited more dicyclohexylurea (0.19 g). Claim:
Evaporation of the filtrate, take up in chloroform
and exhaustive shaking with 0.5 molar Phos. Process for the production of 7-amino-cephalo-] phate buffer pH 7.0 gives after washing (saturated 65 sporic acid esters, characterized by saline solution), drying (sodium sulfate) and Einnet that one diester of cephalosporin C with vapors of the organic phase 0.92 g of crude tert-low molecular weight alkyl alcohols, p-nitrophe-i-butyloxycarbonyl-cephalosporin C-di-p-nitrophenyl nol, 2,4-dinitrophenol, 2,4,6 -Trinitrophenol, Ben- 7 8 zyl alcohol, p-nitrobenzyl alcohol, p-methoxyben- at room temperature, if necessary in counter- zyl alcohol, benzhydrol or ρ, ρ-dimethoxybenz- wart of acetic acid or aqueous acetic acid, hydrol with free amino group in benzene, nitro- if desired together with pyridine, some methane, dioxane or chlorinated hydrocarbon for days and the 7-amino-cephalosporan- substances isolated in a concentration of about 0.2 to 1 ° / ₀ 5 acid esters.