KR830001415B1 - Process for preparing cephalosporin derivatives - Google Patents

Abstract

The solid, storage stable cephalosporin salts I(X=Cl,Br; n=0-6) were prepd. by treating H2NCH2CH2NMe2 with CS2, conveting Me2NCH2CH2NHCS2H to its Me ester, and cyclizing the latter with NaN3 to the tetrazolethiol II. Reaction of the acetoxymethylcephem III with II, followed by treatment with 12N HCl gave 94% pure I(X=Cl, n=1). The MIC of stepilococus aureus FDA 209p and toxicity of mice were 0.39mcg/ml and CD 50>=20 g/kg, resp.

Description

Method for preparing cephalosporin derivative

The present invention relates to a method for producing a solid Sepharoseporin derivative represented by the following general formula (I).

(Wherein X represents chlorine or cancellation and n represents a number from 0 to 6)

It is well known that the compound represented by the following general formula (II) has excellent antibacterial ability in the past, but (West German Publication No. 2461478) Compound (II) has two amino thiazoline groups and dimethylamino group in the molecule. Since it has a basic group and one carboxyl group, there exists a fault which is extremely water solubility, difficult to refine | purify, and hard to say that it is sufficiently stable in storage.

As a result of repeated studies to improve the advantages, the solid cephalosporin derivative (I) is reacted with a compound (II) and an acid represented by two or more moles of formula HX (wherein X is the same as above), if necessary, in the presence of water. The fact that the obtained compound (I) is extremely stable in preservation and that the compound (I) can be obtained as a crystal can almost completely remove impurities which were difficult to remove by the conventional purification method. The present invention was completed based on this finding. That is, the present invention,

(1) Solid Cephalosporin Derivatives (I)

(2) A method for producing a solid cephalosporin derivative (I) characterized by reacting a compound (II) with an acid represented by the formula HX of 2 moles or more in the presence of water if necessary.

n

4인 것은 안정성, 순도에 있어서 가장 바람직하다.In the general formula (I), x represents chlorine or cancellation. The compound of general formula (I) contains water molecules of 6 or less. Among them, six kinds of hydrates, particularly monohydrate to hexahydrate, exist, which may contain 1 mole or less of bonds depending on the production conditions, and any of them are collected as crystals. Anhydrides are usually amorphous powders, but may have a bonding water of one or less. All of these belong to the scope of the present invention. 0.1 as the range of n

n

4 is the most preferable in stability and purity.

The present invention is carried out by reacting a raw material compound (II) with an acid represented by two or more moles of formula HX in the presence of water if necessary. The starting compound (II) is, for example, a solution obtained by removing most of the impurities from the synthesis reaction mixture liquid or the reaction mixture of the starting compound (II), a powder obtained by mixing the solution with a poorly soluble solvent, or the solution. Powder obtained by concentrated drying or lyophilization is used. The starting compound (II) can be used in the present reaction as alkali metal salts such as zwitter ions and sodium.

As the acid represented by the formula HX, hydrochloric acid and hydrobromic acid are used. When the amount of acid used is less than 2 moles with respect to 1 mole of raw material (II), it is quite difficult to obtain uniform crystals. Since decomposition of II) takes place, it is usually 2-10 moles, preferably 2-6 moles. The reaction is usually carried out in the presence of water or an organic solvent or a mixed solvent thereof so as to proceed advantageously in terms of operability, yield, purification effect and the like. As the organic solvent used in the reaction, for example, ethanol, n-propanol, isopropanol, butanol, isobutanol, acetone, methyl ethyl ketone, acetonitrile, tetrahydrofuran, dioxane and the like (I) are poorly soluble and In addition to solvents that can be dissolved in water, these solvents and the target (I) are readily soluble and can also be dissolved in water, for example methanol, dimethyl sulfoxide, formamide, N, N-dimethylformamide, N, Mixed solvents with solvents such as N-dimethylacetamide, methyl cellulose solution, or these solvents and the target (I) are poorly soluble and difficult to dissolve in water, for example, ethyl acetate, ether, dichloro A mixed solvent with a solvent such as methane or chloroform is used. Among them, for example, a mixed solvent of water and acetone, ethanol, n-propanol, i-propanol, methyl ethyl ketone or tetrahydrofuran is very suitable. The reaction temperature is usually carried out in the temperature range of -10-40 ° C, preferably 0-30 ° C. Below -10 ℃, the crystal growth rate is slow and cooling is required during operation, and above 40 ℃, decomposition reaction of raw material (II) occurs, which is disadvantageous. The reaction time varies depending on the purity of the raw material (II) used and the type of impurities contained. However, when the raw material (II) having a purity of 80% or more purified by column chromatography using a conventional purification method such as an adsorbent or the like is used. It is 30 minutes-24 hours, Preferably it is 30 minutes-4 hours. Semi-product (I) is isolated by filtration or centrifugation, a freezing precursor, or the like. When the organic solvent is contained in the isolated compound (I), the organic solvent is removed and dried as necessary.

For a preferred embodiment of the present invention,

(A) Compound (I) is dissolved in the solvent, a calculated amount of HX (X is as described above) is added, and the reaction product is lyophilized. When this is dried again in the presence of a dehydrating agent such as silica gel and phosphoric anhydride under reduced pressure, anhydride of compound (I) is obtained.

(B) The compound (II) is dissolved in water or a mixture of water and the hydrophilic organic solvent, HX is added, and the organic solvent is less likely to dissolve in the solution obtained after the reaction is completed. Slowly add until starting, then leave it for an hour or several hours as it is, and then slowly add an organic solvent, which is hard to dissolve again, until new crystals do not precipitate. The precipitated crystals are collected by means of filtration and centrifugation.

The crystal thus obtained usually has a small amount of an organic solvent although it is usually 3-4 hydrate. Although the degree of hydration differs according to conditions, such as a crystallization temperature and a solvent, it is a 3-10 hydrate normally and may contain a small amount of organic solvents sometimes. Normally, the organic solvent is removed when dried under reduced pressure, but depending on the solvent, it may not be removed under reduced pressure drying. To remove such an organic solvent, this crystal is brought into contact with water vapor to about 1 mole of anhydride of compound (I). Absorb water up to 5-10 moles. When the water-containing one is kept under reduced pressure (about 0-20 mmHg), the water is removed, and crystals of hexahydrate, pentahydrate, tetrahydrate and trihydrate are obtained in sequence.

When the trihydrate crystal is left again under the above reduced pressure in the presence of a dehydrating agent such as phosphoric anhydride, dihydrate crystal, monohydrate crystal, and anhydride are sequentially obtained. In the course of this change, less than one mole of water removed from the molecular compound may adhere to the molecular compound, or may be a mixture of different hydrates.

(C) When the above-mentioned lyophilized product is brought into contact with water vapor, water vapor is absorbed, crystallized as a hydrate and dried, thereby obtaining tetrahydrate, trihydrate, dihydrate, monohydrate, and anhydride sequentially.

Thus obtained crystal of target (I) was confirmed to have 2 moles of HX per 1 mole of raw material (II) by elemental analysis, titration, etc., and according to the microscope, polarizing microscope, X-ray diffraction pattern, etc. Besides being confirmed to be crystalline, a narrow and sharp peak different from the amorphous powder in the infrared absorption spectrum can be seen. In addition, if the moisture contained in the target (I) is reduced to 1 mol or less by heating to dryness, the crystallinity may already be lost in the X-ray diffraction pattern, but in this case, the interference color (crystallinity) may be Is observed.

The objective (I) of the present invention is a desired pH or ion type using an amorphous powder obtained by crystallization or drying as it is, or in combination with a non-toxic alkali or alkali salt such as sodium bicarbonate, sodium carbonate, trisodium phosphate, or the like. It may be adjusted to ionic strength and used as a preparation for injection or oral use.

For example, a solution obtained by dissolving a target (I) in an aqueous solution of sodium carbonate (hereinafter referred to as C solution) can be used as an external disinfectant such as, for example, a disinfectant such as a surgical instrument, a hospital room, and a drinking water. For example, for warm-blooded animals such as humans, mice, large mice, and dogs, gram-positive groups (e.g., Staphylococcus aureus, etc.) and gram-negative groups (e.g., Escherichia coli, Klebsiella pneumoniae) , Proteus vulgaris, Proteus molani, etc.) is administered by intramuscular injection or intravenous injection. When used as an external disinfectant for surgical instruments, 100 r / ml aqueous solution can be prepared in the form of anhydrous target (I) and dispersed in surgical instruments, and urinary tract caused by human or rat Escherichia coli infection. For infectious diseases, C solution is administered intravenously by dividing about 5-50mg / kg three times a day in terms of anhydrous target (I).

Reference Example 1

Antimicrobial Activity (MIC) and Toxicity of Target (I) in Anhydrous State

(1) Antibacterial Spectrum (Agar Dilution Method)

Staphylococcus aureus FDA 209p 0.39mcg / ml

Staphylococcus aureus 1840 0.78mcg / ml

Escherichia coli-NIHT JC-2 0.2mcg / ml

Escherichia coli 0-111 0.05mcg / ml

Escherichia coli T-7 1.56mcg / ml

Klebsiella pneumoniae DT 0.1mcg / ml

Proteus vulgaris IFO 3988 1.56mcg / ml

Proteus molgari IFO 3848 0.39mcg / ml

(2) Acute toxicity (rat, intraperitoneal administration)

20g/kgCD50

20g / kg

However, acute toxicity was tested with a 1: 1 (mole) mixture of the target (I) and sodium carbonate. The target compound (I) of the present invention can also take tautomers by tautomerization as shown in the following formula.

The presence of this type of compound has been studied in various ways, and in the literature, in all cases, the tooth deodorant foam is used [Acta. Acta Crystallographica Vol. 27, 326p (1971), and in other cases, toothbrushes and foams are taken [Chemistry and Industry, 1966, 1634p. According to various measurement results, it is thought that dental zorine and foam are stabilized by the contribution of hydrogen bonds, which are represented by the following formulas, to take advantage of dental zorine and foam.

However, this kind of equilibrium may be changed fluidly depending on the defrosting situation of the target compound (I), for example, the liquidity of the solvent, the polarity of the solvent, and the temperature.

Therefore, the naming of the target compound (I) may be any form, but in the present invention, the target compound (I) is teemed. Name it as a form.

Example 1

를 첨가냉각하고 2황화탄소 400g과 에테르 4.0

의 혼합액을 18-23℃로 한시간 걸려서 적가하고, 또 동온도로 한시간 교반하여 석출되는 2-(N,N-디메틸아미노)에틸아민카르보디치오산의 결정을 여취(濾取)하여 건조시킨다.(1) ether 2.4 to 400 g of 2- (N, N-dimethylamino) ethylamine

Cool and add 400 g of carbon disulfide and ether 4.0

The mixed solution of was added dropwise at 18-23 ° C. over 1 hour, and the crystal of 2- (N, N-dimethylamino) ethylaminecarbodiic acid precipitated by stirring at the same temperature for 1 hour was filtered and dried.

Yield 695 g, yield 93.3%, melting point 156-157 °

을 첨가하여 교반하면서 8-13℃로 IN-KOH 4.32

을 30-40분간 적가하고, 또 0-5℃로 요도드화메틸 668g, 아세톤 6.68

의 혼합액을 30-40분간 적가하고, 동온에서 30분간 교반하였다. 감압하에 아세톤을 유거(溜去) 한후 수층을 초산에틸 3

, 2

로 추출하고, 초산에틸층을 포화식염수 2

로 세착하고, 망초(芒硝)로 건조후 농축시켜서 얻은 결정을 N-헥산 500ml를 첨가하여 재결정하고, S-메틸-[2-(N,N-디메틸아미노)] 에틸아민카르보디치오에이트 575g,수율 75.5%, 융점 61-62℃를 얻었다. 이 결정 520g에 에탄올 1.05

, 아자이드화나트륨(NaN₃)190g, 순수한 물 2.1

을 첨가하고, 3시간 가열, 환열시켰다. S-메틸-[2-N,N-디메틸아미노)] 에틸아민카르보디치에이트의 결정 52g의 에탄올 100ml 용액을 더 첨가하여 한시간 환류시킨다. 20℃로 냉각하고 순수한 물 2,0

을 첨가하여 질소기류중 농염산으로 pH 2-2.5로 하였다. 감압으로 에탄올을 유거하고 암버얼라이트(Amberlite) IR-120(H형)[토마 &허스사제]에 통과시켜서 순수한 물로 산성이 소실할 때까지 세척하였다. 5% 암모니아수의 용출하는 분획을 농축하고 정출한 1-[2-(N,N-디메틸아미노)에틸]-5-메르캅토-1H-테트라졸 350g을 얻었다. 수율 69.3%, 융점 218-219℃. NMR(D₂O 중등몰의 NaHCO₃를 첨가하여 측정, τ) : 5.33(2H,t,

Water on this crystal 4.4

IN-KOH 4.32 at 8-13 ° C. with stirring

Was added dropwise for 30-40 minutes, and 668 g of methyl iodide and acetone 6.68 at 0-5 ° C.

The mixture was added dropwise for 30-40 minutes, and stirred at the same temperature for 30 minutes. After distilling off acetone under reduced pressure, the aqueous layer was diluted with ethyl acetate 3

, 2

Extracted with ethyl acetate and saturated saline 2

The resulting crystals were washed with brine, dried over agar, concentrated and recrystallized by addition of 500 ml of N-hexane, 575 g of S-methyl- [2- (N, N-dimethylamino)] ethylaminecarbodizioate, Yield 75.5% and the melting point 61-62 degreeC were obtained. Ethanol 1.05 in 520g of this crystal

, Sodium azide (NaN ₃ ) 190 g, pure water 2.1

Was added and heated and refluxed for 3 hours. S-Methyl- [2-N, N-dimethylamino)] Ethylamine Carbodiciate A further 52 ml of ethanol 100g solution was added to reflux for one hour. Cool to 20 ℃ and pure water 2,0

Was added to pH 2-2.5 with concentrated hydrochloric acid in a nitrogen stream. The ethanol was distilled off under reduced pressure and passed through Amberlite IR-120 (type H) (manufactured by Thomas & Hurs) and washed with pure water until acid disappeared. An eluted fraction of 5% aqueous ammonia was concentrated to obtain 350 g of 1- [2- (N, N-dimethylamino) ethyl] -5-mercapto-1H-tetrazole. Yield 69.3%, melting point 218-219 ° C. NMR (measured by addition of NaHCO ₃ in medium mole of D ₂ O, τ): 5.33 (2H, t,

에 첨가, 교반하면서 (1)에서 얻은 1-[2-(N,N-디메틸아미노)에틸]-5-메르캅토-1H-테트라졸 86.5g 및 탄산수소나트륨 42g을 첨가하였다. 65℃에서 75분간 교반시킨후 10℃로 냉각시켰다. 5N-HCI 250ml을 첨가 pH2.0으로 만들어 생성된 불용물을 여취하고 수세하였다.(2) 206 g of 7β- [2- (2-amino-4-thiazolin-4-yl) acetamide] -3-acetyloxymethyl-3-cepem-4-carboxylic acid

To the mixture was added 86.5 g of 1- [2- (N, N-dimethylamino) ethyl] -5-mercapto-1H-tetrazole obtained in (1) and 42 g of sodium hydrogencarbonate while stirring. After stirring for 75 minutes at 65 ℃ cooled to 10 ℃. 250 ml of 5N-HCI was added to pH 2.0 to filter the resulting insolubles and wash.

로 충진된 컬럼에 통과시켰다. 물60

로 세정하고 20%메탄올을 이어서 40%메탄올로 용출시키고, 목적물을 함유한 분획 11

를 5

까지 농축하고, 염기성 알루미나(300mesh)300g으로 충진된 컬럼 및 압버얼라이트 IR-120(H형)100ml로 충진된 컬럼에 통과시키고 컬럼을 수세했다. 용출액, 세액을 합해서 농축하여 2

로 하였다. 5℃로 냉각하고 활성탄 5g을 첨가 5분간 교반후 여과하여 활성탄을 제거하였다. 여액을 동결건조하고 7β-[2-(2-이미노-4-치아조린-4-일)아세트아미드]-3-[1-[2-N,N-디메틸아미노)에틸]-1H-테트라졸-5-일]치오메틸-3-세펨-4-카르본산 51.2g을 얻었다.Combine filtrate and washings with sodium hydrogencarbonate to pH5.2 Amberlite XAD-II (100-200mesh) 10

It was passed through a column packed with. Water60

Washed with 20% methanol and then eluted with 40% methanol, fraction 11 containing the desired product.

5

The mixture was concentrated to and passed through a column filled with 300 g of basic alumina (300 mesh) and a column filled with 100 ml of Abverolite IR-120 (type H), and the column was washed with water. Concentrate the eluate and the tax solution together 2

It was set as. After cooling to 5 ° C., 5 g of activated carbon was added and stirred for 5 minutes to remove activated carbon. The filtrate was lyophilized and 7β- [2- (2-imino-4-thiazolin-4-yl) acetamide] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetra 51.2 g of sol-5-yl] thiomethyl-3-cepem-4-carboxylic acid was obtained.

NMR (at 60 MHz, D ₂ 0) τ: 3.45 (S, 5-H) chia 4.35 (d, 7-H) 4.88 (d, 6-H) 5.0-6.7 (m, 5 × CH ₂ ) 6.95 ( S, 2 × CH ₃ ).

로 용해하고, 아세톤 1.0

이어서 12N-HCI 150ml을 첨가하였다.(3) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamide] -3- [1- [2-N, N-dimethylamino) ethyl obtained by the method of (2) ] -1H-tetrazol-5-yl] thioacetic-3-cefe-4-carboxylic acid isocratic dried product (263 g (purity measured by high-performance liquid chromatography of this product was 93% in terms of anhydrous) of water 1.0)

Dissolved in acetone 1.0

Then 150 ml of 12N-HCI was added.

를 첨가하고 10℃에서 한시간 교반한 후, 다시 2

의 아세톤을 첨가한시간 교반하여 정출된 결정을 여취하고, 아세톤 500ml로 4회 세정한후 건조시켜서, 상기 카르본산 2염 산염결정 262g을 얻었다. (순도 환산수율 94%).Add 20 g of activated carbon to the solution, stir at 5 ° C. for 10 minutes, and filter out the activated carbon.

Added and stirred at 10 ° C. for one hour, followed by 2

The mixture was stirred for a while to add acetone, and the crystals crystallized were filtered off, washed four times with 500 ml of acetone, and dried to obtain 262 g of the carboxylic acid dihydrochloride crystal. (94% purity conversion).

IR (KBr) cm ^-1 : 1770 (β-lactam)

Elemental analysis _{(C 18 H 23 N 9 O} 4 S 3. 2HCI. H 2 O)

Found C: 34.87 H: 4.52 N: 20.01 S: 15.33 CI: 11.24

Calculated C: 35.06 H: 4.47 N: 20.45 S: 15.60 CI: 11.50

Moisture by the Cal Fitsha method: found 3.15%

Calculated 2.92%

NMR (at 60 MHz, D ₂ 0) τ: 3.20 (S, 5-H) 4.25 (d, 7-H) 4.75 (d, 6-H) 4.9-6.5 (m, 5 × CH ₂ ) 6.85 ( S, 2 × CH ₃ ).

The purity of the product by high-performance liquid chromatography was 99.6% in terms of anhydride, and the X-ray diffraction pattern indicated that the product was a crystal. Conditions for Liquid Chromatography:

Separation tube: Hitachi ion exchange resin 2614, 21mm × 50cm, Separation tube temperature: 50 ℃, Eluent: 0.3M citric acid buffer solution, pH6.5, 0.2ml / min, Pressure: 4.3kg / cm ² Recorder sensitivity: 10mv, Recording speed : 2.5 mm / min.

(4) 10 g of the crystal obtained in (3) was vacuum dried at 35 deg. C in the presence of phosphoric anhydride at a vacuum degree of 2 mmHg for 3 hours to obtain 9.8 g of the carboxylic acid dihydrochloride powder. The moisture measurement value 1.9% by the Karl Fischer method and this product were amorphous powder in the X-ray powder diffraction pattern. Observation by a polarizing microscope showed the interference color, optically anisotropy, and crystallinity when the material stage was rotated using orthogonal Nicole.

(5) 20 g of the crystal obtained in (3) was dissolved in 200 L of water, cooled to 10 DEG C, passed through a column filled with 135 ml of Amberlite IR-45 (type OH) for 30 minutes and washed with 300 ml of water. The effluent was lyophilized to give 16.0 g of 7β- [2- (2-amino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl]- 1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid was obtained. The moisture by the Kalpitsha method of this product was 3.20%, and the purity of the anhydride converted by high-performance liquid chromatography was 99.2%.

Example 2

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido-3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl ] 10 g of thimethyl-3-cepem-4-carboxylic acid was dissolved in 30 ml of water, and 150 ml of methylethylketone and 5.5 ml of 12N-HCl were added with stirring at 5 ° C. After stirring for 3 hours, the mixture was left overnight at 5 ° C., and the crystals thus obtained were filtered off, washed four times with 20 ml of methyl ethyl ketone, and dried to obtain 10.5 g of dihydrochloride crystals of the title carboxylic acid.

Elemental analysis _{(C 18 H 23 N 9 O} 4 S 3. 2HCI. 2H 2 O)

Found C: 33.94 H: 4.80 N: 20.02 CI: 11.15

Calculated C: 34.07 H: 4.61 N: 19.86 CI: 11.17

Moisture by the Calfitsha method: measured value 5.80%

Calculated 5.68%

Example 3

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido-3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl ] 0.53 g of thimethyl-3-cepem-4-carboxylic acid was dissolved in 1.5 ml of water, and 0.7 ml of 5N-HBr aqueous solution was added at 5 ° C.

11 ml of acetone was added to this solution, the mixture was stirred at 10 ° C. for 2 hours, 5 ml of acetone was further added, and the mixture was stirred at 25 ° C. for 1 hour. The crystals thus obtained were filtered off, washed 5 times with 2 ml of acetone, dried in air, and dried under vacuum to obtain 0.56 g of dihydrogen hydrochloride crystals of the title carboxylic acid.

NMR (in 60 MHz, D ₂ 0) τ: 3.22 (S, Chiazoline 5-H) 4.26 (d, 7-H) 4.75 (d, 6-H) 4, 9-6.5 (m, 5 × CH ₂ ) 6.88 (S, 2 × CH ₃ )

It was confirmed that this product was crystallinity as a result of X-ray powder diffraction pattern and polarization microscope observation.

Example 4

(1) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazole 5.26 g of a freeze-dried product of -5-yl] thiomethyl-3-cepem-4-carboxylic acid (this product had a water content of 3.2% as measured by the Karl Fischer method and a binary ion content of 95.5% as determined by high performance liquid chromatography). The carboxylic acid was dissolved in 50 ml of water, cooled to 5 ° C., 18.7 ml of 1N-HCI, and lyophilized. 6.2 g of a powder of 2HCI.1.5H ₂ O was obtained.

Moisture content: (Kalfitsha method) 4.2% (calculated 4.3% of C ₁₈ H ₂₃ N ₉ O ₄ S ₃ .2HCI.1.5H ₂ O)

Elemental Analysis Value (C ₁₈ H ₂₃ N ₉ O ₄ S ₃ .2HCI.1.5H ₂ O)

Found C: 34.21 H: 4.03 N: 19.82 CI: 11.50

Calculated C: 34.56 H: 4.19 N: 20.15 CI: 11.33

Purity: 95.3% (quantified by high performance liquid chromatography, converted into 2HCI brine)

X-ray powder shape: amorphous

Observation by the polarizing microscope: Even when the material band was rotated using orthogonal nicotine, it was amorphous without exhibiting interference colors.

(2) The carboxylic acid was dried by vacuum drying 2 g of the powder obtained in (1) in a desiccator containing silica gel. 1.9 g of ₂ HCI.0.17 H ₂ O powder was obtained.

Moisture content (kalpit _{syabeop) 0.5% (C 18 H 23} N 9 O 4 S 3 .2HCI 0.17 Calcd 0.5% of H ₂ O), purity 95.3% (quantified by high performance liquid chromatography and was converted to the dihydrochloride anhydride), X-ray powder diffraction pattern Amorphous, polarization microscope observation: No interference color.

(3) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] obtained in (1). -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid. 0.5 g of pure water was added to 1 g of the lyophilized product of dihydrochloride, dissolved for 15 hours at 5 ° C, and dried in a desiccator containing phosphoric anhydride to obtain 1.05 g of the dihydrochloride crystal of carboxylic acid.

Elemental Analysis Value (C ₁₈ H ₂₃ N ₉ O ₄ S ₃ .2HCI-2H ₂ O)

Found C: 33.84 H: 4.64 N: 19.71 S: 15.40 CI: 11.29

Calculated C: 34.07 H: 4.61 N: 19.86 S: 15.16 CI: 11.17

Moisture measurement by Kalpitsha method: 6.00% (calculated 5.68%)

_{^{IR (KBr) cm - 1:}} 1770 (β-lactam), a distinctive narrow and sharp absorption in the crystal is shown in 1670,1190,1170.

Example 5

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl with 93% purity in terms of anhydride ] 5.5 g of a freeze-dried product of] -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid was dissolved in 25 ml of water, 25 ml of acetone was added, then 4 ml of 12N-HCI was added, and acetone was further added. After adding 100ml, the mixture was stirred at 15 ° C for one hour. Again 50 ml of acetone were added, stirred for one hour, filtered, and washed four times with 10 ml of acetone. 5.55 g of dihydrochloride crystals of the carboxylic acid were obtained by drying.

(92% purity conversion)

Elemental Analysis Value (C ₁₈ H ₂₃ N ₉ O ₄ S. 2HCI. H ₂ O)

Found C: 34.48 H: 4.48 N: 20.55 S: 15.22 CI: 11.49

Calculated C: 35.06 H: 4.41 N: 20.45 S: 15.60 CI: 11.50

This product appeared to be a crystalline powder in X-ray powder diffraction form, and the purity of anhydride measured by high performance liquid chromatography was 99.4%.

Example 6

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido-3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl ] 19 g of 2N-HCI was added to 6.2 g of a freeze-dried product of thimethyl-3-cepem-4-carboxylic acid dihydrochloride, and the mixture was mixed for 2 hours with 250 ml of ethanol and stirring at 5 ° C. After stirring at 10 ° C. for 2 hours, the precipitate was filtered off, washed four times with 15 ml of ethanol, dried in air, and dried under reduced pressure at 30 ° C. to obtain 5.6 g of the carboxylic acid dihydrochloride crystal.

This product was confirmed to be crystalline as a result of X-ray powder diffraction and polarization microscope observation.

Example 7

를 10℃로 생각하고 활성탄 7.0g을 첨가하여 5분간 교반한 후 활성탄을 여과하여 제거하고 활성탄을 500ml의 물로 세척한 후 여액, 세액을 합하여서 2.38

까지 감압하에 내온 15-17℃에서 농축하고 또 다시 여과세척했다. 여과액 및 세척액은 2.38ℓ이며 상기 카르본산 478g을 함유하고 있었다. 여액에 아세톤 0.2

를 첨가하고 이어서 12N-HCI 170ml을 첨가했다. 다시 7

의 아세톤을 10분간 걸려서 첨가하고, 5-10℃에서 두시간 교반한 후 또 다시 아세톤 7

를 30분 걸려서 첨가하였다. 또 한시간 교반한 후 하룻밤 정치하여 정출된 결정을 여취하고 아세톤 1

로 4회 세척하였다. 본 결정(결정의 일부를 취하여 데시케이터중에서 30분간 실온, 진공도 30mmHg의 조건으로 건조시킨 것은 칼핏샤법에 의하여 측정한 수분함량이 8.9%이고 아세톤 2.2%가 부착되어 있었다. C₁₈H₂₃N₉O₄S₃-2HCI-3H₂O의 수분의 계산치는 8.28%)을 다른 유리필터에 옮겨서 물을 넣은 병에 통과시켜 가습시킨(수온은 25-30℃로 유지함)질소가스를 8

/min의 속도로 6시간 결정층에 통과시켰다. 이 결정 (결정의 일부를 취하여 칼핏샤법에 의하여 측정한 수분은 19.5%였다. C₁₈H₂₃N₉O₄S_3.2HCI·8H₂O의 수분의 계산치는 19.41%, 본품은 아세톤을 전혀 함유하고 있지않고 X선 분말회절형태는 결정성을 나타냈다)을 약 3cm의 두께로 넓히고 30℃에서 5mmHg의 진공도를 1.5시간 건조시켰다.(이것을 일부 취하여 칼핏샤법으로 측정한 수분은 17.2%이었다. C₁₈H₂₃N₉O₄S_3.2HCI. 7H₂O의 수분의 계산치는 17.41%) 또 같은 조건에서 1.5시간 건조시키고( 칼핏샤법에 의한 수분 15.4%) C₁₈H₂₃N₉O₄S_3.2HCI. 6H₂O로서의 수분의 계산치는 15.3%) 또 1.5시간 건조시키고(칼핏샤법에 의한 수분 13.3%C₁₈H₂₃N₉O₄S₃·2HCI·5H₂O의 계산치는 13.08%) 또 1.5시간 건조시키고 (칼핏샤법에 의한 수분 10.5% C₁₈H₂₃N₉O₄S₃·2HCI·4H₂O의 계산치는 10.75%)또 1.5시간 건조하여 525g의 결정을 얻었다.7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5- Aqueous solution of pH 2.0 containing 510 g of thimethyl-3-cepem-4-carboxylic acid.

Is considered to be 10 ℃, add 7.0g of activated carbon, stir for 5 minutes, filter out activated carbon, wash activated carbon with 500ml of water, add filtrate and tax solution, and add 2.38.

The mixture was concentrated under reduced pressure at 15-17 ° C. under reduced pressure and filtered again. The filtrate and wash were 2.38 L and contained 478 g of the carboxylic acid. Acetone 0.2 in the filtrate

Was added followed by 170 ml of 12N-HCI. 7 again

Acetone was added over 10 minutes, stirred at 5-10 ° C. for 2 hours, and then again with acetone 7

Was added over 30 minutes. After stirring for another hour, it was allowed to stand overnight to filter out the crystallized acetone 1

Washed 4 times. The crystals (part of the crystals were dried in a desiccator for 30 minutes at room temperature and a vacuum degree of 30 mmHg) had a water content of 8.9% and acetone 2.2% as measured by the Kalfitsha method. C ₁₈ H ₂₃ N ₉ The calculated water content of O ₄ S ₃ -2HCI-3H ₂ O is 8.28%), transferred to another glass filter, and passed through a bottle filled with water to humidify (keep water temperature at 25-30 ℃).

Passed through the crystal layer for 6 hours at a rate of / min. This crystal (Moisture measured by Kalfitsha method with a part of crystal was 19.5%. C ₁₈ H ₂₃ N ₉ O ₄ S _3. The calculated value of water of 2HCI · 8H ₂ O was 19.41%, and the product contained no acetone. The X-ray powder diffraction pattern showed crystallinity) to a thickness of about 3 cm and dried at a vacuum degree of 5 mmHg at 30 ° C. for 1.5 hours (partially taken and measured by Kalpitsha method was 17.2%. C _{18 H 23 N 9 O 4 S 3.} 2HCI. calculated values of 17.41% of water 7H ₂ O) and 1.5 hours again dried under the same conditions (15.4% moisture by kalpit _{syabeop) C 18 H 23 N 9 O} 4 S 3. 2HCI. The calculated value of water as 6H ₂ O was 15.3%) and dried for another 1.5 hours (the calculated value of 13.3% C ₁₈ H ₂₃ N ₉ O ₄ S ₃ · 2HCI · 5H ₂ O by Calpissha method was 13.08%) and dried for 1.5 hours. (Calculated value of water 10.5% C ₁₈ H ₂₃ N ₉ O ₄ S ₃ .2HCI.4H ₂ O by Kalpitsha method was 10.75%) and dried for 1.5 hours to obtain 525 g of crystals.

Moisture content (Kalfitsha method): 8.50% (Calculated as C ₁₈ H ₂₃ N ₉ O ₄ S ₃ · 2HCI · 3H ₂ O, 8.28%, X-ray powder diffraction type: Crystallinity. CI content (AgNO ₃ titration method) : 10.6% (calculated for C ₁₈ H ₂₃ N ₉ O ₄ .2HCI. 3H ₂ O 10.8%).

(2) The crystals obtained in (1) were dried at 30 DEG C in a vacuum of 2 mmHg in the presence of phosphoric anhydride for 5 hours to obtain 510 g of crystals. Moisture content (Kalfitsha method): 5.7% (calculated C ₁₈ H ₂₃ N ₉ O ₄ S _3. 2HCI. 2H ₂ O 5.68%): X-ray powder diffraction Form: Crystallinity. IR (KBr) cm ^-1 : 1770 (β-lactan), sharp peaks peculiar to crystallization appear at 1670, 1190 (sh.), 1170.

(3) The crystals obtained in (2) were dried at 30 ° C. in a vacuum of 2 mmHg in the presence of phosphoric anhydride for 8 hours to obtain 495 g of crystals. Moisture content (Kalfitsha method): 3.12% (C ₁₈ H ₂₃ N ₉ O ₄ S _3. 2HCI.H ₂ O calculated value 2.92%: Purity in terms of anhydrous (measured by high-performance liquid chromatography in terms of dihydrochloride anhydride). ): 99.5%, X-ray powder diffraction form: Crystallinity was shown.

Elemental analysis _{(C 18 H 23 N 9 O} 4 S 3. 2HCI. H 2 O)

Found C: 34.78 H: 4.51 N: 20.62 S: 15.31 CI: 11.77

Calculated C: 35.06 H: 4.41 N: 20.45 S: 15.60 CI: 11.50

(C=1%, H₂O)=+67.0°; 잔존용매(아세톤) : 50ppm 이하, CI함량(AgNO₃적정법) : 11.4%, 이론치 11.50 ; λmax (H₂O) : 258mμ(ε19.500) ; 생물학적역가 865mcg/ml

(C = 1%, H ₂ O) = + 67.0 °; Residual solvent (acetone): 50 ppm or less, CI content (AgNO ₃ titration method): 11.4%, theoretical value 11.50; λ max (H ₂ O): 258 mμ (ε 19.500); Biological titer 865mcg / ml

The biological titer is 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazole -5-yl] thiomethyl-3-cepem-4-carboxylic acid 2HCI salt was used as a standard and measured by the cup method using B. Subtilis ATCC 6633 as a test bacterium.

The anhydrous positive ion compound has a titer of 1000 mcg / ml and a calculated titer of 878 mcg / ml for a sample containing 2 moles of hydrochloric acid per mole of the zwitterionic compound and no impurities such as other non-binding hydrochloric acid, water, and residual solvent. will be ml.

Example 8

(1) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido-3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol- 5.26 g of a freeze-dried product of 5-yl] thiomethyl-3-cepem-4-carboxylic acid (this product had a moisture content of 3.1% measured by the Calpissha method and a binary ion content of 93.5% measured by high performance liquid chromatography). Dissolve in addition to 20 cc of water. To this solution was added 20 ml of acetone and 4 ml of 10N-HCI. 96 ml of acetone was added for 10 minutes with stirring at 20 ° C., followed by stirring for 2 hours, followed by addition of 72 ml of acetone for 20 minutes, followed by stirring for 30 minutes. The crystals were washed five times with 15 ml of acetone. It dried in air | atmosphere for 2 hours in% humid air, and obtained 6.0 g of wet crystals (this product measured the moisture value by 13% by the Calpissha method. The product was dried under reduced pressure for 2 hours, and the said carboxylic acid 2HCI. 4H ₂ O crystal was obtained. 5.7 g was obtained.

Water content: (Kalfitsha method) 10.9% (calculated for C ₁₈ H ₂₃ N ₉ O ₄ S _3. 2HCI. 0.1H ₂ O 0.3%), X-ray powder diffraction form: Amorphous. Observation on the polarizing microscope: Orthogonal Nicole was used, and when rotating the stage, it showed interference color, optically anisotropic and crystallinity. Purity: 99.6% (quantified by high performance liquid chromatography, and converted into dihydrochloride anhydrous).

Example 9

(1) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazole 5 ml of aqueous solution containing -5-yl] thiomethyl-3-cepem-4-carboxylic acid (this solution is 10.05 g of the carboxylic acid and 15 mg. 1- [2- (N as a result of quantification by high performance liquid chromatography). (N-dimethylamino) ethyl] -5-mercapto-1H-tetrazole and other trace impurities) was added 1.0 ml of 4N-HCI and the solution was stirred in 300 ml of acetone cooled to 5 ° C. Little by little The powdered precipitate was filtered off, washed four times with 5 ml of acetone and dried under reduced pressure to give the carboxylic acid. 1.2 g of 2HCI.1.2H ₂ O powder was obtained.

Moisture content: (Kalfitsha method) 3.45% (calculated C ₁₈ H ₂₃ N ₉ O ₄ S ₃ · 2HCI · 1.2H ₂ O 3.48%), CI content: (AgNO ₂ titration method 11.6% (C ₁₈ H ₂₃ N ₉ O ₄ S ₃ · 2HCI · calculated 11.4% of 1.2H ₂ O) purity: 96.2% (also in terms of high performance liquid chromatography to measure the dihydrochloride anhydride) X-ray powder diffraction pattern: amorphous.

(2) 0.3 g of the powder obtained in (1) was spread over a charare and absorbed overnight at 25 ° C. in a desiccator containing a saturated aqueous NaBr solution. The powder is once hygroscopic and becomes oily and then solidifies.

It was dried under reduced pressure and the carboxylic acid. 2HCI. 0.3 g of 2.3H ₂ O crystals were obtained.

Moisture content: (Kalfitsha method) 6.6% (calculated C ₁₈ H ₂₃ N ₉ O ₄ S _3. 2HCI. 2.3H ₂ O 6.5%)

Purity: 95.2%, CI Content: Elemental Analysis 11.5% (Calculated 11.1%) X-ray powder diffraction Form: Crystallinity.

Example 10

에 첨가하고 -10℃에서 교반하면서 디사이클로헥실아민 59.8g을 5분간 걸려서 첨가하였다. 20분후 4-클로로-3-옥소-브티릴클로리드의 디클로로메탄용액 (1.4mol/kg)154g을 35분간 걸려서 첨가하였다. 35분후 또 22g을 추가하여 20분간 교반하였다. 이 용액에 물 300ml 및 5N-HCI 20ml을 첨가하여 25℃에서 30분 교반한 후 여과하고 수세하였다. 본 여과액 및 세척액을 디클로로메탄으로 세척하고 디콜로로메탄층을 물로 추출한 후 합친 수층에 치오우레아 12.5g을 첨가하고, 탄산수소나트륨으로 pH 5.0로 만들고 실온에서 3시간 방치한 후 10℃에서 하루밤 방치하고 탄산수소나트륨으로 pH 5.2로 만들었다. 이 액을 암버얼라이트 XAD-2 (100-200mesh) 2.5

로 충전된 컬럼에 통과시키고 7.5

의 물로 세척한 후 15%메탄올 4

이어서 40%메탄올 6

로 용출시켜서 6분획으로 하고 제2에서 제6분획의 유효부분을 농축하고 동결건조함으로써 7-β-[2-(2-이미노-4-치아조린-4-일)아세트아미도]-3-[1-[2-N,N-디메틸아미노)에틸]-1H-테트라졸-5-일]치오메틸-3-세펨-4-카르본산37.1g을 얻었다.(1) 42.4 g of 7-amido-3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid Methane 1.1

59.8 g of dicyclohexylamine was added over 5 minutes with stirring at -10 ° C. After 20 minutes, 154 g of 4-chloro-3-oxo-butyryl chloride dichloromethane solution (1.4 mol / kg) was added over 35 minutes. After 35 minutes, another 22 g was added and stirred for 20 minutes. 300 ml of water and 20 ml of 5N-HCI were added to the solution, stirred at 25 ° C. for 30 minutes, filtered, and washed with water. The filtrate and washings were washed with dichloromethane, the dichloromethane layer was extracted with water, 12.5 g of thiourea was added to the combined aqueous layers, the pH was adjusted to 5.0 with sodium hydrogencarbonate, and left at room temperature for 3 hours, followed by overnight at 10 ° C. It was left to pH 5.2 with sodium bicarbonate. This solution is added to Amberlite XAD-2 (100-200mesh) 2.5

Pass through the column filled with

15% methanol 4 after washing with water

40% methanol 6

Eluted with 6 fractions, and the effective portion of the second to sixth fractions was concentrated and lyophilized to obtain 7-β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3 37.1 g of-[1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid was obtained.

(2) 7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] obtained in (1). A freeze-dried product of -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid was treated in the same manner as in (3) and (4) of Example 1 to obtain a powder of the carboxylic acid dihydrochloride. Got it.

Example 11

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [with a mixture of 3 ml of 1.5N-HCI and 3 ml of n-propane was stirred at 10 ° C. 0.53 g of 2-N, N-dimethylamino) ethyl] -1H-tetrazol-5-yl] thiomethyl-3-cepem-4-carboxylic acid, followed by stirring at 10 DEG C for 5 hours with 9 ml of n-propane added thereto. After 9 ml of n-propane was added and left overnight at 5 ℃. The extracted crystals were filtered off and dried in the air to obtain 0.50 g of dihydrochloride crystals of the carboxylic acid.

IR (KBr) cm ^-1 1770 (β-lactam) 1670, 1190, 1170

Elemental Analysis Value (C ₁₈ H ₂₃ N ₉ O ₄ S ₃ · 2HCI · 2H ₂ O)

Found C: 33.81 H: 4.50 N: 20.11 CI: 10.98

Calculated C: 34.07 H: 4.61 N: 19.86 CI: 11.17

[exam]

7β- [2- (2-imino-4-thiazolin-4-yl) acetamido] -3- [1- [2-N, N-dimethylamino) ethyl] -1H-tetrazol-5- Sodium salt, zwitterion, monohydrochloride and dihydrochloride of dimethyl hydrochloride and dihydrochloride obtained according to the present invention (non-crystalline ingredients), dihydrochloride and dihydrochloride crystals Regarding the qualitative powder, each sample was stored at 50 DEG C for 4 weeks in a sealed vial, and the residual ratio was as follows.

Sample moisture residual ratio

Crystals of Example 1- (3) 3.1% 99%

"1- (4) Powder 1.9 98

"4- (2) Lyophilized 0.50 86

Sodium Salt "0.48 76

Positive (bipolar) ion "0.50 54

Lyophilized Monohydrochloride 0.58 73

Claims (1)

A method for producing a crystalline cephalosporin derivative represented by the following general formula (I), characterized by reacting a compound of the following general formula (II) with an acid represented by two or more moles of general formula HX.

In the above formula, x is chlorine or cancellation, n is a number from 0 to 6.