BE856243A - NEW PENICILLIN COMPOUNDS - Google Patents

Description

       

  New penicillin compounds.

  
The present invention relates to novel compounds

  
of penicillin having excellent anti-bacterial activity, products for injections containing the novel penicillin compounds, preparations for injections and a process for the preparation of the penicillin compounds.

  
The new penicillin compounds according to the invention are compounds of the following formula:

  

 <EMI ID = 1.1>


  
(in which R is a formyl or acetyl group), and their

  
pharmaceutically acceptable non-toxic salts.

  
 <EMI ID = 2.1>

  
are salts formed between the penicillin compounds of formula (I) and pharmaceutically acceptable non-toxic inorganic or organic bases and include, for example, non-toxic metal salts of penicillin compounds such as alkali metal salts (for example salts sodium or potassium) and alkaline-metal salts (for example salts ;; calcium or magnesium). The preferred non-toxic salts are the sodium and potassium salts, the sodium salt being particularly suitable.

  
The penicillin compounds of formula (I) exist as two tautomers (keto type and enol type) of for-

  
 <EMI ID = 3.1>

  
Descriptively, these tautomers are inclusive expressed by the keto type of formula (I) below.

  

 <EMI ID = 4.1>


  
Since the carbon atom marked with an asterisk, attributable to the alpha carbon of the p-hydroxyphenylglycine group in formulas (I) and (II), is an asymmetric carbon, the penicillin compounds of the invention expressed by the formula ( I), based on the steric configuration relating to carbon, include a D isomer, an L isomer and a mixture of these isomers (referred to as a D, L mixture). All these isomers and their mixtures are expressed by the formula
(I) given above.

  
Sometimes the penicillin compounds of formula (I) exist as hydrates, and these hydrates are also included in the penicillin compounds of the invention expressed by formula (I).

  
An object of the present invention is to provide novel penicillin compounds having excellent antibacterial activity against Gram-negative bacteria as well as Gram-positive bacteria, and a process for their preparation.

  
Another object of the present invention is to provide novel penicillin compounds having excellent antibacterial activity against Pseudomonas types, including Pseudomonas aeruginosa, bacteria resistant to.

  
 <EMI ID = 5.1>

  
compounds.

  
Yet another object of this invention is to provide injections for intramuscular, intravenous or subcutaneous administration and preparations for the formation of such injections, which contain a novel and selected penicillin compound having a excellent anti-bacterial activity towards the various bacteria mentioned above and having very low toxicity towards warm-blooded animals, including man.

  
Other objects and advantages of the invention will become apparent from the following description.

  
The novel penicillin compounds of the invention are inclusive represented by formula (I) but are individually expressed by the following formulas (Ia) and (Ib).

  
Formula (la).

  

 <EMI ID = 6.1>
 

  
 <EMI ID = 7.1>

  

 <EMI ID = 8.1>


  
The carbon atom marked with an asterisk in each of formulas (Ia) and (Ib) is an asymmetric carbon atom, and based on its steric configuration, it includes a D isomer, an L isomer and a D mixture, L. For convenience, the penicillin compounds of formula (Ia) will sometimes be referred to as “formylpiperazine derivatives”, and penicillin compounds of formula (Ib) as “acetylpiperazine derivatives”.

  
The D-isomers in particular exhibit excellent anti-bacterial activity and low toxicity, among formyl- and acetyl-piperazine derivatives, and are designated as follows:

  
Derived from D-formylpiperazine.

  
 <EMI ID = 9.1>

  
pyrido (2,3-d) pyrimidine-6-carboxamido_7-p-hydroxybenzyl-penicillin.

  
Derived from D-acetylpiperazine.

  
 <EMI ID = 10.1>

  
nicillin.

  
It is known so far (British patent n [deg.] <EMI ID = 11.1>
 <EMI ID = 12.1>
 (where A is a grouped carbocyclic aromatic ring or a six-membered heteroaromatic ring containing a nitrogen atom as a heteroatom, X is a sulfur atom or an oxygen atom and Y is a hydrogen atom, an alkanoyl group with 1 to 8 carbon atoms or an alkoxycarbonyl group with 2 to 5 carbon atoms &#65533;

  
which result from the substitution of the alpha-amino group of Ampicillin with an acyl group containing a heterocyclic ring. The British patent only describes the acylation of Ampicillin and does not describe the acylation of Ampicillin

  
 <EMI ID = 13.1>

  
specific dine-carboxylic (designated by pyridopyrimidine-carboxylic acid group) of the following formula (Ic), in which the 2- position is substituted, by a 4-formyl-1-piperazinyl- or 4-acetyl-1-piperazinyl group as in the present invention:

  

 <EMI ID = 14.1>


  
where R is a formyl or acetyl group.

  
As will be indicated later in Tables 1 to III, the investigations of the authors of the present invention show that the new penicillin compounds of the invention having a part of Amoxicillin or its isomer expressed by

  
 <EMI ID = 15.1>

  
bacterial, particularly vis-à-vis Pseudomonas aeruginosa, clearly superior to that of the penicillin derivatives resulting from the substitution of the alpha-amino group of Ampicillin by a specific pyridopyrimidinecarboxylic acid group of formula (Ie) of the present invention.

  
On the other hand, United States Patent No. [deg.] <EMI ID = 16.1> describes N-acylated Amoxicillin derivatives of the following formula:
 <EMI ID = 17.1>
 wherein ring A is a benzene ring or a 5 or 6 membered heteroaromatic ring containing one or two nitrogen atoms as heteroatoms, on which one or more of the groups may be present

  
lower alkyl, lower alkoxy, lower alkylthio, lower haloalkyl, lower alkylenedioxy, halogen, hydroxyl, nitro, free or protected amino, lower alkylamino, di-lower alkylamino and lower alkanoylamino; X is oxygen or sulfur and Y is hydrogen, lower alkoxycarbonyl or lower alkanoyl.

  
The specification of the Japanese patent placed on public inspection under the n [deg.] 82,683 / 74, corresponding to the patent of the United States of America 4,003,887 indicates very indefinitely that the ring A in the formula ( XI) may have a substituent. The compounds specifically described in this specification are the same as those described in US Pat. No. 4,003,887 which fall within the definition of formula (XI) given above.

  
The new penicillin compounds according to the invention

  
 <EMI ID = 18.1>

  
and in corresponding patents or patent specifications filed in other countries.

  
U.S. Patent 4,003,887 describes an O-acyl derivative of a compound of the following formula:

  

 <EMI ID = 19.1>


  
which results from the acylation of the hydroxyl group on the benzene ring of the Amoxicillin derivative (XIa) (referred to as a 2-dimethylaminopyridopyrimidine derivative) in which the amino group of Amoxicillin is acylated by a pyridopyrimidinecarboxylic acid group substituted with a dimethylamino group at position 2-. The structure of compound (XIa) is closest to the novel penicillin compounds of this invention. However, as indicated in Tables 1 to III, the novel penicillin compounds of the present invention are characterized in that they have very markedly superior anti-bacterial activity, especially against Pseudomonas aeruginosa, and toxicity. for warm-blooded animals very markedly lower than those of compound (XIa).

  
A compound of the following formula is also known (Belgian patent 833,063):

  

 <EMI ID = 20.1>


  
but this one is apparently of a different type.

  
This cephalosporin derivative has an entirely different main structure from that of the penicillin compounds.

  
 <EMI ID = 21.1>

  
dihydro-5-oxopyrido (2,3-d) pyrimidine (designated by the pyrido-pyrimidine part) is substituted by a 1-piperazinyl li- group

  
 <EMI ID = 22.1> the case in the present invention.

  
The authors of the present invention have already previously described that the compounds of formula

  

 <EMI ID = 23.1>


  
 <EMI ID = 24.1>

  
groups consisting of hydrogen, alkyl having 1 to 4 carbon atoms, hydroxyalkyl having 2 to 4 atoms

  
of carbon, benzyl, benzyl substituted by methoxy, phenyl, propargyl or acyl selected from the group consisting of lower alkano.yl, trifluoroacetyl, lower alkoxycarbonyl and lower alkoxycarbonyl substituted by phenyl, and R2 is a part selected from the group consisting of into hydrogen, alkyl having 1 to 4 carbon atoms, alkyl having 2 to

  
4 carbon atoms, substituted by a hydroxy or a halogen, vinyl, allyl or benzyl),

  
have excellent anti-bacterial activity against

  
 <EMI ID = 25.1>

  
Tables I to III, this compound does not exhibit any marked anti-bacterial activity against Gram-positive bacteria, whereas the novel penicillin compounds of this invention have excellent anti-bacterial activity both vis-à-vis - against Gram-positive bacteria than Gram-negative bacteria. It should also be noted that the structure of the novel penicillin compounds of the invention differs from that of the compound because the 2- position of the pyridopyrimidine carboxylic acid group as a structural factor is specifically substituted with a 4-formyl- or 4- group. acetyl-l-piperazinyl, / because the atom

  
 <EMI ID = 26.1>

  
as an ethyl group. Compounds of formula (XIII) in which IL ^ is a hydrogen atom have no substantial anti-bacterial activity, as shown in Table I.

  
Through the study of the novel penicillin compounds of this invention and several other compounds having similar structures, the authors of the present invention were able to find that, because the 2- position of the pyridopyrimidine part is substituted with a group 1 -piperazinyl containing two nitrogen atoms, the penicillin compounds according to the invention, expressed by formula (I) have an anti-bacterial activity, especially vis-à-vis Pseudomonas aeruginosa markedly higher than that of the penicillin compounds resulting from the substitution of the 2- position with a substituted amino group containing a nitrogen atom, for example, a dimethylamino group or a heterocyclic group containing a nitrogen atom, for example, a 1-pyrrolidinyl group;

   and that, because the 4- position of the piperazinyl group is substituted by a formyl or acetyl group, the penicillin compounds according to the invention have very low toxicity to warm-blooded animals while retaining high anti-bacterial activity in vivo.

  
The novel penicillin compounds of formula (I) and their pharmaceutically acceptable non-toxic salts can be produced, for example, by method (a) or

  
method (b) described below.

  
Method (a).

  
Process (a) can be briefly described by the following reaction scheme:

  
 <EMI ID = 27.1>

  

 <EMI ID = 28.1>


  
in which, R in formulas (I) and (II)

  
is as defined above.

  
According to process (a), the novel penicillin compound of formula (I) according to the invention or its pharmaceutically acceptable non-toxic salt is prepared by reacting

  
 <EMI ID = 29.1>

  
5-oxopyrido (2,3-d) pyrimidin-6-carboxylic (generally referred to as "2-substituted pyridopyrimidinecarboxylic acid"; its 4-formyl substituted product is referred to as "2-substituted FP" instead of "2-substituted" "; and its 4-acetyl substituted product is referred to as" substituted 2-AP "instead

  
of "2-substituted"), or its inorganic or organic salt, or its derivative reactive to the carboxyl group, with ltalpha-aminop-hydroxybenzylphenicillin (designated by the abbreviation AHEP) or its inorganic or organic salt or a derivative convertible to AHBP, in an aqueous or non-aqueous medium, then if desired, by hydrolyzing or catalytically hydrogenolysing the resulting product to form a penicillin compound according to the invention and finally, if desired, converting the reaction product into a non-salt. pharmaceutically acceptable toxicant.

  
The carboxyl-reactive derivatives of the 2-substituted pyridopyrimidine carboxylic acid of formula (II) include all reactive derivatives known and used in the field of the production of penicillins or cephalosporins. Specific examples are acid anhydrides formed with acids such as alkylcarbonic acids (eg ethylcarbonic acid, isopropylcarbonic acid, iso- or sec-butylcarbonic acid), alkylcarboxylic acids (eg pivalic acid, penta-

  
 <EMI ID = 30.1>

  
diethylphosphoric acid), and sulfonic acids
(eg methanesulfonic acid); its reactive esters such as p-nitrophenyl ester, trichlorophenyl ester, p-nitrophenylthioester of N-hydroxy-piperidine, N-hydroxysuccinimide ester or N-hydroxyphthalimide ester; its reactive amides such as N-carbonylimidazole or N-carbonyltetrazole; its acid halides such as the acid chloride; and its acid azides.

  
 <EMI ID = 31.1>

  
semi-synthetic penicillins, and can be used in method (a) of this invention. Typical examples of such derivatives are trimethylsilyl ester, trityl ester, p-nitrobenzyl ester and phenacyl ester. It is also possible to use derivatives obtained by protecting the hydroxy group of the benzene ring of AHBP, for example, with an ethoxycarbonyl or benzyloxy-carbonil group.

  
The inorganic or organic salts of 2-substituted pyrido-pyrimidinecarboxylic acid of formula (II) and / or of AHBP of formula (III) include alkali metal salts such as sodium or potassium salts and salts with bases organic like triethylamine or N-ethylmorpholine.

  
The above reaction is carried out at a temperature

  
 <EMI ID = 32.1>

  
solvent, preferably in the presence of a base. The solvent, base, and other reaction conditions are substantially the same as those used in penicillin chemistry.

  
For example, when using an anhydride derived from 2-substituted pyridopyrimidinecarboxylic acid of formula
(II) and ethyl chloroformate, the reaction is carried out under cooling or at room temperature in the presence of a tertiary amine such as triethylamine or N, N-dimethylaniline in an inert solvent such as acetone, tetrahydrofuran, dimethylformamide, chloroform, dichloromethane or hexamethylphosphoramide, a mixture of these inert solvents, water or a hydrous organic solvent.

  
When using a 2-substituted pyridopyrimidinecarboxylic acid N-hydroxysuccinimide ester of the formula

  
 <EMI ID = 33.1>

  
hours, in the presence of a base such as triethylamine, lutidine, sodium hydroxide or sodium carbonate in dimethylformamide, dichloromethane, dioxane, water or a mixture of these solvents.

  
The reaction between 2-substituted pyridopyrimidinecarboxylic acid of formula (II) or its salt and AHBP of formula
(III) or its salt is advantageously carried out in the presence of a condensing agent such as N, N'-dicyclohexylcarbodiimide.

  
When pyridopyrimidinecarboxy acid is reacted

  
 <EMI ID = 34.1>

  
carboxyl with a derivative convertible into AHBP of formula
(III) sometimes, depending on the reaction conditions, a reaction product is formed having a portion of AHBP convertible into the penicillin compound of this invention. For example, this fact occurs when the carboxyl group of the reaction product is in the form of an ester such as the silyl ester or when the hydroxyl group of the benzene ring of the reaction product is protected by a protecting group such as a group. acyl. In this case, the product is subsequently hydrolyzed or catalytically hydrogenolysed in a customary manner known in the field of penicillin chemistry to form the penicillin compound of this invention.

  
If desired, the penicillin compound of formula
(I) thus formed can be converted to its pharmaceutically acceptable non-toxic salt in a customary manner.

  
Since the carbon atom marked with an asterisk attributable to the alpha carbon of the p-hydroxyphenylglycine group in formula (III) in reaction scheme (a) is an asymmetric carbon atom, the AHBP of formula (III) includes a D isomer, an L isomer and a D, L mixture.

  
If the D-isomer of AHBP is used as a starting material in process (a), the corresponding penicillin compound of this invention can be obtained in a D form. If the D, L mixture of AHBP is used as a starting material , a penicillin compound of this invention is obtained in a mixed form D, L. As mentioned above, the novel penicillins of this invention, in D-form, have particularly high anti-bacterial activities.

  
Therefore, in the method (a), the type of the AHBP isomer of formula (III) used as the starting material is selected depending on the desired isomer of the final product.

  
Method (b).

  
The novel penicillin compounds according to the invention can also be produced by method (b) mentioned below.

  
Process (b) can be briefly described by the following reaction scheme (b).

  
 <EMI ID = 35.1>

  
Step 1.

  

 <EMI ID = 36.1>


  
pans which 8. is as defined above.

  
Process (b) schematically given above consists of step 1 and step 2. Step 1 comprises the reaction between 2-substituted pyridopyrimidinecarboxylic acid of formula
(II), its inorganic or organic salt or its derivative reactive with the carboxyl group and a p-hydroxyphenylglycine of formula
(IV), or its inorganic or organic salt or its derivative for

  
 <EMI ID = 37.1>

  
(referred to as 2-substituted pyridopyrimidinecarbonylglycine or PPCG). Step 2 involves the reaction between PPCG, its salt

  
or its derivative reactive with the carboxyl group and 6-aminopenicillanic acid of formula (VI) (denoted by 6-APA), its inorganic or organic salt or its derivative convertible to 6-APA, optionally hydrolysis or catalytic hydrogenolysis of the resulting product to form a penicillin compound of formula (I), and optionally converting the reaction product to its pharmaceutically acceptable non-toxic salt.

  
The inorganic or organic salt or the carboxyl-reactive derivative of 2-substituted pyridopyrimidinecarboxylic acid of formula (II) used in step 1 can be identical to that described in process (a).

The reaction of step 1 can be carried out between

  
-40 and +40 "C., Preferably at a temperature below room temperature, in the same aqueous or non-aqueous solvent as that described above in process (a).

  
The PPCG of formula (V) formed by the reaction of step 1 is a new compound.

  
In step 2, it is preferable to use PPCG in the form of its salt or its derivative reactive with the carboxyl group.

  
These salts and reactive derivatives are the same as those described above with respect to the salts and derivatives reactive to the carboxyl group of the 2-substituted pyrimidopyrimidinecarboxylic acid used in process (a). A derivative of PPCG in which the hydroxyl group of the benzene ring is protected by an acyl group can be used as the starting compound in step 2.

  
The 6-APA salts or the 6-APA convertible derivatives used in step 2 may be substantially identical to the salts or derivatives of AHBP described above in process (a) except for an AHBP derivative in which the group hydroxyl to the benzene ring is protected. In this reaction it is preferable to use 6-APA in the form of such a derivative. ''

  
Preferred 6-APA derivatives are, e.g., its trimethylsilyl ester, its trityl ester, its p-nitrobenzyl ester, its phenacyl ester and its O, Nbistrimethylsilyl derivative.

  
It is particularly advantageous to react the derivative of 6-APA with the salt or the reactive derivative of the carboxyl group of PPCG in an aprotic medium.

  
When 6-APA is used in the form of free carboxylic acid or its salt, the reaction of step 2 can also be carried out in a protic medium or in an aqueous medium.

  
The reaction of step 2 is preferably carried out

  
 <EMI ID = 38.1>

  
preferably in the presence of a base. The solvent, base, and other reaction conditions are substantially the same as those used in penicillin chemistry.

  
For example, when reacting an anhydride derivative of PPCG of formula (V) and an ethyl chloroformate with a derivative of 6-APA, the reaction is carried out under cooling or at room temperature in the presence of a tertiary amine such as triethylamine or N, N-dimethylaniline in an inert solvent such as acetone, tetrahydrofuran, dimethylformamide, chloroform, dichloromethane, hexamethylphosphoramide or a mixture of these inert solvents.

  
When the N-hydroxysuccinimide ester of PPCG is reacted with a derivative convertible to 6-APA, the reaction is carried out between 0 and 20 [deg.] C. for one hour to 2 days in the presence of a base such as triethylamine or: '_ lutidine in dimethylformamide, dichloromethane, dioxane or a mixture thereof.

  
When PPCG of formula (V) or its salt or derivative thereof reactive with the carboxyl group or its derivative having a hydroxyl group protected at the benzene ring is reacted with a derivative convertible to 6-APA, it is sometimes formed, depending on the conditions of the reaction, a reaction product having a portion of 6-APA and / or PPCG convertible into the compound of pe-

  
 <EMI ID = 39.1>

  
 <EMI ID = 40.1>

  
carboxyl- of the reaction product is in the form of an ester such as a silyl ester, or when the hydroxyl group of the benzene ring of the reaction product is protected by a protective group such as an acyl group. In such a case,

  
the reaction product is hydrolyzed or catalytically hydrogenolysed in a manner customary in the field of penicillin chemistry, to obtain the penicillin compound of the invention.

The penicillin compounds obtained by the process

  
(a) or process (b) can be in the form of free carboxylic acid or salt depending on the choice of starting compounds, reaction conditions, etc. The carboxylic acid is converted to a salt by treatment with a basic substance such as sodium 2-ethylhexanoate, sodium hydroxide or sodium carbonate. The salt, on the other hand, is treated with an acidic substance, if desired, to form a free carboxylic acid.

  
The isolation, purification, extraction and recrystallization and other post-treatments of the product are carried out according to the usual procedures used in penicillin chemistry.

  
The starting compound of formula (II) can be obtained by intramolecularly cyclizing a compound of general formula:

  

 <EMI ID = 41.1>


  
 <EMI ID = 42.1>

  
mes of carbon), to form a compound of general formula:

  

 <EMI ID = 43.1>


  
 <EMI ID = 44.1>

  
sant compound (VIII) to form a. compound of formula

  

 <EMI ID = 45.1>


  
and then effecting formylation or acetylation of compound (IX).

  
Formylation or acetylation can be carried out by suspending the compound of formula (IX) in a formylating agent such as a mixture of acetic anhydride and formic acid or a mixture of formic acid and formamide, or in an acetylating agent such as acetic anhydride and heating to 60 - 90 [deg.] C. for 2 to 3 hours.

  
Substituted pyridopyrimidinecarboxylic acid 2-AP can also be obtained by hydrolyzing the compound of formula
(VIII) under moderate conditions in a mixture of hydrochloric acid and ethanol, for example.

  
The intramolecular cyclization reaction of converting the compound of formula (VII) into the compound of formula (VIII) and hydrolysis for the conversion of the compound of formula (VIII) into the compound of formula (IX) can be carried out according to the following procedure. specification of United States Patent 3,887,557.

  
The preparation of the novel penicillin compounds of the invention and their pharmacological properties are described below. The following description consists of five parts.

  
Part I.

  
References 1 to 9 show the preparation of the starting compound of formula (II) used in process (a)
(references 1 to 7), and the preparation of the starting compound of formula - (V) used in process (b) (references 7 and

  
 <EMI ID = 46.1>

  
Part II.

  
Examples 1 to 16 show the preparation of the novel penicillin compounds of the invention by the process
(a) and method (b).

  
Part III.

  
References 10 to 16 show the preparation of compounds which are outside the scope of the invention, but for which no preparation process is known. These references are used to evaluate the pharmacological actions of the penicillin compounds of the invention.

  
Part IV.

  
Examples 17 and 18 show the production of medicaments using the penicillin compounds of the invention.

  
Part V.

  
Examples 19 to 24 and Tables I to V show the pharmacological actions of the penicillin compounds of the invention, of Amoxicillin (known anti-bacterial agent), of penicillin derivatives not included within the scope of the invention and having structures similar to those of penicillin compounds of this invention, etc.

  
At the beginning of each example and each reference in Parts I to III, the summary of the process which is employed is presented by a reaction formula.

  
 <EMI ID = 47.1>

  
References 1 to 9 presenting the process for the preparation of the starting compounds used in the present invention.

  
Reference 1.

  

 <EMI ID = 48.1>


  
A dephenyl ether (16 ml.) Maintained at 250-255 [deg.] C.,

  
 <EMI ID = 49.1>

  
Diethyl pyrimidinyl_7-aminoethylenemanolate (2.0 g.). The mixture was gently refluxed for 10 minutes and then allowed to stand to cool to room temperature. To the mixture was added n-hexane (12 ml.). The resulting precipitate was collected, washed with ethanol and recrystallized from ethanol to give 2-

  
 <EMI ID = 50.1>

  
Ethyl rimidine-6-carboxylate (1.52 g.), m.p. 300-302 [deg.] C.
(decomposition).

  
Reference 2.

  

 <EMI ID = 51.1>


  
A suspension of 5,8-dihydro-2- (4-acetyl-1-piperazi-

  
 <EMI ID = 52.1>

  
(1,000 g.) And concentrated hydrochloric acid (800 ml.) In a mixture of water (1,200 ml.) And ethanol (1,000 ml.)

  
was refluxed for 8 hours and then left to stand overnight. The precipitated crystals were collected, washed with ethanol, and dissolved with heating in

  
an aqueous solution (10 liters) of sodium hydroxide (250 g).

  
 <EMI ID = 53.1>

  
acetic, and left to stand overnight while cooling with ice. The crystalline precipitate was collected, washed with water and dried at 110 [deg.] C. to give

  
 <EMI ID = 54.1> <EMI ID = 55.1>

  
C 52.36 H 4.76 N 25.45 Found C 52.10 H 4.63 N 25.32 Reference 3.

  

 <EMI ID = 56.1>


  
Formic acid (140 ml.) Was added dropwise to acetic anhydride (200 ml.) Cooled on an ice bath. The solution was heated to 50 [deg.] C. during 15

  
 <EMI ID = 57.1>

  
Calculated: C 51.40 H 4.32 N 23.09 Found: C 51.41 H 4.47 N 23.04

  
 <EMI ID = 58.1>

  

 <EMI ID = 59.1>


  
By the same operating mode as that described in

  
 <EMI ID = 60.1> <EMI ID = 61.1>

  
Calculated: C 52.99 H 4.77 N 22.07 Found: C 52.92 H 4.57 H 21.91 Reference 5.

  

 <EMI ID = 62.1>


  
A suspension of 5, Ô-dihydro-2- (4-formyl-1-

  
 <EMI ID = 63.1>

  
(273 g.) And triethylamine (299 ml.) In dry dichloromethane (3,500 ml.) Was stirred well at room temperature for one hour. To the suspension was added dropwise and while cooling to 0-10 [deg.] C. Isopropyl chloroformate (221 g.) And the reaction mixture was stirred for 2 hours. A solution of N-hydroxysuccinimide (207 g.) In dimethylformamide (350 ml.) Was added to the mixture and the resulting mixture was kept at 5 - 10 [deg.] C. for another 2 hours. The crystalline product was collected by filtration and washed successively with water (2,000 ml.) And acetone (2,000 ml.) To give

  
 <EMI ID = 64.1> <EMI ID = 65.1>

  
Reference 6.

  

 <EMI ID = 66.1>


  
A suspension of 5,8-dihydro-2- (4-formyl-1-pipé) acid

  
 <EMI ID = 67.1>

  
and triethylamine (6.1 ml.) in anhydrous dichloromethane
(100 ml.) Was well stirred at room temperature for one hour. To the solution, ethyl chloroformate

  
 <EMI ID = 68.1>

  
The reaction was stirred for one hour at 5-7 [deg.] C. The insoluble substance was removed by filtration and to the filtrate was added a solution of N-hydroxyphthalamide (7.19 g.) In

  
dimethylformamide (10 ml.). The resulting mixture was kept at room temperature for 2 hours. The crystalline product was collected by filtration and washed successively with dichloromethane and acetone to give N-

  
 <EMI ID = 69.1>

  
Found: C 56.03 H 3.54 N 18.59 Reference 7.

  

 <EMI ID = 70.1>


  
 <EMI ID = 71.1>

  
(100 ml.) Was stirred well at room temperature for one hour. In the suspension, ethyl chloroformate

  
 <EMI ID = 72.1>

  
ble was removed by filtration and to the filtrate was added a solution of 2,4,6-trichlorophenol (8.68 g.) in dichloromethane (30 ml.). The reaction mixture was kept at room temperature for 3 hours. The crystalline product was collected by filtration and washed with dichloromethane to give 5,8-dihydro-2- (4-formyl-1-piperazi-

  
 <EMI ID = 73.1>

  
Calculated: C 47.20 H 2.92 N 14.51 Found: C 47.12 H 3.22 N 14.72 Reference 8.

  

 <EMI ID = 74.1>


  
 <EMI ID = 75.1>

  
and triethylamine (5.6 ml.) in dimethylformamide (50 ml.) was stirred at room temperature for one hour.

  
 <EMI ID = 76.1>

  
succinimide (8.0 g.). The mixture was stirred at 5 [deg.] C. for one hour and then stirred at room temperature for a further 3 hours. The resulting precipitate was collected and dissolved in ice water. The aqueous solution was acidified with 10% hydrochloric acid. The precipitated crystals were collected and recrystallized from

  
 <EMI ID = 77.1>

  

 <EMI ID = 78.1>


  
 <EMI ID = 79.1>

  
p-hydroxyphenylacetic (9.0 g.) and N-hydroxysuccinimide
(4.6 g.) In anhydrous dimethylformamide (100 ml.), Di-

  
 <EMI ID = 80.1>

  
The mixture was stirred at -10 to 0 [deg.] C. for one hour and then left to stand overnight at room temperature. The precipitate was removed by filtration and to the filtrate was added ice water (500 ml.). The precipitated crystals were collected and dissolved in acetonitrile.
(50 ml.). The acetonitrile solution was dried over anhydrous magnesium sulfate and was concentrated under reduced pressure. The resulting crystals were collected for

  
 <EMI ID = 81.1>

  

 <EMI ID = 82.1>


  
By the same operating mode as that described in

  
 <EMI ID = 83.1> <EMI ID = 84.1>

  
succinimide.

  
(Part II.) -

  
Examples 1 to 16 showing the production of the penicillin compounds (I) of the invention.

Example 1.

  

 <EMI ID = 85.1>


  
To an aqueous solution (2000 ml.) Of sodium hydroxide (22 g.) Was added successively at 0 - 2 [deg.] C. and in

  
 <EMI ID = 86.1>

  
 <EMI ID = 87.1>

  
g.) and the mixture was stirred for half an hour. A so-

  
 <EMI ID = 88.1>

  
mixture and the resulting mixture was held for an additional hour and was then filtered. To the filtrate was added ice water (3,000 ml.). The solution has been adjusted

  
at pH 2 with 10% hydrochloric acid. The resulting precipitate was collected by filtration, washed twice with ice water (4,000 ml.) And dissolved in 4% sodium hydroxide solution of sufficient volume to adjust the pH to 6.5. The aqueous solution was filtered and the filtrate was lyophilized to give the sodium salt of

  
 <EMI ID = 89.1>

  
 <EMI ID = 90.1>

  
 <EMI ID = 91.1>

  
 <EMI ID = 92.1>

  

 <EMI ID = 93.1>

H

  
 <EMI ID = 94.1>

  
 <EMI ID = 95.1>

  

 <EMI ID = 96.1>


  
 <EMI ID = 97.1>

Example 2.

  

 <EMI ID = 98.1>


  
 <EMI ID = 99.1>

  
reaction was stirred for 1.5 hours. A 30% solution of sodium 2-ethylhexanoate in n-butanol (335 ml.) And acetone (7,000 ml.) Was successively added to the mixture. The resulting precipitate was collected by filtration, washed with acetone and dissolved in ice water. The aqueous solution was acidified with hydrochloric acid to
10% and the precipitated crystals were collected, washed well with water and dissolved in 2% sodium hydroxide solution. The solution was added at pH 6.5 and then filtered. The filtrate was lyophilized to give the salt

  
 <EMI ID = 100.1>

  
zylpenicillin (289 g.).

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 1.

  
 <EMI ID = 101.1>

  

 <EMI ID = 102.1>


  
Has a solution of D- &#65533; -amino-p-hydroxybenzyl-penicillin (4.61 g.) and triethylamine (1.53 ml.) in dim-

  
 <EMI ID = 103.1>

  
6-carbonyloxy_7phthalamide (4.40 g.) And the reaction mixture was stirred at room temperature for one hour. Insoluble substance was removed by filtration and the filtrate.

  
 <EMI ID = 104.1>

  
sodium noate in n-butanol (7.3 ml.) and acetone
(300 ml.). The resulting precipitate was collected by filtration, washed with acetone and dissolved in ice water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The precipitate was collected, washed with water and dissolved in dimethylformamide (60 ml.). The solution was treated with charcoal (1.5 g.) And filtered. To the filtrate was successively added a 30% solution of sodium 2-ethylhexanoate in n-butanol (7 ml.) And acetone (300 ml.). The precipitate was collected and dissolved in ice water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The resulting precipitate was collected, washed with water and dissolved in 2% sodium hydroxide solution to adjust the pH to 6.5.

   The aqueous solution was filtered and lyophilized to

  
 <EMI ID = 105.1>

  
 <EMI ID = 106.1>

  
was identified as being the compound of Example 1.

Example 4.

  

 <EMI ID = 107.1>


  
 <EMI ID = 108.1>

  
line (1.11 g.) and triethylamine (0.38 ml.) in dimethyl-

  
 <EMI ID = 109.1>

  
late (2.16 g.) and the reaction mixture was stirred at room temperature for 2 days. The insoluble substance was removed by filtration and to the filtrate a 30% solution of sodium 2-ethylhexanoate in n-butanol (1.5 ml.) And acetone (150 ml.) Were successively added. The resulting precipitate was collected by filtration, washed with acetone and dissolved in ice water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The precipitate was collected, washed with water and dissolved in dimethylformamide. The solution in dimethylformamide was treated with charcoal and filtered. To the filtrate was successively added a 30% solution of sodium 2-ethylhexanoate in n-butanol and acetone. The precipitate was collected and dissolved in ice water.

   The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The precipitate was collected, washed with water and dissolved in 2% sodium hydroxide solution to adjust the pH.

  
to 6.5. The aqueous solution was filtered and lyophilized to

  
 <EMI ID = 110.1>

  
 <EMI ID = 111.1>

Example 5.

  

 <EMI ID = 112.1>


  
A suspension of 5,8-dihydro-2- (4-formyl-1-,

  
 <EMI ID = 113.1>

  
(90 g.) And triethylamine (42 ml.) In dry dichloromethane (1,000 ml.) Was well stirred at room temperature for one hour. While cooling, isopropyl chloroformate (36.8 g.) Was added to the suspension and the

  
 <EMI ID = 114.1>

  
of triethylamine (46 ml.) in dimethylformamide (350 ml.) was added to the mixture. The resulting mixture was stirred

  
at 5 - 7 [deg.] C. for a further 1.5 hours and then left to stand overnight at -10 [deg.] C.

  
The insoluble substance was removed by filtration and

  
 <EMI ID = 115.1>

  
Sodium 2-ethyl-hexanoate in n-butanol (250 ml.) And acetone (3,000 ml.).

  
The resulting precipitate was collected, by filtration and dissolved in ice-water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The precipitate was washed with water and dissolved in dimethylformamide (380 ml.). After treating the solution with charcoal (20 g.), A 30% solution of sodium 2-ethylhexanoate in butanol was successively added to the solution.
(130 ml.) And acetone (5,000 ml.).

  
The resulting precipitate was collected, washed with acetone and dissolved in ice water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid.

  
The precipitate was collected, washed with water and dissolved in 2% sodium hydroxide solution to adjust the pH to 6.5. The solution was filtered and lyophilized to give

  
 <EMI ID = 116.1>

  
p-hydroxybenzylpenicillin (78.7 g.).

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 1.

Example 6.

  

 <EMI ID = 117.1>
 

  

 <EMI ID = 118.1>


  
By the same operating mode as that described in

  
 <EMI ID = 119.1>

  
and isobutyl chloroformate (79 ml.).

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 1.

Example 7.

  

 <EMI ID = 120.1>


  
By the same operating mode as that described in

  
 <EMI ID = 121.1>

  
carboxamido_7-p-hydroxybenzylpenicillin (29 g.) was obtained from 5, B-dihydro-2- (4-formyl-1-piperazi-

  
 <EMI ID = 122.1>

  
and sec-butyl chloroformate (13.7 g.).

  
 <EMI ID = 123.1>

  
 <EMI ID = 124.1>

  
 <EMI ID = 125.1>

  

 <EMI ID = 126.1>


  
By the same operating mode as that described in

  
 <EMI ID = 127.1>

  
2-ethylhexanoyl (19.5 g.).

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 1.

Example 9.

  

 <EMI ID = 128.1>
 

  
 <EMI ID = 129.1>

  
g.) and triethylamine (1.66 ml.) in anhydrous dimethylformamide (50 ml. was stirred at room temperature for
30 minutes. To the reaction mixture was added ethyl chloroformate (1.14 ml.) At 5-10 [deg.] C. and the mixture was stirred for one hour.

  
On the other hand, a suspension of D-alpha-amino-p-hydro-

  
 <EMI ID = 130.1>

  
of anhydrous magnesium sulfate (2.0 g.) in dimethylformamide (60 ml.) was stirred at room temperature for
30 minutes, and the insoluble substance was filtered. The filtrate was added to the previous suspension, at 5-10 [deg.] C., And the mixture was stirred for a further 2 hours. Insoluble substance was removed by filtration and to the filtrate was added a 20% solution of sodium 2-ethylhexanoate in n-butanol (10 ml.). Then, ethyl ether (500 ml.) Was added thereto. The precipitated crystals were re-collected and dissolved in water (300 ml.). The aqueous solution

  
was acidified with dilute hydrochloric acid. The precipitate was collected, washed with water and dried to

  
 <EMI ID = 131.1>

  
zylpenicillin (3.4 g.) '

  
 <EMI ID = 132.1>

  

 <EMI ID = 133.1>


  
 <EMI ID = 134.1>

  
Calculated: C 49.43 H 5.15 N 15.90 S 4.55 Found: C 49.64 H 4.91 N 15, $ 6 S 4.72

Example 10.

  

 <EMI ID = 135.1>


  
 <EMI ID = 136.1>

  
g.) and triethylamine (4.98 ml.) in dry dichloromethane (180 ml.) was well stirred at room temperature for one hour. To the suspension, ethyl chloroformate (3.42 ml.) Was added dropwise while cooling.

  
 <EMI ID = 137.1>

  
hour. To the reaction mixture was added a solution of Dalpha-amino-p-hydroxybenzylpenicillin (15 g.) And triethylamine (7.5 ml.) In anhydrous dimethylformamide (120 ml.) And the resulting mixture was stirred for 2 hours. The insoluble substance was removed by filtration and to the filtrate a 20% solution of sodium 2-ethylhexanoate in n-butanol (27 ml.) And acetone (1000 ml.) Were successively added. The resulting precipitate was collected by filtration.

  
 <EMI ID = 138.1>

  
Quenched to pH 2 with 10% hydrochloric acid and the precipitate was collected, washed with water and dissolved in dimethylformamide (250 ml). After treatment of the solution in

  
 <EMI ID = 139.1>

  
Sodium 2-ethylhexanoate in n-butanol (27 ml.) And acetone (1,000 ml.) Were successively added to the solution. The precipitate was collected and dissolved in ice water. The aqueous solution was adjusted to pH 2 with 10% hydrochloric acid. The precipitated crystals were collected, washed with water, dissolved in a solution.

  
 <EMI ID = 140.1>

  

 <EMI ID = 141.1>


  
 <EMI ID = 142.1>

  
Found: C 47.59 H 4.95 N 14.56 S 4.02

Example 11.

  

 <EMI ID = 143.1>
 

  

 <EMI ID = 144.1>


  
A suspension of 5,8-dihydro-2- (4-acetyl-1-piperazinyl) -

  
 <EMI ID = 145.1>

  
 <EMI ID = 146.1>

  
thanol (20 ml.), was refluxed at 120 [deg.] C. for 1.5 hours. After cooling, the precipitate was collected, washed with water and then with hot water, and dissolved in 10% sodium hydroxide solution. Insoluble substance was removed by filtration and the filtrate was acidified with dilute hydrochloric acid. The resulting precipitate was collected and recrystallized from dimethyl sulfoxide to

  
 <EMI ID = 147.1>

  
g.) and triethylamine (0.73 ml.) in anhydrous dimethylformamide (20 ml.) was stirred at room temperature for
30 minutes. To the reaction mixture was added ethyl chloroformate (0.5 ml.) At a temperature of -5 to -10 [deg.] C., And the mixture was stirred for one hour.

  
On the other hand, a suspension of D-alpha-amino-p-hydroxybenzylpenicillin (2.10 g.), Triethylamine (1.05 ml.) And anhydrous magnesium sulfate (1.0 g.) In dimethylformamide anhydrous (20 ml.) was stirred at room temperature for 30 minutes, and the insoluble substance was removed by filtration. The filtrate was added to the previous suspension at a temperature of -5 to -10 [deg.] C. and the mixture was stirred for a further two hours. The insoluble substance was removed by filtration and to the filtrate was added a 20% solution of sodium 2-ethylhexanoate in n-butanol (7.5 ml.) And then ethyl ether (200 ml.). has been added. The precipitated crystals were collected and dissolved in sodium bicarbonate solution.

   The aqueous solution was acidified with dilute hydrochloric acid and the precipitate was collected, washed with water and dried to give the mixture.

  
 <EMI ID = 148.1>

  
ne (1.35 g.).

  
 <EMI ID = 149.1>

  
 <EMI ID = 150.1>

  

 <EMI ID = 151.1>


  
 <EMI ID = 152.1>

  
Found: C 46, Si H 4.99 N 14.78 S 4.19

Example 12.

  

 <EMI ID = 153.1>
 

  
By the same operating mode as that described in

  
 <EMI ID = 154.1>

  
pyrimidine-6-carbonyloxo_7succinimide.

  
 <EMI ID = 155.1>

  
was identified as being the compound of Example 10.

Example 13.

  

 <EMI ID = 156.1>


  
By the same operating mode as that described in

  
 <EMI ID = 157.1>

  
sopropyl.

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 10.

Example 14.

  

 <EMI ID = 158.1>


  
 <EMI ID = 159.1>

  
and N, O-bis (trimethylsilyl) acetamide (0.5 ml.) in anhydrous dimethylformamide (10 ml.) was stirred at room temperature for half an hour. To the resulting solution, we

  
 <EMI ID = 160.1>

  
hydroxyphenylacetic (0.9 g.) and the mixture was cooled on an ice bath. To the mixture was added dicyclohexylcarbodiimide (0.6 g.). The reaction mixture was kept at 0 - 5 [deg.] C. for one hour and then was allowed to stand at room temperature for 3 hours. The precipitate was removed by filtration and to the filtrate was added water.

  
 <EMI ID = 161.1>

  
 <EMI ID = 162.1>

  
 <EMI ID = 163.1>

  
sodium. The insoluble substance was removed by filtration and the filtrate was adjusted to pH 2 with hydrochloric acid.

  
 <EMI ID = 164.1>

  
li, washed with water and dissolved in 2% sodium hydroxide solution. The aqueous solution was adjusted to

  
pH 6.5, filtered and lyophilized to give the sodium salt of <EMI ID = 165.1> penicillin. <EMI ID = 166.1>

  

 <EMI ID = 167.1>

Example 15.

  

 <EMI ID = 168.1>


  
To a mixture of 6-aminopenicillanic acid (0.65 g.) And trimethylsilyl chloride (0.76 ml.) In anhydrous dichlorome-thane (20 ml.) Was added triethylamine
(0.83 ml.) At 0-5 [deg.] C. and the mixture was stirred at 0-5 [deg.] C. during

  
 <EMI ID = 169.1>

  
then was left to stand overnight at room temperature. The reaction mixture was extracted with 3% sodium bicarbonate solution and the extract was adjusted to pH 2 with 10% hydrochloric acid.

  
The precipitate was collected and dissolved in a small amount of dimethylformamide. A 30% solution of sodium 2-ethylhexanoate (1 ml.) And then acetone (50 ml.) Were added to the solution. The precipitate was collected and dissolved in ice water. The aqueous solution was

  
 <EMI ID = 170.1>

  
The precipitate was collected, washed with water and dissolved in 2% sodium hydroxide solution. The solution was adjusted to pH 6.5, filtered and lyophilized to give the salt.

  
 <EMI ID = 171.1>

  
p-hydroxybenzylpenicillin.

  
By comparison of the IR and NMR spectra, this compound was identified as being the compound of Example 14.

Example 16.

  

 <EMI ID = 172.1>
 

  
By the same operating mode as that described in ex.

  
 <EMI ID = 173.1>

  
&#65533; Part III).

  
The following references 10 to 16 show the preparation of compounds which are not included in the scope of

  
 <EMI ID = 174.1>

  
in order to evaluate the pharmacological actions of the novel penicillin compounds of the invention.

  
Reference 10.

  

 <EMI ID = 175.1>


  
By the same operating mode as that described in ex.

  
 <EMI ID = 176.1>

  
pyrimidine-6-carboxylic acid and D-alpha-amino-p-hydroxybenzylpenicillin.

  
 <EMI ID = 177.1>

  
>
 <EMI ID = 178.1>
 <EMI ID = 179.1>

  
Found: C 47.95 H 4.96 N 15.19 S 4.53

  
Reference 11.

  

 <EMI ID = 180.1>


  
 <EMI ID = 181.1>

  
triethylamine (0.73 ml.) in a mixture of anhydrous hexamethylphosphoramide (25 ml.) and dichloromethane (5 ml.) was added ethyl chloroformate (0.5 ml.) at a temperature

  
 <EMI ID = 182.1> triethylamine (1.0 ml.) In a mixture of hexamethylphosphoramide (20 ml.) And dichloromethane (10 ml.) And the mixture was stirred for one hour. After adding dichloromethane (40 ml.) And ice-water, the mixture was adjusted to pH 2 with 10% hydrochloric acid. The organic layer was separated, washed with water and extracted with saturated sodium bicarbonate solution. The extract

  
 <EMI ID = 183.1>

  
The precipitate was collected by filtration, washed with

  
 <EMI ID = 184.1>

  

 <EMI ID = 185.1>


  
 <EMI ID = 186.1>

  
Reference 12.

  

 <EMI ID = 187.1>
 

  
By the same operating mode as that described in

  
 <EMI ID = 188.1>

  
boxyl (1.18 g.) and D-alpha-p-hydroxybenzylpenicillin (1.50 g.).

  
 <EMI ID = 189.1>

  
 <EMI ID = 190.1>

  

 <EMI ID = 191.1>


  
Calculated: C 49.12 H 5.23 N 15.42 S 5.04

  
 <EMI ID = 192.1>

  
Reference 13.

  

 <EMI ID = 193.1>


  
 <EMI ID = 194.1>

  
amine (2.5 ml.) in anhydrous dimethylformamide (50 ml.) a

  
 <EMI ID = 195.1>

  
 <EMI ID = 196.1>

  
and a solution of N-hydroxysuccinimide (1.91 g.) in dimethylformamide (20 ml.) was added thereto. The mixture was stirred at room temperature for 1.5 hours. The precipitated crystals were collected and washed with dichloromethane.

  
 <EMI ID = 197.1>

  
 <EMI ID = 198.1>

  
at 300 [deg.] C.

  
To a solution of D-alpha-amino-p-hydroxybenzyl-peni-

  
 <EMI ID = 199.1>

  
(3.65 g.) Was added at 1-2 [deg.] C., And the mixture was stirred at 1-5 [deg.] C. for 1.5 hours. The resulting crystals were re-

  
 <EMI ID = 200.1>

  
in ice water. The aqueous solution was adjusted to pH

  
 <EMI ID = 201.1>

  
was collected, washed with water and dissolved in a sufficient volume of 2% sodium hydroxide solution to adjust the pH to 7. The solution was filtered and lyophilized to give the sodium salt of D -alpha- (2-amino-5,8-di-

  
 <EMI ID = 202.1>

  

 <EMI ID = 203.1>


  
 <EMI ID = 204.1>

  
Found: C 43.30 H 4.26 N 15.14 S 4.98

  
 <EMI ID = 205.1>

  

 <EMI ID = 206.1>
 

  

 <EMI ID = 207.1>


  
By the same procedure as that described in Example 2, the sodium salt of D-alpha- (5,8-dihydro-2-

  
 <EMI ID = 208.1>

  
p-hydroxybenzylpenicillin (1.6 g.) was obtained from D-alpha-amino-p-hydroxybenzylpenicillin (2.61 g.) and

  
 <EMI ID = 209.1>

  
6-carbonyloxy) succinimide (1.8 g.).

  
 <EMI ID = 210.1>

  
 <EMI ID = 211.1>

  

 <EMI ID = 212.1>


  
 <EMI ID = 213.1>

  
Reference 15.

  

 <EMI ID = 214.1>
 

  

 <EMI ID = 215.1>


  
By the same operating mode as that described in

  
 <EMI ID = 216.1>

  
zylpenicillin.

  
 <EMI ID = 217.1>

  
 <EMI ID = 218.1>

  

 <EMI ID = 219.1>


  
 <EMI ID = 220.1>

  
Calculated: C 50.28 H 4.80 N 16.17 S 4.63 Found: C 50.19 H 4.61 N 15.91 S 4.66

  
Reference 16.

  

 <EMI ID = 221.1>
 

  
By the same operating mode as that described in

  
 <EMI ID = 222.1>

  

 <EMI ID = 223.1>


  
 <EMI ID = 224.1>

  
Calculated: C 51.27 H 5.45 N 15.95 S 4.56 Found: C 51.92 H 5.15 N 16.08 S 4.38

  
(Part IV.)

  
Examples 17 and 18 show the preparation of medicaments using the penicillin compounds of the invention.

Example 17.

  
(1) In an aseptic environment, sodium salt of the

  
 <EMI ID = 225.1>

  
ne (251 g.) was dissolved in 2.008 ml. distilled water for injection. The solution was filtered through a Millipore filter
(0.22 micron pore size; product of Millipore Corp., Bedford, U.S.A.): An amount of 2.0 ml. solution was poured into 1,000 vials each time (10 ml capacity) and lyophilized. Each of the vials was then sealed with a rubber stopper and an aluminum cap. We thus obtained vials (No. A) each containing
250 mg. of the active ingredient.

  
In ampoules, we introduced quantities of

  
2.0 ml. of physiological saline solution for injections and sealed to obtain ampoules (No. B). The physiological saline in the ampoules (No. B) was poured into the vials (No. A) to obtain an injection for intravenous administration.

  
(2) Into the vials (No. A) were introduced in amounts of 2.0 ml. distilled water for injections, and the solution

  
 <EMI ID = 226.1>

  
tions (250 ml.). Solutions for continuous infusion were thus prepared.

  
(3) One thousand vials (No. C) each containing 500 mg. of

  
 <EMI ID = 227.1>

  
hydroxybenzylpenicillin were prepared in the same manner as described above except that 502 g was used. of the active ingredient.

  
 <EMI ID = 228.1>

  
(1) In an aspetic medium, sodium salt of

  
 <EMI ID = 229.1>

  
ne (251 g.) was dissolved in 3.012 ml. physiological saline solution for injections. The solution was filtered through the same Millipore filter as that used in the example

  
17. Into 1000 borosilicate glass ampoules (5 ml capacity) were poured 3.0 ml amounts. of the resulting solution, and sealed. Each of the ampoules contained
250 mg. of the active ingredient.

  
(2) One thousand ampoules each containing 500 mg. salt

  
 <EMI ID = 230.1>

  
benzylpenicillin were prepared in the same manner as described above except that 502 g was used. of the active ingredient.

  
 <EMI ID = 231.1>

  
The following Examples 19 to 24 illustrate the pharmacological activities of the penicillin compounds of the invention by comparison with those of the compounds not included within the scope of the invention.

  
Compound 1.

  
 <EMI ID = 232.1>

  
p-hydroxybenzylpenicillin.

  

 <EMI ID = 233.1>


  
(the compound of this invention). Compound 2.

  
 <EMI ID = 234.1>

  
hydroxybenzylpenicillin.

  

 <EMI ID = 235.1>


  
(the compound of this invention). Compound A.

  
 <EMI ID = 236.1>

  
p-hydroxybenzylpenicillin.

  

 <EMI ID = 237.1>


  
(the compound obtained according to reference 10):
Compound B.

  
 <EMI ID = 238.1>

  
benzylpenicillin.

  

 <EMI ID = 239.1>


  
 <EMI ID = 240.1>

  
Compound C.

  
 <EMI ID = 241.1>

  
line.

  

 <EMI ID = 242.1>


  
(The compound obtained according to reference 6.)

  
Compound D.

  
 <EMI ID = 243.1>

  
pyrimidine-6-carboxamido) -p-hydroxybenzylpenicillin.

  

 <EMI ID = 244.1>


  
(The compound obtained according to reference 11.)

  
Compound E.

  
 <EMI ID = 245.1>

  

 <EMI ID = 246.1>


  
(The compound obtained according to reference 12.)

  
Compound F.

  
 <EMI ID = 247.1>

  
zylpenicillin.

  

 <EMI ID = 248.1>


  
(The compound obtained according to reference 14.)

  
Compound G.

  
 <EMI ID = 249.1>

  
penicillin.

  

 <EMI ID = 250.1>


  
Compound H.

  
D-alpha- (4-ethyl-2,3-dioxopiperazine -1-carboxamido) -benzylpenicillin sodium salt (Piperacillin).

  

 <EMI ID = 251.1>


  
(The compound described in Belgian patent 828,692.)

  
Compound J.

  
 <EMI ID = 252.1>

  

 <EMI ID = 253.1>


  
(The compound described in the United States patent

  
 <EMI ID = 254.1>
 <EMI ID = 255.1>
 <EMI ID = 256.1>

  
Compound L.

  
 <EMI ID = 257.1>

  

 <EMI ID = 258.1>


  
(The compound obtained according to reference 4.)

  
Compound M.

  
D-alpha-amino-p-hydroxybenzylpenicillin sodium salt (Amoxicillin).

  

 <EMI ID = 259.1>


  
(The compound described in the United States patent

  
 <EMI ID = 260.1>

Example 19.

  
The minimum inhibition concentrations (/ µg / ml) in vitro are shown in Table I.

  

 <EMI ID = 261.1>


  

 <EMI ID = 262.1>
 

  
 <EMI ID = 263.1>

  
1 Cf

  
 <EMI ID = 264.1>

  
1 CP

  
s N t

  
 <EMI ID = 265.1>

  
 <EMI ID = 266.1>

  
1

  

 <EMI ID = 267.1>

I

  
 <EMI ID = 268.1>

  
 <EMI ID = 269.1>

  
 <EMI ID = 270.1>

  
1- '

  
 <EMI ID = 271.1>

  
 <EMI ID = 272.1>

  
 <EMI ID = 273.1>

  
 <EMI ID = 274.1>

  
0 * 0
(B

O

  
 <EMI ID = 275.1>

  
 <EMI ID = 276.1>

  
 <EMI ID = 277.1>

  
 <EMI ID = 278.1>

  
and gold! -

  
 <EMI ID = 279.1>

  
 <EMI ID = 280.1>

  
 <EMI ID = 281.1>

  
 <EMI ID = 282.1>

  
 <EMI ID = 283.1>

S

  
 <EMI ID = 284.1>

  
0 'o'

  
 <EMI ID = 285.1>

  
 <EMI ID = 286.1>

B

  
&#65533; &#65533; ^

  
 <EMI ID = 287.1> Table I enables the following conclusions to be drawn.

I

  
 <EMI ID = 288.1>

  
apart in the process of the invention do not exhibit substantially any anti-bacterial activity.

  
(2) Compound J described in US Pat. No. 3,887,557 has anti-bacterial activity against Gram-positive bacteria, such as Staphylococcus aureus, Streptococcus pyogenes and Corynebacterium pyogenes, less than that of compounds 1 and 2 of the invention.

  
(3) Amoxicillin, a known and commonly used anti-bacterial agent, which is referred to as "Compound M" in ta-

  
 <EMI ID = 289.1>

  
stantially ineffective against pathogenic Gram-negative bacteria such as Escherichia coli P-l40a which is a clinically isolated strain resistant to Ampicillin and Cephalexin and against other Gram-negative bacteria, especially the lower pathogenic bacteria such as Klebsiella pneumonia, Enterobacter cloacae, Pseudomonas aeruginosa and Serratia marcescens, while the penicillin compounds of the invention show very high antibacterial activity against these bacteria.

  
Example 20 (efficacy in vivo).

  
The contents of the vials (No. A and No. C) prepared in Example 17 were separately dissolved in deionized water and injections having different concentrations were prepared.

  
Compounds A, B, F, G, H and M were dissolved in deionized water. Compounds C, D, E and J were dissolved in aqueous sodium carbonate solution (pH 8 - 9) to prepare injections having different concentrations.

  
Each of the injections was administered to mice under the conditions described below, and the mean effective doses (ED50) obtained are shown in the table.

  
 <EMI ID = 290.1>

  
Experimental conditions.

  
Male mice (ddY) mice weighing approximately 20 g. Infection: Staphylococcus aureus No. 50,774.

  
Intravenous infection with 50 to 100

  
 <EMI ID = 291.1>

  
bacterial in a saline.

  
Escherichia coli P-5101.

  
Intra-peritoneal infection with 50 to

  
 <EMI ID = 292.1>

  
bacterial ion in tryptosoy broth - 4% mucin.

  
Pseudomonas aeruginosa No. 12.

  
Intra-peritoneal infection with 50 to

  
 <EMI ID = 293.1>

  
bacterial ion in tryptosoy broth - 4% mucin. Drug; twice, about 5 minutes and 6 hours later

  
the infection.

  
Observation: Staphylococcus aureus No. 50,774 14 days

  
Escherichia coli P-5101) 7 days Pseudomonas aeruginosa No. 12 Table II.

  
Efficacy in vivo against systemic infection.

  

 <EMI ID = 294.1>


  
Note: The numbers in the table indicate the values

  
 <EMI ID = 295.1>

  
 <EMI ID = 296.1>

  
sc: subcutaneous administration. iv: intravenous administration.

  
Table II enables the following conclusions to be drawn.

  
(1) Compounds 1 and 2 of the invention exhibit excellent anti-bacterial activity in vivo.

  
(2) Compounds 1 and 2 of the invention have a particularly high anti-bacterial activity against Pseudomo-

  
 <EMI ID = 297.1>

  
parenteral administration.

  
(3) The anti-bacterial activities against Pseudomonas aeruginosa of compounds 1 and 2 of the invention when administered parenterally are about 3 to 25 times as high as those of compounds B and C which are derivatives of Ampicillin having the same pyridopyrimidinecarboxylic acid group as the compounds of the invention.

  
(4) The anti-bacterial activities against Pseudomonas aeruginosa of compounds 1 and 2 of the invention when administered parenterally are about 6 to 20 times as high as those of compounds D and E which are penicillin compounds resulting from the substitution at position 2 of the pyridopyrimidine part of compounds 1 and 2 by a 1-pyrrolidinyl or dimethylamino group instead of the 4-substituted 1-piperazinyl group.

  
(5) Compounds 1 and 2 of the invention have, for parenteral administration, an anti-bacterial activity vis-à-vis Pseudomonas aeruginosa very markedly greater than that of compound J (pipemidic acid, known as a drug having an activity particularly against Pseudomonas aeruginosa) and of compound H (Piperacillin). Example 21 (acute toxicity).

  
Injections containing compounds 1 and 2 and compounds A to J, in different concentrations, which were prepared in Example 20, were administered to male mice (ddY) (4 to 8 in each group) in a dose of 0.1 ml:
per 10 g. body weight. The number of dead mice has

  
was counted after a period of 7 days and the value of

  
 <EMI ID = 298.1>

  
results are shown in Table III.

  
 <EMI ID = 299.1>

  
nosa n [deg.] 12 in Table II, i.e. a chemotherapeutic index, was calculated and the results are also shown in Table III.

  
Table-III - Acute toxicity in mice.

  

 <EMI ID = 300.1>


  
 <EMI ID = 301.1>

  
by Behrens-Xaerber.

  
: The asterisks! in the table indicate that the calculation was not possible.

  
The following conclusions can be drawn from Table III.

  
 <EMI ID = 302.1>

  
are about 2 to 10 times as high as those of compounds A to G, and, therefore, they are of high safety.

  
(2) The anti-bacterial activities in vivo of compounds A resulting from the introduction of a methyl group instead of R in formula (I), and of compounds F and G resulting from the substitution at position 2 of the group pyridopyrimidine of the penicillin compound of the invention with a methylamino or amino group are substantially comparable to those of the penicillin compounds of the invention, as shown in

  
 <EMI ID = 303.1>

  
to that of compounds 1 and 2 of the invention, but the anti-bacterial activity in vivo of compound H is equal to approximately 1/20 to 1/17 of that of the penicillin compound of the invention, as indicated in table II.

  
 <EMI ID = 304.1>

  
of the invention are about 3 to 100 times as high as those of compounds B, D, E, F, G and J. This shows that the penicillin compounds of the invention have excellent anti-bacterial activity against screw of Pseudomonas aeruginosa. Example 22 (subacute toxicity).

  
The contents of flasks n [deg.] A and C obtained in Example 17 and compound A were dissolved in deionized water. Each of the resulting solutions was administered to ten female mice (ddY) with an average body weight of approximately
20 g., In an amount of 0.2 to 0.4 ml., Once a day for 7 days. The modes of administration and the doses are given below.

  

 <EMI ID = 305.1>


  
During the test period, the body weight of each mouse was determined. On the seventh day, the mice were hematologically examined. After haematological examination, the mice were killed and the weight of their organs was determined. They have also been observed from a histopathological point of view.

  
After these examinations, the following facts were observed.

  
(1) Almost no abnormality was observed in the group treated with compounds 1 and 2 of the invention with regard to body weight gain, hematological examination and histopathological observation. This shows the great safety of the penicillin compounds 1 and 2 of the invention.

  
(2) The group treated with 1000 mg / kg / day (sc) of Compound A showed a significant difference in body weight gain and liver weight compared to the control group treated with saline. Histopathologic observation showed an abnormality of the kidneys in this group.

  
(3) Even the group treated with 250 mg / kg / day (iv) of Compound A had a difference in body weight gain, and in lung, liver, kidney and cecum weight. significant compared to the control group treated with saline. Histopathological observation also showed an abnormality in the lungs and kidneys.

  
(4) For the reasons given in (2) and (3), the compound

  
A. is considered unacceptable as a drug. Example 23 (level in plasma).

  
The contents of the vials (No. A) obtained in the example
17 were dissolved in deionized water to form solutions of compound 1 at a concentration of 100 mg / ml. The solutions were administered intramuscularly to twenty-four male (ddY) mice with an average body weight of 26 g. At a dose of 25 mg / kg. The plasma level (µg / ml) of Compound 1 was determined and the results are shown in Table 17.

  
The other experimental conditions were as follows, .. '

  
Taking samples.

  
Blood was collected by cardiac puncture from 4 mice, each time 0.25, 0.5, 1, 2, 3 and 4 hours after administration, and the blood samples were centrifuged to separate the plasma.

  
Determination.

  
Drug levels were determined by the thin film cup method using Escherichia coli Kp as an indicator organism. (The plasma was suitably diluted - up to more than 10 fold - with a buffering agent consisting of 0.067 M phosphate having a pH of 7.0. A standard run for calibration was prepared in the same buffering agent.

  
Table IV - Rates in the

  
plasma of compound 1 in mice.

  

 <EMI ID = 306.1>


  
 <EMI ID = 307.1>

  
it can be seen that the level in plasma of the penicillin compound 1 of the invention was in general the same as the MIC (/ µg / ml) against the various bacteria shown in Table I. Therefore, from point In view of the level in plasma, the penicillin compounds according to the invention also have excellent properties as anti-bacterial agents.

  
Example 24 (urinary excretion).

  
The contents of the vials (No. A) obtained in the example
17 were dissolved in deionized water to form solutions of the penicillin compound in a concentration of
25 mg / ml. The solutions were administered in a dose of

  
 <EMI ID = 308.1>

  
average body of 25 g. After given time periods, the urine of the four mice was collected and the concentration of penicillin compound 1 in the urine was determined. This experiment was repeated three times, and the averages of the results obtained are shown in Table V.

  
The other experimental conditions were as follows.

  
Sample taking and preparation.

  
Urine was collected using a metabolic cage 3, 6 and 24 hours after administration. The collected urine was diluted with 0.067 M phosphate buffering agent (pH 7.0) prior to determination, up to 500 times for urine collected between 0 and 3 hours, 100 times for urine collected between 3 and 6 hours and 10 times for urine collected between 6 and 24 hours.

  
Determination.

  
Levels of Compound 1 were determined by the thin-film cup method using Escherichia coli Kp as the indicator microorganism and a standard series for calibration was prepared in 0.067 M phosphate buffering agent (pH 7.0 ).

  
Table V.

  
Urinary excretion in mice.

  
(Compound 1: 25 mg / kg im, single dose.)
 <EMI ID = 309.1>
 Total body weight of the 4 mice; c: concentration

  
 <EMI ID = 310.1>

  
 <EMI ID = 311.1>

  
From Table V, it can be seen that even 3 to 6 hours after administration, the level in urine of the penicillin compound 1 of the invention is higher than its

  
 <EMI ID = 312.1>

  
shown in Table I, and the compound 1 of the invention is also very effective for the treatment of diseases caused by infection of the urinary tract by various bacteria.

  
As shown in Tables I to V, the novel penicillin compounds of the invention exhibit a wide range of excellent anti-bacterial activities against various Gram-positive bacteria and Gram-negative bacteria.

  
 <EMI ID = 313.1>

  
More particularly, the penicillin compounds of the invention exhibit excellent anti-bacterial activity,

  
during parenteral administration, vis-à-vis strains resistant to Ampicillin against which Amoxicillin does not exhibit any anti-bacterial activity. In addition, they exhibit excellent anti-bacterial activity against bacteria of the Pseudomonas type against which Amoxicillin and several other known antibiotics are substantially ineffective. Among the different compounds

  
of penicillin of the invention, the formyl derivative of formula (Ia), particularly its D-isomer, have especially excellent anti-bacterial activity.

  
The penicillin compounds of the invention exhibit

  
in addition a very high security as demonstrated by the values

  
 <EMI ID = 314.1>

  
therapies several tens of times higher than those of known antibiotics, such as known penicillin derivatives having a structure similar to that of the compound of the invention. This great safety is also proven by haematological examination and histopathological examination of laboratory animals after continuous parenteral administration.

  
The penicillin compounds of the invention further exhibit a high level in plasma even several hours after parenteral administration to warm-blooded animals, and maintain a high level in urine for very long periods of time.

  
The new penicillin compounds chosen according to the invention have characteristics very clearly superior to those of the various known anti-bacterial agents, because they have a broad anti-bacterial spectrum, high antibacterial activity, high safety, high levels. high in blood and urine after administration, etc.

  
The authors of the present invention have found that in the use of the penicillin compounds of the invention for warm blooded animals, including humans, parenteral administration is far more advantageous than administration by oral route. For parenteral administration, it is preferable to dissolve the penicillin compound of the invention in a non-toxic liquid medium for injections, and to inject the solution intramuscularly, intravenously or subcutaneously.

  
It is also possible to dissolve, or mix, the penicillin compound of the invention in, or with, a non-toxic liquid medium or an ointment base and apply the solution or mixture directly to the lesion. The compound can also be used as a suppository after mixing or dissolving with, or in, a suppository base.

  
Examples of a non-toxic liquid medium used for the preparation of injections containing the penicillin compound as an active ingredient include sterilized deionized water, physiological saline solution, glucose solution, for injection, Ringer's solution and solution in an amino acid for injection.

  
The penicillin compounds can also be dissolved in other injections and be administered parenterally as such.

  
The dose of the penicillin compound of the invention for administration to humans should be determined according to the age, body weight, and condition of the patient, the manner in which


    

Claims (1)

administration, number of administrations, etc. Usually the dose for adults is 0.1 to 10 g / day, preferably 0.2 to 4 g / day. As already mentioned, from the point in view of their pharmacological properties, the penicillin compounds of the invention are preferably administered parenterally (for example intravenously, intramuscularly or subcutaneous) in the form of an injection dissolved in a pharmaceutically acceptable liquid medium. Preparations for injections containing the penicillin compounds of the invention are, for example, an injection kit consisting of a vial filled with the penicillin compound as the active ingredient and an ampoule containing an aqueous liquid medium capable of dissolving the active ingredient for forming an injection, or an injection prepared by dissolving the active ingredient in a liquid aqueous medium. Liquid aqueous media are those commonly used in preparations of penicillin or cephalosporin for injection, for example, sterilized deionized water containing a known pH adjuster and an osmotic pressure adjuster and, if desired, a stabilizer, If desired, these preparations may additionally contain other ingredients depending on the purpose of the medication. CLAIMS. 1. Compound of penicillin characterized in that it is represented by the formula: <EMI ID = 315.1> (in which R is a formyl or acetyl group), and a pharmaceutically acceptable non-toxic salt thereof. 2. Compound of penicillin characterized in that it is represented by the formula: <EMI ID = 316.1> and a pharmaceutically acceptable non-toxic salt thereof. 3. Compound characterized in that it is D-alpha- <EMI ID = 317.1> pyrimidine-6-carboxamido_7-p-hydroxybenzylpenicillin and a pharmaceutically acceptable non-toxic salt thereof. 4. A compound according to claim 3, characterized in that it is the sodium or potassium salt of D-alpha- <EMI ID = 318.1> rimidine-6-carboxamido-7-p-hydroxybenzylpenicillin. 5. Penicillin compound characterized in that it is represented by the formula: <EMI ID = 319.1> and a pharmaceutically acceptable non-toxic salt thereof. 6. Compound characterized in that it is D-alpha- <EMI ID = 320.1> pyrimidine-6-carboxamido_7-p-hydroxybenzylpenicillin and a pharmaceutically acceptable non-toxic salt thereof. 7. A compound according to claim 6, characterized in that it is the sodium or potassium salt of D-alpha- <EMI ID = 321.1> takes a solution, in a non-toxic liquid medium, of a penicillin compound of the formula: <EMI ID = 322.1> (wherein R is a formyl or acetyl group), or a pharmaceutically acceptable non-toxic salt thereof. <EMI ID = 323.1> characterized in that it comprises a solution, in a non-toxic liquid medium, of a pharmaceutically non-toxic salt <EMI ID = 324.1> xybenzylpenicillin. <EMI ID = 325.1> characterized in that it comprises a solution, in a non-toxic liquid medium, of a pharmaceutically non-toxic salt <EMI ID = 326.1> xybenzylpenicillin. 11. Preparation for injections characterized in that it consists of a vial containing the compound of penicillin or its non-toxic pharmaceutically acceptable salt according to claim 1 and an ampoule containing an injectable non-toxic liquid medium capable of dissolving the compound of penicillin. or its pharmaceutically acceptable non-toxic salt. 12. Preparation for injection according to claim 11, characterized in that the liquid medium contains at least one pharmaceutically acceptable adjuvant. 13. Use of a penicillin compound of the formula: <EMI ID = 327.1> (wherein R is a formyl or acetyl group), and one of non-toxic pharmaceutically acceptable salts as an anti-bacterial agent. 14. Use of a penicillin compound of the formula: <EMI ID = 328.1> and one of its pharmaceutically acceptable non-toxic salts, as an anti-bacterial agent. 15. Use of a penicillin compound of the formula: <EMI ID = 329.1> and one of its pharmaceutically acceptable non-toxic salts, as an anti-bacterial agent. 16. A method for anti-bacterial treatment characterized in that it comprises parenteral administration to a warm-blooded animal of an effective dose of the compound according to claim 1. 17. A process for preparing a penicillin compound of the formula: <EMI ID = 330.1> (in which R is a formyl or acetyl group), or one of its non-toxic pharmaceutically acceptable salts, characterized in that: (A) A compound of formula <EMI ID = 331.1> (where R is as defined above), or its salt or derivative reactive to the carboxyl group, with a 6-alpha-amino-p-hydroxybenzylpenicillin of the formula <EMI ID = 332.1> or its salt or its derivative. (B) Optionally, the reaction product is hydrolyzed or catalytically hydrogenated, and (C) optionally, the reaction product is converted to a pharmaceutically acceptable non-toxic salt. 18. A process for preparing a penicillin compound of the formula <EMI ID = 333.1> (in which R is a formyl group or: acetyl), or a pharmaceutically acceptable non-toxic salt thereof; characterized in that: (A) a compound of formula is reacted: <EMI ID = 334.1> (where R is as defined above), or its salt or derivative reactive to the carboxyl group, with 6-aminopenicillanic acid, or its salt or derivative, (B) optionally, the reaction product is hydrolyzed or catalytically hydrogenated, and (C) optionally, the reaction product is converted to a pharmaceutically acceptable non-toxic salt. <EMI ID = 335.1> terized in that the carboxyl-reactive derivative of the carboxylic acid of formula (II) or (V) is an acid halide, an acid anhydride, an amide or an active ester. 20. The method of claim 17, characterized in that 6-D-alpha-amino-p-hydroxybenzylpenicillin is used as 6-amino-p-hydroxybenzylpenicillin. 21. The method of claim 17 or 2o, charac- <EMI ID = 336.1> mido_7-p-hydroxybenzylpenicillin, or a pharmaceutically acceptable non-toxic salt thereof. 22. The method of claim 17 or 20, character- <EMI ID = 337.1> p-hydroxybenzylpenicillin or a pharmaceutically acceptable non-toxic salt thereof.