CA1103236A - 3-acyloxymethyl-cephem compounds - Google Patents

Abstract

ABSTRACT

An improved process for preparing cephem compounds of the formula:

[I]

, wherein R1 represents hydrogen or an acyl group and R2 stands for a substi-tuted or unsubstituted pyridinium or a residue of a nitrogen containing hetero-cyclic thiol, or a salt thereof, which comprises reacting a compound of the formula;

[II]

, wherein W represents acetonyl or a group represented by -X-COOH or -X-OH
(in which X is a divalent carbon chain which is able to form a five- or six-membered ring with or

Description

rhe presellt lnvention relates to a mcthod Por producing a compound of the formula:

R Nll ~ S ~ [I]

-- ~ N ~ CH2R
COOII

or a pharmaceutically-acceptable salt thereof, wherein Rl represents hydrogeD
or a suitable acyl group and R stands for a pyridinium group optionally sub-,_ .
stituted by me~hyl, carboxyl, carbamoyl, carba~oyl, bromo, sulfo, hydroxymethyl or formyl, or stands for a residue of a nitrogen-containing heterocyclic thiol, which comprises reacting a compound of the formula:

R NH ~ II]

COOH

or a salt thereof, wherein W represents acetonyl or a group represented by -X-COOH or -X-OH (in which X is a divalent carbon chain which is able to form a five- or six-membered ring with -C-O-C- or -C-O-C-O-, which may include a double bond or such an atom or atoms as oxygen, nitrogen or sulfur therein, and on which carbon chain suitable substituent or substituents may be attached) with pyridine or a pyridine substituted as defined for pyridinium above, or with a nitrogen-containing heterocyclic thiol.
Heretofore, it was known that some of the compounds [I] or salts thereof could be produced by reacting a compound of the formula:

RlNH -- .. .. ,~ S
N ~ CH20COCH3 [III]

COOH

. ~i -1- ~,.

.
. ' ` . :
, or a salt thereof wi-th a nuc1eop]lillc cornpo~md.
The present inventors have found that the compound [~ or a salt thereof is produced by the use oE the new compound [IT] or a salt thereof in better yield or/an~ in shorter period reaction time.
The symbol Rl includes hydrogen, phenylacetyl, phenoxyacetyl, 5-amino-5-carboxyvaleryl ~hoseamino or/and carboxyl groups may optionally 6e protected, or any of the groups Eound in the 6- or 7-positions of penicil-lin or cephalosporin derivatives as the case may be. Thus, ~or example, the acyl group ~ may be selected from among aliphatic acyl groups such as Eormyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, cyclopentylcar-bonyl, cyclohexylcarbonyl, cycloheptylcarbonyl, cyclopentylace~yl, cyclohexa-dienylacetyl, etc.; aromatic acyl groups such as benzoyl, p-nitrobenzoyl, toluoyl, naphthoyl, etc.; mono-su~stituted aliphatic acyl groups such as 2-thienylacetyl, cyanoacetyl, acetoacetyl, 4-chloro-3-oxobutyryl, 4-bromo-3-oxo6utyryl, chloroacetyl, bromoacetyl, 4-methylthio-3-oxobutyryl, 4-carbamoyl--methylthio-3-oxobutyryl, ~-phenoxypropionyl, ~-phenoxybutyryl, tetrazolylthio-acetyl, tetrazolylacetyl, p-nitrophenylacetyl, trifluoromethylthioacetyl, trifluoromethylsulfinylacetyl, trifluoromethylsulfonylacetyl, cyanomethylthio-acetyl, thiadiazolylthioacetyl, p-nitrophenylacetyl, (2-pyridyloxy)acetyl, ~2-oxo-4-~hiazolin-4-yl)acetyl, (2-imino-4-thiazolin-4-yl)acetyl, (2-thioxo~4 thiazolin-4-yl)acetyl, 4-pyridylthioacetyl,~(3-sydnone)acetyl, l-pyrazolyl-acetyl, 2-furylacetyl, (2-oxo-3-methylpyridazinyl)thioacetyl, (2-aminomethyl-phenyl)acetyl, ~2-aminomethylcyclohexenyl)acetyl, etc.; di-substituted aliphatic acyl groups such as ~-carboxyphenylacetyl, mandelyl, ~-sulfophenyl-acetyl, ~-sulfo-Cp-aminophenyl)acetyl, phenylglycyl, ~4-hydroxyphenyl)glycyl, (4-metXy~lthiophenyl)glycyl, ~4-methoxyphenyl)glycyl, ~4~methanesulfin~1-pheny~l~glycyl, ~3-methanesulfonamidophenyl)glycyl, l-cyclohexenylglycyl, thi-enylglycyl, furylglycyl, cyclohexadienylglycyl, ~3,4-dihydroxyphenyl)glycyl,

2 -' ' ` ' ' . , ' " ' ' ' etc.; 5-methyl-3-phenyl-~-isoxazolylcarbonyl; 3-~2,6-dichlorophenyl~-5-methyl-~-isoxazolylcarbonyl; and so forth. It should be understood that the above-mentioned groups are only ;llustrative of the acyl groups that are of use for the purposes of this invention, but prefera~le acyl groups may be represented ~y the formula:

R~\
CHCO-wherein R4 stands for acetyl, halogenoacetyl, phenyl, p-hydroxyphenyl, thienyl, 2-imino-4-thiazolin-4-yl, 2-oxo-4-thiazolin-4-yl, tetrazolyl, phenoxy, 3-amino-

3-carboxypropyl, etc. and R5 stands for hydrogen, sulfo, amino, hydroxy, etc.
It should also be understood that any functional groups, e.g. amino or/and carboxyl, in such acyl groups may be suitably protected. Thus, among protective groups for said amino groups are aromatic acyl groups such as phthaloyl, benzoyl, p-nitrobenzoyl, toluoyl, naphthoyl, p-tert-butylbenzoyl, p-tert-butylbenzenesulfonyl, phenylacetyl, benzenesulfonyl, phenoxyacetyl, toluenesulfonyl, chlorobenzoyl, etc.; aliphatic acyl groups such as acetyl, valeryl, capryl, n-decanoyl, acryloyl, pivaloyl, camphorsulfonyl, methanesul-fonyl, chloroa~etyl, etc.; esterified carboxyl groups such as tert-butoxycar-bonyl, ethoxycarbonyl, isobornyloxycarbonyl, phenyloxycarbonyl, trichloro-ethoxycarbonyl, benzyloxycarbonyl, ~-methylsulfonylethoxycarbonyl, etc.;
carbamoyl groups such as methylcarbamoyl, phenylcarbamoyl, naphthylcarbamoyl, etc.; the corresponding thiocarbamoyl groups; 2-methoxycarbonyl~l-methyl-vinyl; and so forth. As protective groups for the carboxyl groups of said acyls Rl and the 4-carboxyl group of the cephem ring, there may be mentioned methyl, ethyl, tert-butyl, tert-amyl, benzyl, p-nitrobenzyl, p-methoxybenzyl, benzhydryl, l-indanyl, phenacyl, phenyl, p-nitrophenyl, methoxymethyl, ethoxymethyl, benzyloxymethyl,acetoxymethyl, pivaloyloxymethyl, ~-methyl-.. . .
. .
. .

, .
, ~' '' ' ' , sulfonylethyl, methylthiomethyl, trityl, ~ trichloroethyl, silyl groups such as trimethylsilyl, dimethylsilyl, etc.; and so forth. These carboxyl groups may also be in the form oE inorganic or organic salts with alkali metals such as lithium, sodium, potassium etc.; alkaline earth metals such as calcium, magnesium; or various amines such as dicyclohexylamine, triethylamine, tri~utylamine, di-n-butylamine, di-n-propylamine and so orth.

The divalent organic residue denoted by X is usually a carbon chain which is able to form a five- or slx-membered ring with -C-0-C- or -C-0-C-0-and which may include a double bond or such an atom or atoms as oxygen, nitrogen or sulfur therein, preferably oxygen. Moreover, the carbon chain may be such that its carbon atoms carry suitable substituent groups. Among examples of such substituents on the carbon chain, there may be mentioned carboxyl, halogen, nitro, alkyl (e.g. methyl, ethyl, propyl, methylene, ethylene), aralkyl ~e.g. benzyl, phenethyl, etc.), aryl (e.g. phenyl, tolyl, etc.), and hydroxy or mercapto groups substituted thereby ~e.g. methoxy, p-chlorophenylthio, etc.). Where two or more such substituents are present, they may form a ring with the carbon chain.
According to the present invention, a substituted or unsubstituted pyridine or a nitrogen containing heterocyclic thiol, employed as a nucleo-philic, is reacted with the compound [II~ or a salt thereof.
As to the nucleophilic compound used for this reaction, any of thecompounds that are able to replace the 3-acetoxy groups of cephalosporins may be used as the nucleophilic compound. The reaction proceeds at a rate rang-ing from 4 to 16 times that of the 3-acetoxy compound and almost quantitative-The nitrogen-containing heterocyclic thiols usually contain one or more nitrogen atoms which may optionally be in the form of oxide or/and which contain such atoms as oxygen or/and sulfur in addition to the nitrogen atom, :, : ' : : ,. - ~ :. .
:. . , . ' :
. .

with or without nuclear substitution. As the common examples of the nitrogen-containing heterocyclic group ot` such a thioL compound, there may be men-tioned pyridyl, N-oxidepyridyl, pyrimidyl, pyridazinyl, N-oxidepyridazinyl, pyrazolyl, diazolyl, thiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,3,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,2,3-oxad:iazoyl, 1,2,4-oxadiazolyl, 1,3,~-oxadiazolyl, 1,2,5-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, lH-~etra-zolyl, 2H-tetrazolyl and so forth. As the subs~ituents on such nitrogen-containing heterocyclic groups, there may be mentioned such monovalent groups as hydroxy~ mercapto, amino, carboxyl, carbamoyl, lower alkyl (for example, methyl, ethyl, trifluoromethyl, propyl, isopropyl, butyl, isobutyl, etc.~, lower alkoxy (for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, etc.), halogens ~e.g. chlorine, bromine, etc.), and various substituent groups as attached through lower alkylene groups, -S-, -N- or other polyvalent groups.
~hen such polyvalent groups are lower alkylene groups, the substituents may for example be hydroxy, mercapto, amino, morpholino, carboxyl, sulfo, carba-moyl, alkoxycarbonyl, lower alkylcarbamoyl, alkoxy, alkyl~hio, alkylsulfonyl, acyloxy, morpholinocarbonyl and so forth. When such a polyvalent group is -S- or -N-, the substituents may be lower alkyls or lower alkylene groups having the aforementioned substituents. When the polyvalent group is -N-, such substituents as carboxyl, alkoxycarbonyl, acyl, carbamoyl, lower alkyl-carbamoyl, etc. may be directly attached. More particularly, there may be mentioned substituted alkyl groups such as carboxymethyl, carbamoylmethyl, N-lower alkylcarbamoylmethyl ~e.g. N,N-dimethylcarbamoylmethyl), hydroxy lower alkyl ~e.g. hydroxymethyl, 2-hydroxyethyl), acyloxy lower alkyl (e.g.
acetoxymethyl, 2-acetoxyethyl), alkoxycarbonylmethyl (e.g. methoxycarbonyl-methyl, hexyloxycarbonylmethyl, octyloxycarbonylmethyl), methylthiomethyl, meth~lsulfonylmethyl, N-lower alkylamino lower alkyl (e.g. N,N-diemthylamino-meth~l, N~N-diemthylaminoethyl, N,N,N-trimethylamminoniumethyl), morpholino-~.~, .
' ' , ù ~

methyl, etc.; su~st-ituted al~ino groups such as lower alkylamino (e.g. methyl-c~minol, sul~o-lower alkylamino ~e.g. 2-sulfoethylamino), hydroxy-lower alkyl-amino Ce.g. hydroxyethylamino), lower alkylamino-lower alkylamino (e.g. 2-dimsthylaminoethylamino, 2-trimethylamminoniumethylamino), acylamino (e.g.
acetylamino), 2-dimethylaminoacetylamino, 2-trimethylammoniumacetylamino, lower alkoxycarbonylamino ~e.g. methoxycar~onylamino), etc.; and substituted thio ~mercapto~ groups such as methylthio, 2-~hydroxyethylthio, 2-acyloxy-ethylthio Ce.g. 2-acetoxyethylthio, 2-phenylacetoxyethylthio, 2-caproyloxy-ethylthio), carboxymethylthio, alkoxycarbonylmethylthio ~e.g. methoxycar-bonylthio, hexyloxycarbonylmethylthio), carbamoylmethylthio, N-lower alkyl-carbamoylmethylthio ~e.g. N,N-dimethylcarbamoylmethylthio), acetylmethylthio, N-lower alkylamino-lower alkylthio (e.g. 2-N,N-dimethylaminoethylthio, 2-N,N, N^trimethylammoniumethylthio), morpholinocarbonylmethylthio, 2-sulfoethylthio and so forth. More particularly, there may be mentioned various heterocyclic thiols such as tetrazolethiol, methyltetrazolethiol, phenyltetrazolethiol, C2-N,N-dimethylaminoethyl)tetrazolethiol, methylthiadiazolethiol, hydroxy-ethylthiothiadiazolethiol, methylthiothiadiazolethiol, thiadiazolethiol, car-bamoylaminothiadiazolethiol, carbamoylmethylthiothiadiazolethiol, thiazolethi-cl, methylthiazolethiol, carboxymethylthiazolethiol, triazolethiol, dimethyl-2a triazole~hiol, pyrazolethiol, ethoxycarbonylmethyltriazolethiol, imidazolethi-ol, methyloxidiazolethiol, pyridinethiol, pyrimidinethiol, methylpyridazinethi-ol, triazinethiol and so forth. In addition, use may also be made of such ni~trogen-containing heterocyclic compounds as aliphatic or aromatic thiols, e.g. methanethiol, ethanethiol, thiophenol, etc.; thiourea and its derivatives such as N-methy~lthiourea, N-meth~l-N'~pyridylthiourea, etc~;~thioamide deriva-tives such as thiosemicarbazide, thioacetamide, thiobenzamide, etc.; sodium thiosulfate, sodium sul~ite, potassium thiocyanate, sodium azide, etc.;
pyridine and pyridine derivatives such as quinoline, picoline, nicotinic acid, '' ' ' : ~ ' ' nicotinamide, isonicotinamide, isonicotlnlc acid hydra~ide, o-broMopyridine, pyridinesulfonic acid, pyridine-m-carbinol ~3-hydroxymethylpyridine), pyridin-aldehyde, quinoline, isoquinoline, e~c.; and such other nitrogen-containing heterocyclic compounds as pyrazine, pyrazinamide (2-carbamoylpyrazine), pyridazine, pyrimidine, imidazole, l-methylimidazole, pyrazol and so forth.
It is also possible to employ carb~n nucleophilic agents toward whlch the 3-position is known to be refractory. As examples of such carbon nucleophilic reagents, there may be mentioned cyanides, p~rrole, substituted pyrrole, indole, acetylene, active methylene compounds, e.g. acetylacetone, aceto-acetic acid esters, malonic acid esters, cyclohexane-1,3-dione, triacetyl-methane and enamine compounds. Alcohols such as methanol, ethanol, propanol, etc. may also be employed in this reaction.
The substitution reaction between such a substituted or unsubstituted pyridine or a nitrogen containing heterocyclic thiol and a compound of formula ~II] is normally conducted in a solvent. While the commonest solvent is water, the hydrophilic organic solvents inert to the reaction, e.g. acetone, tetra-hydrofuran, dimethylformamide, methanol, ethanol, dimethylsulfoxide, etc., and aqueous solvents such as mixtures of water and such reaction-inert, polar solvents as mentioned above are employed with preference.
2Q ~hile compound [II] may be a free compound, it is more advantageous to subject compound [II] to the reaction in the form of an alkali metal salt, e.g. the sodium, potassium or other salt, or an organic amine salt, e.g. the triethylamine, trimethylamine or other amine salt. The nucleophilic compound is also reacted in its free form or as an alkali metal, organic amine or other salt.
The proportion of a substituted or unsubstituted pyridine or a n~trogen containing heterocyclic thiol to be used in this reaction is prefer-ably~ one equivalent or more with respect to compound ~ . While the optimum !

- - 7 .

3~

p~l depends upon the particular nucleophilic compound and compound ~II), the reaction is generall~ carried out prefera~ly under weakly acid to weakly alkaline conditions. The reaction temperature is pre-ferably somewhere ~e-tveen 40C and 70C, although there are no particular limits. Th~ reaction time cannot be specified in general terms, either, for it depends upon the reaction temperature, pH, type of nucleophllic reagent and other factors.
Roughly speaking, however, the reaction goes to completion in ~0 minutes to 2 hours when the reaction temperature is 60C. The reaction may also be carried out in the presence of, as added to the reaction system, an inorganic salt, e.g. the chloride, bromide, iodide, thiocyanide or nitrate of lithium, sodium, potassium, ammonium or the like.
After the above reaction, the end product ~I~ or a salt thereof (prefera~ly a pharmaceutically accepta~le salt thereof) is recovered from the reaction mixture ~y a conventional method.
Of those starting compounds CII~, the compounds in which W is acetonyl can ~e produced by reacting a compound of the general formula:

R NH ~ S ~ [IV]
F- cH2oH
COOH

~herein Rl is hydrogen or an acyl group) with diketene. The diketene may as ~!
~ell ~e used in the form of an adduct with acetone, acetophenone or the like.
This is an equimolar reaction, which means that the desired reaction can be accomplished by using the compound [IV] and diketene in equimolar proportions.
However, to adjust for the possible decomposition of diketene which would take place in the presence of water or alcohol, an excess of diketene may be em-ployed, Normally~this reaction can be successfully conducted by allowing a ~, , , "' '~' ' ~- ' ' ' ' , 3~

3-hydroxymethylcephalosporin [IVl and di]cetene to interact in a suitable in-ert sol~ent at a temperature oF -30C to 40(:. As said suitable inert solvent, there may be employed any of such solvents as dichloromethane, chloroform, di-chloroe~hane, dimethylformamide, dimethylacetamide, tetrahydrofuran, aceto-nitrile, ethyl acetate, acetone, dioxane, ether, etc. and ~arious mixtures of such solvents. The reaction of diketene with 3-hydroxymethylcephalosporin rIV] proceeds at a high rate but in consideration of the fact that the reac-tion rate depends somewhat on the reaction temperature and to ensure that the reaction will be carried to completion, this reaction is usually conducted for la a period of 0.5 to 15 hours. If necessary, an amine such as triethylamine may be added to the reaction system, or where the startlng material [IV] is an alkali metal salt, an equivalent of, for example, triethylamine hydrochloride may be added to effect a salt interchange before the desired reaction takes place.
Where W stands for -X-COOH, the compound [II] can be produced by reacting a 3-hydroxymethylcephalosporin [IV] with compound of the general formula:

\ [V]

~O
~0 ~where X has the same meaning as defined hereinbefore).
2a As specific examples of compound [V], there may be mentioned maleic anhydride, succinic anhydride, phthalic anhydride, glutaric anhydride, diglycolic anhydride, thiodiglycolic anhydride, p-chlorophenylsuccinic an-hydride, methylenesuccinic anhydride, 3-nitrophthalic anhydride, trimellitic anhydride, isatoic anhydride.

.~; - 9 _ - , Generally speaking, thc reaction of co7npound [IV] with compound [V~
is expediently conducted in the presence of a suita~le inert solvent like that used in the reaction described just above. The stoichiometric proportions of reactants, reaction temperature and other conditions of reaction may also be similar to those used for the reaction described.
The compound [II] wherein W is -X-0ll may be produced by reacting a 3-hydroxymethylcephalosporin [IV] with a compound of the general formula:

X 0 [VI]

~0 ~wherein X has the same meaning as defined hereinbefore).
Thus, as examples of compound [VI], there may be mentioned O-car-boxymandelic anhydride, 0-carboxy-~-hydroxypropionic anhydride, 0-carboxy-~-hydroxypropionic anhydride, 0-carboxy-3-methylsalicylic anhydride, 0-carboxy-~-hydroxy-~-phenyl~propionic anhydride, 0-carboxy-~-hydroxy-~-phenyl)pro-pionic anhydride and so forth.
The conditions of reaction between compound [IV] and compound [VI]
are similar to those used for the reactions previously described. Where the starting materical [IV] has an unprotected amino group, the reaction thereof with diketene, compound [V] or compound [VI] may be conducted so that both the 3-hydroxy and the amino group may be simultaneously acylated by the same ZU acyl groups.
The compound [II] wherein Rl is an acyl group can be converted to the corresponding compound wherein Rl is hydrogen by cleaving the 7-acyl group off in a manner conventional per se Ce.g. any of the procedures set forth in Japanese Patent Publication No. 13862/1966 and No. 40899/1970, Japanese Patent ", ~` 10 --~, ~3~3~

App1ication Laicl Open No. 343~7/1972, No. 95292/1975 and No. 96591/1975, Japanese Patent Publication No. 35079/1975, Unitecl States Patent No. 3632578, etc.) Into this compound may be introduced a compound which has previously been mentioned by way o:F example for R1 as the 6- or 7-substituents of penicil-lin or cephalosporin compounds, by previously activating the same in a known manner.
The compounds [IV] include 7-~D-5-amino-5-carboxyvaleramido)-3-hydroxymethyl-3-cephem-4-carboxylic acid which is produced in high titer by fermentation ~United States Patent No. 3,926,726, Nature New Biolody, 246, 154 ~1973))-The present invention is illustrated in further detail below with ref-erence to examples, but it is to be understood that the examples are solely for the purpose of illustration and not to be construed as limitations of the invention, and that many variations may be resorted to without departing from the spirit and scope of the invention. Those examples indicated as reference examples do not illustrate the preparation of compounds in accordance with the invention claimed herein, but are included for the better understanding of the invention. In this specification, "g", "mg", "m~", "cm", Hz", "DMSO", and "decomp." are abbreviations of "gram", "milligram", "milliliter", '7centimeter", "Herz", "dimethylsulfoxide", and "decomposed", respectively. Resins sold under the trade marks "Amberlite" are products manufactured by Rohm & Haas Co. in U.S.A. The products sold under the trade marks "Celite" and "Cephadex"
are marketed by Johns-Manville Sales Corp. and Pharmacia A.B., respectively.
All the temperatures are uncorrected and the percentages are all on the weight basis except specifically defined. The NMR spectra given therein were meas-ured using a Varian Model ~ 100 ~100M~1z) or T60 ~60 M~1z) spectrometer with tetramethylsilane as the internal reference and all ~ values are in ppm.
The symbol s stands for a singlet, d a doublet, t a trip-'' -;1 ,.~,, .:

'~
:- ' ,, ~' :

3~i let, q a quarte~, m a multiplet, and J a coupling constant. rhe pfl adjustment o~ the reaction solution, when needed, is made by adding a diluted aqueous so-lution of sodium hydrogen carbonate or diluted aqueous hydrochloric acid.
Example 1 ~1) In dichloromethane ~50 mQ) was dissolved 7~-~D-5^phthalimido-5-car-boxyvaleramido~-3-hydroxymethyl-3-cephem-4-c2rboxylic acid ditriethylamine salt (7.05 g), followed by ~he addition of diketene ~0.92 g). The reaction was carried out at roo~ temperature for 3 hours. After the reaction had been completed, the solvent was distilled off under reduced pressure and the resi-lQ due was diluted with water and adjusted to pH 6.0 with sodium hydrogen car-bonate. The aqueous solution was washed twice with ethyl acetate, brought do~n to pH 2.0 with 4N-HCQ and extracted three times with ethyl acetate. The ethyl acetate solution was washed with a saturated aqueous solution of sodium chloride, treated with magnesium sulfate and filtered. The solvent was then distilled off under reduced pressure. To the residue was added ether and the resultant powder was collected by filtration, washed with ether and dried.
~e above procedure provided 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (5.48 g) ~yield 93.2%).
IR~KBr~: 3350, 1775, 1740, 1715, 1640, 1530 cm 1 2a NMR~ in d6-DMS0): 1.30 - 2.40~6H,m), 2.17~3H,s), 3.46(2H,ABq,J=18Hz),3.60~2H,s), 4.72~lH,t), 4.90~2H,ABq,J=12Hzl, 5.01~1H,d,J=5Hz), 5.62~1H,dd,J=5 ~ 8Hz), 7.88~4H,s), 8.80~1H,d,J=8Hz) ~2~ ln water (50 mQ) was dissolved 7~-~D-5-phthalimido-5-carboxyvaler-amido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (5.88 g), together with 5-mercapto-1-methyl-lH-tetrazole ~1.50 g) and sodium hydrogen carbonate (2.10 g). Following the addition of sodium chloride ~15.0 g), the solution was adjusted to pH 5.0 and, then, reacted at 60C for 50 minutes. After cool-~'. t.J

"~ ' ~'' , '' ' ' 3~

ing, a saturated aqueous solut-ion Oe socli~ml chlorlde ~!;0 m~) was added and the mixture was adjusted to pll 1.5 with 4N-HC~. The re~ultant solid precipitate was recovered by filtration, washed w:ith a saturated aqueous solution of sodi-um chloride (20 m~) and dissolved in ethyl acetate (100 mQ)-water ~20 mQ).
After separation, the ethyl acetate solu~ion was dried and, following the ad-dition of toluene (50 m~), concentrated. The solid precipitate was recovered by filtration, washed with toluene-ether and dried. The procedure provided 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-(1-methyl-lH-tetrazol-5-yl~thio-methyl-3-cephem-4-carboxylic acid ~5.75 g) (yield 95.5 %).
IR~KBr): 3325, 1780, 1730, 1715, 1650, 1545cm NMR(~ in d6-DMS0): 1.~0 - 1.76~2H,m), 2.0 - 2.4(4H,m), 3.6~(2H,ABq,~=19Hz), 3.93~3H,s), 4.30~2H,ABq,~=15Hz), 4.73~1H,t,J=8Hz), 5.01~1H,d,J=5Hz), 5.62~1H,dd,J=5 ~ 9Hz), 7.85(4H,s), 8.80(d,J=9Hz) ~ 3) In water ~60 mQ) was dissolved 7~-~D-5-phthalimido-S-carboxyvaler-amido~-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~5.88 g) together with 2-(2-hydroxyethylthio)-5-mercapto-1,334-thiadiazole (2.90 g) and sodium bicarbonate ~2.20 g). Following the addition of sodium bromide (30 g), the solution was adjusted to pH 5.5 and reacted at 60C for 50 minutes. After the reaction was completed, the reaction mixture was diluted with water (40 mQ) and adjusted to pH 5Ø The aqueous solution was washed twice with ethyl acetate3 brought down to pH 2.0 with 4N-HCQ and extracted three times with a mixture of ethyl acetate and tetrahydrofuran ~4:1). The organic layer was w~ashed with a saturated aqueous solution of sodium chloride, dried over mag-nesium sulfate, filtered and distilled under reduced pressure to remove the solvent. To the residue was added ethyl acetate-ether and the resultant powder was recovered by -filtration, washed with ether and dried. ~ne procedure pro-vided 7~-CD-5-phthalimido-5-car~oxyvaleramido)-3-c2-hydroxyeth~lthio)-1,3,4-13 ~

., ".,., ~, :

thiadiazol-5-yl)thiomethyl-3-cephem-4-carhoxylic acid (6.56 g) (yleld 96.6 %) IR~KBr): 3325, 1780, 1715, 1645, 1530cm MMR(d6-DMSO): ~ 1.30 - 2.40(6H,m), 3.20 - 3.80(6H,m), 4.27(2H,ABq,J~12Hz),

4.65(1H,t,J=9Hz), 4.96~1H,d,J~5Hz), 5.55(1H,dd,J=5 ~7 8Hz), 7.87~4H,s), 8.70(lH;d,J=8Hz) (4) In a mixture of water (50 mQ) and tetrahydro~uran (30 mQ~ was dis-solved 7~-~D-5-phthalimido-5-carboxyval0ramiclo)-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (5.88 g) together with 2-carbamoylmethylthio-5-mercapto-1,3,4-thiadiazole(2.28 g) and sodium bicarbonate (2.20 g). The solu-tion was adjusted to pH 5.8 and, then, reacted at 60C for 70 minutes. A~ter cooling, water (30 mQ) was added and the aqueous solution was adjusted to pH

5.0, washed twice with ethyl acetate and brought down to pl-l 2.0 with 4N-HCQ.
It was then extracted three times with a solvent mixture o~ ethyl acetate and tetrahydrofuran ~2:1) and the organic layer was washed with a saturated aqueous solution o~ sodium chloride and dried over magnesium sul~ate. The solvent was d~st~lled o~ under reduced pressure and the residue was treated with ethyl acetate. The resultant powder was recovered by ~iltration, washed with ethyl acetate and dried. The procedure provided 7e-~D-5-phthalimido-5-carboxyvaler-amido)-3-~2-carbamoylmethylthio-1,3,4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid (5.83 g) ~yield 84.2 %~
IR~KBr): 3430, 3340, 1776, 1717, 1680, 1535cm NMR~d6-DMSO): ~ 1.30 - 2.40~6H,m), 3.57~2H,br), 4.40~2H,s), 4.32(2H,ABq,J=12Hz), 4.70~1HJt,J=8Hz), 5.0(1H,d,J=5Hz), 5.55(1H,dd,J=5 ~ 8Hz), 7.20(1H,broad), 7.60~1H,broad), 7.86(4H,s), 8.74(lH,d,J=8Hz) Example 2 Cl~ In dichloromethane ~50 mQ) was dissolved 7~-CD-5-benzamido-5~car-b`oxy~aleramIdo~-3-hydroxymethyl-3-cephem-4-carboxylic acid d;triethylamine . 14 -~., salt (6.69 g). At room temperature, triethylamine ~1.01 g) and diketene ~1.68 g~ were added and the r~action was carried out Eor 3.0 hours. After the reac~ion had been completed, the reaction mixture was treated in the same manner as Example 1. The procedure provided 7~-~D-5-benzamido-5-carboxyvaler-amido1-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~5.12 g) ~yield 91.3 %)-IR~KBr): 3350, 1780, 1735, 1720, 1640, 1530cm NMR~ in d6-DMSO): 1.45 - 2.40~6l-l,m), 2.18~3H,s), 3.50~2H,ABq,Jal9llz), 4.34ClH,m), 4.88(2H,ABq,J=13Hz), 5.05(111,d,J=SHz), 5.65~1H,dd,J=5 ~ 9Hz), 7.27 - 8.0~5H,m), ~.46~1H,d, J=8Hz), 8.75~lH,d,J=9Hz) ~ 2) In water ~20 mQ) was dissolved 7~-~D-5-benzamido-5-carboxyvaler-amido)-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~561 mg) together with potassium lodide ~400 mg) and pyridine ~212 mg). The solution was ad-justed to pH 6.5 and, then, reacted at 60C for 50 minutes. Following the addition of ~ater ~10 mQ), the aqueous solution was adjusted to pH 7.0, washed twice with dichloromethane ~5.0 mQ~, readjusted to pH 6.0 and concentrated under reduced pressure. The residue was subjected to column chromatography on Amberlite XAD-2, followed by elution with water and, then, with a mixture of water and methanol. The eluate was concentrated and lyophilized. The procedure provided N-{7~-~D-5-benzamido-5-carboxyvaleramido)-3-cephem-3-ylmethyl~pyridinium-4-carboxylate monosodium salt ~352 mg).
IR~KBr1: 3360, 3250, 1765, 1645, 1630, 1605, 1575, 1530cm NMR~D2O): ~ 1.50 - 2.60~6H,m), 3.14~2H,ABq,J=19Hz), 4.36(1H,m), 5.05~1H,d,J=5Hz), 5.32~2H,A~q,J=15Hz), 5.60~1H,dd,J=5Hz), 7.0 - 9.0~lOH,m) ~3) ~Reference) In water (6.0 mQ) was dissolved 7~-~D-5-benzamido-5-. . .
carboxyvaleram~do)-3-~3_oxobutyryloxy~methyl-3-cephem-4-carbox~lic acid . ..~ ...

', (561 mg) together wlth thlourea (100 mg) and sodium hydrogen carbonate (253 mg). The solution was adjusted to pH 6.5 and reactecl at 60C for 50 minutes.
After cooling, the reactlon mixture was subjected to column chromatography on Amberlite XAD-2, followed by elution with water and water-methanol. The eluate was concentrated and lyophilized. The procedure pro~ided S~ 7~-~D-5-benzamido-5-carboxyvaleramido)-3-cephem-3-ylmethyl~-thiouronium-4~carboxylate monosodium salt (464 mg).
IR(KBr): 3350, 3230, l762, 1645, 1630, 1600, 1580, 1535cm NMR(D20): ~ 1.50 - 2.50(6H,m), 3.20 - 3.80 ~3H,m), 4.4~2H,m), 5.05(1H,d,J=5Hz), 5.53(111,dd,J=5Hz), 7.30 - 7.90~5H,m) Example 3 (Reference) In dichloromethane ~50 mQ) was dissolved 7-~D-5-~p-toluenesulfon-amido)-5-carboxyvaleramido)-3-hydroxymethyl-3-cephem-4-carboxylic acid di-triethylamine salt ~7.29 g) and, at room temperature, triethylamine (0.51 g) and diketene (1.26 g) were added. The reaction was carried out for 4.0 hours, after which time the reaction mixture was treated as in Example 1. The pro-cedure provided l-(D-5-~p-toluenesulfonamido)-5-carboxyvaleramido)-3-(3-oxo-butyryloxy)-methyl-3-cephem-4-carboxylic acid ~5.64 g) (yield 92.3 %).
IR(KBr): 3275, 1780, 1740, 1730, 1715, 1640, 1535cm NMR~ in d6-DMS0): 1.30 - 1.80(4H,m), 2.0 - 2.4~2H,m), 2.16~3H,s), 2.33(3H,s), 3.45(2H,ABq,J=19Hz), 4.91(2H,ABq,J=12Hz), 5.0(1H,d,J=5Hz), 5.57~1H,dd,J=5 ~7 8Hz)l 7.40(4H,m), 7.81(1H,d,J=9Hz), 8.64~1H,d,J=8Hz) ~xample 4 (Reference) In dichloromethane ~50 m~) was dissolved 7-~D-5-(p-tert-butylbenz-amido)-5-carhoxyvaleramido)~3-hydroxymethyl-3-cephem-4-carboxylic acid di-triethylamine salt ~7.35 g) and, at room temperature, triethylamine (1.01 g) and diketene (1.68 g) were added. The reaction was carried out for 2 hours, ~r~/e ~

:

3~

after which time the reaction rnixture was treated as in Example 1. The pro-cedure provided 7-(D-5-~p-tert-butylbenzamido)-5-carboxyvaleramido)-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid ~5.78 g) ~yield 93.8 %).
IR~KBr): 3280, 1780, 1740, 1725, 1710, 1640, 1530cm NMR(~ in d6-DMSO): 1.32~9H,s), 1.50 - 2.40~6H,m), 2.18~3H,s), 3.48(2H,br), 3.57(2~1,s), 4.35(1H,m), 4.94(2H,ABq,J=13Hz), 5.06(1H, d,J~5llz), 5.62(1H,dd,J=5 ~1 8Hz), 7.46(2H,d,J=8Hz), 7.85(2H,d,J=8~1z), 8.35(1H,d,J=8Hz), 8.78~1H,d,J=8Hz) Example 5 ~Re~erence) ~1~ In dichloromethane (50 mQ) was dissolved 7-(D-S-caprylamido-5-carboxyvaleramido)-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt ~6.01 g), followed by the addition of triethylamine (1.01 g) and di-ketene (1.68 g). The reaction was carried out at room temperature for 3 hours, after which time the reaction mixture was treated as in Example 1. The pro-cedure provided 7-(D-5-caprylamido-5-carboxyvaleramido)-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (4.36 g) ~yield 90.3 %).
IR(KBr~: 3320, 1780, 1745, 1725, 1715, 1645, 1535cm 1 MMR(~ in d6~DMS0): 0.70 - 2.40(2 lH,m), 2.22C3H,s), 3.53~2H, broad), 3.60(2H,s), 4.18(1H,m), 4.91(2H,ABq,J=12Hz), 5.04(1H,d,J=5Hz), 5.65(1H,dd,J=5 ~ 8Hz), 7.87(1H,d, J=8Hz), 8.70(1H,d,J=8Hz) Example 6 (Reference) In dic~loromethane (50 m~) was dissolved 7-phenylacetamido-3-hydroxymethy~l-3~cephem-4-carboxylic acid triethylamine salt (4.49 g) and, at room temperature, triethylamine (0.51 g) and diketene (1.26 g) were added.
The reaction was conducted for 2 hours, after which time the solvent was dis-tilled off under reduced pressure. The residue was diluted with water and adjusted to pH 6Ø This aqueous solution was extracted twice with ethyl ~ 17 -3~

acetate, brought down to pll 2.0 wltll 4N-IICQ and extracted three times with ethyl acetate. The ethyl aceta~e solution was washed wikh a saturated aqueous solution of sodium chloride, treated with magnesium sulfate and filtered. The solvent was distilled off under reduced pressure and the residue was recrys-tallized from ethyl acetate-ether. The above procedure provided 7-phenyl-acetamido-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (4.16 g) (yield 9~.3 %) IR(KBr): 3270, 1785, 1745, 1715, 1655, 1540cm 1 NMR(~ in d6-DMS0): 2.14~3H,s), 3.52~6H, broad), 4.86(2H,ABq,~=13Hz), 5.00~lH,d,J=5Hz), 5.63~ I,dd,J=5 ~1 9Hz), 7.22~5H,s), 8.93~lH,d,J=9Hz) Example 7 ~Reference) A mixture of dichloromethane ~300 mQ), triethylamine ~27 mQ) and dimethylaniline ~100 mQ) was previously cooled to 10C, and 7-~D-5-phthalimido-5-carboxyvaleramido)-3-(1-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-car-boxylic acid ~50 g) was dissolved therein. To this solution was added di-chlorodimethylsilane ~36 mQ) whereupon the internal temperature increased to 27C. The mixture was stirred at that temperature for 30 minutes, after which time the internal temperature was brought down to -35C. Then, phosphorus pentachloride ~32.4 g) was added. The mixture was stirred at -25C for 40 minutes and, after cooling to -35C, thioacetamide ~20 g) was added. The mix-ture was further stirred at -20 - -25C for ~0 minutes and, after cooling to -30C, methanol ~200 mQ) was gently added in droplets. Then, at the same tem-perature, sulfur mono-chloride ~17 mQ) was gently added dropwise. The mix-ture was- stirred for 20 minutes, after which time it was diluted with water ~200 mQ~ and brought to pH 3.2 ~ith 4a % aque~us patas~sium carbonate solution.
After s*irring ~or 6a minutes, the resultant crystals were collected by fi~ltrati~on and rinsed with water and acetone. The crude crystals thus ob-- 18 ~
~ ~ .

' ., , ,, ' "' . : . ' ~ ' :

tained were suspenclecl in 10 ~ hydrochloric acid ~230 mQ) and stirred at 30C
for 1 hour. The insolubles were filtered off and the filtrate was cooled to 5-10C and brought to pH 3.3 with potassium carbonate. The iltrate was stir-red for one hour, then the precipitated crystals were collected by Eiltration, rinsed with water and acetone, and dried over phosphorus pentoxide. The pro-cedure provided 7-amino-3-~1-methyltetrazol-5-yl~thiomethyl-3-cephem-4-car-boxylic acid (17.0 g).
IR~KBr): 1795cm NMR~ in D20 NaHC03): 3.61 ~ 3.98(ABq,J=18Hz,2-CH2), 4.21~s,tetrazole-CH3), 5.21~d,J=4.5~1z,6-H), 5.60~d,J=4.5Hz,7-~1) Chlorine gas ~2.8 g) was bubbled through a solution of diketene ~3.3 g) in methylene chloride ~160 mQ) under stirring and cooling to maintain the interanl temperature at -25 to -35C for a period of 100 minutes. Then, the mixture was further stirred at the same temperature for 30 minutes.
Separately, 7-amino-3-~1-methyltetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid ~10.0 g) and dibutylamine ~7.9 g) were dissolved in methylene chloride ~60 mQ) and the solution was cooled to -10C. To this solution, the above reaction mixture was added dropwise under stirring and cooling to maintain an internal temperature of -10 to -20C for 30 mlnutes. The mixture was further 2Q stirred at that temperature for 40 minutes. Thin-layer chromatography of this reaction mixture revealed the presence of 7-~4-chloro-3-oxobu~ylamido) 3-~1-methyltetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid. Thiourea ~4.64 g) was dissolved in this reaction mixture and the internal temperature was in-creased to 17 - 19C. The mixture was stirred at this temperature~ whereupon crystals separated out. The crystals were recovered by suction-filtration, washed with methylene chloride ~30 mQ) and dried. The p:rocedure provided the 2~?-imino-4-thiaz~lin-4~yl)acetamido]-3-~l-methyltetrazol-5yl)thiomethyl-3~ cep~è~ ~4~car~oxylic acid ~12.8 g), melting point: 176 - 180C ~decomp.) ~` 19 -" .

-. . : ,. . :
.. . ..

~' 3~

IR~Ksr): 1762~ 1662 cm NMR(ô in d6-DMS0): 3.39~s,CH2C0), 3.55 ~ 3.77~ABq,Jal8Hz,2-Cll2), 3.90(s,tetrazole 1-C113), 4.21 ~l 4.36~ABq, J=14Hz,3-CH2), 5.03(d,J=51-1z,6-H), 5.66(dd,J=9 HN S
FT 5llz,7-H), 6.23~s,thiazolin 5-11), 6.2-7.1~m,~
8.85~d,J=9Hz,-CONH-) Hh ~ ~Reerence) In dichloromethane ~30 mQ) ~as dissolved 7~-C2-thienylacetamido)-3-hydroxymethyl-3-cephem~4-carboxylic acid triethylamine salt (4.55 g), Eollowed 10 by the addition of succinic anhydride (1.50 g). The mixture was stirred at room temperature for 2 hours. The dichloromethane was distilled off and, following the addition of 3~6 aqueous phosphoric acid solution (100 mQ), the residue ~ras extracted with ethyl acetate (150 mQ). The ethyl acetate layer ~as washed with a saturated aqueous solution o-f sodium chloride ~100 mQ x 2), dried (over magnesium sulfate) and concentrated under reduced pressure. The residue b~as treated with ether and the resultant powder was recovered by filtration, washed oTith ether and dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-(2-thienylacetamido)-3-(3-carboxy-propionyloxy)methyl-3-cephem-4-carboxylic acid (4.00 g).
2QIR(KBr): 1782~-lactam), 1733(-CO2H)cm NMR(~ in d6-DMSO~: 2.50(4H,-CO(CH2)2-), 3.40 F1 3.63~2H,ABq,J=18Hz,2-CH2), 3.75~2H,s,-C_2CONH-), 4.71 FT 5.07(2H,ABq,J=13Hz,3-CH2), 5.07~1H,d,J=5Hz,6~H), 5.68~1H,dd,J-5 FT 8Hz,7-H),

6.~0 ~ 7.30(3H9 ~ ~ ), 9.10~1H,d,~=8Hz,-CONH-) Example 9 (Reference) In dichlorDmethane C25 m~) ~a~ dissolved 7~-~2-thienylacetamldo)-3-hydraxymethy1~3~cephem-4~carboxy~1ic acid triethylamine salt (3.60 g), follo~red ~y addition of phthalic anhydride ~1.80 g). The mixture was stirred ~ 2Q -..~, . .
' ' ~' -,: ` ' ':

~ ~323~

at room temperature for 2 hours, after which time it was treated in a manner similar to that descri~ed in Example 8. The procedure pro~ided 7~-~2-thi-enylacetamido)-3-(2-carboxybenzoyloxy)methyl-3-cephem-4-carboxyl~c acid ~3.44 g)- -l IRtKBr): 1777, 1724, 1650(shoulder)cm NMR(~ in d6-DMSO): 3.50 ~ 3.70~2H,ABq,J=181-1z,2-CH2), 3.77~2H,s,-CH2CO-), 4.91 ~ 5.29~2H,ABq,J=13l-1z,3-CH2~, 5.10~1H,d,J=5}1z,6-H), 5.70~111,dd,J=5 ~, 8Hz,7-1-1), 6.92 ~, 7~32~3HJ [~

7.64 (4H, \h~_ ), 9.12 (lH,d ,J-8Hz,-CONH-) Example 10 (Reference) In dichloromethane ~40 mQ) was dissolved 7~-mandelamido-3-hydroxy-methyl-3-cephem-4-carboxylic acid triethylamine salt (4.65 g), followed by addition of succinic anhydr-lde (1.50 g). The mixture was stirred at room tem-perature lQor 1 hour and a half. After the reaction had been completed, the mixture was treated as in Example 8 to obtain 7~-mandelamido-3-(3-carboxy-propion~loxy)methyl-3-cephem-4-carboxylic acid ~4.45 g).
IR~KBr~: 1776, 1737, 1684~shoulder)cm 1 NMR ~ in d6-DMSO): 2.48 ~4H,-CO(CH2)2-), 3.3 - 3.7 ~2H,2-CH2), 4.69 ~7 5.03(2H,ABqJJ=13Hz,3-CH2), 4.99~1H, ~-CH-), 5.06~1H,d,J=5Hz,6-H), 5.68~1H,dd,J=5 ~, 8Hz,7-H), 7.35~5H, ~ ), 9.31~1H,d,3=8Hz,-CONH-) Example 11 ~Reference) In dichloromethane ~7 mQ) was dissolved 7~-~2-thienylacetamido)-3-hydroxymethyl-3-cephem-4-carboxylic acid triethylamine salt ~0.68 g), followed by~ addi~ti~on of glutar~c anhydride C0.34 g). l~Le mixture was stirred at room ~ 21 -;
?

3~

temperature ~or 2 c~nd a half hours. The dlchloromethane was distilled off and, following the addition oE 3% aqueous phosphoric acid C15 mQ), the residue was extracted with ethyl acetate (25 mQ). The ethyl acetate layer was washed ~ith a saturated aqueous solution of sodium chloride (15 InQ x 2) and dried (over magnesium sulfate). Then, upon dropwise addition of a 2N-solution of sodium 2-ethylhexanoate in isopropyl alcohol (1.50 mQ), there was obtained a powder. This powder was recovered by filtration, washed with ethyl acetate-ether and dried over phosphorus pentoxide. The proc~dure provided 7R-(2-thi-en~lacetamido)-3-(4-carboxybutyryloxy)methyl-3-cephem-4-carboxylic acid di-la sodium salt ~0.42 g).IR~KBr): 1760, 1736(shoulder), 1661, 1609cm NMR(~ in D20): 1.90~2H,-CH2CII2CH2-), 2.28(4H,-CH2CH2CH2~), 3.21 ~7 3.61(2H,ABq,J=18Hz,2-CH2), 3.78(2H,s, ~ ~ C~12- )' 4.4 - 4.9~2H,3-CH2), 4.98(1H,d,J=5Hz,6-H), 5.60~1H,d,J=5Hz,7-H), 6.95 ~ 7.28~3H, ~ ~ ) Exampel 12 (Reference~
In dimeth~lformamide (50 mQ) was dissolved 7~ -5-~p-t-butylben~
amldo)-5-car~oxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid di-triethylamine salt t7.35 g), followed by addition of succinic anhydride ~1.50 g). The mixture was stirred at room temperature for 30 minutes. After the reaction had been completed, 3% aqueous phosphoric acid ~250 mQ) was added, followed by extraction with ethyl acetate (500 mQ). The ethyl acetate layer was washed with water ~250 mQ x 2), dried (over magnesium sulfate) and col_ centrated under reduced pressure. Then, upon addition of ether, there was ob-tained a po~er. Th~s powder was recovered by filtration, washed with ether and d~ied under reduced pressure over phosphoru5 pentoxide- T~ ocedùre ~roy~ded 7~ D~5~p~t~fiut~1~enzamido~carfioxyvaleram~d~]r3?G3~carfioxyprop~

` ~ ~

onyloxy~methyl-3-cephem-4-carboxylic acid (6.20 g).
IR~KBr): 1779, 1732, 1640cm N~ in d6-DMSO): 1.28~9H,s,~ 73 & 2.24~6H,-~CH2)3-), 2.50~4H,-COCll2CH2CO-), 3.38 & 3.63~2H,ABq,J=18llz, 2-CH2), 4.37~1H,-CH-NH-), 4.71 ~ 5.06~2H,ABq,J=13Hz, 3-CH2), 5.04~1H,d,J=5Hz,6-H), 5.66~1H,dd,J=5 & 8Hz, 7-H), 7.44 & 7.81~4H,- ~ _), 8.43~1H,d,J=8Hz, CONH-), 8.80~lH,d,J=8Hz, -CONH-) Example 13 ~Reference) In dimethylformamide ~50 mQ) was dissolved 7~-[D-5-~p-t-butyl-benzamido)-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt ~7.35 g), followed by addition of ph*halic anhydride ~1.63 g). The mixture was stirred at room temperature for 30 minutes, a-fter wh~ch time it was treated as in Example 12. The pr~cedure provided 7~-[D-5-~p-t-butylbenzamido)-5-carboxyvaleramido]-3-~2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid ~6.70 g).
IR~KBr~: 1784, 1726, 1642cm Nh~ ~ in d6-DMSO): 1.27~9H,s,- ~ ), 1.72 & 2.22~6H, -CCH2)3-), 3.46 ~ 3.71(2H,ABq,J=18Hz,2-CH2), 4.37 (lH,-CH-), 4.89 & 5.27~2H,ABq,J=13Hz,3-CH2), 5.08 (lH,d,J=5Hz,6-H), 5.67~1H,dd,J=5 ~ 8Hz,7-H), 7.43 &

7.81~4H,+ ~ ), 7.62~4H, ~ - ), 8.42~1H, d,J=8Hz,-CHNHCO-), 8.80~1H,d,J=8Hz,-CONH-) Example 14 CReference) In dimeth~l~'ormamide C50 m~) was dissolved 7~-[D-5-~p-t-butyl-~ 23 ~ -:- : ~: :

3i~

~enzamido)-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic ac:id ditriethylamine salt ~7.35 g), followed by addition of maleic anhydrlde ~1.49 g). rne mixture was stirred at room temperature for 30 minutes, after which time it was treated as in Example 12. The procedure provided 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido]-3-~3-carboxyacryloyloxy)methyl-3-cephem-4-carboxylic acid ~6.05 g).
IR(KBr): 1780, 1727, 1640 cm NMR~ in d6-DMSO): 1.32(9H,s, ~ ), L-77 ~1 2.26~6H,-(CH2)3-), 3.41 ~ 3.65~2H,ABq,J=18Hz,2-CH2), 4.37(1H, C_- ), 4.80 ~ 5.16~2H,ABq,J=13Hz,3-CH2), NH
5.02(1H,d,J=5Hz,6-H), 5.66ClH,dd,J=5 ~ 8Hz,7-H), 6.34(2H,-CH=CH-), 7.44 ~ 7.82(4H, ~ ),

8.40~lH,d,~=8Hz,-CHN_CO-), 8.80~lH,d,J=8Hz, -CONH-) Example 15 (Reference) In dichloromethane (20 mQ) was dissolved 7~-~D-5-~p-t-butylbenz-amido)-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid di-trlethylamine salt ~1.47 g), followed by addition of p-chlorophenylthiosuc-cinic anhydride Co.51 g). The mixture was stirred at room temperature for 1 hour, after which time it was treated as in Example 8. The procedure pro-vided 7~-[D-5-~p-t-butylbenzamido)-5-carboxyvaleramido]-3-[3-carboxy-3(or 2)-(p-chlorophenylthio)propionyloxy]methyl-3-cephem-4-carboxylic acid (1.50 g).
IR~KBr): 1778, 1728, 1636 cm NMR(~ in d6-DMSO): 1.28~9H,s, ~ ), 1-74 ~ 2-24 ~6H,-~CH2)3-), 2.73~2H,-CH2CH-), 3.3 - 3.8~2H,2 CH2), ~CH-S), 4.40~lH,-C_-NH-), 4.73 ~ 5.07~2H,ABq, r 24 -.

3~

J=1311~,3-C112), 5.02 (11-1,cl,J=511z,6-11), 5.68(1ll,dd, J=5 ~1 8tlz,7-I-I), 7.~2(411,CQ ~ ~ S-), 7.~ ~

7.82(4il,~ ), 8.44(1}1,d,J=8Hz,-C}lNIICO-), 8.82 (lll,d,J=8Hz,-CONH-) Example 16 (Reference) In dichloromethane (50 mQ) was dissolved 7~-~D-5-phthalimido-5-car-boxyvaleramido)-3-hydroxyrnethyl-3-cephem-4-carboxylic açid ditriethylamine salt (7.05 g), followed by addition of succinic anhydride (1.50 g). The mix-ture was stirred at room tempera~ure for 1 hour and ahalf, after which time it was treated as in Example 8. The procedure provided 7,B- (D-5-phthalimido-5-carboxyvaleramido)-3-(3-carboxypropionyloxy)methyl-3-cephem-4-carboxylic acid (5.~3 g)-IR(KBr): 1777, 1710, 1644 cm NMR(~ in d6-DMSO): 1.52 ~ 2.17 (6H,-~CH2)3-), 2.50~4H,-~CH2)2-), 3.3 - 3.8~2H,2-CH2), 4.70 ~, 5.03~2H,ABq,J=13Hz, 3-CH2), 4.72~1H~t,J=7Hz,-CH-), 5.01~1H,d,J=5Hz,6-H), 5.62 ~lH,dd,J=5 ~ 8H~,7-H), 7.86 ~4H,s, ~ ), 8.78 ~lH,d,J=8Hz,-CONH-) Example 17 ~Reference) In dichloromethane ~50 mQ) was dissolved 7~- ~D-5-phthalimido-5-carboxyvaleramido)-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt ~7.05 g), followed by addition of phthalic anhydride ~2.22 g). The mix-ture was stirred at room temperature for 1 hour and a half, after which time it was treated as in Example 8. The procedure provided 7,B-~D-5-phthalimido-5-carboxyvaleramido)-3- ~2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid ~6.39 g).

~ . ~ - , .
. .
-~, , .

3~i IR(KBr): 1772, 1714, 16~l3 cm 1 NMR(~ in d6-DMSO): 1.52 ~7 2.18(6~l9-(C:1l2)3-), 3.42 ~1 3-69(2~1,ABq, J=1811z,2-Ctl2), ~.74~lll,t,J=811z,-Ct1-), 4.89 ~, 5.28(2H,ABq,13Hz,3-C112), 5.06(1~1,d,J=511z,6-tl), 5.66(111,dd,J=5 ~ 811z,7-H), 7.64~4 7.86(4H,s, j~ ~ N~ ), 8.81(~H,d,J=81 ~" .
-CON11-) Example 18 (Reference) 7~ (2-Thienylacetamido)-3- (3-carboxyacryloylo-xy3-methyl-3-cephem-4-carboxylic acid was synthesized as in Example 8.
IR(KBr): 1780, 1725, 1638 cm NMR(d6-DMSO): ~ 3.43 ~, 3.76(2H,ABq,J=18Hz,2-C}12), 3.75(2H,s,-CH2CO-), 4.79 ~, 5.14 (2 H,ABq,J=13Hz,3-CH2), 5.07(lH,d,J=5Hz,6-H), 5.68(1H,dd,J=5 ~ 8Hz,7-H), 6.35(2H,s,-CH=CH-), 6.90 7.29(3H, ~ ~ ), 9.10(lH,d,J=8Hz,-CONH-) S

Example 19 (Reference) To a mixture of 7,~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt (2.20 g) and 3-nitrophihalic anhydride (1.16 g) was added dichloromethane (15 m~) together with triethylamine (0.42 mQ). The resultant solution was stirred at room temperature for 1 hour and a half. After the reaction had been completed, the dichloromethane was distilled off under reduced pressure and 3% aqueous phosphoric acid (120 m~) and ethyl acetate (160 mQ) were added to the residue.
The ethyl acetate laye,r was washed with water (80 mi~ x 2), .ed tover magne-sium sulate) and conc:entrated under reduced pressure. Then, upon addition o ether, there was obtained a powder. This powder was recovered by filtra-~, - 26 -.~: .. ,, , :: ' ~ :
. - -. : : ~- : : , :

,: ~ - :.
,~ : :.: . : :
, :

tion, waslled with ether ancl clried under reduccd pressure over phosphorus pent-oxide. Ihe proceclure provided 7~-LD-S-(p-t-butylbenzamido)--5-carboxyvaler-amido]-3- (2-carboxy-6 (or 3)-nitrobcnzoyloxy)methyl-3-cephem-4-carboxylic acid (2.10 g).
IR(KBr): 1783, 1735, 1640 cm NhlR(~ in d6-DMSO): 1.28(9}1,s,-C(C}13)3), 1.53 ~ 2.23(6H,-(CH2)3-), 3-43 l~
3.70(2H,ABq,J=18}1z,2-CIl2), 4.37(lH,-CH-), 4.96 5.41 (21l,ABq,J=13Hz,3-CH2), 5.07 (lH,d,J=5Hz,6-H), 5.68(1H,dd,J=5 ~1 8Hz,7-H), 7.42 ~, 7.80(411, J=8Hz,-CII-NH-), 8.84(1H,d,J=8Hz,-CONII-) Ex~ (Reference) To a mixture of 7~-(D-5-phthalimido-5-carboxyvaleramido)-3-hydroxy methyl-3-cephem-4-carboxylic acid ditriethylamine salt (2.12 g) and 3-nitro-phthalic anhydride (1.16 g) was added dichloromethane (15 m~) together with triethylamine (0.42 m~). The solution was stirred at room temperature for 1 hour and a half, after which time it was treated as in Example 19. The pro-cedure provided 7~-(D-5-phthalimido-5-carboxyvaleramido)-3-(2-carboxy-6 (or 3)-ni~robenzoyloxy)-methyl-3-cephem-4-carboxylic acid (2.08 g).
IR(KBr): 1775 (shoulder), 1718, 1642 cm NMR(~ in d6-DMSO): 1.54 ~ 2.19(6H,-(CH2)3-), 3-40 ~ 3-68(2H,ABq, J=18Hz,2-CH2), 4.73(1H,t,J=7Hz,-CH-), 4.98 ~
5.42(2H,ABq,J=13Hz,3-CH2), 5.06(1H,d,J=5Hz,6-H), 5.66(1H,dd,J=5 ~ 8Hz,7-H), 7.7 - 8.4(3H, ~ ), 7 86(4H,s, ~J ~ , 8.81(1H,d,J=8HZ,-CONH-) .

~ 3~3~ .

example 21 (Reference) To 7~-[D-S-(p-t-butyLbenzalllido)-5-carboxyvaleramido]-3 hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt (2.20 g) and trimellitic an-hydride (1.15 g) were added dichloromethane (15 mQ) and triethylamine (0.84 mQ) and the resultant solution was stirred at room temperature for 1 hour.
After the reaction had been completed, the mixture was treated in the same manner as Example 19. The procedure provided 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido~-3-(2,4 (or 5)-dicarbo~ybenzoyloxy)me~hyl-3-cephem-4-carboxylic acid (2.14 g).
IR(KBr): 1777, 1724, 1636 cm NMR~ in d6-DMSO): 1.29(9H,s,-C~CH3)3), 1.54 ~1 2.25~6~l,-(CH2)3-), 3.99 ~ 3.72(2H,ABq,J=18Hz,2-CH2), 4.38(111,-CH-), 4.95 ~1 5.30(2H,~Bq,J=13Hz,3-CH2), 5.11(1H,d,J=
5Hz,6 H), 5.70~1H,dd,J=5 ~ 8Hz,7-H), 7.43 ~1 7.82(4H,- ~ + ), 7.7 - 8.3(3H, ~ COOH

8.43~1H,d,J=8Hz,-CH-NH-), 8.84~1H,d,J=8Hz,-CONH-) Example 22 ~Reference) In dimethylformamide ~10 mQ) was suspended desacetylcephalosporin C~2.16 g) and, under cooling with ice, concentrated hydrochloric acid ~0.83 mQ) was added. To the resultant solution was added dimethylformamide ~10 mQ) together with triethylamine ~4.20 mQ) and 3-nitrophthalic anhydride ~3.86 g), and the mixture was stirred at room temperature for 2 hours. Following this reaction, the mixture was diluted with 3% aqueous phosphoric acid ~150 mQ) and extracted with e~hyl acetate (250 mQ x 2). The ethyl acetate layer was washed with water (200 mQ) and a saturated aqueous solutio~ of sodium chloride ~200 mQ~dried ~over magnesium sulfate) and concentrated under reduced pressure.

, :. ::-- ~ ~

3~

~en, upon addition of ether, there was obtained a powder. This powder was recovered by filtration, washed with ether and dried under reduced pressure over phosphorus pentoxide. Tlle procedure provided 7~-[D-5-~2-carboxy-6 (or 3)-nitrobenzamido)-5-carboxyvaleramido~-3-(2-carboxy-6 (or 3)-nitrobenzoyloxy)-methyl-3-cephem-4-carboxylic acid 13.77 g).
IR(KBr): 1780(shoulder), 1729, 1638, 1534, 1348 cm N~IR~ in d6-DMSO): 1.67 ~ 2.23(6H,-(Cll2)3-), 3.60(2}1,2-C112), 4.39(1H,-C11-), 4.97 ~1 5.40(2H,ABq,J=13Hz,3-C}l2), 5.08(1H,d,J=5Hz,6-H), 5.68(1H,dd,J=5 ~ 8Hz,7-H), 7.5 - 8.5(7H, ~ ~ 2x 2 ~7 -C~l-NH-), 8.78(lH,d,J=8Hz,-CONH-) Example 23 (Reference) In dichloromethane (30 mQ) was dissolved 7~-(2-thienylacetamido)-3- `
hydroxymethyl-3-cephem-4-carboxylic acid triethylamine salt (4.55 g), followed by addition of O-carboxymandelic anhydride. (2.38 g). The rnixture was stirred at room temperature for 1 hour, after which time the dichloromethane was dis-tilled off. To the residue was added 3% aqueous phosphoric acid (100 m~) followed by extraction with ethyl acetate (150 mQ). The ethyl acetate layer was washed with a saturated aqueous solution of sodium chloride (100 m~ x 2), dried (over magnesium sulfate) and concentrated under reduced pressure. Then, '.
upon addition of ether, there was obtained a powder. This powder was re-covered by filtration, washed with ether and dried over phosphorus pentoxide under reduced pressure. The procedure provided 7~-(2-thienylacetamido)-3-mandelyloxymethyl-3-cephem-4-carboxylic acid (4.00 g).
IR(KBr): 1778, 1742, 1666 cm NMR(~ in d6-DMSO): 3.24 ~ 3.45(2H,ABq,J=18Hz,2-CH2), 3.74(2H,s,-CH2CONH-~, ~ 29 -.

.:

3~æ3~

~.76 ~, 5.()6~21-~,ABq,J=l311~, 3-CI-12), 5.03(1H,d, J=51-1~,6-11~, 5.l6(1H,s, ;~ CH-), 5.68(111,dd, J=5 ~ 81-1z,7-11), 6.90 ~j 7.34(8rl,

9.09(1~1,d,J=8~1z,-CON11-) Example 2~ (Referel~ce) In dimethylformamide (40 mQ) was dissolved sodium 7~-mandelamido-3-hydroxymethyl-3-cephem-~-carboxylate (3.86 g), followed by addition of O-car-boxymandelic anhydride ~2.67 g). The mixture was stirred at room temperature for 30 minutes, after which 2% H3PO4(150 mQ) was added. The mixture was ex-

10 tracted with ethyl acetate (25~ m~) and the ethyl acetate layer was rinsedwith water (150 m~ x 2), dried (over magnesium sulfate) and concentrated under reduced pressure. Then, upon addition of ether, there was obtained a powder.
This powder was recovered by filtration, washed with ether and dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-mandel-amido-3-mandelyloxymethyl-3-cephem-4-carboxylic acid (3.68 g).
IR(KBr): 1777, 17~5, 1669 cm NMR(~ in d6-DMSO): 3.2 - 3.7 (2H,2-CH2), 4.7 - 5.2(5H,3-CH2,6H ~, -CH-x2), 5.69(lH,7-H), 7.2 - 7.6(lOH, (~)-- x2 8.67 (lH,-CONH-) Example 25 (Re:Eerence) In dimethylformamide (50 mQ) was dissolved 7~-[D-5-(p-t-butylbenz-amido)-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid di-triethylamine salt (7.35 g), followed by the addition of O-carboxymandelic anhydride (2.67 g). The mixture was stirred at room temperature for 30 minutes, after which time 3% aqueous phosphoric acid (250 mQ) was added, followed by extraction with ethyl acetate (500 mQ). The ethyl acetate layer was rinsed with water (250 mQ x 2), dried (over magnesium sulfate) and concentrated under ..

- ' ' ,' ~ : ' reduce~l pressure. Then, upon ~clclition of etiler, -thcre was obtainecl a powder.
This powder was recov~red by fill:ration, washed with ether and dried undcr reduced pressure over phosphorus pentoxide. The procedure provided 7~-[D-5-(p-t-bu-tylbenzamido)-5-carboxyvaleramido] -3-mandelyloxymethyl-3-cephem-4-car-boxylic acid (6.30 g).
IR(KBr): 1778, 1736, 1642 cm NMR(~ in d6-DMSO): 1.29(911,s,-C(C113)3), 1.73 ~ 2.23(611,-(C~12)3-), 3.2 - 3.6(2H,2-CH2), 4.38 (lH,-C1l-NH-), 4.75 ~1 5.04(21l,ABq,J=13Hz,3-CH2), 5.02(1H,d,J=5Hz,6-fl), 5.16(1H,s, ~-CH-), 5.66(1H,dd,J=5 & 8Hz,7-H), 7.2 - 7.5(5H, C~--), 7.43 ~1 7.81(4H, ~/~\~ ), 8.43 ~lH,d,J=8Hz,-CH-NII-), 8.80 (lll,d,J=8Hz, -CONH-) Example 26 (Reference) In dichloromethane (50 m~) was dissolved 73-[D-5-phthalimido-5-carboxyvaleramido-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt (7.05 g), followed by addition of Q-carboxymandelic anhydride (2.38 g).
The mixture was stirred at room temperature for 1 hour, after which time it was treated as in Example 23. The procedure provided 7~-(D-5-phthalimido-5-carboxyvaleramido)-3-mandelyloxymethyl-3-cephem-4-carboxylic acid (6.25 g).
IR(KBr): 1773, 1715, 1647 (shoulder) cm NMR(ô in d6-DMSO): 1.54 ~, 2.22(6H,-(CH2)3-), 3.27 ~1 3.49(2H,ABq,J=18Hz, 2-CH2), 4.73~lH,t,J=7Hz,-CH-N ~ ), 4.74 ~ 5.03(2H, ABq,J=13Hz,3-CH2), 4.98 (lH,d,J=5Hz,6-H), 5.16 (lH,s, ~ tCH-), 5.61(1H,dd,J=5 ~ 8Hz,7-H), 7.2 - 7.5 (5H~ ~ ), 7.86(4H,s, ~\N- ), 8.77 (lH,d,J=8Hz,-CONH-) ~ ~ ~ 3 Exalllple 27 (Reference~
In dichloromethane (75 mQ) was suspended 7~-[D-5-phthalimido-5-car-boxyvaleramido~-3-(2-carboxybellzoyloxy)-methyl-3-cephem-4-carboxylic acid (6.51 g). Then, at a temperature not exceeding 10C, triethylamine (4.20 mQ) was added. To the solution thus obtained was added N,N-dimethylaniline (10.0 mQ) together with dimethyldichlorosilane (4.40 mQ). The mixture was stirred at 20 - 25C for 30 minutes. The mixture was then cooled to -30C, followed by addition of phosphorus pentachloride t4.20 g). It was then re-acted at -25 ~ 2C for 30 minutes, after which time methanol (25 mQ) was added dropwise at a temperature not exceeding -20C. The mixture was reacted at -15 - -10C for 20 minutes and, after the addition of water (50 mQ), was stirred vigorously for 5 minutes. The reaction mixture was separated and the aqueous layer was taken, washed with ethyl acetate and adjusted to pH 3.2 with 40% aqueous potassium carbonate solution. The resultant crystals were collected by filtration, washed with water, 50% aqueous methanol and acetone in the order mentioned and dried. The procedure provided 7~-amino-3-(2-car-boxyben~oyloxy)methyl-3-cephem-4-carboxylic acid (3.43 g).
IR(KBr): 3170, 1798, 1730, 1700, 1615 cm NMR(~ in D20~NaOD): 3.55(2H~ABq,J=18H~), 4.6 - 5.6(4H,m), 7.30 - 7.90(4H,m) Example 28 (Reference) In dichloromethane (70 mQ) was suspended 7~-[D-5-phthalimido-5-car-boxyvaleramido]-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (5.88 g).
Then, at a temperature not exceeding 10C, triethylamine (2.80 mQ) was added, followed by the addition of N,N-dimethylaniline (10.0 mQ) ~nd dimethyldichloro-silane (3.13 mQ). The mixture was stirred at 20 - 25C for 30 minutes, after which phosphorus pentachloride (4.20 g) was added at -30C. The mixture was reacted at -25 _` 2C for 30 minutes, after which time methanol (25 mQ) was , .' ' ' ' ' ' ' ': ,;
' : . . :

: :: . , .

3;;~3~i added dropwise at a temperat~lre not exceeding -20C. 'I'he reaction was further allowed to proceecl a-t -15 - -10C for 20 minutes and, then, water ~50 m~) was added at -15 - -10C, ~ollowed by vigorous stirring for 5 minutes. The water layer was taken, washed with dichloromethane and adjusted to pH 3.5 with a 40% aqueous solution oE potassium carbonate. The resultant crystals were re-covered by filtration, washed with water, 50% aqueous methanol and acetone, and finally dried. The procedure provided 7~-amino-3-t3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (2.84 g).
IR~KBr): 3200, 1800, 1745, 1720, 1622 cm MMR~ in D20~NaOD): 2~27(3HJS)~ 3.48(2H,ABq,J=18~1z), 4.6 - 5.6(4H,m) Example 29 (Reference) In dimethylformamide ~70 mQ) was suspended desacetylcephalosporin C
~13.7 g), followed by the addition of concentrated sulfuric acid (2.4 mQ).
To the resultant solution was added triethylamine (29 mQ), together with N-carboethoxyphthalimide ~8.5 g). The mixture was stirred at 30C for 50 minutes, followed by the addition of succinic anhydride ~3.0 g). The mixture was stirred for 30 minutes3 at the end of which time a further amount ~0.6 g) of succinic anhydride was added. The mixture was further stirred or 1 hour and, then, poured in a cold saturated aqueous solution of sodium chloride ~200 mQ~. The solution was made acidic with phosphoric acid and extracted three times with ethyl acetate. The extract was extracted back into a solu-tion of sodium hydrogen carhonate ~8 g) in water ~150 mQ). The water layer was adjusted to pH 1.7 with phosphoric acid and extracted with a mixture of tetrahydrofuran and dichloromethane (1:4). The extract was dried over mag~
nesium sulfate, filtered and, with the addition of triethylamine (18 mQ), conc~ntrated to dryness. To the residue was added dichloromethane (200 mQ) together ~ith triet~lamine ~6 mQ) and dimeth~laniline (30 mQ). Then, follow-,;

- . - : . .

. ' ' ' ' ' ' , ' - , ' ' -3~3 ing the a~dition of dimethyldichlorosilane (21 mQ)~ the mixture was stirred for 30 minutes, aftcr which it was cooled to -30C and phosphorus penta-chloride (20 g) was ad~ed. The mixturo was stirred at -30~C for 30 ~ninu-tes and, following the addition of methanol (63 mQ), it was further stirred for 30 minutes. Then~ it was diluted with water (120 mQ), brought to pH 3.0 and allowed to cool. The resultant crystals were recovered by fiLtration. The above procedure provided 7-amino-3-(3-carboxypropionyloxy)-methyl-3-cephem-4-carboxylic acid (5.4 g).
IR(KBr): 1802, 1735, 1720(shoulder)cm ~xample 30 (Reference) To 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido]-3-mandelyloxy-methyl-3-cephem-4-carboxylic acid (6.67 g) was added dichloromethane (60 mQ) and, under cooling with ice, dimethylaniline (3.78 mQ) and triethylamine ~4.20 mQ) were added. To the resultant solution was added dim~thyldichloro-silane (3.87 g), followed by stirring at 8 - 15C for 1 hour. Then, at -30DC, dimethylaniline (1.26 mQ) and phosphorus pentachloride (4.17 g) were added.
The mixture was stirred at -30DC - -20C for 2 hours. Then, at -45DC, methanol (30 mQ) was added dropwise over a period of 10 minutes. After the dropwise addition had been completed, the mixture was stirred at -10- -5C for 40 minutes and, then, water (20 mQ) was added dropwise over 5 minu~es. Then, the mixture was adjusted to pH 3.3 with concentrated aqueous ammonia, whereupon a white slurry separated. After an hour of standing under ice-cooling, the precipitate was recovered by filtration, washed with water, methanol and ether in the order mentioned and dried under reduced pressure over phosphorus pent-oxide.
The procedure provided 7~-amino-3-mandelyloxymethyl-3-cephem-4-car-boxylic acid (2.31 g).
IR(KBr): 1800, 1740, 1621 cm .

, 3~3~

Fxample 3I (Reference) To 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido]-3-[2-carboxy-6 (or 3)-nitrobenzoyloxy~methyl-3-cephem-4-carboxylic acid (7.26 g) was added dichloromethane (60 mQ) and, ~mder cooling with ice, dimethylaniline (3.78 m~) and triethylalnine (4.20 mQ) were added. To the resultant solution was added dimethyldichlorosilane (3.87 g), Eollowed by stirring a~ 7 - 15C or 1 hour.
The mixture was cooled to -30C and dimethylaniline (1.26 mQ) and phosphorus pentachloride (4.17 g) were added. The mixture was stirred at -30 - -20C for 2 hours, after which time it was cooled to --45C and methanol (30 mQ) was added dropwise over a period of 15 minutes. After the dropwise addition had been completed, the mixture was stirred at -10 - -8C for 40 minutes and, then, water (20 mQ) was added dropwise over 10 minutes. The mixture was then adjusted to pH 3.4 with concentrated aqueous ammonia7 whereupon a substantially white slurry separated. After 45 minutes' standing under ice-cooling, the precipitate was recovered by filtration and washed with water, methanol and ether in the order mentioned. It was then dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-amino-3-~2-carboxy-6 (or 3)-nitrobenzoyl~x~]methyl-3-cephem-4-carboxylic acid ~2.93 g).
IR~KBr): 1787, 1734, 1614, 1535, 1350 cm Example 32 ~Reference) In water ~48 mQ) was suspended 7~-amino-3-~2-carboxybenzoyloxy) methyl-3-cephem-4-carboxylic acid (7.33 g) and, at 0 - 3C, 2N-sodium hydroxide ~19.5 mQ) was added in small installments, care being used not to allow the pH
to exceed 8.5. Then, following the addition of sodium hydrogen carbonate (3.65 g), a solution of D-~-sulfophenylacetyl chloride (5.0 g) in ethyl acetate (8.8 mQ) was added dropwise at 0 - 5C over a period of 1 hour. After the dropwise addition had been completed, the reaction was carried out at 0 - 5C
for 20 minutes. The reaction mixture was adjusted to pH 5.5 and separated.

~ . :
.. .

.. .. - -, ."' ' '~ ,~ ' ~ .
-; :

~ ? ~

The water layer wa~ ~aken, degassecl and7 after confirming that the pl-l was wi-thin the range of 5.5 to 6.5, ethanol (800 mQ) was adcled over a period of 1 hour. Thereafter, the mixture was stirred for 30 minutes, followed by cool-ing to a temperature not exceeding 5C. The resultant crystals were recovered by filtration, washecl wi-th ethanol-water (10:1) and ethanol in the order men-tioned and dxied. The procedure provided 7~-(D-~-sulfophenylacetamidc)-3-(2-carboxybenzoyloxy)-methyl-3-cephem-4-carboxylic acid trisodium salt (11.6 g~.
IR(KBr): 3350, 1768, 1735, 1670, 1610 cm 1 NMR(~ in D20): 3.44(2H,ABq,J=18Hz), 5.00(2H,ABq,J=1311z), 5.06(1~-1,d,J=5Hz), 5.08(1H,s), 5.67(1H,d,J=5Hz), 7.3 - 7.9(m,9H) Example 33 (Reference) In water (48 mQ) was suspended 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~6.09 g) and, at 0 - 3C, 2N-sodium hydroxide (10.4 mQ) was added in small installments, care being used not to allow the pH to exceed 8.5. Then, following the addition of sodium hydrogen carbonate (3.65 g), a solution of D-~-sulfophenylacetyl chloride (5.0 g) in ethyl acetate (8.8 mQ) was added dropwise at 0 - 5C over a period of 1 hour. After the dropwise addition had been completed, the reaction was carried out at 0 - 5~C
for 20 minutes. Then, the reaction mixture was adjusted to pH 5.5 and separat-ed. The water layer was taken, degassed and, after confirming that the pH waswithin the range of 5.5 to 6.5~ ethanol (800 mQ) was added to this aqueous solution (about 80 mQ). Thereafter, the mixture was stirred for 30 minutes, at the end of which time it was cooled to a temperature not exceeding 5C.
The resultant crystals were recovered by filtration, washed with ethanol-water (10:1) and ethanol, and dried. The procedure provided 7~-(D-~-sulfophenyl-acetamido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid disodium salt (8.8 g).

..~`i , ~ .

IR(KBr): 3300, 1763, 1740, 1680, 1610~ 1~15, 1047 cm o NMR~ in D20): 2.27(3H,s, -CCI-13), 3.29(21l,ABq,J=18l-1z,2-CH2)~ 4.84(2il,ABq, J=13Hz,3-C112), 5.00(111,d,J=5llz,6-H), 5.07~1W,s,-CH- ), S03Na 5.70(1H,d,J-5Hz,7-H), 7.25 - 7.80(5~1,m) Example 34 ~Reference) In dimethylformamide (10 mQ) was dissolved 7~-tD-c~-sulfophenyl-acetamido)-3-hydroxymethyl-3- cephem-4-carboxylic acid disodium salt (940 mg) together with glutaric anhydride (580 mg) and triethylamine (404 mg). The mix-ture was stirred at room temperature for 2 hours, after wh.ich time the dimethyl-formamide was distilled off. To the residue was added a small amount o:E water and the aqueous solution was desalted with Amberlite IR-120(H), adjusted to pH 6.0 with lN-sodium hydroxide solution and lyophilized. The lyophilizate was re-dissolved in water and purified by column chromatography on Amberlite XAD-2. The procedure provided 7~-(D-c~-sulfophenylacetamido)-3-(4-carboxy-butyryloxy)methyl-3-cephem-4-carboxylic acid trisodium salt.
IR(KBr): 1760, 1675, 1620 cm NMR~ in D20): 1.65 - 2.60~6H,m), 3.41(2H,q,2-CH2), 4.83(2H,d,3-CH2), 5.0(1H,s, e3CH- ), 5.08(1H,d,6-H), 5.75(1H~d,7-H), S03Na 7.47(5H,m) Example 35 (Reference) In dimethylformamide (3 mQ) was dissolved 7,~-~D-c~-sulfophenylacet-amido)-3-hydroxymethyl-3-cephem-4-carboxylic acid disodium salt (470 mg) to-gether with succinic anhydride (250 mg) and triethylamine (200 mg). The mix:-ture was stirred at room temperature for 2 hours. It was then diluted with a small amount of water and the dimethylformamide was distilled off under reduced ._A, , " , .
' ', ~' ~, ..
.. .' ` . ' , ,' ' ' ~
.

' : ' '' ' . :

pressure. Ille residue was dissolved by the adclition of water an~ purified by column-chromatography on ~nberlite XAI)--2. I`h~ procedur~ provided 7~-(D-~-sulfophenylacetamido)-3-(3-carboxypropionyloxy)methyl-3-cephem-4-carboxylic acid trisodium salt.
IR(KBr): 1765, 1685, 1600 cm NMR(~ in D20): 2.62(4H,s,-CO(Cll2)2C0-), 3.38(2H,q,2-C}l2), 4-95(2}l,3-CI-l2), 5.10(1}1,s, ~ ~ - C}l- )) 5.21(1}1,d,6-H), 5.91(1il,d,7-ll), ''~ S03Na 7.78(5H,m ~ - ) Example 36 (Reference) In chloroform (5 m~) was dissolved 7~-(D-~-sulfophenylacetamido)-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethylamine salt (480 mg), fol-lowed by the addition of diketene (300 mg). The mixture was stirred at room temperature for 2 hours, after which the solvent was distilled off. The re-sidue was diluted with water and desalted with Amberlite IR-120(H~. The de-salted solution was adjusted to pH 5.7 with lN-sodium hydroxide solution and lyophilized. Yield 400 mg. The lyophilizate was purified by column chromatog-raphy on Amberlite ~AD-2. The procedure provided 7~-(D-~-sulfophenylacetamido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid disodium salt.
In IR and N~IR spectra, this product was found in good agreement with the product obtained in Example 33.
Example 37 (Reference) In dichloromethane (80 m~) was suspended 7~-amino-3-(3-oxobutyryloxy) methyl-3-cephem-4-carboxylic acid (15.7 g). At -10C, triethylamine (10.1 g) was added. To the resultant solution was added a solution of 4-chloro-3-oxo-butyryl chloride (1.41 mMol/g) (44.9 g) dropwise at -20 - -15C over a period of 20 minutes. After the dropwise addition had been completed, the reaction ~' .

~3~3~

was carried out at a telnperature not exceeding -5C for 1 hour. After the reaction, the dichloron~ethcLne was distilled off and the residue ~as dissolved by the addition of tetrahydrofuran (50 mQ), ethyl acetate (100 mQ) and 10%
aqueous phosphoric acid. The resultant solution was separated and the organic layer was taken. The water layer was extracted with a solvent mixture of ethyl acetate-tetrahydrofuran (5:1). The extracts were pooled, washed with a saturated aqueous solution of sodium chloride, dried over magnesium sulfate and filtered. The solven~ was distilled oEf and the residue was treated with ethyl acetate-ether. The resulted powder was recovered by filtration, washed with ether and dried. The procedure provided 7~-(4-chloro-3-oxobutylamido)-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~20.4 g).
IR(KBr): 3325, 1782, 1740, 1732, 1715, 1685, 1650 cm NMR~ in d6-DMSO): 2.14(3H,s), 3.55(4H,s), 3.54(2H,br), 4.44(2H,s), 4.90(2H,ABq,J=13Hz), 5.05(1H,d,J=5Hz), 5.54(1H,q, J=5 ~ 8Hz), 8.96(1H,d,J=8Hz) The dichloromethane solution of 4-chloro-3-oxobutyryl chloride used in th~s reaction was prepared by dissolving diketene (84.0 g) in dichlorometh-ane (420 mQ) and introducing chlorine gas (78.1 g) at -30 - -35C for one hour.
Example 38 (Reference) ~hile a solution of diketene (0.91 g) in dichloromethane (2 m~) was stirred at -40 - -30C, bromine (1.82 g) was added dropwise. Separate- -1~, 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (2.6 g) and triethylamine (1.7 g) were dissolved in dichloromethane (20 mQ) and cooled to -40C. This solution was added to the abov0 reaction mixture.
After stirring for 20 minutes, the mixture was further-stirred under cool-ing with ice. Following the addition of water (7 mQ), phosphoric acid and ethyl acetate~ the mix~ure was stirred vigorously and the organic layer was washed with an aqueous solution of sodium chloride, dried and ~ .

:

decolorized with activated carbon. Th~n, the solvent was distilled of~ and ether was addecl. Tlle procedure provided 7~-~4-bromo-3-oxobutylamido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (2.4 g).
IR(KBr): 1790, 1730, 1645, 1545 cm 1 N~ in d6-DMSO): 2.17(3tl,s,COCH3), 3.60~2H,s,-COC~12CO-), 3.3 - 3.8 ~2H,broad,2-CH2), 4.36~2~l,s,BrCH2CO-), 4.76 ~ 5.06 ~2l-l,ABq,J=12Hz,3-CH2), ';.07~]H,d,J=4.5~z,6-~l), 5.68~1H,dd,J=4.5 ~ 8Hz,7-H), 9.04~1H,d,J=8Hz,-CONH-~
Example 39 ~Re~erence) A solution of diketene ~0.20 mQ) in dichloromethane ~1 mQ) was stirred at -30C, a 1.5 M solution of chlorine in carbon tetrach~oride ~2.0 g) was added dropwise over a period of 10 minutes. The mixkure was further stirred at -25 - -35C for 30 minutes. Separately, 7~-amino-3-~2-carboxy-6 ~or 3)-nitrobenzoyloxyJmethyl-3-cephem-4-carboxylic acid ~0.90 g) and tri-ethylamine (0.84 mQ) were dissolved in dichloromethane (5 mQ) and cooled to -5 --10C. To this solution was added the above reaction mixture dropwise at -20 - -30C over a period of 15 minutes. The mixture was then stirred at ~he same temperature for 45 minutes. Then, the reaction mixture was distilled under reduced pressure and the residue was stirred vigorously with ethyl acetate (25 mQ), tetrahydrofuran ~5 mQ) and 10% aqueous phosphoric acid t20 mQ). The organic layer was washed with a saturated aqueous solution of sodium chloride (15 mQ), dried over sodium sulfate and concentrated under re-duced pressure. Then, upon addition of ether, there was obtained a powder.
This powder was recovered by filtration, washed with ether ~10 mQ) and dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-~4-chloro-3-oxobutylamido)-3-~2-carboxy-6~or3)-nitrobenzoyloxy~methyl-3-cephem-4-carboxylic acid (0.70 g).

.. ~ . ~ .

., , ~ :
'' NMR(~ in d6-DMSO): 3.4 - 3.8(2ll,2-C112), 3.54~2H,s,-C0C~12CO~, 4.52(2H,~,CQC~12-), ~.94 ~, 5~22(2~l~Asq~J=l3}lz~3-cH2)~
5.04(1H,d,J=511z,6-ll), 5.68~1H,dd,J=5 ~ 8 1Iz,7-H), ~' N02 7.7~- 8.5(3H,m, _~ ), 9.04(1~l,d,J=8Hz,-CONH-) Eixample 40 (Reference~ , In acetone [10 m~) was dissolved 7~-t4-chloro-3-oxobukylamido)-3-~3-oxohutyryloxy)methyl-3-cephem-4-carboxylic acid (4.33 g) and, under cooling with ice, water (10 m~) and thiourea (0.84 g~ were added. Then, sodium hydro-gen carbonate (0.84 g~ and water ~10 m~) were further added. The mixture was reacted at room temperature for 5 hours, ater which time it was cooled with ice. The resultant crystals were collected by filtration, washed with water, acetone and ether and dried. The procedure provided 7~-[2-(2-imino-4-thiazolin-4-yl)acetamido~-3-~3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (4.22 g~.
IR(KB ): 1775, 1740, 1710, 1661cm O
NMR~ in d6-DMSO): 2.17(3H,s, -CCH3), 3.38(2H,s,-CH2CONH-~, 3.51(2H,2-CH2), O O
i, 11 .
3.59~2H,s, -CCH2C-), 4.77 ~ 5.06(2H, ABq,J=13Hz,3-CH2), ~
5.05~1H,d,J=5Hz,6-H), 5.69~1H,dd,J=5 ~ 9Hz,7-H), 6.23 ~` -~lH,s,thiazoline-H), 8.82~1H,d,J=9Hz,-CONH-) Example 41 ~Reference) In dimethylformamide ~3.5 m~) was dissolved 7B-[2-~2-imino-4-thi-azolin-4-yl)acetamido]-3-hydroxymethyl-3-cephem-4-carboxylic acid sodium salt ~1.37 g), followed by the addition of O-carboxymandelic anhydride ~0.90 g).
The mixture was stirred at room temperature for one hour, after which time most of the~dimethylformamide was distilled off under reduced pressure. To the re-sidue was added ethyl acetete ~50 mQ), followed by vigorous stirring. The re-sultant powder ~as recovered by filtration, washed with ethyl acetate ~20 m~j, .... .
~ ~, . - ~ . ', :: : : - .
, ~
:: :
- : : .. .. . .

3~

dichloromethane (20 mQ) and ether (20 mQ) in the order mentioned. The pro-cedure provided 7~-[2-(2-imino-4-~hiazolin-4-yl)acetamido]-3-mandelyloxy-methyl-3-cephem-4-carboxylic acid (1.60 g).
IR~KBr): 1780, 1743, 1665, 1643, 1537 cm NMR(~ in d6-DMSO~D20): 3.33 ~ 3.65(2H,ABq,J=18Hz,2-CH2), 3.37~2H,s,-CH2C0-), 4.8 - 5.3(2H,3-CH2), 4.97~1H,d,J=5Hz,6-H), 5.21~1H,s, ~ - C_-), 5.64(1H,d,J=5Hz,7-H), 6.25(lH,s, thiazolin-H), 7.2 - 7.6(5H, - ~ ) Example 42 ~Reference) In N,N-dimethylformamide (20 mQ) was dissolved 7~-~2-thieny]acet-amido)-3-hydroxymethyl-3-cephem-4-carboxylic acid sodium salt ~3.86 g) and, at -5C, triethylamine ~1.40 mQ) and diketene ~1.50 mQ) were added. The re-action was carried out at -5 - 0C for 1 hour, after which time the reaction mixture was poured in ice-water (200 mQ). The mixture was adjusted to pH 2.0 with 4N-hydrochloric acid and extracted with ethyl acetate. The ethyl acetate layer was rinsed with water, diluted with water, brought to pH 7.0 with 5%
aqueous sodium hydrogen carbonate solution and separated. The water layer was taken, concentrated and subjected to column-chromatography on Amberlite XAD-2, elution being carried out with water-methanol. The eluate was lyophilized.
The above procedure provided 7~-(2-thienylacetamido)-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid sodium salt (4.48 g).
IR~KBr): 3300, 1765, 1745, 1670, 1613cm NMR(~ in D2O~: 2.31(3H,s), 3.47(2H,ABq,J=18Hz), 3.85(2H,s,), 4,88~2H, ABq,J=13Hz), 5.08~1H,d,J=5Hz), 5.60~1H,d,J=5Hz), 6.9 - 7.5~3H,m) Example 43 (~eference) In dichloromethane (50 mQ) was dissolved 7~-phenylacetamido-3-.,, ~

~ . . .. .
i ~- . , .
:

~3~3~

hydroxymethyl-3-cephem-4-carboxylic acid triethylamine salt (4.5 g), ollowed by the addition of succinic anhydride (1.5 g). The mix~ure was stirred at room temperature for 5 hours, after which time the solvent was distilled off under reduced pressure. l`hen, Eollowing the addition of water and ethyl acetate, the residue was adjusted ~o pH 2.0 with phosphoric acid. The ethyl acetate layer was dried and concentrated. I'he resultant crystals were collect-ed by filtration (2.9 g). The mother fluid was further concentrated and, after the addition of ether, the concentrate was allow0d to stand, whereupon crystals (1.6 g) were obtained. These crystals were recrystallized from ethyl acetate.
The procedure provided 7~-phenylacetamido-3-(3-carboxypropionyloxy)-methyl-3-cephem-4-carboxylic acid, melting point: 86 - 89C.
IR~KBr): 1800, 1735, 1692, 1660cm NMR~ in d6-DMSO): 2.48(4H,-(CH2)2-), 3.51(4H,2-CH2,-CH2CO-), 4.69 $
5.02(2H,ABq,J=13Hz), 5.00(1H,d,J=5Hz,6-H), 5.63(1H, dd,J=5 $ 9Hz,7-H), 7.23(5H, ~ ), 9.02~1H,d, J=9Hz,-CONH-).
Example 44 (Reference) The reaction procedure of Example 43 was repeated except that ~ phthalic anhydride (2.2 g) was used in place of succinic anhydride. This pro-cedure pro~ided 7~-phenylacetamido-3-(2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid. Melting point: 128 - 123C (ethyl acetate-ether) IR~KBr): 1788, 1731, 1695, 1662cm NMR(~ in d6-DMSO): 3.53(2H,s,-CH2CO-), 3.61(2H,2-CH2), 4.90 ~ 5.27 (2H,ABq,J=13Hz,3-CH2), 5.08(1H,d,J=5Hz,6-H), 5.68(1H,dd,J=5 ~ 8Hz,7-H), 7.25(5H, ~ ), 7.62(4H, ~ ), 9.07(lH,d,J=8Hz,-CONH-) ~; - 43 -:
' ~ 3~

Ex~mplo 45 (Reference~
Dichloromethane ~20 mQ) was added to a mixture of 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (3.14 g) and diemthylacetamide (1.8 g~. Then, under ice-cooling and stirring, phenoxyacetyl chloride (1.8 g) was added. The mixture was stirred for 1 hour, after which the insolubles were filtered off and the filtrate washed w:lth an aqueous solution of sodium chloride. The organic layer was extracted with aqueous sodium hydrogen car-bonate solution. The water layer was made acidic with phosphoric acid and extracted with ethyl acetate. The extract was washed with aqueous sodium chloride solution, dried and distilled to remove the solvent. To the residue was added ether and the resultant powder was recovered by filtration and washed with ether.
The procedure provided 7~-phenoxyacetamido-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid.

IR(KBr): 1788, 1722cm o MMR~ in d6-DMSO): 2.18~3H,s,CCH3), 3.60(2H,2-CH2), 3.66~2H,s,-CCH2C- ), O O
4.67(2H,s,-OCH2-), 4.99(2H,3-CH2), 5.18(1H,d,J=5Hz, 6-H), 5.78(1H,dd,J=5 ~ 8Hz,7-H), 6.8 - 7.7(5H,m), 9.07~lH,d,J=8Hz) Example 46 ~Reerence) In dichloromethane (5 mQ) was suspended 7~-amino-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid ~314 mg) and, at -10C, triethyIamine (0.28 mQ) was added. To the resultant solution was added diketene ~0.2 mQ), and the reaction was carried out at a temperature not exceeding 0C for 2 hours.
After this reaction, the dichloromethane was distilled off and the residue was dissolved in water-ethyl acetate, adjusted to pH 2.0 with 4N-hydrochloric acid and separated. The ethyl acetate layer was taken, diluted with water, ..r ' ` .

,:;

f~ 3~

adjusted to p~l 7.0 with 5% aqueous sodium hydrogen carbonate, and separated.
Tlle water layer was taken, concentrated and subjected to column-chromato-graphy on Amberlite XAD-2, elution being carried out with water-methanol.
The eluate was lyophilized. The above procedure provided 7~-(3-oxobutylamido)-3-~3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid sodium salt 375 mg).
IR(KBr): 3320, 1745, 1660, 1610cm 1 NMR(~ in D2O): 2.27~6H,s), 3.55(2H,ABq,J=18Hz), 4.93(2H,ABq,J=13Hz), 5.15(lH,d,J=5Hz), 5.70(lH,d,J=5Hz) Example 47 (Reference) In acetonitrile (5 mQ) was dissolved 7~-(4-bromo-3-oxobutylamido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid t2.4 g), followed by the addition of thiocarbamic acid O-methyl ester (0.6 g). The mixture was stirred at room temperature overnight and the resultant crystals were recovered by filtration. The procedure provided 7~-[2-(2-oxo-4-thiazolin-4-yl)acetamido]-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid.
IR(KBr): 1780, 1722, 1675, 1629cm NMR(~ in d6-DMSO~: 2.17(3H,s,-COCH3), 3.32(2H,s,-CH2-CONH), 3.41 ~T
3.64(2H,ABq,J=18Hz,2-H), 3.58(2H,s,-COCH2CO-), 4.75 ~7 5.06(2H,ABq,J=13Hz), 5.06(1H,d,J=4.5Hz,6-H), 5.68(1H,dd,J=4.5 ~T 8Hz,7-H), 5.99(1H,s,thiazolin-H~, 8.94(1H,d,J=8Hz,-CONH-), 11.06(1H,thiazolin-NH) Example 48 (Reference) In dichloromethane (7 m~) was suspended 7~-amino-3-(3-carboxy-prop~on~loxy3meth~1-3-cephem-4~carboxylic acid (660 mg), followed by the ad-dition o~ N,N-dimethylacetamide (0.7 m~). Then, under ice-cooling and stir-ring, a solution of (:LH-tetrazol-l-yl)acet~l chloride (294 mg) in dichloro-methane (2 m~) was added. The mixture was stirred at room temperature for 1 hour, after which time it was poured in an aqueous solution of sodium ~ ~.
~. ................................................................... .

~ , . .

~''' . ' " : ' . ' ' ~ : , ,.' . -3~

hydrogen carbonate and separated. The water layer was taken, washed with dichloromethane, made acidic with phosphoric acid and extracted with ethyl acetate. The extract was re-extracted with aqueous sodium hydrogen carbonate solution to bring the desired compound into the aqueous phase. This aqueous solution was purified by column-chromatography on Sephadex LH-20. The frac-tions containing the dominant product were pooled and lyophilized. The pro-cedure provided 7~-[2-(lH-tetrazol-l-yl)acetamido]-3-(3-carboxypropionyloxy)-methyl-3-cephem-4-carboxylic acid disodium salt.
IR(K~r): 1765, 1620cm 1 ~ N~m(~ in D20): 2.60(4H,m-(CH2)2-), 3.44 ~ 3.74~2ll,ABq,J=l7Hz,2-CH2), 5.20(1H,d,J=5Hz,6-H), 5.59(2H,s,NCH2C0-), 5.76(1H,d, J=5~1z,7-H), 9.33(1H,s,t-~trazol-H) Example 49 (Reference) In dichloromethane (10 mQ) was suspended 7~-amino-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (942 mg), followed by the addition of N,N-dimethylacetamide (1 mQ). Then, under ice-cooling and stirring, a solution of ~lH-~etrazol-l-yl)acetyl chloride (441 mg) in dichloromethane (3 mQ) was added. The mixture was further stirred at room temperature for 30 minutes, after which it was poured in an aqueous solution of sodium hydrogen carbonate.
The water layer, i.e. aqueous extract, was purified by column-chromatography on Sephadex LH-20. The fractions rich in the desired product were pooled, concentrated, made acidic with phosphoric acid and extracted with ethyl acetate. The extract was dried, concentrated and treated with ether. The procedure provided 7~-[2-(lH-tetrazol-l-yl)acetamido]-3-~3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid.

IRCKBr): 1782, 1707cm NMRC~ in d6-DMSO): 2.17(3H,s,-CH3), 3.55(2H,broad,2-CH2), ~, .

- .

3~3~

o o Il ~
3.~9(2H,5, -CCH2C-), 4.78 ~, 5.08(2~1,ABq,~=1311z, 3~C~12), 5.09(1H,d,J=5Hz ,6-1-1), 5.34(211,s,NC~12C0-), 5.71(111,dd,J=S ~ 8Hz,7-ll), 9.28(1H,s,tetrazol-H), 9.46(1H,d,J=8Hz,-C()NH-`I
Example 50 (Reference) In dry tetrahydrofuran (30 mQ) was dissolved D~ t-butoxycarbonyl-amino-c~.-(p-hydroxyphenyl)acetic acid (2.68 g), followed by the addition of 2,6-lutidine (1.08 g). While cooling at -:L0C and stirring, ethyl chloro formate (1.08 g) was gently added and the mixture was stirred at -10C for 10 20 minutes. An ice-cooled mixed solution of 7~-amino-3-(3-oxobutyryloxy)-methyl-3-cephem-4-carboxylic acid (3.14 g) and sodium hydrogen carbonate (1.0 g) in water ~30 mQ) was added in a single dose to the above mixture.
The entire mixture was stirred at -3C for 10 minutes and, then, under cool-ing with ice-water, for 2 hours. Following the addition of water (60 m~), the mixture was washed with ethyl acetate ~50 m~) and, under stirring in the presence of ethyl acetate (100 m~), 50% phosphoric acid was gently added so as to bring the pH to 3Ø The ethyl acetate layer was taken, rinsed with water ~100 mQ), dried o~er anhydrous sodium sulfate and treated with a 2N-solution of sodium 2-ethylhexanoate in isopropyl alcohol (6 m~). The result-20 ant precipitate was taken by d~cantation, loosened with ethyl acetate9 re-covered by filtration and dried under reduced pressure over phosphorus pent-oxide. The brown powder thus obtained was purified by column-chromatography on Amberlite XAD-2. The procedure provided 7~-[D-cL-t-butoxycarbonylamino-c~-~p-hydroxyphenyl)acetamido]-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid sodium salt.
IRCKBr): 34ûû, 177a, 168Q, 1610cm N~iRC~ in D2O~: 1.45~9H,s), 2.30~3H,s), 3.43(2H,broad), 5.65(1H,d, ~=5Hz,7-H~, 6.76 - 7.40(4H,m) ~"

..

~ 3~3~

Example 51 -In water (0.3 m~) was dissolved 7~-(D-~-sulEophenylacetamido~-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid disodium salt (278 mg), together with pyridine (60 mg) and KSCN (1.2 g~. The mixture was heated at 60C for 1 hour, after which it was subjectod to column-chromatography on Amberlite XAD-2, elution being carried out ~with water. The fraction rich in the desired product were pooled, lyophilized and recrys~allized ~rom methanol.
The procedure provided 7-(D-~-sulfophenylacetamido)-3-cephem-3-pyridinium-methyl-4-carboxylate sodium salt.
IR~KBr): 1760, 1665, 1610cm NMR(~ in D2O): 2.97, 3.35(2H,ABq,J=18HzJ2-CH2), 5.27, 5.40(2H,3-CH2), 5.07(1H,d,J=5.2Hz,6-H), 5.71(1H,d,~

J=5.2Hz,7-H), 5.10(1H,s, ~ - CH- ), 7.47(5H,m), 8.04, 8.55,8.90(5H, + N ~ )' Example 52 The reaction procedure of Example 51 was repeated using isonicotin-amide ~90 mg~ in place of pyridine and the reaction product was treated in the same manner as Example 51. The procedure provided 7-(D-~-sulfophenyl-acetamido)-3-cephem-3-~4-carbamoylpyridinium)methyl-4 carboxylate sodium salt.
It was purified ~y recrystallization from ethanol-water. Melting point:
175C Cdecomp.) IRCKBr~: 1765, 1692, 1645, 1615, 1029cm NMRC~ in D20): 2.99, 3.56(2H,ABq,J=18Hz,2-CH2), 5.40,5.51(2H,3-CH2), 5.13~1H,d,J=4.8Hz,6-H), 5.73(1H,d,J=4.8Hz,7-H), 5.10ClH,s, ~ -Cl- ), 7.40(5H,m), 8.31, 9.07(4H) S03Na . .
,, .. , ~ , . :

Fxamp c 53 (R~`arcnce) In 50% aq~leo~s ace-tone (8 mR,) was clissolved 7~-(2-thienylacetamido~-3-mandelylo~ymethyl-3-cephem-4-carboxylic acid (0.49 g) together with acetyl-acetone (0.50 g~ and sodium hydrogen carbonate (0.17 g), and the solution was stirred at 60C for 1 hour. The reaction mixture was brought to room tem-perature and most o-~ the acetone was distilled off lmder reduced pressure.
To the residue was added 5% aqueous phosphoric acid solution (10 mQ), to-gether with ethyl acetate ~20 mQ). The ethyl acetate layer was washed with a saturated aqueous solution of sodium chloride ~20 mQ), dried over ~agnesium sulfate and concentrated under reduced pressure. Then, upon addition of ether, there was obtained a powder. This powder was suspended in water ~3 mQ) and dissolved by the addition of sodium hydrogen carbonate. Ihe solution was subjected to column-chromatography on Sephadex LH-20, elution being carried out with water. The desired fractions were pooled and lyophilized. The pro-cedure provided 7~-~2-thienylacetamido)-3-(2-acetyl-3-oxo)butyl-3-cephem-4-carboxylic acid sodium salt (0.28 g). This product was dissolved in water ~10 mQ), followed by the addition of 10% aqueous phosphoric acid ~2 mQ) and ethyl acetate ~20 m~). The ethyl acetate layer was rinsed with water, dried over magnesium sulfate and concentrated under reduced pressure. Then, upon 2Q addition o~ ether, there was obtained a powder. This powder was recovered by ~iltration, Nashed with ether and dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-(2-thienylacetamido)-3-(2-acetyl-3-oxo)-butyl-3-cephem-4-carboxylic acid ~0.23 g).
IR~KBr): 1765, 1718cm NMRC~ in d6-DMSO): 2.13 ~ 2.18(6H,s,(C~CH3)2), 2.6 - 3.1~2H,m,3-CH2), 3.31 ~ 3.56(2H,ABq,2-CH2), 3.75(2H,s,-CH2CO-), 4.14ClH,-CH-), 5.01(1H,d,J=5Hz,6-H), 5.58(1H,dd, ~=5 ~ 8Hz,7-H), 6.92 ~ 7.30(3H, - 49 _ ~,, 3~

9.04(1ll,d,J~8l~ CONII-) Lxample 54 ~Re~erence) In 50% aqueous acetone (40 mQ) was dissolved 7~-(2-thienylacetamido)-3-[2-car~oxy-6 ~or 3~ -nitrobenzoyloxy]methyl-3-cephem-4~carboxylic acid (2.65 g) together with acetylacet~ne (2.42 g) and sodium hydrogen carbonate (1.22 g).
The mixture was stirred at 60C for 1 hour, after which time it was treated by a procedure similar to that described in Ex~ample 53. The procedure yrovided 7~-(2-thienylacetamido)-3-(2-acetyl-3-oxo)butyl-3 cephem-4-carboxylic acid ~1.26 g). In IR and NMR spectra, this product was in good agreement with the product according to Example 53.
Example 55 (Reference) In 50% aqueous acetone (8 mQ) was dissolved 7~-[D-5-(p-t-butylbenz-amido)-5-carboxyvaleramido]-3-[2-carboxy-6 (or 3)- nitrobenzoyloxy]methyl-3-cephem-4-carboxylic acid (0.73 g) together with acetylacetone (0.50 g) and sodium hydrogen carbonate ~0.34 g). The mixture was stirred at 60C for 1 hour, after which it was treated by a procedure similar to that described in Ex~mple 53.
The procedure provided 7~-[D-5-~p-t-butylbenzamido)-5-carboxyvaler-amido]-3-C2-acetyl-3-oxo)butyl-3-cephem-4-carboxylic acid ~0.29 g).
IR(KBr): 1767, 1721, 1655, 1635cm NMRC~ ln d6-DMSO~: 1.3~(9H,s,-C(CH3)3), 1.76 ~ 2.25~6H,-~CH2)3-), 2.6 - 3.1~2H,m,3-CH2), 3.2 - 3.6~2H,2-CH2), 4.13ClH,-CH~COCH3)2), 4.37(lH,-CH-), 4.99~1H,d, J=5Hz,6-H), 5.57~1H,dd,J=5 ~ 8Hz,7-H), 7.43 ~
7.83C4H, ~ + ), 8.40~1H,d~J=8Hz,-C~1-NH-), 8.77~lH,d,J=8Hz,-CONH-) ~., x m~e 56 ~l~efcrerlce) In 50~ aqueous acetone (8 mQ~ was dissolved 7~-[D-5-(p-t-'butylbenz-amido)-5-carboxyvaleramido]-3-mandelyloxymethyl-3-cepllem-4-carboxylic acid (0.67 g), together with acetylacetone (0.50 g) and sodium hydrogen carbonate ~0.25 g) The mixture was stirred at 60C for 1.5 hours, after which it was treated by a procedure similar to that described in Example 53. The procedure provided 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido~-3-(2-acetyl-3-oxo~-butyl-3-cephem-4-carboxylic acid (0.33 g). In IR and NMR spectra, this pro-duct was in agreement with the product o'btained in Example 55.
Example 57 (Reference) In 50% aqueous acetone (l~t mQ) was dissolved 7~-[D-5-(~-k-butyl-benzamido)-5-carboxyvaleramido]-3-[2-carboxy-6 (or 3) -nitrobenzoyloxy]methyl-3-cephem-4-carboxylic acid (0.73 g), pyrrole (0.20 g) and sodium hydrogen carbonate (0.25 g). The mixture was stirred at 60C for 1 hour, after which it was brought down to room temperature and most of the acetone was distilled off under reduced pressure. Then, 5% aqueous phosphoric acid solution (15 mQ) and ethyl acetate (30 mQ) were added. The ethyl acetate layer was washed with a saturated aqueous solution of sodium chloride (20 mQ), dried (over magnesium sulfate) and concentrated under reduced pressure. Then, upon addition of ether, there was obtained a powder. This powder was suspended in water (3 mQ) and dissolved by the addition of sodium hydrogen carbonate (0.17 g). The solution was subjected to column-chromatography on Sephadex LH-20 (250 mQ), elution being carried out with water. The fractions containing the desired product were pooled and lyophilized. The above procedure provided 7~-~D-5-~p-t-butylbenzamido)-5-carboxyvaleramido]-3-(2-pyrrolyl)methyl-3-cephem-~-carboxylic acid disodium salt (0.29 g).
IR~KBr): 1760, 1600cm NMR~ in D20): 1.23~9H,s,-C~CH3)3), 1.86 ~ 2.~2~6H,-~CH2)3-), 2-86 ":., .

3.25(211,ABtl,J=181lz,2-C112), 3.42 ~1 3.73(21-1,ABq,J=15~lz, 3-C112~, 4.51(1~1~-CII-), 4.96(1~ l,J=5~1z,6-~1), 5.61(11l, d,J=5~-1z,7-H), 5.93(111,pyrrole--3-~1), 6.07(111,pyrrole-4-1l), 6.77(111,pyrrole-5-H), 7.38 ~ 7.76~4H, ~+ ), 7.80(1~1,pyrrole-1-H~.
Example 58 (Reference) In 50% aqueous acetone (14 mQ,) was dissolved 7~-[D-5-(p-t-butyl-benzamido)-5-carboxyvaleramido]-3-[2-carboxy-6 ~or 3) -nitrobenzoyloxy]-methyl-3-cephem-4-carboxylic acid ~0.73 g~, together with N-methylpyrrole (0.24 g) and sodium hydrogen carbonate ~0.25 g). ï'he mixture was stirred at 60C for 1 hour. After the reaction had been completed, the mixture was treated in the same manner as Example 57. The procedure provided 7~-[D-5-~p-t-butylbenzamido)-5-carboxyvaleramido]-3-~N-methylpyrrol-2--yl)methyl-3-cephem-4-carboxylic acid disodium salt ~0.21 g).
IR(KBr): 1757, 1597cm NMR~ in D20): 1.21~9H,s,-C~CH3)3), 1.87 F7 2.45(6H,-(CH2)3-), 2.79 Fr 3.07(2H,ABq,J=18Hz,2-CH2), 3.46~3H,s,N-CH3), 3.51 ~
3.87(2H,ABq,J=15Hz,3-CH2), 4.51~1H,-CH-), 4.91~1H,d, J=5Hz,6-H), 5.58~1H,d,J=5Hz,7-H), 5.86~1H,pyrrole-3-H), 2Q 5.99~1H,pyrrole-4-H), 6.61(lH,pyrrole-5-H), 7.39 7.79~4H,~+) Example 59 ~Reference) In 50% aqueous acetone ~12 m~) was dissolved 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-mendelyloxymethyl-3-cephem-4-carboxylic acid ~0.64 g), together with indole (0.35 g) and sodium hydrogen carbonate ~0.17 g). The mixture was stirred at 60C for 1 hour. Following the completion of the re-action, the mixture was treated by a procedure similar to tha~ describe in : ~ -:: -~3~;~6 Example 57. The procedure provided 7~-(D-5-phthalimiclo-5-carboxyvaleramido)-3-(3-indolyl)methyl-3-cephem-~l-car~oxylic acid disodium salt (0.24 g).
IR(KBr): 1758, 1702, 1600cm MMR(~ in D20): 1.68 ~ 2.26(6H,-(C~l2)3-), 2.~5 ~1 2.82(2H,ABq,J=18Hz, 2-CH2), 3.61 ~ 3.86(2H,ABq,J=15Hz,3-CH2), 4.6 - 4.9 (2H,-CH- ~ 6-H), 5.~6(1H,d,J=5Hz,7-H), 7.0-7.8(10H, T ~ ~ N-Bxample 60 (Reference) In 50% aqueous acetone (14 mQ) was dissolved 72-(D-5-phthalimido-5-carboxyvaleramido)-3-[2-carboxy-6 (or 3)- nitrobenzoyloxy]methyl-3-cephem-4-carboxylic acid (0.70 g), together with indole (0.35 g) and sodium hydrogen carbonate ~0.25 g). The mixture was stirred at 60C for 45 minutes and, after the reaction had been completed, it was treated by a procedure simllar to that described in Example 57. The procedure provided 1~-(D-5-phthalimido-5-carboxyvaleramido)-3-~3-indolyl)methyl-3-cephem-4-carboxylic acid disodium salt C0.22 g). In IR spectrum, this product was found to agree with the product obtained in Example 59.
Example 61 (Reference) In water (7 mQ) was dissolved 7~-[D-5-(p-t-butylbenzamido)-5-car-boxyvaleramido~-3-[2-carboxy-6 (or 3)- nitrobenzoyloxy]methyl-3-cephem-4-carboxylic acid (0.73 g)~ sodium azide (0.26 g) and sodium hydrogen carbonate ~0.25 g). The mixture was stirred at 60C for 40 minutes. The reaction mix-ture was brought to room temperature and, following the addition of 10% aque-ous phosphoric acid (10 mQ), extracted with ethyl acetate ~30 mQ). The eth~l acetate layer was washed with a saturated solution o~ sodium chloride C20 m~, dried over magnesium sulfate and concentrated under reduced pressure.
'~

- 53 ~
~,.

: ` .

3~

'I'hen, upon addition of ether, th~re was obtained a powder. 'I'his powder was suspended in water (3 m~) and dissolved by the addition of sodium hydrogen carbonate (0.17 g). This solution was subjected to column-chromatography on Sephadex LH-20 (250 mQ), elu-tion being carried out with water. The fractions containing the desired product were pooled and lyophilized.
The procedure provided 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaler-amido]-3-azidomethyl-3-cephem-4-carboxylic acid disodium salt ~0.36 g).
IR~KBr): 2100, 1766, 1606cm NMR(~ in D20): 1.32(9H,s,-C(CH3)3), 1.92 ~ 2.48(6H,-(Cll2)3-), 3.05 ~
3.53 (2H,ABq,J=18Hz,2-CH2), 4.03 ~ 4.20(2H,ABq,J=13Hz, 3-CH2), 4.52(1H,-CH-), 5.08(1H,d,J=5Hz,6-H), 5.68(1H, d J=5Hz 7-H) 7.56 ~ 7.86(4H, ~9~, Example 62 In phosphate buffer (40 m~) of pH 6.4 was dissolved 7-[2-(2-imino-4-thiazolin-4-yl)acetamido]-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~908 mg), together with a nitrogen-containing heterocyclic thiol (2.2 mMol) and sodium hydrogen carbonate (336 mg). The solution was stirred at 60C for 1 hour, after which it was concentrated under reduced pressure to about 20 mQ. The concentrate was subjected to column-chromatography on Amberlite XAD-2, elution being carried out with water, 5% ethanol and 10%
ethanol in the order men~ioned. The ~ractions containing the desired product were pooled and lyophilized to obtain the corresponding one of the following compounds. The reaction yield figure was the value determined by liquid chromatography immediately following the reaction.
~1) 7~[2-~2-Imino-4-thiazolin-4-yl)acetamido]-3-(2-carboxymethyl-1,3, 4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid disodium salt; ' yield 85%
IRCKBr); 1761cm . ~

3~3~

NMR(~ in D20): 3.56 l'l 3.92~2l-1,ABq,J=1811z,2-C~12), 3.76(2~1,s,-CH2CO), 4.16(2~1,s,-CI-12C0), 4.20 ~7 4.62~2~1,ABq,J=1311z,3-CH2), 5.24(1~1,d,J=511z,6-11), 5.79(111,d,J=51-1z,7-H), 6.65(11 s,thiazolin-5-H).
(2)7-[2-(2-Imino-4-thiazolin-4-yl)acetamido]-3-(3-hydroxymethyl-4-methy~l-1,2,4-triazol-5-yl)thiome~hyl-3-cep}lem-4-carboxylic acid sodium salt;
yield 82%.
IR~KBr): 1760cm NMR(~ in D2O): 3.40 ~ 3.82(2H,ABq,J=18Hz,2-CH2), 3.62~2H,s,-CH2C0), 3.74(3H,s,-CH3), 3.72 ~ 4.34~2H,ABq,J=1311z,3-C112), 4.82(2H,s,CH20H), 5.08(1H,d,J=5Hz,6-H), 5.64(1H,d, J=5Hz,7-H), 6.52(1H,s,thiazolin-5-H).
Example 63 ~i, In 50 mQ of water was dissolved 5.61 g of 7-[D-5-(benzamido)-adipinamido]-3-(3-oxobutyryloxy)me~hyl-3-cephem-4-carboxylic acid, together with 1.50 g of 5-mercapto-1-methyl-lH-tetrazole and 2.20 g of sodium bi-carbonate. After the pH was adjusted to 5.2, the reaction was carried out at 6QC for 50 minutes. After cooling, 100 mQ of a saturated aqueous solution ~--of sodium chloride was added and the pH was adjusted to pH 1.5 with 4N-HCQ.
20 The solid precipitate was recovered by filtration~ rinsed with 20 mQ of a saturated aqueous solution of sodium chloride and dissolved in 100 mQ of ethyl acetate-tetrahydrofuran (2:1) and 20 mQ of water. The organic layer was dried and the solvent was distilled o~f under reduced pressure. To the re-sidue was added ether-ethyl acetate and the resultant powder was recovered by ;~
filtration, rmsed with ether and dried. The procedure provided 5.45 g (yield 94.8%) of 7-[D-5-benzamido-5-carboxyvaleramido] -3-(1-methyl-lH- ;
tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid.
IR~Br~ 3340, 1783, 1730, 1645, 1535cm ~ .:

.
~:

, 3~

NMR(~ in d6-DMSO~: 1.50 - 2.0(411,m), 2.05 - 2.45(211,m), 3.70(2HJbroad, 3.93(3H,s,N CH3), 4.15 - 4.55(3H,m), 5.10(111,d,J=5Hz, 6-1l), 5.66(1il,dd,J-5 ~ 9Hz,7-H), 7.32 - 7.97[5H,m, ~ ), 8.43(1~1,d,J=811z,-CONII-), 8.73(1H,d, J=9Hz,-CONII-) Example 64 In 50 mQ of water was d:issolved 6.11 g of 7-[D-5-(p-toluenesulfon-amido)adipinamido]-3-(3-oxobutyryloxy)me~hyl-3-cephem-4-carboxylic acid, to-gether with 1.50 g of 5-mercapto-1-methyl-lH-tetrazole and Z.20 g of sodium bicarbonate. After the solution was adjusted to pH 5.0, the reaction was con-ducted at 60C for 50 minutes. Following the reaction, the reaction mixture was treated in the same manner as Example 63. The procedure provided 5.96 g ~yield 95.1%) of 7-[D-5-~p-toluenesulfonamido)-5-carboxyvaleramido]-3-(1-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid.
IR~KBr): 3275, 1780, 1727, 1635, 1535cm NMR~ in d6-DMSO): 1.45 - 1.78(4H,m), 2.0 - 2.3(2H,m), 2.41(3H,s,-CH3), .
3.71~2H,broad,2-CH2~, 3.95~3H,s,-NCH3), 4.28(2H,broad), 5.06(1H,d~J=5Hz,6-H), 5.62(1H,dd,J-5.~ ~ 9.OHz,7-H), 7.47(4H,m, ~ ), 7.91~1H,d,J=9.OHz), 8.68~1H,d, J=9.OHz) Example 65 In 50 mQ o ~ater was dissolved 6.17 g of 7-[D-5-~p-tert-butyl-benzamido)-5-carboxyvaleramido]-3-(3-oxobutyryloxy)methyl-3-cephem-4-car-boxylic acid, together with 1.74 g of 5-mercapto-1-methyl-lH-tetrazole and 2.50 g of sodium bicarbonate. After the solution was adjusted to pH 5.0, the reaction was conducted at 60C for 50 minutes. Following the reaction, the mixture was treated in the same manner as Example 63 to obtain 6.02 g : :
, ~ ~ . . . : ::

:a ~ 9~

(yield 95.3%) of 7-(D-5-(p-tert-buty1bcnzamido)-5-carboxyvalerclrnido)-3-(l-methyl-lll-tetrazo1-5-y1)thiomethy1-3-cephem-4-carboxy1ic acld.
IR~KBr): 3350, 1780, 1725, 1643, 1532, 1504 cm NMR(~ in d6-DMSO): 1.30(9~1,s,-C(CH3)3), 1.50 - 2.45(6~l,m,-(CH2)3-), 3.64~2H,broad,2-CH2), 3.93(3H,s,-NCH3), 4.27(2H, broad,3-C}12), 4.36(1H,m,-CII-), 5.0(1HJd,J=5Hz,6-H), 5.62~lHIdd,J=5.0 ~7 8.OHz,7-1l), 7.35(2H,d,J=8.OHz), 7.70(2H,d,J=8Hz~, 8.36~LH,d,J=8.0Hz,-CONH-), 8.76~lH,d,J=8.OHz,-CONH-) n ~, "' In 50 mQ of water was dissolved 4.83 g of 7-[D-5-~caprylamido)-5-carboxyvaleramido]-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxrlic acid, to-gether with 1.74 g of 5-mercapto-1-methyl-lH-tetrazole and 250 g of sodium bicarbonate. After the solution was adjusted to pH 5.0 and 35.0 g of sodium bromide was added, the reaction was conducted at 60C for 45 minutes.
Following this reaction period, the reaction mixture was treated in the same manner as Example 63. ~he procedure provided 4.78 g ~yield 96.3%) of 7-[D-5-~caprylamido)-5-carboxyvaleramido]-3-~1-methyl lH-tetrazol-5-yl)thio-methyl-3-cephem-4-carboxylic acid.
2a IR~KBr); 3300, 1775, 1725, 1655, 1640, 1545, 1533cm NMR~ in d6-DMSO): 0.60 - 240~21H,m), 3.70~2H,broad,2-CH2), 3.95~3H,s,NCH3), 4.15~lH,m), 4.27(2H,broad,3-CH2), 5.03~1H,d,J=5Hz,6-H), 5.65~1H,dd,J=5.0 ~ 8.0Hz,7-H), 7.97(lH,d,J=8.OHz,-CONH-), 8~76(lHgd~J=8~OHz~-CONH-) Exam~e 67 In 50 m~ of water was dissolved 4.32 g of 7-phenylacetamido-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid, together with 1.50 g of 5-mercapto-1-methyL-lH-tetrazole and 1.68 g of sodium bicarbonate and the reaction was conducted at 60C for 50 minutes. After cooling, the reaction . - 57 -~3~

mixture was adjusted to p1l 5.0 and washed Wit]l ethyl acetate. It was then brought down to p~l 2.0 and extrac-ted three times with ethyl acetate. rlhe ethyl acetate solution was washed with a saturated aqueous solution of sodium chloride, treated with magnesium sulfate, filtered and distilled under reduced pressure to remove the solvent. The residue was recrystallized from ethyl acetate-ether. The procedure provided 4.29 g (yield 96.2%) of 7-phenylacet-amido-3-(1-methyl-lil-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid.
IR(KBr): 3270, 1785l 1733, 1662, 1628, 1542cm NMR(~ in d6-DMSO): 3.55(2H,s,-CH2CO-), 3.60(2H,broad,2-Cll2), 3.92(3H,s,NCH3), 4.26(2H,broad,3-CH2), 5.00(1ll,d, J=5.OHz,6-l-1), 5.60(lH,dd,J=5.0 ~, 8.0Hz 7 7-}l), 7.23 (5H,s, ~-- ), 8.98(1H,d,J=8Hz,-CONH-) Example 68 In 10 mQ of water was dissolved 561 mg o-f 7-[D-5-benzamido-5-car.-boxyvaleramido]-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid, together with 252 mg of sodium bicarbonate and 212 mg of 2-methylthio-5-mercapto-1,3, 4-thiadiazole. The reaction was conducted at 60C for 50 minutes and, after cooling, the reaction mixture was washed with ethyl acetate and freeze-dried.
The resultant solid was dissolved in a small quantity of methanol and treated 20 with acetone. The crystals formed were collected by filtration and rinsed with ether. The procedure provided 614 mg of 7-[D-5-benzamido-5-carboxyvaler-amido]-3 E2-methylthio-1,3,4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid disodium salt.
IR(KBr): 3400, 1766, 1640, 1600, 1530cm in D20): 1.40 - 2.55(6H,m,-(CH2)3-), 2-68(3H,s,SCH3), 3.33~2H,ABq,J=18Hz,2-CH2), 4.14(2H,ABq,J=14Hz,3-CH2), 4.30~1H,m,-C~H-), 5.01(1H,d,J=4.5Hz,6-H), 5.58~1H,d, , ' ~L~ 3~
~. ~v~

J=4.5llz,7-ll), 7.25 - 7.95(5~-l,m,~

~xample 69 _ In 10 mQ of water was dissolved 561 mg of 7-[D-5-benza~ido-5-car-boxyvaleramido]-5-(3-oxobutyryLoxy)methyl-3-cephem-4-carboxylic acid, to-gether with 252 mg of sodium carbonate and 224 mg of 5-mercapto-2-ekhoxycar-bonylmethyl-lH-1,3,4-triazole. The reaction was conducted at 60C for 50 minutes and, after cooling, the reaction mixture was treated in ~he same manner as Example 68. The procedure provided 642 mg of 7-[D-5-benzamido-5-carboxyvaleramido]-3-(2-ethoxycarbonylmethyl-lH,1,3,4-triazol-5-yl~thiomethyl-3-cephem-4-carboxylic acid disodium salt.
IR~KBr): 3400, 3280, 1765, 1745, 1640, 1603, 1535cm MMR(~ in D20): 1.20~3H,t,J=8.OHz,-CH2CH3), 1.50 - 2.50(6H,m,-(CH2)3-), 3.32(2H,ABq,J=19Hz,2-CH2), 3.80 - 4.50(7H,m), 4.95(1H,d, J=4.5Hz,6-H), 5.52(1H,d,J=4.5Hz,7-H), 7.20 - 7.90 ~5H,m, ~ ).

Example 70 In 12 mQ of water was dissolved 1.27 g of 7~-[D-5-(p-t-butylbenz-amido)-5-carboxyvaleramido]-3-~3-carboxypropionyloxy)methyl-3-cephem-4-car-boxylic acid, together with 0.25 g of 5-mercapto-1-methyl-lH-tetrazole and 0.68 g of sodium hydrogen carbonateg followed by the stirring for one hour and a half at 60C. The reaction solution was allowed to be cooled at the room temperature and added 30 mQ of 4% aqueous solution of phosphoric acid, followed by extracting with 60 mQ of ethyl acetate. The ethyl acetate layer was washed with Q saturated aqueous solution of sodium chloride (40 mQ x 2) and dried over magnesium sulfate, followed by condensation under reduced pressure. The condensate was added ether and the resultant powder was re-covered by ~iltration~ washed with ether and dried over phosphorous pentoxide .

}3~

nder reduced pressure to give l.02 g oE 7~-[D-5-(p-t-butylbenzamldo~-5-car-boxyvaLeramido]-3-(l-methyl-1l-l-tetrazol-5-yl~thiomethyl-3-ccphem-4-carboxylic acid.
In IR and NMR spectra, this product was Eound in good agreement with the product obtained in Example 65.
Example 71 In 12 mQ of water were dissolved 1.26 g of 7~-[D-5-(p-t-butylbenz-amido)-5-carboxyvaleramido]-3-(3-carboxyacryloxy)methyl-3-cephem-4-carboxylic acid, 0.25 g of 5-mercapto-1-methyl-lH-tetrazole and 0.68 g of sod;um hydrogen lQ carbonate, and the solution was stirred for 2 hours at 60C, followed by the treatment in the same manner as Example 70. The procedure provided 0.88 g of 7~-[D-5-(p-t-butylbenzamido~-5-carboxyvaleramido]-3-~l-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid, of which IR and NMR were found in good agreement with that of the product obtained in Example 65.
Example 72 The reaction of Example 71 was repeated employing 1.36 g of 7~-[D-5-~p-t-butylbenzamido)-5-carboxyvaleramido]-3-~2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid. The procedure provided 0.97 g of 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido]-3-(1-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid,of which IR and NMR were found in good agreement wi~h that of the product obtained in Example 65.
Example 73 In 12 mQ of water were dissolved 1.55 g of 7~-[D-5-~p-t-butylbenz-amido)-5-carboxyvaleramido]-3-[3-carboxy-3 ~or 2~-~p-chlorophenylthio~propi-onyloxy]methyl_3-cephem-4-carboxylic acid, 0.25 g of 5-mercapto-1-methyl-lH-tetrazole and 0.68 g of sodium hydrogen carbonate, and the solution was stirred ~or one hour and a hal~ at 60C, followed by the treatment in the same manner as Example 70. The procedure provided 0.99 g of 7~-[D-5-~p-t-' .: :

, . ' ' ' . ;, :::
: .. , 3i~

butyl-benzamido~-5-carboxyvaleramido]-3~ methyl-1l-l~tetrazol-5-yl~thiomethyl-3-cephem-4 carboxylic acid, of which IR and NMR were Eound in good agreement with that of the product obtained in Example 65.
Example 74 The reaction of Example 73 was repeated employing 1.21 g of 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-(3-carboxypropionyloxy)methyl-3-cephem-4-carboxylic acid. The procedure provided 1.00 g of 7~-~D-5-phthalimido-5-car-boxyvaleramido)-3-~1-methyl-lH-te~razol-5-yl)thiomethyl-3-cephem-4-carboxylic acid, of which IR and NMR were found in good agreement with that of the product obtained in Example 1 ~2).
Example 75 The reaction of Example 73 was repeated employing 1.30 g of 7~-(D-5-phthalimido-5-carboxyvaleramido)-3-~2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid. The procedure provided 0.94 g of 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-~1-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-car-boxylic acid, of which IR and NMR were found in good agreement with that of the product obtained in Example 1 ~2).
Example 76 In water ~6 mQ) was dissolved 7~-[D-5-~2-carboxy-6(or 3)-nitrobenz-amido)-5-carboxyvaleramido]-3-~2-carboxy-6(or 3)-nitrobenzoyloxy)methyl-3-cephem-4-carboxylic acid ~0.78 g) together with 5-mercapto-1-methyl-lH-tetra-zole ~0.12 g) and sodium hydrogen carbonate ~0.42 g). The solution was stir-red at 60~C for 30 minutes~ after which ~ime it was treated as in Example 70.
The procedure provided 7~-[D-5-~2-carboxy-6~or 3)-nitrobenzamido)-5-carboxy-valeramido]-3-~1-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid ~0.50 g).
IROKBr): 1782, 1731, 1645, 1537, 1351cm 1 MMR~d6-DMSO): ~ 1.73 ~ 2.26~6H,-~CH2)3-), 3.69(2H,2-CH2), 3.94~3H,s, , , :, '., " ". ' ' ' ' .

3~

, N-C113~, ~1.32(21-1,3-C112), 1.52(11-1,-C1-1-), 5.06(111~d,J=5~1z,6-~1), 5.67(l1-1Jd1JJ-5 Fl 81-1æJ7~i1~, 7.6 - 8.4 (411J~
~.
-C~l-N~I-), 8.79(l11JdJJ=811z,-CON~1-) In water (6 m~) was dissolved 7~-~D-5-(p-t-butyl-benzamido~-5-carboxyvalerclmldo]-3-(2-carboxy-6(or3)-nitrobenzoyloxy~methyl-3~cephem-4-carboxylic acid (0.73 g~ together with 5-mercapto-1-methyl-lH-tetrazole ~0.12 g) and sodium hydrogen carbonate (0.34 g~. The mixture was stirred at 60C
for 30 minutesJ aeter which time it was treated as in Example 70. The pro-lO cedure provided 7~-[D-5-(p-t-butylbenzamido~ -5-carboxyvaleramido]-3-(l-methyl-lH-tetrazol-5-~rl)thiomethyl-3-cephem-4-carboxylic acid (0.53 g~.
In IR(KBr~ and NMR(d6-DMSO~ spectraJ this compound was in good agreement with the product obtained in Example 65.
Example 78 The reaction of Example 76 was repeated employing 0.73 g of 7~-[D-5-~p-t-butylbenzamido~-5-carboxyvaleramido]-3-(2,4(or 5~-dicarboxybenzoyl-oxy)methyl-3-cephem-4-carboxylic acid to give 0.52 g of 7~-[D--5-(p-t-butyl-benzamido)-5-carboxyvaleramido]-3-(l-methyl-l~1-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid. In IR(KBr~ and NMR(d6-DMSO~ spectra, this compound 20 was in good agreement with the product in Exclmple 65.
Example 79 In water ~6 m~ was dissolved 7~-(D-5-phthalimido-5-carboxyvaler-amido~-3-(2-carboxy-6(or 3~-nitrobenzoyloxy~methyl-3-cephem-4-carboxylic acid (3.70 g) together with 5-mercapto-2-methyl-1,3,4-thiadiazole ~0.13 g) and sodium hydrogen carbonate (0.34 g). The solution was stirred at 60C for 30 minutes, after which time it was treated as in Example 65. The procedure provided 7~-~D-5-phthalimido-5-carboxyvaleramido)-3-(2-methyl-l,3,4-thiadiazol-- . ''; ~

: , .

3~

5-yl~thiomethyl-3~cephel7l-4-carboxylic acid (O.Sl g).
IR ~KBr): 1773(sh~, 1715, 1648(sh) cnl 1 NMR(d6-DhlSO~: ~ 1.53 ~7 2.15(6H,-(CH2~3-)~ 2.67~3H,s,-C1l3~ 3.45 ~, 3.72(2H7ABq,J=18}1z,2-CII2), 4.19 F7 4.50(2H,ABq,J~1311z, 3-CH2), ~.72 ~lH,t,Ja7llz,-CH-), 5.02(1}l,d,J=511z,6-H), 5.61(1H,dd,J=5 ~ 8}1z,7-H), 7.87(4}1,s, ~N-8.74 (lH,d,J=8Hz,-CONH-) Example 80 The reaction of Example 79 Was repeated ~mploying 0.73 g of 7~-10 ~D-5-~p-t-butylbenzamido) -5-carboxyvaleramido]-3-~2-carboxy-6tor 3) -nitro-benzoylox~)methyl-3-cephem-4-car7Ooxylic acid to ~ive 0.55 g of 7~-[D-5-(p-t-butylbenzamido)-5-carboxyvaleramido] -3-(2-methyl-1,3,4-thiadiazol-5-yl)thio-methyl-3-cephem-4-carboxylic acid.
IR(KBr): 1780, 1728~ 1644cm NMR~d6-DMSO): ~ 1.28(9H,s,-C ~CH3)3), 1.74 ~7 2-23 ~6H,-~CH2)3-), 2.66~3H,s,-CH3), 3.50 ~7 3.75(2H,ABq,J=18Hz,2-CH2), 4.20 ~7 4.50(2H,ABq,J=l3Hz,3-CH2), 4.39~1H,-CH-), 5.05(1H,d,J=5Hz,6-H), 5.65(1H,dd,J=5 ~7 8Hz,7-H), 7.44 ~7 7.80(4H,~ ), 8.42~1H,d,J=8Hz,-CH-N_-) 8.80(1H,d,J=8Hz,-CONH-) Example 81 ln water (5 m~) was dissolved 7~-~D-mandelamido)-3-(3-carboxy~
propionyloxy)methyl-3-cephem-4-carboxylic acid (0.46 g) together with 5-mer-capto-l-methyl-lH-tetrazole (0.12 g) and sodium hydrogen carbonate (0.25 g).
The solution was stirred at 60C for 1 hour and a half. After cooling in the air, the reaction mixture was subjected to column chromatography on Amberlite XAD-2, elution being carried out with water and, then, with a solvent mixture .
" ~ ' , .', ~ ' ' ., ., ' ' ` ' ' ' ' , - . .
:: ;
. : . .
.

$

oE water and methanoL. Tlle Eractions contalning the desired compound are pooled, concentrated and lyophilized. I'he procedure provided 7~-(D-mandel-amido)-3-(1-Tnethyl-lll-tetrazol-5-yl)thlomethyl-3-cephem-4-carboxylic acid sodium salt (0.31 g) IR(KBr): 1761, 1675, 1604cm NMR~D20): ~ 3.25 ~ 3.68(2H,ABq,J=18~1z,2-C1l2), 3~95(3H,s,,N-CH3), 4-02 f'T 4.29(2H,ABq,J=1311æ,3-Cll2), 4.97(1H,d,J=511z,6-H), 5.18(1H,s,-CH-), 5~51(11-1,d,J=51lz,7-H), 7.37(51l,s, ~_) In water (5 m~) was dissolved 7~-(2-thienylacetamido)-3-(2-carboxy-benzoyloxy)methyl-3-cephem-4-carboxylic acid (0.50 g) together with sodium hydrogen carbonate (0.17 g), potassium iodide (0.40 g) and pyridine (0.21 g).
The solution was adjusted to pH 6.5 and, then, stirred at 60C Eor one hour and a half. After cooling in the air, the reaction mixture was subjected to column chromatography on Amberlite XAD-2, elution being carried out with water and, then, with a solvent mixture of water and methanol. The fractions con-taining the desired product were pooled, concentrated and lyophilized.
The procedure provided 7~-(2-thienylacetamido)-3 (l-pyridylmethyl)-3-ceiphem-4-carboxylic acid betaine (0.23 g).
IR~KBr): 1763, 1698, 1617 cm NMR~D20): ~ 3.17 ~ 3.67~2H,ABq,J=17Hz,2-CH2), 3.38~2H,s,-CH2CO-), 5.l9(1H,d,J=5Hz,6-H), 5.41 ~ 5.67~2H,ABq,J=l'lHz,3-C112), 5.75(1H,d,J=5Hz,7-H), 7.01 ~ 7.28~3H, ~ 3~ ), 8.15 8.62 ~ 9.04~5H, ~ ) N

, -: ' ': '' Example 83 The followinx compo~mds were syn~hesized b~J proceclures similar to that described in Example 79.
(1) 7~-~D-5-Phthal:imido-5-carboxyvaleramido)-3-[2-(2-hydroxyethylthio)-1,3,4-thiadiazol-5-yl]thiomethyl-3-cephem-4-carboxylic acid.
IR(KBr): 3325, 1780, 1715, 1645, 1530cm NMR~d6-DMSO): ~ 1.30 - 2.40(m,6H), 3.20 - 3.80~m,6H), 4.27(AB-q 2H,J=12Hz), 4.65(t,1H,J=9Hz), 4.96(d,1H,J=5Hz) J
5.55(q,1H,J=5 ~ 811z)~ 7.87(s,4H), 8.70(d,1H,J=8Hz) ~2) 7~-(D-S-Phthalimido-5-carboxyvaleramido)-3-~2-carbamoylmethylthio-1,3,4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid IR~KBr): 3430, 3340, 1776, 1717, 1680, 1535 cm NMR(d6-DMSO): ~ 1.30 - 2.40(m,6H~, 3.57(broad;2H~, 4.40(s,211~, 4.32(AB-q, 2H,J=12Hz~, 4.70(t~1H,J=8.0Hz~, 5.0(d,1H, J=5Hz~, 5.55(q,lH,J=5 ~ 8Hz~, 7.20(br.lH), 7.60(br.lH), 7.86(s,4H), 8.74(d,lH,J=5Hz) Example 84 7~-(2-Thienylacetamido)-3-(1-methyl-lH-tetrazol-~-yl~hiom~thyl-3-cephem-4-carboxylic acid was prepared by the procedure similar to that de-scribed in Example 81. Yield 0.37 g starting from 0.50 g of 7~-~2-thienylacet-amido)-3-~2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid.
IR(KBr): 1776, 1734, 1672cm NMR~d6-DMSO): ~ 3.56 ~ 3.78~2H,AB~,J=18Hz,2-CH2), 3.73~2H,s,-CH2CO-), 3.92(3H,s,~ N-CH3), 4.21 ~ 4.37~2H,ABq,J=13Hz,3-CH2), 5.05(1H,d,J=5Hz,6-H), 5.66(1H,dd,J=5 ~ 8Hz,7-H), 6.90 7.29~3H, ~ ~ ), 9.10(1H,d,J=8Hz,-CONH-) ExamplP 85 In water ~12 m~) was dissolved 7~-[D-5-(p-t-butyl-benzamido)-5-,.

3~

carboxyvaleramiclo]-3-mandelyloxynlethyl-3-cephem-4-cclrboxylic acid (1.33 g) together with 5-mercapto-1-methyl-1l-l-tetrazole (0.25 g) and sodium hydrogen carbonate (0.51 g). The mixture was stirred at 60C for 30 minutes, after which time it was brought down to room temperature. ~ollowing the addition of 4 % aqueous phosphoric acid solution ~30 mQ), the reaction mixture was extracted with ethyl acetate (60 mQ). The ethyl acetate layer was washed with a saturated aqueous solution of sodium chloride (40 mQ x 2), dried (over magnesium sulfate) and concentrated under reduced pressure. 'I'hen, upon addi-tion of ether, there was obtained a powder. This powder was recovered by filtration, washed with ether and dried under reduced pressure over phosphorus pentoxide. The procedure provided 7~-[D-5-(p-t-butylbenzamido)-5-carboxy-valeramido]-3-(1-methyl-lH-tetrazol-S-yl)thiomethyl-3-cephem-4-carboxylic acid ~1.14 g).
In IR(KBr) and NMR(d6-DMS0) spectra, this compound was good agree-ment with the product in Example 65.
Example 86 In water (12 mQ) was dissolved 7~-(D-5-phthalimido-5-carboxyvaler-amido)-3-mandelyloxymethyl-3-cephem-4-carboxylic acid ~1.27 g) together with 5-mercapto-1-methyl-lH-tetrazole (0.25 g) and sodium hydrogen carbonate (0.51 g). The solution was stirred at 60C for 30 minutes. Af~er the reaction had been completed, ~he reaction mixture was treated in the same manner as Example 85. The procedure provided 7~-(D-5-phthalimido-5-carboxyvaleramido)-3-(l-methyl-lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid (1.12 g).
In IR(KBr) and NMR~d6-DMSO) spectra, this compound was good agree-ment with the product in Example 1 ~3).
Example 87 In water ~5 mQ) was dissolved 7~-(D-mandelamido)-3-mandelyloxy-meth~l-3-cephem-4-carboxylic acid ~0.50 g) together with 5-mercapto-1-methyl-: ; . ~ :

: , lll-tetr~zole (0.12 g) and sodium hydrogen carhonate (0.17 g). The solution was stirred at 60C for 30 minutes. AEter cooling in the air, the reaction mixture was subjected to column chromatography on Amberlite XAD-2, elution being carried out with water and a solvent mixture of water and methanol.
The fractions containing the desired product were pooled, concentrated and lyophilized. The procedure provided 7~-(D-mandelamido)-3-(1-methyl~lH-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid sodium salt tO.34 g~.
In IR and NMR spectra, this compound was good agreement with the product in Example 81.
Example 88 In water (5 mQ) was dissolved 7~-~2-thienylacetamido)-3-mandelyloxy-methyl-3-cephem-4-carboxylic acid (488 mg) together with sodium hydrogen car-bonate ~84 mg), potassium iodide (400 mg) and pyridine (212 mg). The solution was adjusted to pH 6.5 and reacted at 60C for 45 minutes. After cooling, the reaction mixture was subjected to column chromatography on Amberlite XAD-2, elution being carried out with water and, then, a solvent mixture of water and methanol. The fractions containing the desired product were pooled, concentrat-ed and lyophilized. The procedure provided 7~-(2-thienylacetamido)-3-(1-pyridylmethyl)-3-cephem-4-carboxylic acid betaine (250 mg).
2a In IR and N~IR spectra, this compound was good agreement with the product in Example 82.
Example 89 In water (4 mQ) was dissolved 7~-(2-thienylacetamido)-3-mandelyloxy-methyl-3-cephem-4-carboxylic acid (0.49 g) together with 5-mercapto-1-methyl-lH-tetrazole (0.12 g) and sodium hydrogen carbonate (0.17 g). The solution was stirred at 60C for 30 minutes. After the reaction had been completed, the reaction mixture was treated by a procedure similar to that described in Example ~5. The procedure provided 7~-(2-thienylacetamido)-3-(l-methyl-lH-.

tetraæol-S-yl)thionlethyl-3-cephein-4-carhoxylic acicl (0.40 g).
In IR and NMR spectra, thls compo~lnd WclS good agreement with the praduct in Example 8~.
Lxample 90 lhe following compounds were synthesized by procedures similar to those described in Example 85 and 86.
(1) 7~-[D-5-~p-t-Butylbenzamido)-S-carboxyvaleramido~-3-(2-methyl-1,3,4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid. Yield 84 %.
In IR and NMR spectra, this compo~md was good agreement with the product in Example 80.
(2) 7~-(D-5-Phthalimido-5-carboxyvaleramido)-3-(2-methyl-1,3,4-thia-diazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid. Yield 86 %.
In IR and NMR spectra, this compound was good agreement with the product in Example 79.
~3) 7~-~D-5-Phthalimido-S-carboxyvaleramido)-3-[2-(2-hydroxyethylthio)-1,3,4-thiadiazol-5-yl]thiomethyl-3-cophem-4-carboxylic acid. Yield 81 %.
In IR and NMR spectra, this compound was good agreement with the product in Example 83 (1).
C4) 7~-(D-5-Phthalimido-5-carboxyvaleramido)-3-(2-carbamoylmethylthio-2Q 1,3,4-thiadia~ol-5-yl)thiomethyl-3-cephem-4-carboxylic acid. Yield 88 %.
In IR and NMR spectra, this compound was good agreement with the product obtained in Example 83 (2~.
Examp~ 91 In water (30 mQ) were dissolved 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~3.14 g), sodium hydrogen carbonate (1.84 g) and 5-mercapto-1-methyl~lH~tetrazole ~1.4 g) and the solution was adjusted i*s pH to 5.5, followed by stirring and heating at 60aC for one hour. After cool-ing, the reaction solution was washed with dichloromethane (20 mQ) and the ~ .

... . . ' ~ :~

,~
' ~ ' ' ' ~' .' ' ' ' :

aqueous layer was adjusted to p~l 3.3, -Eollowed by stirring for one hour under ice-cooling. Ihe resultant precipitates werc collectcd by filtration and washed with water, methanol and acetone in this order, ollowed by drying to give 7~-amino-3-(1-methyl-1~l-tetrazol-5-yl)thiomethyl-3-cephem-4-carboxylic acid ~2.7 g).
IR~KBr): 1795cm NMR~ in D20~NallCO3): 3.61 ~ 3.98~2H,ABq,J=18Hz,2-CH2), 4.21(s,-NCl13), 5.21(d,J=4.5~1z,6-ll), 5.60(d,J=4.51-lz,7-H) ~xample 92 In water (30 mQ) were dissolved 7~-amino-3-~3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid ~3.14 g), sodium hydrogen carbonate ~0.84 g) and 5-mercapto-1-[2-(N,N-dimethylamino)ethyl]-lH-tetrazole (2.60 g) and the re-sultant solution was stirred for 60 minutes at 55~C. After cooling, acetone (15 mQ) was added to the reaction solution and the mixture is passed through a column of active alumina (10 g). The column was washed with water-acetone (1:1) (30 mQ) and the washing was combined with the eluate, followed by dis-tilling off acetone under reduced pressure. To the residual solution was added Amberlite IR-120 (acid form) (6.0 mQ) and the mixture was stirred for 30 min-utes under ice-cooling. The insolubles were filtered off and the filtrate was condensed. The condensate was added dropwise into ethanol about 30 times volume of the condensate and precipitated solid was collected by filtration, followed by washing with ethanol and drying to give 7~-amino-3-1-[2-~N,N-dimethylamino)ethyl]-lH-tetrazol-5-yl3thiomethyl-3-cephem-4-carboxylic acid ~3.28 g~-IR~KBr): 3450, 1780, 1620, 1540cm 1 NMR(~ in D20): 3.07~6H,s), 3.70~2H,ABq,J=17Hz), 3.85~2H,t,J=6Hz), 4.25~2H,ABq,J=12Hz), 4.8 - 5.2~4H,m) . .

3~;

E a_y~
In water (30 mQ) was dissolved 7~-amino-3-(3-oxobutyryloxy)met}lyl-3-cephem-4-carboxylic acid (3.14 g) together with sodiwm hydrogen carbonate ~1.84 g) and 2-mercapto-5-methyl-1,3,~-thiadiazole (1.6 g) and the resultant solution was adjusted i-ts pH to 6.4, followed by stirring or one hour at 60C. After coollng, the reaction solution was washed with dichloromethane and pH of the aqueous layer was adjusted to 3.5 under ice-cooling, followed by stirring for one hour. The precipitated materials were collected by filtra-tion and washed with water, methanol and acetone in this order, ~ollowed by drying to give 7~-amino-3-(2-methyl-1,3,4-thiadia~ol-5-yl)thiomethyl-3-cephem-4-carboxylic acid (2.9 g).
IR~KBr): 1795 cm NMR(~ in D20~NaHC03): 2.87(3H,s,thiadiazole -CH3) J 3.53 ~ 3.95~2H,ABq, J=18Hz,2-CH2), 4.10 ~1 4.46(2H,ABq,J=13Hz,3-CH2), 5.17(1H,d,J=4.5Hz,6-H), 5.58(1H,d,J=4.5Hz,7-H) Example 94 In water ~1 mQ) containing sodium salt o~ 5-mercapto-lH-1,2,3-triazole (120 mg) and sodium hydroxide (40 mg) was dissolved under ice-cooling 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (282 mg) and to the resultant solution was added lN-HCQ to adjust its pH to 5.5 under stirring, followed by further stirring for one hour at 55C. To the reaction solution was added methanol (5 mQ) and the mixture was allowed to cool to the room temperature. The cooled mixture is adjusted its pH to 3.9 by adding lN-HCQ
under stirring and the resultant mixture was ~urther stirred for one hour under ice-cooling. The precipitated insolubles were collected by filtration and washed with water and methanol in this order. The insolubles were dried naturally and then over phosphorus pentoxide to give 7~-amino-3-(lH-1,2,3-triazol-5-yl)-thiomethyl-3-cephem-4-carboxylic acid (190 mg).

.,, 3~

IR(KBr): 1800, 1525cm _.x~
In water (1 m~) containing sodium hydrogen carbonate (84 mg) was dissolved 7~-amino-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (314 mg) and to the resultant solution were added isonicotinamide (185 mg) and potassium iodide (1.0 g)~ followecl by stirring for one hour at 55C. To the reaction mixture was added ethanol (20 mQ) lmder stirring and resulting pre-cipitates were collected by filtration, fol]owed by washing with ethanol.
After natural drying, the brown power was dissolved in water (3 m~) and the solution was chromatographed on column packed with Amberlite XAD-2. The eluate was freeze-dried to give 7~-amino-3-(4-carbamoylpyridinium)methyl-4-carboxylate (150 mg).
IR(KBr): 3500, 1760, 1600cm Example 96 In a mixture of tetrahydrofuran (5 mQ) and water ~10 m~) were dis-solved 7~-(2-thienylacetamido)-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid (876 mg) and sodium sulfite (504 mg) and the resultant solution was stir-red for 120 minutes at 60C. To the reaction mixture was added water (10 mQ) and the mixture was adjusted its pH to 7.2 with 2N-HCQ, ollowed by condansa-tion under reduced pressure. ~The residue was subjected to column-chromatog-raphy of silica-gel and eluted with acetonitrile-water (7:1 - 5:1). The frac-tions containing the objective compound were combined and the mixture was sub-jected to distillation of acetonitrile. The residue was chromatographed on column packed with Amberlite XAD-2 and eluted with water and then water-methanol. The fractions containing the objective compound were combined and the mixture was condensed under reduced pressure, followed by freeze-drying to give disodium salt of 7~-(2-thienylacetamido)-3-sulfomethyl-3-cephem-4-carboxylic acid (310 mg).

:~ .

3~

IR(KBr): 3~50, 1760, lh65, 1605~ 1190, lO55cm NMR(~ in D20): 3.67(2H~Bq,J=17llz), 3.92(2H,s), 4.16(2H,~Bq,J=L6Hz~, 5.20~111,d,J=5~-1z), 5.64(111,d,J=5Hz), 7.05 ~ 7.40(3H,m).
F.x~
In 50 mQ of dlchloromethane was dlssolved 7.05 g of 7-[D-phthalimido-5-carboxyvaleramido]-3-hydroxymethyl-3-cephem-4-carboxylic acid ditriethyl-amine salt and, at 0C, triethylamine (1.5 mQ) was added to the solution, followed by dropping diketene (2.0 mQ) during 10 minutes at -5 - 0C The mixture was further stirred for 50 minutes at -5 - 0C and added 40 mQ of water, followed by adjusting its pll to 6.0 with 2N-HCQ. ~e aqueous layer was washed with dichloromethane (10 mQ) and 2.25 g of 5-mercapto-1-[2-(N,N-dimethylamino)ethyl]-lH-tetrazole was added thereto, followed by adjusting to pH 5.5. The solution was stirred at 60DC for 40 minutes, and then the in-solubles were filtered off. To the filtrate were added sodium chloride (15.0 g) and a saturated aqueous solution of sodium chloride (50 mQ) and the mixture was adjusted to pH 2.0 with 4N-HCQ. The solid materials precipitated were collected by filtration and washed with a saturated aqueous solution of sodium chloride and water in this order, -followed by drying to give 6.75 g of 7~-(D-5-phthalimido-5-carboxyvaleramido)-3~ l-[2-(N,N-dimethylamino)ethyl]-lH-tetrazol-5-yl} thiomethyl-3-cephem-4-carboxylic acid hydrochloride.
IR(KBr): 3370, 1775, 1715, 1640cm NMR~ in d6-DMS0): 1.30 - 2.40~6H,m), 3.5 - 4.8(9H,m), 5.04~lH,d, J=5Hz), 5.60~1H,q,J=5.8Hz), 7.90~4H,s), 8.86 (lH,d,J=8Hz) Example 98 In a phosphate bufer solution of pH 6.4 ~3 m~) were dissolved 5-mercapto-2-methyl-1,3,4-thiadiazole (79 mg), sodium hydrogen carbonate t50 mg) and 7~-[2-~lH-tetrazo:L-l-yl)acetamido]-3-~3-carboxypropionyloxy)methyl-3-.. . .

: :

:
' ~ ' cephem-4-carboxylic ncid disoclium sal-t ~243 mg) and the resultant solution was heated for one hour at 60C. After cooling, the reac-tion solution was concentrated under red~lced pressure and the residue was subjected to column-chromatography on Sephadex l.H-20 (250 m~), elution being carried out with water. The fractions containing the desired product were pooled and lyophil-ized. The procedure provided sodium 7~-[2-(1ll-tetrazol-1-yl)acetamido]-3-~2-methyl-1,3,4-thiadiazol-5-yl)thiomethyl-3-cephem-4-carboxylate.
NMR~ in D20): 2.78(3H,s,-CH3), 3.60(2H,ABq,J=18llz,2-CH2), 4.25(211,ABq, J=13Hz,3-CH2), 5.12(1H,d,J=4.5Hz,6-H), 5.58(2H,s,-CH2C0-), 5.70(1H,d,J=4.5Hz,7-H), 9.15(1H,s,tetrazole-H) Example 99 In a phosphate buEfer of pH 6.4 (3 mQ) were dissolved sodium 7~-[D-c~-t-butoxycarbonylamino-c~ hydroxr~ph~yl)acetamido]-3-~3-oxobutyryloxy) methyl-3-cephem-4-carboxylate (280 mg) and sodium salt of 5-mercapto-lH-1,2, 3-triazole ~120 mg) and the resulting solution was heated at 60C for 60 minutes. After cooling down to room temperature, the reaction solution was chromatographed on a column packed with Amberlite XAD-2 and the combined elu-ates containing the desired product was freeze-dried to give sodium 7~-[D-CL-t-bu~oxycarbonylamino-c~-(p-hydroxyphenyl)acetamido]-3-(lH-1,2,3-triazol-5-yl)-20 thiomethyl-3-cephem-4-carboxylate.
IR~KBr): 3400, 1762J 1678cm NMR~D20): ~ 1.46[9H,s,-C(CH3)3], 3.00 - 4.12(4H,m,2-CH2,3-CH2), 5.00~1H,d,J=4.5Hz,6-H), 5.60~1H,d,J=4.5Hz,7-H), 6.78 -7.42(4H,m, ~ ), 7.71(lH,s,triazole-4-H) Thus obtained sodium 7g-[D-c~-t-butoxycarbonylamino-c~-(p-hydroxy-phenyl)acetamido]-3-~lH-1,2j3-triazol-5-yl)-thiomethyl-3-cephem-4-carboxylate ~175 mg) was dissolved in ormic acid ~4 mQ) and the resultant solution was stirred for 2 hours at room temperature. The reaction solution was subjected .

~323~

to distillation under red~lced pressure and then to azeotropy (3 times) with toluene to remove formic acid, followed by dry:Lng over phosphorous pentoxide over night. Thus obtained foamy material was stirred with water~methanol ~8:2) ~15 mQ) and the mixture was subjected to ~iltration, treatment with ac-tivated carbon and then filtration using Celite. The filtrate was freeæe-dried to give 7-[D~ amino-~-(p-hydroxyphenyl)acetam~do]-3-~lH-1,2,3-triazol-5-yl~thiomethyl-3-cephem-4-carboxylic acid. This product was in good agree-ment with the standard in thin-layer chromatography and liquid chromatography.
~ le lO0 In water ~10 mQ) were dissolved 7R-[2-~2-imino-4-thiazolin-4-yl)-acetamido]-3-(3-oxobutyryloxy)methyl-3-eephem-4-carboxylic acid ~908 mg), 5-mercapto-1-[2-(N,N-dimethylamino)ethyl]-lH-tetrazole ~450 mg) and sodium hydrogen carbonate (168 mg) and the resultant solution was heated at 55C for 60 minutes. The reaction solution was revealed to contain 7~-[2-(2-imino-4-thiazolin-4-yl)acetamido]-3~ 1-[2-~N,N-dimethylamino)ethyl]-lH-tetrazol-5-yl}
thiomethyl-3-cephem-4-carboxylic acid in 81 % yield relative to the starting cephalosporin by liquid chromatography~ The reaction solution was adjusted its pH to 5.8 and purified by column-chromatography using Amberlite XAD-2.
IR~KBr): 1765cm NMR~D2O): ~ 3.06~6H,s,-N~CH3)2), 3.5 - 4.8~10H,m), 5.12~1H,d,J=5H~,6-H~
5.65~1H,d,J=5Hz,7-H), 6.62~1H,s,thiazolin-H) Example 101 In dimethylformamide ~16.0 mQ) was suspended mono-sodium salt mono-hydrate of deacetylcephalosporin C~4.13 g~ and the resultant suspension was dissolved by adding concentrated hydrochloric acid (1.66 mQ) at a temperature lower than 0C~ followed by adding dimethylformamide ~16.0 mQ), triethylamine (4.90 m~) and phthalic anhydride ~2.96 g) in this order. The resultant mix-ture was stirred for one hour and a half at 20C and the reaction solution was 3~
poured onto a mixture oE an aqueous solution of sodium chloride (200 m~) and dichloromethane (40 m~). The mixture was ad~usted its pH to 6.5 and the aque-ous layer was separated. The layer was washed wi-th dichloromethane and ex-tracted by a mixed solution ot` ethyL acetate-tetrahydrofuran (3~ 50 m~ x 3) and the combined extract was washed with a saturated aqueous solution of sodium chloride, followed by drying over magnesium sulfate. The dried extract was concentrated under reduced pressure and ether was added to the concentrate.
The procedure provided 7~-[D-5-~2-carboxybenzamido)-5-carboxyvaleramido]-3-(2-carboxybenzoyloxy)methyl-3-cephem-4-carboxylic acid (6.22 g).
IR(KBr): 1780, 1735, 1725, 1715, 1640cm l NMR(~ in D6-DMS0): 1.40-2.40(6H,m)9 2.62(2H,ABq,J=18Hz), 4.35(1H,m), 5.09(2H,ABq,J=13Hz), 5.10(lH,d,J=5Hz), 5.72~1H,dd,J=5 ~ 8Hz), 7.30-7.9(8H,m), 8.53(1H,d,J=8Hz), 8.82(1H,d, J-8Hz).
Example 102 Following the procedure of Example 6, 7~-[2-(2-amino-4-thiazolin-4-yl)-aoetamido]-3-hydroxymethyl-3-cephem-4-carboxylic acid triethylamine salt is reacted with diketene to yield 7~-~2-(2-amino-4-thiazolin-4-yl)-acetamido]-3-(3-oxobutyryloxy)methyl-3-cephem-4-carboxylic acid.

- 75 ~
~ .

.
: .

':

,

Claims (15)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:

1. A process for producing a cephalosporin antibiotic of the formula:

or a pharmaceutically-acceptable salt thereof, wherein R1 represents hydrogen or a suitable acyl group and R2 stands for a pyridinium group optionally sub-stituted by methyl, carboxyl, carbamoyl, carbazoyl, bromo, sulfo, hydroxymethyl or formyl, or stands for a residue of a nitrogen-containing heterocyclic thiol which comprises reacting a compound of the formula:

or a salt thereof, wherein W represents acetonyl or a group represented by -X-COOH or -X-OH in which X is a divalent carbon chain which is able to form a five- or six-membered ring with or , which may include a double bond or such an atom or atoms as oxygen, nitrogen or sulfur therein, and on which carbon chain suitable substituent or substituents may be attached, with pyridine or a pyridine substituted as defined for pyridinium above, or with a nitrogen-containing heterocyclic thiol, and where required converting any cephalosporin antibiotic thus produced into a pharmaceutically-acceptable salt.

2. A process as claimed in claim 1, wherein tho symbol X is selected from the group consisting of -CH=CH-, -CH2CH2-, , -CH2CH2CH2-, -CH2OCH2-, -CH2SCH2-, , , , , , , CH3-?H-, ?H2-?H2, , and .

3. A process as claimed in claim 1 wherein the symbol R1 represents an acyl group of the formula:

wherein R4 represents acetyl, a halogenoacetyl, phenyl, p-hydroxyphenyl, thienyl, 2-imino-4-thiazolin-4-yl, 2-oxo-4-thiazolin-4-yl, tetrazolyl, phenoxy or 3-amino-3-carboxypropyl and R5 represents hydrogen, sulfo, amino or hydroxy, and each of amino and carboxyl group in R4 and R5 may be protected.

4. A process as claimed in claim 3, wherein the protective group of amino group is selected from the group consisting of phthaloyl, benzoyl, o-carboxybenzoyl, p-nitrobenzoyl, toluoyl, naphthoyl, p-tert-butylbenzoyl, p-tert-butylbenzenesulfonyl, phenylacetyl, benzenesulfonyl, phenoxyacetyl, toluenesulfonyl, chlorobenzoyl, acetyl, valeryl, capryl, n-decanoyl, acryloyl, pivaroyl, camphorsulfonyl, methanesulfonyl, chloroacetyl, tert-butoxycarbonyll ethoxycarbonyl, isobornyloxycarbonyl, phenyloxycarbonyl, trichloroethoxycarbonyl, benzyloxycarbonyl, .beta.-methylsulfonylethoxycarbonyl, methylcarbamoyl, phenylcarbamoyl, naphthylcarbamoyl, and 2-methoxycarbonyl-1-methylvinyl.

5. A process as claimed in claim 1, wherein R1 is D-5-amino-5-carboxy-valeryl of which amino group is protected and W is acetonyl.

6. A process as claimed in claim 1, wherein R1 is D-5-phthalimido-5-carboxyvaleryl and W is acetonyl.

7. A process as claimed in claim 1, wherein R1 is D-.alpha.-sulfophenyl-acetyl and W is acetonyl.

8. A process as claimed in claim 1, wherein R1 is 2-(2-imino-4-thia-zolin-4-yl)acetyl and W is acetonyl.

9. A process as claimed in claim 1, wherein R1 is D-.alpha.-sulfophenyl-acetyl, W is acetonyl, R2 is 4-carbamoylpyridinium and isonicotinamide is employed as the substituted pyridine.

10. A process as claimed in claim 1, wherein R1 is D-5-phthalimido-5-carboxyvaleryl, W is acetonyl, R2 is {1-[2-(N,N-dimethylamino)ethyl]-1H-tetrazol-5-yl} thio and 5-mercapto-1-[2-(N,N-dimethylamino)ethyll-1H-tetrazole is employed as the nitrogen-containing heterocyclic thiol.

11. A process as claimed in claim 1, wherein R1 is 2-(2-imino-4-thia-zolin-4-yl)acetyl, W is acetonyl, R2 is {1-[2-(N,N-dimethylamino)ethyl]-1H
tetrazol-5-yl} thio and 5-mercapto 1-[2-(N,N-dimethylamino)ethyl]-1H-tetrazole is employed as the nitrogen-containing heterocyclic thiol.

12. A process as claimed in claim 1, wherein R1 is D-5-phthalamido-5-carboxyvaleryl, W is acetonyl, R2 is (1-methyl-1H-tetrazol-5-yl)thio, and 5-mercapto-1-methyl-1H-tetrazole is employed as the nitrogen-containing heterocyclic thiol.

13. A process for producing a cephalosporin antibiotic of the formula:

or a salt thereof, wherein R1 represents hydrogen or a suitable acyl group and R2 stands for a residue of a nucleophilic compound selected from nitrogen-containing heterocyclic thiols and nitrogen-containing heterocyclic compounds which comprises reacting a compound of the formula:

or a salt thereof, wherein W represents an acetonyl group or a group rep-resented by -X-COOH or -X-OH (X is an organic residue), with a nucleophilic compound as defined above.

14. A process as claimed in claim 13, wherein the symbol X is selected from the group consisting of -CH=CH-, -CH2CH2-, , -CH2CH2CH2--CH2OCH2-, -CH2SCH2-, , , , , , , CH3-?H- , , , and ,

15. A process as claimed in claim 13 or 14, wherein R stands for the residue of a nitrogen-containing heterocyclic thiol selected from the group consisting of pyridyl thiol, N-oxide pyridyl thiol, pyrimidyl thiol, pyridaz-inyl thiol, N-oxidepyridazinyl thiol, pyrazolyl thiol, diazolyl thiol, thiazol-yl thiol, thiazolyl thiol, 1, 2, 3-thiadiazolyl thiol, 1, 2, 4-thiadiazolyl thiol, 1, 3, 4-thiadiazolyl thiol, 1, 2, 5-thiadiazolyl thiol, 1, 2, 3-oxa-diazolylthiol, 1, 2, 4-oxadiazolyl thiol, 1, 3, 4-oxadiazolyl thiol, 1,2, 5-oxadiazolyl thiol, 1, 2, 3-triazolyl thiol, 1, 2, 4-triazolyl thiol, 1H-tetrazolyl thiol and 2H-tetrazolyl thiol.