CA1164863A - Analogs of lincomycin and clindamycin - Google Patents

Abstract

ABSTRACT OF THE DISCLOSURE

Novel useful analogs of the well known antibiotics lincomycin and clindamycin. These analogs are prepared by condensing a cyclic acid with a sugar amine and are represen-ted by the general formula:

wherein R1, which can be singly or multiply substituted in any position of the pyridine ring not already substituted by R2, is hydrogen, alkyl and substituted alkyl, wherein the alkyl portion is from 1 to 8 carbon atoms, and isomeric forms thereof, cycloalkyl and substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl and substituted phenyl or -(CH2)m-OH, -(CH2)m-NR4R5, and isomeric forms there-of, wherein m is an integer of from 1 to 8, and R4 and R5 are hydrogen or alkyl of from 1 to 8 carbon atoms and isomeric forms thereof; and wherein R2, which can be singly substituted in any position of the pyridine ring not already substituted by R1, is:

-?-X
and X is the amino function of 7(R)-hydroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-hydroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(R)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(S)-methoxy-methyl 1-thio-.alpha.-linco-saminide, 7-deoxy-7(S)-(methylthio)-methyl 1-thio-.alpha.-linco-saminide, 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl 1-thio-.alpha.-lincosaminide or 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl 1-thio-.alpha.-lincosaminide.

Description

1 16~863 This application is the first of three divisional applications of copending application serial no. 362,485, filed October 16, 1~80.

BACKGROUND OF THE INVENTION
The characteristics and preparation of the anti-biotic lincomycin are disclosed in U.S. Patent 3,086,912.
Clindamycin is disclosed in U.S. Patent 3,496,163. These antibiotics have been extensively used as medicines in humans and animals. A number of patents world-wide have issued concerning these antibiotics and a variety of derivatives thereof.
Lincomycin has the following structural formula I ÇH3 H N~ H
- ~ ~ H04-H
H ~ C N ~

~ SEH3 H OH

Clindamycin has the following structural formula cb/~

1 l6~863 e, N\IH H-CHCl H

H~SCH3 H OH

BRIEF SUMMARY OF THE INVENTION
This application relates to and claims novel and useful compounds of the general formula:

~ N ~ I

wherein Rl, which can be singly or multiply substituted in any position of the pyridine ring not already substituted by R2, is hydrogen, alkyl and substituted alkyl, wherein the alkyl portion is from 1 to 8 carbon atoms, and isomeric forms thereof, cycloalkyl and substituted cycloalkyl, substituted oxygen, substi.tuted nitrogen, halogen, phenyl and substituted ( 2)m OH, (CH~)m-NR4R5, and isomeric forms thereof, wherein m is an integer of from 1 to 8, and R~ and R5 are hydrogen or alkyl of from 1 to 8 carbon atoms and isomeric forms thereof; wherein R2, which can be si.ngly substituted in any position of the pyridine ring not already substituted i~
cb/~?

1 l64863 by ~1' is:

o --C--X
and X is the amino funetion of 7(R)-hydroxy-methyl l-thio-~-lincosaminide, 7(S)-hydroxy-methyl l-thio-a-lineosaminide, 7(S)-halo-methyl l-thio-~-lincosaminide, 7(R)-halo-methyl l-thio-~-lincosaminide, 7(S)-methoxy-methyl l-thio-~-lincosaminide, 7-deoxy-7(S)-~methylthio)-methyl l-thio-~-lincosaminide, 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl l-thio-~-lincosaminide or 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl l-thio-~lincosaminide.
This application also relates to novel and useful eompounds, which are claimed in the above noted parent application, of the general formula:

II

wherein Rl, which ean be singly or multiply substituted in the 2, 3, 4, 5, 6, 7, 8 or 9 position of the ring not already substituted by R2, is as defined above; wherein R2, which can be singly substituted in any position of the ring not already substitut~d by Rl, is as defined above; wherein R3 is hydrogen, methyl, ethyl or C2H5OH; and wherein n is an integer of from 1 to 4.
This applieation further relates to novel and useful compounds, which are claimed in a second divisional application, cb~

1 16~863 serial no. ~14,644, filed November 1, 198~r of the gelleral formula:

>~ X
\ B / VI

wherein A, B and E are nitrogen, oxygen, sulfur or CRlRl;
wherein Rl, which can be.singly or multiply attached to any ring carbon atom not already substituted by R2, is as defined above; and wherein R2, which can be attached to any ring carbon, not already substituted by Rl, or nitrogen atom, is as defined above.
This application still further relates to novel and useful com~ounds, which are claimed in a third divisional application, serial.no, 414,645, filed November 1, 1982, of the general formula:

D ~ R
R2~
E ~ ~ VII

wherein A, B, D and E are nitrogen, oxygen, sulfur or CRlRl;
wherein Rl, which can be singly or multiply attached to any ring carbon atom not already substituted by R2, is as defined above; and wherein R2, which can be attached to any ring carbon, not already substituted by Rl, or nitrogen atom, is as defined above.
Compounds of particular importance are of the general formula:

cb/~

I l6~863 [~ ~
INR ~ III

wherein Rl is in the 4 position and is alkyl of from 1 to 8 carbon atoms and isomeric forms thereof; wherein R2 is in the

2 or 3 position and is otherwise as defined above; and wherein R3 is as defined above.
Important precursor compounds of the above have the general formula:
~ nl ~ N ~ IV

wherein Rl and R2 are as defined immediately above.
The phaLmaceuticallY accep~ble acid addition salts of the above noted compounds including the 2-phosphates and 2-palmitates, wherein the substituent is attached to the oxygen atom at the 2 position of the sugar ring of the above compounds, are also described and claimed in this and the above noted related applications.
The synthesis of the novel analogs described herein can be shown in exemplary form as follows:

cb/ ~

1 l64863 HC-Cl ~ O HC-Cl C-o~ ~ NHZ--CH > ~ H
H ~ O HO ~ ~

~ S-CH3 ~ S-CH3 C2Hs k ~ CH3 ~ N 1 C-NH- H

V~

,, nH

The wavy lines denote either the D-cis or L-cis isomer.
An alternate procedure which may be used to synthesize the novel analogs described herein can be shown in exemplary form as follows:

Hs ~ Hg N 1 C//OH + 7-C1-MT~ ~ ~ N ~ //7-Cl-MT~
H H

The wavy line denotes either the D-cis, L-cis, D-trans or L-trans structures.

O' cb/ ~

The L-cis structure ~N lc _ N H--C H X~ 2 0 HO~ ~.1 ~AS''c1~3 V
OH

has been shown to be 5 to 10 times more active than clindamycin against _ aureus and S. hemolyticus in laboratory mice.
An isomer of V may be isolated from the above reaction and is presumed to be the D-cis compound, VA.
The D-cis structure is not as potent an antibacterial agent as the L-cis compound.

C=2H5 J ~O H C - C 1 ~S-CH3 V A
OH

DETAILED DESCRIPTION OF THE INVE.NTION

Upon reacting an amino acid of the formula -6a-. ~
cb/,"~

C-OH

N

wherein R1, which can be singly or multiply substituted in any position of the pyridine ring not already substituted by //
-C-OH , is selected from the group consisting of hydrogen, alkyl and substi-tuted alkyl wherein the alkyl portion is from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof, cycloalkyl and substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl and substituted phenyl; -(CH2)m-OH, -(CH2)m-NR4R5, and isomeric forms thereof, wherein m is an integer of from l to 8, inclusive, R4 and R5 are H or alkyl of from l to 8 carbon atoms, inclusive, and isomeric forms thereof, wherein O
-C-OH
which can be singly substituted in any position of the pyridine ring not already substituted by R1, with a sugar amine compound selected from the group consisting of 7(R)-hydroxy-methyl l-thio-a-lincos-aminide, 7(S)-hydroxy-methyl l-thio-a-lincosaminide~ 7(S)-halo-2S methyl l-thio-a-lincosaminide~ 7(R)-halo-methyl l-thio-a-lincos-aminide, 7(S)-methoxy-methyl l-thio-a-lincosaminide~ 7-deoxy-7(S)-(methylthio)-methyl l-thio-a-lincosaminide~ 7-deoxy-7(S)-(2-hydroxy-ethylthio)-methyl l-thio-a-lincosaminide, and 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl l-thio-a-lincosaminide; there are obtained novel and useful compounds of formula I.
Upon reacting an amino acid of the formula ~ N ~
I

ll6486~
-8- . 3809A

wherein R1 and the position of substitution of -C-OH are as defined above; wherein R3 is selected from the group consisting of H, CH3, C2H5, and -CH2-CH2-OH; wherein n is an integer of from 1 to 4, inclusive, with a sugar amine compound, as defined above, there are obtained novel and useful compounds of formula II.
Upon reacting an acid of the formulae HO-C ~ R
E A
\ 8 ~
HO c~f B

wherein A, B, D and L are selected from the group consisting of nitrogen, oxygen, sulfur and CR1R1; R1 is as defined previously and can be attached to any ring carbon or nitrog~Jn atom; R1 can be multiply attached to any ring carbon atom; -C-OH can be attached to any ring carbon or nitrogen atom, with a sugar amine compound selected from the group as defined above, there are obtained novel and useful compounds of formulae VI and VII.
MTL is methyl l-thio-a-lincosaminide of the formula HO-C-H

H~ ~ ~

OH
epi-MTL is methyl 7(S)-7-deoxy-7-hydroxy-1-thio-~-lincos-aminide of the fonnula c~3 H-C-OH
H2N ~H
~0 He/l \¦

OH

7-Cl-MTL is methyl 7(S)-7-deoxy-7-chloro-1-thio-a-lincosaminide of the formula H-C-Cl HO ~ O
V ' \i ~ CH3 OH
epi-7-C1-MTL is methyl 7(R)-7-deoxy-7-chloro-1-thio-~-lincosaminide of the formula fH3 Cl-C-H

HO ~ ~

~ CH3 OH
7~S)-methoxyrmethyl l -th~O-a-l incos~m~nide c~n be shown as follows (See U.S. 3,702,322, Example 1, Part B-l):

H-f-O-CH3 ~ S-CH3 With reference to the foltowing formula~ 7-deoxy-7(S)-(methylthio)-methyl l-thio-a-lincosaminide exists when R
is CH3; 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl l-thio--lincosaminide exists when R is -CH2-CH2-OH; and, 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl l-thio--lincos-aminide exists when R is -CH2-CH2-CH2-OH (See ~.S. 3,915,954, Examples 1, 10 and 31):

H-C-S-R

H ~ 0~

OH.
The hydroxy and halo groups at the 7 position of the above formulas can be shown as follows ~I~Y

H ~ 0 ~ CH3 ll64a63 wherein Y ;s selected from the group consisting of 7(R)-hydroxy, 7(S)-hydroxy, 7(S)- halo , and 7(R)- halo When a pyridine acyl group is used, the resulting analo~ can be reduced to give a mixture of the corres-ponding saturated compounds, one of which is the L-cis isomer. Other compounds which may be present include the L-trans, D-cis, and D-trans isomers. Generally, for any of the compounds described herein, the reduced form is more antibacterially-active than the unsaturated precursor. The use of a piperidine acyl group gives analogs existing as D-cis, L-cis, D-trans, and L-trans isomers. Again, the L-cis isomer has been found to be more anti-bacterially active.
The general method used herein to prepare the novel analogs is the well known process wherein an appropriate acid is coupled with an appropriate sugar amine.
C'~xed Carboxylic Acid Anhydride Procedure," Chemistry of The Amino Acids, Vol. 2, p. 970, John Wiley and Sons, Inc.
1961.) When the acid is unsaturated, the resulting unsaturated analog can be catalytically reduced under standard conditions to prepare the saturated analog. For example, the reduction can be conducted using the follow-ing conditions:
H2 at 5 to 5~ psi Catalyst - platin;um oxide (PtO2) Solvent - H20 or H20 + MeOH, or H20 + FtOH
HCl - 10~ excess Time - 24 to 48 hours As used ~ereTn, alkyl of 1 ~o 8 car~on ato~s~?
~nclusive, and isomeric forms thereof, includes methyl, - ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, and branched chain isomers thereof.
Substituted alkyl means the above alkyl compounds in which one or more of the hydrogen atoms has been replaced by a halogen, i.e., Cl, Br, F, and I, oxygen, hydroxyl, amine (primary), amine ~secondary-alkyl substituted by alkyl as above), amine (tertiary-alkyl substituted by alkyl as above), sulfur, -SH, and phenyl. Exemplary compounds are l-fl uoroethyl . l-chloroethyl, 2-fl uoroethyl, 2-chloroethyl, l-bromopropyl, 2-iodopropyl, l-chlorobutyl, 4-fluorobutyl, and 4-chlorobutyl.
Cycl oalkyl means cyclopropyl, cyclobutyl, cyclopentyl, cycl ohexyl, cycloheptyl and cyclooctyl .
Substituted cycloalkyl means a cycloalkyl substituted as above for substituted alkyl. Exemplary compounds are 2-cyclopropylethylg

3-cyclobutylpropyl, 4-cyclopentylbu~yl, and 4-cyclohexylbutyl.
Aromatic means phenyl and substituted phenyl wherein one or more of the hydrogen atoms has been repl aced by a halogen, as above, hydroxyl, amine (primary, secondary, and tertiary with the latter two alkyl substituted as above), -SH, and phenyl . Exempl ary compounds are p-bromophenyl, m-iodophenyl, o-chlorophenyl, p-ethyl phenyl, m-propylphenyl, o-methylphenyl, and p-octylphenyl.
As detailed infra, the compounds of the invention can be phosphorylated to give the 2-phosphate, and acylated to give the 2-palmitate ~"hich are both anti-bacteri al ly-acti ve in vivo.
Substituted oxygen means oxygen substituted by an alkyl of from 1 to 8 carbons, inclusive, aryl, and substituted aryl.
Substituted nitrogen means nitrogen substituted by an acyl of from 2 to 18 carbons, a monoalkyl of 1-8 carbons, inclusive, and a dialkyl, wherein the alkyl is from 1 to 8 carbons, inclusive, includ-ing the isomeric forms for all acyl and alkyl groups.
Halo means chloro, bromo, iodo, or fluoro.
Exemplary sources for the amino acids used as starting materials herein are as follows:
1. Heterocyclic Compounds, Vol. 1, John Wiley and Sons, Inc., 1950. This source describes the preparation of halogen and alkyl substituted amino acids.
2. Chem. Abstracts:
- 81 - 105223A - alkyl and cycloalkyl 81 - 152243S - alkyl and halogen substituted ~2 - 170746H - halogen substituted 85 - 46322Q - dihalo substituted 85 - 177258W - dihalo substituted _ - 116928X - dihalo substituted 81 - 3737d - phenyl substituted 78 - 58201t - phenyl substituted 76 - 126800y - tetrahalo substituted 82 - 11036K - bromo subst;tuted 83 - 27119W - bromo substituted 84 - 16613X - bromo substituted 78 - 123494G - bromo substBtuted 1 0 -CH2CH2=CH-CH3 ~,, 84 - 135488Y - I ll ~N "`~OOH

lS CH3-O
81 - 151951J - l ll ~ Br~ ~ ~ COOH

Br-CH
81 - 77809a - ~ ~
N COOH

84 - 30918G - ~
COOH
C2Hs-O~,~ O-C2Hs ~ ~ ~ COOH

I
7g - l9109Y
~ ~ COOH

1164~63 81 - 3737D - ~3 s l [~COOH

81 - 135964K -~Of~COOH

- 1 77349B -~f OOH
~N

~COOH

2 0 7 8 - 71 8 6 5 G - 1 ,!J

~ COOH
8t - 1 35964K -~LO~N~J

C OOH

83 - 1 47397G - \ _~L
~N Cl 82 ` 11036K - ~NH2 ~ COOH

116~863 ~Cl 84 - 11 6928X - ClJ~N~ COOh C2Hs-O~\~ Q-CzH5 81 - 331 39C -~NJJ~:OOH

Cl Cl~ Cl Cl N COOH
~C2H5 Cs~
79 - 11 5449b -~ ~ .
N COOH

IyH-CH3 Cl ~Cl 67 - 63229K - I~ ~
N COOH

68 - 1 04926b ~

_ - 59048Z ~
N' ` COOH

` ` 1164863 71 - 124907M H-~ CO-OH
s L

- OH

IO H-N~co oH
I

~, lS 68 - 59465N ~Nl C-O-CH3 This compound can be hydrolyzed to the acid by means well known in the art, which acid can then be reduced, also by means well known in the art.

86 - 106501e l~<CH3 g~ N C_N

This compound can be hydrolyzed to the acid by means well known in the art. The resul t-ing acid then can be N-demethylated by the procedures disclosed in U.S. Patent 3,583,972.

69 - 67282M CH3-l~ 1l ~N C-O-C2H5 1164~3 This compound can be hydrolyzed to the acid by means well known in the art.

- 70297-14-Z CH3-~

This compound can be hydrolyzed to the acid by means well known in the art. Also, one or both of the N-CH3 groups can be removed from the resulting acid by following the procedures disclosed in U.S. Patent 3,583,972.

I q 90- - 168488X O~NI~ C-O-C4Hg CH3 f -H

This compound can be hydrolyzed to the acid by means well known in the art. The result-ing acid can be converted to the following compound ~ 01 ~N~ C-OH
H

by methods disclosed in U.S. Patent 3,583,972.

85 - 142995G NHz~CHz~

H COOH

1 16Q~63 - 1g- 3~09 81 - 1 5202~SCH3 CH CH2-CH2~
1H ~ 1 S ~ COOH

- 110156M ,~1~

I H
OH

Compounds having free NH2 or OH groups will have to have these groups protected before being condensed with 15 the amino sugar. PrQtection of such groups is well known in the art. See Protective Groups in Organic Chemistry, J. F. W. McOmie, Plenum Publishing Co., Ltd., 1973.

3. Jour. Chem. Soc. 1969 - 2134 -Various H-alkyl substituted pyridines C--N ~-R CH2R

~ + R-MgX

(commerciall~
available) R = alkyl, branched alkyl and cycloalk~l

4. Jour. Chem. Soc. 1969 - 934 -R R R R R

NH~ ~ r ~ C3N ~ C-OH

~ 3~0 The following examples are illustrative of the process and products of the invention, but are not to be construed as limiting. All percentages are by weight and all solvent mixture proportions are by volume unless otherwise noted.
Example I - 4-Cis-ethyl-~-pipecolic acid amide of 7-Cl-MTL HCl (U-57,930E - Compound V)-. _ PART I

2~S HCl l2H5 CH3 0 ~ 7-Cl-MTL ~ O HC-Cl C-~H `~N ~-NH~H
~~\1 ~ S-CH3 OH

A solution of 67 9 (0.357 moles) of t~e amino acid'HCl (C.A. 51, 1643a, 1957) and 71.5 9 (0.714 moles) of triethylamine dissolved in 2.5 liters of acetonitrile is cooled to 10 C and 47.6 ~ (0.354 moles) of isobutyl^
chloroformate added in one portion. This mixture (Solution A) is stirred at 10 C for 1 ~our. Solution B is made up by dissolving 97,7 g ~0.357 moles) of 7-Cl-MTL (J. Med. Chem., 12-780, 1969, B. J. Magerlein and F.
Kagan) in a warm mixture of 1500 ml of acetone and lSOO
ml of H20. Solut;on B is cooled to 30 C and added in one -portion to Solution A. The reaction is stirred at 25 C
for 18 hours and the acetone and acetnoitrile removed under vacuum. rhe white, mushy residue is filtered and the crystalline material collected and dried to give 95 g of pure product. Workup of the filtrate (chromatography) 1 16~863 -20~ 3809A
gave another 10 9 of product. The oYerall yield is 73g.
Anal. Calcd. for C17H25ClN205S: C, 50.42; H, 6.22; N, 6.92; S, 7.92; Cl, 8.7~.
. Found: C, 50.67; H, 6.40; N, 6.64; S, 7~90; Cl, 8.70.
aCHCl3 ~C, 1.0) ~ 293 PART II

~s C2Hs CH3 15~ -7-Cl-MTL ~ O HC-Cl XH20 HCl H H~
~OH
\L__~/S-CH3 V I ~ H

A mixture of 4.05 9 (0.01 mole) of starting material, 40 ml of water, 60 ml of methanol, 1.0 ml of 37~ HCl and 8.0 g of PTO2 catalyst were reduced on a Parr hydrogenator at 50 p.s.i. for 3 hours. Analysis of the reaction mixture by TLC on silica gel plates in a system composed of CHCl3:methanol (6:1) showed that, all of the starting material was gone and that two more polar materials were present in a ratio of about 1:1. The reaction was filtered to remove the catalyst and the filtrate concentrated under vacuum to give a,white crystalline mush. This was filtered and the filtrate saved. The white s~lid, which was the most polar of the two products observed upon TLC of the reduction mixture, was recr~stalli~ed from water to give the desired product, U-57,930E, m.p, 222-224, in a yield of from 25 to 35~.
Anal. Calcd. for C~7H32Cl2N205S: C, 45.63; H, 7.21; N, -21-.. 3809A
6.26; S, 7.17; Cl, 15.35.
Found: C, 45.77; ~, 7.44; N, 6.39; S, 7.21; Cl, 16.17.
aH20 (C, 1.0) f 176 The absolute configuration and sterochemistry of V was established by X-ray crystallography.
U-57,930E, tested in comparison with clindamycin, has the following antimicrobial spectra:
Ta~le I
The Minimal Inhibitory Ooncentration of U-57,930E
and Clindamycin Ys. Aero~ic Bacteria.

~IC (yq/ml~
Orqanism UC Clindamycin U-57,930E

Staphylococcus 6685 >25 >25 .

aureus 6686 - .S .20 6687 .025 .20 6688 >25 >25 6689 .05 .78 . ~690. .OZS .20 6691 .10 ZO
6692 >2~ . . >25 6693 .05 .78 6694 >25 >25 6695 .10 .39 6696 .10 .39 6675 .05 39 76 .05 .10 746 <.OS .05 571 .20 .78 1 1~48~3

5~0 .20 .39 Staphylococcus 719 .~0 .20 epidermidis 3389 .10 .20 Streptococcus faecalis 694 25 6.25 Streptococcus pyogenes 152 <.012 <.012 Streptococcus viridans 153 <.012 .05 871 <.012 <.012 Diplococcus pneumoniae I 41 <.012 <.012 Oiplococcus pneumoniae II3213 <.012 <.012 Escherichia coli45 50 >50 Proteus vulgaris93 >50 >50 I e b-s 1 e1la pneumoniae 58 6.25 >50 Salmonella schottmueller;126 >50 ~50 ..... Pseudomonas aeruginosa 95 >50 >50 35 The procedure for the above test is as follows;

The Minimal Inhibitory Concentrat;on (MIC's) o~ both compounds Vs. aerobic bacteria is determined usinq a standard microplate b`ro~h-dilution method. Brain`Heart Infusion (BHI - Difco) broth medium is used, and the plates are incubated at 37 C for 20 hrs.
S. aureus UC 6685-6696 are clinical isolates which are resistant to one or ~ore commercial antibiotics.
"UC" is a registered trademark of The Upjohn Co~pany Culture Collection. These cultures can be obtained from The Upjohn Company in Kalamazoo, Michigan, upon request.
Table II
The ~inimal Inhibitory Concentration of Clindamycin and U-57930E vs. Gram-Positive and Gram-NegatiYe Anaerobic 8acteria MIC(~g/ml_ Organism UC Clindamycin U-57930E

8acteroides'' fraqilis 6513 0.06 0.12 Z 6428 0.06 0.25 6864 3.9 2.0 686Z 7.8 15.6 8acteroides thetaiotaomicron6512 2.0 0.5 8acteroides distasonis 6518 0.12 <0.03 Bacteroides melaninoqenicus6326 0.06 0.06 Clostridi'um perfri'nqens 241 0~06 0.l2 6509 0,06 0.l2 Clostridium -novyi B 6329 0.06 0.l2 --2~- 3809A
Clostridium tertiu~ 6508 7.8 7.8 Clostridium cadaveris 6510 <0.03 0.06 -Clostridium sordellii 6505 2.0 0.5 Clostridium tentani 6521 <0-03 <0~03 Clostridium botulinum A 6506 0.25 <0.03 Cl ostri dium bife rme ntans 6507 0.50 0.06 Cl ostri dium~
d;fficile 6834 7.8 3.9 ,.
cu 6857 250 125 6858 3.9 3.9 6861 3.9 2.0 Propionibacterium acnes 6564 0.06 0.12 6575 <û.û3 0.06 Eubacterium limosum 6515 2.0 2.û

Eubacterium 1 entum 6522 0.50 1.0 Actinomyces naeslundii 5920 0.25 0.25 Fusobacterium nucleatum 6516 0.12 0.12 6324 0.06 0.06 Fusobacterium varium 6052 15.6 3.9 .

Fusobacterium necrophorum 6568 0.06 0.06 Peptococcus asaccharo1yticus 6214 0.50 0.25 Peptococcus magnus 6258 0.06 0.06 Peptococcu~
aerogenes 6319 <0.03 0.06 Peptostreptococcus anaerobius 6321 0.12 0.12 The procedure for the above test is as follows:
Serial two-fold dilutions of drug are prepared in l.0 ml ~5 volumes of Schaedler 8roth, and ~.0 ml of molten (47 C) Wil~ens-Chalgren Agar Medium, infra,is added to the antibiotic-supplemented broth. After mixing with the antibiotic, the agar is poured into lO0 mm x 20 mm petri dishes. The dishes are allowed to stand on th~ bench overnight prior to inoculation.
Cultures are streaked on Wilkens-Chalgren Agar, and grown for 48 hours at 37 C in a BBL Anaerobe Jar. Growth from the plate is harYested, and a cell suspension is made in Schaedler broth to equal the turbidity of a 0.5 ~cFarland Standard ( lO8 cellslml). The suspension is pipetted into the wells of a Steers replicator, and ~pprox. 1-2 ~1 is deli~ered to the surface of the a~ar 1 16~863 pla~es. After allowing a few minutes for the inoculum to dry, the plates are placed in a BBL Anaerobe Jar (atmos-phere of 85~o N, 10% H, 5% CO2) and incubated at 37 C for 72 hours.
The Minimal Inhibitory Concentration (MIC) is read as the least amount of drug that inhibits growth. A very faint film of growth, or <3 colonies is considered negative.
Wilkins-Chalgren Agar Medium Dispense the following ingredients and dissolve in 1000 ml distilled water. The pH should be 7.0 - 7.2.
Trypticase 10 9 Gelysate 10 g Yeast Extract 5 g lS Glucose 1 9 NaCl . 5 g L-Arginine-Free Base 1 g Py-ruvic Acid-Sodiùm Sa1t I g Agar 15 9 Add Heme and +Vitamin K1 solutions to yield final concentrations of 5 yg/ml Hemin and 0.5 g/ml K1 Autoclave at 121 C for 15 minutes aerobically.
Heme Stock - 0.~ 9 Hemin ~ 10 ml 1 N NaOH f 990 ` ml H2 Autoclave at 121 C for 12 minutes.
Add 10 ml stock per liter of medium.
+ Vitamin K Stock - .05 ml Yit. K1 solution ~ 20 ml 95% ethanol Filter sterilize Add 0.2 ml stock per liter of medium The I.P. LD50 of U-57,930E in the mouse was found to be 592 mg/kg. This value is the resulting mean of two separate and identical LD50 determinations. This value is approximately 2 times the LD50 for clindamycin HCl.
The LD50 value should be interpreted as indicating that the acute r.P. toxicity of U-57,930E is approximately one-half that of clindamycin HCl.

116~86~

In vivo: Mouse Protection Test Orqanism & RouteClindamycin U-57930A Ratio _ aureus Subcut. 5.7(4.2-7.8)* <<5 Oral 12.3(8.8-17.3) 1-5 ca.10 S. hemolyticus Subcut. 2.3(1.6-3.3) .25 ca.10 Subcut. 3.3(2.6-4.2) .25(0.2-0.33~ 13 Oral 12.3(10.2-14.8) 2.9 (2.0-4.1) 4.2 K. pneumoniae >320 >320 *CDso ' s as mg/kg The procedure for the above test is as follows:
Mouse-protection Tests: Groups of 10 standard laboratory mice (CF-l Mice) weighing 18-20 gm were infected with approximately 100 LD50's of standardized bacterial ce.ll suspensions which had been maintained frozen at -170 C. Immediately before use, the suspen-sions were thawed quickly and properly diluted. Infec-tion was via the intraperitoneal route.
Treatment of the infected groups was begun immediate-ly and continued once per day for 4 days (first 24 hr period = 1). Groups of untreated infected mice served as virulence controls for the culture.
Seven days after the treatment regimen was begun the surviving animals were sacrificed and the median protect-ive dose of the antibiotic calculated on the basis of mortality rates in the treatment groups. The median protective dose and its 95X confidence interval were calculated according to the method of Spearmen & Karber as programmed on a 360 digital computer.
Also isolated from Example I, Part II, is compound Y A. This material is obtained as follows:
The filtrate which was saved from Part II was con-centrated to dryness under vacuum, the residue converted to its free base and chromatographed over silica gel using ` ` ` ` 1164863 CHCl3:methanol (6:1) as the eluting solvent. In this manner the least polar materia1 mentioned in Part II was obtained. It was converted to its HCl salt and recrystal-lized from acetone and water. This isomer is tentatively being assigned structure Y A.

C2Hs ~ ~ C~l~ NH_ Cll H HO ~ ~

V A ~ -CH3 OH
Epimerization of the carbonyl function attached to the piperidine ring of V and V A may be accomplished by methods wel~ known to those skilled in the art. The trans isomers V B and V C produced by these epimeriza-tionS may be isolated by conventional procedures such ascrystallization or chromatography.

C2Hs V eDimerize~ ~ " //0 V B

Ç2Hs V A epimeriZ~ ~ ~ VC
H C-7-Cl-M~

Alternatively, V and V A may be hydrolized to give the amino acids V D and V E which may then be epimerized by methods well known to those skilled in the art to V F and V &, respectively.
The amino acids V F and V G may be coupled with any of 1 16486~

the lincosaminides described earlier.

S C2Hs Hydrol i ze C) e,~ 'eri ze N C-OH N ~7-OH
Y D H Y F H

C2Hs C2Hs V Hydrolize ~ ~ epimerize ~ ~
N~ P-OH ~ N J ;~-OH
H H
VE V G

Thé D-cis isomer (V A) of U-57,930E has an anti-bacterial spectrum when tested on 8Hl broth as described previously in Table 1.
Organism UC No. MIC (~g/ml) _ aureus 76 250 S. fecalis 694 ~1000 5. pyoqenes 152 62.5 D. pneumoniae 41 62.5 E. coli 45 ~1000 K. pneumoniae 58 >1000 S. schottmuelleri 126 >1000 Ps. aeruqinosa 95 >1000 Example 2 - Other Analo~s bf 7-Cl-MTL.
By following the procedures of Example 1. but substituting the amino acid with the ~ollowing amino acids there are prepared the corresponding novel antibacterially-active analogs as their free bases or acid addition salts. The latter can be prepared by methods well-known to ~hose skilled in the art.
Amino Acid Analog U-45,863 C-OH
U-46,138 HO-~=O

~ ~ U-46,137 O U-46,337 - C-OH (Fast isomer on TLC-N MeOH:CHCl 95:5. Run on H silica gel plates) O U-46,465 N ~ C-OH (Slow isomer on TLC) H

8 U-46,699 C-OH (Slow isomer on TLC, I prepared from U-46,465) - 1 16~863 H5C2 ~LC-OH U-45,656 C2~l5 ~L U-45,652 C-OH

C2Hs~L U-46,701 C~Hsr~~O U-60,481 C-OH
N

~,~ U-44,469 ` N -OH

bH 2 C-OH U-45,657 Cl Example 3 - Analoqs of MTL
By follow;ng the procedures of Example 1, but substitùting the amino acid with the following amino acids and substituting MTL (J. Am. Chem. Soc., 89-2448, 1967 W. Schroeder, B. Bannister and H. Hoeksema) for 7-Cl-MTL, there are prepared the corresponding novel anti-bacterially-active analogs:
Amino Acid Analogs -U-46,136 ~ C-OH

U-45,653 C-OH
2~
U-60,493 (fast ~ isomer TLC) ~ N ~ -OH

~ U-60,492 (s~ow ~ ~ O isomer TLC) N C-OH

Example 4 - Analog~ 0~ ~Pi'-MT~
By following the procedures of Example 1, but substituting the amino acid with the following amino acids, and substituting epi-MTL (J. Chem. Soc. Perkin I

., 1 18~863 1974, p. 360-8, 8annjster) for 7-Cl-~lTL there are prepared the corresponding novel antibacterially-activ.e anal~gs;
~ Analoq ~ o U-46,135 N

F2Hs U-.45,6~9 Cpd. A

2 O ~C -OH

~_ Cpd. B.
O
ll ~ N ~ __C-OH

Exampl e 5 ~ Ana l o~s o~ epi ~7-CI ,~lTl~
8y following the procedures. of Exa~ple 1, but su~stituting the amino acid with the following aminQ
acids, and substituting epi-7-Cl-MTL for 7-Cl-MTL, there are prepared the correspond.ing novel antibacterially-acti ve an al ogs 1 1~4863 -3~- 3809A
Amino Acid Analog /~ .
C p d . C
~ ~C-OH

C 2H s ~ 1I Cpd, D
~ ~C -OH

: C p d . E
-OH
H

~2Hs ~ O
Cpd. F
~ N ~ ^-~-OH
H

Epi-7-Cl-MTL can be prepared b~ the procedure used to prepare 7-Cl-MTL with the exception that the starting material is epi-MT~ instead of MTL, Chemical and p~rsical c~aracterization o~ most of the compounds of Examples 2~5 are as follows~

~ 164863 - 3 5 ~ 38 O9A
C 11 N S Cl aD Mp.

U-45,863 1 47.80 5.62 7.44 8.51 9.41 CHCll -2 47.76 5.54 7.35 8.76 9.34 +286 96-100 U-46,138 1 47.80 5~62 7.44 8.51 9.41 - -2 47.30 5.74 6.91 8.51 9.44 U-46,137 1 47.80 5.62 7.44 8.51 9.41 EtOH
2 47.58 5.74 7.56 8.53 9.49 +216 189-lgO

U-46,136 1 50.26 6.19 7.82 8.95 MeOH
2 49.16 5.86 8.01 g.l5 +194 199-201 U-46,135 1 50.26 6.19 7.82 8.95 MeOH
2 50.50 6.19 7.98 9.27 +269~ 97-100 U-46,337A 1 42.96 6.73 6.68 7.65 16.91 EtOH
2 42.73 6.86 6.52 7.73 16.68 +206 220-30 U-46,465E 1 47.05 7.11 7.32 8.38 9.26 EtOH
2 46.63 7.37 7.12 8.47 9.33 +231 180-3 25U-46,699E 1 44.34 6.98 6.47 7.40 16.36 H20 2 44.79 7.24 6.25 7.36 16.42 +172 229-234 U-45,656 1 50.42 6.22 6.92 7.92 8.76 CHC13 2 50.85 6.39 6.72 7.54 8.93 ~2S0 U-45,652 1 50.42 6.22 6.92 7.92 8.76 CHC13 2 51.~3 6.40 6.65 7.56 8.02 +273 U-45,653 1 52.83 6.78 7.25 8.30 - M~OH
2 53.83 7.08 7.39 8.14 - +203 U-45,659 1 52.83 6.78 7.25 8.30 - CHC13 2 52.77 6.70 7.34 8.55 - +295 . -36- 3809A
U-46,701A 1 46.857.43 6.07 6.9515.37 2 - - _ _ _ U-44,469E 1 44.066.96 6.05 6.9215.30 2 44.836.68 6.07 6.7215.49 U-45,657 1 43.804.90 6.81 7.8017.24 MeOH
2 43.524.93 6.82 7.8217.41 +181 105-130 1 - CALCD.
2 - FOUND' Example 6'- Fusaric Ac~'d'Ami'de bf'7-Chloro-MTL.

H9C ~
OH7-Cl MTL

HgC4 ~ d HC-Cl C - NH~H
H ~ 0 ~ OH
~ S-CH3 (U-55~5Bl) OH
By following the procedure of Example 1, ~ut sub~ti-tuting the amino acid with fusaric acid, there is obtained U-55,581, Anal. Calcd. for C19H29ClN205S: C, 52,70; ~, 6.75; N,

6.47; S, 7~41; Cl, 8.19.
Found: C, 52.15; H, 6.65; N, 6~36; S, 7,21; Cl,

7.9~.
Example 7 - 4-Cis-n-Butyl -L-Pipecol ic Acid Amide of 7-Cl-MTL or U-60,970E

1~&4863 C4H9C4~9 C-7-Cl-~L ~ C/NH-CH (~)H20 H ~ 0~

OH
~C17H32Cl2N205S-(X)H20) A mixture of 4.0 9 (0.0093 mole) of starting material, 40 ml of water, 40 ml of methanol, 2 ml of 37% HCl, and

8.0 9. PtO2 catalyst were reduced on a Parr hydrogenator at 50 psi for 18 hours. The reaction was filtered to remove the catalyst and the filtrate concentrated under vacuum to give an amber oil. The oil was dissolved in 20 ml. of a 2:1 so1ution of CHCl3 and methanol and enough triethylamine added to neutralize the HCl pre-sent. This solution was then chromatographed over silica gel using a solvent system composed of CHCl3:methanol (2:1). Two main product fractions are obtained. The fractions contain;ng the faster moving material were pooled and evaporated under vacuum to give a white solid, fraction A. The fractions containing the slower moving material were pooled and evaporated under vacuum to give a white solid, fraction B. Fraction B was dissolved in a small amount of H20 and enough 37% HCl added to make the pH 2. Crystallization occurred. The solid was collected and recrystallized from H20 to give white crystals of the desired product, U-60,970E, m.p. 224-226 in a yield of 25-35%.
Anal. Calcd. for C17H32Cl2N205S: C, 47.99; H, 7.63; N, 5.89; S, 6.75; Cl, 14.92.
Found: C9 47.97; H, 7.42, N, 6.23; S, 6.90; Cl, 14.87.

~164863 -38- ~ 380gA
~leOH + 178 ~C, 1.0) CMR analysis supports the proposed structure.
S The Minimal Inhibitory Concentration (MIC) in ~g/ml of U-60,970E against var;ous bacter;a is as follows:
Or~anism UC# MIC
_ aureus 76 0.125 570 0.25 746 0.062 S. faecalis 694 0.25 S. pyogenes 152 0.008 D. pneumoniae 41 0.016 E. _oli 45 31.2 lS K. pneu~loniae 58 7.8 S. schottmuelleri 126 31.2 Ps. aeru~inosa 95 >125 The test procedure is as disclosed in Example 1.
U-60,970E was also tested in vivo in standard lab-oratory mice which were experimentally infected with bacteria. The test was conducted in comparison with U-57,930E. The following results show that U-60,970E
is significantly more active in vivo against D. pneumoniae I and III than U-57,930E. Against S. aureus and S.
hemolyticus U-60,970E demonstrates essentially the same activity as U-57,930E.
..

~1648~3 _39_ 3809A
__ ~ .. . . ._ _ , _ r~ '`I ~
~0 1~ N ~ CO
,~
a~
I u~ In ~O <~
.- u7 1~ .. c~l a S_ r-- ~ l _~ O ~ _ CO C~J C`J ~ C~l r-- r-- r~ ~ r~ r~
I C~l~0d'~) ~t O .
I_ O~
O
G ~0 ~ ~r ~ l ~ .. . .
--~ ~ .- o o o, F ~J In N ~ '~
C ~ O U~
C ~ ~) . .
r ~ . O O O
~ O ~ __ _ _, O t_~
~ ~ c~l 1~ ~ cn C~ _ ~ U~
V~ . . . .
_ r- O <~ O
15 ~, _ _ :~ C~ d c~i C~J C~l . C~l . ,_ .
U7 . ,~
oo U~
o ., ~ ' o ' o . . .~
t C~ In _ ~ _ C o _ _ 2 0.~ G~ C~J ~ ~ J
r~ .-- . . .
T ~ 00 r-- ,_ ,_ IIJ
a~ ~ ~ ~ ,_ r~ .-- a .-- .--u~ In r~ 00 V~ . . . ~ ~O
O ~
o a~ J 0 ~

~, r C C~.l 1~ r~ 1 ~J ~8 . .
~ O O r-- , D _ .~
~n o ~ i _ _ _ I_ 3 0 c ~ ~o , ._ ,_ 35 .E~ ~

c E ~ ¦
c ~ ol o Example 8 - 4-Cis-n-Butyl-D-Pipecolic Acid Amide of 7-Cl-MTL or U-61,734E

C4Hg ,C4Hg ~ C-7-Cl-MTL ~ ~ C~ NH-CH
H

OH
(C,7H32ClN205S (X)H20j Fraction A from the preceding experiment was converted to its HCl salt in the same manner as described for fraction B. A
25-35% yield of product was obtained whose CMR spectrum - was essentially identical to that obtained from fraction B.
Example 9 - Preparation of a_Compound in Which the Amino~
acid Portion Contains a Heteroatom In a 5-Membered Rin~

I Hydrolysis>
CH3~h~ ~ C// CH3- ~ -OH

7-Cl-MTL

CH3- ~ C// HC-Cl I ~ NH - CH

CH3 ~ SCH3 OH

~ 3809A
The aminoacid ester (see C.A. 69 - 67282M) may be hydrolyzed to the free acid by methods well known to those skilled in the art (acid or basic hydrolysis may be used). It may be obtained in the form of the HCl 5 salt or the zwitterion. The coupling of the aminoacid HCl with 7-Cl-MTL is accomplished in the same manner as described in Example 1, except that 67.7 9. (0.357 moles) of the aminoacid is used. After workup, as described in Example 1, the crude product may be purified via chroma-tography over silica gel and the product fractions com-bined and converted to the HCl salt.
Example 10 - Preparation of a Compound in l~hich the Amino-acid Portion Contains a Heteroatom in a 6-Membered Ring CH3 ~ Hydrolysis~ ~ O

O`NC-O-CH3 ~ C-OH

Cl Cl CH3 +
CH3 ~ HC-Cl 7-Cl-MTL

3 ~ -CH, Cl OH
The aminoacid ester (see C.A. 68 - 59465N) may be hydrolyzed to the free acid by methods well known to those skilled in the art (acid or bas-c hydrolysis may be used). It may be obtained in the form of the HCl salt I 16486~

-42-` 3809A
or the zwitterion. The coupling of the aminoacid-HCl with 7-Cl-MTL is accomplished in the same manner as described in Example 1 except that 103.6 9. ~0.357 moles) of the aminoacid is used. After workup, as described in Example 1, the crude product may be purified via chroma-tography over silica gel and the product fractions com-bined and converted to the HCl salt.
Example 11 - 2-Phosphate Analo~s The 2-phosphate analog of the compounds prepared in Examples l-lOcan be prepared by procedures well-known to those skilled in the art. By obvious appropriate modifi-cation, the procedure disclosed in U.S. Patent 3,487,C68 may be used. 8asically, any procedure would first involve the protection of vulnerable groups by methods well-known to those skille-d in the art--which would then be removed upon com-pletion of the phosphorylation.
Example 12 - 2-Palmitate Analoqs The 2-palmitate analog of the compounds prepared in Examples l-lOcan be prepared by procedures well-known to those skilled ;n the art. By obv;ous appropriate modifi-cation, the procedure disclosed in U.S. Patent 3,580,904 may be used. Basically, any procedure would first involve the protection of vulnerable groups by methods well-known to those skilled in the art which would then be removed upon com-pletion of the acyJation with palmitoyl chloride.
The Minimal Inhibitory Concentration (MIC's) of arepresentative number of the compounds prepared.~in Examples 2-8 follows. The test procedure is as given supra.

1~6486'3 ` 43 3809A
,. ~-o~ ~' C~ U~ o o o o o o ~Y
~o ~- , ~ ~ o o o o o o l ~
U~ _ _ _ _ ~ _ ~ _ r~ r~ ~ _ ~
O~ Xy~ O O O ~ O O O O O O ~
~ ~ S S U~ LS~ O ~) O O O O O O ~) 'r ~_3_~_~C ~ c~J ~1 O rA
~r Lf~ _ __ _ __ __ _ I

O ~q ~I U~ ~ O O O O O O Q
r~ / ~ -r D C~l C~l l O O O O O O l a~
~ ~ ~D U~ O O O O O O=t~
~ _ ~ ~ ~ A _ _ I~
s _ _ _ _ _ _ I 1-1--1~ o~ ~ o o o o o o o o ol ol olo El ~ ~ o o o o o o o o ol ol ol o l S_ ~ " O o o u~ O O O O 01 ol ol n ~ ~ U~ __ r~ r~ ~ r~ ~_ ~_ El~ ~ ~ ~ A

-I ' u~ ~ o c o o c~ o olol olo gl o oO O g gl gl ol 0~ g o=~
E l-n ,_ I ,_ ~ ,_ , I r~ I r~ I r I ~-- ~~
-~ 1~ ~ I ~ ~ A ~ ~ ~L~
'l l l l _ l l l l ol O ol O ol o ol ol ol ol o r ¦ ~g-- g l ~ ¦ ~ ~ o l o l ~ J
- I ~ l ~l A ~¦ ¦ ~¦ A¦ ~
+ , ! ---I I, I
I~D X Ln~ I I I I I
~u~ ~ C~ ol o ol ol ol ol D ~ ~1 'I ~1 ol o ol ol gl ol ' o-~l l l -I ~ 1'-1''1'^1 x ~ . . l _ 1,, 1 ~ lI ~ 1 0 I ~ I a~ l I ~
'`I ~1 '-I ,1 'I ~ ~ "I ~
~1 ''I ~1 ,~ , ~:r ~ l l -v7 ¦ ~ I ~ ~ s l o l ~ 1 ~;1 ~1 ~1 ~1 ~ I ~1 .o~I El ~1 ~I E
Q~ l o l ~ 1 ~ _ c~ ~ 1 o l s_ s~ I El ~1 ~
~1 ~1 ~I aJI ~1 ~ ~ Cl ~>I ~1 C
~ '1 1' ~ ~
I v71 ~I.v~l ~1 ~1 ~n ~ c~ ~CI vll .~1 ~

_ --- .1, a~ ~ o ol oi o ol 'l v s_ I o ol~lo 0~ I j9 3E
D X ~ ~ a o ~ __ o o o o o c ~_ _ ~:I ~ V . O O O O O V o E 1--~ O O O O O O O O O O O Q) _I
c~ ~ o o o u~ O o o o o o ~ ~ E ~
~ C.T O O O C~J O O O O O O C~J V~ O
~0 ~ _ r _ r~ r~ _ A A r~ _ l ~ C
C~ O O O O O O O O O O O
r~ f-- O O O O O O O O O O O
^ ~ ~ O O O O O O O O O O O
~ _~ r--r- ~ ~ _ ,_ r . ,_ C)o _ _ _ _ _ _ A _ A A _ J O O O O O O O O O O O
E ~ ~ o o o o o o o o o o o _ ~ ~ z o o o o o o o o o o o L a D ~ ~ ~--A _ r~ _ r~ r~ r~ ~_ ~ ~_ O_ ~ ~ l~ O O O O O O O O O O O ll ~_ E ~o ~ O o ~o roo--O . ~o O ,oo--O ~o O _ .
2 _ _ A __ _ ~ _ _~r _ :~
~ ~ ~ ~ ~ V ~ ~ ~ ~0=~~
~ ~ oO O In 0=~

. ~. . I I I 1~ - ~ ~ ~ ~ ~ _ x IIIIl ~1 l I 1 1'`~1 1 ~ I I
I I ~ l 0 ~ -I
I I 1 ~1 ~01 ~ ~_~ ~
~ol ,_1 `I 'I ~.1 '~ " S-l =' ~1 '`I'~ I ,1 'I In ~-~ ~ -I a)l ~1 ~1 01 ul ~1 cr ~ aJI o ,al ~I ~l ~1 ~ I u~ ,_ ~1 c o E I ~ o ~1 ~ 1 ~o E ~1 ~ E l a~ l ~1 01 ~1 ~_ cr: ~ 01 S_ ~1 , I < I s_l El ~1 _ _ aJ ~1 ~ ~1 v~ o ~ e ~ o 1 ~648~3 MIC in mcq/ml U-Number and Structure _ U-60,493 U-60,492 U-~0,481 ~ ~ R ~ X
.
S. aureus UC 76 1000 1000 2.0 _ .
S. aureus UC 570 1000 >1000 3.9 S aureus UC 746 250 250 2.0 S h~nolyticus UC 152 7.8 15.6 ~1.0 St. faecal;s UC 694 1000 >1000 31.2 S. lutea UC 130 ~. coli UC 45 _ >1000 ~1000 500 -P. vulgaris UC 93 >1000 >1000 1000 K pneumoniae UC S8 ~1000 >1000 250 S S cb~ t t~l e I 1 e r i DC : Z 6 > 1 000 > 1 000 1000 Ps. aeruginosa UC 95 ~1000 >1000 >1000 O. pneumoniae UC 41 15.6 31.2 Fast Slow isomer isomer on TLC on TLC IR &
NMR OK
, O O
X=l-7-Cl-MTL Y=C-MTL
. R=C2H5 . . - .
Since the compounds of the subject invention are active against various Gram-positive and Gram-negative microbes, they can be used in various environments to 3~ inhibit such microbes. For example, they can be used as disinfectants to inhibit S. aureus on washed and stacked food utensils contaminated with this bacterium. They also can be used as disinfectants on various dental and medical - equipment contaminated with S. aureus. Further, the compounds of the invention can be used as bacteriostatic rinses for laundered clothes, and for impregnating papers ~ 6- 3~09A
and fabrics; and, they are also use~ul f~r suppressing the growth or sensitive organisms in plate assays and other microbiological media.
The compounds of the subject invention exist in the protonated or non-protonated forms according to the pH of the environment. When the protonated form is intended, the compounds exist as pharmaceutically-acceptable acid-addition salts, and when the non-protonated form is intended, the compounds exist as the free base, The free bases can be converted to stable acid-addition saits by neutralizing the free base with the appropriate acid, about pH 7.0, and advantageously to about pH 2 to pH 6.
Suitable acids for this purpose include hydrochloric, sulfuric, phosphoric, thiocyanic, fluosilicic, hexafluoro-arsenic, hexafluorophosphoric, acetic, succinic, citric,lactic, maleic, fumaric, pamoic, cholic, palmitic, mucic, camphoric, glutaric, glycolic, phthalic, tartaric? lauric, stearic, salicylic, 3-phenylsalicylic, 5-phenylsalicylic, 3-methylglutaric, o.rthosulfobenzoic, cyclohexanesulfamic, cyclopentanepropionic, 1,2-cyclohexanedicarboxylic, 4-cyclohexanecarboxylic, octadecenylsuccinic, octenyl- -succinic, methanesulfonic, helianthic, Reinecke's, dimethyldithiocarbamic, hexadecylsulfamic, octadecyl-sulfamic, sorbic, monochloroacetic, undecylenic, 4'-hydroxyazobenzene-4-sulfonic, octadecylsulfuric, picric, benzoic, cinnamic, and like acids.
The acid-addit;on salts can be used for the same purposes as the free base or they can be employed to up-grade the same. For example, the free base can be con-verted to a water-insolub1e salt, such as the picrate, which can be subjected to purification procedures, for example, solvent extractions and washings,.chromatography, fractional liquid-liquid extractions, and crystallization, and then used to regenerate the free base form by treat-ment with alkall or to make a different salt by meta-thesis. Or the free base can be converted to a water-soluble salt, such as the hydrochloride or sulfate, and .

1 164g63 the aqueous solution of the salt extracted with various water-immiscible solvents before regenerating the free base form by treatment of the thus-extracted acid solu-tion, or converted to another salt by metathesis.
In addition to the antibacterial uses, disclosed above, the free bases can be used as buffers or as ant-acids, The thiocyanic acid addition salt when condensed with formaldehyde forms resinous materials useful as pickling inhibitors according to U.S, patent Nos.
2,425,320 and 2,606,155. T~e free bases also ma~e good vehicles for toxic acids. For example, the fluosilicic acid addition salts are useful as mothproofing agents according to U.S, patent Nos. 1,915,334 and 2~075,359 and the hexafluoroarsenic acid and hexafluorophosphoric acid addition salts are useful as parasiticides according to U.S. patent Nos. 3,122,536 and 3,122,552, The compounds of t~e su~ject invention are useful as antibacterial agents in suitahle compositions, These compositions are preferabl~ presented for administration to humans and animals in unit dosage forms, such as tablets, capsules, pills, powders, granules, sterile parenteral solutions or suspensi:ons, and ~ral solutions or suspensions, and oil-water emuls~ons containing suit-able quantities of the active compound in the form of the ~ree ~ase? or its pharmacologicall~ accepta~le salts.
For oral administration, either solid or fluid unit dosage forms can be prepared. For preparing solid compo-sitions such as ta~lets, the principal actiye ingredient is mixed with conventional ingredients such as talc, magnesium stearate, dicalcium phosphate, magnesium aluminum silicate, calcium sulfate, starch, lactose, acacia, methylcellulose, and functionally similar mater-ials as pharmaceutical diluents or carriers. The tablets can be laminated or otherwise compounded to provide a dosage form affording the advantage of prolonged or delayed action or predeterm~ned successive action of the enclosed medication. For example, the tablet can ~ 16486~
-4~ 3809A
comprise an inner dosage and an outer dosd~e component, the latter being in the form of an envelope over the former. The two components can be separated by an enteric layer which serves to resist disintegration in the stomach and permits the inner component to pass intact into the duodenum or to be delayed in release. A Yariety of materials can be used for such enteric layers or coat-ings, such materials including a num~er of polymeric acids or mixture of polymeric acids with suc~ materials as shellac, cetyl alcohol, cellulose acetate phthalate, styrene maleic acid copol~mer and the like. Alternatively, the two component system can ~e utilized for preparing .
tablets containing two or more incompati~le active ingredients. Wafers are prepared in the same manner as tablets, differing only in shape and the inclusion of sucrose or other sweetener and flavor. In their s;mplest embodiment, capsules, like tablets, are prepared by mix-ing the compound of the formulation with an inert pharma-ceutical diluent and filling and mixture into a ~ard gelatin capsule of appropriate size. In another embodi-ment, capsules are prepared ~y filling hard gelatin cap-sules with polymeric acid coated ~eads containing the active ~ompound. Soft gelatin capsules are prepared by machine encapsulation of a slurry of the active compound with an acceptable vegetable oil, light liquid petrolatumor other inert oil.
Fluid unit dosage forms for oral administration such as syrups, elixirs, and suspensions can be prepared, The water-soluble forms of the active compound can be dis-solved in an aqueous vehicle together with sugar,aromatic flavoring agents and preservatives to form a syrup. An elixir is prepared by using a h~dro-alcoholic (ethanol) vehicle with suitable sweeteners such as sucrose together with an aromatic flavoring agent. Suspensions can be prepared of the insoluble forms with a syrup vehicle with the aid of a suspending agent such as acacia, tragacanth, methylcellulose and the like, ~ 164863 -49- 38~9A
Topical ointments can be prepared by d;spersing the active compound in a suitable ointment base such as petrolatum, lanolin, polyethylene glycols, mixtures there-of, and the like. Advantageously, the compound is finely divided by means of a colloid mil7 utilizing light liquid petrolatum as a levigating agent prior to dispersing in the ointment base. Topical creams and lotions are pre-pared by dispersing the compound in the oil phase prior to the emulsification of the oil phase in water.
For parenteral administration, fluid unit dosage ~orms are prepared utilizing the active compound and a sterile vehicle, water being preferred. T~e active com-pound, depending on the form and concentration used, can be either suspended or dissolved in the vehicle. In pre-paring solutions, a water-soluble form of the active compound can ~e dissolved in water for injection and filter steril.ized before filling into a suitable vial or ampul and se~ling. Advantageously adjuvants such as a local anesthetic, preservative and buffering agents can be dissolved in the vehicle. To enhance the stability, the composition can 5e frozen after filling into the vial and the water removed under vacuum. The dry lyophilized powder is then sealed in the vial and an accompanying vial of water for injection is supplied to reconstitute the powder prior to use. Parenteral suspensions are prepared in substantially the same manner except that the active compound is suspended in the vehicle instead of being dissolved and sterilization cannot be accomplished by filtration. The active compound can be sterilized by exposure to ethylene oxide before suspendin~ the sterile veh;cle. Advantageously, a surfactant or wetting agent is included in the composition to facilitate uniform distribution of the active compound.
Tne term unit dosage form as used in t~e specifica-tion and claims refers to physically discrete units suit-able as unitary dosages for human subjects and animals, each uni~ containing a predetermined quantity of active material calculated to produce the desired therapeutic e~fect in association with the required pharmaceutical diluent, carrier or vehicle. The specifications for the novel unit dosaQe forms of t~is invention are dictated by and directly dependent on (a) the unique characteris-tics of the active. material and the particular thera-peutic effect to be achieved, and (b) the limitations inherent in the art of compounding such an active material for therapeutic use i.n humans and animals, as disclosed in detail in this specification, these ~eing features of the present invention. Examples of suita~le unit dosage forms in accord with this invention are tablets, capsules, pills, troches, supposito.ries, powder packets, granules, wafers, cachets, teaspoonfuls, tablespoonfuls, dropperfuls, ampuls, lS vials, segregated.multiples of any of t~e foregoing, and other forms as herein described, In addition to t~e adminjstration of the active com-pound as the princ.ipal active ingredient of compositions ~or the treatment of t~e conditions descri~ed herein, the 2~ said compound can ~e included with other types of com-pounds to o~tain advantageous com~inations of properties, Such combina.tions i.nclude t~e active compound with anti-biotics suc~ as spectinom~cin, chloramphenicol, novo-~iocin, di~ydranovobiocin, tetracyclines (e,g., tetra-cy-cline, oxytetracycline and c~.lortetracycline) ? peni-cillins, erythromycin ? kanamycin, streptomycin ? neomycin ?
polymyxin, ~acftractn, nystatin, filipin ? fuma~illin and endomy~cin to broaden the ~acterial spectrum of the compo-sition and for synergistic action against particular bacteria.;. steroids having anti-inflammatory activity such as hydrocortisone~ prednisolone, 6a-meth~lprednisolone, 6-fluoroprednisolone and the lfke; analgesics such as aspirin, sodium salicylate (acetylsalicylic acid)-anhydride, N-acetyl-p-aminophenyl and salicylamide, antihistamines, such as chlorpheniramine maieate, diphenylhydramine, promet~azine, pyrathiazine, and the like; sulfas, such as sulfadiazine, sulfamethazine, -51- 3~09A
sulfamera~ine sulfacetamide, sulfadimethyloxazole?
sulfamethi 201 e, and the like; antifungals, such as undecylenic acid, sodium propionate, salicylanilide, sod'ium caprylate, and hexetidine; and the vitamins.
S The dosage of the active compound for treatment depends on route of administration; the age, weight, and condition of the patient; and the particular disease to be treated. A dosage schedule of from about 15 to SOO
mg., 1 to 4 times dail~ (every six hours), embraces the effective range for the treatment of most conditions for which the compositions are effective~ For ch~ldren, the dosage is calculated on the basis of 15 to 30 mg,lkg./day to be administered every six ~ours.
The active compound is compounded with a suita~le pharmaceutical carrier in unit dosage form for convenient and effective administration. In the preferred embodi-ments of this invention, the dosage units contain the compound in. 15, 30, 50, 125, 250 and 500 mg. amounts for systemic treatment; in 0.25, 0.5, 1, 2 and 5% amounts for topical or localized treatment; and 5 to 65~ w/v for parenteral treatment. The dosage of compositions contain-ing the active compound and one or more other actiYe ingredients is to be determined with reference to the usual dosage of each such ingredient.
The following examples are i'llustrative of the best mod6 contemplated by the inventor for carrying out his invention and are not to be construed as limiting.
The examples use U-57,930E or U-60,970E as the active compound, but it'should be understood that this is only exemplary of the other active compounds of the subject invention.
As distinguished from the previous examples, the follow-ing are Compos i tion Exampl es.
Composition Exampl e 1 Capsul es One thousand two-piece hard gelatin capsules for ora use, each containing 250 mg. of U-57,930E or U-60,970E are prepared from the following types and amounts of materials:

Gm.
U-57,930E or U-60,970E 250 Corn starch 100 Talc 75 Magnesium stearate 25 The materials are thoroughly mixed and then encap-sulated in the usual manner.
The foregoi ng capsules are useful for the systemic treatment of infection in adult humans by the oral 10 administration of 1 capsule every 4 hours.
Using the procedure above, capsules are similarly prepared containing U-57,930E or U-60,970E in 15, 30, 50, 125 and 500 mg. amounts by substituting 15, 30, 50, 125, and 500 gm of U-57,930E or U-60,970E for the 250 gm used above.
15 Composition Example 2 Capsules One thousand two-piece hard gelatin capsules for oral use, each containing 200 mg. of U-57,930E or U-60,970E and 250 mg. of tetracycline hydrochloride, are prepared from the follow-Ing types and amounts of ingredients;
Gln.
U-57,930E or U-6Q,970E 200 Tetrac~cline hydrochloride250 Tal c 75 Magnesium stearate 25 The ingredients are thoroughly mixed and then encap-sulated in the usual manner, The foregoing capsules are useful for t~e systemic treatment of infection in adul t humans ~y the oral - 3Q administration of 1 capsule ever~ 6 hours~
Using the procedure above, capsules are similarly prepared containing U-57,930E or U-60,970E and each of the following antibi otics -i-n p~ace- of--~e-trac~cline by substi tuting 250 gm. of such other antibiotic for tetracycline: chlor-35 amphenicol, oxytetrac~cline, chlortetracycline~ fumag;llin7erythromycin, streptomycin, dihydronovobiocin and novo-biocîn. When a penic~llin, such as potassium penicillin 3~09AG, ;s to be used in place of tetracycline, 250,000 uni-ts per capsule is employed.
Such combination products are usefu1 for the system-ic treatment of mixed infections in adult humans by the S oral administration of 1 capsule every 6 hours.
Composition Example 3 Tablets One thousand tablets for oral use, each containing 500 mg. of U-57,930E or U-60,970E are prePared from the following types and amounts of materials:
Gm.
ll-57,930E or U-60,970E 500 Lactose 125 Corn starch 65 Magnesium stearate 25 Light liquid petrolatum 3 The ingredients are thoroughly mixed and slugged.
The slugs are broken down by forcing through a number sixteen screen. The resulting granules are then com-pressed into tablets, each tablet containing 500 mg. of U-57,930E or U-60,970E.
The foregoing tablets are useful for systemic treat-ment of infections in adult humans by oral administration of 1 tablet every 4 hours.
Using the above procedure, except for reducing the amount of U-57,930E or U-60,970E to 250 gm., tablets containing 250 mg. of U-57,930E or U-60,970E are prepared.
Composition Example 4 Tablets One thousand oral tablets, each containing 250 mg.
of U-57,930E or U-~0,970E, and a total of 250 mg. (83.3 mg. each) of sulfadiazine, sulfamerazine, and sulfamethazine, are pre-pared from the followi'ng types~a~nd'''amounts of materials:
~m.
U-57,930E or U-60,97~E 250 Sul fadiazine 83.3 Sulfamerazine 83.3 11648~
-5~- 3809A
Sulfamethazine 83.3 Lactose 50 Corn starch 50 Calcium stearate 25 Light liquid petrolatum 5 The intredients are thoroughly mixed and slugged.
The slubs are broken down by forcing through a number sixteen screen. The resulting granules are then com-pressed into tablets, each containing 250 mg. of U-57,930E
or U-60,970E and a total of 250 mg. (83.3 mg. each) of sulfadiazine, . . .
sulfamerazine, and sulfa~ethazine.
The foregoing tablets are useful for systemic treatment of infe~tions by the oral administration of 4 tablets first and then 1 every six hours.
For the treatment of urinary infections, the triple sulfas in the above formulation is advantageously replaced by 250 gm. of sulfamethylthiadiazole or 250 gm.
of sul facetamide.
Composition Example 5 Oral Syrup One thousand cc. of an aqueous suspension for oral use, containing in each 5 cc. dose, one-half gram of total sulfas and 250 mg. of U-57,930E or U-60,970E is prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U 6Q,~70E 50 Sul fadiazine 33.3 Sulfamerazine 33.3 Sulfamethazine 33.3 Citric acid 2 Benzoic acid Sucrose 700 Tragacanth 5 Lemon oil 2 ccO
Deionized water, q.s. 1,000 cc.
The citric acid, benzoic acid, sucrose, tragacanth, and lemon oil are dispersed in sufficient water to make -55- 3~09A
~50 cc. of solution. T~le U-57,930E or U-60,970E and finely powdered sulfas are stirred into the syrup until uniformly dis-tributed. Sufficient water is added to make 1 ,000 cc.
The composition so prepared is useful in the systemic 5 treatment of pneumonia in adult humans at a dose of 1 teaspoonful 4 times a day.
Composition Example 6 Parenteral Solution A sterile aqueous solution for intrarnuscular use, containing in 1 cc. 200 mg. of U-57,930E or U-60,970E is prepared from the following types and amounts of materials:
Gm.
U-57,930E or U-60,970E 200 Lidocaine hydrochloride 4 Methyl paraben 2.5 Water for injection, q.s. 1,000 cc.
The ingredients are dissolved in water and the solu-tion sterilized by filtration. The sterile so7ution is filled into vials and the vials sealed.
20 Composition Example 7 Parenteral Preparation A sterile aqueous solution for intramuscular use, containing in 1 cc. 200 mg. of U-57,930E and U-60,970E and 400 mg. of spectinomycin sulfate, is prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 200 Spectinomycin sulfate 400 Lactose 50 Water for injection, q.s. 1,000 cc.
The U-57,930E or U-60,970E, spectinomycin sulfate and lactose are dissolved in the water and the solution sterilized by filtration. The sterile solution, in the amount of 2 cc., is aseptically filled into sterile vials and frozen. The 35 water is removed under high vacuum and the vials contain-ing the lyophilized powder are sealed. Just prior to use, sufficient sterile water for injection to make 2 cc. of -~6- 38Q9A
solution is added to the vial.
Composition Example 8 Topical Ointment One thousand gm. of 0.25% ointment is prepared from 5 the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 2.5 Zinc oxide 50 Calamine 50 Liquid petrolatum (heavy) 250 Wool fat 200 White petrolatum, q.s. 1 ,000 gm.
The white petrolatum and wool fat are melted and 100 gm. of liquid petrolatum added thereto. The U-57,930E or U-60,970E, zinc oxide and calamine are added to the remaining liquid petrolatum and the mixture milled until the powders are finely divided and uniformly dispersed. The powder mix-ture is stirred into the white petrolatum mixture and stirring continued until the ointment congeals.
The foregoing ointment is usefully applied topically to the skin of mammals for the treatment of infection.
The foregoing composition can be prepared by omitting the zinc oxide and calamine.
following the procedure above, ointments are similarly prepared cont~ining U-57,930E or U-60,970E in 0.5, 1, 2, and 5%
amounts by substituting 5, 10, 20 and 50 gm. of U-57,930E or U-60,970E for the 2.5 gm. used above.
Composi tion Example 9 Cream One thousand gm. of a vaginal cream are prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U~60,970E 50 Tegacid Regular1 150 Spermaceti 100 Propylene glycol 50 Polysorbate 80 5 1 16486~
~57~ 3~09A
,~lethylparaben Deioni~ed water, q.s. 1,000 gm.
' Sel,-emulsifying glyceryl monostearate from Goldschmidt Chemical Corporation, New York, N~Yo The Tegacid and spermaceti are melted together at a temperature of 70-80 C. The methylparaben is dissolved in about 500 gm. of water and the propylene glycol, Polysorbate 80, and U-57,930E or U-60,970E are added in turn, main-taining a temperature of 75-80 C. The methylparaben mixture is added slowly to the Tegacid and spermaceti melt, with constant stirring. The addition ii continued for at least 30 minutes with continued stirring until the temperature has dropped to 40-45 C. The pH of the final cream is adjusted to 3.5 by incorporating 2.5 gm. of citric acid and 0.2 g. of dibasic sodium phosphate dis-solved in about 50 gm. of water. Finally, sufficient water is added to bring the final weight to 1,000 gm. and the preparation st;rred to maintain homogeneity until cooled and congealed.
The foregoing composition is useful for the treat-ment o~ vaginal infections in humans.
Composition Example 10 Ointment, Ophthalmic One thousand gm. of an ophthalmic ointment containing 0.5% U-57,930E or U-60,970E are prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 5 Bacitracin 12.2 Polymyxin B sulfate (10,000 units/mg.) Light liquid petrolatum 250 Wool fat 200 White petrolatum, q.s. 1,000 gm.
The solid ingredients are finely divided by means of an air micron;zer and added to the light liquid petrolatum.
The mixture is passed through a colloid mill to uniformly -5~- 3~09A
distribute the micro~ ed particles. The wool fat and white petro1atum are melted together, strained, and the te~perature adjusted to 45-50 C. The liquid petrolatum slurry is added and the ointment stirred until congealed.
S Suitably the ointment is packaged in one dram ophthalmic tubes.
The foregoing ointment is usefully applied to the eye for treat~ent of localized infection in humans and other animals Advantageously the foregoing composition can contain 5 gm. (0.5%) of methylprednisolone for the treatment of inflammation, and, alternatively, the bacitracin and polymyxin 8 sulfate can be omitted.
Composition Example 11 Eye-Ear Drops One thousand cc. of a sterile aqueous solution for eye or ear use containing 10 mg. of U-57,930E or U-60,970E and 5 mg.
of methylprednisolone in each cc. is prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 10 Methylprednisolone phosphate sodium 5 Sodium citrate 4.5 Sodium bisulfite Polyethylene glycol 4000 120 Myristyl-y-picolinium chloride 0.2 Polyvinylpyrrolidone Deionized water, q.s. ad 1000 cc.
The ingredients are dissolved in the water and the resulting solution is sterilized by filtration. The solution is aseptically filled into sterile dropper containers.
The composition so prepared is useful in the topical treatment of inflammation and infection of the eye and ear as well as other sensitive tissues o~ the animal body.
Composition Example 12 Troches 1 16486~

Ten thousand troches are prepared from the follo~Ying types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E lOO
Neomycin sul fate 50 Polymyxin B sùlfate (lO,OOO
units/mg.) Ethyl aminobenzoate 50 Calcium stearate 150 Powdered sucrose, q.s. 5,000 gm.
The powdered materials are mixed thoroughly and then compressed into half gram troches following the usual techniques for the preparation of compressed tablets.
The troches are held in the mouth and allowed to dissolve slowly to provide treatment for the mouth and throat of humans.
Composition Example 13 Suppository, Rectal One thousand suppositories, each weighing 2.5 gm. and containing lOO mg. of U-57,930E or U-60,970E are prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E lOO
Polymyxin B sulfate (lO,OOO
units/mg.) 1.25 Methylprednisolone Ethyl aminobenzoate 75 Zinc oxide 62.5 Propylene glycol 162.5 Polyethylene glycol 4,000 q.s.
2,500 gm.
T~e U-57,930E or U-60,970E, polymyxin B sulfate, methylpredniso-lone, ethyl aminobenzoate, and zinc oxide are added to the propylene glycol and the mixture milled until the 35 powders are finely divided and uniformly dispersed. The polyethyl ene glycol 4000 is melted and the propylene glycol dispersion added slowly with stirring. The 1 16486~
- ` -60- 3~09A
suspension is poured into unchill~d molds at 40 C.
The composition is allowed to cool and solidify and then removed from the mold and each suppository foil wrapped.
S The foregoing suppositories are inserted rectally for local treatment of inflammation and infection.
Alternatively, the foregoing composition can be pre-pared omitting the steroid.
Composition ExamPle 14 Mastitis Ointment One thousand gm. of an ointment for the treatment of mastitis in dairy cattle is prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 25 Methylprednisolone acetate 0.5 Light liquid petrolatum 300 Chlo-robutanol, anhydrous 5 Polysorbate 80 5 2% Aluminum monostearate-peanut oil gel 400 White petrolatum, q.s. 1000 gm.
The U-57,930E or U-60,970E and methylprednisolone acetate are~
milled with the light liquid petrolatum until finely divided and uniformly dispersed. The chlorobutanol, polysorbate 80, peanut oil gel and white petrolatum are heated to 120 F. to form a melt and the liquid petrolatum dispersion stirred in. With continued stirring, the dis-persion is allowed to cool (and congeal) to room tempera-ture and is filled into disposable mastitis syringes in10 gm. doses.
Composition Example 15 Animal Feed One thousand gm. of a feed mix is prepared from the following types and amounts of ingredients:
Gm.
U-57,930E or U-60,970E 10 ` 1164~6~
-61- 3~09A
Soybean meal ~
Fish meal 400 Whéat germ oil 50 Sorghum molasses 140 S The ingredients are mixed together and pressed into pellets. The composition can be fed to laboratory animals, i.e., rats, mice, guinea pigs, and hamsters for prophylaxis during shipping.
For other animals such as poultry, e.g., chickens, ducks, turkeys, and geese, the composition can be added to the animal's regular feed in an amount calculated to give the desired dose of U-57,930Eor U-60,970E
Compos_tion Example 16 Following the procedure of each of the preceding Composition Examples 1-15, inclusive, each antibacterially-active compound of the subject invention is substituted in an equivalent amount for the U-57,930E or U-60,970E shown in the example to p~ovide therapeutic properties.
Similarly, each of the above free base compounds can be used in the form of a pharmaceutically (or pharmaco-Iogically) acceptable acid addition salt, e.g., hydro-chloride, sulfate, nitrate, phosphate, citrate, lactate, acetate, tartrate and succinate.
Further, the 2-phosphate andtor 2-palmitate of each of the above antibacterially-active invention compounds can be substituted as the active ingredient to provide compositions having therapeutic properties.

Claims (4)

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

1. A process for preparing a compound of the general formula:

wherein R1, which can be singly or multiply substituted in the 2, 3, 4, 5 or 6 position of the pyridine ring not already suhstituted by R2, is selected from the group consisting of hydrogen, alkyl and substituted alkyl, wherein the alkyl portion is from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof, cycloalkyl and substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl and substituted phenyl, -(CH2)m-OH, -(CH2)m-NR4R5, and isomeric forms thereof, wherein m is an integer of from 1 to 8, inclusive, and R4 and R5 are selected from the group consisting of hydrogen, alkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof, and wherein the substituents are: for alkyl one or more groups selected from F, Cl, Br, I, =O, -OH, -NH2, secondary-alkyl amine, tertiary-alkyl amine, wherein alkyl is as defined above, S, -SH
and phenyl, for cycloalkyl as defined for alkyl, for oxygen a group selected from alkyl, as defined above, and aryl, for nitrogen a group selected from acyl of from 2 to 18 carbon atoms, inclusive, monoalkyl and dialkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms of the acyl and alkyl groups, and for phenyl one or more groups selected from F, Cl, Br, I, -OH, -NH2, secondary-alkyl amine, tertiary-alkyl amine, wherein alkyl is as defined above, -SH and phenyl; and wherein R2, which can be singly substituted in any position of the pyridine ring not already sub-stituted by R1, is:

-?-X
and X is the amino function of a compound selected from the group consisting of 7(R)-hydroxy-methyl 1-thio-a-lincosaminide, 7(S)-hydroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(R)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(S)-methoxy-methyl 1-thio-.alpha.-lincosaminide, 7-deoxy-7(S)-(methylthio)-methyl 1-thio-.alpha.-lincosaminide, 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl 1-thio-.alpha.-lincosaminide and 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl 1-thio-.alpha.-lincosaminide; which comprises reacting an amino acid of the general formula:

wherein R1 is as defined above, and the -OOOH group is singly substituted in any position of the pyridine ring not already substituted by R1, with a sugar amine compound selected from the group consisting of 7(R)-hydroxy-methyl 1-thio-a-lincosaminide, 7(S)-hydroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(R)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(S)-methoxy-methyl 1-thio-.alpha.-lincosaminide, 7-deoxy-7(S)-(methylthio)-methyl 1-thio-.alpha.-lincosaminide, 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl 1-thio-.alpha.-lincosaminide and 7-deoxy-7(S)-(3-hydroxypropylthiol-methyl 1-thio-.alpha.-lincosaminide; and, when required, preparing a pharmaceutically acceptable acid-addition salt.

2. A process according to claim 1, wherein R1 is in the 4 position and is alkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof; and wherein R2 is in the 2 or 3 position.

3. A compound of the general formula:

wherein R1, which can be singly or multiply substituted in the 2, 3, 4, 5 or 6 position of the pyridine ring not already substituted by R2, is selected from the group consisting of hydrogen, alkyl and substituted alkyl, wherein the alkyl portion is from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof, cycloalkyl and substituted cycloalkyl, substituted oxygen, substituted nitrogen, halogen, phenyl and substituted phenyl, -(CH2)m-OH, -(CH2)m-NR4R5 and isomeric forms thereof, wherein m is an integer of from 1 to 8, inclusive, and R4 and R5 are selected from the group consisting of hydrogen, alkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof, and wherein the substituents are: for alkyl one or more groups selected from F, Cl, Br, I, =O, -OH, -NH2, secondary-alkyl amine, tertiary-alkyl amine, wherein alkyl is as defined above, S, -SH and phenyl, for cycloalkyl as defined for alkyl, for oxygen a group selected from alkyl, as defined above, and aryl, for nitrogen a group selected from acyl of from 2 to 18 carbon atoms, inclusive, monoalkyl and dialkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms of the acyl and alkyl groups, and for phenyl one or more groups selected from F, Cl, Br, I, -OH, -NH2, secondary-alkyl amine, tertiary-alkyl amine, wherein alkyl is as defined above, -SH and phenyl; and wherein R2, which can be singly substituted in any position of the pyridine ring not already substituted by R1, is:

-?-X

and X is the amino function of a compound selected from the group consisting of 7(R)-hydroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-hyrdroxy-methyl 1-thio-.alpha.-lincosaminide, 7(S)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(R)-halo-methyl 1-thio-.alpha.-lincosaminide, 7(S)-methoxy-methyl 1-thio-.alpha.-lincosaminide, 7-deoxy-7(S)-(methylthio)-methyl 1-thio-.alpha.-lineosaminide, 7-deoxy-7(S)-(2-hydroxyethylthio)-methyl 1-thio-.alpha.-lineosaminide and 7-deoxy-7(S)-(3-hydroxypropylthio)-methyl 1-thio-.alpha.-lincosaminide;
and the pharmaceutically acceptable acid-addition salts thereof;
when produced by the process defined in claim 1 or an obvious chemical equivalent thereof.

4. A compound according to claim 3, wherein R1 is in the 4 position and is alkyl of from 1 to 8 carbon atoms, inclusive, and isomeric forms thereof; and wherein R2 is in the 2 or 3 position; and the pharmaceutically acceptable acid-addition salts thereof; when produced by the process defined in claim 2 or an obvious chemical equivalent thereof.