IL28781A - 7-halo-7-deoxythiolincosaminides and process for preparing same - Google Patents

Description

7-HACO-7-PE0XYTHtOLIN008AMtmPE8 AND PROCESS FOR PREPARINO SAME 1980A-1 This invention relates to novel compounds and to processes for preparing them, and is particularly directed to 7-halo-T-deoxythiol incosaminides and to processes whereby they and like compounds are produced.

The novel compounds of the invention can be represented by the following structural formula: wherein R is alkyl of not, more than 20 carbon atoms, advantageously not more than 8 carbon atoms, cycloalkyl of from 5 to not more than 8 carbon atoms, and aralkyl of not more than 12 carbon atoms, advantageously not more than 8 carbon atoms.

Examples of alkyl of not more than 20 carbon atoms are methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, penta-decyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof. Examples of cycloalkyl are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-methyl cyclopentyl , 2,2-dimethy lcyclobutyl , methylcyclobutyl, and 3-cyclopentylpropyl . Examples of aralkyl are benzyl, phenethyl, a-phenyl propyl , and -naphthy lmethy 1.

The novel compounds of the Invention, Formula I, as well as other related compounds, can be prepared by replacing by chlorine, the 7-hydroxy of a compound of the formula: 198 OA- 1 The replacement is effected advantageously by mixing the start i ng compound of Formula II with a Rydon reagent and heating. In the process R can be any radical which Is non-reactive with triphenylphosphinedichloride but for preparing compounds of the invention, R is as given above.

The starting compounds of Formula II are prepared by hydrazi no lysis of a compound of the formula: wherein R is as given above and Ac is the acyl radical of a 4-substi tuted-L-2-pyrrol idi necarboxyl lc acid of the formula-: while HR»1 and HR2 arejalkyl wherein t a-nd-¾¾-6f-e al kyl idene/of not more than 20 carbon atoms (including methylene), advantageously of not more than 1980A-1 and aralkyl, respectively, carbon atoms, and a ral ky l I dene/of not more than 12 carbon atoms advantageously not more than 8 carbon atoms, and 3 is hydrogen or H 2. The hydraz i nol ysi s can be effected as described in U. Patent 3,179,565.

The compounds of Formula I are also useful as intermediates for producing 7-chloro-7-deoxyl I ncomyc? n and isomers and analogs thereof. Such compounds are prepared by acylating a compound of Formula I with a -substi tuted-L-2-pyr rol i di necar-boxylic acid of Formula A or B. This acylation can be effected by procedures already well known In the art of acylating amino sugars. The starting acids trans- -ethy 1 ; and 4-propyl -L-2-pyrrol tdlnecarboxyl i c acid can be obtained by hydrolysis of N-demethyl H neomycin B and li neomycin 0, respectively; and the N-methyl analogs from lincomycin B and lincomycin, respectively. These and analogous 4-subst i tuted-L-2-pyrrol i d i necarboxyl i c acids of Formulas A and B can be prepared by reacting a 4 -oxo compound of the formula: wherein Z is a protective hydrocarby loxycarbon 1 group which is removable by hydrogenol ysl s, trttyl (I.e., tr iphenylmethy 1 ), di phenyl (p-methoxyphenyl )methy 1 , bis- (p-methoxypheny 1 )phenyl -methyl, benzyl, or p-ni trobenzy I with a Wi ttig agent, e,g., an al kyl idenetri phenyl phosphorane [see e.g., Wittig et al., Ber., 3 , 138 (l954)j Trlppett, Quarterly Reviews, XVII, No. 4, p. o6 (1963)]. Examples of hydrocarbyl oxycarbonyl groups (Z) are tertiary-butoxycarbonyl j benzy loxycarbony) groups of the formula 198 OA -1 wherein X is hydrogen, nitro, methoxy, chloro, or bromo, for example, ca rbobenzoxy, p-ni trpcarbobenzoxy, p-bromo-, and p-chlorocarbobenzoxy ; and pheny+oxycarbony 1 groups of the formula wherein Xx is hydrogen, al lyl, or alkyl of not more than 4 carbon atoms such as phen^pxycarbony 1 , p- tol yloxycarbony 1 , p-ethy 1 pheny+pxycarbony 1 , and p-a 11 y 1 pheoyH oxyca rbony 1 and the like.

In carrying out this process the 4-oxo-L-2-pyr rol i di ne-carboxyl ic acid (Formula C) is added to a freshly prepared Wittig reagent. The Wittig reagents herein used can be general ly represented by the following formula: wherein Rx is as given above. These Wittig reagents are prepared by reacting an alkyl, cycloalkyl, or aral kyl triphenyl -phosphoni um ha 1 ide with a base such as sodamide, or sodium or potassium hydride, or the sodium or potassium metalate of di-methyl sul foxi de and the like. For example, the elimination of hydrogen halide from al kyl t riphenyl phosphoni um halide, produces al kyl idenetriphenylphosphorane. [The preparation of phosphor-anes is discussed in detail by Trippett, Quart. Rev. XVI I, No. 4, p. 406 (1965)]. The reaction is generally carried out in an organic solvent, such as benzene, toluene, ether, dimethyl-sulfoxide, tetrahydrofuran, or the like, at temperatures be- 1980A-1 The thus-obtained product, a -al kyl idene-, -cyc) oa 1 kyl i dene- , or -aral kyl idene-l-protected-L-prol ine which has the fol lowing formula: is recovered from the reaction mixture in a conventional manner, general ly by extraction from aqueous solutions of the reaction mixture. The crude product can be purified by conventional means, such as recrysta 11 i zat i on, chromatography, or formation and recrystall i zati on of easi ly formed derivatives such as amine salts of the amino acid, e.g., the d i cylohexy 1 ami ne salt, and the l ike, and liberating the amino acids from such compounds. By hydrogenat i ng an acid of Formula D in the presence of a catalyst, e.g., platinum, which is effective to saturate a double bond, but which is ineffective to effect hydrogenol ys i s, a compound of the fol lowing formula: is obtained. Platinum deposi ted on a carrier, e*g., carbon or an anion exchange resin l ike Dowex-1, a cross-l inked polystyrene trimethyl benzyl ammoni urn resin in the hydroxide cycle is suitable. If desired, the compounds of Formula I can be acylated with acids of Formula C, D, or E to form compounds IC, ID, and IE, respectively. Compound IC can then be converted to compound ID by treatment with a Wittig reagent and compound ID hydrogen- 1980A-1 genation, both of the acid D and the acylate ID, gives a mixture of ci s and t rans epimers which, if desired, can be separated by counter current distribution or chromatography. The starting acids of Formula B in which 3 is hydrogen are obtai ned when an acid of Formula D or E is subjected to hydrogenol ys i s over a pal ladium catalyst, e.g., pal ladium on carbon. Likewise, compounds of Formula ID and IE are converted to compounds of Formula IB in which R3 is hydrogen by the same process. The starting acids of Formula A are obtained by treating an acid of Form-ula D or Formula E with hydrogen bromide in acetic acid to remove the Z group. Compounds of Formula ID and IE are converted to compounds of Formula IB by the same process.

Some of the starting compounds of Formula I II are obtained biosynthet i ca 11 y. Lincomycin, methyl 6,8-di deoxy-6- (trans-1-methyl - -propyl -L-2-pyrrol id ί necarboxamido)-!- thio-D-erythro-a-D-galacto-octop ranos ide, is obtained as an elaboration product of a 1 i ncomyci n-produci ng actinomycete according to U. S. Patent 3,086,912. It has the following structural formula: wherein R and R3 are methyl and Rj.H is propyl. Lincomycin B methyl 6, 8- d i deoxy-6- (t rans- 1 -methyl - -ethyl-L-2-pyrrol idine-carboxamido)-!- thi o-D-erythro-g-D-galacto-octopyranos i de (Form ula IV wherein R and R3 are methyl and -RXH is ethyl) also is an elaboration product of the same microorganism when cultured 1980A-1 Lincomycin C, ethyl 6, 8- di deoxy-6- (trans-1 -methyl - - propyl -L-2-pyrrol i di neca rboxam? do) -1- thi o-D-ery thro- ct-D-ga lac to-octopyrano-side (Formula IV wherein R is ethyl, -RtH is propyl, and R3 is methyl) is obtained when the process of U. S. Patent 3*086,912 is carried out in the presence of added ethionine. Lincomycin D, methyl 6,8-d?deoxy-6- (trans-4-propyl -L-2-pyrrol idineca rboxam i do) -1- thi o-D-erythro-ct-D-gal acto-octopyranos ? de (Formula IV wherein R is methyl, -RXH is propyl, and R3 is hydrogen) is obtained when the fermentation of U. S. Patent 3,086,912 is carried out in the presence of added ct-MTL, methyl 6-amino-6,8-di deoxy-D-erythro-Ί- thi o-ct-D- gal acto-octopyranos I de, a compound of Formula III obtained by the hydrazi nol ysi s of lincomycin.

Methyl 6,8-dideoxy-6- (trans-4-ethyl -L-2-pyrrol idi neca rboxam i do )- 1- thi o-D-er thro- g-D-ga lac to-octopyranos ide (Formula IV wherein R is methyl, -RtH is ethyl and R3 is hydrogen) is also produced when ct-MTL is added to the fermentation of U. S. Patent 3,086,912. Similarly, lincomycin K, ethyl 6,8-d i deoxy-6- (trans -4- propyl -L- 2- pyrrol 1 di neca rboxami do) -1-th i o-D-ery thro-ct-D- gal acto-octopy-ranoside (Formula IV wherein R is ethyl, -RxH is propyl, and R3 is hydrogen) is produced when the fermentation of U. S. Patent 3,086,912 is carried out in the presence of added a-ETL, ethyl 6-ami no-6,8-d ?deoxy-D-erythro-ct- thi o-D-ga 1 acto-octopyranos i de, a compound of Formula III obtained by the hydrazinolysis of l incomycin C. Ethyl 6,8-d ? deoxy-6- (trans-4-ethy 1 -L-2-pyrrol i -d i neca rboxam i do) -1 - th i o-D-erythro- ct-D-gal acto-octopyranos i de (Formula IV wherein R is ethyl, -Rj.H is ethyl, and R3 is hydrogen) is also obtained when a-ETL is added to the fermentation of U. S. Patent 3,086,912.

Other starting compounds of Formula I I I where Ac and R are as given above and having both the D-erythro and L- threo configurations are described in Belgium Patent 667,948, February 198 OA- 1 Li neomycin or any of the starting compounds of Formula III which has the D-erythro configuration can be converted to the L-threo configuration by converting the 7-hydroxy group to a 7-oxo group and then back again to a 7-hydroxy group. A suitable procedure for this purpose is illustrated in the following sequen For example, li neomycin on treatment with acetone in the presence of p- tol uenesul foni c acid is converted to -0- i sopropyl i dene-lincomycin which on oxidation wi th chromic oxide gives 7-oxo-3* -0- i sopropyl idenel i ncomyci n (methyl 6,8-dideoxy-3> -0- i sopropyl idene-6- (trans -1-methyl -4- propyl -L-2-pyrrol idi necarboxamίdo)-l- thio-D-gl ycero«a-D-ga lacto-octanopyranos-7-ulos ? de which on treatment with sodium borohyd ride is converted to 7-epi 1 i ncomyci n (methyl 6,8-di deoxy-6- (trans -1-methyl -4-propyl -L-2-pyrrol idi ne-ca rboxam ? do ) - 1 - thio-L- threo- -D-ga lacto-octopyranos ide) . Any of the starting compounds of Formula II having a D-erythro configur 198 OA -1 by this procedure, As the biosynthetic-produced 1 i neomycins, as well as the amino sugars deri ed therefrom (starting compounds of Formula II-), are either methyl or ethyl t hi ogl ycos i des, it is sometimes desirable to convert them to higher or lower glycosides. It is thus sometimes desirable to convert any of the compounds of Formulas II or HI to higher or lower glycosides. This can effectively be accomplished by reacting the compound to be converted wi th a me rcaptan of the Formula Ra$H where e is an alkyl group of not more than 20 carbon atoms, but one different from R n with a ormula: wherein Ac is hydrogen or acyl, which on treatment with acid and/or on heating is recyclized to give a compound of the foi l di rect 1 y to any of the starting products of Formula III. The resulting products can be subject 1980A-1 for each mole of starting compound, though more or less can be used. Advantageously a slight excess, say from 5 to 20$ excess, of bromine is used. The bromine initially replaces the RS- group and the resulting intermediate hydrolyzes to the sugar in which the pyranose from XII la is in equilibrium wi th the aldose from XII lb. In the presence of acid, e.g., hydrochloric acid or other strong non-oxidizing acid such as p-toluene sulfonic acid, and sulfonic acid type anion exchange resins, the mercaptan ReSH reacts with the sugar XIII to form the thiogl ycos ide XIV. Con-comitantly some diacetal of Formula IX may be formed which after separation can be cyclized as described above to form more of the desired thioglycoside XIV.

The mechanism by which triphenylphosphinedihal ide effects the substitution of the 7- hydroxy by halogen Is not fully under-stood. The mechanism is such, however, that a change In configuration results. Thus, a 7-hydroxy compound of the D-erythro configuration yields a 7-halo compound of the L-threo onfigure t ion.

Triphenylphosphi nedi chloride is formed by the addition of chlorine to tri phenyl phosphi ne. Rydon et al., J. Chem. Soc, 2224 (1955); Ibid, 228l (195*0; Ibid, 303 (1956). The triphenylphosphi nedi chlor? de can be formed in situ by addition of chlorine to a solution of the triphenylphosphi rte in an inert solvent such as acetoni tri le or dimethyl ormam Ide, or it can be isolated as a separate entity. In either case the reaction with the starting thiol incosaminide is effected by contacting the triphenylphosphinedichloride therewith in an inert solvent, e.g., acetoni tri le or dimethyl formam ide, until the desired substitution of the 7-hydroxy Is obtained. The reaction takes place at ordinary temperature, though gentle heating can be effected if desired. Advantageously the temperature is mai tained between 1980A-1 the reaction mixture by well known techniques such as filtration, solvent extraction, etc. The reaction mixture advantageously is treated with methanol to destroy any excess triphenylphosphi ne-dichloride, filtered to remove any solid such as tri phenyl phos-phine oxide, formed in the reaction, and then treated to recover the product. The methanol can be added either before or after the filtration. Advantageously the treated and filtered reaction mixture is evaporated to dryness and purified by solvent extraction and/or chromatography.

The compounds of Formula I exist either in the protonated or non-protonated forms according to the pH of the environment. When the protonated form is intended the compound is qualified as the acid addition salt; when the non-protonated form is intended it is qualified as the free base. The free bases can be converted to stable acid-addition salts by neutralizing the free base with the appropriate acid to below about pH 7.0, and advanr tageously to about pH 2 to pH 6. Suitable acids for this purpose include hydrochloric, sulfuric, phosphoric, thiocyanic, fluosilicic, hexaf 1 uoroa rsen i c, hexaf 1 uorophosphor i c, acetic, succinic, citric, lactic, maleic, fumaric, pamoic, cholic, pal -mi. tic, mucic, camphoric, glutaric, glycol ic, phthal ic, tartaric, lauric, stearic, salicylic, 3-phenyl sal i cyl i c, 5-phenyl sal i cy 1 i c, 3-methyl gl utari c, orthosul f obenzoi c, cycl ohexanesul fam i c, cyclo-pentanepropioni c, 1, 2-cyclohexaned icarboxyl i c, -cyclohexane-carboxyl ic, octadecenyl succi ni c, octeny 1 succi ni c, methanesul-fonic, benzenesul foni c, helianthic, Rei necke1 s, dimethyldithio-carbamic, cyclohexyl sul fami c, hexadecyl sul fami c, octadecyl sul -famic, sorbic, monochloroacet i c, undecylenic, 4' -hydroxyazoben-zene- -sul foni c, octyldecyl sul furi c, picric, benzoic, cinnamic, and 1 i ke aci ds .

The acid-addition salts can be used for the same purposes 1980A-1 For example, the free base can be converted to an insoluble salt, such as the picrate, which can be subjected to purification procedures, for example, solvent extractions and washings, chromatography, fractional 1 i qu i d- 1 i qu i d extractions, and crys-tal l ization and then used to regenerate the free base form by treatment with alkal i or to make a different salt by metathesis. Or the free base can be converted to a water-soluble salt, such as the hydrochloride or sulfate and the aqueous solution of the salt extracted with various water- immi sci b 1 e solvents before regenerating the free base form by treatment of the thus-extracted acid solution or converted to another salt by metathesis.

The free bases of Formula I can be used as a buffer or as an antacid. The compounds of Formula I react with isocyanates to form urethanes and can be used to modify polyurethane resins. The thiocyanic acid addition salt when condensed with formaldehyde forms resinous materials useful as p i ckl i ng i nhi bi tors according to U. S. Patents 2,425,320 and 2,6o6,155. The free bases also make good vehicles for toxic acids. For example, the fluosil icic acid addition salts are useful as mothproofing agents according to U. S. Patents 1,915, 33^ and 2,075,359 and the hexaf 1 uo roar sen i c acid and hexaf 1 uorophosphor i c acid addition salts are useful as parasiticides according to U. S. Patents 3,122,536 and 3,122,552.

The following examples are illustrative of the process and products of the present invention but are not to be construed as limiting. The parts and percentages are by weight and the solvent ratios are by volume unless otherwise specified.

EXAMPLE 1 7-Chloro-7-deoxy-g-methyl thiol i ncosami ni de 198 OA -1 To a suspension of 197.2 g. of tri henyl phosphi ne in 1.5 1. of anhydrous acetoni t r i 1 e was added 52.5 g. of chlorine. With sti rring, 18.75 g. of methyl a- thi ol i ncosami n ide (U. S. Patent 5*179*565) was added. After 2.5 hrs. at ambient temperature, 50 ml. of methanol was added. The mixture was concentrated to a thick syrup. The concentrate was diluted with methylene chloride and extracted three times with water. The aqueous extracts were washed twice with methylene' chloride. The extracts were made alkal ine with sodium hydroxide and extracted repeatedly with methylene chloride. The organic extract was dried and evaporated under vacuum. The residue was chromato-graphed over 1.1 kg. of sil ica gel using chloroform-methanol ( :l) for elution. The major fraction selected on the basis of its TLC (thin layer chromatography) profile weighed 4.4 g, Recrysta 11 ization from methanol -water afforded 2.73 g. of 1, o m.p. 178-181 .

Anal . Calcd. for C9H18C1N04S: C, 39-77; H, 6.67; N, 5.I6; S, 11.80; 0, 13.05.

Found: C, 39.91; H, 7.02; N, 5-57; S, 11.99; CI, 13-33. EXAMPLE 2 198 OA -1 d (U. ), 1.23 ml. of triethyl amine and 600 mg. of i sobuty 1 chl oroformate o in 80 ml . of ace ton i tri le was sti rred at -5 for 15 minutes. A solution of 1.09 g. of 7-chloro-7-deoxy-a-methyl hiol i ncosami n-ide in 20 ml. of water and 20 ml, of acetone was added. The mixture was stirred at ambient temperature for 2 hrs. The mixture was evaporated and the residue extracted with methylene chloride. Evaporation of the solvent yielded 6l3 mg. of oil. This oil was chromatographed over 100 g. of silica gel using chl oroforrri methanol (7:l) for elution. The major fraction, 380 mg., was recognized as 7-chl oro-7-deoxy 1 i ncomyci n by TLC. It was dissolved in acetone, acidified with dilute Hcl and evaporated. The residue was crystallized from absolute ethanol to o give 180 mg. of 7-ehloro-7-deoxyl i ncomyc i n, m.p. 158-I0O , identical by infrared data with a known sample of 7-chloro-7-deoxy-l I neomycin.

EXAMPLE 3 Part Α-3· Methyl N- (l-carbobenzoxy- - t rans and cis-propyl -L-prol yl -7-chl oro-7-deoxy-c.- thi ol i ncosami nide) 1980A-1 Ci s and t rans-l-carbobenzoxy-4-p ropy 1 - L-prol j ne (2.35 g.) was dissolved in 150 ml. of acetonitrile containing 1.12 ml. of tr iethylami ne. The solution was cooled to 0 and .18 ml. of o i sobutyl chloroformate added. After 10 minutes at 0 , a solution of 2.17 9· of methyl -7-chloro-7-deoxy- -thioU ncosami ni de in 40 ml. of acetonitrile and 40 ml. of water was added. The mixture was stirred for 2 hours at ambient temperature and the solvent distilled i n vacuo to yield a crystalline residue.

The crystals were collected by filtration, washed, and dried to yield 3·36 g. of methyl (carbobenzoxy- - trans and ci s-propyl -L-prol yl )-7-chloro-7-deoxy-a-thiol i ncosami nide), m.p„ o 18O-I85 . A portion was recrysta 11 i zed several times from o ethanol . It melted at 189-192 , Anal . Calcd. for C25H37CIN2O7S: C, 55.08; H, 6.84; CI, 6.51; N, 5.14.

Found: C, 5^.80; H, 7-15; CI, 6.59; N, 5.l6.

Part B-3. Methyl N-( -trans and ci s-propyl -L-prol yl ) - 7-chloro-7-deoxy-q7yth?Ol incosaminide hydrochloride A portion/of the crude product from Part A-3 was d solved In 50 ml. of methanol and 0,5 g. of 10# palladium charcoal added. The mixture was shaken under 3 lbs. of hydrogen pressure for hrs. TLC showed partial hydrogenolysls. An additional 0.5 g. of catalyst was added and hydrogenat I on continued for l8 hrs. The catalyst was removed by filtration.

The residue was chroma tog raphed over silica gel, and the more polar fraction collected. It weighed 185 mg. It was converted to the hydrochloride in the usual manner affording 150 mg. of o crystals, m.p. 220-222 C, assaying about four times as active as lincomycin.

Another portion of the crude product (22.9 g.) was djs-solved In 500 ml. of methanol and 6.0 g. of 10$ palladium on charcoal added. The mixture was shaken under 35 lbs. of hydrogen pressure for 18 hrs. The catalyst was removed by filtration and the clear liquid phase was evaporated. The solid crude product was converted to the hydrochloride. Crystallization of the crude product from acetone-water afforded 15.08 g. of cry- o stals, m.p. 218-223 dec. ecrystal 1 izat ion from water yielded • ° ° an analytical sample, m.p. 228-23* dec, [ ]D + 159 · Calcd. for CtTl SCla: C, 45.63 H, 7.21; N, 6.26.

Found: (Corrected for 3·91# HsO) C, 45.85; H, 7.51; N, 5.86.

The fact that this compound has equal or greater activity K (N-demethy 11 i ncomyci η) is much less active than li neomycin. Furthermore, i t has unexpected Gram-negati e activity. Also, the isolated trans- i somer is about 8 times as active as I i neomycin against Gram-positive bacteria and 8 or more times as active against Gram-negative bacteria.

Part C . l-Carbobenzoxy-4-propyl ? dene-L-prol ine and di cyclohexy lami ne salt.

Sodium hydride (3.8 g. ) was warmed with 75 ml. of di- o methylsul foxide at a temperature of 70-75 C. until the reaction o was complete. After cooling to 20 C, 30.8 g. of propyl tri-phenyl phosphoni urn bromide was added, and the resulting red solution was stirred for 30 minutes to insure complete reaction. A solution of 5.2 g. of 4-keto-l-ca rbobenzoxy-L-prol i ne in 15 ml. of dimethyl sulfoxide was added over a period of 15 minutes, ' o and the resulting mixture was stirred for 20 minutes at 2o C. o and then at 70 C. for 4 hours. The reaction mixture was cooled, 100 ml. of 5# aqueous potassium bicarbonate and 100 ml. of water added, and filtered. The filtrate was washed twice with 150 ml. portions of ether and the ether was discarded after back-extract-i ng with bicarbonate. The bicarbonate solutions were combined, diluted with 200 ml. of water, and aci di f ied wi th 4N hydrochloric acid. The acidified mixture was extracted with three 200-ml . portions of ether. The combined ether extracts were washed with three 50-ml. portions of saturated aqueous sodium bisulfite, then with water and dried over anhydrous sodium sulfate. Evaporation of the solvent gave 5.7 g. of a sol id residue which was 1-carbobenzoxy - -propyl i dene-L-prol i ne.

This residue was dissolved in 18 ml. of acetonitrile and treated with 2.8 ml. of di cyclohexylami ne. The crystal l ine o di cyclohexylami ne salt, 5.2 g. (55# yield), melted at 154-157 C. After three recrystal 1 i zati ons from acetonitrile, an analy- rotation of [a]Q -8 (c = Ο.3898 g. /100 ml., CHC13).

Anal. Calcd. for C28H42N204: C, 71Λ5; H, 9.00; N, 5.95.

Found: C, 71.77; H, 9-39; N, 5.1.

Part D-3. Cis- and trans-l-carbobenzoxy -propyl -L-prol i ne.

Ten grams of the amine salt of Part C-3 was shaken with ether and 2 potassium hydroxide. The aqueous layer was separated and acidified. Extraction with methylene chloride led to the i sol at ion 6 g. of oily acid. A mi ture of 2 g. of thi s oil and 800 mg. of 7# platinum on Dowex-1 catalyst in 50 ml . of methanol was shaken under 40 lbs. of hydrogen pressure for 17 hr. The catalyst was removed by filtration and the solvent d i st i 1 led ]_n vacuo, leavi ng a res i due of 2 g. of oil. Thin-layer chromatography, using a methanol-5# ammonium hydroxide system and permanganate -per iodate indicator spray indicated that the double bond was hydrogenated . Ninhydrin gave a negative test. This product resisted crystal l ization and was used without purification in Part A-3.

EXAMPL Tr iphenylphosphi ne (39·3 g.) was added to 300 ml. of acetoni t ri 1 e and 0 ml . of solvent distil led. The solution o was cooled to 20 and 8.1 ml. of bromine in 10 ml. of aceto- o nitrile added dropwise, keeping the temperature below 0 by 1980A-1 crystallized from the reaction mixture. Methyl a-thiol i ncosam- inide, 10 g., was added and the reaction mixture stirred at ambient temperature for l8 hrs. To the deeply colored solution, 20 ml. of methanol was added. After stirring for 10 min., the solvent was distilled i n vacuo. The residue was partitioned between 50 ml. of water and 100 ml. of chloroform. The chloroform was extracted three times with dilute acid. The combined aqueous extract was made alkaline with potassium hydroxide and o stirred at 10 for 1 hr. Filtration afforded a dark residue which was dissolved in 70 ml. of hot ethanol . Crystals were deposited from this solution on cooling. Recrystal 1 i zat ion from ml. of ethanol gave 150 mg. of methyl 7-bromo-7-deoxy-a- o o thiol incosaminide, m.p. l6j-l66 (sinter l4l ).

Anal . Calcd. for C8HieBrN04S: Br, 25.17.

Found: Br, 24,58.

EXAMPLE 5 Part A-5. 7-Bromo-7-deoxy-N-demethyl -N-carbobenzoxy- 11 neomycin (trans) and 7-bromo-7-deoxy-N-demethyl -N-carboben- zoxyal lol i ncomyci n (cis).

To a solution of 1 mg. of l-c rbobenzoxy-4- trans and ci s-propyl -L-prol i ne (Part D-3) in 15 ml. of acetonltrile was o added 0.64 g. of triethylami ne. The solution was cooled to 7 and 0.079 ml. of i sobutyl chloroformate was added. After stir 1980A-1 deoxy-α- thiol i cosami ni de i n 8 ml . of acetone and 2 ml . of water was added. The reaction mixture was stirred for 2 hrs.

The solvent was distilled in vacuo and the residue crystallized from hot acetone to give 150 mg. of a mixture of 7-bromo~7-deoxy- N-demethyl -N-carbobenzoxy 1 i ncomyci n, and 7-bromo-7-deoxy-N-de- o methyl -N-carbobenzoxyal lol i ncomyci n, m.p. l44-l46 C. dec. An o additional quantity, m.p. 139-1^5 > was obtained by concentrating the mother liquor.

The material possessed a satisfactory infrared curve showing amide absorption at l675 cm."1 and 1550 cm."1. It was not further purified but used directly in the next step.

Part B-5. 7-Bromo-7-deoxy-N-demethyl 1 i ncomyci n hydrochloride and 7-bromo-7-deoxy-N-demethylal lol i ncomyci n hydrochlor- A mixture of 120 mg. of the 144-146 C. material of Part A-5 and 60 mg. of 10$ palladium on charcoal in 8 ml. of methanol was shaken under 40 lbs. of hydrogen pressure for 5 hrs. The catalyst was removed by filtration and the residue evaporated to dryness. The residue was dissolved in a few ml. of acetone and acidified with dilute HC1. Crystalline 7-bromo-7-deoxy-N-demethy 1 1 i ncomyci n and 7-bromo-7-deoxy-N-demethylal lol i neomycin hydrochlorides formed on refrigeration of the solution. Standard curve assay vs. S. lutea showed 0.5 x the activity of li neomycin. 1980A-1 By substituting the methyl a- thiol i ncosami ni de by other alkyl or by cycloalkyl or aralkyl a-thiol incosam.i nides wherein alkyl is ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl , dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, and eicosyl and the isomeric forms thereof; cycloalkyl is cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-methylcyclo-pentyl, 2, 3-dimethyl cyclobutyl , 2-methylcyclobutyl , and 3-cyclo-pentyl propyl ; and aralkyl is benzyl, phenethyl, 3-phenyl propyl , and ct-naphthylmethyl , the corresponding alkyl, cycloalkyl, and aral kyl 7-halo-7-deoxy-a-thiol i ncosami nides; al kyl , cycloal kyl , and aral kyl 7-halo-6,7>8- tri deoxy-6- (l-c rbobenzoxy- -propyl -L-2 -pyrrol idi necarboxami do) -1-th io-L- threo-a-D-gal acto-octopyrano-s ides; and al kyl , cycloalkyl, and aralkyl 7-halo-6,7>8-trideoxy-6- (4- propyl -L - -pyrrol idi necar boxami do) -1-thio-L- threo-a-D-gal ac to- octopyranos ides are obtained. For example, by substituting the methyl a-thiol i ncosami ni des by ethyl, propyl, butyl, pentyl, and hexyl a-thiol i ncosami n i des, there are obtained ethyl 7-hal o-6,7,8-trideoxy-6- (l-carbobenzoxy-4-propyl -L-2-pyrrol idi ne ca rboxam i do) - 1 - 1 h i o- L - th reo- a-D - ga 1 acto- oc topy ranos ? de, propyl 7-halo-6,7>8-trideoxy-6- (l-carbobenzoxy-4-propyl -L-2-pyrrol idi ne-ca rboxam t do) -1- th io-L- threo-a-D-gal acto- octopyranos i de, butyl 7-hal o-6,7» 8- tr i deoxy-6- (l-carbobenzoxy - -propyl -L-2-pyrrol idi necarboxami do) -1-th io-L- dTre^-a-D-^Tacto-octopy ranos i de, pentyl 7-halo-6,7*8-trideoxy-6- (l-carbobenzoxy-4-propyl -L-2-pyrrol idine-ca rboxam ido)-l-thio-L -threo- a- D -gal acto- oc top y ra nos i de , hexy 1 7- ha lo-6,7*8-tr i deoxy-6- (l-carbobenzoxy-4-propy 1 -L-2-pyrrol i di necarboxamido)-!- 1 hi o-L- threo-a-D-gal acto-octopy ranos ide, ethyl 7-halo-6,7*8-trideoxy-6- (4-propyl -L-2-pyrrol idi necarboxami do) -1-t hi o-L- threo-a-D-gal acto-octopy ranos ide, propyl 7-halo-6,7>8-trideoxy-6- (4-propyl -L-2-pyrrol i d i necarboxamido)-!- thi o-L- threo- 1980A-1 Anal . Galcd. for CgiH_uCl gO@$i C, 51,99} H, 8,52; N, 5,78; §, 6.6l.

Found: C, 50.64; H, 8.32; N, 5.68; S, 6.41. [a]35 + 150° (c = 0.4070 In f½0)< D By substituting the 2«propanethlo1 by cycl ohexahethiol , the S-cyclohexyl analog of 1 i ncomyc! n Is obtained.

An l . Calcd. for Csah gCI l½06S: C, 54.04; H, 8.48; , 5.48; S, 6.27; CI, 6. 4.

Foundi C, 53.57; H, 8,74; N, 5,37; S, 6.25; CI, 6.66. ss o t«] Q + 123 (c = 0.9952 In H20) By substituting the 2-propanethlol by other al kyl mercaptans, for example, ethyl, propyl, pentyl , hexyl , heptyl , octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl , hexadecyl, heptadecyl , octadecyl , nonadecyl, and elcosyl mercaptans and the isomeric forms thereof and l ike alky) mercaptans containing not more than 20 carbon atomsj by other cycloalkyl mercaptans, for example, cyclopropyl , cyclobutyl , cyclopentyl, cycloheptyl, cyclooctyl, 2-methyl cyclopentyl , 2,3-dlmethylcyclo-butyl, 2-methy lcyclobutyl , and 3- cyclopentyl propyl mercaptans and like cycloalkyl mercaptans conta i nl ng from to not more than 8 carbon atoms; or by aralkyl mercaptans, for example, benzyl, phenethyl, 3-phenyl ropyl , and 1-naphth Imethyl mercaptans and l ike aralkyl mercaptans contal ni ng not more than 12 carbon atoms, the correspondi g alkyl, cycloalkyl, and aralkyl a-thiol i ncosaml nides are obtained* By substituting the 11 ncomyci n by epl - 11 ncomyci ri, the epi* thiol 1 ncosaml nl des are obtained. The epi -1 I ncomyci n Is prepared as fol lows: A solution of 9-8 g. of li neomycin In 150 ml, of acetone is added to a solution of 9«8 g. of p-tol uenesul foni c acid mono-hydrate in 10 ml. of acetone with good stirring and avoidance 198 OA- 1 temperature for 1 hour, after which 100 ml, of anhydrous ether is added and stirring is continued in an ice-bath for 0.5 hour, o The mixture is fil tered and the sol i d is d r i ed j_n vacuo at 50 C. ; yield, 13*55 g> (85.5$) of 3, 4-0- i sopropyl i denel i ncomyc i n £- tol uenesul fonate. An additional 1.15 g. (7 ' Λ$) can be recovered from the mother liquors by adding 350 ml . of anhydrous ether to the mother liquor from the previous filtering operation and chill ing the solution for 1 hour. The l4.5 g. so obtained are suspended in 200 ml. of ether and shaken vigorously with 125 ml . of $ potassium bicarbonate solution. The aqueous layer is back-extracted with two 100-ml . portions of ether. The ether extracts are washed with 50 ml, of saturated sodium chloride solution and then filtered through anhydrous sodium sulfate.

The ether is evaporated under vacuum, leaving 7.9 g. (73· 1$) of 3>^-0-i sopropyl idenel i neomycin which is dissolved in 25 ml , of ether acetate and concentrated to about 10 to 15 ml. The concentrate is allowed to stand at room temperature for several hours and then refrigerated overnight. The crystals are filtered from the solution and washed sparingly with cold ethyl acetate; yield, 4.55 g. (42*2$) of 3*^-0-i sopropyl idenel i neomycin having o a melting point of 126-128 C, and an optical rotation of o [a] D 101-102 (c, 1, methylene chloride).

To a solution of 6 g. (0.0135 moles) of i sopropyl i dene-lincomycin in 75 nil . of pyridine was added 12 g. (excess) chromic o oxide. The solution warms up about 20 C. After one hour the mixture was added to a solution containing 250 ml . each of ethyl ether and ethyl acetate. This was then filtered and evaporated to a syrup, 8.4 g. This syrup was distributed in a 500-trans-fer countercur rent distribution using the system, watenethyl acetate: ethanol : cyclohexane (l:l:l:l). i soprop l i denel i ncom ci n was isolated as the eak fraction from 198 OA- 1 Ana 1. Calcd. for 021Η3βΝ20β$: C, 56.72; H, 8.16; N, 6.30; S, 7.21.

Found: C, 56.37; H, 7.62; N, 6.51; S, 6.84.

To 1.6 g. of Craig-pure 7-dehydro-3* -0- i sopropyl i dene-II neomycin in 75 ml. of methanol was added 400 mg. of sodium borohydride. After 1.5 hr. this solution was evaporated to dryness on a rotary evaporator. The residue was added to 25 ml. of water and extracted three times with 25 ml. each of methylene chloride. The extract was back-washed with 15 ml, of water, then dried over magnesium chloride and evaporated to dryness.

The residue, 1.4 g., was distributed in a 500-transfer counter-current distribution using the solvent system, water:ethyl acetate: ethanol : cyclohexane (1:1:1:1), and a single peak which fit the theoretical was observed at K = 1.05. The material in tubes 240 to 280 was isolated as a syrup.

Anal . Calcd. for C21H38N2O6S: C, 56.47; H, 8.58; N, 6.27; S, 7.I8.

Found: C, 56.24; H, 8.54; N, 6.13; S, 7.01.

Thin-layer chromatography (TLC) showed that this material consisted of two substances. One was 3*4-0- ί sopropyl idenel i neomycin; the other 3*4-0- i sopropyl i dene-epi 1 i ncomyci n, which moved s 1 i ght 1 y s lower .

The last mentioned syrup was stored at room temperature hrs. in a sol ut ion contai ni ng 60 ml. of 0.25 IN hydrochloric o acid and 40 ml. of ethanol. It was then kept at 0 C. for 4 days. Following neutral ization wi th sodium bicarbonate, it was evaporated to 25 ml., then extracted with chloroform. The extract was washed with a little water and dried over magnesium sulfate, then evaporated to a residue. Thin layer chromatography of the residue showed two substances, both of which were active against 1 utea. The residue was chromatographed on a l4" x 3/4" 198 OA -1 Patent 2,393*625) column which was eluted gradiently with solvent which varied continuously from 100$ Skellysolve B (technical hex-ane) to 100$ acetone. The total volume was 5000 ml.

The two compounds were thus separated: Fraction I: Tubes -6 (40 ml . cuts) Ep i 1 ΐ ncomyc i n.

Assay 50 meg. /ml .

Anal . Calcd. for CieH3 206S: C, 50.92; H, 8.55; N, 6.60; S, 7.56.

Found: C, 50.19; H, 7-91; N, 6.05; S, 6.42.

Fraction I I: Tubes 73-104. Lincomycin.

Assay 950 mcg./mg.

Claims (1)

1. 98OA-1 HAVING NOW particularly described and ascertained the nature of our said invention and in what manner the same is to be performed, we declare that what v/e claim is : -1- A thiol i ncosami nide of the formula: wherein R is alkyl of not more than 20 carbon atoms, cycloalkyl of from 5 to not more than 8 carbon atoms, or aralkyl of not more than 8 carbon atoms and halo is chlorine or bromine. -2- A thi ol i ncosami ni de of claim 1 in which halo is chlorine and R is alkyl of not more than 20 carbon atoms. -3- A thiol i ncosami nide of claim 1 in which halo is bromine and R is alkyl of not more than 20 carbon atoms. -4- A thiol i ncosami ni de of claim 2 in which R is methyl. -5- A thiol i ncosami ni de of claim 3 in which R is methyl. -6- A thiol i ncosami nide of claim 1 having the formula: wherein R is alkyl of not more than 20 carbon atoms and halo is chlorine or bromine. 198 OA- 1 A thi ol i ncosami ni de of claim 6 in which halo is chlorine. -8- A thi ol i ncosam i ni de of claim 7 in which R is methyl. -9- A thiol ? ncosaminide of claim 6, in which halo is bromine. -10- A thiol i ncosaminide of claim 9 in which R is methyl. -11- A thiol i ncosami nide of claim 1 having the formula: wherein R is alkyl of not more than 20 carbon atoms. -12- A thi ol i ncosami ni de of claim 11 in which halo is chlor- 1 ne. -13- A thiol i ncosami ni de of claim 12 in which R is methyl. -14- A thiol i ncosami ni de of claim 11 in which halo is bromine. -15- A thiol i ncosami ni de of claim 14 in which R is methyl. -16- A process for making a th i ol i n cosami n i de of the formula: 1980A-1 „.* wherein R is alkyl of not more than 20 carbon atoms, cycloalkyl of from 3 to not more than 8 carbon atoms, or aralkyl of not more than 8 carbon atoms and halo is chlorine or bromine which comprises reacting in an inert solvent tri pheny 1 phosph i ned i ch 1 or i de or tri pheny 1 phosph i nedi b romi de -wi th a compound of the formula: wherein R is as given above. -17- The process of claim 16 in which the reaction is conducted temperature between about 20° C. and about 55° C. -18- The process of claim 16 or 17 in which the reaction product is treated wi th methanol and filtered to remove any solid, such as tr i phen yl phosph ine oxide, formed in the reaction. DATED THIS 28th