AU705004B2 - 2,3-dihydro-benzofuranol derivatives and process for resolving the same - Google Patents

Description

AUSTRALIA

Patents Act COMPLETE

SPECIFICATION

(ORIGINAL)

Class Int. Class Application Number: Lodged: Complete Specification Lodged: Accepted: Published: Priority Related Art: Name of Applicant: Hoechst Marion Roussel, Inc.

I Actual Inventor(s): Gilbert Marciniak Richard A. Schnettler Timothy A. Ayers Damian J. Krysan Address for Service: t "PHILLIPS ORMONDE

FITZPATRICK

Patent and Trade Mark Attorneys 367 Collins Street Melbourne 3000 AUSTRALIA Invention Title: 2 3 -DIHYDRO-BENZOFURANOL DERIVATIVES AND PROCESS

FOR

RESOLVING THE SAME Ovr Ref 551904 POF Code: 1432/1432 The following statement is a full description of this invention, including the best method of performing it known to applicant(s): -1a 2,3-D'HYDRO BENZOFURANOL DERIVATIVES AND PROCESS FOR RESOLVING THE SAME BACKGROUND OF THE INVENTION This application is a divisional application from parent application 49209/96 the entire content of which is incorporated herein by reference.

This invention relates to novel compounds produced during the synthesis of 2,3-dihydro-ben;ofuranol derivatives and a process for optically resolving S. 10 many of these compounds. The optical resolution of these compounds is I "important as the intermediate resolved is a crucial intermediate in the synthesis of the compounds in the parent application. The production of optically pure intermediates therefore facilitates the production of optically pure compounds of the parent application.

The parent application of this application relates to a novel process for preparing 2,3-dihydro-benzofuranol derivatives.

The 2,3-dihydro-benzofuranol derivatives marifest the property of being free radical scavengers. Disease cond;:ions capable of being ameliorated by A free radical scavengers are, for example, stroke, nervous system trauma or 20 reperfusion damage as more fully described in Patent Application WO 93/20057, filed March 10, 1993 and U.S. counterpart U.S. Serial No.

08/318,633, U.S. filing dats December 22, 1994, which is incorporated herein by reference.

More specifically the parent application of this application relates to a novel process for preparing 2,3-dihydro-benzofuranol derivatives of the formula

(I)

i 1D 2 c \wofrNNYLYspECNsGL£y,^3aDC^ 2including the stereoisomers, enantiomers, optically active and racemic mixtures thereof, or their pharmaceutically acceptable salts thereof, wherein

R

2 is C 1 4 alkyl each R 2 moiety being independently 1 4 aky r oh 2 moieties, when taken together with the ca'rbon atom to which they are attached, form a C 5 -6 cyclic hydrocarbyl moiety;

R

4 iS C 1 6 alkyl;

R

5 is H or C(O)R with R being H or C 1 g alkyl;

R

6 is C 1 6 alkyl;

R

7 is H or c.

6 alkyl; X is COORB, CH 2 0H, halomethyl, C(O)A or CH 2

A;

A is NR 7 R9, -N9R 6

R

6

R

6 -Qe, pyrroiidino, piperidino, morpholino, rN-"N Rl

YC

R

8 is H, C1..

6 alkyl, or -(CH 2 with m being 2,3 or 4; R9g is H, C 1 4 alkyl,--CH 2

(R

11 n is 1, 2, 3 or 4, p is 1, 2, or 3; Rio is H, C 8 alkyl, C 6 alkenyl1, C 4 6 cycloalkyl, cyclohe-' ylmethyl, hydroxyalkyl (C 2 6 :dihydr(Dxyalkyl (C 3 6

C

2 9 acyloxyalkyl (C 2 6

C

1 4 alkoxyalkyl (Cl 1 6 t being 0, 1 or 2, or pyrimidinyl, with the proviso that when Y is other than H then Rio is H; Y is H, CH 3 or COOR 7

R

11 is H, C 1 4 alkoxy, C 1 4 alkyl or halogeno;

R

12 is ortho C 1 4 alkoxy, ortho C 1 4 alkyl or p-halo; and Q is a halide, or sulfonate ion e.9S0 3 RI with R 1 being H,

C

1 6 alkyl, aryl or aralkyl.

iN '4 6 6

:I

a i

I

-3- SUMMARY OF THE INVENTION In one aspect the present invention provides a process for resolving into its optically active isomers compound of formula (9) F"4 PgO F" R"R"2 0

R"

7 wherein

R"

2

R"

4

R"

6 and R"7 are methyl, Pg is hydrogen or is a suitable protecting group, comprising the steps of: reacting compound of formula with lipase! candida cylindracea and vinyl acetate, separating the individual isomers, optionally deprotecting the 5-hydroxy group In a further aspect the present invention provides the compound hydroxy-3-methylene-2,2,4,6,7-pentamethyl-2,3-dihydro-benzofuran.

In an even further aspect the present invention provides the compound 5-benzyloxy-3-hydroxymethyl- 2 2 4 ,6,7-pentamethyl-2,3-dihydro-1 -benzofuran.

In yet an even further aspect the present invention provides the compound 3-hydroxymethyl-5-methoxy-2,2,4,6,7-pentamethyl-2,3-dihydro-1benzofuran.

As used herein in this application: 1C n~rwoRo\r~cNvmsP~c~Ncni~ -4 the term "alkyl" means univalent radical It includes the straight and branched chain saturated aliphatic hydrocarbyl moieties having the indicated number of carbon atoms. For e.ample, the terms "CI- 9 alkyl" and "CI-8 alkyl" refer to a saturated straight or branched chain hydrocarbon radicals having from one to nine and one to eight carbon atoms respectively, preferably having one to six ("C 1 6 alkyl") and, more preferably having one to four carbon atoms ("C1- 4 alkyi"). Included within the scope of this term are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tertiary butyl, pentyl, isopentyl, hexyl, 2,3dimethyl-2-butyl, heptyl, 2,2-dimethyl-3-pentyl, 2-methyl- A hexyl, octyl, 4-methyl-3-heptyl, nonyl and the like.

Likewise, C 1 -6 alkyl preferably has C1- 4 alkyl. All C 1 -4 15 alkyls, including the foregoing preferences, can have 1, 2, 3, ot 4 carbons in any arrangement.

e"° the term "alkylene" means saturated divalent alkane radical Likewise the term "alkylene" includes 20 straight or branched-chain moieties. Some examples of branched-chain alkylene moieties are ethylethylene, 2-

A

methyltrimethylene, 2,2-dimethyltrimethylene, and so on.

:For example, C 3 alkylene can mean

I

-CH

2

-CH

2

-CH

2 or or -CH,-CH- or -CH-CH 2 C C o

CH

3 CH3 CH3 the term "alkenyl" means unsaturated univalent radical. It includes the straight and branched chain unsaturated aliphatic hydrocarbyl moieties having thp indicated number of carbons. For example, the term "C 2 -6 alkenyl" refers to an unsaturated straight or branched ,i chain hydrocarbon radical having from two to six carbon atoms. Included in the scope of this term are ethenyl, propenyl, 2-methyl-2-propenyl, butenyl and the like; r ::ti the designation or -CO- refurs to a carbonyl group of the fcrmula: The term -C(O)R includes those carbonyl moieties wherein R is H or C1-9 alkyl moiety, embracing, for example, formyl, methylcarbonyl, ethylcarbonyl, propylcarbonyl, and the like. The term -COOR includes those alkoxycarbonyl moieties wherein R is H or C1- 6 alkyl moiety embracing, for example, methoxycarbonyl, ethoxycarbonyl, t-butyloxy- I carbonyl, and the like. Alkoxycarbonyl wherein R is not H are also called esters; the NR 7

R

8 moieties include the amino, mono and disubstituted amines with R 7 and Ra being as defined: the term "Bn" refers to a benzyl functionality of the formula:

CH

2 the term "aralkyl" refers to moieties of formula (CH m wherein m=1,2,3 or 4

-(CH

2

)M

including benzyl, phenylethyl, phenylpropyl or phenylbutyl moieties; the phenyl moieties of which may bear 1,2,3 substituents selected from the group consisting of C1- 4 i alkoxy (preferably methoxy), C-4 alkyl (preferably Smethyl) or halogen (preferably chloro but including bromo and iodo) at the position ortho, meta or, para; 6 the mono and di-hydroxy substituted alkyl moieties are thc3e moieties wherein the alkyl moiety can bear one or two OH groups (other than two hydroxy groups on one carbon atom), preferably moieties bearing a hydroxy group on a terminal carbon atom;

C

2 9 acyloxy alkylene (C2- 6 are those compounds wherein the acyloxy moiety has 2 to 9 carbon atoms and the alkylene moiety has 2 to 6 carbon atoms such as exemplified by

-CH

2

CH

2

-OC(O)CH

3 :1 the -C 2 6 alkylene-O-(CH2 2 -4 OH moieties have respectively a divalent 2-6 carbon atom moiety attached to an oxygen The oxygen is also attached to a 2-4 carbon 15 moiety terminating in a hydroxy moiety, one example is

-CH

2

CH

2

OCH

2

CH

2

CH

2

OH;

piperidino refers to compound of formula: 6 2 5 3 4 pyrrolidino refers to :)mpound of formula: piperazino refers to compound of formula: 6 2 6 2 5 3; 4 (nco morpholino refers to compound of formula: C

K

~i j. -7

OH

tlie designation:

-(R

OH

refers to hydroquinone, substituted hydroquinone and it is understood that R can be attached in any of the 2,3.5, or 6 positions; 4

(R)

the designation I 3 7 1 refers to a benzofuran derivatives, substituted o benzofuran and it is understood that R can be "attached in any of the 2,3,4,5,6, or 7 position; 15 benzofuranol derivative refers to benzofuran derivative; and \4 the designation 6- I 2 20 7 refers to a benzofuranone, substituted benzofuranone and it is understood that R can be attached in any of the 2,4,5,6, or 7 positions.

The designation refers to a bond that protrudes forward out of the plane of the page.

The designation refers to a bond that protrudes backward out of the plane of the page.

The term "pharmaceutically acceptable salts" include those acid addition salts derived by reaction with acids, for example, hydrochloric, hydrobromic, sulfuric, nitric or phosphoric acids and such organic carboxylic acids as acetic, propionic, glycolic, maleic, tartaric, citric, salicylic, 2-acetyloxybenzoic acids or organic sulfonic a. ~T~rafl.$~ A ~a.A 1 C o r i o ur eo oo o 1 oo o o i:l Yo b i it ,:i .s

L,,I,

i: i ii i i -ii Ib ~fY;il -8acids such as methanesulfonic, 4-toluenesulfonic and naphthalenesulfonic acids. Of course other acids well known to the pharmaceutical art may also be utilized. The term "pharmaceutically acceptable salts" may also include hydrates.

Stereoisomers of compounds as discussed herein is a general term for all isomers of these compounds that differ only in the orientation of their atoms in space. It includes geometric (cis/trans) isomers, and isomers of compounds with more than one chiral center that are not mirror images of one another 10 (diastereomers or diastereoisomers). The term "enantiomer" refers to two stereoisomers that are non superimposable mirror images of one another. The term "chiral center" refers to a carbon atom to which four different groups are attached. The nomenclature R/S is used as described in IUPAC-IUB Joint Commission on Biochemical Nomenclature, Eur.J.Biochem. 138: 9-37 (1984).

15 A chiral material may either contain an equal amount of the R and S isomers in which case it is called "racemic" or it may not contain equal amounts of R and 3 isomer in which case it is called "optically active", or "nonracemic".

A mixture may be resolved or isolated according to conventional and 20 standard procedures well known in the art, chromatographic separation on chiral stationary phase, use of optically active esters, fractional crystallization of addition salts formed by reagents used for that purpose, as described in "Enantiomers, Racemates, and resolutions", J. Jacques, A. Collet, and S.H.

Wilen, Wiley (1981), enzymatic resolution and the like. The term "resolution" 25 means separation of a racemic mixture into its optically active compounds. In addition, enantiomers may be prepared by utilizing enantioselective or asymmetric synthesis -which are well known by a person of ordinary skill in the art. The term "enantioselective" or "asymmetric" means the ability to produce a product in an optically active form.

CWk'N=RDENMhPSFECNWM9W575ODOC s r r r r o~ .ill:C o r~ jl oi r oo :il:ts o o ":il o i a -iS ,tl: r ij r '1 -9- It is understood that the compounds discussed herein may exist in a variety of stereoisomeric configurations.

The term "enantiomeric excess" or ee refers to the percent by which one enantiomer, El, is in excess in a mixture of the two enantiomers, El plus E2, such that; (El E2) x 100% ee (El E2) 10 the term refers to the plus enantiomer, refers to the minus enantiomer.

"Pg" means a suitable protecting group. "Protected hydroxy" means protecting group (Pg) attached to the oxygen of the hydroxy group in place of H. Suitable protecting groups can be found in T. W. Greene and P. Wuts, 15 Protective groups in organic synthesis, 2nd. ed., John Wiley Sons Inc., New York (1991), incorporated herein by reference. For convenience in schemes Pg may also be a hydrogen atom.

Prior to the disclosure in the parent application compounds of formula (I) 20 had been synthesized by the process depicted in the following reaction SCHEME I as disclosed in Patent Application W093/20057, filed March 1993.

i -i i

~I

a

I

I:

I

1 a j C TwNWORDnUEI nnM\PECNKi6SSS750LDOC ui i e i' V

F-

1 10 SCHEME Previous synthesis Protection R4 PgO 3 Pg

R

6 R7 Formation of 6-member ring PgO Ringcontraction R2 COOH Reduction -R2 (4) Halogenation and Substitution

(I)

Fries rearrangement of substituted acrylic acid diesters of hydroquinones at elevated temperatures such as 120-150 0 C gives the 6-member ring (protected 6-hydroxy-3,4-dihydro-l,2H-benzopyran-4-one) Ringcontraction of the resulting enolizable ketone with thallium(III)nitrate in trimethylorthoformate/methanol gives compound The acid moiety of is then reduced to its corresponding alcohol, the so-produced alcohol is converted to a halogen which is then substituted by an r .y, 4. P 1 -Il- 11 amino groul to give che desired 2,3-dihydro-benzofuranol This synthesis uses in the ring contraccion step thallium 'III) nitrate salts which have to be handled with great care because of their high toxicity. Moreover, the 5 use of these salts can induce problems of treating the Siwaste, the solvent and all he material contacting the K'l salts. EDue to all these inconveniences the large scale I preparation of 2,3-dihydro-benzofuranol derivatives may be impractical.

The parent application of this application discloses a new process which circumvents the ring 1" contraction step and the use of thallium(III)nitrate salts- This new process, using a Friedel-Crafts reaction which will be defined, gives the new intermediate 5-member ring from the starting hydroquinone directly instead of the LJ." 6-member ring previously obtained SCHEME I. This new intermediate is used to obtain the stereoisomers or the mixture of the 2,3-dihydro-benzofuranol derivatives D Racemate or optically active de:ivatives may be V obtained as depicted in the following SCHEME .I.

Scheme II, Step A: In '-he present invention the aryl used in the Friedel- Crafts reaction is a substituted hydroquinone of formula wherein R 4 R6 and R 7 are defined above.

HO

S..OH..

R6 2,6-dimethylhydroquinone (R 7 is hydrogen) and 2,3,5trimethylhydroquinone are commercially available. Other substituted hydroquinone may be easily synthesized t W L I I I I U LI t 0 n y a r o x y g r o u p 2. A Process according to claim 1 wherein said reaction Of compound J0~ with lipaselcandida cylndrecee and vinyl acetate is carried out in ether.

3 .A proces~s according to claim 2 wherein said etheristbylmhyehr

A

-12 using well known methods in the art such as for example disclosed in tlMethoden der Organischen Chemie" Houben Weyl, band VII/3a chinone, teil 1 and disclosed by J.T. Gupton et al.

Org. Chemn. 1983, 48, 2933-2936) which are incorporated T, 5 herein by reference.

Hydrdxy groups of the trialkylhydroquinone are optionally protected using suitable protecting groups (Pg) to give protected hydroquinone 13). Many protective groups used usually for alcohols are applicable to phenol. Ethers and esters are the most common protective groups used.

as for example methyl, cyclohexyl, isopropyl, t-butyl, or methoxyr.athyl, benzyloxymethyl, 2-(trimethylsilyl)ethoxymethyl, tetrahydropyranyl, allyl, benzyl, or silyl ethers such as trimethylsilyl, t-Butyldimethylsilyl. Ester means forming esters (-OCOR) such as for example an aceate

(-OCOCH

3 levulinate (CH 3

COCH

2

CH

2

CO

2 pivaloate *0((CH 3 3 CC0 2 benzoate (-0C0C 6 11 5 cart1:nates such as methyl carbonate (-OCOCH 3 aryl carbon'ate, benzyl carbonate

(-OCOCH

2

C

6

H

5 carbamates (-OCONHR) phosphinates such as f.dime thy lphosphonylI ester ((CH 3 2 sulfonates such as methylsulfonate or mesyl (-OSO 2

CH

3 or toluene sulfonate or tosyl S0 2

C

6

H

4 -p-CH 3 *1 13 SCS.EM.E I I R4 Hyd roxy Protection 1) Fried el-Crafts Reaction 'OPg Hyd roxy Protection Pg, Hyd-rob oration! oxidation Step D: O lefination R2 PgO R2

R

6 r R7 SCHIIE IIl RACEMATE or OPTICALLY ACT WE COMPOUNDS by resoluticn of the alcohol (9) It is found that the protective groups, may influence the formation of the five member ring. The alkyl g,.oups as protecting groups are preferred and more ,?r-:ferably the methyl group is chosen.

Preferablyr the trialkyihydroquinone is protected 14 using a common reaction such as treating trialkylhydroquinone with dimethylsulfata cr methyliodide in the presence of a base such as potassium carbonate, sodium hydroxide, potassium hydroxide in a solvent such as acetone, alcohols (for example methanol, ethanol) preferably under reflux. The protected hydroquinone is isolated according'to well known procedure in The art.

Scheme II, Step B: The new intermediate is obtained by using a Friedel--Crafts acylation as disclosed in Methodender se** OrganischenChemie (Houden-Weyl, VII/2a teil I, 1973); or in Friedel-Crafts and related reactions (Interscience, New York, 1963too* 15 1964), which are incorporated herein by reference. The Friedel-Crafts reaction involves the reaction between an aryl and an acyl halide, a carboxylic acid, an anhydride or a ketene in presence of a catalyst. For the acyl halide all four halides (Cl, Br, I, F) can be used. In the 20 present invention, the reagent to accomplish the formation of is preferably a 2-halogeno-2-(C1-4)alkyl(C1-6) acylhalide or a 2-halogeno-2-(C 1 -4)alkyl(CI- 6 )acid of I s formula R 2

-C(W)(R

2 )C(O)V wherein W is hydrogen or halogen .2 such as iodide, bromide, chloride or fluoride and more preferably bromide or chloride and V is halogen as defined above or hydroxy As fc example when R 2 is methyl or ethyl, respectively the 2-bromo-2-methylpropionic bromide or 2-bromo-2ethylpropionic bromide are commercially av-ilable. When R 2 is propyl, commercially available 2-propylpentanoic acid can be converted to a 2-halogeno-2-(C 1 -4)alkyl(1C-6) acid by replacing the a-hydrogen with a halide. More preferably the a-hydrogen is replaced by iodide, bromide or chloride.

The "a-hydrogen" means the hydrogen attached to the carbon t directly adjacent to the carbonyl function. The a-hydrogen can be replaced by bromide or chloride by well-known

IN

-i 1 1 1

RR

R7 rr-~h~nvoRouPnn~sp~asc~Pfii saLax, jj: I 15 et S S methods in the art for example using bromine or chlorine with a phosphorus halide as a catalyst (the reaction is known as the Hell-Volhard-Zelinskii reaction, and chlorosulfuric acid may be also used as a catalyst to obtain the carboxylic acid a-iodinated, as well as chlorinated or brominated), using N-bromosuccinimide or Nchlorosuccinimide and bromic acid or chloric acid. A carboxylic acid may be a-chlorinated using cuprous chloride in po2ar inert solvents. An acyl chloride can be Qiodinated with iodine and a trace of iodic acid.

The Friedel-Crafts reaction is accomplished most commonly in a solvent such as dichloromethane, dichloroethane, tetrachloroethane, chlorobenzene, nitromethane or ca.bon disulfide or without any solvent.

Catalysts are Lewis acids. "Lewis acids" are species with a vacant orbital. The most common catalysts are ferric chloride, iodine, zinc chloride, aluminium chloride and iron and more preferably aluminum chloride or ferric chloride is used. Preferably the catalyst is used at a ratio of 0.1 to 2 per mole of reagent.

More preferably the hydroquinone is treated with R2-C(halogeno)(R 2 )C(O)halide in a solvent such as dichloromethane, dichloroethane, tetrachloroethane for example in the presence of Lewis acid catalysts (aluminum chloride or ferric chloride) at a range of temperature between -100C to 1000C. The so-produced benzofuranone is isolated and optionally deprotected by common methods well 0 known in the art.

A byproduct 1,4-di-(2-halogeno-2-alkyl-alkylacetoxy)- 2,3,5-trialkylhydroquinone may be formed during the reaction and the mixture may require a supplemental step of saponification to obtain the desired product. Therefore the mixture is treated by basic conditions such as potassium hydroxide or sodium hydroxide in a mixture of

I.I

H';1 1> ~:iif Ei~

I

:Yq!u i- 16 r solvent such as aqueous methanol/tetrahydrofuran at a range of temperature of 40 to 80oC. More preferably an aqueous solution of sodium hydroxide is added to the crude product dissolved in methanol/tetrahydrofuran 1/1 and the reaction is conducted at 600C for 3 to 5 hours. The new benzofuranone is isolated by standard methods.

Scheme II, Step C The 5-hydroxy group of the so-produced benzofuranone is protected. Suitable protecting groups as described previously are used. More preferably reagents such as 2methyl-proprionylhaliod, methylhalide or benzylhalide are used. More preferably 2-methyl-proprionylchloride is added to a solution of benzofuranone in solvent such as for example dichloromethane and the mixture is stirred at a range of temperature between -50C to 10C under inert atmosphere. The protected compound is isolated by extraction with quantitative yield and can be used without further purification for the next step.

Alternatively, more preferably benzylbromide or benzylchloride is added to a solution of benzofuranone (6) in solvent such as for example acetone, dichloromethane, tetrahydrofuran, dimethylformamide or, dimethylulfoxide in presence of a base such as potassium carbonate, potassium hydroxide, sodium hydride or, sodium amide. More preferably, benzylbromide is added to a solution of benzofuranone in acetone in the presence of potassium carbonate and the mixture is stirred at a range of temperature of 5°C to 65 0 C. The protected compound is isolated by filtration or standard methods well known in the art.

:d I a 'i (1 t 6 r :x i\ :I I I Ir 1 ,is t~LDcc 1 ff-^ 'i l :ir i

I

!iil

I

Il i L 17 Scheme II, Step D: *00* r*00 *0* *0 00

S.

0 0 0 *r 00* 0 00 C The ketone group of the benzofuranone is converted to the exo-methylene group (exo-methylene group means divalent C 1 radical which is attached as a side chain rather than included into a ring) using methods known in the art such as for example the method known as the Wittig reaction or a 2-step process involving alkylation with methyllithium or a methyl magnesium halide reagent followed by acidic catalyzed elimination of the tertiary alcohol.

In the Wittig reaction ketone group of the benzofuranone is treated with a phosphorus ylid (also called phosphorane which means a substance in which a 15 carbanion is attached to a heteroatom with a high degree of positive charge, i.e. -C--X to give an olefin as disclosed by Johnson in YlidChemistry (Academic Press, New York, 1966) which is incorporated herein by reference. Phosphorus ylides are usually prepared by treatment of a phopshonium salt with a base, and phosphonium salts are commercially available or usually prepared from a phosphine and an alkyl halide. Phosphonium salts are most often converted to ylides by treatment with strong bases such as for example butyllithium, sodium or potassium amide, hydride or alkoxide. Solvents such as for example tetrahydrofuran are generally used. The reaction is performed at a range of temperature between -5 0 C to 35oC under inert atmosphere.

More preferably, alkoxide such as for example potassium tert-butoxide is added portionwise at about OOC under inert atmosphere to a suspension of benzofuranone and methylphosphonium halide in dry tetrahydrofuran. The mixture is then treated as known in the art to give the product in good yield. The quality of the alkoxide such as potassium tert-butoxide is important to improve the yield of olefin Alternatively, in the alkylation/elimination process 00.

of 00 000 0e *5 2' i+i r~ T a 1 lawgzz- c4 18 ketone is treated by a methylmagnesium halide such as methylmagnesium chloride, methylmagnesium bromide or methylmagnesium iodide. More preferably, the ketone is treated by methylmagnesium chloride in ether solvents such as for example ether, tetrahydrofuran at a range of temperature from -5 0 C to 50 0 C to give an intermediate tertiary alcohol. Addition of acid such as concentrated sulfuric acid causes elimination to give the desired olefin which is further purified by well-known methods in the art such as for example by crystallization.

Scheme II, Step E: ~Conversion of exo-methylene of in the methyl- 15 alcohol group can be accomplished using hydroboration/ oxidation. Olefins are treated with borane in ether solvents. Usually, borane complexes such as with tetrahydrofuran, dimethylsulfide, or tertiary amine which are commercially available are used. Borane can also be prepared insitu by well known methods in the art, by reacting sodium borohydride and boron trifluoride.

More preferably, the olefin is treated with a solution of borane-methyl sulfide complex at about OOC under 5 inert atmosphere in solvents such as for example I chloroform, dichloromethane, cr ethers such as diethylether, tert-butyl methyl ether, tetrahydrofuran.

Borane adds to the olefin to form an intermediate which is oxidized. The so-produced organoborane can be oxidized to the primary alcohol (primary alcohol means an alcohol in which the carbon attached to the hydroxy group is linked to one or no alkyl group and at least two hydrogen atoms) with sodium hydroxide-hydrogen peroxide as known in the art.

The so-produced alcohol can be utilized without further purification. i *H A I* a41 -te -c; i- i t

I~

I;aet i: bi-'l~ r, V m: r 19 Scheme III, Step A: o a~ 4 a In general, the racemic 3-hydroxymethyl-benzofuran (9) may be resolved or isolated according to conventional and standard procedures well known in the art, e.g., chromatographic separation on chiral stationary phase, use of optically active esters, fractional crystallization of addition salts formed by reagents used for that purpose, enzymatic resolution and the like. More preferably the 3hydroxymethyl-benzofuran is resolved using enzymatic resolution. More preferably enzymatic transacylation, wherein one enantiomer is reactive and acylated, and the other one remains unchanged, is utilized to resolve the alcohol.

Enzymes gc.terally employed are lipases from microorganisms like Candidacylindracea, Rhizopusarrhizus, Chromobacterium viscosum, Pseudomonas cepecia, Mucor miehei or, Asperigillusniger, or from mammalian liver, like porcine 20 pancreatic lipase (PPL), or enzymes from Boehringer Mannheim Chirazyme L-l, L-2, L-3, L-5, or L-6. The enzyme may be used as a crude extract or in purified form, sometimes entrapped in sepharose or in chromosorb as a solid support. Acylations are run as transesterifications of the alcohol with esters such as for example methyl acetate, acetic anhydride, vinyl acetate, isoprenyl acetate, 2,2,2-trifluoroethyl acetate with the enzyme in organic solvents such as for example ethers such as ether, t-butyl methyl ether, tetrahydrofuran or other solvents such as benzene. According to the present invention, 3-hydroxymethyl benzofuran is resolved using lipase from Candida cylindracea microorganism. More preferably the reaction is performed using vinyl acetate in ether solvent such as for example t-butyl methyl ether at room temperature or a range of temperature from 0°C to 50°C.

i ii r:

L

1, r' d ~1 i u~4i,-- !,e ii i~i g i t i i; I 1.

-i;a Ih i.

i:-i i: 6 i i i :l i: l l .c, '~Fl~f "a: i,

I

I

.w r r o r i ri-iii.

,I:

b~V

-I'

r.

S I(~ p ;s 1: v :ii Cit I a

CI

CC

20 The optically active acetyl de:ivative of and the unreactive alcohol may be isolated by procedures well known in the art. For example, the mixture is filtered, concentrated under reduced pressure to constant weight and the residue is chromatographed on silica gel to give the acylated isomer and the unchanged isomer. Other methods such as for example HPLC (High Purification Liquid Chromatography) or crystallization may also be used. The acetyl isomer may be deesterified by well known procedures in the art wherein for exampl:2 the acetyl isomer is dissolved in methanol; and treated by basic conditions such as for example potassium carbonate at a range of temperature from 150C to 60 0 C. The recovered desired optically active alcohol may be purified by procedures 15 well known in the art, such as crystallization.

The undesired optically active alcohol may be recycled one or more times. More preferably, the hydroxy group of the 3-hydroxymethyl is transformed into a leaving 20 group and more preferably into a mesylate by standards methods known in the art. Elimination of the so-produced leaving group leads to the olefin which can be injected into the process. Elimination can be c ried out using procedures well known to one of ordin°-oy skill in the art.

More preferably, the leaving group is eliminated in basic conditions such as for example potassium tert-butoxide in tetrahydrofuran at room temperature.

The 5-hydroxy of compound may be further deprotected using common methods well known by one skilled in the art. More preferably if isobutyryl has been used to protect the 5-hydroxy, basic conditions such as sodium hydroxide in a mixture of solvents such as water/methanol/tetrahydrofuran can be used at 70 0 C-85 0

C.

I

SI

.4 '4

-I.

44;

I

.ti (I* -1 3 acetic, propionic, glycolic, maleic, tartaric, citric, salicylic, 2-acetyloxybenzoic acids or organic sulfonic 21 SCHEM4E III Pgo PgO beC 0..

(S-9) 47

'A

4 ~it '4'4' &V

C

Does 9, OH

A

Step B Hydroxy conversion to a leaving group PgO Step C Substitution of Lg Scheme III, Step B: The primary alcohol of 3-hydroxymethyl-2,3-dihydrobenzofuran derivative obtained is transformed into a leaving group (Lg) which means a group which can be easily substituted by a nucleophile. Leaving groups are for example tosylate, brosylate, nosylate, mesylate, triflatQ, nonaf late, tresylate or halides.

4, 'V CA \WINWORDLENYMISPECNKM9575DI.DOC 22 More preferably, the hydroxy group is converted into a halide or a mesylate. When the hydroxy group is converted to a halide the most common reagents utilized are for example halogen acids or thionyl halide, phosphorus pentahalide, phosphorus trihalide, phopshoryl halide, trialkyl phopsphorylhalide, triphenylphosphine halide and the like (wherein halide means halogen such as chloride bromide (Br) or iodide More preferably, the hydroxy group is converted into a bromide by using triphenylphosphine-bromine which is prepared insitu in solvent such as dichloromethane at a range of temperature from -5 0 C tc 100C. The alcohol is added to this mixture at such temperature and then allowed 15 to warm to room temperature. The mixture treated as known in the art gives the product (10) in quantitative yield.

Alternatively, the hydroxy group is converted to a mesylate. The reaction may be performed in basic condition such as in pyridine at room temperature or, more preferably in tetrahydrofuran in presence of a base such as triethylamine at a range of temperature -5 0 C to 20 0 C for example.

25 Scheme III, Step C: To obtain the final compound of formula wherein X is CH 2 A, A being as defined previously, the leaving group is substituted by the desired amino -NR 7 RB, pyrrolidino, piperidino, morpholino or piperazino group.

The desired amines HNR 7

R

8 are commercially available or easily synthesized using well known methods in the art as for example described in Comprehensive Organic Chemistry (Chapter 1.3, Synthesis of amines and ammonium salts, Trost-Flemming, Pergamon Press, 1991), which is incorporated moll% '*a ~i

;L

II:

23 herein by reference. The most common reactions involve reaction between the desired alkyl halide and ammonia: R-X NH3 o R-NH 2

+HX

More preferably fcr the synthesis of a primary amine reduction'of an azide obtained by substitution of an alkylhalide or the reaction known as Gabriel reaction, see for example E.F.V. Seeven and K. Tumbell, Chem.Rev.

1988,88, 297 (incorporated herein by reference) involving reaction of a phthalimide ion with an appropriate alkylating reagent and subsequent removal of the phthaloyl group are used. Pyrrolidine, piperidine, morpholine, piperazine and N-methyl piperazine, 2-methylpiperazine, piperazinylformic acid are commercially available.

The substituted piperazine of formula HN~ N-Rlo :i

:I

;1 i

.II

i 1 sucan be easily synthesized by common methods as described above for the amines HNR 7 Rs.

The piperazine of the above formula wherein Y is COOR 7 can be easily synchesized by esterifying the commercially available piperazinylformic acid with the desired alkyl reagent by using common esterification methods well known by one ordinary skilled in the art.

The substitution of the leaving group of compound by the desired amino group may be performed by procedures well known in the art such as for example in acetonitrile, dimethyl formamide, methanol, ethanol or isopropanol under reflux temperature. After extraction, the final product can be either isolated using column chromatography or by crystallization. Crystallization seems to give better yield than column chromatography. Optionally the 5-hydroxy s I: J "r-

.I

x- .t'i 9 jli;tl- y d~aa au j B~CB~ ~t Ta*- -24group may be deprotected according to well known methods in the a For the resolution the 5-hydroxy group is preferably protected by an ester group such as an acetate group this may need a previous deprotection if the 5-hydroxy group is protected by an alkyl group such as a methyl or benzyl.

The deprotection of the hydroxy group protected as a methoxy is performed using common reagents of deprotection, which are for example trimethyl silyliodide, boron tribromide, boron trifluoride, trimethylsilyl methylsulfide or trimethylsilyl phenylsulfide, aluminium halide (halide being chloride or bromide), according to procedures well known in the art.

Ic The deprotection of hydroxy group protected as a benzyloxy is commonly performed by catalytic or chemical reduction, using for example palladium on carbon in ethanol, sodium in ammonia or ethanol, trimethyl S, silyliodide in dichloromethane, and others.

ONIl 0 ti 1 ;W N< Ft UE NY CP C t~e 9 S7 0t OCJi 1 2, 6-dime thyihyd roqui none (R 7 is hydrogen) and 2,3,5trimethylhydroquinone are commercially available. Other substituted hydroquinone may be easily synthesized 4

I

A

:1 *1 j I

,L

LI 25 SCHEME TV: Synthesis of optically active 2,2,4,6,7pentamethyl-3-t (4-methylpiperazino)-methyl I- 2, 3-dihydro-l-benzofuran-5-ol using alcohol resolution BnBr Base BnO (99%) 1. MeMgCI 2. H 2 S0 4 (99%) (6VI1) (7V11) 1. BH 3 .DMS BnO.

2. NaOH/ H 2 02 (87%) Lipase/ OH1- Candida cylindracea vinyl acetate (9V11) (8V1) Acetylated R-(9VI1), ee 87% 9V I) R-(9 VII), 65%, ep 98.4 -26 SCHEME TV.: (contiLnued) BnO 1,MsCI BnO (99%)0 R-(9VI1) R-(1 OVII) N-methylpiperazine HO-N

N-

2. H 2 Pd/C0 0**t (68%) 2 HCI L1 I Ii _._i(.iii-iii=l i r! i, i ~ior j

I

'4 i '1 27 The followinq examples present typical syntheses as described in Schemes II and IV. rhese examples are understood to be illustrative only and are not intended to limit the scope of the present invention in any way. As used herein, the following terms have the indicated mernings: refers to grams; "mmol" refers to millimoles; "mL" refers to millimeters; "bp" refers to boiling point; "mp" refers to melting point; refers to degrees Celsius; "mm Hg" refers to millimeters of mercury; "Pa" refers to pascals; "lL" refers to microliters; "pg" refers to micrograms; "iM" refers to micromolar; "TLC" refers to thin layer chromatography; refers to molarity; refers to normal; "[a]D 20 refers to specific rotation of the D line of sodium at 20°C obtained in a 1 decimeter cell; "GC" refers to gas chromatography; and "Rf" refers to retention factor.

C:WINWORDn'ENNYMPECNKG4SS750I.DOC Ij_-I ;i

I

i i

E

i -7-

I

S C 'CCCi 4 4rl~ t 28 EXAMPLE 1 1,4-DIMETHOXY-2,3,5-TRIMETHYLHYDROQUINONE

H

3

CO

OCH

3 (3 A mixture of trimethylhydroquinone (60.87 g, 0.4 mol), dimethylsulfate (151.36 g, 1.2 mol) and potassium carbonate (221 g, 1.6 mol) in acetone (1.6 L) is refluxed for three days under nitrogen. After cooling, 10% sodium hydroxide (400 mL) is added and most of the acetone is evaporated.

The black mixture is taken up in heptane (800 mL), the organic phase is separated and washed with 10% sodium hydroxide (2x200 mL), water (200 mL) and brine (200 mL).

The solvent is dried (magnesium sulfate) and evaporated under reduced pressure to give a yellow oil. Purification on a small pad of silica gel eluting with heptane/ethyl acetate 95:5 gave 57.4 g of 1,4-dimethoxy-2,3,5trimethylhydroquinone as a colorless oil which slowly crystallized.

To avoid the formation of large amount of 2,3,5trimethyl-l,4-benzoquinone, nitrogen is previously bubbled for 30 min into acetone. Compound 1,4-dimethoxy-2,3,5trimethylhydroquinone may be purified by distillation.

Ct ;i i j

I

the mixture is treated by basic conditions such as potassium hydroxide or sodium hydroxide in a mixture of K ?:I

L::

I, UNNIMM I. r I;~s: i ;1 29 EXAMPLE 2 5-HYDROXY-2,2 4,6 7-PENTAMETHYL-2,3-DIHYDRO-1-BENZOFURAN-3-

ONE

So a a S Cr a (6) Aluminium chloride (25 g, 188 mmol) is added portionwise at 0 0 C and under nitrogen to a solution of 1,4dimethoxy-2,3,5-trimethylhydroquinone (33.83 g, 188 mmol) and 2-bromo-2-methylpropionylbromide (129.46 g, 563 mmol) in tetrachloroethane (188 mL). The dark solution is then 15 heated at 70 0 C until completion of the reaction (3-5 days) as indicated by TLC (heptane/ethyl acetate 90:10). The reaction is quenched by careful addition of ice. The black mixture is acidified to pH 1 with concentrated hydrochloric acid and extracted with dichloromethane (2x150 mL). The organic phase is washed with water (150 mL), 10% potassium bicarbonate (2x150 mL), dried (magnesium sulfate) and evaporated to dryness. The residue (106 g) is triturated in heptane in the aim to precipitate the 1,4-di-(2-bromo-2methylpropionoxy)-2,3,5-trimethylhydroquinone formed during the reaction and filtered off (30.15 The filtrate is evaporated to dryness and the residue (66.37 g) is passed through a small pad of silica gel eluting with heptane/ethyl acetate 95:5 to give 39.78 g of crude 5-(2bromo-2-methylpropionoxy)-2,2,4,6,7-pentamethyl-2,3dihydro-l-benzofuran-3-one (Rf=0.4 heptane/ethyl acetate 90:10). The yellow solid is dissolved in a mixture of methanol/tetrahydrofuran (400 mL, 1:1) and sodium hydroxide g, 500 mmol) in water (100 mL) is added dropwise under nitrogen. The solution is stirred for 4 hrs at 60 0 C and overnight at room temperature. The black mixture is then acidified with concentrated hydrochloric acid. Most of the solvent is evaporated under reduced pressure and the 1 i ii b: ii: i ~i i i f 4'M4 3i 0 residue is taken up n ethyl acetate (300 mL). The organic phase is wasbed with water (150 mL), 10% sodium bicarbonate (2x150 mL), brine, dried (magnesium sulfate) and evaporated to dryness to give 22.39 g of the crude 2,2,4,6,7-pentamethyl-2,3-dihydro--l-benzofuran-3-one as a yellow po wder. A sample is recrystallized in heptane/diisopropyloxide, mp=142*C-144'C. Rf=0.29 (heptane/ ethyl acetate 80:20) EXAMPLE 3 5-(2-THYLPROPINOXY) 7-PENTAMETHYL-2, 3-DIHYDRO-l- BENZQFURA-N- 3-ONE

I

Y-01 (7) A solution of 2-methylpropionylchloride (isobutyrylchloride, 8.05 g, 75.55 nunol) in dichloromethane (10 mL' is added dropwise at 0 0 C and under nitrogen to a solution of hydroxy-2, 2, 4,6, 7-pentamethyl-2, 3-3ihydro-l-benzofuran-3one (12.8 g, 58.11 mmol) and pyridine (5.97 y, 61 mrnol) in dichloromethane (58 mLj. The ice bath -s removed and the mixture is stirred for 2 hrs at room temperature. Water mL) is added and the organic phase is washed with 2N hydrochloric acid (100 mL), water (100 mL), 10% sodium bicarbonate (100 mL) and brine. The solution is dried (magnesium sulfate) and evaporated to dryness to give 17 g (100%) of 5-(2-methylpropionoxy)-2,4!,4,6,7-pentamethyl-2,3dihydro-l-benzofuran-3-one as an oil which is used without purification for the next step. Rf=0.5 (heptane/ethyl 90:10) w 4

EXAIV

C2-METBYLPROPIO] 2, 3-DIHYDRO-l-BENZOFURAN 0 ILE 4 NOXY)-2, 2,4,6 ,7-PENTAM.ETHYL- Ciii 0 A S C 44 it 4 Ci (8) Potassium tert-butoxide (1.53 g, 13.68 mmol) is added portionwise at 0 0 C and under nitrogen to a suspension of methy'ltriphenylphosphonium bromide (4.9 g, 13.68 mrnol) in 15 dry tetrahydrofuran (57 mL) and the reaction mixture is stirred for 1 hr at room temperature. The 5-(2-methylpropionoxy ,7-pentamethyl-2, 3-dihydro-l-benzofuran- 3-one (3.31 g, 11.4 mmol) in dry tetrahydrofuran (20 mL) is dropwise added at 0 0 C to the yellow suspension and the 20 reaction is stirred overnight at room temperature. Water is added and most of tetrahydrofuran is evaporated under reduced pressure. The residue is taken up in ethyl acetate, washed with brine, dr 4 ,d (magnesium sulfate) and the solvent is evaporate~l to dryness. Purification by flash chromatography eluting with heptane/ethyl acetate 95:5 and then 90:10 gave 2.8 g of 3-methylene-5-(2methylpropionoxy)-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran as a yellow oil. Rf=0.79 (heptane/ethyl acetate 70:30) rI i -32 EXAMPLE 5-HYDROXY-3-HYDROXYMETHYL-2,2r4,6,7-PENTAMETHYL-2,3- DIHYDRO-1-BENZOFURAN

HO

OH

9 10M borane dimethylsulfide complex (6.1 mL, 61 mmol) is added dropwise at 0 0 C and under nitrogen to a solution of 3methylene-5-(2-methylpropionoxy)-2,2,4,6,7-pentamethyl-2,3- I 15 dihydro-l-benzofuran (11.7 g, 40.57 mmol) in tetrahydrofuran (40 mL) and the solution is stirred at room temperature for 3 hrs. Water (10 mL) is carefully added followed by addition of 3N Sodium hydroxide (30 mL) and hydrogen peroxide (10.1 mL). After stirring 2 hrs at room temperature, most of tetrahydrofuran is evaporated and the residue is extracted with ethyl acetate. The organic phase is washed with 10% sodium sulfite (10 mL), water (100 mL), brine dried (magnesium sulfate) and evaporated to dryness to give 13.29 g of a mixture of 5-hydroxy-3-hydroxymethyl- 2,2,4,6,7-pentamethyl-2,3-dihydro-l-benzofuran (Rf=0.37 heptane/ethyl acetate 50:50) and of the corresponding isobutyryl ester (Rf=0.57 heptane/ethyl acetate 50:50). j The residue is then treated for 2 hrs at 800C with Sodium hydroxide (6.5 g, 162 mmol) irA xture of water/ methanol/tetrahydrofuran 40:20: In hydrochloric acid is added ur.til pH 1 and most of the so. ent is removed under reduced pressure. The residue is taken up in ethyl acetate and the organic phase is washed with water, brine, dried S(magnesium sulfate) and evaporated to dryness.

Purification by flash chromatography eluting with heptane/ethyl acetate 80:20 to 50:50 gave 8.05 g of 5-hydroxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3r ade ni H1admoto h o etisrmvdudr'' dihydro-l-berizofuran as a yellow oil which slowly crystallized. A sample is recrystallized from ethyl acetate/heptane, mp=89-90 0

C.

EXAMPLE 6 3-BRQMQMETHYL-2, 2,4,*6 ,7-PENTAMETHYL-2, 3-DIHYDRO-l- BENZQFtIRAN- HO B 0 CO(Deprotected To an ice-cooled solution of triphenylphosphine (41.89 g, 160 mmol) in dichloromethane (120 mL) is added dropwise a solution of bromine (24.33 g, 152 minol) in dichloro-methane (40 mL) and the resulting mixture is stirred at 0CC for I hr giving a white precipitate free of bromine coloration. To this mixture- is added alcohol hydroxy-3--hydroxymethyl-2, 2, 4,6, 7-pentamethyl-2 3-dihydrol-benzofuran (34.26 g, 145 mmol) and the resulting solution is allowed to warm to room temperature and stirred for 18 hrs. The :-olution is concentrated to a small volume and is chromatographed i.n silica gel using dichloromethane/hexane 1:2 as eluent. Fractions containing the product are combined and evaporated to give 43.28 g (99 of 3-bromomethyl-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran-5-ol as an oil. A sample is recrystallized from ethyl acetate/heptane, mp 79 0 C-800C.

354 ;tI -34- EXAMPLE 7 5-METHOXY--2, 2,4,6 ,7-PENTAMETHYL-2, 3-DIHYDRO-1-BENZOFEJRAN-3-

ONE

CH

3 0 0O 0 (7) A mixture of phenol 5-hydroxy-2,2,4,6,7-pentamethy'L- 2,3-dihydro-l-benzofuran-3-one (7.16 g, 32.54 mmol), (6.16 g, 48.B mmol) and potassium carbonate (13.3 9, 97.63 mmol) in acetone (160 mL) is refluxed under nitrogen for 3 days. After cooling, 3N sodium hydroxide (100 mL) is added anid most of the acetone is evaporated *linder reduced pressure. The mixture is extracted with etliyl acetate (200 mL) and the organic layer is washed with 3N sodium hydroxide (2x100 tnt), water, brine, dried (magnesium sulfate)and evaporated to dryness to give 7.52 g of 5-methoxy-24,2,4,6,7-pentamethyl-2,3-dihydro-l- 77 benzofuran-3-one as a yellow solid which is used without purification for the next step. Rf=0.4 (heptane/ethyl acetate 90:10)

VI

EXAMPLE 8 5-METHOXY-3-METHYLENE-2,2,4,6,7-PENTAMETHYL-2,3-DIHYDRO-1-

BENZOFURAN

CH

3 0 0 (8 Potassium tert-butoxide (4 g, 35.5 mmol) is added portionwise at OoC and under nitrogen to a suspension of methyltriphenylphosphonium bromide (12.7 g, 35.5 mmol) in dry tetrahydrofuran (120 mL) and the reaction mixture is stirred for 1 hr at room temperature. The 2,2,4,6,7-pentamethyl-i,3-dihydro-l-benzofuran-3-one (5.55 g, 23.7 mmol) in dry tetrahydrofuran (40 mL) is added A, 20 dropwise at OOC to the yellow suspension and the reaction is stirred overnight at room temperature. Water (50 mL) is i carefully added and most of the solvent is evaporated under reduced pressure. The residue is taken in ethyl acetate (200 mL), washed with brine, dried (magnesium sulfate) and evaporated to dryness. Purification through a small pad of silica gel using dichloromethane as a solvent gave 5.47 g 5-methoxy-3-methylene-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran as a yellow oil. Rf=0.48 (heptane/ethyl acetate 90:10).

II 2.

Trost-Flemming, Pergamon Press, 1991), which is incorporated ~BIP~CC~I-L-" IIIICII 1:

L"

I

36 EXAMPLE 9 3-HYDROXYMETHYL-5-METHOXY-2,2,4,6,7-PENTAMETHYL-2 3- DIHYDRO-1-BENZOOFRAN r ~rO a~Q rr ij sr o r '8, 8 o in 1.4 ,r ~a I o ~i a: r rl (9 10M borane dimethylsulfide complex (2.08 mL, 20.8 mmol) is added dropwise at 0 0 C and under nitrogen to a solution of 5-methoxy-3-methylene-2,2,4,6,7-pentamethyl-2, 3 -dihydro-lbenzofuran (4.03 g, 17.35 mmol) in dry tetrahydrofuran mL). The solution is stirred at room temperature for 2 hrs. Water (10 mL) is carefully added to the solution at 0°C, followed by addition of 3N sodium hydroxide (5.78 mL) and hydrogen peroxide (5.78 mL). After 2 hrs. at 20 room temperature, most of the tetrahydrofuran is evaporated to dryness and the residue is extracted with ethyl acetate (2x100 mL). The combined organic phases are washed with 10% sodium sulfite, water, brine, dried (magnesium sulfate) and evaporated to dryness to give 4.2 g of crude alcohol.

25 Purification by flash chromatography eluting with heptane/ethyl acetate 70:30 and then 60:40 gave 4 g (92%) 3-hydroxymethyl-5-methoxy-2,2,4,6,7-pentamethyl-2,3dihydro-l-benzofuran of as a white powder.

A sample is recrystallized from hexane, mp=79-81 0

C.

Rf=0.28 (heptane/ethyl acetate 70:30).

I I I~ t iiI I ~19ich':ii i-I yield than column chromatography. Optionally the i'*w

M^'M^

j^i IIBli~FeaspwllaYllm*- ii ,r s

I,

1~ r: a

Z

,ae o o

I

or or i~c~g~i~ r

P

6 'i :ij 37 EXAMPLE 5-BENZYLOXY-2, 2,4,6,7-PENTAMETHYL-2, 3-DIHYDRO-1-BENZOFURAN- 3-ONE (7 Potassium carbonate (720 g) is added to a solution of 5-hydroxy-2,2,4,6,7-pentamethyl-2,3-dihydro-l-benzofuran-3one (453 g, 2.1 mol) in acetone (2 A solution of benzyl bromide (423 g, 2.5 mmol) in acetone (200 mL) is added portionwise over a period of 10 min. A slight exotherm is observed. After 3 hrs, the mixture is heated lo reflux.

After 39 hrs, TLC shows complete conversion to product.

The mixture is cooled to 50 0 C and filtered using 1.5 L of ethyl acetate to remove the solids from the flask. The T0 solids are washed with ethyl acetate (1.5 The filtrate is concentrated. The resulting solids are dissolved in ethyl acetate (7 This solution is washed with water, dried (magn sium sulfate) and concentrated. The resulting solids are placed on a tray for air-drying. After 2 days, 5-benzyloxy-2,2,4,6,7-pentamethyl-2,3-dihydro-l-benzofuran- 3-one is collected (638g, mp=114°C-1150C.

ir t- ;t /1 v Op MM -38- EXAMPLE 1 5-BENZYLOXY--3-M4ETHYLENE-2, 2,4,16,7-PENTAMETSYL-2, 3-DIHYDROl-EBENZOFtIRAN BnO 0 (8) A solution of methylmagnesium chloride 3.0 M (800 niL, 2.4 mol) in tetrahydrofuran s added to a solution of benzyloxy- ,2.-4,6,7-pentamethyl-2,3-dihydro-l-benzofuran-3one (500 g, 1.6 mol) in tetrahydrofuran at 0 0 C over a period of one hr. The mixture is allowed to warm to room temperature. After 15 hrs, TLC and GC showed complete conversion to 5-benzyloxy-2,2,3,4,6,7-hexanethyl-2,3dihydro-l-benzofuran-3-ol. The mixture is cooled to O 0 C and a saturated solution of ammonium chloride (350 mL) is added Pvery carefully. Concentrated sulfuric acid (300 niL) is A added dropwise over 1 hr. TLC showed conversion to -benzyloxy-3-methylene-2,2,4,6,7-pentamethyl-2,3-dihydro- -benzofuran. Water (1.5 ethyl acetate (1.5 and a solution of ammonium chloride (1 L) are added to dissolve the salts. The organic phase is dried (magnesium Eulfata and concentrated. The crude oil is transferred to crystallization dish using minimal ethyl acetate and seeded. Complete crystallization is obtained in ca.

30 min. The solid is placed on a tray and allowed to aiLdry. After 2 days 5-benzyloxy-3-methylene-2,2,4,6,7pentamethyl-2,3-dihydro-l-benzofuran (492 g, 99%) is collected.

mp=55-570C.

13 F a:N 0: -I a F^l 1 4! ^i sr' i c ^n T" aM 39 EXAMPLE 12 5-BENZYLOXY-3-HYDROXYMETHYL-2 ,2,4 6, 7-PENTAMETHYL-2 3- DIHYDRO-1-BENZOFURAN (9 A solution of complex borane-dimethyl sulfide (2.0 M) in tetrahydrofuran (950 mL, 1.9 mol) is added over 2 hrs 15 to a solution of 5-benzyloxy-3-methylene-2,2,4,6,7pentamethyl-2,3-dihydro-l-benzofuran (492 g, 1.6 mol) in tetrahydrofuran (1.6 L) under nitrogen and cooled with an ice bath. The pot temperature is maintained between 0°C- The solution is allowed to warm to rooi.i temperature.

20 After 15 hrs, the solution is cooled with an ice bath and water (900 mL) is carefully added (hydrogen evolution ceased after ca. 30 mL of water has been introduced). A solution of sodium hydroxide (3.0 M, 530 mL) is added over min. maintaining the pot temperature below 10 0 C. A solution of hydrogen peroxide 30% (530 mL) is introduced keeping the pot temperature below 20 0 C. After 3 hrs, water (1 ethyl acetate (1 L) are added. Many solids formed (Note upon acidification of the aqueous waste, all of the so2ids dissolved easily. Perhaps acidification here might better dissolve the salts). The organic phase is separated and the aqueous phase is extracted with ethyl acetate. The combined organic phases are dried and concentrated. The oily residue is poured into hexane (1 L) using hexane (800 mL). White crystals are formed. The solids are scraped from the side and swirled to increase the crystallization. The solid is collected and air dried to give benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3p r i: i! r.

i; k 1 13 i' ii r P~ i

Y:-

liP j:t r i 40

C..

0 44 dihydro-l-benzofuran (371 The mother liquor is boiled down to 700 mL and charcoal is added. After filtering through celite, seed crystals are added. Nitrogen is ulown on the solution to evaporate the hexane. The oil is washed off the solid using hexane. The solid is collected and washed thoroughly with hexane to provide 5-benzyloxy-3hydroxymethyl-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran (28 The mother liquor is concentrated. The resulting oil is plug filtered using 1.8 L of gravity silica gel collecting 4 (500 mL) fractions of 100% hexane, 6 (500 mL) fractions of 5% ethyl acetate/hexane, 4 (500 mL) fractions of 10% ethyl acetate/ hexane, and 4 (500 mL) fractions of 20% ethyl acetate/hexane. The fractions containing the desired product are concentrated. The oil is transferred to an erlenmeyer using hexan% (300 mL).

After standing overnight, 5-benzyloxy-3-hydroxymethyl- 2,2,4,6,7-pentamethyl-2,3-dihydro-l-benzofuran (42 g) is collected. The mixed fractions and the mother liquor from above are concentrated and plug filtered through 1 L of gravity silica gel collecting 2 (250 mL) fractions of 100% hexane, 8 (250 mL) fractions of 5% ethyl acetate/hexane, (250 mL) fractions of 10% ethyl acetate/hexane, (250 mL) fractions of 20% ethyl acetate/hexane. Fractions containing the desired product are combined. The residue 25 is dissolved in hexane (100 mL). After standing overnight, g of 5-benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl- 2,3-dihydro-l-benzofuran are collected. Total of 5-benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3dihydro-l-benzofuran collected is (451 g, 87%).

S

k C\WNWORDLJENNYMSPECNM95575TDLDOC -41- EXAMPLE 13 S-5-BENZYLOXY-3-HYDROXYMETHYL-2, 2,4,6, 7-PENTAMETHYL-2, 3- DIHYDRO-1-BENZOFURAN BnO O H 0 R (9) *1 41 -BA mixture of 5-benzyloxy-3-hydroxymethyl-2,2,4,6,7- Dentamethyl-2,3-dihydro-l-benzofuran (40.1 g 0.12 mol), i i and stirred for 24 hrs. The mixture is filtered and the filtrate is concentrated (60 0 C/15 tor) to constant weight.

o The residue is chromatographed over silica gel (hexane:ethyl acetate; 4:1) and the first fractions 20 contained the desired acetate (19.7 g, e.e. 87%, Rm pasconfiguration). The so-produced acetate (19.7 g, g; 0.053 mol) is dissolved in 400 L methanol and treated with potassium carbonate (2.0 g, 0.014 mol). The mixture is fstirred at room temperature for 5 hrs. (TLC; no starting material present), and then the solvent is evaporated (60oC/15 tor). The residue is taken up in ether/water and the ether extract is washed with brine and dried over magnesium sulfate. The solvent is evaporated to give 17.0 g oily residue which is dissolved in 800 mL hexane.

Crystallization occurred ove a 48 hrs period. The solvent is derrnted from the crysta mass and without disturbing *the crystal mass cold hexn e (150 mL) is added and gently swirled. The he xane is decanted and an additional 150 mL of cold hexane is added and the R-5-brnzyloxy-3hydroxymethyl-2,2,4,6,7-pentamethyl-2,3-dihydro-1- Sagbenzofuran is collected and dried to g (27%) 0 g oly rsid ue whh is d o20 7.9 in 0methanol)e.

Sthe rystal mass cold hexne

D

150 m) is added and gently M ill a -41 -42- Anal.Calc'd for C 2 1H 25 0 3 C,77.27; H,8.03.

Found; C, 77.35; H, 7.96.

Proce.3s for recycling the S-5-benzyloxy-3-hydroxymethyl- 2,2,4,6,7-pcntamethyl-2, 3-dihydro-l-benzofural.

Mesyl chlor'ide (13.3 g, 116 mmol) is added portionwise over min. to a solution of S-5-benzyloxy--3-hydroxymethyl- 2,2,4,6,7-pentamethyl-2,3-dihydro-l-benzofuran (31.6 g, 97 mol) and triethylamine (11.8 g, 116 rnmol) in tetrahydrofuran (300 mL) at 0 0 C. The mixture is allowed to *0 warm to room temperature. Aftar 3 hrs, potassium tertbutoxide (39 g, 348 mmol) in tetrahydrofuran (200 mL) is added over 30 min. The solution is allowe,3 to warm to room temperature. After 1 hr, water and ethyl acetate are added. The organic phase was washed with brine, dried (n.agnesium sulfate), and coiLcentrated. The oil was transferred to a crystallization dish using minimal hexane and seeded. Complete crystallization occurred after in.The solid is allowed to air dry overnight to give 30.2 g of 5-benzyloxy-3--methylene-2,2,4,6,7- ****pentamethyl-2,3-dihydro-l-benzofuran and recycle. Total4 yield of 5-benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl- ***2,3-dihydro-l-benzofuran after 3 recycles a3 uvernignt at room temperature. *rne D.Lau-. IuLLL.UL ai acidified with concentrated hydrochloric acid. Most of the solvent is evaporated under reduced pressure and the 1<1*~ 8-,"I 43 EXAMPLE 14 R-5-BENZYLOXY-3-(METHANESULFONATO)-2,3-DIHYDRO-2,2,4,6,7- PENTAMETHYL-1-BENZOFURAN BnO.

Su..

P..

o a, S* 0 a 9.

0 o* p o 040 i -i R (10 Methanesulfonyl chloride (8.4 g, 74 mmol) is added portionwise over 15 min to a solution of R-5-benzyloxy-3hydroxymethyl-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran (20 g, 61 mmol) and triethylamine (7.5 g, 74 mmol) in tetrahydrofuran (200 mL) at 0°C. After 30 min, the mixture is allowed to warm to room temperature. After 20 3 hrs, the mixture (now a slurry of a white solid) is poured into 5% hydrochloric acid (200 mL) using ethyl acetate (100 mL). The organic phase is washed with brine, dried (magnesium sulfate) and concentrated to give benzyloxy-3-(methanesulfc .aco)-2,3-dihydro-2,2,4,6,7pentamethyl-l-benzofuran-3-methanol as a white solid (24.7g, mp=122-123 OC, 20 (methanol) +10.5.

1 i l~il pur.ritcation ror Lne nex. e. b)-u.j acetate 90:10) I: i: i-a

~I

B

-F e 44 EXAMPLE R-5-HYDROXY-3-[(4-METHYLPIPERAZINO)-METHYL -2,3-DIHYDRO- 2,2,4,6,7-PENTAMETHYL-1-BENZOFURAN DIHYDROCHLORIDE HYDRATE Ca S a a.o 4 a 43 0

S.D

Aaa~ a aear Xe *a a a'l~ i;1

/-N

N N- 2 HCI. R (I A mixture of R-5-benzyloxy-3-(methanesulfonato)-2,3- 15 dihydro-2,2,4,6,7-pentamethyl-l-benzofuran (13.3 g, 33 mmol) and 4-methylpiperazine (6.6 q, 66 mmol), and potassium carbonate (18 g, 0.13 mol) in acetonitrile (200 mL) is heated under reflux for 18 hrs. The mixture is cooled to room temperature and concentrated. The residue is dissolved in water/chloroform. The aqueous phase is extracted with chloroform. The combined organic phases are dried (magnesium sulfate) and concentrated to give crude R-5-hydroxy-3-[(4-methylpiperazino)-methyl]-2,3-dihydro- 2,2,4,6,7-pentamethyl-l-benzofuran (16 The crude 25 product is dissolved in ethanol (50 mL) and acetic acid mL) and added to 1.0 g of 10% palladium on carbon in a Parr bottle. This mixture is placed on a Parr shaker under 345 kPa of hydrogen for 18 hrs. The catalyst is removed by filtration through celite and the filtrate is concentrated.

TLC showed starting material still present. The residue is dissolved in ethanol (50 mL) and acetic acid (50 mL) and added to 2.0 g of 10% palladium on carbon in a Parr bottle.

After 18 hrs, the mixture is filtered through celite and concentrated. The 1 H NMR of the crude product showed complete debenzylation. To the crude product is added a dilute solution of hydrochloric acid (8 mL of concentrated hydrochloric acid in 20 mL of water) followed by ethanol i i I I A' ,L i b i: r~ 411b8 ii

I'

.I

1

J

I

i it 45 mL). The solution is concentrated to dryness. The residue is dissolved in hot isopropanol (100 mL) and ca. 1 g of charcoal is added. After filtering through celite, the solution is allowed to stand for 2 days. A white solid is collected, washed with isopropanol, and allowed to air-dry for two days. After 2 days in vacuum oven at 45°C, R-5-hydroxy-3-[(4-methylpiperazino)-methyl]-2,3dihydro-2,2,4,6,7-pentamethyl-1-benzofuran dihydrochloride hydrate (9.0 g, 68%) is collected. [a] 2 0 (water) +20.9.

o C o CC 6 a 0 C Co..

a 0C C Co4C C o oa o 00CC ooa o a o o uD o o 1C )000 CC C C C *9 1 09 The compounds of the parent application are free radical scavengers as 10 disclosed in Patent Application W093/20057, filed March 10, 1993 and U.S.

counterpart 5,721,233 filed December 22, 1994. Free radical reactions have been implicated in the pathology of more than 50 human diseases. Radicals and other reactive oxygen species are formed constantly in the human body both by deliberate synthesis by activated phagocytes) and by chemical 15 side-reactions. They are removed by enzymic and non enzymic antioxidant defense systems. Oxidative stress, occurring when anti-oxidant defenses are inadequate, can damage lipids, proteins, carbohydrates and DNA. A few clinical conditions are caused by oxidative stress, but more often the stress results from the disease and can make a significant contribution to the disease 20 pathology. For a more detailed review see B. Halliwell in Drugs, 1991, 42, 569- 605.

There is a growing body of information that suggests a pathophysiologic role of oxygen free-radical-mediated lipid peroxidation following central nervous 25 system trauma or stroke, either ischemic or hemorrhagic. A reduction in C.\IANWRDUEtnMMsPECNXMSSS0DOC

I

ki 1t 41r..

S01 i IJ~i u.L-dcej ana evaporated to aryness.

Purification by flash chromatography eluting with heptane/ethyl acetate 80:20 to 50:50 gave 8.05 g of 5-hydroxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3- :4 -9 i:

I

s..

3% ;6 46 cerebral tissue concentration of endogenous antioxidants has been observed, as well as an increase in lipid peroxidation products. Inhibitors of brain lipid peroxidation counteract and reduce cerebral tissue damage, as well as to prolong life of traumatized animals. These findings have been reviewed by E.D. Hall and J.M. Braughler in Free Radical Biologyand Medicine, 1989, 6, 303-313 and elsewhere. M. Miyamoto et al., Pharmacc' Exp. Ther., 1989, 250, 1132) report that neurotoxicity due to excessive glutamic acid release is similarly reduced by antioxidants.

They suggest the use of agents that inhibit brain lipid peroxidation for treatment of neurodegenerative diseases such as Huntington's and Alzheimer's disease in which excessive glutamic acid release has been observed. M.R.

Hori et al., (Chem.Pharm. Bull. 1991, 39, 367) report on anti-amnesic activity of brain lipid peroxidation inhibitors in rats.

The role of oxygen free radicals in Parkinson's disease has been reviewed recently (Fre Radical Biol. Med., 1991, 161-169) and a free radical scavenger has been tested clinically with some success (Fundam.Clin.Pharmacol. 1988,2,1- 12).

Ischemia followed by reperfusion causes formation of oxygen-derived free radicals and increased lipid peroxidation and results in tissue injury. Administration of free radical scavengers to animals subjected to ischemia/reperfusion reduces these effects in heart, lung, kidney, pancreas, brain and other tissues.

The process of inflammation is also known to involve the release of superoxide radicals from phagocytic cells which cause some of the symptoms of rheumatoid arthritis and other inflammatory diseases such as ulcerative collitis. Free radicals scavengers, such as the compounds of this invention, are also useful in treatment of these diseases.

it i A 47 Smoke inhalation leads to lung injury due to an increase in pulmonary microvasculature and pulmonary edema.

This process is accompanied by increased lipid peroxidation in lung tissue. An inhibitor of lipid peroxidation was shown to reduce these symptoms in animals subjected to hot sawdust smoke by Z. Min et al., (J.Med.Cell. PLA, 1990, 176-80). They suggest the use of antioxidants in treatment of smoke inhalation-lung injury, adult respiratory distress syndrome and emphysema.

j Reactive oxygen species also play a role in the formation of foam cells in artherosclerotic plaques (reviewed by D. Steinberg et al., NewEngl.J. Med., 1989, 320, 915-924) and the free radical scavenger probucol has a i5 marked antiartherosclerotic effect in hyperlipidemic rabbits (Carew et al., Proc. Natl. Acad. Sci. Usa, 1987, 84 4iS 7725-7729). Degenerative retinal damage and diabetogenic retinopathy have also been listed as target for treatment with free radical scavengers (cf. J.W. Baynes, Diabetes,1991, 20 40,405-412; S.P. Wolff etal., Free Rad. Bid. Med., 1991, 10, 339-352).

The compounds may be also useful in the treatments of cancers, and degenerative diseases related to aging, stroke, and head trauma, since oxygen-derived free radicals have been identified among causative factors for reviews, see B. Halliwell and C. Gutteridge, Biochem.J., 1984, 219, 1-14; TINS 1985, 22-6. Antioxidants have also been shown to be useful in the treatment of cataracts, FreeRad.Biol. Med., 12:251-261 (1992).

SIn vitro and invivo activity for the compounds of this invention may be determined by the use of standard assays which demonstrate the free radical scavenginq property, S- affinity for cardiac tissue and cardioprotective properties, as well as by comparison with agents known to be effective for these purposes. Exemplary of the assay useful for determining the free-radical scavenging property 48 of the compounds of this invention is by the in vitro inhibition of lipid peroxidation in rat brain homogenates.

The free radical scavenging properties of the compounds may readily be evaluated using standard and recognized procedures utilized in the art. For example the free radical scavenging property may be evaluated by an assay wherein superoxide radicals are generated by 4 mU of xanthine oxidase in the presence of 0.1 mM xanthine and detected by reduction of 40 pm nitro blue tetrazolium (NBT) to the diformazan dye in a spectrophotometric assay as described by C. Beauchamp and I. Fridovick, (Analyt.Biochem.

1971, 44, 276-287). The 30 U of superoxide dismutase inhibited this reduction by 90% which is due to superoxide H 15 radicals. In the presence of a superoxide scavenger (test compound) there is a competition for the superoxide radical and thus a reduction in the color formation of NBT demonstrates the superoxide radical scavenging property of the test compound.

Inhibiting the process of lipid peroxidation may be assayed using tissue homogeneates for measuring the antioxidant activity of biological fluids by the methodology of J. Stocks et al., (Clin.Sci. Mol. Med.,1974,47, 215-222), wherein a brain tissue homogeneate of treated adult Sprague Dawley rats is utilized.

Samples of total volume 1 mL of diluted brain homogenate and with the scavenger at an appropriate dilution are incubated. Non-incubated samples are taken as back-ground. Controls are run without scavenger and a sample containing only buffer is taken as blank. After incubation at 37 0 C for 30 min, 200 pL of 35% perchloric acid Sis added, the samples centrifuged and 800 pL of the i 5 supernatants mixed with 200 pL of thiobarbituric acid reactive material is developed at 100 0 C in a boiling water bath for 15 min, and absorbance read at 532 nm.

it I" r I~ i -1; 49 9 9 9 9**ea* 9~ *994 a ea.

9 p a a if a.

a *i ;;i fs~ For exvivo inhibition of tissue including heart or brain tissue, lipid peroxidation in mice may be utilized to demonstrate the ability of the compounds to penetrate and act as free radical scavengers in the brain. This assay involves pretreatment of male CDI mice by subcutaneous administration of the test compound. One hour later the brains are excised, homogenized 1+9 in 20 mM potassium phosphate buffer at pH 7.3 (0.24 M KC1) and incubated at 1/100 concentration in 1 mL of buffer at 37°C for 30-120 min. At the end of the 'ncubation 200 pL of perchloric acid is added and proteins removed by centrifugation. To 800 mL of the supernatant are added 200 pL of 1% TBA and the samples are treated to 100 0 C for 15 15 min. The TBA-adduct is extracted into 2 times 1 mL of n-butanol. The fluorescence is measured at an excitation wavelength of 515 nm and an emission wavelength of 553 nm against a standard prepared from malondialdehyde dimethylacetal.

Stimulated human leukocytes release radicals and other oxygen metabolites, which, during inflammation, act as a mict~bidl agents. At the same time, they release proteolytic enzymes, such as elastase, which are also microbicidal but potentially threaten the connective tissue of the hcst. An endogenous al-proteinase inhibitor (a 1 Pi) normally protects the host tissue from protelytic digestion. The axPi is however, inactivated by the leukocyte-derived oxidants. Antagonism of aiPi is an indication of the disclosed radical scavengers. The concentration needed to protect 50% of the elastase inhibitory capacity of aiPi (PCs0) depends on the amount of stimulated leukocytes present.

Method: The procedure described by Skosey and Chow was followed (see J.L. Skosey and D.C. Chow inHandbookof Methods for OxygenRadica Research (Greenwald, ed.) 1985, 413-416,

I

iii~: :li

I~

i I:Ii" il; I;ilI,.

i i ii Fi 4 i: ,0 e As A 9 50 CRC Press, Boca Raton). In short, human aiPi was incubated with zymosan-stimulated human peripheral-blood leukocytes in the presence or the absence of the scavengers. The amount of aiPi protected from oxidative inactivation was determined by its residual elastase inhibitory capacity.

The relevance to inflammation matter has been reviewed by Weiss (see S.J. Weiss, N. England J. Med. 1989, 320, 365-376).

Lung emphysema is associated with a genetic defect in aiPi; the disease is further enhanced by oxidants inhaled during cigarette smoking, which leads to oxidative inactivation of alPi in the lung tissue (see J. Travis and G.S. Salvesen, Annu. Rev. Biochem., 1983, 52, 655-709). Oxidized alPi has also been isolated from rheumatoid synovial fluid (see P.S.

Wong and J. Travis, Biochem.BiophysRoc. Commun. 1980, 06, 1440-1454). The degradation of hyaluronic acid, a macromolecule accounting for the viscosity of synovial fluid, is triggered by superoxyl radicals released from human leukocytes invitro (see R.A. Greenwald and S.A. Moak, 20 Inflammation, 1986, 10, 15-30). Furthermore, nonsteroidal anti-inflamatory drugs were shown to inhibit the release of superoxyl radicals from leukocytes (see H. Strom and I. Ahnfelt-Ronne, Agents andActions, 1989, 26, 235-237 and M. Roch-Arveiller, V. Revelant, D. Pharm Huy, L. Maman, J. Fontagne J.R.J. Sorenson and J.P. Giroud, Agentsand Actions, 1990, 31, 65-71), and 5-aminosalicylic acid may exert its therapeutic activity in inflammatory bowel disease by radical scavenger mechanism (see I. Ahnfelt- Ronne, O.H. Nielsen, A. Christensen, E. Langholz, V. Binder and P. Riis, Gastroenterology, 1990, 98, 1162-1169).

Therefore, it is believed that the compounds of this invention may be useful in the mentioned pathologic situations and that inflammatory bowel disease may be a special target. An immune stimulatory effect of antioxidants has also been reported in that they enhanced lymphocyte activity Anderson and P.T. Lukey, Ann.N.Y.

Acad.Sci., 1987,498, 229-247) invitro in the presence of 'y i; :1 j 3~ 1 i n- i-- 1 0000.

a I r r 51 triggered leukocytes, and exvivo after pretreatment of human.

volunteers.

Thus, using standard and well known methodology, as well as by comparison with known compounds found useful, it is to be found that the compounds are free radical scavengers useful in the prevention and treatment of such disease states related to neurotoxicity due to excessive glutamine release, to Huntington's disease, Alzheimer's disease and other cognitive dysfunctions, memory, learning and attention deficits), amnesia, and Parkinson's disease, as well as treatment and prevention of tissue damage in heart, lung, kidney, pancreas and brain tissues induced by ischemia/reperfusion, and to allay acute blood loss due to hemorrhagic shock.

The compounds of the parent application are of particular interest in treating patients with stroke, nervous system trauma, and reperfusion damage. As used herein, these terms have the following m-anings: a) stroke means cerebrovascular disease which includes cerebral insufficiency due to transient disturbances of blood flow, infarction, and arteriovenous malformation which causes symptoms of mass lesion, infarction or hemorrhage.

b) nervous system trauma means injury to the head or spine. For example, injury can occur from skull or spine penetration or from rapid brain acceleration or deceleration which injures tissue at the point of impact, at its opposite pole or within the frontal or temporal lobes. Injury may consist of nerve tissue, blood vessels and /or meninges damage resulting in neural disruption, ischemia and/or edema.; and I f 4i- 'i iT ii' ;li: n; i :~4 ii"~i~ 52

:I

ao rar n r o o r~ o i D ou e o o r r i*: o r os i:: j6ii r r i~r r i:

I

i i?

_J

:,z c) reperfusion damage means the damage that occurs in any blood-deprived tissue, anywhere in the body, upon reintroduction of the blood supply. For example, reperfusion of an ischemic area of the myocardium or the cerebrum.

The compounds of the parent application can be utilized both prophylatically and therapeutically. The amount of active ingredient for therapeutic administration can vary over a wide rance and is dependent upon such factors as the species of mammal to be treated, its age, health, sex, weight, natulre and severity of the condition being treated.

The term "patient" refers to a warm-blooded animal such 15 as, for example, rats, mice, dogs, cats, guinea pigs, primates and humans. Generally, a therapeutically effective amount of the active ingredient to be administered will range from about 0.1 mg/kg to 30 mg/kg of body weight per day. For prophylatic administration, 20 corresponding lower doses can be utilized. Preferably, the compounds of the present invention will be administered to the patient in combination with a pharmaceutically acceptable carrier which is any substance which aids in the administration of the compound without substantially 25 affecting its therapeutic properties.

Most preferably, the compounds are administered intravenously particularly under crisis situations wherein it is essential that the therapeutic agent be gotten to its site of action as quickly as possible, such as in those emergency conditions caused by coronary infarction, stroke and surgical interventions, conditions which can cause severe reperfusion damage.

The compounds of this invention can be orally administered, preferably using more active ingredient per day than when parenterally administered, preferably taking d u..Ly resiaue is pourea into nexanre i u J (800 mL). White crystals are formed. The solids are scraped from the side and swirled to increase the crystallization. The solid is collected and air dried to give benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3rt 2W C J

I

earlP- ifc-. J t T T O T O M3 t roaecasi^^ 1 53 o divided doses 3 to 4 times per day. Preferably, enteral administration in post "crisis" situations, particularly after being released from hospitalized conditions. The compounds can be used in standard dosage unit forms such as tablets, capsules, dragees, lozenges, elixirs, emulsions, suspensions, and in cases wherein topical application is preferred by suppository or sub-lingual administration.

Tablets and capsules containing from 100 mg to 400 mg of active ingredient are preferred modes of enteral administration. Of course, in the trea ment of inflammation the preferred method of administration is by depot injection directly to the situs of the inflammation area with follow-up enteral means of administration.

In preparing solid dose forms such as tablets, the active ingredient is generally blended with conventional pnarmaceutical carriers or excipients such as gelatin, various starches, lactose, calcium phosphate or powdered sugar. The term rharmaceutical carrier as used herein also includes lubricants employed to improve the flow of tablet grannulations and which prevent adhesion of tablet material to the surfaces of tablet dies and punches. Suitable lubricants include for example, talc staric acid, calcium stearate, magnesium stearate and zinc stearate. Also included within the definition of a pharmaceutical carrier 25 as used herein, are disintegrating agents added to assist the breakup and dissolution of tablets following administration, as well as coloring and/or flavoring agents to enhance the aesthetic qualities of the tablets and make them more acceptable to the patient.

Suitable liquid excipients for the preparation of liquid dosage unit forms include water and alcohols such as ethanol, benzyl alcohol and the polyethylene glycols, either with or without the addition of a surfactant. In general, the preferred liquid excipients, particularly for injectable preparations, include water, physiological and saline solutions, dextrose and glycol solutions such as an

I

*55555 5* 0 3 r r q t dr" j 54 aqueous propylene glycol or polyethylene glycol solutions.

In order to minimize or eliminate irritation at the site of injection, such compositions ray contain a non-ionic surfactant having a hydrophile-lipophile balance (HLB) of from about 12 to about 17. The quantity of surfactant in such formulations ranges from about 5 to 15% by weight.

The surfa6tant can be single component having the aboveidentified HLB, or a mixture of two or more components having the desired HLB. Illustrative of surfactants useful in parenteral formulations are the class of polyoxyethylene Ssorbitan fatty acid esters as, for example, sorbitan monooleate and the high molecular weights adducts of ethylene oxide with a hydrophobic base, formed by the condensation of propylene oxide with propylene glycol. In 1 certain topical and parenteral preparations, various oils can be utilized as carrier or excipients. Illustrative of such oils are minerals oils, glyceride oils such as lard oil, cod liver oil, peanut oil, sesame oil, corn oil and soybean oil. For insoluble compounds, suspending agents S' 20 may be added as well as agents to control the viscosity, as •2 for example, magnesium aluminum silicate or carboxymethylcellulose. In addition to these excipients, buffers, S: preservatives and emulsifying agents may be also be added.

Typical enema preparation of the retention type enema utilize small volumes, generally much less than about 150 mL for an adult, typically volumes of only a few millilite:s are preferred. Excipients and solvents for use in retention anemas should, of course, be selected so s to avoid colonic irritation and should be also selected so as to minimize absorption of the various agents.

The compounds of the parent invention can also be administered topically. This can be accomplished by simply preparing a solution of the compound to be administered, preferably using a solvent known to promote transdermal absorption such as ethanol or dimethyl sulfoxide (DMSO) with or without other excipients. preferably topical of cold hexane is added and the R-5-bnnzyloxy-3hydroxymethyl-2,2,4,6,7-pentamethyl-2,3-dihydro-lbenzofuran is collected and dried to give 10.9 g (27%) fite solid 98.4%, [a] 2 0 7.9 in methanol).

f g 3 eq

I.

h i.

I

administration will be accomplished using a patch either of the reservoir and porous membrane type or of a solid matrix variety.

Some suitable transdermal devices are described in U.S.

Pat. Nos. 3,742,951, 3,797,494 3,996,934, and 4,031,894.

These devices generally contain a backing member which defines one of its face surfaces, an active agent permeable adhesive layer defining the other face surface and at least one reservoir containing the active agent interposed between the face surfaces. Alternatively, the active agent may be contained in a plurality of microcapsules distributed throughout the permeable adhesive layer. In either case, the active agent is delivered continuously from the reservoir or r.icrocapsules through a membrane into the active agent permeable adhesive, which is in a contact with the skin or mucosa of the recipient. If the active agent is absorbed through the skin, a controlled and predetermined flow of the active agent is administrated to the recipient. In the case of microcapsules, the encapsuling agent may also function as the membrane.

In another device for transdermallly administering the compounds in accordance with the parent invention, the pharmaceutically active compound is contained in a matrix from which it is delivered in the desired gradual, constant and controlled rate. The matrix is permeable to the releLse of the compound through diffusion or microporous flow. The release is rate controlling. Such a system, which requires no membrane is described in U.S. Pat.

No. 3,921,636. At least two types of release are possible in these systems. Release by diffusion occurs when the matrix is non-porous. The pharmaceutically effective compound dissolves in and diffuses through the matrix itself. Release by microporous flow occurs when the pharmaceutically effective compound is transported through a liquid phase in the pores of the matrix.

i Ir a, ai f'l"l :i i

A

56 The compounds of the parent application may be incorporated into an aerosol preparation by means commonly known to those skilled in the art. The aerosol preparation may be prepared for use as a topical aerosol or may be prepared for inhalation. The aerosol preparation may be in the form of a solution or a suspension and may contain other ingredients such as solvents, propellants and/or dispersing agents. Typical examples of aerosol preparations are shown in Remington's Pharmaceutical Sciences, 18th ed., Mack Publishing Company, Easton Pennsylvania, pp. 1694-1712 (1990) incorporated herein by reference.

As it is true for most classes of compounds suitable for use as therapeutic agents certain subclasses and certain specific compounds are more preferred than others. In this instance it is preferred that the R 2 R4, R 6 and R 7 moieties be methyl. Preferably R 5 is H or an acyi moiety including formyl and acetyl. X is preferably CH2A. A is preferably -N N-Rio ;Y s preferably H: Rio is preferably C1-6 alkyl, more

Y

preferably Cj.3 alkyl and more preferably methyl. Other preferred forms of Rio are acyloxyalkylene, especially -CH 2

-O-C(O)CH

3 hydroxyalkyl (C2 especially

-(CH

2 2 -OH, and pyrimidinyl Throughout the description and claims nf the specification the word "comprise" and variations of the word, such as "comprising" and "comprises" is not intended to exclude other additives, components, integers or steps.

i

Claims (7)

1. A process for resolving into its optically active isomers compound of formula (9) R"4 POH RR"7 (9) wherein R"2, R"4, R"6 and R"7 are methyl, Pg is hydrogen or is a suitable protecting 10 group, comprising the steps of: reacting compound of formula with lipase/ candida cylindracea arid vinyl acetate, separating the individual isomers, 15 optionally deprotecting the 5-hydroxy group.

2. A process according to claim 1 wherein said reaction of compound (9) with lipase/candida cylindracea and vinyl acetate is carried out i ether.

3. A process according to claim 2 wherein said ether is t-butyl-methyl ether.

4. A process according to any one of the preceding claims wherein said reaction is carried out at a temperature of from 0°C to The compound 5-hydroxy-3-methylene-2,2,4,6,7-pentamethyl-2,3- dihydro-benzofuran.

6. The compound 5-benzyloxy-3-hydroxymethyl-2,2,4,6,7-pentamethyl-2,3- dihydro-1-benzofuran.

7. The compound 3-hydroxymethyl-5-methoxy-2,2,4,6,7-pentamethyl-2,3- dihydro-1-benzofuran. WI hiORD' NYM\ SECN 575.0 C-'VAwRDENc NYMPECN 1\85575DI.DC00 -58

8. A process according to claim 1 substantially as hereinbefore described with reference to any of the examples. DATED: 22 September, 1998 PHILLIPS ORMONDE FITZPATRICK Atnso HOCS MAIO:RUSEL.IC a- I V A 448 41Apoesacrigt li ussatal shribfr ecie WLI.LULI UIi aL i LLA t-L t- affinity for cardiac tissue and cardioprotective properties, as well as by comparison with agents known to be effective for these purposes. Exemplary of the assay useful for determining the free-radical scavenging property *I i~ ur~trr=lrm~auurr ux~~ r; p li ABSTRACT A process for resolving into its optically active isomers compound of formula (9) o 0 0 S wherein R" 2 R" 4 R" 6 and R" 7 are methyl, Pg is hydrogen or is a suitable protecting group, comprising the steps of: reacting compound of formula with lipase/ candida cylindracea and vinyl acetate, separating the individual isomers, 15 optionally deprotecting the 5-hydroxy group. CMwtiOINNVNSP NWas75OiLDOC P i: d i~ Y.