Patent Application: US-58532996-A

Abstract:
an oxazolyl enaminone is provided having the formula : ## str1 ## wherein r 1 is selected from the group consisting of a branched or unbranched alkyl groups containing from 1 to 4 carbon atoms and r 2 is selected from the group consisting of a branched or unbranched alkyl group containing from 1 to 4 carbon atoms , a chloro group , a trifluoromethyl group and a trifluoromethoxy group . also provided are pharmaceutically accepatable salts and isomers of the above , and pharmaceutical compositions containing the same .

Description:
isoxazole enaminones are synthesized according to the following scheme . isoxazoles are condensed with the respective β - hydroxy keto esters , 1a - c , as previously reported [ 1a , 1b , 1f , 9 ]. prerequisite β - hydroxy keto esters , 1a - c , are synthesized via a condensation of ethyl crotonate with the respective acetoacetic esters as reported by friary and coworkers for the synthesis of the 4 - carbo - tert - butoxy - 5 - methylcyclohexane - 1 , 3 - dione , 1c [ 10 ], and recently reported by scott et al . [ 1f ]. the subsequent condensation reaction of the β - hydroxy keto esters , 1a - c , with 3 - amino - 5 - methyl - isoxazole , 2 , proceeds smoothly to provide easily recrystallizable products , 3 a - c . ## str5 ## where a , r = tert - bu ; b , r = et ; c , r = me the starting isoxazole , 2 , employed in the current study has a hetero ring like that of muscimol , 5 . however , it should be noted that in the current series , a lipophilic methyl group replaces the highly ionic and hydrophilic 2 - moiety , while an amino group replaces the hydroxyl moiety . the compounds of the present invention can be readily provided in the form of pharmaceutically acceptable salts . suitable pharmaceutically acceptable salts include salts derived from inorganic or organic acids . typical examples of pharmaceutically acceptable salts include salts derived from inorganic acids such as hydrochloric acid , hydrobromic acid , sulfuric acid , phosphoric acid , nitric acid and the like , and organic acids such as acetic acid , propionic acid , glycolic acid , oxalic acid , pyruvic acid , malic acid , succinic acid , malonic acid , maleic acid , fumaric acid , citric acid , tartaric acid , mandelic acid , cinnamic acid , benzoic acid , p - toluenesulfonic acid , salicylic acid , ethane sulfonic acid , methanesulfonic acid and so forth . the formation of such salts is well within the abilities of those of ordinary skill in the art . for each utility and indication , the amount of ingredient required will depend upon a number of factors including the severity of the condition to be treated and the identity of the recipient , and will ultimately be at the discretion of the attendant physician or veterinarian . in general however , for each of these utilities and indications , a suitable effective dose will preferably be in the range 0 . 1 to 250 mg per kilogram bodyweight of recipient per day , more preferably in the range 0 . 1 to 10 mg per kilogram bodyweight per day . while it is possible for the active ingredients to be administered alone it is preferable to present them as pharmaceutical formulations . the formulations , both for veterinary and for human use , of the present invention comprise at least one active ingredient , as defined above , together with one or more acceptable carriers thereof and optionally other therapeutic ingredients . the carrier ( s ) must be &# 34 ; acceptable &# 34 ; in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof . the formulations include those suitable for oral , rectal , nasal , topical ( including buccal and sublingual ), vaginal or parenteral ( including subcutaneous , intramuscular , intravenous , intradermal , intrathecal and epidural ) administration . the formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy . such methods include a step of bringing into association the active ingredient with the carrier which constitutes one or more accessory ingredients . in general , the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both , and then , if necessary , shaping the product into tablet form , for example . formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules , cachets or tablets each containing a predetermined amount of the active ingredient ; as a powder or granules ; as a solution or a suspension in an aqueous liquid or a non - aqueous liquid ; or as a bolus , electuary or paste . a tablet may be made by compression or molding , optionally with one or more pharmaceutically acceptable excipients . compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free - flowing form such as a powder or granules , optionally mixed with a binder , lubricant , inert diluent , preservative , surface active or dispersing agent . molded tablets may be made by molding a mixture of the powdered compound moistened with an inert liquid diluent in a suitable machine . the tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein . for topical applications , the formulations are preferably applied as an ointment or cream . when formulated in an ointment , the active ingredients may be employed with either a paraffinic or water - miscible ointment base . alternatively , the active ingredients may be formulated in a cream with an oil - in - water cream base . if desired , the aqueous phase of the cream base may include , for example , at least 30 % w / w of a polyhydric alcohol , i . e ., an alcohol having two or more hydroxyl groups such as propylene glycol , butane 1 , 3 - diol , mannitol , sorbitol , glycerol and polyethylene glycol and mixtures thereof . the topical formulations may desirably include a compound which enhances absorption or penetration of the active ingredient through the skin or other affected areas . examples of such dermal penetration enhancers include dimethylsulfoxide and related analogues . formulations suitable for topical administration to the eye also include eye drops wherein the active ingredient is dissolved ( or suspended ) in a suitable carrier , especially in aqueous solvent for the active ingredient . formulations suitable for topical administration in the mouth include lozenges containing the active ingredient , preferably in a flavored base , usually sucrose and acacia or tragacanth ; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin , or sucrose and acacia ; and mouthwashes comprising the active ingredient in a suitable liquid carrier . formulations for rectal administration may be presented as a suppository with a suitable base comprising , for example , cocoa butter or a stearate . formulations suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size , for example , in the range of from about 20 to about 500 microns , which is administered by rapid inhalation through the nasal passage . suitable formulations wherein the carrier is a liquid , for administration as , for example , a nasal spray or as nasal drops , include aqueous or oily solutions of the active ingredient . formulations suitable for vaginal administration may be presented as pessaries , tampons , creams , gels , pastes , foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate . formulations suitable for parenteral administration include aqueous and non - aqueous sterile injection solutions which may contain anti - oxidants , buffers , bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient ; and aqueous and non - aqueous sterile suspensions which may include suspending agents and thickening agents . the formulations may be presented in unit - dose or multi - dose containers , for example , sealed ampoules and vials , and may be stored in a freeze - dried ( lyophilized ) condition requiring only the addition of the sterile liquid carrier , for example , water for injection , immediately prior to use . extemporaneous injection solutions and suspensions may be prepared from sterile powders , granules and tablets of the kind previously described . formulations for intramuscular administration are particularly preferred . preferred unit dosage formulations are those containing a daily dose or daily sub - dose , as herein above recited , or an appropriate fraction thereof , of an active ingredient . it should be understood that , in addition to the ingredients particularly mentioned above , the formulations of this invention may include other agents conventional in the art having regard to the type of formulation in question . for example , those suitable for oral administration may include flavoring agents . the present invention further provides veterinary compositions containing at least one active ingredient as above defined together with a veterinary carrier thereof . veterinary carriers are materials useful for the purpose of administering the composition and may be solid , liquid or gaseous materials useful for the purpose of administering the composition and are otherwise inert or acceptable in the veterinary art and are compatible with the active ingredient . these veterinary compositions may be administered orally , parenterally or by any other desired route . for oral administration , the compositions can be in the form of a tablet , granule drench , paste , cachet , capsule or feed supplement . granules may be made by the well known techniques of wet granulation , precompression or slugging . they can be administered to animals via an inert liquid vehicle so as to form a drench , or in a suspension with water or oil base . preferably , further accessory ingredients , such as a dispensing agent , are included . additional formulation information can be found , for example , in u . s . pat . no . 5 , 079 , 252 . cyclic enaminone esters of the isoxazole series are shown in table i . high field ( 1 h and 13 c ) nuclear magnetic resonance ( nmr ) analyses on the reported compounds were consistent with the assigned structures . the 1 h nmr studies on these compounds proved interesting and unique and differed significantly in the vinyl proton signal from methyl 4 -[( p - chlorophenyl ) amino ]- 6 - methyl - 2 - oxocyclohex - 3 - en - 1 - oate , ( add 196022 ) [ 1c , 1d ], 4 ( table ii ), the prototypic anticonvulsant in the series . whereas the vinyl proton of 4 occurred at δ5 . 48 ppm , the isoxazolyl enaminones herein reported occurred between about δ 6 . 01 - 6 . 02 for the c 3 and about δ6 . 23 - 6 . 25 ppm for the c 4 isoxazolyl protons , respectively . the unequivocal assignment of these protons was provided by 1d noe analysis of 3b . on irradiating the c 5 methyl on the isoxazole ring , only the c 4 , proton should show a noe , which in fact occurred . the 1d noe spectrum showed a methyl singlet at about δ2 . 25 ppm and a respective proton noe at about δ6 . 02 ppm . in compound 8 , methyl 4 &# 39 ;-[( 4 &# 39 ;- chloro - 2 &# 39 ;- pyridinyl ) amino ]- 6 - methyl - 2 - oxocyclohex - 3 - en - 1 - oate [ 1b , 1d ], the vinyl proton appeared at δ6 . 82 ppm and indicated a deshielding effect , indicative of hydrogen bonding . this was further supported by the acidic nh proton of 8 ( δ9 . 57 ppm , saturated solution ), being more deshielded than 4 ( δ7 . 96 ppm , saturated solution ) [ 1a ]. ## str6 ## table i__________________________________________________________________________ . sup . 13 c correlation chart for compounds 3 and 4 . 3a 3b 3c 4__________________________________________________________________________isoxazole ( c . sub . 4 &# 39 ;) ch . sub . 3 11 . 83 11 . 73 11 . 78 -- me ( c . sub . 6 ) ch . sub . 3 19 . 11 19 . 07 19 . 20 19 . 27c . sub . 6 ch 31 . 82 31 . 62 31 . 64 31 . 65c . sub . 5 ch . sub . 2 34 . 71 34 . 60 34 . 60 35 . 07c . sub . 1 ch 60 . 66 59 . 74 59 . 74 60 . 00 ch 95 . 80 95 . 71 95 . 77 96 . 79 ( 94 . 64 ) ch 104 . 25 103 . 98 103 . 95 c 157 . 15 157 . 31 157 . 45 c 159 . 54 159 . 42 159 . 47 161 . 33 * c 168 . 75 168 . 67 168 . 78ketone ( c . sub . 2 ) c 169 . 77 170 . 39 170 . 97 171 . 16ester co 193 . 13 192 . 70 192 . 70 191 . 11 roco 27 . 77 ( q ), 14 . 04 ( q ), 51 . 43 51 . 35 80 . 02 ( s ) 59 . 90 ( t ) 124 . 64 ( d ) 128 . 46 ( q ) 129 . 14 ( d ) 137 . 65 ( q ) __________________________________________________________________________ table ii__________________________________________________________________________ . sup . 1 h correlation chart for compounds 3c and 4 . ## str7 ## compound a b c d e f g__________________________________________________________________________4 1 . 08 2 . 22 - 2 . 98 3 . 75 5 . 48 6 . 86 7 . 20 - 7 . 50 -- ( d , 3h ) ( m , 3h ) ( s , 3h ) ( s , 1h ) ( bs , 1h ) ( m , 5h ) 3 . 08 ( d , j = 11 . 00 hz ) 3c 0 . 98 2 . 38 - 2 . 48 3 . 64 6 . 02 9 . 64 6 . 24 2 . 36 ( d , 3h ) ( m , 3h ) ( s , 1h ) ( s , 1h ) ( bs , 1h ) ( s , 1h ) ( s , 3h ) 3 . 14 ( d , 1h ) __________________________________________________________________________ preliminary pharmacological testing of the compounds listed in table i have been provided using procedures that have been described [ 11 ]. there are three initial tests : maximal electroshock seizure ( mes ); subcutaneous pentylenetetrazol ( scmet ); and neurologic toxicity ( tox ) in mice . intraperitoneal administration of the test compounds was as a suspension in 0 . 5 % methylcellulose . these phase i data are shown in table iii . as previously reported [ 1b ], due to species specificity , all class 1 and 2 active enaminone analogs ( activity at 100 mg / kg or less and activity at doses greater than 100 mg / kg but less than 300 mg / kg , respectively ), are subsequently evaluated for oral ( po ) activity ( phases via and vib ) in the rat , which has been previously shown to be more sensitive to these compounds . as noted from table iii , each of the compounds is a class 1 analog , all displaying activity in the mes intraperitoneal evaluation at 100 mg / kg at 30 min , and 3c is also active at that dose in the 4 h evaluation . the most potent analog , however , is 3b , which is active in the mes evaluation at 30 mg / kg . 3c is also active in the scmet evaluation at 30 mg / kg , at 30 min , however at 300 mg / kg , the animal exhibits continuous seizure activity . it should be noted that 3c is the only compound shown to cause motor impairment at 300 mg / kg , either at 30 min or 4 h , while none of these compounds exhibits toxicity at the lower dosages . thus , compounds 3b and 3c were further evaluated in a phase via study to ascertain their po activity in rats . table iv details this analysis and reveals that these compounds are also active in rats at 30 mg / kg while displaying minimal motor impairment over the 4 h evaluation . compound 3b , due to its greater overall potency in both phase i and phase via evaluations , was further evaluated in phase vib to quantitate its activity and toxicity . additionally , a special phase ii intraperitoneal rat evaluation of each of these analogs was performed . this data is shown in table v and compared to compound 4 , which was previously reported [ 1b ]. the mes evaluations , at 10 mg / kg were followed at 15 , and 30 min , and at 1 , 2 , 4 and 6 h , while the toxicity evaluations , at 100 mg / kg ( except for 3a , which was evaluated at 56 mg / kg ), were observed at the six time periods previously indicated and , in addition , at 8 and at 24 h . compound 3b displays activity at 15 min , and at 1 h , while 3c also shows activity at 15 min , as well as at 1 and 2 h . compound 3a shows no protection at 10 mg / kg . the toxicity evaluation indicates that none of the isoxazoles exhibits motor impairment throughout the test period . further , none of the animals exhibits toxicity after 24 h . table iii__________________________________________________________________________anticonvulsant screening project ( asp ) - phase i test results dose , scmet ,. sup . a scmet ,. sup . a mes ,. sup . b mes ,. sup . b tox ,. sup . c tox ,. sup . c aspcompound mg / kg 30 min 4 h 30 min 4 h 30 min 4 h classification . sup . d__________________________________________________________________________3a 100 0 / 1 0 / 1 2 / 3 0 / 3 0 / 8 0 / 4 1 300 0 / 1 0 / 1 1 / 1 1 / 1 0 / 4 0 / 23b 30 0 / 1 0 / 1 1 / 1 0 / 1 0 / 4 0 / 2 1 100 0 / 1 0 / 1 2 / 3 0 / 3 0 / 8 0 / 4 300 0 / 1 0 / 1 1 / 1 0 / 1 0 / 4 0 / 23c 30 1 / 5 0 / 1 0 / 1 0 / 1 0 / 4 0 / 2 1 100 0 / 1 0 / 1 1 / 3 1 / 6 0 / 8 0 / 4 300 . sup . 0 / 1 . sup . e 0 / 1 1 / 1 0 / 4 1 / 4 0 / 2__________________________________________________________________________ . sup . a subcutaneous pentylenetetrazol test ( number of animals protected / number of animals tested ). . sup . b maximal electroshock test ( number of animals protected / number of animals tested ). . sup . c rotorod toxicity ( number of animals exhibiting toxicity / number of animals tested ). . sup . d classifications are : 1anticonvulsant activity at 100 mg / kg or less 2anticonvulsant activity at doses greater than 100 mg / kg ; 3no anticonvulsant activity at doses up to and including 300 mg / kg . . sup . e continuous seizure activity . table iv______________________________________phase via oral rat data for compounds 3b and 3c . dose , time , compound mg / kg h mes tox______________________________________3b 30 0 . 25 1 / 4 0 / 4 0 . 50 2 / 4 0 / 4 1 . 00 2 / 4 0 / 4 2 . 00 1 / 4 0 / 4 4 . 00 4 / 4 0 / 43c 30 0 . 25 2 / 4 0 / 4 0 . 50 2 / 4 0 / 4 1 . 00 1 / 4 0 / 4 2 . 00 3 / 4 0 / 4 4 . 00 1 / 4 0 / 4______________________________________ table v______________________________________phase ii intraperitoneal rat toxicity data of compounds 3a , 3b , 3c and 4 . mes tox10 mg / kg 100 mg / kgtime ( h ) 3a 3b 3c 4 3a . sup . c 3b 3c 4______________________________________0 . 25 0 / 4 3 / 4 1 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 0 / 80 . 50 0 / 4 0 / 4 0 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 3 / 81 . 00 0 / 4 0 / 4 1 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 6 / 82 . 00 0 / 4 0 / 4 1 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 7 / 84 . 00 0 / 4 0 / 4 0 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 4 / 86 . 00 0 / 4 0 / 4 0 / 4 nd . sup . d 0 / 8 0 / 8 0 / 8 1 / 88 . 00 nd . sup . d nd . sup . d nd . sup . d nd . sup . d 0 / 8 0 / 8 0 / 8 0 / 824 . 00 nd . sup . d nd . sup . d nd . sup . d nd . sup . d 0 / 8 0 / 8 0 / 8 . sup . 2 / 8 . sup . c______________________________________ . sup . c rotorod toxicity taken at 56 mg / kg . . sup . d nd = not determined . . sup . e two animals died . melting points were determined on a thomas - hoover capillary melting point apparatus and are uncorrected . ir spectra were recorded on samples in nujol , as diluted chloroform solutions in matched sodium chloride cells , or neat with a perkin - elmer 1330 spectrophotometer . 1 h nmr spectra were recorded on a general electric qe 300 - mhz spectrometer in deuterated solvents using tetramethylsilane as an internal reference . 13 c nmr spectra were recorded from the dmso - d 6 ( septet at δ39 . 50 ± 0 . 02 ppm ) on the same instrument in the broadband decoupled mode . signal multiplicities were separately determined by standard distortionless enhancement by polarization transfer ( dept ) [ 12 , 13 ] experiments . coupling patterns are described as follows : s , singlet ; bs , broad singlet ; d , doublet ; t , triplet ; q , quartet ; m , multiplet ; and 1h , 2h , 3h , etc . as the number of hydrogens integrated within a given coupling pattern . the chemical shifts and the coupling constants were rounded off to one decimal place . tlc analysis employed a n - buoh - hoac - h 2 o ( 5 : 4 : 1 ) or ( 5 : 1 : 4 , upper layer ) elution solvent mixture and 5 × 10 - cm or 5 × 20 - cm fluorescent plates ( whatman silica gel 60a ). elemental analyses ( c , h , and n ) were performed by schwarzkopf microanalytical laboratory , woodside , n . y . where analyses are indicated only by the symbols of the elements , analytical results were within 0 . 4 % of the theoretical values . 4 - carbo - tert - butoxy - 5 - methylcyclohexane - 1 , 3 - dione , 1a [ 10 ], 4 - carbo - ethoxy - 5 - methylcyclohexane - 1 , 3 - dione , 1b [ 9 ], and 4 - carbomethoxy - 5 - methylcyclohexane - 1 , 3 - dione , 1c [ 1a ], were prepared by literature methods as indicated . 3 - amino - 5 - methyl isoxazole , 2 , was obtained from fluka chemika ag ( germany ) and used without further purification . a mixture of 4 - carbo - tert - butoxy - 5 - methylcyclohexane - 1 , 3 - dione , 1a , ( 6 . 11 g , 27 mmole , mp 145 °- 146 ° c .) ( lit . 130 °- 131 . 5 ° c . 10 , c , h ) and 3 - amino - 5 - methylisoxazole , 2 , ( 3 . 24 g , 33 mmole ) were added to a mixture of absolute etoh ( 100 ml ) and etoac ( 100 ml ) and the solution refluxed and stirred for 6 h . evaporation under reduced pressure yielded a yellow solid which was recrystallized three times from etoac to yield 3 . 57 g ( 43 %) of tert - butyl 4 -[( 5 &# 39 ;- methyl )- 3 &# 39 ;- isoxazolylamino ]- 6 - methyl - 2 - oxocyclohex - 3 - en - 1 - oate , 3a , white crystals ; mp 197 °- 201 ° c . ( with effervescence ) ( c , h , n ); 1 h nmr ( dmso - d 6 ): δ1 . 00 ( d , 3h , j = 6 . 0 hz ), 1 . 42 ( s , 9h ), 2 . 35 ( s , 3h ), 2 . 44 ( m , 2h ), 2 . 51 ( m , 1h ), 2 . 93 ( d , 1h , j = 11 . 5 hz ), 6 . 01 ( s , 1h ), 6 . 23 ( s , 1h ), 9 . 58 ( bs , 1h ); 13 c ( dmso - d 6 ): δ11 . 83 ( q ), 19 . 11 ( q ), 27 . 77 ( q ), 31 . 82 ( d ), 34 . 71 ( t ), 60 . 66 ( d ), 80 . 02 ( s ), 95 . 80 ( d ), 104 . 25 ( d ), 157 . 11 ( s ), 159 . 54 ( s ), 168 . 75 ( s ), 169 . 77 ( s ), 193 . 13 ( s ). in a procedure like that for 3a , ethyl 4 -[( 5 &# 39 ;- methyl )- 3 &# 39 ;- isoxazolylamino ]- 6 - methyl - 2 - oxocyclohex - 3 - en - 1 - oate , 3b , was produced from 1b and 2 after three recrystallizations from etoh : yield 5 . 68 g ( 76 %) of white crystals ; mp 195 °- 197 ° c . ( c , h , n ); 1 h nmr ( dmso - d 6 ): δ1 . 00 ( d , 3h , j = 5 . 7 hz ), 1 . 20 ( t , 3h , j = 7 . 1 hz ), 2 . 35 ( s , 3h ), 2 . 38 - 2 . 55 ( m , 3h ), 3 . 09 ( d , 1h , j = 11 . hz ), 4 . 12 ( q , 2h , j = 7 . 1 hz ), 6 . 02 ( s , 1h ), 6 . 25 ( s , 1h ), 9 . 63 ( bs , 1h ); 13 c nmr ( dmso - d 6 ): δ11 . 73 ( q ), 14 . 04 ( q ), 19 . 07 ( q ), 31 . 62 ( d ), 34 . 60 ( t ), 59 . 74 ( d ), 59 . 90 ( t ), 95 . 71 ( d ), 103 . 98 ( d ), 157 . 31 ( s ), 159 . 42 ( s ), 168 . 67 ( s ), 170 . 39 ( s ), 192 . 70 ( s ). 1c and 2 were used to produce methyl 4 -[( 5 &# 39 ;- methyl )- 3 &# 39 ;- isoxazolylamino ]- 6 - methyl - 2 - oxocyclohex - 3 - en - 1 - oate 3c in a procedure like that for 3a : yield 4 . 31 g ( 58 %) of white crystals ; mp 220 °- 222 ° c . ( c , h , n ); 1 h nmr ( dmso d 6 ): δ0 . 98 ( d , 3h , j = 5 . 9 hz ), 2 . 36 ( s , 3h ), 2 . 38 - 2 . 48 ( m , 3h ), 3 . 14 ( d , 1h , j = 11 . 6 hz ), 3 . 64 ( s , 3h ), 6 . 02 ( s , 1h ), 6 . 24 ( s , 1h ), 9 . 64 ( bs , 1h ); 13 c nmr ( dmso - d 6 ): δ11 . 78 ( q ), 19 . 20 ( q ), 31 . 64 ( d ), 34 . 64 ( t ), 51 . 43 ( q ), 59 . 74 ( d ), 95 . 77 ( d ), 103 . 95 ( d ), 157 . 45 ( s ), 159 . 47 ( s ), 168 . 78 ( s ), 170 . 97 ( s ), 192 . 70 ( s ). initial evaluations for anticonvulsant activity included phases i , ii , via and vib test procedures which have been described [ 11 ]. these tests were performed either in male carworth farms no . 1 ( cf1 ) mice or male sprague - dawley rats . phase i , a qualitative anticonvulsant intraperitoneal evaluation in mice included three tests : maximal electroshock ( mes ), subcutaneous ( scmet ), and the rotorod test for neurological toxicity ( tox ). in the mes test , maximal electroshock seizures are elicited with a 60 - cycle alternating current of 50 ma intensity ( 5 - 7 times that necessary to elicit minimal electroshock seizures ) delivered via corneal electrodes for 0 . 2 seconds . a drop of 0 . 9 % saline is placed on the eye prior to application of the electrodes in order to prevent the death of the animal . abolition of the hind limb tonic extension component of the seizure is defined as protection and results are expressed as the number of animals protected / the number of animals tested . in the sc met test , eighty - five mg / kg of pentylenetetrazol , which induces seizures in more than 95 % of mice , is administered as a 0 . 5 % solution subcutaneously in the posterior midline . the animal is observed for 30 minutes . failure to observe even a threshold seizure ( a single episode of clonic spasms of at least 5 seconds duration ) is defined as protection and the results are expressed as the number of animals protected / the number of animals tested . the rotorod test is used to evaluate neurotoxicity . the animal is placed on a 1 inch diameter knurled plastic rod rotating at 6 rpm . normal mice can remain on a rod rotating at this speed indefinitely . neurologic toxicity is defined as the failure of the animal to remain on the rod for 1 minute and the results are expressed as the number of animals protected / the number of animals tested . compounds were suspended in 0 . 5 % aqueous methylcellulose and were administered at three dosage levels ( 30 , 100 , and 300 mg / kg ) with anticonvulsant activity and motor impairment noted 30 min and 4 h after administration . these phase i data are shown in table iii . phase via was a similar qualitative evaluation to the phase i evaluation , however the test drug was administered orally in rats utilizing the three tests noted previously . these data are shown in table iv . the phase ii test quantitated the anticonvulsant activity and motor impairment observed for the most promising compounds in phase i and phase via respectively . phase ii quantified data in cf1 mice by intraperitoneal administration . a special intraperitoneal test in rats was performed at 10 mg / kg for activity and at 100 mg / kg for toxicity . data on these results is shown in table v above . 1 . 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