NOTES
January 19TO Experimental Section5
2,5-Dimethoxy-4-methyl-p-nitrostyrene.-Amixture of .i.4 g (30 mmoles) of 2,5-dimet,hoxy-p-tolualdehyde, 2.5 g of ?;H40Ac, 2.5 ml of CH,KOz, and 25 ml of C6H6was refluxed for 20 hr, during which time HzO was azeotroped with a Dean-St,ark titbe. After cooling, the resulting solution was washed successively with H,O (two 25-ml portions), saturated solution of SaHSOs itm-0 2.5-nil portions), and H20 (two 25-ml portions). The CsH6 layer was dried (XasSOd) and evaporated in vacuo leaviiig 6.0 g ( 9 0 5 ) of yellow solid, mp 111-112'. Recrystallization from C&-C;H,s (1:2) gave 5.3 g (797,), mp 118-119". This melting point remained unchanged upon another recrystallization. .-i ICnH13NO4) d. C, H, ?J. 2,5-Dimethoxy-4-methyl-p-phenethylamine(4).-To a stirred suspension of 3.0 g (80 mmoles) of LiAlHl in 50 ml of T H F was added a solution of 4.4 g (18 mmoles) of 2,j-dimethoxy4-niethyl-B-iiitro..tyrene in 50 ml of T H F . The mixture was refluxed for 1 hr, cooled in ice, and treated with a mixture of H2O and T H F to decompose excess LiAlH,. The resulting mixture was filtered and the filter cake was extracted with T H F . The combined T H F solution was evaporated in vacuo leaving 3.7 g of oily product. -1solution of this oil in 25 ml of Et20 was treated with Et2O-HC1 to precipitate 3.4 g (837,) of the hydrochloride salt, mp 200-203". Recrystallization from E t O H gave 1.8 g, mp 212-213'. Addition of E t 2 0 to the filtrate yielded i 3 0 mg, mp 211-213". The total yield was 627,. Anal. (CH HiSClNOz) C , H, S . In a separat,e run distillation of free amine yielded 59% of a liquid, bp 95-105' 10.15mm), n Z 31.5385. ~ 2,5-Dimethoxy-N,N,4-trimethyl-p-phenethylamine(6).T o 14.0 g (0.3 mole) of formic acid, cooled in ice-HzO, was added dropwise 3.0 g (0.016 mole) of 2,S-dimethoxy-4-methylphenethylamine (4),then 3.6 g (0.12 mole) of formalin in 10-ml portions. The mixtiire was refluxed for 5 hr. After cooling to room temperature, 7 nil of concentrated HC1 was added and the resulting solution wa. evaporated in uacuo leaving an oil. This oil was dissolved in 2.5 ml of H?O and extracted with CHCl, (two 25-ml portions). The aqueous layer was made basic with 2 A T NaOH and extracted with Et20 (three 25-ml portions). The Et20 extracts containing the product was concentrated to about 25 ml. Addition of EtzO-HC1 to this solution precipitated the amine hydrochloride, yield 2.2 g mp 165-167". Recrystallization from EtOH-Et,O gave l . i g (42Yc), mp 168-169". Anal. (C13H??Cl?;Os) C, H , S .
2,5-Dimethoxy-N,4-dimethyl-~-phenethylamine (5).A mixt,iire of 3.b g (30 mmoles) of 2,5-dimethoxy-4-methyl-Pphenethylamine i4), 4.2 g (40 mmoles) of benzaldehyde, and 15 nil of C6H6 was reflused for 30 min and then subjected to distillation until the temperature reached 100". To the remaining viscous liquid was added Jowly a solution of 5.4 g (40 mmoles) of hIe2S04 in 20 ml of C6H6. The mixture was first heated until the reaction began. For several minutes no further heat was applied; then the mixture \\-as refluxed for 30 min. Xext, 20 ml of water was added and refluxing was continued for an additional 30 min. The aqueous phase was separated, extracted with C6H6(three 25nil portions), made basic with 2-V NaOH, and again extracted with C6H6 (three 2.5-ml portions). The combined C ~ H extracts G were dried ( S a 8 0 2 then evaporated in vacuo. Distillation of the residue gave 4.9 g i,i9%) of product, bp 96-99' (0.075 mm), n% 1.5278. When a solution of t.his product in 50 ml of Et20was treated with Et,O-HCl, a hydrochloride salt precipitated, yield , mp 130-1.5l0. Recrystallizat,ion from E t O H gave mp l.i0-1.il0. Anal. (C1zH2oClXO2)C, H, N. imethoxy-N,4-dimethylamphetamine (2).-The procedure was the ,hanie as described for the preparation of 2,5dimethoxy-N,4-dimethyl-p-phenethylamine( 5 ) . The free amine was obtained in a i 3 C , yield, bp 79" (0.075 mm) to 82" (0.05 mm), II% 1.5210. When a solution of this product in Et20 was mixed with Et20-HC1, the hyd salt separated as an oil at first aiid then solidified; yield p 122-123'. For purification, the hydrochloride balt w d in a small amount of E t O H aiid slowly precipitated . In this fashion pure 2, mp 125-126', was obtained in 4 6 7 yield. Anal. (C1,H&lN02) C, 13, s. re taken on a Mel-Temp apparatus and are corare indicated only by symbols of the elements or functions, analytical results obtained for those elements or functions were vitiiin zt0.4YG of the theoretical values.
135
Pharmacology. Conditioned Behavioral (VI) Test.--Adult male Sprague-Dawley rats were trained to press a lever in a n operant condit'ioning chamber on a variable-interval 2-mi11 (vi 2 ' ) schedule of food reinforcement. -445-mg Noyes pellet was delivered to the animal following each lever press on an average of every 2 min. This procedure produces a stable base line of responses from day t,o day. The animals were maintained on 22 hr of food deprivation and run daily for 1 hr. Immediately prior to each test session, two animals were given randomly assigned intraperitioneal doses of each compound in aqueous solution. The effect on performance was determined by calculating the per cent change in total response from the predrug session using the following formula: 5 change = [(predrug-drug)/predrug] x 100. Test sessions were given following 2 days of 10% or less change in performance. Dose-response relationships were obtained for each compound by averaging the results for the two animals. The dose which produced a 5070 decrease in response rate (ED:,) was extrapolated from these curves. Effect on Barbiturate Sleeping Time.-Vice were injected intraperitoneally with 50 pmoles/ kg of compounds in 307@ propylene glycol. A4fter5 min, sodium pentobarbital (40 mg,'kg) in saline was given via the same route. Controls were first given 30% propylene glycol then pentobarbital in saline. The presleeping time and sleeping time (loss of righting reflex) were recorded and treated statistically.
Acknowledgment.-The authors wish to thank Nrs. Faye Wilson for her technical assistance.
Substituted Quinazolonephenoxyethylhydrazines as Monoamine Oxidase Inhibitors'" S U R E N DS. RPIRMAR ~ . i l i ~R. C. A R O R I ' ~ Departmmt of Pharmacology and Therapeutzcs, King George's Medzcal Collegr, Lucknow Cniversity, Lucknow 3 , India Received April W9, 1969 Revised Manuscript Receiz~edBugust 19, 1969
The effectiveness of the chain length in inhibiting the enzyme monoamine oxidase (L20)was reflected by the pronounced inhibition observed with phenoxyalkylhydrazines having two or four CH2 groups as compared to those possessing three, five, or six CH, groups.2 Furthermore, anticonvulsant properties exhibited by quinazolones3 and various 3 l h O inhibitors4 led us to synthesize substituted quinazolonephenoxyethylhydrazines (Table I) and to determine their ability to inhibit JLAO. All quinazolonephenoxyethylhydrazineswere found to inhibit RIA0 activity of isolated rat liver mitochondria during oxidative deamination of tyramine by rat liver homogenate using kynuramine as the substrate (Table 11). The use of cyanide and semicarbazide during manometric determination of 1190 activity" was avoided in experiments using tyramine as the substrate since 0 2 uptake has been shown to reflect true enzyme activity in washed mitochondrial prepara(1) (a) This investigation was supported in part with the financial assistance ohtained from t h e Council of Scientific and Industrial Research. New Delhi, and t h e Indian Council of Xedical Research, Keiv Delhi. (1,) Research Fellon of the Sational Institute of Sciences, New Delhi. (2) D . J. Drain, J. G. B. Hoives, R . Lazare, A. If.Saloman. R. Shadbolt, and H. W.R . Williams. J . .Wed. Chem., 6, 63 (1963). (3) h l . L. Gujral, P. K. Saxena, and R . P. Kohli, I n d i a n J . .Wed. Res., 46, 207 (1957). (4) D . J. Prockop. P. a. Shore, and B. R. Brodie, .inn. S. Y . Acud. Sci., 80, 643 (1959). (5) K. H. Creasey, Biochem. J . , 64, l i 8 (1956).
tioiis.6 The degree of 11-40 inhibition was fouiid to incrc;iic significantly ti!, the i~itroductionof a wb>tituent at position 6 of the quinazolone nucleus during oxidative deamination of either tjximine or kynuramine (TiLble 11). Further increase in the degree of JLIO irihibitioii w i b observed with compounds posses\irig wbstituents at both positions 6 and 8 of the quinazolone iiucleu\. Such a pattern of cwzyme inhibition n-:tz :LI\O reflected by the Iio values determined for thebe illliibitors using t j n m i n e :\\ the subjtrate. The rrl:iti~~c tdectronegativity of the halogen substituent attached :it position 6 of the quinazolone nucleus w a b found to liar-c no effect o n the degree of enzyme' inhibitioii. Thebe cl~iinazolonephenox?.ethylh!.drazir~e~ were fouiid to he weal< inhibitors of 1IXO tis compared to tranj 1q promitic. :ind /3-phcnJ lihoprop!-lh!.drazine.6
Experimental Section Anthranilic acids were hyiithesized accordiiig t o ihe inethiicl~ reported in the literatiire: ,~-chloroanthrauili~,~ 5-hrunioaiithra~iilic,~3-iodoaiith~aiiilic,~:i,5-dichloroanthranili(.,i a i i d :',,~-dibronioaiithra~iiIic acid.8 Acetanthranils, synthesized by refluxirig a niixt ru'e of 8 1 1 appropriate anthranilic acid (1 mole) atid 2 moles of .4cd),In w e r e iised without further purification. QuinazolonephenoxyethylBromides.-~~iii~azoloiie~ were .-yiit hesized by heating equimolar proportioils of appropriate ace(aiithranil and p-amiiiophenox).eth?l bromide1] as reported eai.1ier.I" The various qiiiiiazolonepheiio~yeth3.l bloniides were characterized by their sharp melting poiiit and elemeutal aiial>-?ei (Table I ) . Quinazolonephenoxyethy1hydrazines.-A mixture uf a11 iippropriate quinazolonephenoxyethyl bromide (1 mole) aiid 2.5 ( 0 ) S. 9.Parmar. Riorhem. I'hurmacol., 15, 1 4 Y i 11966). ( 7 ) hI. R I . Endicott. 13. 13, .\Iden, a n d hl. L.Sherrill, J . .4m. Chrrri. S o r . , 6 8 , 1303 (1946). ( 8 ) -1.S.JT-lieelrr and \V. hi. Oats, ihu/., 3.2, i 7 0 (1Y10). ( ! I ) C. J . litemme a n d J. 11. Hunter, d . O r g . Chem.. 6 , 2 2 i t l \ M O J . ' 10) S. 3. Parmar and K. C . 4rora. Can. J . C h e m . , 46, 2519 (1968). I 1 I ) 1 3 . c'. .\rora. 1'11.11. TtieBia, Lucknow Vniversity, Lucknon-, l!l6X.
mole. of ?j€12?;11?. I I z O [ 9 9 - 1 0 0 ~j ' ~iii wlhulute EtOII \\:I. 1'1,fliixed fur 15-20 hi.'" After distilling esceas EtOH the hydr:+ziilr~i n-hich separated out on c d i i i g were filtered a i d iw (EtOH). They are recorded iri Table I. Determination of Monoamine Oxidase Activity. ( a ) Manometric Method.-1lAO activity of isolated rat liver m i ~ ~ i c ~ h i ~ ~ i i l r i a wan determined by the cuiivent ioiial IVarhiirg maii(~inet riv t c ~ v t i iiiqiie iising tyramine as the d x t r a t e . 6 ( b ) Spectrophotofluorometric Method.---llA( ) j i c t ivi1.v { i f ]'at liver homogenate (.j w/v) in 0.25 d l sucrose \va> detei~nii~ic~ti rpectrophotoflriororIiet I' 11y usiiig kynriramiiie a s thc ..iili-trnte.'2 The reaction rnixt w e , iii filial curicelit ration, c~oii;i\lt d of 83 m.M phosphate buffer (pH 7.4) and 0.2 ml of rat liver homogeiiate. CZuinazolonephenos3.ethylh~dra~ilies(:< X 1 .I! 1) kyniiramine (100 p g ) , and 1120 were added t o d j i i - 1 the final rolriine to 3 nil. The reaction waq ron :r . Pro Foundation. S e w Y o ~ kaiid i i f the University of -\I;itiitoba. IVinnipeg, for 6eIlC'l'l Jt IS gifts of :in Aniiiico Ho\\.muti spectrophcitiifil~orcirn~~ti~~' :tilt1 lqxuramine, respect iv(>Ij-. J
