Aust. J. Chem., 1989, 42, 1759-68

Imidazo[ l,2-blpyridazines. VIII* Syntheses and Central Nervous System Activities of Some 6-Benzylamino(and methoxybenzy1amino)- 3-methoxy-2-phenyl(substituted phenyl or pyridinyl)imidazo[l,2-blpyridazines

Gordon B. B ~ r l i n , ~ Les P. ~ a v i e s ~ and Maria M. L. NguA

A Division of Neuroscience, John Curtin School of Medical Research, Australian National University, G.P.O. Box 334, Canberra, A.C.T. 2601.

Visual Sciences Group, Research School of Biological Sciences, Australian National University, G.P.O. Box 475, Canberra, A.C.T. 2601.

Abstract

Syntheses and I C ~ O values for the displacement of 3 ~ - d i a z e p a m from rat brain plasma membrane are reported for a series of 6-benzylamino(and methoxybenzylamino)-3-methoxy- 2-phenyl(substituted phenyl and pyridinyl)imidazo[l,2-blpyridazines (and their 6-anilino and 6-phenethylamino analogues). The results are compared with those reported previously (by us) for the 6-chloro, 6-phenoxy, 6-benzylthio, and 6-benzyloxy compounds.

In the imidazo[l,2-blpyridazine ring system, 6-(0- and m-methoxybenzylamino) groups were found to be beneficial to binding activ~ty; elght compounds have been prepared with 1 ~ 5 0

values less than 2 nM (cf. 3~-d iazepam with l c s o of 4 . 2 nM). 2-(p-Aminopheny1)-3-methoxy- 6-(m-methoxybenzylamino)imidazo[l,2-b]pyridazine exhibited the highest activity with I C ~ O

1 . 0 nM.

Introduction

In previous parts of this series we have reported the synthesis and systematic structure-activity studies of derivatives of the imidazo[l,2-blpyridazine ring system and their displacement of 3~-d iazepam from rat brain membranes. More recently, this has involved an examination of 6-chloro-,l 6 - ~ h e n o x y - , ~ 6-ben~yl th io- ,~ 6-(alkoxy- and methylthio-phenoxy and methoxybenzy1thio)-,4 and 6-benzyloxy-3-methoxy-3-arylimidazo[l,2-blpyridazines.5 In this paper we describe similar studies of 6-benzylamino(and methoxybenzy1amino)-3-methoxy- 2-phenyl(substituted phenyl and pyridinyl)imidazo[l,2-blpyridazines.

Syntheses

The 6-anilino-3-methoxy-2-(p-methylphenyl)imidazo[l,2-b]pyridazine (1; R =

Me) required as a reference compound in this work was prepared from the known 3-

* Part VII, Aust. J. Chem., 1989 42, 1749.

Barlin, G. B., Davies, L. P., and Ngu, M. M. L., Aust. J. Chem., 1988, 41, 1149. * Barlin, G. B., Davies, L. P., and Ngu, M. M. L., Aust. J. Chem., 1988, 41, 1735.

Barlin, G. B., Davies, L. P., Ireland, S. J., and Ngu, M. M. L., Aust. J. Chem., 1989, 42, 1133. Barlin, G. B., Davies, L. P., Ireland, S. J. , and Ngu, M. M. L., Aust. J. Chem., 1989, 42, 1735. Barlin, G. B., Davies, L. P., and Ngu, M. M. L., Aust. J. Chem., 1989, 42, 1749.

G. B. Barlin, L. P. Davies and M. M. L. Ngu

anilino-6-chloropyridazine through 3-anilino-6-hydrazinopyridazine which was catalytically reduced to 6-anilinopyridazin-3-amine. This latter compound condensed with p-methylphenylglyoxal to give the hydroxy compound (1; R = H ) and methylation with diazomethane afforded the 0-methyl ether (1; R = Me). In our hands 3-anilino-6-chloropyridazine with ethanolic ammonia did not give 6-anilinopyridazin-3-amine directly.

Table 1. Results for the displacement of 3~-diazepam from rat brain membranes by substituted imidazo[l,2-blpyridazines

1 ~ 5 0 values (nM) [in the presence of 1 0 0 ~ ~ y-aminobutyric acid (CABA) (see Experimental section for details)] are the concentrations required to displace 50% of specific 3 ~ - d i a z e p a m binding to rat brain membrane preparations, by using standard assay conditions described in ref. 1. For those compounds which were assayed in more than one experiment mean 1 ~ 5 0 values?s.e.m. (standard error of the mean) are given with the number of assays in

parentheses

Compound number

Substituted imidazo[l,2-blpyridazine

lcso (or percentage displacement)*

CBLD-2 10 6-NHPh-3-OMe-2-C6H4Me-p (not significant at 1000 nM)

C B L D - ~ ~ ~ 6-NHCHzPh-3-OMe-2-Ph 9 CBLD-228 6-NHCHzCHzPh-3-OMe-2-Ph 800 CBLD-2 57 6-NHCH2 CsHqOMe-0-3-OMe-2-Ph 1 . 8 CBLD-3 15 6-NHCHzCsH40Me-o-3-OMe-2-C6H4Me-p 5 .5 CBLD-309 6-NHCHzCsH40Me-o-3-0Me-2-C6H4C1-p 16 GBLD-3 1 2 6-NHCHzC6H40Me-o-3-OMe-2-C6H4F-p 2.221 .3(2) GBLD-323 6-NHcHzCsH40Me-o-3-OMe-2-C6H4N02-m 3 0 GBLD-324 6-NHCHzCfjH40Me-0-3-OMe-2-C6H4N02-p (20.6% at 10 n ~ ) CBLD-3 10 ~ - N H C H Z C ~ H ~ ~ M ~ - O - ~ - O M ~ - ~ - C ~ H ~ N - B ~ 3 CBLD-2 3 1 6-NHCHzC6H40Me-m-3-0Me-2-Ph 2 .4(2) GBLD-308 6-NHCHzCsH40Me-m-3-0Me-2-C6H4Me-p 3 .2 t0 .3 (2 ) GBLD-283 6-NHCHzCsH40Me-m-3-OMe-2-C6H4F-o 1 . 5 GBLD-2 16 6-NHCH2CsH40Me-rn-3-OMe-2-C6H4F-m 15 CBLD-274 6-NHCH2CsH40Me-m-3-OMe-2-C6H4F-p 1 . 5 t 0 . S(2) CBLD-272 6-NHCHzCsH40Me-m-3-0Me-2-CmF3-m 5 5 CBLD-281 6-NHCHzCsH40Me-rn-3-OMe-2-C6H4CF3-p 24 CBLD-307 ~ - N H C H Z C I ~ H ~ O M ~ - ~ - ~ - O M ~ - ~ - C ~ H ~ N O ~ - ~ 2.5+0.5(3) CBLD-306 6-NHCH2CsH40Me-m-3-0Me-2-C6H4N02-p 13 .0 GBLD-3 13 6-NHCHzCsH40Me-m-3-OMe-2-C6H4NH2-m 1 .8tO. 4(3) GBLD-3 14 6-NHCHzCsH40Me-m-3-OMe-2-C6H4NH2-p 1 .010.1(2) GBLD-271 ~ - N H C H Z C ~ H ~ O M ~ - ~ ~ - ~ - O M ~ - ~ - C ~ H ~ N - ~ ~ (21 .7% at 30 n ~ ) G B L D - ~ ~ ~ 6-NHCHzCsH40Me-m-3-0Me-2-C5H4N-P 2 .IrtO. 6(3) GBLD-2 58 6-NHCH2C6H40Me-m-3-OMe-2-C5H4N-y 5 . 8 GBLD-253 ~-NHCHZC~H~(OM~);!-~~,~-~-OM~-~-P~ 3 . 4

Diazepam 4.3-+0.3(3)

A Percentage displacement at the concentration specified. C5H4N = pyridinyl.

The 6-benzylamino- 3 -methoxy - 2 -phenyl(and pyridinyl)imidazo[l,2-blpyrid- azines were instead prepared from 6-chloropyridazin-3-amine 2-oxide (2; X = C1) by replacement of the relatively more reactive 6-chloro substituent with the relevant benzylamine to give the 6-benzylaminopyridazin-3-amine 2-oxide (2; XcNHCH2Ar). The last named compounds condensed readily with phenacyl bromide, a-bromoacetylpyridines and analogous compounds to give the hydroxy compounds (3; R = H) and like ( I ; R = H) these were methylated readily with diazomethane to give compounds (3; R= Me).

3-Methoxy-6-phenethylamino-2-phenylimidazo[l,2-b]pyridazine was prepared in an analogous manner.

Biological Activity

The imidazo[l,2-blpyridazines reported in this paper were examined for their ability to displace 3H-diazepam bound to rat brain synaptic plasma membranes. Details of the test procedure have been reported in our earlier paper1 and the results for this series are recorded in Table 1. Standard errors of the mean (s.e.m.) are shown for multiple determinations.

6-Benzylamino-3-methoxy-2-phenylimidazo[l,2-b]pyridazine (GBLD-222, 1c50 9) was more effective in the displacement of 3~ -d i azepam than was its 6-benzyloxy5 or 6-benzylthio3 analogues (GBLD-158 and 137 with ICSO values of 20 and 25 respectively). It was also significantly more active than its 6-anilino and 6-phenethylamino analogues (GBLD-210 and 228 which showed no significant displacement at 1000 nM and an 1c50 value of 800 nM respectively). These differences, particularly for the 6-anilino compound, were much larger than in the series 6 -~heny l th io - ,~ 6-benzylthi0-~ and 6-phenethylthio-3-methoxy-2- phenylimidazo[l,2-b]pyridazines3 (GBLD-1 29, 137 and 143 with I C ~ O values 11 7, 25 and 666 respectively). The 6-phenoxy4 analogue (GBLD-163; IC50 1120) was also more active than the 6-anilino compound (GBLD-210).

Methoxy groups in the aromatic substituent at the 6-position, as observed p r e v i o u ~ l y , ~ * ~ engendered significant activation. Thus the 6-(0-methoxy-, m- methoxy- and 3',41-dimethoxy-benzylamino) compounds GBLD-257, 231 and 253 had ICSO values of 1 . 8 , 2 . 9 and 3 - 4 nM respectively, i.e. 3-5 times more active than the 6-benzylamino analogue GBLD-222 ( 1 ~ 5 0 9 n ~ ) . Previously observed effects varied from approximately three times in the 6-benzylthio4 and 6-benzyloxy5 compounds to approximately 15 times in the 6-phenoxy4 series.

Amongst the 6-(2'-methoxybenzylamino) compounds, a 2-(p-methylphenyl) substituent as in GBLD-3 15 was deactivating relative to its 2-phenyl analogue GBLD-257 by approximately threefold. This is a similar effect but greater in magnitude than that observed in the 6-benzyloxy s e r i e ~ . ~ Likewise the 24p- chlorophenyl) compound GBLD-309 was less active by approximately eightfold and compares with a deactivation by approximately twofold in the 6-benzylthio ~ e r i e s . ~

The 2-(p-fluorophenyl) compound GBLD-3 12 ( 1 ~ 5 0 2 2 n ~ ) had approximately the same activity as its 2-phenyl analogue GBLD-257 ( 1 ~ 5 ~ 1 . 8 n ~ ) whereas in the 6-chloro, 6-(0-methoxyphenoxy), 6-(0-methoxybenzylthio) and 640- methoxybenzyloxy) series, the 2-(p-fluorophenyl) substituent increased activity (relative to the 2-phenyl analogue).

~ a v i e s , L. P., unpublished data.

G. B. Barlin, L. P. Davies and M. M. L. Ngu

The 24m- and p-nitrophenyl) compounds GBLD-323 and 324, as observed in previous series, were less active than their 2-phenyl analogue GBLD-257. Substitution of the 2-phenyl group in GBLD-257 by a pyridin-3-yl group as in GBLD- 310 was slightly detrimental to activity ( 1 ~ 5 0 values 1 - 8 and 3 nM respectively).

In the series of 6-(m-methoxybenzylamino) compounds, the 2-(p-methylphenyl) compound GBLD-308 ( 1 ~ ~ 0 3 .2 n ~ ) had approximately the same activity as the parent (GBLD-231; 1C50 2 . 9 n ~ ) and thus differs from its effect in the 640- methoxybenzylamino) series reported above. Of the 2-(fluorophenyl) compounds both the o- and p-isomers (GBLD-283 and 274,1~50 1 5 nM for both) were more active than GBLD-23 1 but the m-isomer (GBLD-216; 1 ~ 5 0 15 n ~ ) was considerably less active. Higher activity by 2-(p-fluorophenyl) compounds has been noted previously in the series of 6-chloro,l 6-(0-metho~yphenoxy),~ 6-(m-methoxybenzylthio) and 6-(m-metho~ybenzyloxy)~ compounds whereas 2-(0-fluoropheny1)-3-methoxy-6- (0-methoxyphenoxy)imidazo[l,2-blpyridazin (GBLD-282; 1 ~ 5 0 148 n ~ ) was half as active as its 2-phenyl analogue (GBLD-167; I C ~ O 70 n ~ ) . In other series5 the 24m- fluorophenyl) group usually had little effect (relative to the 2-phenyl analogue).

The 2-(m- and p-trifluoromethylphenyl) compounds GBLD-272 and 281 were less active than GBLD-231 and this is consistent with results from other series.*r5

Whereas the 2-(p-nitrophenyl) compound GBLD-306 was less active than GBLD-231, the 2-(m-nitropheny1)isomer (GBLD-307; 1 ~ 5 0 2 - 5 n ~ ) was slightly more active. The 2-(m- and p-aminophenyl) compounds GBLD-313 and 314 ( 1 ~ 5 0 values 1 . 8 and 1 - 0 nM respectively) were approximately twofold more active than GBLD-231 and this is qualitatively similar to the effect in other derivatives of this ring ~ y s t e m . ~ ~ ~ ~ ~

Unlike the results in the 6-(0-methoxybenzylamino) series, the 3-methoxy-6- (m-methoxybenzy1amino)-2-(pyridin-3'-yl) compound GBLD-254 (ICSO 2 . 1 n ~ ) was more active than GBLD-231 whereas the 2-(pyridin-2/-yl and pyridin-4/-yl) isomers were less active. The general superiority of the 2-(pyridin-3/-yl) isomer has been noted previously.*

Experimental

All compounds were examined for the presence of impurities by thin-layer chromatography on alumina and silica and by I H n.m.r. spectroscopy. Solids for analysis were dried at 80-100" and 0 . 1 mmHg for 3-6 h unless otherwise specified. Melting points are uncorrected and were taken in open Pyrex capillaries. Analyses were performed by the Australian National University Analytical Services Unit. 'H n.m.r. spectra were generally recorded at 90 MHz and 30" with a Jeol FX9OQ Fourier-transform spectrometer, with tetramethylsilane (in CDC13 or CD3SOCD3) as internal standard.

This compound was prepared from 3,6-dichloropyridazine and aniline in ethanol at reflux. It had m.p. 185-186" (from cyclohexane) (lit. 1 9 0 0 , ~ 191.2-192°8). I H n.m.r. (CDC13) 6 7.10, d, 54,s 9 HZ, H4(5); 7 .26, d, J4,5 9 HZ, H 5(4); 7.30-7.50, complex, Ph.

A mixture of 3-anilino-6-chloropyridazine ( 2 . 0 g) and hydrazine hydrate (40 ml) was refluxed for 5 h. The hydrazine was evaporated under reduced pressure and the residue

Kumagai, M., Nippon Kagaku Zasshi, 1961, 82, 226 (Chem. A b s t ~ , 1962, 56, 10139). Relyea, D. I., Riddell, J. A., and Tawney, P. O., J. Med. Chem., 1963, 6, 807.

Imidazo[l,2-blpyridazines. VIII

suspended in water and the product (0.70 g) collected. It was recrystallized from aqueous methanol to give 3-anilino-6-hydrazinopyridazine, m.p. 204-207" (Found, for a sample dried at 40°/0. 1 mmHg for 4 h: C, 59.4; H, 5.7: N, 34 7. CloHllNs requires C, 59.7; H, 5 -5 ; N, 34.5%). 'H n.m.r. (CD3SOCD3) 6 4.1, br, NH2; 6 .78, d, J4,5 9 HZ, H5(4); 6 .95, d , J4,5 9 HZ, H 4 6 ) ; 6.98-7.69, complex, Ph; 7 .79, br, NH.

3-Anilino-6-hydrazinopyridazine (0 .2 g) in methanol (150 ml), with a little water, was shaken with hydrogen over Raney nickel at room temperature and pressure for 6 h. The catalyst was filtered off and the filtrate evaporated to give a solid (0 .1 g). 'H n.m.r. (CD3SOCD3) 6 5.71, br, NH2; 6.77, d , J4.5 9 HZ, H 5(4); 6 .97, d , J4,5 9 HZ, H4(5); 6.87-7.83, complex, Ph; 8.65, br, NH.

A portion of this product with aqueous picric acid gave a yellow precipitate which recrystallized from ethanol to give 6-anilinopyridazin-3-amine picrate, m.p. >250° (Found, for a sample dried at 50°/0 0. 1 mmHg for 6 h: C, 46.5; H, 3.3; N , 23 -4 . C14H13N707 requires C, 46.3; H, 3 .2; N, 23.6%).

A mixture of 6-anilinopyridazin-3-amine (0 .20 g), 4-methylphenylglyoxalg (0 .18 g) and concentrated hydrochloric acid (0 .3 ml) in ethanol (25 ml) was refluxed for 4 h.

After cooling, the solvent was evaporated to give a dark red residue which was stirred with excess ethereal diazomethane at 0° , and then at 20" overnight. The mixture was evaporated and the residue was subjected to t.1.c. (alumina; chloroform) and the band showing light-blue fluorescence under the 254-nm lamp at low RF was collected and extracted with chloroform to give the product (0.04 g) which was recrystallized from a mixture of methanol and cyclohexane to give the title compound, m.p. 228-230" (Found: C, 72.5; H, 5 .8; N, 16.5. C20H18N40.1/6H20 requires C, 72.1; H, 5 .5; N, 16.8%). 'H n.m.r. (CDCI3) 6 2.39, s, Me; 4 - 14, s, 3-OMe; 6 .64, d, J7,8 9 HZ, H7; 6 .77, br, NH; 7.11-8.03, complex, H 2/,3',5',6' and Ph; 7.65, d, J7,8 9 HZ, H8.

6-Chloropyridazin-3-amine 2-oxidelo (0 .2 g) and benzylamine (2 .0 g) were heated in a screw-top Teflon-lined stainless steel reaction vessel at 160' for 20 h. After cooling, chloroform (2 ml) was added to the brown residue and the mixture subjected to column chromatography (alumina; chloroform) and the product eluted with ethanol and methanol, respectively. The light-brown residue was precipitated from cold chloroform with ether and the yellow precipitate (0.18 g) recrystallized from acetone to give golden brown crystals of the title compound (0.15 g), m.p. 189-191" (Found: C, 61.4; H, 5 .7; N, 25.9. CllHlzN40 requires C, 61.1; H, 5 .6; N, 25.9%). 'H n.m.r. (CD3SOCD3) 6 4.32, d, JCH,NH 6 Hz, CH2; 5.85, br, NH2; 6 .52, d , 7 .07, d, J4,5 9Hz , H4,5; 7.30, br, Ph.

6-Chloropyridazin-3-amine 2-oxide4 (0 .3 g) and phenethylamine (2.4 g) were heated in a screw-top reaction vessel at 160' for 16 h. The reaction mixture was poured into cold ether and the light-yellow precipitate filtered off. It was recrystallized from chloroform to glve yellow crystals of the title compound (0 .3 g), m.p. 170-173" (Found: C, 62.5; H, 6 . 1 ; N, 24.1. C12H14N40 requires C, 62.6; H, 6 . 1 ; N, 24.3%). 'H n.m.r. (CD3SOCD3) 6 2.80, t, J ~ H , N H 6 . 5 Hz, CHzPh: 3 .29, m, CH2NH; 5.84, br, NH2; 6 .45, d, 7.05, d , 54,s ~ H z , H4,5; 6 .53, br t, JCH,NH 6 . 5 HZ, NH; 7.25, br, Ph.

This compound was prepared from 6-chloropyridazin-3-amine 2-oxide (0 .8g) and 2- methoxybenzylamine (4 .8 g) as for the benzylamino analogue. The product was subjected to column chromatography (alumina; acetone and the product eluted with ethanol and

Kimura, S., Miyagi, Y., and Goto, R., Bull. Chem. Soc. Jpn., 1966, 39, 1333. lo Itai, T., and Nakashima, T., Chem. Pharm. Bull., 1962, 10, 936.

G. B. Barlin, L. P. Davies and M. M. L. Ngu

methanol). It was recrystallized from propanol and ethyl acetate to give yellow crystals of the title compound (0 .7g) , m.p. 204-206" (Found: C, 58-9 ; H, 6 .0 ; N, 22.9. ClzH14N402 requires C, 58.5; H, 5 .7 ; N, 22.8%). 'H n.m.r. (CDCI3) 6 3.81, s, OMe; 4.27, d , JCH,NH 6 HZ, CH2; 5 .9 , br, NH2; 6 .56, d, 7.08, d , J4,5 9 Hz, H4,5; 6.74-7.31, complex, H3/,4',5',6'.

This product was prepared as for its isomer above, purified by column chromatography (alumina; chloroform, eluted with ethanol) and recrystallized from a mixture of propanol and acetone to give the title compound (63%), m.p. 161-163" (Found: C, 58.7; H, 6 .0 ; N, 23.0. C12H14N402 requires C, 58.5; H, 5 .7 ; N, 22.8%). 'H n.m.r. (CD3SOCD3) 6 3.72, s, OMe; 4 .29, d , JCH,NH 6 HZ, CH2; 5.8, br, NH2; 6.52, d, 7.07, d , J4,5 9 Hz, H4,5; 6.71-7.32, complex, H 2/,4/,5',6/.

In a similar manner, from 6-chloropyridazin-3-amine 2-oxide and 3',41-dimethoxybenzy1- amine (160") was prepared the title compound (34%), m.p. 152-154' (from a mixture of acetone and ethyl acetate) (Found: C, 56.7; H, 6 .1 ; N, 20.2. C13H16N403 requires C, 5 6 5 ; H, 5 .8; N, 20.3%). 'H n.m.r. (CD3SOCD3) 6 3.71, s, 3 .72 , s, 2xOMe; 4.42, d, JCH,NH 5 .5 HZ, CH2; 5 .84, br, NH2; 6 . 50, d , 7.06, d, J4-5 9 HZ, H4,5; 6.76-6.95, complex, H2',5',6'.

6-Benzylamino-3-methoxy-2-phenylimidazo[l 2 - b p y r d a z i e (GBLD-222) and Related Compounds

A mixture of 6-benzylaminopyridazin-3-amine 2-oxide (0 .08 g), phenacyl bromide (0 .07 g) and ethanol (6 ml) was refluxed with stirring for 4 h. The solvent was evaporated and the brown residue methylated with excess ethereal diazomethane at O o , then at 20" overnight. The solvent was evaporated and the residue subjected to t.1.c. (alumina; chloroform, then ether) and the product (0 .03 g) was recrystallized from light petroleum (b.p. 60-80") to give the title compound, m.p. 132-134' (Found: C, 73.0; H, 5 .7; N, 16.8. C20H18N40 requires C, 72.7; H, 5.5; N, 17.0%). 'H n.m.r. (CDC13) 6 4.04, s, OMe; 4 .59, d , JCH,NH ~ H z , CH2; 6 .37 , d, J7,8 9 HZ, H 7; 7.36-8.12, complex, 2xPh; 7.53, d , 57,s 9 HZ, H 8.

In a similar manner were prepared the following compounds. 3-Methoxy-6-phenethylamino-2-phenylimidazo[l,2-b]pyridazine ( ~ ~ ~ ~ - 2 2 8 ) . - T h e crude prod-

uct was subjected to t.1.c. (alumina; chloroform) and recrystallized from light petroleum (b.p. 60-80") to give the title compound (15%), m.p. 109-1 11" (Found, for a sample dried at 8O0/0. 1 mmHg for 8 h: C, 73.1; H, 5 .9; N, 16.0. C21H20N40 requires C, 73 .2 ; H, 5 .9; N, 16.3%). 'H n.m.r. (CDC13) 6 2.99, t, JCH,NH 6 . 5 HZ, CH2Ph; 3 .69, q, JCH,NH 6 . 5 HZ, CHzN; 4 .15 , s, OMe; 4 .50, t, JCH,NH 6 . 5 HZ, NH; 6 .27 , d, J7,8 9 HZ, H 7; 7.23-8.15, complex, 2xPh; 7.50, d, J7,8 9 HZ, H8.

3 -Methoxy -6-(2'-methoxybenzy1amino)-2-phenylimidazo[l 2 - b p y r i d a z i e (cs~~-257).-The crude product was subjected to t.1.c. (alumina; chloroform) and recrystallized from a mixture of chloroform and cyclohexane to give the title compound (35%), m.p. 132-134" (Found: C, 69.9; H, 5 .7; N, 15 ~ 4 . C21H20N402 requires C, 70.0; H, 5 .6 ; N, 15.5%). 'H n.m.r. (CDC13) 6 3.88, s, 2'-OMe; 4 .11 , s, 3-OMe; 4 .59, d, J c H , ~ ~ ~ H z , CH2; 5 .10, t, JCH,NH ~ H z , NH; 6 .35, d, J7,8 9 Hz, H 7; 6.83-8.15, complex, H3',4/,5/,6' and Ph; 7 .48, d , J7,8 9 HZ, H8.

3 -Methoxy - 6 - (3' -methoxybenzylamino) - 2-phenylimidazo[l, 2-blpyridazine (cs~~-231).-The crude product was subjected to t.1.c. (alumina; chloroform) and recrystal4ized from cyclohexane and also chloroform/cyclohexane to give the title compound (35%), m.p. 157-159' (Found: C, 69.7; H, 5 . 8 ; N, 15.3 . C21H20N402 requires C, 70.0; H, 5 .6; N , 15.5%). 'H n.m.r. (CDC13) 6 3.80, s, 3'-OMe; 4 .06, s, 3-OMe; 4 .58, d , J c H , ~ ~ 5 HZ, CH2; 5 .0 , t , JCH,NH 5 HZ, NH; 6 .36, d , J7.8 9 HZ, H7; 6.79-8.14, complex, H2',4',S,6' and Ph; 7 .53, d , J7.8 9 HZ, H8.

6-(3',4'-Dimethoxybenzy1amino)-3-methoxy-2-pheny1imidazo[1,2-b]pyridazine (GELD- 2531.-The crude product from this preparation was applied to an alumina t.1.c. plate and developed twice with chloroform, and then recrystallized from a mixture of acetone and cyclohexane to give off-white crystals of the title compound (48%), m.p. 162-164" (Found: C, 67.7; H, 5 . 7 ; N, 14.1 . C22H22N403 requires C, 67.7; H, 5 .7 ; N, 14.3%). 'H n.m.r. (CDC13) 6 3.85, s, 2xOMe; 4 .08 , s, 3-OMe; 4 .50, d , JCH,NH 5 HZ, CHZ; 5.00, t, JCH,NH 5 HZ, NH; 6 .36, d , J7,8 9 HZ, H 7; 6.75-8.13, complex, H 2/,5',6/ and Ph; 7 .49, d, J7,8 9 HZ, H 8.

Imidazo[l,2-blpyridazines. VIH

3-Methoxy-6-(31-methoxybenzylamino)-2-(pyridin-2-yl)imidazo[1,2-b]pyridazine (GBLD-271) and Related Compounds

A mixture of 6-(31-methoxybenzylamino)pyridazin-3-amine 2-oxide (0.25 g), 2-bromoacetyl- pyridine hydrobromide4 and sodium hydrogen carbonate (0.08 g) in ethanol (10 ml) was refluxed for 8 h. After cooling, excess cold ethereal diazomethane was added and the mixture stirred at 0" and then at 20" overnight. This mixture was evaporated and the residue subjected to t.1.c. (alumina; chloroform/acetone, 3 : 1) and the product (0 ~ 0 5 g) recrystallized from a mixture of acetone and cyclohexane to give the title compound, m.p. 134-135" (Found: C, 66.8; H, 5 .3; N, 19.3 . C20HlgN502 requires C, 66.5; H, 5 .3 ; N, 19.4%). 'H n.m.r. (CDCI3) 6 3.76, s, 3'-OMe; 4.10, s, 3-OMe; 4 .55 , d , JCH,NH 5 . 5 HZ, CH2; 5.16, t, JCH,NH 5 .5 HZ, NH; 6 .42 , d , 57,s 9 HZ, H 7; 6.64-8.71, complex, H21,41,5',61 and pyridin-2"-yl; 7.51, d, J7,8 9 HZ, H 8.

In a similar manner the two pyridinyl isomers listed below were prepared. 3 - Methoxy - 6 - (3' -methoxybenzylamino) - 2 - (pyridin - 3" -yl)imidazo[l , 2 -b]pyridazine (GBLD-

254).--The crude product from the reaction of 6-(31-methoxybenzylamino)pyridazin-3-amine 2-oxide and 3-bromoacetylpyridine hydrobromide4 followed by methylation was subjected to t.1.c. (alumina; chloroform/acetone, 1 : 1) and recrystallized from chloroform/cyclohexane/light petroleum (b.p. 40-60') to give the title compound (25%), m.p. 188-189" (Found: C, 64.6; H, 5 .4 ; N, 18.8. C20H19N502.0.5H20 requires C, 64.8; H, 5 .4; N, 18.9%). I H n.m.r. (CDC13) 6 3.81, s, 3'-OMe; 4.08, s, 3-OMe; 4 .57, d, JCH,NH 5 .5 HZ, CH2; 4 .9 , br, NH; 6 .42, d, J7.8

9 Hz, H 7; 6.77-9.31, complex, H 2',4',5',6' and pyridin-3"-yl; 7.56, d, J7,8 9 Hz, H 8. 3 - Methoxy -6-(3' -methoxybenzylamino)-2 -(pyridin -4'' -yl)imidazo[l ,2-bJpyridazine (GBLD-

258).-The product from the reaction of 6-(3'-methoxybenzylamino)pyridazin-3-amine 2-oxide with 4-bromoacetylpyridine hydrobromide4 and subsequent methylation was subjected to t.1.c. (alumina; chloroform/acetone, 3 : 1 and alumina; ethyl acetate) and recrystallized from acetone/cyclohexane and toluene/cyclohexane to give the title compound, m.p. 137-139" (Found: C, 66.5; H, 5 .3; N, 19.3. C20H19N502 requires C, 66.5; H, 5 .3; N, 19.4%). ' H n.m.r. (CDC13) 6 3.81, s, 3'-OMe; 4 .11, s, 3-OMe; 4.58, d, JCH,NH 5 . 5 HZ, CH2; 4 .7 , br, NH; 6 .43 , d , J7.8 9 HZ, H7; 6.78-8.65, complex, H2',4/,5',6' and pyridln-4"-yl; 7 .58, d , J7,8 9 Hz, H 8.

2-(2~-Fluorophenyl)-3-methoxy-6-(31~-methoxybenzylamino)imidazo[l,2-b]pyridazine (GBLD-283) a n d Related Compounds

a-~romo-2-fluoroacetophenone4 (0.184 g) was added to a solution of 6-(3'-methoxybenzyl- amino)pyridazin-3-amine 2-oxide (0 .2 g) in ethanol ( 8 . 0 ml) and the mixture refluxed with stirring for 9 h, then evaporated under reduced pressure.

The residue was dissolved in ethanol (2 .0 ml), ethereal diazomethane (from 2 . 0 g nitrosomethylurea) added and the mixture stirred at 0" and at 20" overnight. The solvent was evaporated and the product subjected to t.1.c. (alumina; chloroform) and recrystallized from benzene/cyclohexane to give the title compound (0.079 g), m.p. 133-1 34" (Found, for a sample dried at 100" for 6 h: C, 67 .0 ; H, 4 .9; N , 14.5. C21HlgFN402 requires C, 66 .7 ; H, 5 .1 ; N, 14.8%). I H n.m.r. (CDC13) 6 3.81, s, 3/'-OMe; 4 .05, s, 3-OMe; 4.57, d , JCH.NH 5 HZ, CH2: 4 .8 , t, JCH,NH 5 HZ, NH; 6 .40, d, J7,8 9 Hz, H 7; 6.80-7.90, complex, H3',4',5',6',2",4",5",6"; 7 .58, d, 57.8 ~ H z , H8.

In a similar manner the following compounds were prepared. 2-(3'-Fluorophenyl)-3-methoxy-6-(3"-methoxybenzylamino)imidazo[1,2 -b.lpyridazine (GBLD-

216).-The title compound, prepared from 6-(3"-methoxybenzylamino)pyridazin-3-am 2- oxide (0 .25 g) and cx-bromo-3'-fluoroacetophenone11 (0.22 g), as light-yellow needles (0.095 g) had m.p. 159-161" (Found: C, 66.7; H, 5 .3; N, 14.7. C21HlgFN402 requires C, 66.7; H, 5 .1; N, 14.8%). IH n.m.r. (CDC13) 6 3 .80 , s, 3"-OMe; 4 .06, s, 3-OMe; 4 .07 , d , J 5 .5 Hz, CH2; 4 .88, t , J 5 .5 Hz, NH; 6 .39, d, 57,s 9 HZ, H7; 6.76-7.91, complex, H2~,4',5/,6',2",4",5",6"; 7 .52, d , 5 7 3 9 HZ, H 8.

2-(4'-Fluoropheny1)-3-methoxy-6-(3" - methoxybenzylamino)imidazo[l , 2 -b]pyridazine (GBLD- 274).-The crude product from 6-(3'-methoxybenzylamino)pyridazin-3-amie 2-oxide (0 .2 g)

" Lutz, R. E., Allison, R. K., Ashburn, G., Bailey, P. S., Clark, M. T., Codington, J . F., Deinet, A. J., Freek, J. A., Jordan, R. H., Leake, N. H., Martin, T. A., Nicodemus, K. C., Rowlett, R. J., Jr, Shearer, N. H., Jr, Smith, J. D., and Wilson, J. W., 111, J. Org. Chem., 1947, 12, 617.

G. B. Barlin, L. P. Davies and M. M. L. Ngu

and a-bromo-4-fluoroacetophenoneH with subsequent methylation gave, after t.1.c. (alumina; chloroform) and recrystallization from toluene/cyclohexane, the title compound (0 -050 g), m.p. 161-163" (Found, for a sample dried at 80°/0. 1 mmHg for 12 h: C, 67.2; H, 5 .1 ; N, 14.5. C21H19FN402 requires C, 66.7; H, 5 .1; N, 14.8%). 'H n.m.r. (CDC13) 6 3.79, s, 3'-OMe; 4.04, s, 3-OMe; 4 .56 , d , J C H , N H 5 HZ, CH2; 4 .91, t, J C H , N H 5 HZ, NH; 6 .37, d, J 7 , g

9 Hz, H 7; 6 .76-8 .12, complex, H 21,31,51,61,211,411,511,611; 7.5 1, d, J 7 , g 9 HZ, H 8. 3-Methoxy-6-(3'-methoxybenzylamino)-2-(31'-trifluoromethylphenyl)imidazo[l,2-b1pyridazine

(cs~~-272).-6-(3~-Methoxybenzylamino)pyridazin-3-amine 2-oxide (0.25 g) and a-bromo-3-tri- f l ~ o r o m e t h y l a c e t o p h e n o n e ~ ~ ~ ~ (0.28 g) as above eventually gave yellow crystals of title compound (0.12 g), m.p. 144-145" (from toluene/cyclohexane) (Found: C, 62.0; H, 4 . 6 ; N, 13.0. C22HlgF3N402 requires C, 61.7; H, 4 .5 ; N, 13.1%). 'H n.m.r. (CDC13) 6 3.79, s, 3'-OMe; 4 .07, s, 3-OMe; 4 .56, d , J C H , N H 5 HZ, CH2; 4 .95, t , J C H , N H 5 HZ, NH; 6 .39 , d, J 7 , g

9 Hz, H 7; 6.75-8.44, complex, H 21,41,51,6',211,411,511,6"; 7.53, d, J 7 , g 9 Hz, H 8. 3-Methoxy-6-(3'-methoxybenzyIamin0)-2-(4~' -tri~uoromethylphenyl)imidazo[l,2-blpyridazine

(cs~~-281).-In a similar manner, from 6-(31-methoxybenzylamino)pyridazin-3-amine 2-oxide (0 .23 g) and a-bromo-4-trifluoromethylacetophenone4~12 was prepared the title compound (0.140 g), m.p. 151-152" (Found: C, 61.9; H, 4 .6; N, 13.0 . C22HlgF3N402 requires C, 61.7; H, 4 .5; F, 13.3%). ' H n.m.r. (CDC13) 6 3.80, s, 3'-OMe; 4 .07, s, 3-OMe; 4.57, d , JcHSNH 5 .5 HZ, CH2; 4 .86 , t, J C H , N H 5 - 5 HZ, NH; 6 .41, d, J 7 , g 9 HZ, H 7; 6.77-7.38, complex, H2',4',5',6'; 7.55, d, J 7 , g 9Hz, H8; 7.65, d, 8 .19, d, J21~,3~/ 9Hz , H2//,3",5",6".

A mixture of 6-(3'-methoxybenzylamino)pyridazin-3-amine 2-oxide (0.25 g) and a-bromo- 4-methyla~etophenone'~ (0.22 g) in ethanol (10 ml) was refluxed for 5 h. After cooling, the orange precipitate (0 .23 g) was filtered off, washed with water and ether, and dried under vacuum.

This solid was stirred overnight with excess ethereal diazomethane at 0" and then at 20". The mixture was evaporated and the residue subjected to t.1.c. (alumina; chloroform) and the product was recrystallized from a mixture of toluene and cyclohexane to give light-brown crystals of the title compound (0,085 g), m.p. 148-1 50" (Found: C, 70.9; H, 6 .1; N, 14.9. C22H22N402 requires C, 70.6; H, 5 .9; N, 15 .O%). 'H n.m.r. (CDC13) 6 2.35, s, Me; 3.74, s, 3'-OMe; 4 .01, s, 3-OMe; 4 .51, d, J C H , N H 5 . 5 HZ, CH2; 5.29, t , J C H , N H 5.5 HZ, NH; 6 .31, d, 37,s 9 Hz, H7; 6.75-8.01, complex, H2',41,5',61,2u,311,51i,611; 7.42, d , J 7 , 8 9 Hz, H8.

The crude product from 6-(2'-methoxybenzylamino)pyridazin-3-amine 2-oxide (0.25 g) and a-bromo-4-methylacetophenone (0 .22 g) followed by methylation gave, after t.1.c. (chloroform/cyclohexane, 3 : 7) and recrystallization from toluene/light petroleum (b.p. 40-6O0), the title compound (0.04 g), m.p. 136-138" (Found, for a sample dried at 90e/0. 2 mmHg for 8 h: C, 70.8; H, 5.8; N, 15.0. C22H22N402 requires C, 70.6; H, 5 .9; N, 15.0%). 'H n.m.r. (CDC13) 6 2.36, s, Me; 3.82, s, 2'-OMe; 4 .07, S, 3-OMe; 4 .57, d, JCHrNH 5 . 5 HZ, CH2; 5 .16, t, J C H J H 5 .5 HZ, NH; 6 .32, d , J 7 , g 9 HZ, H 7; 6.81-8.02, complex, H 3',4',5',6',2",3",5",6"; 7.45, d, J 7 , 8 9 Hz, H 8.

This compound was prepared from 6-(2'-methoxybenzylamino)pyridazin-3-amine 2-oxide (0- 25 g) and cl-bromo-4-chloroacetophenone12 (0.24 g) as described above. The title compound (0.1 g) had m.p. 131-133" (from toluene/cyclohexane) (Found: C, 63.7; H, 4 . 9 ; N, 13.9. C21HigCIN402 requires C, 63.9; H, 4 . 9 ; N, 14.2%). 'H n.m.r. (CDC13) 6 3.81, s, 2"-OMe; 4.08, s, 3-OMe; 4 .56, d , J C H , N H 5.5 HZ, CH2: 5 .22, t, J C H , N H 5 .5 HZ, NH; 6 .34, d , J 7 , g 9 HZ, H 7, 6.80-7-32, complex, H 3",4",5",6"; 7.42, d, J 7 , g 9 HZ, H8; 7.35, d , 7.99, d , J211,3fl 9 HZ, H 2',3',5',6'.

l2 Blank, B., DiTullio, N. W., Deviney, L., Roberts, J. T., and Saunders, H. L., J. Med. Chem., 1975, 18, 952.

3(and 4)-Nitroacetophenone were brominated with bromine in acetic acid by a method similar to that reported in ref. 13. The a-bromo-3-nitroacetophenone was recrystallized from acetic acid and had m.p. 88-90" [IH n.m.r. (CDC13) 6 4 .50 , s, CH2; 7.66-8.81, complex, H 2,4,5,6.1

The a-bromo-4-nitroacetophenone was recrystallized from glacial acetic and had m.p. 95-97' ['H n.m.r. (CDC13) 6 4.48, s, CH2; 8.16, d , 8 .36, d , 52.3 9Hz , H2,3,5,6].

3-Methoxy-6-(3'-methoxybenzylamino)-2-(3-nitrophenyl)imidazo[l,2-bpyridazine (GBLD-307) and Related Compounds

A mixture of 6-(3'-methoxybenzy1amino)pyridazin-3-amine 2-oxide (0.37 g) and oc-bromo- 3-nitroacetophenone (0 .42 g) in ethanol (15 ml) was refluxed for 0 . 5 h. After cooling to room temperature, sodium hydrogen carbonate (0 .07 g) was added. After effervescence ceased, the mixture was brought to reflux for 3 . 5 h.

After cooling, water (10 ml) was added and the pH adjusted to 7. After chilling in ice, the brown precipitate ( 0 . 5 g) was filtered off, washed with water, ether and dried. This solid was added to a cold solution of ethereal diazomethane and the mixture stirred at 0" and 20" overnight. The crude product was subjected to column chromatography (alumina; chloroform) and the product (0 .1 g) was recrystallized from methanol to give yellow crystals of the title compound (0.095 g), m.p. 186-188" (Found: C, 61.9; H, 4 . 6 ; N, 17.1. C21HlgN504 requires C, 62.2; H, 4 .7 ; N, 17.3%). 'H n.m.r. (CDC13) 6 3.78, s, 3'-OMe; 4.11, s , 3-OMe; 4 .57, d , JCH,NH 5 . 5 HZ, CH2; 5.20, t , JCH,NH 5 .5 HZ, NH; 6 .45 , d , 17,s 9 HZ, H7; 6.76-8.88, complex, H 2',4',5',6',2",4u,5u,6'1; 7 .50, d, 57.8 9 Hz, H 8.

In a similar manner the following compounds were prepared. 3 - Methoxy - 6 - (3' -methoxybenzylamino) -2-(4"-nitrophenyl)imidazo[l,2-b]pyridazie (GBLD-

306).-The title compound (13%) was obtained as orange crystals, m.p. 208-210" (from chloroform/methanol) (Found: C, 62.4; H, 4 .5 ; N, 17.2 . C21HlgN504 requires C, 62.2; H, 4 . 7 ; N, 1 7 3%). H n.m.r. (CDCI3) 6 3.81, s, 3'-OMe; 4.11, s, 3-OMe; 4 .58, d, JCH,NH 5 .5 HZ, CH2; 5.20, t, JCH,NH 5 .5 HZ, NH; 6 .45 , d, 57,s 9 HZ, H7, 6 .81-738 , complex, H2',4',5',6'; 7.57, d , J7,8 ~ H z , H8; 8 .24, b, H2",3",5",6".

3 - Methoxy -6-(2' -methoxybenzylamino)-243 "-nitrophenyl)imidazo[l,2-blpyridazine (GELD- 323).-The crude product from 6-(2'-methoxybenzylamino)pyridazin-3-amine 2-oxide (0.49 g) and cu-bromo-3-nitroacetophenone (0.54 g) followed by methylation gave, after t.1.c. (alumina; chloroform) and recrystallization from ethanol/light petroleum (b.p. 40-6O0), 2 : 1, yellow crystals of the title compound (0.075 g), m.p. 174-176" (Found: C, 62.1; H, 5 .0; N, 17.3 . C21HlgN504 requires C, 62.2; H, 4 .7; N , 17.3%). 'H n.m.r. (CDC13) 6 3 .84, s, 2'-OMe; 4.17, s, 3-OMe; 4 .58, d , JCH,NH 5 . 5 HZ, CH2; 5.25, t, JCH,NH 5 .5 HZ, NH; 6 .42, d, ]7,8 9 HZ, H7; 6.83-8.89, complex, H 3/,4/,51,6i,2/',411,51i,61i.

3- Methoxy -6-(2' - m e t h o x y b e n z y l a m i n o ) - 2 - ( 4 " - n i t r o p h e n y [ i z i e (CBLD- 324).-In a similar manner, was prepared the title compound (25%), m.p. 163-165" [from ethanol/light petroleum (b.p. 40-6O0), 3 : 11 (Found: C, 62 3; H, 4 .7 ; N, 17.3 . C21HlgN504 requires C, 62 .2; H, 4 .7 ; N, 17.3%). lH n.m.r. (CDC13) 6 3.86, s, 2'-OMe; 4 .16, s, 3-OMe; 4.59, d , JCH,NH 5 .5 HZ, CHZ; 5.26, t , JCH,NH 5 .5 HZ, NH; 6 . 4 3 , d, 57.8 9 HZ, H7; 6.85-7.32, complex, H3/,4',5',6'; 7.46, d , 17.8 9 HZ, H8; 8 .2 , br, H2",3u,5u,6/i.

2-(4'-Fluoropheny1)-3-methoxy-6-(2'' -methoxybenzylamino)imidazo[l , 2 -b]pyridazine (GELD- 312).-6-(2'-Methoxybenzylamino)pyridazin-mine 2-oxide (0 .25 g) and a-bromo-4- f l u o r o a ~ e t o p h e n o n e ~ ~ (0.22 g) as above gave the title compound (0.05 g) which had m.p. 135-136" (from toluene/cyclohexane) (Found: C, 66.3; H, 5 .3 ; N, 14.6. C21H19FN402 requires C, 66.6; H, 5 .1 ; N, 14 .a%). lH n.m.r. (CDC13) 6 3.84, s, 2"-OMe; 4 .09, s, 3-OMe; 4 .58, d, JCH,NH 5 . 5 H z , CH2; 5.13, t, JCH,NH 5 .5 HZ, NH; 6 .35, d , 17,s ~ H z , H7, 6.82-8.12, complex, H2',31,5',6/,3/',4fl,5u,6u; 7.45, d , J7,8 9 HZ, H8.

3' - Methoxy - 6-(2' -methoxybenzylamino) -2-(pyridin-3u-yl)imidazo[l,2-b]pyridazine (GBLD- 3101.-When 6-(2'-methoxybenzylamino)pyridazin-3-amine 2-oxide (0.37 g) was condensed with 3-bromoacetylpyridine hydrobromide4 (0.42 g) as above it eventually gave the title compound (0.060 g), m.p. 170-171" (from toluene/cyclohexane) (Found: C, 66.3; H, 5 .4 ;

l3 Long, L. M., and Troutman, H. D., 1. Am. Chem. Soc., 1951, 73, 542.

G. B. Barlin, L. P. Davies and M. M. L. Ngu

N, 19.3 . CzoHigN502 requires, C, 66.5; H, 5 .3; N, 19.4%). 'H n.m.r. (CDC13) 6 3.85, s, 2'-OMe; 4 .12, s, 3-OMe; 4.59, d, JCH,NH 5.5 HZ, CH2; 5 .35, t, JCH,NH 5 .5 HZ, NH; 6 -41 , d, J7.8 9 HZ, H7; 6.83-9.31, complex, H2',41,51,61,211,4",511,6"; 7.48, d, J7,g 9 HZ, H8.

To a rapidly stirred mixture of reduced iron powder (0.08 g), methanol (5 ml), water (1 ml) and concentrated hydrochloric acid (0.24 ml), was added a solution o'f 3-methoxy- 6-(31-methoxybenzylamino)-2-(311-nitrophenyl)imidazo[l,2-b]pyridazine (0 . O S g) in methanol (20 ml), while the temperature was maintained at 85-90". After stirring at the same temperature for 3 h, the mixture was filtered and the solid washed with hot methanol. The combined filtrates were evaporated, the residue was diluted with water and adjusted to pH 7, and the mixture extracted with chloroform. The extract gave an orange residue which was subjected to t.1.c. (alumina; chloroform) to give a brown product (0.03 g) which was recrystallized from toluene to give yellow crystals of the title compound (0.015 g), m.p. 84-86" (Found, for a sample dried at 60°/0. 1 mmHg for 12 h: C, 65.7; H, 5.6; N, 18.3. C21H21N502.0.5H20 requires C, 65.6; H, 5 .8; N, 18.2%). 1~ n.m.r. (CDC13) 6 3 . 4 , br, NH2; 3.78, s, 3/'-OMe; 4.02, s , 3-OMe; 4.54, d , JCH,NH 6 HZ, CH2; 5.00, t, JCH,NH 6 HZ, NH; 6 .34, d, 37.8 ~ H z , H7; 6.53-7.45, complex, H21,41,5',6',2",411,511,6"; 7.48, d, J7,g ~ H z , H8.

In a similar manner to above the title compound (0 .02 g) was prepared as dark brown crystals, m.p. 70-77' (from toluene/cyclohexane) (Found, for a sample dried at 60°/0. 1 mmHg for 6 h: C, 66.4; H, 6 .0; N, 18.2 . C Z ~ H ~ I N ~ O ~ . O . ~ ~ H ~ O requires C, 66.4; H, 5.7; N, 18.4%). 'H n.m.r. (CDC13) 6 3.4, br, NH2; 3.77, s, 3"-OMe; 4 .00, s, 3-OMe; 4 .53, d, JCH,NH 6 HZ, CH2; 5.06, t, JCH,NH 6 HZ, NH; 6 .33 , d , J7,8 ~ H z , H7; 6 .73 , d , 7.86, d, J ~ " , ~ I I 9 HZ, H 2',3',5',6'; 6.65-7.34, complex, H 211,411,S11,611; 7.46, d , J7.g 9 HZ, H 8.

Biological Testing: Receptor Binding Assays

The biological testing of the compounds for their ability to displace 3 ~ - d i a z e p a m from rat brain membrane preparations was carried out as described p r e v i ~ u s l y , ~ and the results are reported in Table 1. The lc5o values were usually determined in a single experiment with 3 or 4 concentrations of test compound, selected to span the predicted icso value; within each experiment assays were performed in triplicate. In all cases, the correlation coefficients of the lines of best-fit to log-logit curves were greater than 0.96.

Acknowledgments

We thank Dr D. J. Brown for helpful discussion and Dr M. D. Fenn for the lH n.m.r. spectra. One of us (M.M.L.N.) thanks this University for support as a scholar.

Manuscript received 13 February 1989