5706
LILLIANLI-YENU'ANGAND MADELEINE AI. JOUILIE
molecules and the bulk material. We have obtained preliminary evidence that denaturation by alkali or acid treatments for brief intervals deforms the DNA sufficiently for the formation of the lead complex. IVe have not yet investigated the pre-
[CONTRIBUTION FROM THE
1'01. 79
cipitability of the products of enzymatic hydrolysis by DNase. Other cations may substitute for lead ion in the test. The effects of other cations or anions are under investigation. BERKELEY,CALIFORNIA
DEPARTMENT OF CIIEMISTRY
O F THE UNIVERSITY OF PRN?iSYI.VANIA]
Synthesis of Bis-benzimidazoles BY LILLIANLI-YENWANGAND NADELETNE M. JOUI,LI@ RECEIVED MAY13, 1957 A number of new bis-benzimidazoles have been prepared. Substituents have been placed on both benzene rings of the benzimidazole nuclei. The chain linking the two benzimidazole units has been varied from simple alkane chains t o substituted alkane chains or aryl chains (benzene ring). It was found t h a t polyphosphoric acid is a useful medium for preparing bis-benzimidazoles. In difficult cases the use of a little concentrated sulfuric acid as a catalyst was beneficial.
The benzimidazole nucleus has been of consider- further modification was made. It was found that able interest since it was noted that benzimidazole the bis-benzimidazoles could be conveniently iioinhibits the growth of certain yeasts and bacteria.2 lated as their dihydrochlorides b y cooling the reThe discovery of 5,6-dimethylbenzimidazoleas a action solution after the refluxing period. The isounit in vitamin Blz has increased this interest. A lation of the dihydrochloride was preferable to neunumber of alkyl benzimidazoles have been tested tralization of the whole solution because in general and found t o have some anti-vitamin BIZactivity the dihydrochloride of the diamine is more soluble and some have been reported to have anti-viral ac- than the dihydrochloride of the bis compound. tivity also. T h a t structural modifications can pro- Thus a separation of most of the excess diamine duce marked effects on physiological activity has can be accomplished in this step. The dihydrobeen shown b y the test data on the substituted chloride was then treated with hot bicarbonate sobenzimidazoles and naphthoquinone imidazoles lution or aqueous ammonia to remove any heniiwhich have been synthesized in this L a b ~ r a t o r y . ~midazolylalkylcarboxylic acid which would norSome of these compounds exerted strong inhibitory mally be present. action not only against purine and €312 requiring This modification of Phillips' method gave fair tnicroorganisms but against folic acid-requiring yields in most cases. It failed, however, when used organisms as well. to condense 3-nitro-5-chloro-o-phenylenediaiiii11e Structural modifications of the benzimidazole with succinic, adipic and suberic acids. The polynucleus can be carried out in several ways. I n the phosphoric acid method, recently reported by present inveqtigation methods were developed to Hein, Alheim and Leavitt for the preparation of synthesize new bis-benzimidazoles where the two simple benzimidazoles, was then tried.' benzimidazole nuclei are united through their 2The polyphosphoric acid method not only worked positions either directly or through one or more for 3-nitro-5-chloro-o-phenylenediamine with suratonis. Such systems can be modified not only by cinic, adipic and suberic acids b u t was found to be changing the nature and the number of the con- convenient and fairly general giving about the necting atoms b u t by changing the nature of the cane yields as the modified Phillips method. The substituents in the benzimidazole nuclei as well. polyphosphoric acid method failed in one case, with h review cif the literature revealed the fact that a 5-nitro-o-phenylenediamine. number of his-benzimidazoles have been reported I n the preparation of the bis-benziniidazolyletlihut apparently none of them have been screened anes, although 3,4-diaminotoluene and 4-chloro-ofor physiological activity. In the present study, phenylenediamine were condensed with succinic greatest emphasis was placed on the preparation of acid to give fair yields of the bis compounds, it was 5ubstituted his-benzimidazolylalkanes although a found preferable to use succinic anhydride instead of few other types were also prepared. the free acid for the other substituted diamines. Phillips' method for the preparation of simple As the alkane chain between the two benziinibenzimidazoles is well known.5 The same proce- dazole rings increased in length, the preparation of (lure can be used to prepare bis-benzimidazoles by bis-benzimidazoles became more difficult. The use refluxing two moles of diamine with one mole of a of polypliosphoric acid was the best method for the dibasic acid in 4 N hydrochloric acid. The method condensation of o-phenylenediamines with the was modified by Shriner and Upson by using a longer chain dibasic acids. Substituted bis-benzimuch longer reflux period.6 In the present work a midazolyloctanes were not easy to obtain even by (1) To whom all inquiries should be addressed. this method until it was noted that the addition of (2) D. W. U'oolley, J . Bid. Chem., 162, 225 (1944). a sinall amount of concentrated sulfuric acid facili( 3 ) I. T a m m , K . I'olkers and F. L. Horsfall, J . E x p t l . M e d . , 98, 219, tated the reaction. Although this niodification was 229, 24.5 (1953). not tried with all of the other dibasic acids, it would f4) Progress Report, July 1955-Jan. 1956, U.S.P.H.S. Cknnt C 2189, University of Pennsylvania, Philadelphia 4, Pa. probably be helpful in those cases also. ( 5 ) sf. A. Phillips, J . Chem. SOL.,2393 (1928). (6) K. L. Shriller and R. \V. Upson, THISJ O U R N A L , 63,2277 (1941).
(7) 11 W. Hein, R J. Alheim and A. A. Leavltt, rbzd., 79, 427 (1957).
Nov. 5 , 1957
SYNTHESIS OF
5707
BIS-BENZIMIDAZOLES
TABLE I BIS BENZIMIDAZOLES
SubstitueFts R R H C€It H NO; H CI H CnHrO N H t C1 H CHI H Nor H CI H NHz H NHz.2HCI NO1 Cl NHr CI H CHI NO, H H C1 NOS CI H CHI H C1 H H H CHr H CtHrO c1 H H NO2 H H H
C1
30 31 36 42 35 45 35 29 38 65 25 25 28 30 33 10 40 40 44 42 35 39 30 33
292-296 288-29of 247-250f 221-223 243-245 204-207 2825 226-228 208-210 245f 218-221 222-225 218f 250-256/ 200
-Calcd.C 74.45 54.55 58.03 68.54 53.19 75.44 56.84 60.17 67.50 46.36 48.13 55.55 76.25 58.81 62.03 50.32 76.97 63.61 65.31 67.06 62.80 52.91 49.99 67.07
20
365f
63.34
Yield,
Connecting unit CHxCHz CHzCHa CHzCHz CHrCHz CHaCHz (CHdr (CH2)4 (CHa)r (CHa)r (CHdr (CHa)r (CHdr (CHds (CHds (CHds (CHds (CHds (CHn)s CHOHCHOH CHOHCHOH CHOHCHOH CHOHCHOH CHOHCHOH
%d
!
M.P., OC . 265-268 290-293 270-273 225-228 256-258 235-236 267-269 276-278
-
Anal)rses. %
H 6.25 3.44 3.65 6.33 3.90 6.96 4.34 4.49 6.29 5.19 3.41 4.66 7.57 4.94 5.21 3.80 8.07 5.82 4.80 5.63 5.80 3.53 3.15 5.63
N C 19.30 23.58 16.92 15.96 23.26 17.60 22.05 15.59 26.23 18.02 18.71 21.59 16.18 20.57 14.47 17.61 14.96 13.49 19.04 17.38 14.65 15.43 21.87 17.38
3.19
14.77
1
,
..
...
21.42
...
19.62
.. .
... ... ...
30.42 15.78 18.22
... ...
18.31 14.85
...
... ... ... ...
19.53
,..
9.940 18.70
--
C 74.60 54.59 57.88 68.79 53.23 75.62 56.79 59.91 67.41 46.32 48.33 55.35 76.07 58.59 61.87 50.12 76.96 63.54 65.35 66.87 62.97 52.71 49.89 66.99
Found -H N C I 6 . 3 8 19.44 ... 3 . 5 3 23.77 ... 3 . 5 1 16.78 21.22 6 . 3 7 15.69 ... 4.04 23.39 19.70 7 . 1 1 17.45 4 . 3 2 22.05 ... 4.65 15.38 ... 6.16 26.00 .. 5.11 18.00 30.52 3 . 5 5 18.54 15.54 4.87 21.47 18.21 7.76 16.36 4 . 9 1 20.64 . 5 . 2 7 14.57 18.19 3 . 7 1 17.57 14.72 8 . 2 1 14.82 5 . 8 7 13.52 ... 5 . 0 8 19.06 ... 5 . 4 9 17.21 ... 6.02 14.56 ... 3 . 5 8 15.59 19.62 ... 3 . 0 8 21.84 5 . 5 0 17.33 10.06@
63.36
3.22
...
...
..
...
a Prepared .y method A. Prepare1 by rnetho B. Listed but not fully described in Swiss Patent. products. e Decomposes indefinitely 1 1-225". I Iecomposes. 0 Sulfur, 7'.
14.80
18.63
Yields of pure
A modified Phillips procedure was also used for was then cooled, diluted with water and allowed to stand The precipitate was removed by filtration, the preparation of 1,2-bis-benzimidazolyl-l12-eth-overnight. extracted with hot, dilute ammonium hydroxide and dried. anediols and 2,2'-(thiodiethylene)-bi~-benzimida- The product was recrystallized from ethanol-water. zoles. The polyphosphoric acid method was not The Modified Phillips Method. Method B. 1,Z-Bis(6-nitro- 2-benzimidazoly1)-ethane.-4- Nitro-o-phenylenetried in these cases. For the preparation of 2,2'-o-phenylenebis-(6- diamine (7.65 g., 0.05 mole) and 3.05 g. (0.025 mole) of succinic acid were refluxed for ten hours in 50 ml. of 4 N chlorobenzimidazole) only the polyphosphoric acid hydrochloric acid. Succinic anhydride may be used in method was successful. This is in agreement with place of succinic acid. The reaction mixture was cooled arid the observation of others that Phillips' method is the crystalline dihydrochloride which separated was renot a successful one for the condensation of o- moved by filtration. It was extracted with hot dilute ammonium hydroxide and washed with water. The product phenylenediamines with aromatic acids. was recrystallized from ethylene glycol with the aid of deCrippa, Perronato and Sacchetti reported the colorizing carbon. In all other cases where method B was formation of bis- (2 ,2 '-benzimidazolyl) -thiourea used, the product was recrystallized from ethanol-water from 2-aminobenzimidazole when the latter was with the aid of decolorizing carbon, unless otherwise noted. 1,Z-Bis-(6-ethoxy-Z-benzimidazolyl)-ethane.-In this heated with carbon disulfide in dry ethanol.8 case the bis derivative was made from 3-amino-4-nitroThe reaction was repeated, in the present study, phenetole without isolating t h e intermediate diamine. 3but the thiourea derivative was not obtained. Amino-4nitrophenetole (9.1 g., 0.05 mole) was dissolved The only product isolated proved to be ethyl N- in 50 ml. of 4 N hydrochloric acid, 1 g. of 5y0palladium-onalumina was added and hydrogenation carried out in a Parr benzimidazolyl thiocarbamate. apparatus. The catalyst was removed and 2.5 g. (0.025
Experimental
All of the melting points reported were taken in an apparatus similar t o the one described by Wagner and Meyer.9 The values are uncorrected. Preparation of Bis-benzimidazolyl Ethane. Polyphosphoric Acid Method. Method A. l,Z-Bis-(6-methyl-Zbenzimidazolyl)-ethane.-3,4-Diaminotoluene (6.10 g., 0.05 mole) was mixed with 2.98 g. (0.025 mole) of succinic acid and the mixture poured into 30 ml. of polyphosphoric acid which had been preheated to 100'. T h e mixture was stirred and heated a t 150' for three hours. T h e reaction mixture (8) G. B. Crippa, G. Perronato and G. Sacchetti, Gasa. chim. ifaf,, 66, 38 (1935). (9) E. C. Wagner and J. F. Meyer, I n d . Esg. Chem., Anal. Ed., 10, 584 (1938).
mole) of succinic anhydride and 2 ml. of concentrated sulfuric acid were added. Method B was followed from this point. 1,Z-Bis-(7-amino-5-chloro-2-benzimidazolyl )-ethane This bis derivative was prepared from 3-nitro-5-chloro-ophenylenediamine b y t h e procedure described for 1,2-bis(6-ethoxy-2-benzimidazolyl)-ethane. I n this case the crude dihydrochloride was treated with hot sodium bicarbonate solution in place of aqueous ammonia. It was washed with water and dried. Attempts to recrystallize the product from ethylene glycol gave a gum which changed t o a crystalline solid when treated with anhydrous acetone. (7-Nitro-5-chloro-2-benzimidazolyl)-3-propanoicAcid,Attempts to prepare 1,2-bis-(7-nitro-5-chloro-2-benzimidazoly1)-ethane from 3-nitrc-5-chloro-o-phenylenediaminewere unsuccessful with both methods A and B. The only prod-
.-
5705
PRICETRUITT, JAMES ERWIN COOPER, 111, AND FRANK M. U'OOD,JR.
Vol. 79
uct t h a t could be isolated was tlie benzimidazolylpropionic Preparation of Bis-benzimidazolylhexanes.-Suberic acid acid. Since this is a new compound its preparation is iri- \vas the starting organic acid for this series. cluded. Method -4 was used, b u t in this case the polyBis-benzimidazolyloctanes were prepared from sebacic phosphoric acid was preheated to 150" and after adding the x i d and the appropriate diamines. diamine t h e misture x a s kept a t this temperature for five 1,8-Bis-( 6-chloro-2-benzimidazolyl)-octane .--In this case, hours. On cooling the reaction rnisture and dilutiug with 1 inl. of concentrated sulfuric acid was added t o the polywater a tar formed from which no identifiable product could phosphoric acid and the reaction mixture was stirred for be obtained. T h e filtrate from the tar was neutralized and five hours at 150'. t h e yellow product which formed was removed and washed Preparation of Bis-benzimidazolyl-1,Z-ethanedio1s.-The with mater. It was recrystallized from ethanol-water with stafting materials mere tartaric acid and o-phenylenedithe aid of decolorizing carbon, yield 4376, m.p. 240-242'. amines. 1,2-Bis-( 6-ethoxy-2-benzimidazolyl)-l,Z-ethanediol~ v d b A n d . Calcd. for CloHsS30aCl: C, 44.52; H, 2.99; K, 15.58; C1, 13.15. Found: C , 44.68; H, 3.12; K, 18.02; p r e p x e d from 3-amino-4nitrophenetole and d-tartaric acid by the procedure described for making 1,2-bis-( 6-ethoxy-2C1, 13.26. Preparation of Bis-benzimidazolylbutanes. 1,4-Bis-(2- benziinidazoly1)-ethane. 1,2-Bis-(6-nitro-2-benzimidazolyl)-1,2-ethanediol was rebenzimidazoly1)-butane.-This compound was iiiltde earlier by Shriller a n d Upson from o-plienylenedial7iine and adipic crystallized from ethylene glycol-water with the aid of tlcacid. It was made in this work, by both inetliuds X and B colorizing carbon. Preparation of 2,2'-(Thiodiethylene)-bis-benzimidazole.t o get samples for t h e screening program. Tlic yields n-cre This compound was prepared from o-phenylenediamine and around 40;: b y both methods. 1,4-Bis-(6-amino-2-benzimidazolyl)-butane.--1 ,l-Bis-(F- 3,3'-d ithiopropionic acid .lo Z,Z'-o-Phenylenebis-(6-chlorobenzimidazole) w a s prcnitro-2-benzimidazolyl)-butane(3.8 g., 0.01 mole) was a n d phthalic anhyadded to 20 ml. of 4 N hydrochloric acid and 0.5 g. of 5% pared from 4-chloro-o-phenylenediamine dride. The crude product was washed with hot 95y0ethanol. palladium-on-alumilia added. TVhen the hydrogenation was recrvstallized from N-dimeth- yield) . was complete, 50 nil. of 4 N hydrochloric acid was added T h e residue (43% and t h e solution warmed. T h e catalyst was removed and vlformamide. PreDaration of Ethvl N-Benzimidazolv1thiocarbamate.tlie dihydrocliluride separatcd from the filtrate 011 cooling. It was recrystallized from 4 S hydrochloric acid. Method 2-Amii;obenzimidazole (13.3 g.) was mixed with 40 ml. of carbon disulfide and 30 ml. of absolute ethanol. T h e re13 w a i followed from this poiiit. action mixture was refluxed on a ?team-bath for 50 hours 1,4-Bis-(7-nitro-5-chloro-Z-benzimidazolyl)-butane .-Tiii.; compound was prepared by method A. The tcmpei-aturc .tiid the product was isolated and purified as described by Crippa, et nZ.* T h e analytical data indicated t h a t the com~ n kept s a t 130' for five hours. Tlic crude product was pound mas a thiocarbamate rather than thc bis derivative estracted with hot 105; sodium bicarbonate solution, dried reported in reference 8. T h e yield was 407,, 1n.p. 202" and recrystallized from ethj-lene glycol. AmZ. Calcd. for C l ~ H ~ l S B S OC:, 51.27; 13, 5.01; 1,4-Bis-(7-amino -5-chloro-Z-benzimidazolyl)-butane. --This bis-benzimidazole was prepared from 3-tiitrc1-5-cIilor1)- S, 19.00; S, 14.50. Found: C, 54.54; -IT, 3.15; N, 19.09; o-phenylenecliainine by reducing it in 4 S hydrocliloric s, 14.64. acid over pall:idiurn-on-:iluiniii:i. The filtrate from tlic (10) T h e 3,3'-dithiopropionic acid was obtained from t h e American catayst was refluxed with aciipic acid for ten hours a s in Cyanamid Co. t h e ureuaration of 1. ~ - b i ~ - i i - a n l i n o - ~ - c ~ i ~ ~ ~ r ~ ~ - ~ - ~ c l l z i l ~ l i d a -
[COSTRIBUTION FROX THE CHEMICAL
LABORATORY O F NORTH
T E X A S STATE COLLEGE]
Benzo [d]pyrido [a] benzimidazole-4,9-quinone BY PRICETRUITT, JAMES ERWINCOOPER, 111,' AND FRANK M. WOOD,J R . ~ RECEIVED MAY25, 1957 The reactioii of 2-arninopyridine and 2-acetamid~~-3-chloro-l,~-uaplitlio~uinone gave the title compound. T h e same substance was obtained from the reaction of 2-;iminopyridine and 2,3-dichloro-1,4-naphthoquinone.
2-Xceta1~iido-3-cliloro-l,4-11aplitlioquiiiorie reacts action with 2-chloroacetamido-3-chloro-1,4-naphreadily with primary amines t o yield 2-acetamido- thoquinone, an orange product (111) was obtained, m.p. 306'. It did not contain chlorine. A mixed 3-alkyl(aryl)amino- 1,4naphthoquinones.3 , 4 Since Calandra and Adamsj had reported that melting point determination proved that the two the reaction of 2-aminopyridine and 2,3-dichloro- substances melting a t 306' were identical. The reaction of 2-aminopyridine (11) and 2,3-di1,4-naphthoquinone yielded a product described as (IV) in refluxing ethanol 2-(2-pyridylamino)-3 -chloro- 1,4-naphthoquinone, chloro-1,4-naphthoquinone it seemed reasonable that 2-aminopyridine should did indeed give 2-chloro-3-(2-pyridylamino)-1,4naphthoquinone (I-),m.p. 275-27G0, as Calandra react with 2-acetamido-3-chloro-1,4-naphthoquinone to give 2-acetaniido-3- (3-pyridylamino)- 1,4- and Xdams i n d i ~ a t e d . ~ naphthoquinone. When 2-acetamido-3-cliloro-l,4- X suspension of V in glacial acetic acid gave I11 naphthoquinone (I) and 2-amiiiopyridine (11) were when heated and cooled. A solution of I and I1 were refluxed for 12 hr. in heated together in refluxing butanol, an orange product (111),m.p. 30B", was obtained. It did not methanol and VI was obtained as orange crystals, contain chlorine. Again, when the 2-acetamido-3- m.p. 21fi0, along with unidentified red platelets. chloro-l,4-naphthoquinonewas replaced in the re- VI was identified as 2-chloro-3-hydroxy-l,4-naphthoquinone.'j These workers reported VI t o melt f I ) S a t i n i i a l Science I'rerluctoral f'ello\v, 1B54-1935 a t 215+'. VI reacted with aniline to yield VII, ( 2 ) Research Fellow of Parke, Davis Sr Co., 1'350-1953. 1n.p. 1%'. 2-Anilino-3-hydroxy-1,4-naphthoqui( 3 ) K. Fries and K. Billig, Bcv., 58, 1128 (1925). ( 4 ) J . R . IS. IIoover a n d A. R . Day, THISJocRx,\r., 76, 11 18 none is reported to melt a t 183°.6 (1'354). ( 5 ) J. C. Calandra and E. C. Adams, ibid., 72, 4804 (lY50).
((j)
T Zincke and C. Gerland, B e ? , 20, 3622 (1867)
