July 1965
ASTITUMOR FLUORENE DERIVATIVES
Hyperchromicity (Table It).-Alkaline digestion could not be used for those compounds with a 3-methyluracil residue since this moiety undergoes degradation in alkali.1° Hyperchromicity was determined by measuring t,he optical density a t the hmaxof a solution of the diribonucleoside phosphate at’ p H 6.0 before and after digestion with purified snake venom diesterase (Worthington Biochemicals). The digestion was carried out as follows. To 2 nil. of an aqueous solution of the diribonucleoside phosphate was added 0.2 ml. of 0.1 Jf Tris buffer ( p H 8.2), 0.025 ml. of 0.1 .\I l I g S 0 4 solution, and 0.1 ml. of a freshly prepared enzyme solution ( 5 mg./ml.). Incubation was carried out a t 37” in a jacketed compartment of a Cary Model 14 spectrophotometer reand was judged complete when the optical density at A,, mained steady for several hours. I n one experiment &methyluridylyl-(3’+5’)-5-methyluridine was hydrolyzed with 0.5 -\NaOH and the hyperchromicity measured a t p H 6.0 was similar to that obtained by use of the enzyme. I n separate experiments the effectiveness of the enzyme treatment was tested on more concentrated samples of each of the diribonucleoside phosphates under similar ronditions. The products of the digestion were sepmtted by electrophoresis and in no rase could t,he unchanged diribonucleohide phosphate be detected. Ribonuclease Treatment.-To the diribonucleoside phosphate (0.2mg.) in 100 h of water was added 25 h of 0.1 ill Tris buffer solution p H 7.0 and 25 A of pancreatic ribonuclease solution (1 mg./ml. of Worthington crystalline, Code R), and the solution
49 1
was incubated at 37” for 15 hr. The reaction mixture was examined by means of elect,rophoresis. The following compounds were not hydrolyzed: 3-methyluridglyl-( 3’-.5’)-3-methyluridine, 3-methyluridylyl-i3‘+5’)uridine, and 5-methyluridyly1-( 2 ’-5’)-5-methyluridine. The following compounds were completely hydroyzed to the 3’ nucleotide and nucleoside: uridylyl-(3’+5’)-3-11iethyluridine and %methyluridylyl-( 3’+5’)-5-methyluridine. When a mixture of equal parts of 5-methgluridylyl-(2’-+5‘)-5methyluridine and 5-met~hyluridylyl-(3’+5’)-5-methyluridine was subjected to these hydrolytic conditions, no breakdown products were detected.
Acknowledgments.-The
authors wish to thank Dr.
J. J. Fox for samples of 1-sylofuraiiosylthymine and 1-arabinofuranosylthyniine. They would also like to acknowledge the continued interest and encouragement of Dr. C. A. Sichol in this program. The authors would like to thank the Cancer Chemotherapy Sational Service Center for a gift of chemicals. This research was partially supported b y a grant from the Kational Cancer Institute of the United States Public Health Service (CA-05697).
Derivatives of Fluorene. XXI. New Halogenofluorenes. 11.’” Further Potential Antitumor Agents HSI-LUNGP A S
.4SD
T. LLOYDFLETCHER’~
Chemistry Research Laboratory of the Department of Surgery, Unibersity of Washington School qf Medtcine, Seattle, Jlrashinylon
98105
Received February 15, 1965 Further polyhalofluorene derivatives have been found effective against. certain tumors in mice, and data are On the basis of present data, the most active compresented for some of the compounds reported (Adenocarcinoma 75,5), N-2-( 7-bromopounds in this group are: N-2-(1,3-dichloro-7-1iitrofluorenyl)acetamide 1,3,4-trichlnrofluoreiiyl)acetamide(Sarcoma 180), Pi-2-( 1,3,7-trichlorofluorenyl)acetamide(Cloudman melanoma), 1,3-dibromo-7-nitro-2-fluorenamine (Adencarcinoma and Sarcoma), 1,3,7-tribromo-2-fluorenamine (Adenocarcinoma), and X-24 1,3,4,6,7-pentachlorofluorenyl)acetamide(Sarcoma). The position of t’he chlorine atom, designated earlier1&as 4(?) on the basis of infrared absorption, is confirmed by synthesis. Some compounds we described as 2,3,7-trisubstituted fluorenes are shown t o be 2,4,7-derivatives, in agreement with recent reports,2,3 but still in marked disagreement with an earlier paper.‘ 2,7-Dibronio-, -dichloro-, and -difluorofluorenes and the three corresponding 9-oxofluorenes are all nitrated at the 4-position. Chlorination of N-2-( 4chlorofluorenyl)acetaniide, S-2-fluorenylformamide, and ethyl N-2-fluorenylcarbamate, as found earlier la with ?r’-2-fluorenylacetamide, leads to the 1,3,4,7-tetrachloi-o compounds. 2,3,7-Trichlorofluorene is prepared by three routes. Acetylation of some relatively weak amines by merely shaking a t room temperature with glacial acetic acid is described. Bromination of Tu’-247-bromofluoreny1)acetamide wit,h N-bromosuccinimide in dimethylformamide gives the 3-bromo derivative.
Our previous report in this seriesla included a number of polyhalogenated fluorene derivatives which show antitumor activity in animals. In expanding this area, in a search for further conipourids with biological activity, we have first confirmed the supposed 4-position for a number of these substances. In all cases in the previous paperla the position designation 4(?) is, in fact, 4. In addition to this niirior point, we wish, in the present paper, to clarify a discrepancy in the literature and, in so doing, correct a position designation in the names of a number of the reported compounds in the previous paper. In brief, eight di- and trichlorofluo(1) l a ) For Halogenofluorenes. I. see H.-L. Pan and T. L. Fletcher, J . .lied. ( ‘ h e m . , 7 , ;31 (1964). (h) Slipported in part h y a grant (Ca-017111 froill t h e rs‘ational Cancer Institute, h-ational Institutes of Healtil. a n d i n p a r t hj- Research Career Development Auard d-Ii3-G~I-14,SYI (T. I.. F , ) , 1 2 ) 11.. Schidlo a n d A . Bieglita, Chem. Ber., 96, 2595 (1963). ( 3 ) .%. Sieglitz, i b d , 97, 3392 (1964). (4) I n particular see paragraph 2 and footnotes 6-8 in ref. l a .
TABLE I No.‘b
Correct name
XIX XX XXI LII LIII LXXIII
9-Bromo-2,4,i-trichlorofluorene 2,4,7-Trichlorofluoren-9-ol 9-0xo-2,4,7-trichlorofluorene 4-$mino-2, 7-dirhlorofluoren-9-01 2,7-Dichloro-Q-oxo-4-fluorenamine 2,4,7-Trichlorofluorene In text only 2,7-Dichloro-4-nitrofluorene I n text only 2,7-Dichloro-4-fluorenamine See ref. la, Table 11, or text.
rene derivatives (shown in Table I), nanied 2,3,7substituted fluorenes, are 2,4,7-substituted fluorenes. As noted,la there are discrepancies apparent in comparing some of our conipounds with those of the same iianie described by Tiretov, et aL4 The latter claimed to have obtained the 3-nitro derivative, b y nitrating 2,7-dichlorofluorene, and from this 2,3,7-trichloro-
NO2
NH,
x = ('1,
I
E'
c1
X = H,C1, F
fluorciie. We fiiid, however, that riot only 2,T-dichlorofluorcwc hiit also the 2,7-dibronio and -difluoro derivat ivva as wrll as thv i*orresponding 2,7-dihalo-!3-osofluorcnes nitrate i i i the 4-position. While this itoil.; was i n progrcss, further coufirination of the above appeared*: in the latter, 4-carboxyfluoreiie aiid 4-chloro-
We leaned heady oii infrared spcct ra'" t o support our belief that 111aiiy of our tri-, tet ra-, mid peiitachlorofluoreiic~delivat ivc5 \\ere suhstitutcd iii thci 4-posit io11 1)yoiic of the c.liloriiic~atoilla. 111 this report n e p r c ~ 1 1 1 vhciiiical cvidciiide tliat proves this l o bc true as sho\vIi iii
Scheiiic 111. Syiithesis of ~--acetaiiiido-4-chloro-
ANTITUMOR FLUORENE DERIVATIVES
July 1965
493
2,7-Dichloro-4-nitro-9-oxofluorene. A.-2,7-Dichloro-9-oxofluorene7 (30 g., 0.12 mole) was added portionwise to a stirred mixture of acetic acid (600 ml.) and 90% HNO3 (600 ml.). The
mixture was heated to 50", then concentrated H2SOa (120 ml.) was introduced a t such a rate that the temperature remained below 55'. After all the sulfuric acid had been added, the mixture was stirred a t 50-55' for 10 min. and cooled. Water dilution, filtration, and recrystallization from acetone gave 29 g. ( 8 2 % ) ; m.p. 187-187.5' (lit.z m.p. 187"); vmag 1724 (ketone C=O), 840 (two adjacent hydrogens) cm.-l. Anal. Calcd. for CIaH,Cl?N03: C, 53.09; H, 1.71. Found: C, 53.25; H, 1.57. B.-2,7-L)ichloro-4-nitrofluorene was oxidized in acetic acid with sodium dichromate giving 98% of material, n1.p. 186-187". The melting point was not depressed when mixed with the product obtained in A. 2,7-Difluoro-4-nitro-9-oxofluorene.A.-2,7-Difluoro-9-uxofluorenes8 was nitrated as described above giving a iiC;, yield; n1.p. 168.5-169' (acetone); vmax 1724 (ketone C=O), 835 (two adjacent hydrogens) cm.-'. Anal. Calcd. for C13HjF?N03: C, 59.78; H, 1.93; X, 5.36. Found: C, 60.01; H, 1.82; N , 5.69, 5.49. B.-2,i-Difluoro-4-nitrofluorene was oxidized n i t h dichroniate to give a 95y0 yield of the product in A. 2,7-Difluoro-9-0~0-4-fluorenamine.-Thenitro compound was reduced with SnC12.2H20 and concentrated HCI giving an 80cjb yield, m.p. 220-221" (toluene). Anal. Calcd. for ClaH7F2NO: C, 6i.53; H, 3.05. Found: C, 67.79; H, 2.99. 9-0~0-2,4,7-trifluorofluorene.-The preceding aniine was converted to the trifluoro compound by a Schiemann reaction. The melting point and mixture melting point with an authentic samplegbwere the same and infrared spectra were identical. 2,7-Dibromo-4-nitro-9-oxofluorene. A.-2,7-Dibromo-9-oxofluorene was nitrated in the same manner as the 2,7-dichloro and 2,i-difluoro analogs giving a 10% yield; m.p. and m.ni.p. (with the following product) 196-197"; vmax 1724 (ketone C=O), 838 (two adjacent hydrogens) cm.-1. B.-2,i-Dibromo-4-nitrofluorene was oxidized as described for the preceding analogs giving a 98% yield, n1.p. 196-107" (toluene-ethanol). Anal. Calcd. for Cl3HjBr2NO3:N, 3.66. Found: N, 3.52. N-4-( 2,7-Dichloro-9-oxofluorenyl)acetamide.-2,i-I~ichloro-l1o~o-4-fluorenamine'~ (see Table I ) was acetylated in the usual manner, m.p. 312-313" (acetic acid). Anal. Calcd. for ClaH,Cl,N02: C, 58.85; H, 2.96; N, 4.58. Found: C, 58.82; H, 2.94; N,4.69. 2,7-Dichloro-4-nitrofluorene.-2,7-Dichlorofluorene1*(25.5 g., 0.11 mole) was suspended in acetic acid (450 ml.). To the stirred suspension, heated a t 3 5 O , a mixture of concentrated HSS04(25 ml.) and HN08 ( d 1.42) ( 2 5 ml.) was added dropwise over a period of 20 min. The suspension was stirred for 5 more min., then heated a t 70" for 15 min. and cooled. The product was filtered, washed, and dried giving 28 g. (94'/), n1.p. 180181". Recrystallization from acetic acid gave an analytical sample, m.p. 180.5-181° (lit.2m.p.179-180'). Anal. Calcd. for Cl3HiCl2NO?:N, 5.00. Found: N , 4.76. 2,7-Difluoro-4-nitrofluorene.-2,7-~ifluorofluorenegwas nitrated as described above giving 87-9253 yields, m.p. 136-137" (methanol). Anal. Calcd. for C13H,F1N02: C, 63.16; H, 2.85; N , 5.67. Found: C, 63.01; H,2.93; N, 5.61. 2,7-Dibromo-4-nitrofluorene.-2,7-l~ibro~iiofl~orene~~ wits also nitrated ( 7 7 7 4 , m.p. 194.5-195.5' (t'oluene-ethanol). Anal. Calcd. for ClaHiBr2N02:C, 42.31; H, 1.90; Br,43.31; N , 3.80. Found: C, 42.17; H, 1.90; Br, 43.28; N, 3.96. 2,7-Difluoro-4-fluorenamine.-2,7-Difluoro-4-nitrofluorene (2.5 g.) was dissolved in a mixture of toluene (100 nil.) and 95% ethanol (250 ml.) and reduced with 8 5 5 hydrazine hydrate (5
( 5 ) Ng. Ph. Euu-Hoi, Rec. trao. chim., 73, 197 (1Y51); T. H. Chao and L. P. Cipriani, J . Ore. C h e m . , 26, 1079 (1961); we are indebted t o 11. J. S a n i k u n g for pointing out this reaction. (6) All melting points below 250° were taken on a Fisher-Johns block and are corrected t o standards. Melting points above 2.50' were taken with a Hoover capillary melting point apparatus and are uncorrected. Analyses were performed b y Schwarzkopf Microanalytical Laboratory, Woodside, N . Y., and b y A . Bernhardt, Miilheim (Ruhr). T h e interpretation of infrared absorption spectra (obtained on a Beokman IR-5, 1.6-2 mg. of substance/300 mg. of KBr) is based on L. J. Bellaniy, "The Infrared Spectra of Complex Rlolecules," 2nd E d . , John Wiley and Sons, Inc., New York, N. Y., 1958. All chlorinations were carried out using a 3Iatl1eson 620 BMV flou-meter. (7) J. Schmidt and H. Wagner, Ann., 337, 147 (1912).
( 8 ) (a, Made b y 31.J . Kainkung of tliis laborators h y sodiuiii dicliroiiiate oxidation of 2,7-difuorofluoreneQt o give 70% of pure conipound. ni.1). 203206'. Anal. Calcd. for ClaHsFzO: C, 72.22: H , 2.80. Found: C , 72.51; H , 2.89. (b) 11. .J. Namkung, T. L. Fletcher, and W.€1. Wetzel, J . M e d . C h e m . , 8 , 551 (1965). (9) Made by M.J. Namkung from 7-fluoro-2-fluorenaniine by a Hchiernann decomposition of the diazonium fluoborate, wliicli was prepared in aqueous tetrahydrofuran [see T. I,. Fletcher and AI. J. Namkung, Chem. Ind. (London), 179 (196l)l. in boiling xylene (72% over-all). Sublimation a t 75' (1 mm.) followed b y recrystallization from methanol-u-ater gave the product, SI. Berkovic [Israel J . Chem., 1, 1 (1963)l reports 1n.p. 8 3 O . m.p. 82-82.5'. (IO) A . Pictet and L. Ramseyer, Ber., 44, 2486 (1911).
fluorene and 2-acetamido-4-chloro-7-fluorofluorene followed by chlorination gave the same compounds as we described earlier made by a different route. Chlorination of 2-formaniidofluorene and ethyl S-2fluorenylcarbamate gives tetrachloro derivatives with the chloriries most likely in the 1-, 3-, 4-, and 7-positions as in the chlorination of 2-acetaniidofl~orerie.~"Infrared spectra of these compounds show similar absorptions in the 800-900-em-' region, indicating that two adjacent hydrogens are present in these niolecules studied.'" Antitumor activities of some of the polyhalogenated fluoreries are shown in Table 11. The most active compounds are substituted with chlorine or bromine a t the 1-and 3-, or 1-, 3-, and 4-positions and with halo, including fluoro, or nitro a t the 7-position of 2-aminoor 2-acetaniidofluorene. Whether blocking at both positions ortho to the 2-amino group is necessary for activity or for lowering toxicity is not yet known, but we are preparing compounds with only one of the ortho positions blocked so that this question can be explored. The first of these compounds, soon to be tested, was made from S-2-(7-bron1ofluorenyl)acetaniide by brominating in dimethylformaniide with I\-bromosuccininiide.: =Zn alternate route is also shown in Scheine IT'. Acetylation of two relatively weak amines, by merely shaking a t room teniperature in glacial acetic acid, is described. SCHEME
11
One further question worthy of examination, of greater significance than for this series of compounds alone, is the relation between carcinogenicity (a wellestablished property of 2-acetamidofluorene arid some of its derivatives) arid tumor inhibition.
Experimentale
Confidence 1
'.,
!l!l.
7
!),j1 1
{I!) , 7
!,$I
7
!)!I.7
S!l I
SlSU
5180
99. 7
x 180
9!). 7
Cai55
AXTITUMOR FLUORESE DERIVATIVES
July 196;
495 Tumor 9
Tumor systemb
I'luorene
Daily dose, mg./kg.
12 6 3 1 5
Tumor mg
xt..
.
t.
decrease,
YC
Survivors
7/10 7/10 9/10 10/10
324/1436 619/1436 907/ 1436 1221/1436
Specificity testC Confidence, Index
78 57 37 15 2.0
8180
125
6/6
112/520
isR
Ca7.55
1920 1920 960 30 1.5 120 60 30
10/10 9/10 10/10 10/10 10/10 lO/lO 10/10 10/10
414/957 195/957 180/9.57 238/957 315/957
57 SO 82 76 68 53 62 59
960 480 240 120
10/10 10/10 10/10 10/10
69 74 56
+Xi)
m/io
"0 120 GO 960 480 240 120
10/10 10/10 10/10 9/10 9/10 lop0 9 /lo
,502/1596 420/159G 709/1596 000 / 1,506 256/1596 265/1396 '1" ,iJ/l596 :194/1596 413/1596 392/1596 377/1596 423/1596
4t59/95i
368/957 397/957
2.0
44
84 84 83 76 c w
10
i6 c-
i t
74 1.6
2,7-Diamino-l,3-dibromo-
S180
500
6/6
135/520
-13 -e
2-Trifluoroacet amido-3-bromo-7-nitro-9-oxo-d
s91
200
8/10
121/510
r I
9-0xo-2,4,7-trichloro-d
S180
300 333 222 333 222 149 98 147 98 66 43
4/6 6/6 6/6 5/6 6/6 6/6 6/6 6/6 6/6 5/6 6/6
175/770 197/7iO 238/770 458/1395 620/1395 828 11393 625/1395 333,906 454/906 591/906 692/906
-
1 1,
ih r-
i .-I
70 68 56 41
56 64 50 33 24
2.2 123 5 /6 20/520 97' a The screening data in this table were kindly supplied by the Cancer Cheniotherapy Xational Service Center, National hstitutes of Health, Bethesda, l I d . ilssays were performed according to specifications established by CCYSC as reported in Cancer Chemotherapy Sarcoma 180 was t,ested in random bred Albino mice; Adenocarcinoma 755 and Cloudman llelanoma S9l were Rept., 25, 1 (1962). This is an extension of data reported earlier.ln E Passed st,age 3 of tested i l l HDF, mice. c See footiiote d i n Table I11 of ref. la. Passed stage 2 of sequential testing. Confirmation test. sequential testing. 1 Activity confirmed. Q
nil.) and a small amount of Rsney nickelll to give a quantitative Found: C,44.42: H, 1.85; K,4.25. vield of amine. Recrystallization from ethanol-water gave an N-Acetyl derivative had m.p. 332-333" (toluene). malytical sample, m.p. 124.5-125.5O. Anal. Calcd. for ClIHSBr2S02:C, 45.60; H, 2.30; Br, =InaZ. Calcd. for C,?HSF2X:C, 71.88; H, 4.18; N, 6.43. 40.46; S , 3.55. Found: C, 45.41; H, 2.62: Br, 40.41; N, 3.70. Found: C, 71.74; H, 4.26: N, 6.64. 2,4-Dichloro-7-nitro-9-oxofluorene. A.-This ketone was 2.7-Dibromo-4-fluorenamine.-2.7-Dibroino-4-ni trofluorene synthesized from methyl 2-hromo-5-ni trohenzoatel2 and 2,4( 7 . 4 9.) was redured in the sanie wa?; giving 6.7 g. (99yc),n1.p. dichloroiodobenzeiie b y an Ullrnann procedure as described 189.5-190.5° (ethanol). below, giving a 23Tc yield; m.p. 230-230.5' (benzene): vmax Anal. Calcd. for C,,H9Br&: C. 46.05: H, 2.68: N. 4.13. 1724 (ketone C=O), 844 (two adjacent hydrogens) cm.-'. Found: C, 45.84; H, 2.64; Ki4.34. ' Anal. Calcd. for CI3H,CI,N0g: C, 53.09: H, 1.71: C1, 24.11: N-Acetyl derivative had m.p. 267.5-26s" (ethanol). N, 4.76. Found: C, 53.20, 53.19; H, 1.81, 1.66; CI, 24.04: 4nal. Calcd. for C11H11Br2NO:C, 47.28; H, 2.91; S , 3.68. N, 4.92. Found: C, 46.87; H, 2.94; N, 3.89. B.-2,4-Dichloro-9-oxofluorene ( 1 g.) was added in several 2,7-Dibromo-9-oxo-4-fluorenamine.-2,7-Dibromo-4-nitro-9- portions to a mixture of 907' HNO, (5 ml.) and acetic acid ( 5 ml.) oxofluorene was reduced with SnClt .2H20 and concentrated at 3.5'. Concentrated H2S04 ( 1 r i d . ) was then added slowly HCl giving a Sur{, yield, m.p. 253.5-254.5" (toluene). with the temperature held a t 40-45". After 5 niin. o f stirring Ilnal. Calcd. for C13HiBr2NO: C, 44.23: H, 2 . 0 0 ; N, 3.97. ,
I
(11) T. I,. Fletcher and 31. J. S a m k u n g , J . Org. C'hem., '23, 680 (1938)
(12) A . E. Holleman and 13. R. del3ruyn. Rec. / m i ' . chim., 2 0 , 206 (1901); R. E. Buckles, R. Filler, and L. Hilfman, J . Org. Chrm., 17, 233 (1952).
July 1965
ANTITUMOR FLUOREXE DENIVATIVES
173")] and anhydrous FeC13 ( I g.) were dissolved in acetic acid (300 ml.), with stirring, and chlorine was bubbled in a t a rate of 200 ml./min. for 2 hr. a t 60-70". After 3 hr. of stirring a t this temperature, the mixture was cooled to room temperature, and the product was filtered off and recrystallized from acetic acid and then from toluene giving 9.3 g. (537,), m.p. 271.5-273.5'. Another recrystallization from toluene gave an analytical sample: m.p. 273.5-274.5'; vmax 3205 (amide S-H), 1667 (amide I), 818 (two adjacent hydrogens) cm.-l. -4naZ. Calcd. for C14H,C1,KO: C, 48.45; H, 2.03; C1, 40.87; N , 4 . 0 4 . Found: C, 48.69; H,2.21; C1, 40.78; N, 4.15. Ethyl N-2-Fluorenylcarbamate.-2-Fluorenamine (36.2 g., 0.2 mole) was dissolved in dimethyl sulfoxide (250 ml.), and Xa2COs (21.2 g., 0.2 mole) in water (100 ml.) was added. The mixture was cooled to < S o and ethyl chloroformate (24 g., 0.22 mole) was added dropwise with rapid stirring over a period of 1 hr. The reaction mixt,ure was stirred 1 hr. more at 0-5" and a third hour at room temperature, and diluted with water. The product was recrystallized from methanol-water giving 46.7 g. (927,), m.p. 119-120.5". Recrystallizat,ion from methanol gave an analytical sample, n1.p. 120-121 '. Anal. Calcd. for C16H15N02:?;, 5.53. Found: N, 5.46. Ethyl N-2-(1,3,4,7-Tetrachlorofluorenyl)carbamate.--Eth~l N-2-fluorenylcarhamate (12.i g., 0.05 mole) was chlorinated by a procedure similar to the one described for the fluorenylformamide. The product was filtered off, after dilution with n-ater, and recrgst,allized from ethanol giving 12 g. (61%)) m.p. 203.5205'. After recrystallizations from ethanol, acetic acid, benzene, and finally methanol, the product melted a t 193", resolidified, and remelted a t 206-208.5'; vmSx3226 (amide N-H), 1700 (ester C=O), 820 (two adjacent hydrogens) cm.-l. Anal. Calcd. for Ci~HIIC1~N02: C, 49.14; H , 2.84; C1, 36.26: N , 3.58. Found: C, 49.15; H, 2.90: C1, 36.03; N, 3.67. 3,7-Dibromo-9-0~0-2-fluorenamine.-To7-bromo-9-oxo-2fluorenamine20 (2.7 g., 0.01 mole) in dimethyl sulfoxide (50 ml.) 48yc HBr (1.8 ml. excess) was added.21 The mixture was heated under reflux at 95-100' for 1 hr. and cooled. iifter water dilution the mixture was made alkaline with XH,OH, and the product was isolated and recrystallized from toluene giving 2.6 g. (74%,); m.p. 254-255'; vmax liO9 (ketone C=O), 818 (two adjacent hydrogens) cm.-1. Anal. Calcd. for Cl,H7Br2NO: C, 44.23; H, 2.00; N, 3.97. Found: C, 44.46; H,2.13; K,4.14. 3,7-Di bromo-2-fluorenamine.--3,i-Dibromo-9-oxo-2-fluorenamine (0.9g.) was refluxed for 6.5 hr. in a mixture of 2,2'oxydiethanol (60 ml.) and 85% hydrazine hydrate (10 ml.)l6 and diluted with water. The product, 0.7 g. (88%), m.p. 144.5146.5', was recrystallized from ethanol-water giving an analytical sample, m.p. 146.5-147.5'. Anal. Calcd. for CI3HgBrgX: C, 46.05; H, 2.68. Found: C, 46.22; H, 2.97. N-2-(3,7-Dibromofluorenyl)acetamide. A.-3,7-Dibromo-2fluorenamine (0.2 g.) and acetic acid (15 ml.) were shaken a t room temperature22unt,il crystallization of the product took place. The excess acetic acid was removed by evaporation and the (20) C . Courtot, A n n . Chim. (Paris), 14, 5 (1930). (21) T. L. Fletcher, RI. J. Namkung, and H.-L. Pan, Cheni. I n d . (Loxido~i), 660 (1957).
497
product, 0.2 g., was recrystallized from ethanol: m.p. 276.52 r1 4 . 5 " : umRI 3245 (amide N-H), 1656 (amide I), 824 (two adjarent hydrogens) cm.-'. dnal. Calcd. for Cl5Hi1Br2KO:Br, 41.94; K j3.68. Found: Br, 41.84; N , 3.81. B.-N-2-( 7-Bromofluoreny1)Itcetamide ( 3 g., 0.01 mole) and Sbromosuccinimide ( 2 g., 0.011 mole) were dissolved in dimethyl formaniide5 (20 ml.) by warming. The solution was set aside in the dark for 2 hr. and then the precipitated producst was filtered off and rerrystallized from acetic arid giving 2.7 g. (71%). There was no melting point depression when the product was mixed with the compound obtained in A. 2,4-Dichloro-9-oxofluorene.-4-C hloro-Y-oxo-2-fluorenaniine ( 2 g.) was diazotized in concentrated HC1 (100 ml.) a t 0-5' with NaK02 ( 1 g.). The diazonium chloride was treated with a solution of CuCl (excess) in concentrated HC1 (20 nil.). The product was chromatographed in benzene (alumina) and recrystallized from benzene-ligroin ( d 0.68-0.70) giving 1.6 g. ( 7 3 : ; ) : 111.p. 152-1.53': vmRX l i 1 8 (ketone C=O), 731 (four adjacent hydrogens) cm. -1. -Anal. Calcd. for C18HsC1?0: C, 62.68; H,2.43; C1, 28.47. Found: C, 62.53: H, 2.46; C1, 28.33. 2,7-Diamino-l,3-dibromofluorene.-l,3-l~ibronio-~-ni t ro-2fiuoren!"el;' was reduced in a mixture of t'oluene and ethanol by Raney nicakel and hydrazine hydratell giving a R O T yield, m.p. 214-215' (ethanol), umax 810 (two adjacent hydrogens) cn- .-1. .-lnal. Calcd. for C,3HloBr,N,: K , 7.90. Found: N, 7.88. N-742-Amino-l,3-dibromofluorenyl )acetamide.-Acetylation of the above amine gave a 100% yield; m.p. 258.5-259" (ethanol); vmllX 1661 (amide I ) ,823 (two adjacent hydrogens) cm.-l. .-lnal. Calcd. for CljH12Br2N20: Br, 40.35; N, 7.07. Found: Br,39.93; N,7.18,7.12. 1,3-Dibromo-7-phenylazo-2-fluorenamine.-2,7-~iamino-l,3dibromofluorene was condensed a t room temperature with nitrosolienzene in a mixture of acetic acid and dimethyl sulfoxide. The product was chromatographed (benzene) on alumina and recryst,allized from benzene-petroleum ether (b.p. 30-60") giving a SOYc yield, m.p. 214-215". .Anal. Calcd. for CI9Hl3Br2N3:C, 51.50; H, 2.96; S , 9.48. Found: C, 51.93; H, 3.03: N, 9.60. 7-Chloro-l,3-dibromo-2-fluorenamine.-~-Chloro-2-fluorenaminez0(6.5 g., 0.03 mole) was dissolved in dimethyl sulfoxide21 (60 ml.). To the stirred solution 48% HBr (7.9 ml.) was added. The mixt,ure was heated at 9G100" for 1 hr., diluted with water, and made basic with XH40H. The product was isolated and recrystallized from toluene-ligroin giving 9.9 g. ( 8 8 % ) , m.p. 198-199 O . Anal. Calcd. for C13H8Br2ClN:Br, 42.79; N, 3.75. Found: Br, 42.51; S , 4.01. L
Acknowledgment.--We wish to thank Sorma Saimy and Carol-Ann Cole for assistance in the preparation of some of the starting materials and for running the infrared spectra. (22) Tius facile, room terriperature acetylation, 111 higli 3 ield, ]>itshccn obseri ed here previously for such a weak amine as 9-oxu-Z-Auurenaniine.
