G 05 T.\BI,EI1 P H I R ~ I I C O L O G I C A LSCREENING
Compd I I1 I11 IV V vI VI1 VI11 IX X XI XI1 XI11 XIV
XV SVI XVII XVIII
XIX XX XXI 3Iorphine'HCl Phenylbutazone Hydroc hlorothiazide Dehydrocholic acid Chlorprops mide
Approx LDao (mouse), mg /ka ip
1120-1 230 560-630 1090-1 2 10 >so0 540-610 580-620 510-590 290-320 550-620 45-70 1100-1260 5T0-640 580-820 590-650 280-310 1140-1220 410-450 :390-410 1170-1250 590-630 > I600
Analgesic act. (mouse) Increase of reaction time, mg/kg
'lo"
1P
14 21 37 59 .X 8 38 58 10 28 12 23 34 50 14 16 34 36 13 28 9 6i 61
200 100 100 200 200 100 200 200 200 25 100 200 200 200 50 200 100 30 100 100 100 5 100
Hot plate test, 1 hr after treatment. hr after treatment. e Equimolar doses. Q
Antipyretic act. (rat) Antiinflammatory Max activity (rat) temp Inhibition deof edema, mg/kg crease, ma/ke: Ip OCC 5% ip
22 47 3T Inactive 26 50 Inactive 37 30 Inactive 45 Inactive 38 Inactive 18 Inactive
200 100 100 200 200 100 200 200 200 25 100 200 200 200 50 200 100 50 100 100 100
2 2 1 4 2 2 2 2 2 4 1 4 3 2 3 1 3 4 0 8 2 9 2 4 2 7 3 3 0 9 2 1 1 3 0 i 1 6 2 7 1 7
200 100 100 200 200 100 200 200 200 25 100 200 200 200 50 200 30 100 100 100 100
18
100
1 6
100
30 13 13 25
17
RESVLTR
Iliuretic activity (rat) Test volt,/ control mdkp vol PO Inactive 1.15 Inactive 1 29 1 19 1.1.5 1.12 Inactive Inactive 1.30 Inactive 1.27 1.13 1.14 Inactive Inactive 1.18 Inactive Inactive Inactire Inactive
1.56
50 50 50 50 50
.5 0 50 50 50 50 50 50 50 50 50 50 50 50 50 50 30
Choleretic activity (rat) Increase of bile flow, 7cd my'kge 1 hr 2 hr id 32
18
Hypoglycemic activity (rat) Blood sugar decrease, m g / k g
'Zf Inactive
i3
14
Inactive
Inactive
80
Inactive
36
83
9i 134 92
39 131 78
28
43
120
92
42
100
83 87
Inactive 13 :3 5
100 100 50
38 Inactive
50 100
Inactive Inactive Inactive 11
50 100
12 Inactive
100
(4) P. Iheresidue, giving a solid product which, on crystallization from ethanolligroin ( b p 75-120°), gave colorless crystals, mp 2277228". Pharmacology.-The acute toxicity, and antiinflammatory, analgesic, and diuretic activit,ies were investigated according to the techniques previously described.'& The antipyretic action was studied in rats made pyretic by brewer's yeast, according to Smith and Hamb0urger.l The activity on the choleresis was investigated in rats, using the biliary fistula technique of 3larazziUberti and Turba.j The hypoglycemic action was measured in rats, according to the procedure described by CeriotL6 The antibacterial and antifungal activities were measured against Micrococcus pyogenes z,ar. aureus ATCC 6538 P, Bacillus subtilis ATCC 6633, Escherichia coli McLeod ATCC 10,536, Salmonella typhi T 30 Roma 31 507, and Candida albicans ATCC 10,231, using the serial dilution technique.' All compounds were administered as the hydrochlorides in aqueous solution. Morphine, phenylbutazone, hydrochlorothiazide, dehydrocholic acid, and chlorpropamide were used as standards for comparing the analgesic, antiinflammatory-antipyretic, diuretic, choleretic, and hypoglycemic activities, respectively .
PO
100
d
50
S-aliie+ a t 1 and 2
Effect of Organic Compounds on Reproductive Processes. 11. Alkylating Agents Derived from Various a,w-Alkplenediols \T. A. SKIWER, J. H4Y-FORD, T . E. SHELLESBERGER, A\D
IT. T . COLWELI.
Life Scaences Research, Stanford Research Znstzticte, .lienlo P a r k , Calzfornia Receaved January 10, 1966
A program of synthesis of alkylating agents having related carrier moieties but with a variety of alkylating functions has been under way in our laboratories. These compounds are being evaluated for their effect on reproduction in the housefly (Musca doiizesfia L.), mice, and ,Japanese quail. Previous studies have shown that ~~,~'-bis(aziridiriylacetyl)-l,8-octaniethyle1iediamine' inhibits reproduction in the housefly at 1 arid 0.1% concentration when added to their feed. I t was of interest to see whether similar conipounds derived from a,@-alkanediols would also affect their reproduction. The compounds synthesized (2-21) are shown in Table I. The bisbronioacetyl esters (2, 6, 10, and 14) were best prepared by the addition of diol to chilled bronioacetyl bromide. The iodo derivatives (3, 7, 11, and 15) mere prepared from the bromo compounds using S a 1 in acetone. The aziridinyl derivatives (4, 8, arid 12) were derived froin the bronio compounds by the addi(1) IT-. A. skinner, H. C. Tone, T E. Shellenberger, and G. \V Nevell, J . ,\fed. Chvm , 8, 647 (1965).
SCI-T,C02(CII?),,O?CCI1,s 2,n = G : S 3,n =
=
131.
=
J
6:s
=
?rT
4,n
=
(1
6, ti 7, n
=
8 :S =
€31.
10, n
= 10:
=
8 ;S =
I
11,
71
=
10;
8,71
=
s: s
x
12, 11
=
10:
/'/
=
\~:
1 ' \
~
9, n =
I',
5 , r1 = (i;S =ClTy?;
s:
s
=
(:lTzx
'
)//
=
I3r
15,
S
=
I
16,
S
=
S
=
Rr I
13, 71
=
10;
I'\ ',
s
=
/,'
CIIYX
1 ~
CIL( C I I?),o,ccr-r=cri 17
CfI,(CET,) OL!CIIiS
S
=
s=s
\\'
14,
S S
I'/
C i r , ( c i r ? ) , o , C c I ~ ? c € -, T ~ ~ 'I
\
18
\i
(:r12-cIIc0,(cH2),,o~cCrI=cr-T. 19, n 20, n
= 0 = 8 21, n = 1 0
tiori of aziridirie in tetrahydrofuran to the bromo derivative in the same solvent using potassium carbonate as an acid acceptor. The iodo derivatives gave results in this displacemerit reaction similar to those obtained with the bromo compounds. The reported yields (Table I) reflect the difficulty encountered in purifying the aziridiriyl compounds. The crude yields were generally good (ca. SO%), but u good chromatographir proredure was riot discovered and considerablr 10s.; was encountered owing to the formation of polymer
July 1966 niole) of bromoacetyl bromide, under nitrogen, was added 5.0 g (0.034 mole) of 1,s-octanediol over 25 min. The mixture was stirred a t room temperature 2.5 hr. Ether was then added and the solution was treated with excess solid K?CO,. The mixture was then filtered, the filtrate was washed with 5coK & 0 3 and dried, and the ether was removed in vacuo to yield 10.14 g (77%) of solid, nip 33-35", Rf (CHC13) 0.49 (silica gel G). Method B. 0,O'-Bis( iodoacety1)-1,s-octamethylenediol(7).A solution of 5.0 g (0.013 mole) of 6 in 80 ml of acetone was added to a solution of 15.8 g (0.105 mole) of KaI in 160 nil of acetone. After addition was complete, the reaction mixture was refluxed o n a steam bath for 2 hr. The mixture was then cooled, ice was added, and the crystalline product was collected by filtration. An additional portion of product was obtained cia ether extraction of the filtrate to yield a combined total of 5.52 g ( 8 7 5 ) of 7, nip 25.5-26.5'; Rf (CHC13) 0.55 (silica gel G). Method C. O,O'-Bis(aziridinylacetyl)-1,6-hexamethylenediol (4).-In a thoroughly dried flask under a ?j, atmosphere was placed 2.32 g (0.0168 mile) of K&03 and 1.0 g (0.0028 mole) of 2 in 40 ml of tetrahydrofuran ( T H F ) . This mixture was cooled to 0" and treated with a solution of 0.72 g (0.017 mole) of aziridirie over a 5-min period. The reaction mixture was stirred at 0' for 3 hr then allowed t,o warm to room temperature and stirred for 15 hr. Et,her was then added, the mixture was filtered, and the solvent was removed from the filtrate in uucuo, keeping the temperature below 40", to yield a liquid which later solidified. Crystallization from petroleum ether (bp 30-60")-ether gave 0.081 g (11%) of product, mp 58-62, which partly decomposed to a polymeric mass over a period of several days. An analytical sample was prepared by formation of the chloroethylamine hydrochloride. The aziridine 4 (90 mg) was dissolved in 10 ml of ethanol. The solution was then cooled to 0" and satiirated with HC1. After removal of the solvent, the white residiie was crystallized from ethanol-ether, mp 170-174'. Method D. O,O'-Bis(acrylyl)-l,6-hexanediol(19).-In a dried flask under a 8, atmosphere was placed 5.9 g (0.05 mole) of 1,6hexanediol dissolved in 200 ml of T H F and 27.6 g ( 0 . 2 niole) of K2C03. The mixture was cooled to 0" and 13.6 g (0.15 mole) of acrylyl chloride was added. The reaction was allowed to warm to room temperature and stirred for 62 hr. The mixture was then filtered and the solvent was removed from the filtrate. The residue was washed well with ether and filtered again, and the ether was removed from the combined organic solution to yield 6.9 g of crude product. This material was chromatographed on silica gel using 4:1 petroleiim ether-ether elution. Three grams (277,) of clear, colorless product was obtained; infrared : : :A ( H ) 5.80 (C=O, ester), 6.1 and 6.2 doublet (C=C, acrylyl). Method E. 0,O'-Bis( aziridinylpropionyl)-1,6-hexanediol(5). -Compound 19 (0.75 g, 0.0032 mole) dissolved in 10 nil of T H F was placed in a dry flask under X?. Aziridine (0.916 g, 0.021 mole) was slowly added and the reaction mixture was stirred for 62 hr. Aliquots were periodically taken and the infrared spectrum was checked for t,he presence of remaining double-bond absorption. The solvent was then removed in vacuo, keeping the temperature below 40", to yield a solid, 0.88 g (goyc),mp 30-34", R, (CHC1,) 0.67 (alumina). An analytical sample was prepared by twice recrystallizing from pentane: mp 34-36". The nmr and infrared spectra were in agreement with the assigned structure. Method F. 04Aziridinvlacetvl)hexanol il6~.-Conmound 14 (9.7 g, 0.044 mole) was added over 10 min to a stirred sblution of 3.18 g (0.074 mole) of aziridine in 16 ml of triethylamine cooled to 0". The reaction was held at 0" for 50 hr. The copious precipitate of triethylamine salts was then filtered off and washed with ether. After removal of the ether In m " o the remnant was distilled, bp 71-80" (0.15 mm), t o yield 4.8 g (60%) of colorless product. This procedure is similar to that of Bestian.2 Tsou, Hoegerle, and Sn5 have discussed the infrared spectral characteristics of some aziridyl derivatives related to those prepared in this paper. m e are in agreement with their findings that a weak peak or shoulder from aboiit 3.30-3.35 p is characteristic of these addiicts. The nmr spectra of the various series were also definitive for structure identification. The triplet from the alcohol methylene nest t o oxygen a t 4.0-4.2 ppm was chosen as the basis for integral determinations. The acetyl methylene was characteristically (5) IC. C Tsou, K. Hoeperle, and H. C. F. Su, J . >Wed. Chem., 6 , 435 (1963).
NOTES
607
a singlet a t 2.9-3.1 ppm. The two propionyl methylenes from the acrylyl adducts overlapped to form a poorly defined triplet a t 2.4-2.5 ppm. All of the aziridinyl adducts possessed sharp multiplets (ea. 1.7 and 1.1 ppm) up- and downfield from the remainder of the methylene absorption. Each multiplet contained about half of the integral area reqnired for the four aziridinyl methylene protons.
Acknowledgment.-This work was supported by U. S. Public Health Service Grant GlI-11491. We wish to thank T'. Tovar for assistance with the biological studies.
Nitrogen M u s t a r d Type Derivatives of Thiophene and Benzo [ b l t h i o p h e n e DAVID A. SHIRLEY ASD GEORGE R. BELL,JR. Department of Chemistry, Cniversity of Tennessee, Knoxville, Tennessee Received Sovember 17 , 1965
This paper reports the synthesis for anticancer evaluation of a series of "nitrogen mustard" derivatives (I) of thiophene and benzo [blthiophene ; specific cases RCH?CHzN(CH?CHzCl)?
I
include those in which R = 2-thienyl, 3-thienyl, 2benzo [blthienyl, and 3-benzo [blthienyl. The only prior report of nitrogen mustards related to the above types describes' the synthesis of S,n'-bis(2chloroethyl)-2-thienylaniirie. Similar derivatives of furan and tetrahydrofuran were reported by Landing, et u Z . , ~ arid of thiazole by Jlikhailov, et aL3 The synthetic route employed for our synthesis is indicated in Scheme I. Yield ranges for the four SCHEME I RbIgX or RLi
TsOCHSCHzCl
+
RCH?CH?Cl
(48-807c)
Ts
=
I1
p-toluenesulfonyl
examples of R are given in parentheses along the arrows. Samples of I V for biological testing were converted t o the hydrochloride salts in 6&85% yields. I n the formation of I1 in the cases in which R = 2thienyl and 2-benzo[b]thienyl, it was planned t o use the organolithium reagents (RLi) readily formed in high yield by metalation of the parent heterocycle with n-butyllithiun~.~This was a satisfactory route for formation of 2-(2-chloroethyl) benzo [blthiophene; however, 2-thienyllithium and 2-chloroethyl p-toluenesulfonate gave as the only major product 2-thienyl p-tolyl sulfone. S o 2-(2-~hloroethyl)thiophenecould be isolated from this system even though several variations of (1) E. Wilson and AI. Tishler, J . Am. C h e m . Sac., 73, 3635 (1951). ( 2 ) R. H. Landing, A . Goldin, H. A. S o e , B. Goldberg, and D. AI. Sliapiro, Cancer, 2, 1055 (1949). (3) n. h1. Mikhailov, V. P. Bronovitakaye, and J. K. P l a t o r a , Zh. Obshch. Khim., 26, 3445 (1956); Chem. Abstr., 51, 9588 (1957). (4) (a) H. Gilman and D. A. Shirley, J . Am. C h e m . Soc., 71, 1870 (1949); (b) U. A. Shirley and M. D. Cameron, ibid., 73, 2788 (1950).
