92 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA J o m l ofNarwal Prodvrts Vol. 54, No. I , pp. 92-97, Jan-Fcb 1991 zyxwvutsr zyxwvuts zyxwvu zyxwv zyxwv NEW AVARONE AND AVAROL DERIVATIVES FROM THE MARINE SPONGE DYSZDEA‘CZNEREA zyxwvutsrqponmlkjihgf S. HIRSCH,A. RIJDI, Y . KASHMAN,+ Sackkr Faculty of Ewrrt Sciences, Scbool of Chemistry and Y.LOYA D@arhnentof Zoology, Tel Aviv Uniurrrity, Ramat Aviv 69978, Tel Aviv, Israel Ass~~~cr.--Six new avarol and avarone derivatives, 3‘-hydroxyavarone [3}, 3’,6‘dihydroxyavarone [4],6’-hydroxyavarol[5], 6’-acetoxyavarol[61,6’-acetoxyavaronem,and 6’hydroxy4‘-rnethoxyavarone 181, are reported from the Red Sea sponge Dysidu cinma. The structures of the new compounds were determined by spectroscopic data, mainly 1D and 2D n m r measurements. The absolute configurations of 5,6, and 7, and most likely also of 5 4 , and 8, were established on the basis of cd measurements to be the same as that of avarol. Several of the new compounds are cytotoxic, possess antimicrobial activities, and have anti-HIV-1 reverse transcriptase activities; the most active is compound 8. In search of biologically active marine natural products we have found that the CH,CI, extract of Dyidus cinerea (Keller) (family Dysideidae, order Dictyoceratida) collected in the Gulf of Eilat, the Red Sea, possesses anti-HIV-1 reverse transcriptase activity. A strong inhibition of the HIV-1 reverse transcriptase enzymatic functions was tracked down to a fraction containing quinones. This report describes the structure of the new compounds, while the biological activity will be published elsewhere (16). A variety of compounds having a quinone or a hydroquinone moiety attached to a sesquiterpene unit have been isolated from marine algae (1-3) and sponges (4-9, 13). Among the sponge metabolites are avarol El] (5,6) and ilimaquinone 121 (8), which possess the rearranged drimane skeleton. zy 15 1 2 11 PR 3 R,=OH, R,=R,=H 5 R=H,R,=OH 4 R,=R3=OH, R2=H 7 R,=R,=H, R,=OAc 6 R=H,R,=OAc 9 R=R,=OAc 8 R,=H, R2=OMe, R,=OH 10 zyxwvut zyxwv zyxwvu zyxwv zyxwvu zyxw zyxw zyxw Jan-Feb 19911 Hirsch et al. : Avarone and Avarol Derivatives 93 Specimens of the D.cinerea sponge were frozen shortly after collection at 15-20m near Ras Zaatir, in the Gulf of Eilat. The CH,CI, extract of the freezedried specimens was chromatographed in CHC13-MeOH-hexane (1: 1:2) over Sephadex LH-20, then over a Si gel column eluted with hexane and increasing percentage of EtOAc, and again over Sephadex LH-20 prepared and eluted with CHCI3-hexane (7:3) to afford six new compounds 3-8.Starting from 30 g of one of the specimecs of the dry sponge we obtained compounds 3 (100 mg), 4 (150 mg), 5 (100 mg), 6 (100 mg), 7 (20 mg), and 8 (55 mg) (the relative amounts changed remarkably from one collection to another). Compound 3 showed hydroxyl (3400 cm- ') and carbonyl (1640 cm- *)absorptions in the ir spectrum and uv absorptions (MeOH) at 256 (E 10200)and 284 (E 3300) compatible with a hydroxyquinone structure (10). In its 'H-nmr spectrum compound 3 showed four methyl signals, three singlets (6 0.85, 1.00, and 1.53ppm) and one doublet (6 0.93), an AB quartet (6 2.51 and 2.65 ppm,J= 13.5 Hz), a vinyl proton (6 5.15), two aromatic protons (6 6.76 and 6.75), and one exchangeable proton (6 7.05). The I3C-nmr spectrum (Table 1)showed 2 1 carbons including two carbonyls at 182.8 and 187.7 ppm, characteristic for a quinone (lo), and six additional sp2 carbons. The molecular formula C, *HZBO3 (cims and elemental analysis) and the above-mentioned spectral features clearly indicated that compound 3 contains a hydroxyquinone moiety TABLE1. Cornoarison of '3C-nmr Shifts for Cornmunds 1-8.' Compound Carbon 3b c-1 . . . . . . . c-2 . . . . . . . c-3 . . . . . . . c-4 . . . . . . . 19.9 26.9 120.7 144.0 43.4 35.9 27.9 38.1 38.5 48.2 32.6 17.2 17.7 20.1 18.1 120.7 187.7 152.7 139.9 182.8 131.3 I 4' 19.9 5 6d 7 tr 20.4 27.3 120.5 144.0 43.2 35.8 28.8 38.8 38.7 49.2 35.8 16.4 18.2 20.1 18.2 113.2 153.3 120.6 105.8 131.3 155.1 20.3 27.3 120.6 144.4 42.7 36.0 28.6 39.1 38.6 49.8 36.0 16.2 18.1 20.0 18.1 116.2 153.6 119.6 106.9 132.1 147.0 20.5 26.8 120.7 144.0 44.1 36.2 27.9 38.2 38.8 49.2 35.9 16.9 18.0 20.1 18.1 120.7 185.5 137.5 134.5 181.0 150.8 19.9 27.9 120.7 144.0 43.7 36.0 28.0 38.2 38.5 48.4 32.8 17.1 17.7 20.1 18.0 120.8 187.3 109.0 155.8 178.5 151.3 I 1' 2 19.5 26.4 120.5 144.5 41.8 37.0 27.6 36.0 38.2 45.8 35.7 17.1 17.5 19.8 17.8 129.1 153.3 111.1 111.0 153.1 119.4 33.0 28.6 36.7 153.4 43.3 28.6 28.0 38.2 40.5 50.2 32.4 17.3 17.9 20.6 102.6 117.4 182.4 161.8 102.1 182.4 160.5 zyxwvu c-5 . . . . . . . ...... . . ... .. . C-6 c-7 c-8 . . . . . . . ... .... c-10 . . . . . . . c-11 . . . . . . . c-9 c-12 C-13 C- 14 C-15 c-1' c-2' c-3' c-4' c-5' C-6' ... .... . . . . . . . ...... . . . . . . . . ... .... . . . . . . . ... .. .. . . . . . . . . . . . . . . . . . . . . . 'Assignments 27.0 120.6 144.1 43.5 36.0 28.1 38.4 38.5 48.6 33.2 17.0 17.6 20.1 18.0 121.8 185.4 152.7 130.6 181.8 144.5 the carbon resonance lines were determined by CH-correlation experiments ( J = 140, 10, and 7 Hz), in CDCI,. b n g range CH-correlations observed ((3"s): 3/2a, 15; 4/10,15; 5/14; 6/10; 8/68,11,13; 91 10,11,13; 10/6f3,11,12; 11/12; 1'111; 2'/4', 6'111; 3'/11; 5'/4',6'. 'Long range CH-correlations observed (UH's): 1'111; 2'/4',11; 3'/4'; 5'/4'; 6'/4',11. dLong range CH-correlations observed (UH's): 1'/3', 11; 2'/4', 11; 5'13'; 6'/4', 11. T h e reported carbon chemical shifts of the quinone moiety of isospongiaquinone are (Uti,):1'/ 117.8, 2'/182.4, 3'/161.8, 4'1102.0, 5'1182.0 and 6'/153.4. The following CH-correlations were observed (UH's): l ' / l l ; 2'/11; 3'/4'-OMe; 5'/3'; 6'/11. 'The '3C-nmr chemical shifts are of avarol dimethyl ether. 94 zyxw zy zyxwvuts zy Journal of Natural Products Wol. 54, No. 1 linked to a sesquiterpene portion, similar to that of avarol [l}(1). Comparison of the 13C-nmrdata (Table 1) suggested for 3 the same sesquiterpene unit as in 1.The proposed structure was also supported by the mass spectrum of 3 which showed, besides the [M 3H)+ peak characteristic for quinones (1l), dominant fragments at mlz 191 [C&23 -the methylated decalin moiety]+ and at m/z 140 [C,H503 3H -the quinone together with the benzylic methylene)+. Further unequivocal proof for the suggested 3 '-hydroxyavarone structure was obtained from long range CH correlations (shown in Table 1). The latter experiment also established the 3 ' position for the phenolic OH group by means of the C-3' to H2-11correlation (the 6' position for the hydroxyl being excluded as the two quinone protons appear as singlets in the nmr spectrum). The relative stereochemistry of the chiral centers (C-5, -8, -9, and - 10)was assigned as indicated on the basis of the almost identical '3C-nmr spectrum of the sesquiterpene moiety of 3 and the same portion in avarol 117 (5,6). The stereochemistry was confirmed by an nOe experiment. Irradiation of Me- 12 gave enhancements of H- la (2%), Me-13 (2%), Me-14 (4%), and H- 11(4%). Irradiation of Me-13 gave an enhancement of H-78 (2%) and H - l l ( 3 % ) , while irradiation of Me-14 gave enhancements of H-lp (4%), H-68 (7%), H-7P (3%), and Me-12 (5%). The 5S,8S,9R, 10s absolute configuration ofavarol(6,12) is also suggested for compound 3 on the basis of the absolute configuration of compound 6 , as determined by cd measurements. Comparison of the 'H- and especially the 13C-nmrspectra (Table 1)of compounds 3-8 clearly indicates that all six compounds have the same decalin moiety as avarol, with the differences between the various compounds being in the quinone portion. Compound 4, C,,H,,O4, has an additional hydroxyl on the quinone moiety, mlz 153 [c+o4]+ (lo%), the base peak being the decalin fragment at m/z 191 [C14H23} (100%). A singlet at 6 6.73 and an additional exchangeabletwo-proton signal at 7.10 ppm could be seen in the 'H-nmr spectrum. The distinction between three possible isomeric structures (the quinones with the single proton at the ortho, meta, or para position to the linking C- 11methylene) in favor of the 3',6'-dihydroxy isomer was based on the long range CH correlations (shown in Table 1). In the case of the 3',4'- or 4',6'-dihydroxy isomers, theabove correlations required two to four bond correlations against the common two and three bond ones in the case of the proposed 3',6'-isomer. Additional support for the proposed 3',6' structure came from the absence of a 'J correlation between the single quinone proton and C- 11, required in case of the 3',4'-dihydroxy isomer, and from the formation of a different dimethoxy derivative from the one obtained from compound 8 , on methylation with CH2N2. Compound 6 , C23H3204 mlz 373, WH]+ (9%), is slightly more polar than the other isolated compounds. The 'H-nmr spectrum of 6 showed, in addition to the sesquiterpene portion, one acetate (6 2.25 s) and two ortho-coupled aromatic protons (6 6.80 d and 6.13 d , J = 8.7 Hz). From the 13C-nmr spectrum it was clear that compound 6 is a 1,2-disubstituted hydroquinone. Upon acetylation compound 6 afforded triacetate 9, and oxidation with A g 2 0 gave quinone 7 (9). The latter compound was identical with natural compound 7. Another closely related, more polar compound was 5 , C21H3003, mlz 330 [MI+. According to the nmr data (Table 1and Experimental) it was clear that 5 is deacetyl-6. Indeed, acetylation of 5 afforded the same triacetate 9 as 6. Similarly to compounds 3 and 4, compounds 5 , 6 , and 7 also gave strong mlz 191 peaks in the ci mass spectra. Compounds 6 and 7 also showed strong peaks at mlz 181 (2 1% and 26% respectively) for the acetoxy methylene aromatic unit qH,04. In case o f 6 the latter fragment is the + + zyxwvu zy zyxwvu zyxw zyxwv Jan-Feb 1991) zyxwv zyxw zyxwvu zyxwv zyxwvut zyxwv Hirsch et al. : Avarone and Avarol Derivatives 95 acetoxy hydroxyquinone moiety, and in case of 7 it is the quinone with an additional two protons. [The addition of 1-3 protons to quinones in the mass spectrometer is well known (1l).)The base peak of 5 was d z 140 [C,H,03)'. The long range CH correlations measured for compound 6 (shown in Table 1) are in full agreement with the suggested structure. Hydroboration of compound 9 followed by oxidative workup yielded the 3-keto derivative 10.Compound 10 showed a negative Cotton effect at 290 nm for the n H n * transition of the carbonyl, with the same sign and magnitude as the Cotton effect of the 3-keto derivative of avarol(6). Hence, the absolute configurations of 5 , 6 , and 7have to be the same as that of avarol. If a common biogenetic route is assumed, the absolute configurations of 3 , 4 , and 8 are also the Same as that of avarol. The last compound, 8, C22H30O4, also possesses the same tricyclic carboxylic skeleton as compounds 1-7(Table 1, Experimental). A singlet at 6 5.8 1, an exchangeable hydroxyl at 6 7.50 s, and a methoxyl at 6 3.80 s clearly pointed to a pentasubstituted quinone. As with compounds 3 , 4 , and 7,the uv spectrum of the yellowish solution of 8 suggested that the major tautomer of all these compounds is the one with the ortho-hydroxy-para-quinone structure [rather than the tautomeric ortho-quinone, which should give a red solution with absorptions up to A = 450nm (15)). This, of course, does not exclude a possible condensation of these compounds with ophenylenediamine. Indeed, compounds 4 and 8 as well as ilimaquinone 12) (obtained by us from a Smenospongia sp.) yielded, as judged by the development of a cherry-red color, an heterocyclic adduct by reaction with o-phenylenediamine. In the case of ilimaquinone, which was obtained in larger amounts, we have isolated the condensation product 11. An nOe enhancement of 10% between the methoxy group and the single quinone proton in 8 determined their vicinal position. Thus, compound 8 has to be either the 6'-hydroxy-3'-methoxy or the 6'-hydroxy-4'-methoxy isomer. The first of the latter two compounds is a known marine natural product, isospongiaquinone (7). As the 13Cdata of the quinone ring of isospongiaquinone differ substantially from those of compound 8(Table l),compound 8has to be the 6'-hydroxy-4'-methoxy isomer. As with the former compounds, the nmr assignments were achieved from COSY and CH correlation experiments. The CH correlations (shown in Table 1)were in full agreement with the suggested substitution pattern of the quinone. Small scale methylation of 8 with CH,N, gave as expected a dimethoxyavarone (6,,, 3.73 sand 3.68 s). Compounds 3, 4 , and especially 8 (but not 5 , 6, and 7) showed activity as inhibitors of HIV- 1 reverse transcriptase; these results are described more fully elsewhere (16). Avarol 11) was previously found to inhibit the growth of H N ; however, it was suggested that the mechanism of this activity does not involve the inhibition of the enzyme reverse transcriptase ( 14). Compounds 3-8 are cytotoxic, the IC,,'s against P388 mouse leukemia cells being 0.6, 1.2, <0.6, 10, >20 pg/ml for compounds 3, 4 , 6 , 7,and 8 , respectively. The compounds have also modest antifungal activity against Candiab albicans (MIC's of 12.5, 50, 12.5 pg/ml were measured for 3 , 4 , and 6, respectively). z zyxwvut EXPERMENTAL Ir s p x r a were recorded for solutions in CHCI, on a Perkin-Elmer Model 177, uv spectra for solutions in MeOH with a Varian Cary 219 spectrophotometer, and nmr spectra for solutions in CDCI, with a Bruker AM-360spectrometer (TMSas internal reference) equipped with an hpect 3000 computer. M w spectra were measured with a Finnigan 4020 quadrupole spectrometer equipped with a data system. The cd w e was recorded with a Jaxo 5OOC spectropolarimeter. Rotations were measured for solutions in MeOH . 96 zyxwvutsr zyxw zyx zyxwvut zyxw zyxwvut zyxw Wol. 5 4 , N o . 1 Journal of Natural Products zyxwvu zyxwvut C O L L E ~ OAND N EXTRACIION.-D. cinnra was collected at depths of 15-20 m near Ras Zaatir, in the Gulfof Eilat, the Red Sea. The specimens were frozen immediately after collection. The freeze-dried organism (30 g) was then extracted with CH2C12to give after evaporation a gum (2 g). A voucher specimen was kept in our collection in Tel Aviv, TA-YK 2465. 3 . - h oil: [ a 1 2 5+45.0° ~ (c= 0.060, CHCI,); cims mlz [M + 3H7+ 331 (loo%), COMPOUND [CI4H,,O3]+ 236 (lo%), [Cl4H2,1+ 191 (61%), [C,H8031+ 140 (5%). Found C 76.60, H 8.42; C21H2803requires C 76.79, H 8.59; v max 3400 br, 2850, 1630, 1350 cm-'; A rnax (MeOH) 256 (10200), 284 (3300);6 , (CDCI,) 7.05 (s, 3'-0H, exchangeable by D20), 6.75 (s, HA'), 6.75 (s, H-6'), 5.15 (bn,H-3), 2.65(d, H-11,]= 13.5), 2.51(d, H-ll,]= 13.51, 2.04(ba, H-l,]=6.1), 2.02(m, H-2), 1.85 (m, H-2) 1.63 (dt, H-6,]= 12.6, 3.11, 1.53 (4H, bn, H-1, Me-15), 1.36 (2H, m, H-7h 1.25t(H-8,]=6.2), 1.04(m,H-10), 1.03(m,H-6), 1.00(s,Me-14),0.93(d,Me-13,]=6.2),0.85 (I, Me-12); 8, (CDCI,) see Table 1. COMPOUND 4.-An oil: +65.6" (c=O.O32, CHCI,); cims d z [M-%H121+ 236 (45%), [CI4H2,]+ 191 (100%), [C,H50J+ 153 (10%);Y max 3400-3300 br, 2850, 1640, 1350 cm-'; A rnax (M~oH)256 (11400), 282 (8140); S, (CDCI,) 7.1 (s, 2 X OH, exchangeble by D,O), 6.73 (I, HA'), 5.14(b~,H-3),2.60(d,H-ll,]=13.3),2.57(d,H-ll,J=13.3),1.98(m,H-2), 1.96(m,H-U, 1.88 (m, H-2), 1.65(dt,H-6,]= 12.7, 3.2), 1.55(m,H-loverlapswithMe-15), 1.54(s,Me-15), 1.38(m, 2H. H-7), 1.30(t, H-8,]=6.2), 1.15 (m, H-6, overlapswith H-lo), 1.13 (m, H-lo), 1.01 (s, Me-14, 0.93 (d, Me,]= 5.91, 0.83 (s, Me-12); 6, (CDCI,) see Table 1. zyxwvutsr COMPOUND5 . - h oil: [ ~ ] ~ ~ ~ + 6 0(c=0.02, .0" MeOH); eims de [MH1+ 330 (IO%), [c14H2,]+ 191 (20961, [C,H8O31+ 140 (100%);Y rnax 3600-3300 br, 2900 cm-'; Amax (MeOH) 242 (2750), 294 (2200), 385 (500); 6 , (CDCI,) 6.58 (d, H-4',]=8.4), 6.19 (d, H-3',]=8.4), 5.50(bn, OH), 5.20(ba, H-3),4.70(bn,OH),4.50(bn,OH), 1.55s, 1.02s, 0.90s, 0.86(dJ=6.4)4xMe; 6, (CDCI,) see Table 1. COMPOUND6 . - h oil: [ ~ ] ~ ~ ~ + 1 8(c=O.44, .9" CHCI,); cims d z [MH}+ 373 (9%), [C21H!,@3]+ 331 (2561, [C,,H3102]+ 315 (6%), [C&&i+ 191 (IO%) I,~ , O J + 181 (21%); Y mag 3600-3300 br, 2900, 1720, 1425, 1175 cm-'; A max (MeOH) 298 (4000);6 , (CDCI,) 6.80 (4H- 3',]=8.7),6.31(d,H-4',]=8.7),5.35(bn,OH,exchangeablebyD,O),5.15(ba,H-3),5.00(OH, bn, exchangeable by D,O), 2.75 (d, H-ll,]= 14.2), 2.67 (d, H-ll,]= 14.2), 2.25 (s, 6'-OAc), 2.03 (2H,m,H-2), 1.95(rn,H-l), 1.65(m,H-8), 1.61(ddd,H-6,]=12.3,4.3,3.0), 1.56(4H,m,H-l, overlaps with Me-15), 1.55 (bn, Me-15), 1.37 (m, H-IO), 1.35 (2H, m, H-7), 1.12 (dt, H-6,1= 12.3, 8.5), 1.01 (s, Me-14), 0.89(s, Me-12), 0.83 (d, Me-l3,]=6.5); 6,(CDCI3)seeTable 1. 7.-h oil: [u]~'D-4.2" (c=0.018, CHCI,); cims d z [M+ 3H]+ 373 (12%), COMPOUND [C,,HBO31+ 329 (43%), [C,,H,,]+ 191 (loo%), [CgH9041+ 181 (26%); Y rnax 1720, 1630, 1350 crn-'; A rnax (MeOH) 250 (10300), 330 (550); 6 , (CDCI,) 6.80 (d, H-3',1= 10.1)*6.76 (d, H-4'1 ]~10.1),5.10(ba,H-3),2.62(d,H-11,]~13.5),2.45(d,H-ll,]~13.5),2.30~~6'-0A~~,2.0~m, H-2), 1.87 (m, H-1), 1.63 (2H, m, H-8, H-6), 1.58 (4H, brs, H-1, Me-15), 1.36 (3H, m, H-6, H-7h 1.10(dt, H-6,]= 12.3,8.5), l.OO(s, Me-14), 0.90(d, Me-13,]= 5.6), 0.86(~,Me-I2);6~(CDC13)see Table 1. [m+ COMPOUND&-An oil: [a]25D +75" (c=O.@O, CHCI,); c h d z 358 (4%), [C&23]+ 191 (30%), [C8HIo0J+ 170 (100%);Y max 3200,2900, 1670, 1640, 1620, 1350, 1130cm-'; A mag (~eOH)256(12700), 294(54O);6,(CDCl3)7.50(ba, 6'-0H, exchangeablebyD20),5.81(s, H-3'), 5.14 (brs, H-3), 3.80 (3H, s, 3'-OMe), 2.59 (d, H-ll,]= 13.51, 2.52 (d, H-ll,]= 13.51, 2.10 (2H, m, H-2), 2.0 (m, H-I), 1.65 (2H, m, H-8, H-6, not resolved), 1.54 (4H, bn, H-1, Me-15), 1.34 (3H, m, H-6, H-7), 1.09(m, H-6), 1.05 (s, Me-14), 0.96(d, Me-13,]=5.7), 0.84(s, Me-l2);6,(CDCl3)see Table 1. OXIDATIONOF COMPOUND 6 TO 7.-To a solution of 8 mg of 6 in 40 ml of Et,O was added Ag20 (80mg), and the heterogeneous m i m e was stirred overnight at room temperature. The reaction mixture was filtered and the solvent evaporated to give 4 mg of a yellow oil. The ir, ms, and 'H nmr of this product were identical with those of naturally occurring 7. ACEMATION OF 5 AND 6 TO COMPOUND 9.-A solution of 15 mg of 5 or 6 in 1 mJ of pyridine and 1ml of Ac20 was allowed to stand overnight at room temperature, and then the mixture was diluted with H,O and extracted with CHCI,. The CHCI, solution was washed with H 2 0and dried over MgSO4. Evaporation gave 11 mg ofpure 9: cims [C13H140d+266 (l%), [C&2J+ 191 (100%); v max 1760, 12 10 cm- 6 , (CDCI,) 7.09 (d, H-4' ,] = 8.9), 7.0 1 (d, H-3 ',] = 8.9),5.18 (bn,H-3), 2.68 (d, H- 11 ]=13.5),2.36(d,H-ll,]=13.5),2.26(6H,s,2xOAc~,2.04~3H,s,OAc~, 1.85(m,H-1), 1.66 '; I zyxwvu zyxwvu zyxw zyxwvut Jan-Feb 199 11 Hirsch et al. : Avarone and Avarol Derivatives 97 zyxwvut zyxwvu zyxwvu (2H, m, H-8, H-6), 1.58 (4H, bn, H-1, Me-15), 1.34 (3H, m, H-6, H-7). 1.14 (m, H-6), 1.05 (3H, s, Me-14), 0.86(3H, s, Me-12), 0.70(3H, d, Me-13,]=6.3). OXIDATIVE HYDROBOUTION OF 9 TO AFFORDCOMPOUND 10.-To a solution of triacetate 9 (20 mg) in dry Et,O (2 ml), NaBH, (12 mg) and ZnC1, (3.5 mg) were added, followed by a solution of boron trifluoride-etherate (0.1 ml) in dry Et,O (1 ml),and the mixnVewas stirred at room temperature for 2 h. An excess ofJones reagent was added, and the mixture was refluxed for 2 hand then diluted with H 2 0 . The product was extracted with Et20, and the combined Et20 extracts were washed with H,O, aqueous NaHCO,, and H,O again, dried over MgS0,, and evaporated to give ketone 10 (4 mg): cims [M - C2H202]+ 398 (2%), [C13Hl,0d+ 267 (loo%), IC&230,1+ 207 (20%); Y f n a ~1760, 1705 cm-'; 6,(CDCI,)7.09(d,H-4',]=8.9), 7.01(d,H-3',]=8.9), 2.68(d, H-ll,]=13.5), 2.46(d, H-ll,]= 13.5), 2.36 (6H, s, 2 X OAc), 2.26 (3H, s, OAc), 2.10 (3H, m, H-2, HA), 1.90 (m, H-l), 1.66(2H, m, H-8, H-6), 1.58 (m, H- 1) 1.34 (3H, m, H-6, H-7), 1.14 (m, H-6), 1.05 (3H, s, Me- 14), 0.89(3H, d, Me-4,]=6.8), 0.84(3H, s, Me-12), 0.70(3H, d, Me-13,]=6.3);cd(MeOH)IB2&l -4910. COMPOUND11.-A solution of 10 mg of 2 (0.028 mmol) and 3.4 mg (0.03 1 m o l ) ofphenylene diamine in 10 ml MeOH was allowed to stand overnight at room temperature. The solvent was evaporated and the reaction mixture purified on a Sephades LH-20 column eluted with hexane-CHC1, (3:7). Compound 11was achieved as a cherry-red oil: cims [MI+ 430 (I%), tCI7H2d+230 (46%), ICI3Hl,,N21+ 194 (10%); [CI5Hz5]+205 (3%), [CI4H2,]+ 191 (32%); A max(MeOH) 230 (23740), 294 (8500), 320 (9200), 380 (2600), 460 (1080); 6 , (Me,CO-dd 8.26 bn, 7.08 d (]=7.9), 7.05 t (]=7.9), 6.85 d (/=8.2), 6.67t(]=8.3), 5 . 2 2 ~ , 4 . 3 9 ~ , 4 . 3 8 ~ , 2 . 4 7 d ( ] = 1 32.37d(]=13.9), .9), 1.10s, 1.0d (]= 5.6), 0.85 s. ACKNOWLEDGMENTS This work ws supported financially by HBOI,Ft. Pierce, Florida. We are grateful to D n . P. McCarthy and N. Burres at HBOI for the bioassays. We thank Prof. J. Vacelet, Marseille, for the identification of the sponge. LITERATURE CITED 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. W. Fenical, J.J. Sims, D. Squatrito, R.M. Wing, andP. Radlick,]. Org. Cbem., 38,2383(1973). W. Fenical and 0. McConnell, Expcrimriu, 31, 1004 (1975). G. Cimino, S. De Stefano, W . Fenical, L. Minale, and J.J. Sims, EwpCrmriu, 31, 1250 (1975). G. Cimino, S. De Stefano, and L. 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Hizi and S. Loya, FEBSLrtt., 269 (l), 131 (1990). zyxwvutsrq Rereid I3 Apil I 990 View publication stats