to date , dozens of simple monoindole derivatives were reported from marine sources , such as sponges [ 15 ] , ascidians , bryozoans , bacteria , and fungi . some of these metabolites were reported to exhibit antibacterial , antifungal , and auxin   activities . in our previous study on cytotoxic compounds from the marine sponge , we isolated a series of bisindole alkaloids . in our continuing search for cytotoxic metabolites from the same sponge compounds 12 and 46 were isolated for the first time from a natural source although they were previously reported as synthetic intermediates ( figure 1 ) . the molecular formula was established as c11h8brno3 on the basis of the eims and nmr data . in the lreims of 1 , a ( m ) ion cluster was observed at m / z 281/283 in the ratio of 1:1 that is characteristic of a monobrominated compound . analysis of the h , c , cosy , hmbc , and hsqc data , along with comparison of chemical shift values with those of known indole alkaloids , allowed us to establish a 6-bromoindol-3-yl residue as a partial structure of 1 . the singlet at h 8.45 ( h-2 ) , and a spin system comprised of signals at h 8.07 ( 1h , d , j=8.0 hz , h-4 ) , 7.40 ( 1h , dd , j=8.0 , 2.0 hz , h-5 ) , and 7.73 ( 1h , d , j=2.0 , h-7 ) indicated the presence of a 6-bromoindol-3-yl moiety ( table 1 ) . long - range correlations from h-4 ( h 8.07 ) to c-3 ( c 112.5 ) and c-6 ( c 116.2 ) , along with the cosy correlation between h-4 and h-5 , and the long - range correlations from h-5 ( h 7.40 ) to c-3a ( c 124.8 ) and c-7 ( 115.5 ) strongly suggested the presence of a 6-bromoindol-3-yl moiety . the nmr signals at c 178.2 ( c-8 ) , c 164.0 ( c-9 ) , and h 3.89 ( -och3 , 3h ) , along with the hmbc correlations of -och3/c-9 , suggested an oxoacetic acid methyl ester moiety . the eims fragments at m / z 194/196 , corresponding to c8h5brn , corroborated the presence of a bromoindole group . these fragments , along with the fragments at m / z 222/224 revealed the presence of a 3-carbonyl - bromoindole group , and established the connectivity between the 6-bromoindole moiety and the oxoacetic acid methyl ester moiety ( figure 2 ) . therefore , compound 1 was defined as ( 6-bromo-1h - indol-3-yl ) oxoacetic acid methyl ester . compound 1 was known as an intermediate in the synthesis of some marine natural products , such as didemnimides a and b , whereas it has not been reported from a natural source . pyruvic acid derivatives are unusual natural products , and most of indole pyruvic acid derivatives were isolated from marine sponges [ 1416 ] and ascidians . the molecular formula was established as c11h9no3 on the basis of the fabms and nmr data . in the lrfabms of 2 , the main difference from compound 1 was lack of bromine atom on the indole ring . therefore , compound 2 was defined as ( 1h - indol-3-yl ) oxoacetic acid methyl ester . compound 2 was known as an intermediate in the synthesis of natural products , such as didemnimides a and b , rebeccamycin , and 11-dechlororebeccamycin , whereas it has not been reported as a natural product . the molecular formula was established as c11h9no4 on the basis of the eims and nmr data . in the lreims of 1 , a ( m ) ion the main difference from compound 2 was an additional hydroxyl group on the indole ring . a singlet at h 8.22 ( 1h , s , h-2 ) , and a spin system comprised of signals at h 7.82 ( 1h , d , j=8.0 , h-4 ) , 6.74 ( 1h , dd , j=8.0 , 2.0 , h-5 ) , and 6.87 ( 1h , d , j=2.0 , h-7 ) , were observed in h nmr spectrum . the hmbc correlations from h-2 ( h 8.22 ) , h-5 ( h 6.74 ) , and h-7 ( h 6.87 ) to c-3a ( c 118.5 ) , from h-2 to c-3 ( c 112.5 ) and c-7a ( c 138.5 ) , and from h-5 ( h 6.74 ) to c-6 ( c 154.4 ) , indicated the presence of a 6-hydroxyindol-3-yl moiety . the eims fragments at m / z 132 and 160 corroborated the proposed structure ( figure 2 ) . therefore , compound 3 was defined as ( 6-hydroxy-1h - indol-3-yl ) oxoacetic acid methyl ester . to the best of our knowledge , compound 3 has not been reported previously either from a natural source or as a synthetic product . the molecular formula was established as c10h8n2o2 on the basis of the eims and nmr data . in the lreims of 3 , a ( m ) the main difference from compound 2 was the presence of an oxoacetamide moiety instead of the oxoacetic acid methyl ester moiety . the c signals at c 182.9 ( c-8 ) and c 165.9 ( c-9 ) , the h singlets at h 8.06 and h 7.69 ( each 1h , -nh2 ) ( tables 1 and 2 ) , along with the long - range correlation between -nh2 ( h 7.69 ) and c-8 ( c 182.9 ) , established an oxoacetamide moiety . the eims fragments at m / z 116 and 144 revealed the presence of a 3-carbonylindole group , and established the connectivity between the oxoacetamide moiety and the indole moiety ( figure 2 ) . thus , compound 4 was defined as ( 1h - indol-3-yl ) oxoacetamide , which was also known as an intermediate in the synthesis of some marine natural products , such as arborescidines   and dihydrohamacanthins , but has not been isolated previously from a natural source . the molecular formula was established as c10h7brn2o2 on the basis of the eims and nmr data . in the eims data of 5 , the main difference from compound 4 was an additional bromine atom on the indole ring . the fragments at m / z 194/196 and 222/224 revealed the presence of 3-carbonyl - bromoindole group ( figure 2 ) . therefore , compound 5 was defined as ( 6-bromo-1h - indol-3-yl ) oxoacetamide , which was also reported as an intermediate in the synthesis of some natural products , such as arborescidines , dihydrohamacanthins , but has not been isolated from a natural source . the molecular formula was established as c10h9no3 on the basis of the eims and nmr data . in the lreims of 6 , a [ m ] analysis of the h , c , cosy , hmbc , and hsqc data , allowed us to establish a 6-hydroxyindol residue as a partial structure of 6 . the long - range correlation from h-2 ( h 7.86 , 1h , s ) and -och3 ( h 3.76 , 3h , s ) to c-8 ( c 164.8 ) established the presence of a formic acid methyl ester and the connectivity between the 6-hydroxyindol moiety and the carboxylic acid methyl ester . the eims fragments at m / z 132 and 160 corroborated the proposed structure ( figure 1 ) . therefore , compound 6 was defined as ( 6-hydroxy-1h - indol-3-yl ) carboxylic acid methyl ester , which was known as an intermediate in the organic synthesis of a 5-ht4 receptor antagonist , but has not been reported from a natural source . according to the ms and nmr data of 7 , the main difference from 6 was lack of a hydroxyl group in the indole moiety . the ms and nmr data of 7 matched well with reported data , and was identified as ( 1h - indol-3-yl ) carboxylic acid methyl ester which was previously reported from marine - derived bacteria   and fungi , and red alga , with cytotoxicity against k562 human chronic leukemia ( mic s 14.0 g / ml ) . it is expected that ( 1h - indol-3-yl ) oxoacetamide derivatives serve as intermediate for the biogenesis of co - occurring bisindole alkaloids , topsentins and hamacanthins   ( scheme 1 ) . schiff base formation between amino and carbonyl groups may ( either via a or b ) leads to the genesis of hamacanthin a ( i ) and topsentin ( ii ) skeletons . n bond ( c ) in the topsentin skeleton , and successive schiff base formation between newly generated amino group and the intact carbonyl group may lead to a genesis of hamacanthin b skeleton ( iii ) . compounds 1 , 2 , and 47 were evaluated for cytotoxicity against a panel of five human solid tumor cell lines . compound 7 showed weak cytotoxicity to human lung cancer , human ovarian cancer , human skin cancer , human cns cancer , and human colon cancer with ed50 values 24.1 , 13.4 , 15.2 , 26.2 , and 4.85 g / ml , respectively , while other compounds did not show significant activity ( ed50>30g / ml ) . the ed50 values of doxorubicin against these tumor cell lines in the same experiment were 0.02 , 0.14 , 0.03 , 0.04 , and 0.10g / ml , respectively . h and c nmr spectra were recorded on a varian unity 300 and varian inova 500 instruments . chemical shifts were reported with reference to the respective residual solvent or deuterated solvent peaks ( h 2.5 and c 39.5 for dmso - d6 ) . fabms data were obtained on a jeol jms sx-102a ; eims data were obtained on a shimadzu qp5050 . hplc was performed with an ymc ods - h80 column ( 250  10 mm i.d . , 4 m , 80  ) and c18 - 5e shodex packed column ( 250  10 mm i.d . , 5 m , 100  ) using a shodex ri-71 detector . the sponges were collected by hand using scuba ( 20 m depth ) in october 2002 , off the coast of jeju island , korea . the two sponges were separated and only spongosorites sp . was subjected to chemical analysis . the morphology of the sponge was described elsewhere . a voucher specimen ( registry no . evaluation was performed at korea research institute of chemical technology . the frozen sponge ( 0.8 kg ) the meoh extract showed significant toxicity to brine shrimp larvae ( ld50 23.7 g / ml ) . aqueous meoh fraction was subjected to a reversed - phase flash column chromatography ( ymc gel ods - a , 60  , 230 mesh ) with a stepped gradient solvent system of 60 to 100% meoh / h2o to afford 16 fractions . fraction 2 ( 0.80 g ) , one of the bioactive fractions ( ld50 33.9 g / ml ) , was subjected to a reversed - phase hplc ( ymc ods - h80 column ) eluting with 75% meoh to afford 13 sub - fractions . compound 1 ( 0.95 mg ) was obtained by separation of the sub - fraction 28 on a reversed - phase hplc eluting with 58% mecn . compound 2 ( 2.2 mg ) was obtained by separation of the sub - fraction 2 - 2 on a reversed - phase hplc eluting with 35% mecn . the sub - fraction 2 - 1 was subjected to successive reversed - phase hplc ( ymc ods - h80 column ) eluting with 38% mecn , and further purification with 43% mecn ( c18 - 5e shodex packed column ) to afford compounds 3 ( 0.4 mg ) , 4 ( 0.78 mg ) and 6 ( 0.62 mg ) . compounds 5 ( 1.2 mg ) and 7 ( 3.3 mg ) were obtained by separation of sub - fractions 25 and 24 , respectively , on a reversed - phase hplc ( shodex c18 m10e column ) eluting with 42% mecn . ( 6-bromo-1h - indol-3-yl ) oxoacetic acid methyl ester ( 1 ) : yellow amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 281/283 ( m ) . ( 1h - indol-3-yl)oxoacetic acid methyl ester ( 2 ) : yellow amorphous powder ; ir ( film ) max 3206 ( br ) , 1727 , 1615 cm ; uv ( meoh ) max ( log  ) 362 ( 3.11 ) , 262 ( 3.03 ) nm ; h nmr data , see table 1 ; c nmr data , see table 2 ; lrfabms m / z 204 ( m + h ) . ( 6-hydroxy-1h - indol-3-yl ) oxoacetic acid methyl ester ( 3 ) : yellow amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 219 ( m ) . ( 1h - indol-3-yl ) oxoacetamide ( 4 ) : white amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 188 ( m ) . ( 6-bromo-1h - indol-3-yl ) oxoacetamide ( 5 ) : yellow amorphous powder ; ir ( film ) max 3386 , 3211 , 1663 , 1591 , 1572 , 1407 cm ; uv ( meoh ) max ( log  ) 320 ( 2.61 ) , 275 ( 2.75 ) , 258 ( 2.73 ) , 212 ( 3.21 ) nm ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 266/268 ( m ) . ( 6-hydroxy-1h - indol-3-yl ) carboxylic acid methyl ester ( 6 ) : colorless oil ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 191 ( m ) . ( 1h - indol-3-yl ) carboxylic acid methyl ester ( 7 ) : yellow amorphous powder ; ir ( film ) max 3255 ( br ) , 1693 , 1620 , 1591 , 1531 , 1444 , 1197 cm ; uv ( meoh ) max ( log  ) 349 ( 2.62 ) , 240 ( 2.75 ) ; lreims m / z 175 ( m ) . a panel of five human solid tumor cell lines , human lung cancer , human ovarian cancer , human skin cancer , human cns cancer , and human colon cancer , were used to screen cytotoxicity of the compounds based on an established protocol . h and c nmr spectra were recorded on a varian unity 300 and varian inova 500 instruments . chemical shifts were reported with reference to the respective residual solvent or deuterated solvent peaks ( h 2.5 and c 39.5 for dmso - d6 ) . fabms data were obtained on a jeol jms sx-102a ; eims data were obtained on a shimadzu qp5050 . hplc was performed with an ymc ods - h80 column ( 250  10 mm i.d . , 4 m , 80  ) and c18 - 5e shodex packed column ( 250  10 mm i.d . , 5 m , 100  ) using a shodex ri-71 detector . the sponges were collected by hand using scuba ( 20 m depth ) in october 2002 , off the coast of jeju island , korea . the two sponges were separated and only spongosorites sp . was subjected to chemical analysis . the morphology of the sponge was described elsewhere . a voucher specimen ( registry no . evaluation was performed at korea research institute of chemical technology . the frozen sponge ( 0.8 kg ) the meoh extract showed significant toxicity to brine shrimp larvae ( ld50 23.7 g / ml ) . aqueous meoh fraction was subjected to a reversed - phase flash column chromatography ( ymc gel ods - a , 60  , 230 mesh ) with a stepped gradient solvent system of 60 to 100% meoh / h2o to afford 16 fractions . fraction 2 ( 0.80 g ) , one of the bioactive fractions ( ld50 33.9 g / ml ) , was subjected to a reversed - phase hplc ( ymc ods - h80 column ) eluting with 75% meoh to afford 13 sub - fractions . compound 1 ( 0.95 mg ) was obtained by separation of the sub - fraction 28 on a reversed - phase hplc eluting with 58% mecn . compound 2 ( 2.2 mg ) was obtained by separation of the sub - fraction 2 - 2 on a reversed - phase hplc eluting with 35% mecn . the sub - fraction 2 - 1 was subjected to successive reversed - phase hplc ( ymc ods - h80 column ) eluting with 38% mecn , and further purification with 43% mecn ( c18 - 5e shodex packed column ) to afford compounds 3 ( 0.4 mg ) , 4 ( 0.78 mg ) and 6 ( 0.62 mg ) . compounds 5 ( 1.2 mg ) and 7 ( 3.3 mg ) were obtained by separation of sub - fractions 25 and 24 , respectively , on a reversed - phase hplc ( shodex c18 m10e column ) eluting with 42% mecn . ( 6-bromo-1h - indol-3-yl ) oxoacetic acid methyl ester ( 1 ) : yellow amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 281/283 ( m ) . ( 1h - indol-3-yl)oxoacetic acid methyl ester ( 2 ) : yellow amorphous powder ; ir ( film ) max 3206 ( br ) , 1727 , 1615 cm ; uv ( meoh ) max ( log  ) 362 ( 3.11 ) , 262 ( 3.03 ) nm ; h nmr data , see table 1 ; c nmr data , see table 2 ; lrfabms m / z 204 ( m + h ) . ( 6-hydroxy-1h - indol-3-yl ) oxoacetic acid methyl ester ( 3 ) : yellow amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 219 ( m ) . ( 1h - indol-3-yl ) oxoacetamide ( 4 ) : white amorphous powder ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 188 ( m ) . ( 6-bromo-1h - indol-3-yl ) oxoacetamide ( 5 ) : yellow amorphous powder ; ir ( film ) max 3386 , 3211 , 1663 , 1591 , 1572 , 1407 cm ; uv ( meoh ) max ( log  ) 320 ( 2.61 ) , 275 ( 2.75 ) , 258 ( 2.73 ) , 212 ( 3.21 ) nm ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 266/268 ( m ) . ( 6-hydroxy-1h - indol-3-yl ) carboxylic acid methyl ester ( 6 ) : colorless oil ; h nmr data , see table 1 ; c nmr data , see table 2 ; lreims m / z 191 ( m ) . ( 1h - indol-3-yl ) carboxylic acid methyl ester ( 7 ) : yellow amorphous powder ; ir ( film ) max 3255 ( br ) , 1693 , 1620 , 1591 , 1531 , 1444 , 1197 cm ; uv ( meoh ) max ( log  ) 349 ( 2.62 ) , 240 ( 2.75 ) ; lreims m / z 175 ( m ) . a panel of five human solid tumor cell lines , human lung cancer , human ovarian cancer , human skin cancer , human cns cancer , and human colon cancer , were used to screen cytotoxicity of the compounds based on an established protocol .
seven ( 17 ) monoindole derivatives were isolated from the meoh extract of a marine sponge spongosorites sp . by bioactivity - guided fractionation . 
 the planar structures were established on the basis of nmr and ms spectroscopic analyses . 
 compounds 15 are unique indole pyruvic acid derivatives . compounds 12 and 46 
 are isolated for the first time from a natural source although they were previously reported as synthetic intermediates . 
 compound 3 was defined as a new compound . 
 co - occurring bisindoles such as hamacanthins and topsentins might be biosynthesized by condensation of two units of these compounds . 
 the compounds were tested for cytotoxicity against a panel of five human solid tumor cell lines , and compound 7 displayed weak activity .