4 Bioactive Marine Natural Products gave an unsaturated glycol.Laureatin was finally assigned structure(7)on the basis of chemical degradation studies and NMR spectroscopic data.Other brominated metabolites which have been isolated from Laurencia nipponica, and chlorofucin.The total syntheses of (+)-prela tinand ()urall have e recently. Laureatin(7)and isolaureatin exhibit significant larvicidal activity (ICso)0.06 and 0.50 ppm,respectively,in mosquitos.Brominated compounds isolated from marine algae,particularly bromophenols,are toxic and due to this they are not of clinical value. 3.3 Nitrogen Hetero clics Marine algae had yielded nitrogen containing heterocyclic compounds.Of these the most interesting compounds are domoic acid(8)and the kainic acid. H COOH CH2COOH -COOH Domoic acid (8)(CIsHzINO).m.p.217C (dec.):[alp-109.6 [H2O]an anthelmintic agent was first isolated from the alga Chondria armata. The acid had UV242 nm (log E 4.42).Catalytic reduction of the compound ith P-Ogave.Acetylation of the compound gave N- acetyl derivative,m.p.1C[6[H]:24 nm (og.48) Domoic acid showed marked anthelmintic activity.It was found to be very effective inexpelling ascaris and pinworms without any observable side effects. 3.4 Kainic Acids In Asia,the dried red alga Digenea simplex is widely used as an anthelmintic It is found very effective in the treatment of ascariasis.30 In the Mediterranean. extract of the alga Corallina officinalis is also used for the same purpose. Kainic acids as the active principles had been isolated from thes e algae.Of 高 by its synthesis.32 The stereochemistry of a-kainic acid is shown in(9). -COOH COOH
4 Bioactive Marine Natural Products gave an unsaturated glycol. Laureatin was finally assigned structure (7) on the basis of chemical degradation studies and NMR spectroscopic data. Other brominated metabolites which have been isolated from Laurencia nipponica, are prelaureatin, laurallene, isolaurallene, bromofucin, and chlorofucin. The total syntheses of (+)-prelaureatin and (+)-laurallene have been achieved recently.24 Laureatin (7) and isolaureatin exhibit significant larvicidal activity (IC50) 0.06 and 0.50 ppm, respectively, in mosquitos. Brominated compounds isolated from marine algae, particularly bromophenols, are toxic and due to this they are not of clinical value. 3.3 Nitrogen Heterocyclics Marine algae had yielded nitrogen containing heterocyclic compounds. Of these the most interesting compounds are domoic acid (8) and the kainic acid. Domoic acid (8) (C15H21NO6), m.p. 217°C (dec.): [α]D – 109.6° [H2O] an anthelmintic agent was first isolated from the alga Chondria armata. 25-29 The acid had UV λmax 242 nm (log ε 4.42). Catalytic reduction of the compound with Pt-O2 gave tetrahydrodomoic acid. Acetylation of the compound gave Nacetyl derivative, m.p. 121°C; [α]D–56° [H2O]; λmax 242 nm (log ε 4.48). Domoic acid showed marked anthelmintic activity. It was found to be very effective in expelling ascaris and pinworms without any observable side effects. 3.4 Kainic Acids In Asia, the dried red alga Digenea simplex is widely used as an anthelmintic. It is found very effective in the treatment of ascariasis.30 In the Mediterranean, extract of the alga Corallina officinalis is also used for the same purpose. Kainic acids as the active principles had been isolated from these algae. Of the kainic acids, α-kainic acid was the most active constituent. The structure (9) for α-kainic acid had been assigned by degradation studies31 and confirmed by its synthesis.32 The stereochemistry of α-kainic acid is shown in (9).33 9 8
Bioactive Metabolites of Marine Algae,Fungi and Bacteria 5 Isomers of a-kainic acid had been isolated are y-allo-kainic acid (10)4 and y-kaini ic acid lactone (1) kainic acid and L-a-allo-o-kainic acid are configurational isomers.In o- kainic acid the substituents at C-2 and C-3 and at C-3 and C-4 are trans and cis.respectively.In o-allokainic acid configurations at both the centres are trans.a-Kainic acid lactone was considered to be an artifact.36 a-Kainic acid had been found effective in the treatm iasis with a single dose of 5 to 10 mg per adult resulting in a 40 to 70%reduction in the population of instestinal parasitic worms.a-Allokainic acid was found to have far less anthelmintic activity.Several preparations of kainic acids are available in the market.including 'Digenin'and'Helminal'(The Merck Index,1968).This ents on of the few ins ances in which clinically useful pharmaceutical product has been is ated from marine source COOH COOH 3.5 Guanidine Derivatives Certain shellfish periodically become poisonous to humans.It is now well established that the substance responsible is produced by a marine plankton, Gonyaulax catenella.At certain unpredictable time the red plankton multiply nd cause“red tide”Although nany fishes are killed by this "red tide' mussels and clam and concent e the toxic prn .thus bec min laskan butter clam. Cm the ale ll (12). Saxitoxin (CIoHoN-O)when heated with P/HI in acetic acid,gave weakly basic compound I,CsHoN2O(m.p.100-102C).NMR analysis of Iindicates
Bioactive Metabolites of Marine Algae, Fungi and Bacteria 5 Isomers of α-kainic acid had been isolated from alga Digenea. The isomers isolated are γ-allo-kainic acid (10) 34 and γ-kainic acid lactone (11).35 L-α- kainic acid and L-α-allo-α-kainic acid are configurational isomers. In α- kainic acid the substituents at C-2 and C-3 and at C-3 and C-4 are trans and cis, respectively. In α-allokainic acid configurations at both the centres are trans. α-Kainic acid lactone was considered to be an artifact.36 α-Kainic acid had been found effective in the treatment of ascariasis, with a single dose of 5 to 10 mg per adult resulting in a 40 to 70% reduction in the population of instestinal parasitic worms. α-Allokainic acid was found to have far less anthelmintic activity. Several preparations of kainic acids are available in the market, including ‘Digenin’ and ‘Helminal’ (The Merck Index, 1968). This represents one of the few instances in which clinically useful pharmaceutical product has been isolated from marine source. 3.5 Guanidine Derivatives Certain shellfish periodically become poisonous to humans. It is now well established that the substance responsible is produced by a marine plankton, Gonyaulax catenella. At certain unpredictable time the red plankton multiply and cause “red tide”. Although many fishes are killed by this “red tide”, mussels and clams survive and concentrate the toxic principles, thus becoming poisonous to humans. The toxin isolated from the Alaskan butter clam, Californian mussel37 and the alga Gonyaulax catenella38-40 is called saxitoxin (12). Saxitoxin (C10H19N7O4) when heated with P/HI in acetic acid, gave weakly basic compound I, C8H10N2O (m.p. 100-102°C), NMR analysis of I indicates 10 11 12 13 1 2 10 11 9 7 3 4 5 6 8
6 Bioactive Marine Natural Products the presence of one C-CH group.2 On oxidation with potassium permanganate,urea and guanidinoacetic acid were obtained.Hydrogenation of I in the rese f patinumd (00me hydrogen absorption) gave II.CsH4N2O (m. 129-131C)which also contain d one C- CH. hydrolysis of II edto the strony basic. hygroscopic oily diamine III,CH6N2 and on heating with Pd-C.III resulted in the formation of a substance which readily gave a positive Ehrlich test for pyrroles.On the basis of these data,it was concluded that III was a pyrrolidine and II was a saturated cvclic urea.This conclusion was fully supported on and its stro infrared itst absorp he structure (1oed th basis of degradation studies and spectroscopic analysis.Saxitoxin blocks nerve conduction by specifically interfering with the intital increase in sodium permeability of the membrane.The symptoms caused by the toxin include peripheral paralysis.Inextreme cases,complete loss of strength in the muscles and finally death occurred which is ca sed due to respiratory failure.Saxitoxin is absorbed from the gastro-intestinal tract.It produced no major vascular action.The oral LDso for toxin in various species of animals is reported.In man death had occurred following ingestion of as little as I mg of toxin.+ The toxic compounds from marine algae appear to have biomedical potential. The compounds with neurotropic effects may yield important drugs 3.6 Phenazine Derivatives The marine alga Caulerpa lamourourii is widely distributed in the Phillippines The upper branches are eaten as a'salad'.despite their peppery and astringent taste.However,the alga is found toxic to some individuals.Chemical investigation of the alga had furnished caulerpicine,caulerpin,cholesterol. araero-itosteroa Caulerpn had s een isol fom Caulerpa serlarioides.C.racems var.claviferand ceic from C.racemosa. Caulerpin (13)(C24HIsN2O4)(M*398)red prisms m.p.317C had ma 222.270.292.317nm(e50.000.27.000.29.000and35.000:IR bands at 1684,1631 and 1613 cm-suggesting the resence of carbonyl functions in conjugation with aroma The MR of the ound indicated the presence of 18 protons t 6.17 (6H.2 OMe).2.4-3.0 (8H.m) 1.76(2H,s)and 1.36(2H,s).Aromatic protons signal at t 2.4-3.0 and 1.79 and the IR bands at 730 and 920 cm-suggested the presence of two identically substituted aromatic ring systems.This was substantiated by elimination of 26m units(CH=CH)i s spectrum aulerpin,c aulerpinic acid and decarboxy caulerpin acid.Caulerpin containe two me oxy groups in the form of o,B-unsaturated methyl ether group [Vmax 1685 cm NMR t6.17 (6H)].Its mass spectrum supported the assignment m/z 398(M*),366 (Mt-MeoH,338(366-C0).306(338-MeOH).339M-C0Me)and280 (M+-2CO2Me).The M*peak in the mass spectrum was the base peak
6 Bioactive Marine Natural Products the presence of one C—CH3 group.41,42 On oxidation with potassium permanganate, urea and guanidinoacetic acid were obtained. Hydrogenation of I in the presence of platinum oxide (200 mole % hydrogen absorption) gave II, C8H14N2O (m.p. 129-131°C) which also contained one C—CH3 group. Strong acid hydrolysis of II led to the strongly basic, and highly hygroscopic oily diamine III, C7H16N2 and on heating with Pd-C, III resulted in the formation of a substance which readily gave a positive Ehrlich test for pyrroles. On the basis of these data, it was concluded that III was a pyrrolidine and II was a saturated cyclic urea. This conclusion was fully supported with its ultraviolet absorption and its strong infrared absorption at 3410 and 1635 cm–1 in chloroform. The structure (12) to saxitoxin was assigned on the basis of degradation studies and spectroscopic analysis. Saxitoxin blocks nerve conduction by specifically interfering with the intital increase in sodium permeability of the membrane. The symptoms caused by the toxin include peripheral paralysis. In extreme cases, complete loss of strength in the muscles and finally death occurred which is caused due to respiratory failure.43 Saxitoxin is absorbed from the gastro-intestinal tract. It produced no major vascular action. The oral LD50 for toxin in various species of animals is reported. In man death had occurred following ingestion of as little as 1 mg of toxin.44 The toxic compounds from marine algae appear to have biomedical potential. The compounds with neurotropic effects may yield important drugs. 3.6 Phenazine Derivatives The marine alga Caulerpa lamourouxii is widely distributed in the Phillippines. The upper branches are eaten as a ‘salad’, despite their peppery and astringent taste. However, the alga is found toxic to some individuals. Chemical investigation of the alga had furnished caulerpicine, caulerpin, cholesterol, taraxerol, β-sitosterol and palmitic acid.45 Caulerpin had also been isolated from Caulerpa sertularioides, C. racemosa var. clavifera46 and caulerpicin from C. racemosa. 47 Caulerpin (13) (C24H18N2O4) (M+ 398) red prisms m.p. 317°C had λmax 222, 270, 292, 317 nm (ε 50,000, 27,000, 29,000 and 35,000); IR bands at 1684, 1631 and 1613 cm–1 suggesting the presence of carbonyl functions in conjugation with aromatic system. The NMR spectrum of the compound indicated the presence of 18 protons τ 6.17 (6H, 2 OMe), 2.4-3.0 (8H, m), 1.76 (2H, s) and 1.36 (2H, s). Aromatic protons signal at τ 2.4-3.0 and 1.79 and the IR bands at 730 and 920 cm–1 suggested the presence of two identically substituted aromatic ring systems. This was substantiated by elimination of 26 mass units (CH=CH) in the mass spectrum of caulerpin, caulerpinic acid and decarboxy caulerpin acid. Caulerpin contained two methoxy groups in the form of α,β-unsaturated methyl ether group [νmax 1685 cm–1; NMR τ 6.17 (6H)]. Its mass spectrum supported the assignment m/z 398 (M+ ), 366 (M+ –MeOH), 338 (366–CO), 306 (338–MeOH), 339 (M+ –CO2Me) and 280 (M+–2CO2Me). The M+ peak in the mass spectrum was the base peak
Bioactive Metabolites of Marine Algae,Fungi and Bacteria 7 to secondary amino groups.The functions were conjugated with the two methoxy carbonyl groups as indicated by the low frequency carbonyl absorption (1685 cm-).The methoxy carbonyl groups were placed at the two a-positions of the two naphthalene rings oni tthe NHgroup s at the B. ositions This arrangemen for the stro hydrogen bonding ofthe -NH protons.Caulerpinic acid when heated with copper bronze in quino 200-210C vielded a decarboxylated compound m.p.>300C.(M282).On the basis of these studies caulerpin was assigned the structure o.B- dihydrodibenolb ilphenazine-5.12-dicarboxvlate (13)The stability of the comp und was state ed to fav structu ather than the isomer Caulerpin caus ed a n anesthetic action when placed in the m outh which resulted in numbness of the lips and tongue.In some people it produced toxic effects.The toxic syndrome had been reported to be some what similar to that produced by ciguatera fish poisoning. H.CO OCH 3.7 Amino Acids and Amines Extracts of the marine algae Laminaria angustata and Chondria amata ar reported to contain agents with hypotensive and other pharmacologica om?umher f mlae e Te campud e chaneted Me 14 Laminine was isolated from water extracts of Laminaria angustata by amberlite ion exchange resin,IR-120in acidic form and subsequent formation of reineckate and oxalate salts.The other amino acids isolated from this
Bioactive Metabolites of Marine Algae, Fungi and Bacteria 7 Saponification of caulerpin with alcoholic KOH yielded caulerpinic acid (C22H14N2O4) (M+ 370). The two exchangeable protons at τ 1.36 were due to secondary amino groups. The functions were conjugated with the two methoxy carbonyl groups as indicated by the low frequency carbonyl absorption (1685 cm–1). The methoxy carbonyl groups were placed at the two α-positions of the two naphthalene rings conjugated with the NH groups at the β-positions. This arrangement accounted for the strong hydrogen bonding of the –NH protons. Caulerpinic acid when heated with copper bronze in quinoline at 200-210°C yielded a decarboxylated compound m.p. >300°C, (M+ 282). On the basis of these studies caulerpin was assigned the structure α,β- dihydrodibeno[b,i]phenazine-5,12-dicarboxylate (13).48 The stability of the compound was stated to favour the linear structure rather than the geometrical isomer. Caulerpin caused a mild anesthetic action when placed in the mouth, which resulted in numbness of the lips and tongue. In some people it produced toxic effects. The toxic syndrome had been reported to be some what similar to that produced by ciguatera fish poisoning. 3.7 Amino Acids and Amines Extracts of the marine algae Laminaria angustata and Chondria amata are reported to contain agents with hypotensive and other pharmacological properties. Laminine (14), a choline like basic amino acid had been isolated from a number of marine algae.49,50 The compound had been characterised as trimethyl(5-amino-5-carboxypentyl)ammonium oxalate (14). Several syntheses of laminine are reported.51 Laminine was isolated from water extracts of Laminaria angustata by amberlite ion exchange resin, IR-120 in acidic form and subsequent formation of reineckate and oxalate salts. The other amino acids isolated from this 13 14
8 Bioactive Marine Natural Products source were:L-lysine,L-arginine,ethanolamine and choline.Laminine monocitrate was found to have a transitory hypotensive effect.Laminine,in general,depressed the contraction of excited smooth muscles.Laminine e had an LDs in mouse,394 mg/kg.It is considered to be the widespread occurrence of volatile amines,such as methylamine, dimethylamine,trimethylamine,ethylamine,propylamine,isobutylamine, isoamylamine,2-phenylethylamine and 2-methylmercapto propylamine in red,green and brown algae.It is mentioned that biological activities of some of the extracts of the marine algae may be due to the presence of these amines 3.8 Sterols The presence of sterols in algae was first established by Heilbron et al33 and later by Tsuda et al.34 Gibbons et alss established the presence of 22- dehyd cholesterol and demosterol in red algae.However,lat the sterol content of red a were more varied th n had b sho Idler et al examined some species of red algae and found tha the three species contained C27,C2s and C29 sterols.An interesting feature of their result was the considerable variation in sterols content of four different samples of the alga Rhodymenia palmata.The percentage of demosterol,for exa varied from 97.2 to 7.7%in the mixture of sterols. cholesterol was detected in the concentration as high as97.3%and as low as 2.1%.Cholesterol was again found the major sterol of Rhodophyta.Four species of algae,Rhodymenia palmata,Porphyra purpurea,P.umbilicalis and Halosaccion ramentaceum were found to contain desmosterol as the Japanese and British investigators,only one sterol was detected in 25 species. while nine were found containing two sterols.Meunier et als had given a comparative data of 14 species of Rhodophyta.All the species examined were found to contain cholesterol (15)as the maior sterol except hyoned sciformis in which 22-dehydrocholesterol (16)was detected in the highest centration.Hypnea japonica was anothe example in which 22. dehydrocholesterol was present as the major sterol
8 Bioactive Marine Natural Products source were: L-lysine, L-arginine, ethanolamine and choline. Laminine monocitrate was found to have a transitory hypotensive effect. Laminine, in general, depressed the contraction of excited smooth muscles. Laminine monocitrate had an LD50 in mouse, 394 mg/kg. It is considered to be a potentially useful pharmacological agent. Steiner and Hartmann52 had reported the widespread occurrence of volatile amines, such as methylamine, dimethylamine, trimethylamine, ethylamine, propylamine, isobutylamine, isoamylamine, 2-phenylethylamine and 2-methylmercapto propylamine in red, green and brown algae. It is mentioned that biological activities of some of the extracts of the marine algae may be due to the presence of these amines. 3.8 Sterols The presence of sterols in algae was first established by Heilbron et al53 and later by Tsuda et al.54 Gibbons et al55 established the presence of 22- dehydrocholesterol and demosterol in red algae. However, later investigations showed that the sterol content of red algae were more varied than had been believed.56 Idler et al57 examined some species of red algae and found that the three species contained C27, C28 and C29 sterols. An interesting feature of their result was the considerable variation in sterols content of four different samples of the alga Rhodymenia palmata. The percentage of demosterol, for example, varied from 97.2 to 7.7% in the mixture of sterols. Similary, cholesterol was detected in the concentration as high as 97.3% and as low as 2.1%. Cholesterol was again found the major sterol of Rhodophyta. Four species of algae, Rhodymenia palmata, Porphyra purpurea, P. umbilicalis and Halosaccion ramentaceum were found to contain desmosterol as the main sterol. However, Hypnea japonica was the only alga having 22- dehydrocholesterol as the major sterol. Of the 34 algae investigated by the Japanese and British investigators, only one sterol was detected in 25 species, while nine were found containing two sterols. Meunier et al58 had given a comparative data of 14 species of Rhodophyta. All the species examined were found to contain cholesterol (15) as the major sterol except Hypnea musciformis in which 22-dehydrocholesterol (16) was detected in the highest concentration. Hypnea japonica was another example in which 22- dehydrocholesterol was present as the major sterol. 15 16 3 19 18 22 19 18 21 20 22 23 24