Acylated and deacylated saponins of Quillaja saponaria mixture as adjuvants for the FML-vaccine against visceral leishmaniasis.

The adjuvant of the FML-vaccine against murine and canine visceral leishmaniasis, the Riedel de Haen saponin mixture, was fractionated by ion exchange chromatography on DEAE-cellulose to afford one TLC homogeneous Quillaja saponaria Molina QS21 saponin fraction (18.0%), a mixture of two deacylsaponins (19.4%), sucrose (39.9%), sucrose and glucose (19.7%), rutin (0.8%) and quercetin (2.2%), that were identified by comparison of 1H and 13C NMR spectroscopy. The QS21 shows the typical aldehyde group in C-23 (65% equatorial) and a normonoterpene moiety acylated in C-28. The deacylsaponins show the aldehyde group but do not have the normonoterpene moiety. Balb/c mice were vaccinated with 150 microg of FML antigen of Leishmania donovani and 100 microg of each obtained fraction and further challenged by infection with 10(8) amastigotes of Leishmania chagasi. The safety analysis and the effect on humoral and cellular immune responses and in clinical signs showed that the QS21 saponin and the deacylsaponins are the most active adjuvant compounds of the Riedel the Haen saponin mixture. Both induced the highest and non-significantly different increases in DTH, CD4+ T lymphocytes in spleen, IFN-gamma in vitro, body weight gain and the most pronounced reduction of parasite burden in liver (95% for QS21 and 86% for deacylsaponins; p>0.05). While the QS21 showed mild toxicity, significant adjuvant effect on the anti-FML humoral response before and after infection, and decrease in liver relative weight, the deacylsaponins showed no toxicity, less haemolysis and antibody and DTH responses increased mainly after infection, still inducing a stronger Leishmania-specific in vitro splenocyte proliferation. Our results confirm in the Riedel de Haen saponin extract the presence of deacylsaponins normonoterpene-deprivated which are non-toxic and capable of inducing a specific and strong immunoprotective response in vaccination against murine visceral leishmaniasis.

An anti-inflammatory and immunomodulatory polysaccharide from Orbignya phalerata.

A polysaccharide, a glucan with mean M(r) of 1.0 x 10(6) (MP1), was isolated from the mesocarp of fruits of Orbignya phalerata. Chemical and spectroscopic studies indicated that MP1 has a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with (3-->4), (4-->6), and with (3-->6) branching points. MP1 enhanced phagocytosis in vivo and exhibited anti-inflammatory activity.

Biochanin A triglycoside from Andira inermis.

A new isoflavonol triglycoside, biochanin A 7-O-beta-D-apiofuranosyl-(1-->5)-beta-D-apiofuranosyl-(1-->6 )-beta-D- glucopyranoside (1), was isolated from Andira inermis roots in addition to the known compounds genistein 7-O-beta-D-apiofuranosyl-(1-->6)-beta-D-glucopyranoside and lanceolarin.

Anti-inflammatory and immunologically active polysaccharides of Periandra mediterranea.

Three polysaccharides, glucans with mean M(r)'s of 1.5 x 10(5), 3.6 x 10(4) and 2.1 x 10(4), were isolated from dried roots of Periandra mediterranea by fractionation on Sephacryl S-300 HR and Sephadex G-25. Chemical and spectroscopic studies indicated that they have a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with both (3-->4) and (4-->6) branching points. The polysaccharides enhance phagocytosis in vivo, and exhibit anti-inflammatory activity.

Flavonol glycosides from Costus spicatus.

Two flavonol diglycosides, tamarixetin 3-O-neohesperidoside, kaempferide 3-O-neohesperidoside and the known quercetin 3-O-neohesperidoside, together with six other known flavonoids were isolated from the leaves of Costus spicatus and their structures were elucidated by a combination of spectroscopic and chemical methods. The flavonol diglycosides were evaluated for inhibitory activity of nitric oxide production by activated macrophages (Fig. 1).

Immunologically active polysaccharides from Centrosema pubescens.

Two polysaccharides with mean M(r)s of 2.0 x 10(6) and 3.75 x 10(5), were isolated from powdered seeds of Centrosema pubescens by fractionation on Sephacryl S-300 HR. Chemical and spectroscopic studies indicated that they have a backbone chain composed of beta-(1-->4)-linked D-galactopyranose residues having branches composed of alpha-(1-->5)-linked L-arabinofuranose residues at position 6 of D-galactose of the backbone chain. The polysaccharides showed reticuloendothelial system-potentiating activity in a carbon clearance test.

Flavonol glycosides from leaves of Costus spiralis.

A new flavonol diglycoside, 3,5-dihydroxy-7,4'-dimethoxyflavone 3-O-neohesperidoside (1), together with four known flavonol 3-O-neohesperidosides were isolated from the leaves of Costus spiralis and their structures were elucidated by a combination of spectroscopic methods and chemical reactions.

A furostanol glycoside from rhizomes of Costus spicatus.

A new furostanol glycoside was isolated from the rhizomes of Costus spicatus. Its structure was established as (3 beta,22 alpha,25R)-26-(beta -D-glucopyranosyloxy)-22-methoxyfurost-5-en-3-yl O-D-apio-beta-D-furanosyl-(1-->2)-O-[6-deoxy-alpha-L-mannopyranosy l-(1-->4)]- beta-D-glucopyranoside. The structural identification was performed using detailed analysis of 1H and 13C NMR spectra including 2D NMR spectroscopic techniques (COSY, HETCOR and COLOC) and chemical conversions.

A new steroidal saponin from the rhizomes of Costus spicatus.

A new steroidal saponin has been isolated from the rhizomes of Costus spicatus and its structure was elucidated as (3 beta, 22 alpha, 25R) -26-(beta-D-glucopyranosyloxy)-2-methoxyfurost-5-en-3-yl O-D-apio-beta-D-furanosyl-(1-->4)-O-[alpha-L-rhamnopyranosyl-(1--> 2)]- beta-D-glucopyranoside by means of IR, MS, NMR and chemical evidence.

[Utilization of the waste of sisal industry in the control of mosquito larvae]. / O aproveitamento do resíduo da indústria do sisal no controle de larvas de mosquitos.

The aim of this research was to utilize the waste residues of sisal fiber separation from Agave sisalana leaves to develop a larvicide for the combat of mosquito transmitting tropical diseases. Larvae of Aedes aegypti and Culex quinquefasciatus were exposed to different concentrations of the Agave extract for 24 hours to determine lethal concentrations. The LC50 for A. aegypti was 322 ppm and the LC50 for C. quinquefasciatus was 183 ppm. To detect the active substances, saponins were investigated. It was found that the various components of the extract were effective in eliminating the larvae. Under field conditions, this formulation can probably be used at 100 ppm, which causes 100% mortality of C. quinquefasciatus larvae after 3-4 days. The product is not recommended for use against A. aegypti due to the necessity for high concentrations and to the fact that the larvae of this species live frequently on drinking water. To avoid fermentation, Agave extract should be used in a dehydrated form which also represent a good formulation for practical use.

Haemolytic activities of plant saponins and adjuvants. Effect of Periandra mediterranea saponin on the humoral response to the FML antigen of Leishmania donovani.

An 87.7% (P < 0.01) and 84% (P < 0.001) of protection against visceral leishmaniasis was achieved in CB hamsters and Balb/c mice, respectively, with saponin combined to the fucose-mannose ligand of Leishmania donovani (FML). However, an undesirable haemolytic effect was described for several saponins. Aiming to improve the formulation with FML/saponin, we comparatively analysed the haemolytic potential of recently characterized plant saponins and currently used adjuvants. The haemolytic activity of steroidic saponins from Agave sisalana; Smilax officinalis as well as commercial saponin (Riedel De Haën's), was higher than that of triterpenoid ones (Bredemeyera floribunda; Periandra mediterranea) and the Freund's complete adjuvant. The concentration resulting in 50% haemolysis was 500 micrograms ml-1 for aluminum hydroxide. The low haemolytic effect of P. mediterranea saponin was abolished by removal of its glycidic moiety and its sapogenin fraction as well as the Freund's Incomplete Adjuvant were non-haemolytic within this range. Furthermore, the adjuvant effect of three doses of P. mediterranea saponin injected with the FML antigen of L. donovani, was assayed in mice, either by the intraperitoneal (i.p.) or the subcutaneous (s.c.) route. The anti-FML IgG antibody levels increased and detectable levels were observed up to 3 months in the s.c. group. The response was expanded in both groups after an injection with a fourth vaccine dose. The IgG response showed increased levels of IgG2a only in the i.p. group, while IgG2b and IgG1 but not IgG3 antibodies were higher than controls in both groups. In conclusion, the results suggest that the recently described triterpenoid fractions of P. mediterranea can be safely used as adjuvant with low or non-haemolytic effect.

A new triterpenoid saponin, bredemeyeroside B. from the roots of Bredemeyera floribunda.

A new triterpenoid saponin named bredemeyeroside B has been isolated from the roots of Bredemeyera floribunda and its structure elucidated as 3-O-beta-D-glucopyranosyl-2 beta, 3 beta, 27-trihydroxyolean-12-en-23,28-dioic acid 28-O-beta-D-xylopyranosyl-(1-->4)-[beta-D-apiofuranosyl-(1-->3)]-a lpha -L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1-->3] -beta-D-fucopyranoside by means of MS, NMR and chemical evidence. In addition rutin was also isolated. In laboratory tests bredemeyeroside B showed snake venom antidote activity.

Steroidal saponins from Smilax officinalis.

Three new steroidal saponins were isolated from the rhizomes of Smilax officinalis. The structures of these saponins were established by extensive spectral data, hydrolysis and chemical correlation as sarsasapogenin 3-O-beta-D-glucopyranosyl-(1-->4)-[alpha-L-arabinopyranosyl-(1-->6 )-beta- D-glucopyranoside, neotigogenin 3-O-beta-D-glucopyranosyl-(1-->4)-[alpha-L-arabinopyranosyl-(1-->6 )]-beta- D-glucopyranoside and 25S-spirostan-6 beta-ol 3-O-beta-D-glucopyranosyl-(1-->4)-[alpha-L-arabinopyranosyl-(1-->6 )]-beta- D-glucopyranoside. Acid hydrolysis of the latter compound gave a sapogenin which has a new orientation of an hydroxyl on the steroidal skeleton. A route is proposed for the biogenesis of the latter sapogenin which is an uncommon steroidal aglycone.

Alkamides from Cissampelos glaberrima.

Two alkamides, deca-2E,4E-dienoic acid isobutylamide (1) and octa-2E,4E-dienoic acid isobutylamide (2), were isolated from the roots of Cissampelos glaberrima (Menispermaceae). Their structures were established by spectral and physical methods. Traces of two new alkamides, decen-2-oic acid isobutylamide (3) and decanoic acid isobutylamide (4), were identified by mass spectrometry.

Pharmacological screening of plants recommended by folk medicine as snake venom antidotes; IV. Protection against jararaca venom by isolated constituents.

Fifteen compounds, isolated from plants reputed as snake venom antidotes, belonging to different classes of natural products, were shown to protect mice to a significant degree against the lethal action of the venom of Bothrops jararaca snakes. Administration was by the oral route, one hour prior to envenomation. The substances are nitrogen-free, low-molecular-weight compounds for which some kind of biodynamic activity has previously been reported. The fact that they are mostly trivial, naturally-occurring compounds should explain why plants used as snake-bite antidotes are so widely distributed over the plant kingdom.

Pharmacological Screening of Plants Recommended by Folk Medicine as Snake Venom Antidotes; IV. Protection against Jararaca Venom by Isolated Constituents1.

Fifteen Compounds, isolated from plants reputed as snake venom antidotes, belonging to different classes of natural products, were shown to protect mice to a significant degree against the lethal action of the venom of BOTHROPS JARARACA snakes. Administration was by the oral route, one hour prior to envenomation. The substances are nitrogen-free, low-molecular-weight compounds for which some kind of biodynamic activity has previously been reported. The fact that they are mostly trivial, naturally-occurring compounds should explain why plants used as snake-bite antidotes are so widely distributed over the plant kingdom.

Neutralization of lethal and myotoxic activities of South American rattlesnake venom by extracts and constituents of the plant Eclipta prostrata (Asteraceae).

Ethanolic extracts of the aerial parts of Eclipta prostrata L. (Asteraceae) neutralized the lethal activity of the venom of South American rattlesnake (Crotalus durissus terrificus) when mixed in vitro before i.p. injection into adult Swiss mice. Samples of ethanolic extract corresponding to 1.8 mg of dry extract per animal neutralized up to four lethal doses of the venom (LD50 = 0.08 micrograms venom/g animal). Three substances isolated from the plant--wedelolactone (0.54 mg/animal), sitosterol (2.3 mg/animal) and stigmasterol (2.3 mg/animal)--were able to neutralize three lethal doses of the venom. Aqueous extracts of the plant inhibited the release of creatine kinase from isolated rat muscle exposed to the crude venom. The protection conferred against the myotoxic effects of the venom could be demonstrated also in vivo, when the venom was preincubated with the extract prior to injection into mice.