Supercritical Fluid Extraction of Pyrrolidine Alkaloid from Leaves of Piper amalago L.

Supercritical fluid extraction was used to extract the alkaloid N-[7-(3',4'-methylenedioxyphenyl)-2(Z),4(Z)-heptadienoyl]pyrrolidine from leaves of Piper amalago L. A three-level orthogonal array design matrix, OAD OA9(34), was used for optimization of the parameters of supercritical extraction of the alkaloid, employing supercritical carbon dioxide: extraction time (20, 40, and 60 min), temperature (40, 50, and 60°C), pressure (150, 200, and 250 bar), and the use of cosolvents (ethanol, methanol, and propyleneglycol). All parameters had significant effect on the alkaloid yield. The alkaloid yield after 60 min of extraction without cosolvents at 9 different conditions (32) in terms of temperature (40, 50, and 60°C) and pressure (150, 200, and 250 bar) was also evaluated. The optimal yield (≈3.8 mg g-1) was obtained with supercritical CO2 + methanol (5% v : v) at 40°C and 200 bar for 60 min of extraction.

Antileishmanial activity of isolated triterpenoids from Pourouma guianensis.

The inhibiting activity of triterpenoids isolated from the methanolic extract of Pourouma guianensis (Moraceae) leaves is described for promastigotes and intracellular amastigotes of Leishmania amazonensis. Whereas the fractions containing apigenin, friedelin, epi-friedelinol, arjunolic acid, hyptatic acid B, stigmasterol and sitosterol were of no or relatively low inhibitory activity, fractions containing tormentic acid, 2alpha,3beta-dihydroxyursan-12-en-28-oic acid, 2alpha,3beta-dihydroxyolean-12-en-28-oic acid, oleanolic acid and ursolic acid were very potent in inhibiting promastigote growth at 100 microg/ml. Of the eleven isolated compounds, however, only ursolic acid and oleanolic acid showed high activity against intracellular amastigotes (IC50 value = 27 microg/ml and 11 microg/ml, respectively), which was superior to the control drug Glucantime (IC50 value = 83 microg/ml). The antileishmanial activity of oleanolic acid was directed against the parasite and not due to activation of nitric oxide intermediates by macrophages, but this triterpenoid also significantly inhibited the phagocytic capacity of those cells at concentrations above 40 microg/ml, indicating a cytotoxic effect. These results indicate that Pourouma guianensis contains many triterpenoids and some, such as ursolic and oleanolic acids, may serve as lead compounds for new antileishmanial drugs, but chemical modifications may be necessary to avoid unselective cytotoxicity.

Toxicological analysis and effectiveness of oral Kalanchoe pinnata on a human case of cutaneous leishmaniasis.

Leishmaniasis is an extremely difficult disease to treat. Previously, it was shown that oral Kalanchoe pinnata (Kp) leaf extract is strongly effective against murine leishmaniasis. Here, it is shown that the serum levels of alanine-aminotransferase (ALT), aspartate-aminotransferase (AST), urea and alkaline phosphatase were unchanged in mice orally treated with supraoptimal Kp doses for 30 days, indicating the absence of chronic toxicity to the liver, heart or kidney. Additionally, evidence is presented that human leishmaniasis may also be controlled with oral Kp. A 36-year-old man with an active cutaneous leishmaniasis was orally treated with 30 g wet weight of Kp leaves/day for 14 days. During the Kp treatment, the lesion stopped growing and slightly decreased. No adverse reactions or toxicity was observed. This study reports for the first time that Kalanchoe pinnata contains substances potentially active and safe for the oral treatment of human cutaneous leishmaniasis.

Selective effect of 2',6'-dihydroxy-4'-methoxychalcone isolated from Piper aduncum on Leishmania amazonensis.

2',6'-Dihydroxy-4'-methoxychalcone (DMC) was purified from the dichloromethane extract of Piper aduncum inflorescences. DMC showed significant activity in vitro against promastigotes and intracellular amastigotes of Leishmania amazonensis, with 50% effective doses of 0.5 and 24 micrograms/ml, respectively. Its inhibitory effect on amastigotes is apparently a direct effect on the parasites and is not due to activation of the nitrogen oxidative metabolism of macrophages, since the production of nitric oxide by both unstimulated and recombinant gamma interferon-stimulated macrophages was decreased rather than increased with DMC. The phagocytic activity of macrophages was functioning normally even with DMC concentrations as high as 80 micrograms/ml, as seen by electron microscopy and by the uptake of fluorescein isothiocyanate-labeled beads. Ultrastructural studies also showed that in the presence of DMC the mitochondria of promastigotes were enlarged and disorganized. Despite destruction of intracellular amastigotes, no disarrangement of macrophage organelles were observed, even at 80 micrograms of DMC/ml. These observations suggest that DMC is selectively toxic to the parasites. Its simple structure may well enable it to serve as a new lead compound for the synthesis of novel antileishmanial drugs.