Evaluation of the toxicological effects of Neltuma alpataco (Prosopis alpataco) pod alkaloid extract.

The pods of Neltuma spp. have shown potential as a source of protein and energy in livestock. However, prolonged consumption of some of these species can lead to neurological symptoms in ruminants. This study aimed to determine the alkaloid content, as well as the in vitro and in vivo effects of an alkaloid-enriched extract (AEE) from N. alpataco pods. High performance liquid chromatography-high resolution mass spectrometry (HPLC-HRMS) identified juliprosine and juliprosopine as primary alkaloids, with juliprosine being most abundant. AEE from N. alpataco demonstrated dose-dependent cytotoxicity on glioma cells after 48 h, with a 50% cytotoxic concentration (CC50) of 24.69 µg/mL. However, the release of LDH was observed only at the highest tested concentration, indicating cellular damage. Further examination through phase-contrast microscopy and dual acridine orange/ethidium bromide fluorescence staining revealed morphological changes consistent with an apoptotic mechanism of cell death, ultimately leading to secondary necrosis. Finally, the LD50 after intraperitoneal injection in mice was determined to be 12.98 mg/kg. Taken together, these findings demonstrated for the first time the in vivo and in vitro toxicity of the AEE from N. alpataco pods.

Ipomoea carnea alkaloid extract vs swainsonine: A comparative study on cytotoxic activity against glial cells.

The consumption of Ipomoea carnea produces a neurological syndrome in animals. The toxic principles of I. carnea are the alkaloids swainsonine (SW) and calystegines B1, B2, B3 and C1. In this study, we investigated the cytotoxicity of an alkaloid extract of Ipomoea carnea (AEE) and natural swainsonine (SW) isolated from Astragalus lentiginosus (25-1000 µM of SW) for 48 h in a glioma cell line. Although the natural SW did not induce any changes in cell viability, the AEE exhibited a dose dependent cytotoxic effect and release of lactate dehydrogenase (LDH) indicative of cytolysis. In order to evaluate the morphological changes involved, cells were examined using phase contrast and fluorescence microscopy with acridine orange-ethidium bromide staining. The AEE caused a cell death compatible with necrosis, whereas exposure to 1000 µM of SW resulted in cytoplasmic vacuolation. Immunocytochemical studies revealed that astrocytes treated with 150 µM of AEE from I. carnea or 1000 µM of SW exhibited morphological characteristics of cell activation. These findings suggest that swainsonine would not be the only component present in the AEE of I. carnea responsible for in vitro cytotoxicity. Calystegines might also play a role in acting synergistically and triggering cell death through necrosis.

Cytotoxic activity induced by the alkaloid extract from Ipomoea carnea on primary murine mixed glial cultures.

The prolonged consumption of Ipomoea carnea produces neurologic symptoms in animals and a typical histological lesion, cytoplasmic vacuolization, especially in neurons. The toxic principles of I. carnea are the alkaloids swainsonine and calystegines B1, B2, B3 and C1. In this study, primary brain cultures from newborn mouse containing mixed glial cells were utilized. These cells were exposed to Ipomoea extracts containing between 0 and 250 µM swainsonine for 48 h. Morphological changes were investigated through Phase Contrast microscopy and Rosenfeld's staining. The extract induced cytoplasmic vacuolization in astrocytes and microglia in a dose dependent manner, being more evident when cultures were exposed to 250 µM of swainsonine. In addition, acridine orange staining evidenced an increase in the number of lysosomes in both microglia and astrocytes cells. Consistent with this, scanning electron microscopy also showed that both types of cells presented morphological characteristics of cell activation. Ultrastructurally, cells showed vacuoles filled with amorphous material and surrounded by a single membrane and also multilayer membranes. Taken together, these findings suggest that swainsonine along with calystegines, are probably responsible for the activation of glial cells due to a possible lysosomal dysfunction and therefore intracellular storage. Our results demonstrate that this in vitro glial cell model is a very good alternative to in vivo studies that require several weeks of animal intoxication to observe similar neurotoxic effects.