Neutral and ionic forms of EPI (A).

<p>Potentiometric pKa determination (B). Illustration of the pKa values of EPI predicted using the ChemAxon method (C). This graph shows the variation in the of distribution species vs pH values, and indicates that the ChemAxon predicted pKa values are respectively pKa1 = 6.40 and pKa2 = 13.90 (C). DSC (D) and DSC-TGA (E) data illustrating the thermal degradation that occurs around 220°C in a single step.</p

Effects of EPI against adult <i>S</i>. <i>mansoni</i> BH strain (45 days post infection).

<p>Values are presented as mean± SEM. Number of animals/group = 10. Group treated with 100 mg /kg 3 animals died.</p><p>Asterisks indicate that difference compared to the control was significant with *** P<0.001, **P<0.01 and *P<0.05.</p><p>Effects of EPI against adult <i>S</i>. <i>mansoni</i> BH strain (45 days post infection).</p

Effects of EPI against juvenile <i>S</i>. <i>mansoni</i> BH strain (treated 21 days post infection).

<p>Values are presented as mean ± SEM. Number of animals/group = 8.</p><p>Asterisks indicate that difference compared to the control was significant with ***P<0.001, **P<0.01 and *P<0.05.</p><p>Effects of EPI against juvenile <i>S</i>. <i>mansoni</i> BH strain (treated 21 days post infection).</p

Histopathological study of organs sections of different groups of mice.

<p>Comparison between the histology of: liver (A,B,C), spleen (D,E,F), lung (G,H,I), kidney (J,K,L), and brain (M,N,O) obtained from Swiss mice (n = 4) treated with 0.0 (first column), 530 (second column) or 8000 (third column) mg/kg of EPI. The first and the second column refer to mice killed 7 days after assay and the third one represent organs from animals that died due to the administration of higher concentration of the drug. Light microscopy evaluation of these organs showed severe morphological changes in the spleen, lung, kidney and brain, but not in the parenchyma of liver in the animals treated with 8000 mg/kg of EPI. Focal alterations in the red pulp were observed in the spleen of animals treated with 530 mg/kg of the drug. Control group presented no morphological changes in the organs.</p

Photomicrographs of hepatic granulomas of mice liver 45 days post infection.

<p>A and B: Liver of treated infected animals (40 mg/kg). Note the necrotic-exudative phase with loose inflammatory infiltrate (macrophages, neutrophils and eosinophils) and areas with necrotic cells around the egg. C and D: Liver of untreated infected animals, the granulomas were found in the periportal area. Different stages of evolution of the granuloma, including granulomatous pre-exudative, necrotic-exudative, productive and healing by fibrosis were observed. Arrows indicate the presence of inflammatory cells.</p

EPI bond distances obtained through x-ray diffraction (Experimental) and DFT results (Calculated).

<p>Atom labels accordingly to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0066702#pone-0066702-g004" target="_blank">Figure 4</a>.</p

Effect of treatment with EPI on oviposition by adults, 15 days post-treatment (45 days post infection).

<p>Number of animals/group = 10. Group treated with 100 mg /kg 3 animals died.</p><p>Asterisks indicate that difference compared to the control was significant with *** P<0.001</p><p>Effect of treatment with EPI on oviposition by adults, 15 days post-treatment (45 days post infection).</p

Effect of treatment with EPI on relative organ weight.

<p>Asterisks indicate that difference compared to the control was significant with **P<0.01 and *P<0.05.</p><p>Effect of treatment with EPI on relative organ weight.</p

Analytical HPLC used LiChrospher 60 RP column and eluted with potassium phosphate

<p>. (A) Standard EPI (20 µg/mL), (B) Standard pilocarpine (50 µg/mL), (C) “cultivated jaborandi leaves” solution, resulted from first extraction step, (D) “cultivated jaborandi acid” solution, obtained EPI under salt form, (E) Solution of “crude EPI” with some impurities as pilocarpine and other alkaloids, (F) last step of isolation showing EPI >98% purity.</p

Granuloma diameter (μm) in the liver of the mice from treated and untreated adult worm-infected groups 45 day post infection.

<p>Difference was significant ** <i>P</i><0.01.</p><p>Granuloma diameter (μm) in the liver of the mice from treated and untreated adult worm-infected groups 45 day post infection.</p