Correction: Inhibitory Effects of Hydroethanolic Leaf Extracts of Kalanchoe brasiliensis and Kalanchoe pinnata (Crassulaceae) against Local Effects Induced by Bothrops jararaca Snake Venom.

[This corrects the article DOI: 10.1371/journal.pone.0168658.]

Inhibitory Effects of Hydroethanolic Leaf Extracts of <i>Kalanchoe brasiliensis</i> and <i>Kalanchoe pinnata</i> (Crassulaceae) against Local Effects Induced by <i>Bothrops jararaca</i> Snake Venom

<div><p>The species <i>Kalanchoe brasiliensis</i> and <i>Kalanchoe pinnata</i>, both known popularly as “Saião,” are used interchangeably in traditional medicine for their antiophidic properties. Studies evaluating the anti-venom activity of these species are scarce. This study aims to characterize the chemical constituents and evaluate the inhibitory effects of hydroethanolic leaf extracts of <i>K</i>. <i>brasiliensis</i> and <i>K</i>. <i>pinnata</i> against local effects induced by <i>Bothrops jararaca</i> snake venom. Thin Layer Chromatography (TLC) and High Performance Liquid Chromatography coupled with Diode Array Detection and Electrospray Mass Spectrometry (HPLC-DAD-MS/MS) were performed for characterization of chemical markers of the extracts from these species. For antiophidic activity evaluation, <i>B</i>. <i>jararaca</i> venom-induced paw edema and skin hemorrhage in mice were evaluated. In both models, hydroethanolic extracts (125–500 mg/kg) were administered intraperitoneally in different protocols. Inhibition of phospholipase enzymatic activity of <i>B</i>. <i>jararaca</i> was evaluated. The HPLC-DAD-MS/MS chromatographic profile of extracts showed some particularities in the chemical profile of the two species. <i>K</i>. <i>brasileinsis</i> exhibited major peaks that have UV spectra similar to flavonoid glycosides derived from patuletin and eupafolin, while <i>K</i>. <i>pinnata</i> showed UV spectra similar to flavonoids glycosides derived from quercetin and kaempferol. Both extracts significantly reduced the hemorrhagic activity of <i>B</i>. <i>jararaca</i> venom in pre-treatment protocol, reaching about 40% of inhibition, while only <i>K</i>. <i>pinnata</i> was active in post-treatment protocol (about 30% of inhibition). In the antiedematogenic activity, only <i>K</i>. <i>pinnata</i> was active, inhibiting about 66% and 30% in pre and post-treatment protocols, respectively. Both extracts inhibited phospholipase activity; however, <i>K</i>. <i>pinnata</i> was more active. In conclusion, the results indicate the potential antiophidic activity of <i>Kalanchoe</i> species against local effects induced by <i>B</i>. <i>jararaca</i> snake venom, suggesting their potential use as a new source of bioactive molecules against bothropic venom.</p></div

Inhibition of the edematogenic activity of <i>B</i>. <i>jararaca</i> venom (BjV) by extracts of <i>K</i>. <i>brasiliensis</i> (Kb) (A) and <i>K</i>. <i>pinnata</i> (Kp) (B) in post-treatment protocol.

<p>BjV was injected i.pl. in the right hind paw of mice 5 min before treatment with extract (500 mg/kg, i.p.). Paw thickness was measured during 120 min after venom injection. Edema was expressed as the increase in paw thickness calculated, by the percentage difference between the paw thickness after (at respective time) and before (basal values) venom injection. The points represent the mean ± SEM (n = 5). *p<0.05 and **p<0.01 compared to the group receiving venom alone along with the i.p. injection of 5% castor oil in PBS (Two-way ANOVA followed by the Bonferroni test).</p

HPLC-DAD-MS chromatograms of <i>K</i>. <i>brasiliensis</i> (A) and <i>K</i>. <i>pinnata</i> (B) HE extracts.

<p>The extracts were chromatographed on a Phenomenex RP-18 column (250 x 4.6 mm, 5 um) and the column was eluted with a gradient of acetonitrile (ACN) in 3% acetic acid at a flow rate of 0.7 mL/min. The elution profile was monitored at 340 nm. Peaks 6–8, 11–17, 22 and 23 were considered to be major peaks in panel A, and peaks 4 and 11–13 were considered to be major peaks in panel B.</p

Effect of extracts of <i>K</i>. <i>brasiliensis</i> (Kb) (A) and <i>K</i>. <i>pinnata</i> (Kp) (B) on MPO levels in paw edema induced by <i>B</i>. <i>jararaca</i> venom (BjV) in pre-treatment protocol.

<p>LP: left paw (basal control; no injection of venom, extract or dexamethasone). UMPO: a unit of MPO, defined as the equivalent to the consumption of 1μmol of hydrogen peroxide per minute. The columns represent the mean ± SEM (n = 5), **p <0.01 and ***p<0.001 compared to the control group (i.pl. injection of BjV with i.p. administration of 5% castor oil in PBS) after Tukey’s test (one-way ANOVA followed by the Tukey’s test).</p

Inhibition of the enzymatic and biological activities of <i>B</i>. <i>jararaca</i> venom by hydroethanolic extracts of <i>K</i>. <i>brasiliensis</i> and <i>K</i>. <i>pinnata</i>.

<p>Inhibition of the enzymatic and biological activities of <i>B</i>. <i>jararaca</i> venom by hydroethanolic extracts of <i>K</i>. <i>brasiliensis</i> and <i>K</i>. <i>pinnata</i>.</p

Inhibition of <i>B</i>. <i>jararaca</i> (BjV) venom-induced hemoglobin accumulation by extracts of <i>K</i>. <i>brasiliensis</i> (Kb) (A) and <i>K</i>. <i>pinnata</i> (Kp) (B) in post-treatment protocol.

<p>The percentage of activity presented was calculated as: [(Hemoglobin content in animals receiving venom plus extract ÷ Hemoglobin content in animals receiving venom alone) x 100]. Values expressed as mean ± SEM (n = 5). *p<0.05 and ***p<0.001 compared to venom alone (BjV) (100% of activity) (one-way ANOVA followed by the Tukey’s test).</p

Inhibition of the hemorrhagic activity of <i>B</i>. <i>jararaca</i> (BjV) venom by extracts of <i>K</i>. <i>brasiliensis</i> (Kb) (A) and <i>K</i>. <i>pinnata</i> (Kp) (B) in pre-treatment protocol.

<p>BjV was injected s.c. in the ventral region of mice pre-treated i.p. with different doses of extracts. Three hours after venom injection, the skin was removed and weighed. The columns represent the mean ± SEM (n = 5). *<0.05 and **p<0.01 compared to venom alone (one-way ANOVA followed by the Tukey’s test).</p

Inhibition of PLA₂ activity of <i>B</i>. <i>jararaca</i> venom by a hydroethanolic extract from <i>K</i>. <i>brasiliensis</i> (Kb) (A) and <i>K</i>. <i>pinnata</i> (Kp) (B).

<p>The points are the mean ± SEM (n = 3). Note that the error bars are smaller than the symbols.</p