This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. This article has been cited by other articles in PMC. Photography by Jacyra A. Figure S3: co-TLC analysis, it was possible to observe the presence of the flavonoids apigenin Rf 0,64, fluorescent green color , luteolin Rf 0,64, fluorescent yellow color , orientin Rf 0,64, fluorescent yellow color , isoorientin Rf 0,64, fluorescent yellow color , and vitexin Rf 0,64, fluorescent green color in the extract.

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Published26 Oct Abstract Snakebites are a serious worldwide public health problem. The specific treatment consists of antivenom serum therapy, which has some limitations such as inability to neutralize local effects, difficult access in some regions, risk of immunological reactions, and high cost. Thus, the search for alternative therapies to treat snakebites is relevant. Jatropha mollissima Euphorbiaceae is a medicinal plant popularly used in folk medicine as an antiophidic remedy.

Therefore, this study aims to evaluate the effect of the aqueous leaf extract from J. High Performance Liquid Chromatography with Diode Array Detection analysis and Mass Spectrometry analysis of aqueous leaf extract confirmed the presence of the flavonoids isoschaftoside, schaftoside, isoorientin, orientin, vitexin, and isovitexin. Local skin hemorrhage, local edema, leukocyte migration, and myotoxicity were significantly inhibited by the extract.

These results demonstrate that J. Introduction Snakebites represent a serious worldwide public health and social problem because of their high frequency, morbi-mortality, and sequelae left in the victims. Moreover, accidents caused by snakes are considered a neglected disease mainly in Africa, Latin America, Asia, and Oceania [ 1 , 2 ]. Data indicate that, worldwide, more than 5 million people suffer snakebites every year, resulting in 25, to , deaths and leaving approximately , people with permanent disabilities [ 1 ].

In Brazil, an estimated number of 25, snakebites occur per year [ 3 ]. The Bothrops snakes have high complexity and variation in the protein composition of their venom. This variation is due to factors such as diet, age, seasonal variation, sexual dimorphism, and geographical origin, which occurs within the species, interfamily, intergenus, interspecies, intersubspecies, and intraspecies [ 5 , 6 ].

The pathophysiological process of the snake envenoming is complex and includes the combined action of several toxins such as snake venom metalloproteinases SVMPs , snake venom serine proteinases SVSPs , phospholipases A2 PLA2 , hyaluronidases, bradykinin-releasing enzymes, lectins, L-amino oxidases, and pharmacological mediators [ 7 , 8 ]. Bothrops envenomation causes mainly immediate local tissue damage including pain, edema, local hemorrhage, and myonecrosis and systemic effects cardiovascular alterations, coagulation, and renal alterations [ 9 , 10 ].

Currently, the intravenously antivenom serum therapy is the only specific treatment for snakebites. The antivenom, however, has some limitations such as difficult access in some regions, risk of immunological reactions including serum sickness , high cost, and limited effectiveness in the improvement of the local tissue damage [ 11 , 12 ].

Administration of the antivenom may prevent death but does not prevent local tissue damage and resultant disabilities. The low inhibition of local effects due to the delay of receiving the serum or due to the low efficacy is the leading cause of amputations, which can lead to serious social, economic, and health negative impacts, given that most victims live in rural areas [ 13 ]. So, the search for new complementary therapies to treat snakebites has become of utmost importance.

In this scenario, the use of many medicinal plants has been an old practice in folk medicine against snakebites, especially in tropical and subtropical regions such as Africa, Asia, and South America [ 14 , 15 ]. The Jatropha L. In Africa, Asia, and Latin America, the Jatropha species are used not only in traditional medicine to cure various ailments, but also as ornamental plants and energy crops [ 17 ].

Several known species from genus Jatropha have been reported for their medicinal uses, chemical constituents, and biological activities such as Jatropha curcas, Jatropha elliptica, Jatropha gossypifolia, and Jatropha mollissima [ 17 , 18 ]. This plant is endemic in the semiarid region of Northeastern Brazil. In folk medicine, it is widely used for various purposes, especially as antiophidic [ 19 — 21 ] and anti-inflammatory [ 22 ] remedies, healing [ 20 ], veterinary vermifuge [ 19 ], and treatment of renal inflammation and loss of appetite [ 20 ].

Some studies have shown that this species has antioxidant [ 23 ], antimicrobial [ 24 ], and antihelmintic activities [ 25 ]. Thus, the aim of this work was to evaluate the ability of the aqueous leaf extract of J. Our results indicate that J. Materials and Methods 2. All other reagents and solvents used were of analytical grade. The water used was purified by reverse osmosis.

The botanical identification of the material was performed by Dr. After identification and confirmation of the plant species, the leaves were dried at room temperature, triturated with an industrial blender, and stored in hermetically sealed bottles until used for aqueous extract preparation.

Snake Venom Lyophilized B. The venom was weighed and dissolved with phosphate buffer saline PBS and the protein content quantified by the Bradford method [ 26 ]. Animals Male and female Swiss albino mice 30—35 g , 6—8 weeks of age, used in this study were maintained under standard environmental conditions with free access to water and food.

On the day of the experiment, the animals were placed in the experimental room for at least one hour prior to tests, for acclimation. The total number of animals used was Preparation of the Aqueous Extract from the Leaves of J.

The aqueous extract obtained after vacuum filtration was freeze-dried and dissolved in PBS at adequate concentrations for the biological assays.

The chromatograms were analyzed under nm UV light and then sprayed with specific chromogenic agents according to the class of compounds investigated Dragendorff reagent, natural reagent A, ferric chloride, and sulfuric vanillin and heating.

The retention factor, behavior, and color of the spots were compared with the chromatographic profiles of the reference substances. Flow elution was 0. The aqueous leaf extract of J. The lyophilized aqueous extract of J. The final concentration of the extract was 2. Their retention time and ultraviolet spectra were obtained for chromatogram peaks at and nm, with the acquisition of UV spectra in the range of to nm.

The extract was prepared in triplicate and analyzed. To confirm the presence of the standards in the aqueous extract of J. After total dissolution and prior to analysis, the samples and solvents were filtered through a 0.

The mobile phase was comprised of acetic acid 0. The mass spectrometer source parameters were set as follows: capillary voltage at 3.

The electrospray ionization ESI source was operated in the positive and negative ionization mode. The data were acquired using amplitudes of 0. The data were processed through Bruker Compass Data Analysis 4. After 30 min, the animals received a subcutaneous s. After photo documentation of the produced hemorrhagic halos, the hemorrhagic skin was removed and weighed.

The group in which animals received s. Another group that received s. Inhibition of the Edematogenic Activity The edematogenic activity of B. Groups of 5 animals were treated with different doses of J. After 30 min, the animals received an intraplantar i. A group of animals that received i. Another group that received i. After 30 min, the animals received by i. After 6 hours B. Inhibition of the Myotoxic Activity The myotoxic activity of B.

After 30 min, all the animals received an intramuscular i. A group in which animals received i. Results 3. Phytochemical Analysis of the Aqueous Extract of J. Thin Layer Chromatography TLC Profile For phytochemical analysis by TLC, the aqueous extract was fractionated by liquid-liquid partition to obtain fractions with different polarities, thus facilitating the chromatographic analysis of the compounds.

Moreover, the TLC analysis was performed with different developers in order to identify the classes of compounds presented in the aqueous extract of J. The chromatograms show spots suggestive of the presence of phenolic compounds, flavonoids, and saponins Figure S2. By co-TLC analysis, it was possible to observe the presence of the flavonoids apigenin Rf 0. For confirmation of presence of such substances, analyses were performed by HPLC.

It is possible to observe that J. Among them, most have UV spectra similar to glycosylated flavonoid derivatives from apigenin nm II band and nm I band and luteolin and nm II band and nm I band [ 30 ]. Glycosylated derivatives from these two flavonoids, orientin and isoorientin derived from luteolin and vitexin and isovitexin derived from apigenin , have similar absorption II bands for these aglycones, differing mainly by the maximum absorption of the I band.

The peaks 3, 4, 5, and 6 had their identity confirmed as, respectively, isoorientin tR Peaks 1 tR Four compounds were identified as isoorientin peak 3 , orientin peak 4 , vitexin peak 5 , and isovitexin peak 6. Applying the systematic analyses carried out by Ferreres and coworkers [ 31 , 32 ], it was possible to identify the compounds as schaftoside and isoschaftoside flavonoid, respectively. According to such report, preferential fragmentation is of the sugar moiety at the 6-C rather than the 8-C position.

Considering the preferential fragmentation at the C-6 position, chromatographic signal 1 corresponded to the isoschaftoside flavonoid and chromatographic signal 2 corresponded to the schaftoside flavonoid.

Furthermore, peaks 3, 4, 5, and 6 were confirmed as isoorientin, orientin, vitexin, and isovitexin as suggested by the coinjection performed with these standards see Section 3. All the signals present in Table 1 are in full agreement with the previous published data.










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