Wound tissue remodeling by latex exudate of Himatanthus drasticus: A plant species used in Brazilian folk medicine.

This work investigated the healing properties of proteins extracted of latex (HdLP) on excisional wounds. Cell toxicity of HdLP was investigated carried out in murine fibroblasts after incubation with HdLP (12.5-100 µg/ml). The dermal irritability test was performed to evaluate dermal reactions. The wounds were performed and treated with vehicle or HdLP (0.5 %, 1.0 %, and 2.0 %). The macroscopic parameters, histological analysis and measurement of inflammatory markers and mediators were evaluated. HdLP did not exhibit cytotoxicity and did not induce skin irritation. HdLP stimulated the release of IL-1ß at the beginning of the inflammatory phase. This effect probably favored the earlier release of IL-10 by macrophages, during the proliferative phase. The shortening and completeness of healing were characterized by fibroblast proliferation and the presence of newly synthesized collagen fibers. This was accompanied by well-organized re-epithelialization. The involvement of latex proteins in this activity is reported for the first time.

Successful Pre-Clinical Management of Irinotecan-Debilitated Animals: A Protein- Based Accessory Phytomedicine.

BACKGROUND: Calotropis procera is a laticiferous plant (Apocynaceae) found in tropical regions all over the world. The ultrastructural characteristics of laticifers, their restricted distribution among different taxonomic groups, and in some species in each clade, as peptidases from latex, make them very attractive for biological analysis. OBJECTIVE: The study aims to investigate the effects of LP-PII-IAA (laticifer protein (LP) sub-fraction II (PII) of C. procera presenting an iodoacetamide-inhibited cysteine proteinase activity) on irinotecan-induced intestinal mucositis, a serious adverse effect of this medicine for the treatment of cancer. METHODS: LP-PII-IAA is composed of closely related isoforms (90%) of peptidases derived from catalysis and an osmotin protein (5%). Animals receiving co-administration of LP-PII-IAA presented a significant decrease in mortality, absence of diarrhea, histological preservation, and normalization of intestinal functions. RESULTS: Clinical homeostasis was accompanied by a reduction in MPO activity and declined levels of IL-1ß, IL-6 and KC, while the IL-10 level increased in LP-PII-IAA-treated animals. COX-2 and NF-kB immunostaining was reduced and the levels of oxidative markers (GSH, MDA) were normalized in animals that received LP-PII-IAA. CONCLUSION: We suggest that peptidases from the latex of Calotropis procera were instrumental in the suppression of the adverse clinical and physiological effects of irinotecan.

Anti-inflammatory latex proteins of the medicinal plant Calotropis procera: a promising alternative for oral mucositis treatment.

OBJECTIVE AND DESIGN: Oral mucositis (OM) is an intense inflammatory reaction progressing to tissue damage and ulceration. The medicinal uses of Calotropis procera are supported by anti-inflammatory capacity. PII-IAA, a highly homogenous cocktail of laticifer proteins (LP) prepared from the latex of C. procera, with recognized pharmacological properties was tested to treat OM. MATERIALS AND SUBJECTS: Male Golden Sirius hamsters were used in all treatments. TREATMENT: The latex protein samples were injected i.p. (5 mg/Kg) 24 h before mucositis induction (mechanical trauma) and 24 h later. METHODS: Histology, cytokine measurements [ELISA], and macroscopic evaluation [scores] were performed. RESULTS: PII-IAA eliminated OM, accompanied by total disappearance of myeloperoxidase activity and release of IL-1b, as well as reduced TNF-a. Oxidative stress was relieved by PII-IAA treatment, as revealed by MDA and GSH measurements. PII-IAA also reduced the expression of adhesion molecules (ICAM-1) and Iba-1, two important markers of inflammation, indicating modulatory effects. Histological analyses of the cheek epithelium revealed greater deposition of type I collagen fibers in animals given PII-IAA compared with the control group. This performance was only reached when LPPII was treated with iodoacetamide (IAA), an irreversible inhibitor of proteolytic activity of cysteine proteases. The endogenous proteolytic activity of LPPII induced adverse effects in animals. Candidate proteins involved in the phytomodulatory activity are proposed. CONCLUSIONS: Therapy was successful in treating OM with the laticifer protein fraction, containing peptidases and osmotin, from Calotropis procera. The effective candidate from the latex proteins for therapeutic use is PII-IAA.

Chemical profiling of secondary metabolites from Himatanthus drasticus (Mart.) Plumel latex with inhibitory action against the enzymes α-amylase and α-glucosidase: In vitro and in silico assays.

ETHNOPHARMACOLOGICAL RELEVANCE: Himatanthus drasticus is an important medicinal plant whose latex is traditionally used in Northeast Brazil to treat various diseases, including diabetes. The use of α-amylase and α-glucosidase inhibitors can be an effective strategy to modulate levels of postprandial hyperglycemia via control of starch metabolism. AIMS OF THE STUDY: This study aimed to verify if H. drasticus latex has inhibitory activity against enzymes linked to type 2 diabetes, besides chemically characterizing the metabolites responsible for such activities. In addition, in silico analysis was performed to support the traditional claim of possible antidiabetic activity of this latex. MATERIALS AND METHODS: Latex from H. drasticus stems was sequentially partitioned with n-hexane (FHDH), CHCl3 (FHDC) and EtOH (FHDHA). Wash extraction of the FHDHA fraction was performed to obtain the other extract fractions. The FHDHA was submitted to chromatography in a SPE C18 cartridge using gradient elution with MeOH/H2O to produce five fractions: FHDHA1, FHDHA2, FHDHA3, FHDHA4 and FHDHA5. The FHDHA1 was subjected to semi-preparative reverse phase HPLC. Lineweaver-Burk plots were used to investigate the kinetic parameters of α-amylase and α-glucosidase inhibitory activity. The interactions between plumieride and porcine pancreatic α-amylase and α-glucosidase were analyzed through an in silico molecular docking study. RESULTS: Phytochemical identification of compounds present in the FHDHA fraction of H. drasticus latex was possible by 1H, 13C NMR analysis and mass spectrometry, and the results were compared with the literature. The identified compounds were α-ethyl glucoside, protocatechuic acid, 3-O-caffeoylquinic acid, 15-demethylplumieride acid, 5-O-caffeoylquinic acid, caffeic acid, vanillic acid, plumieride, and catechin. The inhibition results of the fractions tested against α-amylase and α-glucosidase showed inhibitory activities dependent on the increase of fractions and compound concentrations. The IC50 results obtained from FHDHA, FHDHA1 and plumieride fractions against α-amylase were 36.46, 72.61, 33.87 µg/mL respectively. The IC50 of plumieride was the closest to that of acarbose (22.52 µg/mL), a result similar to that obtained for α-glucosidase. The type of inhibition was competitive for both enzymes. CONCLUSIONS: There was strong inhibition of α-amylase and α-glucosidase by FHDHA, FHDHA1 and plumieride, suggesting that these enzymes slow glucose absorption.

A phytomodulatory hydrogel with enhanced healing effects.

The healing performance of a hydrogel composed of hemicelluloses extracted from seeds of Caesalpinia pulcherrima (Fabaceae) and mixed with phytomodulatory proteins obtained from the latex of Calotropis procera was characterized on excisional wounds. The hydrogel did not induce dermal irritability. When topically used on excisional wounds, the hydrogel enhanced healing by wound contraction. Histology and the measurement of inflammatory mediators (myeloperoxidase, interleukin-1ß, and interleukin-6) suggested that the inflammatory phase of the healing process was intensified, stimulating fibroplasia and neovascularization (proliferative phase) and tissue remodeling by increasing new collagen fiber deposition. In addition, reduction on levels of malondialdehyde in the groups that the hydrogel was applied suggested that the oxidative stress was reduced. The hydrogel performed better than the reference drug used, as revealed by the extended thickness of the remodeled epithelium.

Latex proteins from Calotropis procera: Toxicity and immunological tolerance revisited.

Many thousands of plants are disseminated worldwide in traditional and folk medicines based on the belief that their leaves, roots, seeds, bark or secretions, when adequately handled, can treat, alleviate or ameliorate numerous disease symptoms. Calotropis procera (Apocynaceae) is a popular medicinal plant and the claims of this shrub's phytomedicinal properties have been scientifically validated. In this study, further prospects towards the in vivo toxicity and oral immunological tolerance of phytomodulatory proteins isolated from the latex of C. procera are reported. Acute toxicity was determined in mice by oral and intraperitoneal administration of latex proteins (LP) and was followed behavioral, hematological and histological analyses. Oral immunological tolerance to LP was assessed by intraperitoneal immunization in mice that had received LP orally before. Animals given 5000 mg/kg orally exhibited only discrete behavioral alterations and augmentation of monocytes. Death was not notified 14 days after exposure. However, all animals receiving LP 150 mg/kg by i.p. died in 1 h. Death (20%) was documented when LP (75 mg/kg) was given in the peritoneum and signs of harmful effects were observed in the survivors (80%). Oral immunological tolerance was observed in animals previously given LP orally, when they were further immunized/challenged with peritoneal exposure to different doses of LP. This was confirmed by the lowering of IgE and IgG in the serum, IL-4 and IFN-γ in spleen homogenates and the absence of anaphylaxis signs. It is therefore concluded that LP exhibited quite discrete adverse effects when orally administrated at higher concentrations and this route of administration did not stimulate adverse immunological reactions. Instead it was observed immunological tolerance. The present study contributes very important information concerning the safe use of C. procera as a phytotherapeutic agent.

Cytotoxicity against tumor cell lines and anti-inflammatory properties of chitinases from Calotropis procera latex.

The role of chitinases from the latex of medicinal shrub Calotropis procera on viability of tumor cell lines and inflammation was investigated. Soluble latex proteins were fractionated in a CM Sepharose Fast-Flow Column and the major peak (LPp1) subjected to ion exchange chromatography using a Mono-Q column coupled to an FPLC system. In a first series of experiments, immortalized macrophages were cultured with LPp1 for 24 h. Then, cytotoxicity of chitinase isoforms (LPp1-P1 to P6) was evaluated against HCT-116 (colon carcinoma), OVCAR-8 (ovarian carcinoma), and SF-295 (glioblastoma) tumor cell lines in 96-well plates. Cytotoxic chitinases had its anti-inflammatory potential assessed through the mouse peritonitis model. We have shown that LPp1 was not toxic to macrophages at dosages lower than 125 µg/mL but induced high messenger RNA expression of IL-6, IL1-ß, TNF-α, and iNOs. On the other hand, chitinase isoform LPp1-P4 retained all LPp1 cytotoxic activities against the tumor cell lines with IC50 ranging from 1.2 to 2.9 µg/mL. The intravenous administration of LPp1-P4 to mouse impaired neutrophil infiltration into the peritoneal cavity induced by carrageenan. Although the contents of pro-inflammatory cytokines IL-6, TNF-α, and IL1-ß were high in the bloodstreams, such effect was reverted by administration of iNOs inhibitors NG-nitro-L-arginine methyl ester and aminoguanidine. We conclude that chitinase isoform LPp1-P4 was highly cytotoxic to tumor cell lines and capable to reduce inflammation by an iNOs-derived NO mechanism.

Wound healing modulation by a latex protein-containing polyvinyl alcohol biomembrane.

In a previous study, we performed the chemical characterization of a polyvinyl alcohol (PVA) membrane supplemented with latex proteins (LP) displaying wound healing activity, and its efficacy as a delivery system was demonstrated. Here, we report on aspects of the mechanism underlying the performance of the PVA-latex protein biomembrane on wound healing. LP-PVA, but not PVA, induced more intense leukocyte (neutrophil) migration and mast cell degranulation during the inflammatory phase of the cicatricial process. Likewise, LP-PVA induced an increase in key markers and mediators of the inflammatory response (myeloperoxidase activity, nitric oxide, TNF, and IL-1ß). These results demonstrated that LP-PVA significantly accelerates the early phase of the inflammatory process by upregulating cytokine release. This remarkable effect improves the subsequent phases of the healing process. The polyvinyl alcohol membrane was fully absorbed as an inert support while LP was shown to be active. It is therefore concluded that the LP-PVA is a suitable bioresource for biomedical engineering.

Effects on fibrinogen, fibrin, and blood coagulation of proteolytic extracts from fruits of Pseudananas macrodontes, Bromelia balansae, and B. hieronymi (Bromeliaceae) in comparison with bromelain.

Extracts rich in cysteine proteases obtained from fruits of Pseudananas macrodontes (Pm), Bromelia balansae (Bb), and B. hieronymi (Bh) have previously shown an anti-inflammatory effect on animal models. Given the close relationship between hemostasis and inflammation, it is attractive to investigate therapeutic agents capable of modulating both systems. The aim of this work was to study the effect of Pm, Bb, and Bh on fibrin(ogen) and blood coagulation compared with stem bromelain (Bro). Action on fibrinogen was electrophoretically and spectrophotometrically evaluated, fibrinolytic activity was measured both electrophoretically and by the fibrin plate assay, and the effect on blood coagulation was studied by conventional coagulation tests (PT and APPT). All extracts showed the same proteolytic preference for fibrinogen subunits, that is Aα > Bß, whereas γ was partially hydrolyzed by 100-fold concentration increase. Unlike Bro, cysteine proteases of Pm, Bb, and Bh increased absorbance at 540 nm of fibrinogen solution, suggesting thrombin-like activity, which was time-dependent and reached maximum values at lower concentration. All extracts showed the same proteolytic preference for fibrin subunits; however Pm, Bb, and Bh showed lower fibrinolytic activity than Bro at the assayed concentrations. Although Bb acted only as anticoagulant, Pm, Bh, and unexpectedly Bro showed dual action on blood coagulation: at low concentration showed procoagulant effect and at high concentration anticoagulant effect. Results reveal new plant species as potential sources of pharmacological agents for the treatment of a wide range of hemostatic disorders as well as to wound healing.

Proteomic analysis and purification of an unusual germin-like protein with proteolytic activity in the latex of Thevetia peruviana.

MAIN CONCLUSION: The latex from Thevetia peruviana is rich in plant defense proteins, including a 120 kDa cysteine peptidase with structural characteristics similar to germin-like proteins. More than 20,000 plant species produce latex, including Apocynaceae, Sapotaceae, Papaveraceae and Euphorbiaceae. To better understand the physiological role played by latex fluids, a proteomic analysis of Thevetia peruviana (Pers.) Schum latex was performed using two-dimensional gel electrophoresis and mass spectrometry. A total of 33 proteins (86 %) were identified, including storage proteins, a peptidase inhibitor, cysteine peptidases, peroxidases and osmotins. An unusual cysteine peptidase, termed peruvianin-I, was purified from the latex by a single chromatographic step involving gel filtration. The enzyme (glycoprotein) was inhibited by E-64 and iodoacetamide and exhibited high specific activity towards azocasein (K m 17.6 µM), with an optimal pH and temperature of 5.0-6.0 and 25-37 °C, respectively. Gel filtration chromatography, two-dimensional gel electrophoresis, and mass spectrometry revealed that peruvianin-I possesses 120 kDa, pI 4.0, and six subunits (20 kDa). A unique N-terminal amino acid sequence was obtained to oligomer and monomers of peruvianin-I (1ADPGPLQDFCLADLNSPLFINGYPCRNPALAISDDF36). High-resolution images from atomic force microscopy showed the homohexameric structure of peruvianin-I may be organized as a trimer of dimers that form a central channel similar to germin-like proteins. Peruvianin-I exhibited no oxalate oxidase and superoxide dismutase activity or antifungal effects. Peruvianin-I represents the first germin-like protein (GLP) with cysteine peptidase activity, an activity unknown in the GLP family so far.

Comparison of immunomodulatory properties of mannose-binding lectins from Canavalia brasiliensis and Cratylia argentea in a mice model of Salmonella infection.

The immunomodulatory properties of mannose-binding lectins ConBr (Canavalia brasiliensis) and CFL (Cratylia argentea) were investigated comparatively in a model of Salmonella infection. The lectins were intraperitoneally (i.p.) administered to mice daily for three days before the bacterial challenge with Salmonella enterica Ser. Typhimurium (0.2 mL i.p.; 10(7) CFU/mL). In vivo assays have shown that both lectins induced a significant leukocyte infiltration into the peritoneal cavity of uninfected mice, which was higher in the CFL group 3 days post-infection. Total and differential cell counts in the bloodstreams have shown uninfected animals pretreated with ConBr and CFL exhibited accentuated lymphopenia. Conversely, there was an increasing population of lymphocytes following 3 days post-infection in mice pretreated with both lectins. In addition, the bacterial burden was significantly reduced into the peritoneal cavity, bloodstreams, spleen and the liver in these mice. The lectins did not induce the release of pro- or anti-inflammatory cytokines into the peritoneal fluid of uninfected animals. However, following infection, the release of TNF-α and IL-10 in the peritoneal fluid were down-regulated in mice pretreated with both lectins whereas IL-1 was only reduced in mice pretreated with ConBr. Uninfected animals pretreated with CFL exhibited high nitric oxide (NO) content in the peritoneal fluid, which was decreased after infection in comparison to ConBr group. The lectins did not alter the serum levels of NO in uninfected mice but treatments with ConBr significantly reduced the NO content in infected animals in comparison to CFL group 24h after the bacterial challenge. Survival experiments have shown survival rates ranging from 70% to 100% in mice that received CFL or ConBr. On the other hand, untreated mice (PBS group) died 1-6 days after infection. We conclude that ConBr and CFL are prospective phytotherapeutics capable of modulate the cascade of pro-inflammatory plus regulatory cytokines and nitric oxide release derived from systemic infections.

Antitumor effect of laticifer proteins of Himatanthus drasticus (Mart.) Plumel - Apocynaceae.

ETHNOPHARMACOLOGICAL RELEVANCE: Himatanthus drasticus (Mart.) Plumel - Apocynaceae is a medicinal plant popularly known as Janaguba. Its bark and latex have been used by the public for cancer treatment, among other medicinal uses. However, there is almost no scientific research report on its medicinal properties. AIM OF THE STUDY: The aim of this study was to investigate the antitumor effects of Himatanthus drasticus latex proteins (HdLP) in experimental models. MATERIALS AND METHODS: The in vitro cytotoxic activity of the HdLP was determined on cultured tumor cells. HdLP was also tested for its ability to induce lysis of mouse erythrocytes. In vivo antitumor activity was assessed in two experimental models, Sarcoma 180 and Walker 256 carcinosarcoma. Additionally, its effects on the immunological system were also investigated. RESULTS: HdLP did not show any significant in vitro cytotoxic effect at experimental exposure levels. When intraperitoneally administered, HdLP was active against both in vivo experimental tumors. However, it was inactive by oral administration. The histopathological analysis indicates that the liver and kidney were only weakly affected by HdLP treatment. It was also demonstrated that HdLP acts as an immunomodulatory agent, increasing the production of OVA-specific antibodies. Additionally, it increased relative spleen weight and the incidence of megakaryocyte colonies. CONCLUSION: In summary, HdLP has some interesting anticancer activity that could be associated with its immunostimulating properties.

Protective effect of proteins derived from the latex of Calotropis procera against inflammatory hyperalgesia in monoarthritic rats.

Calotropis procera (family: Apocynaceae) is a plant growing in the wild and has been used in the traditional medicinal system for the treatment of various diseases. The plant produces milky latex that possesses potent antiinflammatory and analgesic properties. In present study the non-dialysable protein fraction isolated from the latex (LP) of this plant was evaluated for its efficacy against inflammation in rats where paw edema was induced by sub-plantar injection of carrageenin or monoarthritis was induced by intra-articular injection of Freund's complete adjuvant (FCA). The effect of LP was evaluated on edema volume in the paw model and on joint diameter, stair climbing ability, motility, dorsal flexion pain, levels of oxidative stress markers and joint histology in arthritis model. The protection afforded by LP was compared with that of standard antiinflammatory drug, diclofenac (5 mg/kg). LP exhibited a dose-dependent antiinflammatory effect and produced 32% and 60% inhibition of paw edema at 10 and 25 mg/kg doses and 12% and 36% inhibition of joint inflammation at 50 and 150 mg/kg doses. The protective effect of LP was associated with normalization of joint functions, histology and levels of oxidative stress markers in joint tissue. The findings of this study suggest that the protein fraction of latex of Calotropis procera has the potential to relieve inflammation and pain associated with various arthritic conditions.

Proteins from latex of Calotropis procera prevent septic shock due to lethal infection by Salmonella enterica serovar Typhimurium.

AIM OF THE STUDY: The latex of Calotropis procera has been used in traditional medicine to treat different inflammatory diseases. The anti-inflammatory activity of latex proteins (LP) has been well documented using different inflammatory models. In this work the anti-inflammatory protein fraction was evaluated in a true inflammatory process by inducing a lethal experimental infection in the murine model caused by Salmonella enterica Subsp. enterica serovar Typhimurium. MATERIALS AND METHODS: Experimental Swiss mice were given 0.2 ml of LP (30 or 60 mg/kg) by the intraperitoneal route 24 h before or after lethal challenge (0.2 ml) containing 10(6) CFU/ml of Salmonella Typhimurium using the same route of administration. RESULTS: All the control animals succumbed to infection within 6 days. When given before bacterial inoculums LP prevented the death of mice, which remained in observation until day 28. Even, LP-treated animals exhibited only discrete signs of infection which disappeared latter. LP fraction was also protective when given orally or by subcutaneous route. Histopathological examination revealed that necrosis and inflammatory infiltrates were similar in both the experimental and control groups on days 1 and 5 after infection. LP activity did not clear Salmonella Typhimurium, which was still present in the spleen at approximately 10(4) cells/g of organ 28 days after challenge. However, no bacteria were detected in the liver at this stage. LP did not inhibit bacterial growth in culture medium at all. In the early stages of infection bacteria population was similar in organs and in the peritoneal fluid but drastically reduced in blood. Titration of TNF-alpha in serum revealed no differences between experimental and control groups on days 1 and 5 days after infection while IL-12 was only discretely diminished in serum of experimental animals on day 5. Moreover, cultured macrophages treated with LP and stimulated by LPS released significantly less IL-1beta. CONCLUSIONS: LP-treated mice did not succumb to septic shock when submitted to a lethal infection. LP did not exhibit in vitro bactericidal activity. It is thought that protection of LP-treated mice against Salmonella Typhimurium possibly involves down-regulation of pro-inflammatory cytokines (other than TNF-alpha). LP inhibited IL-1beta release in cultured macrophages and discretely reduced IL-12 in serum of animals given LP. Results reported here support the folk use of latex to treat skin infections by topic application.

Involvement of NO in the inhibitory effect of Calotropis procera latex protein fractions on leukocyte rolling, adhesion and infiltration in rat peritonitis model.

AIM OF THE STUDY: The latex of Calotropis procera has been used in the traditional medicinal system for the treatment of leprosy, ulcers, tumors, piles and diseases of liver, spleen, abdomen and toothache. It comprises of a non-dialyzable protein fraction (LP) that exhibits anti-inflammatory properties and a dialyzable fraction (DF) exhibiting pro-inflammatory properties. The present study was carried out to evaluate the effect of LP sub-fractions on neutrophil functions and nociception in rodent models and to elucidate the mediatory role of nitric oxide (NO). MATERIAL AND METHODS: The LP was subjected to ion exchange chromatography and the effect of its three sub-fractions (LP(PI), LP(PII) and LP(PIII)) thus obtained was evaluated on leukocyte functions in the rat peritonitis model and on nociception in the mouse model. RESULTS: LP sub-fractions exhibit distinct protein profile and produce a significant decrease in the carrageenan and DF induced neutrophil influx and exhibit anti-nociceptive property. The LP and its sub-fractions produced a marked reduction in the number of rolling and adherent leukocytes in the mesenteric microvasculature as revealed by intravital microscopy. The anti-inflammatory effect of LP(PI), the most potent anti-inflammatory fraction of LP, was accompanied by an increase in the serum levels of NO. Further, our study shows that NO is also involved in the inhibitory effect of LP(PI) on neutrophil influx. CONCLUSIONS: Our study shows that LP fraction of Calotropis procera comprises of three distinct sets of proteins exhibiting anti-inflammatory and anti-nociceptive properties of which LP(PI) was most potent in inhibiting neutrophil functions and its effects are mediated through NO production.

Latex proteins from the plant Calotropis procera are partially digested upon in vitro enzymatic action and are not immunologically detected in fecal material.

Soluble proteins from the latex of Calotropis procera (LP) were investigated in vitro and in vivo for digestibility as the latex has previously been shown to produce considerable toxic effects on animals. The latex is also an important biologically active compound that displays antiinflammatory and antidiarrhea properties. The proteins were digested by the action of trypsin, pepsin or chemotrypsin as revealed by gel filtration and SDS-PAGE analysis. Furthermore, the full LP digestion was easily achieved by protease treatment. Rabbit polyclonal antibodies raised against LP failed to detect cross-reactive molecules in fecal material of experimental rats following 35 consecutive days of LP consumption in water. Similar patterns of electrophoresis were observed for the negligible amounts of protein observed in the fecal extracts of control and test animals. No death or toxic effects were observed among animals. Taken together these results suggest that harmful and toxic effects on animals of the latex from C. procera are present in its rubber and low molecular weight fractions rather than its protein content.

Antinociceptive activity of Calotropis procera latex in mice.

This work evaluated the antinociceptive effect of proteins from the Calotropis procera (Asclepiadaceae) latex using three different experimental models of nociception in mice. The latex protein fraction administered intraperitoneally in male mice at the doses of 12.5, 25 and 50 mg/kg showed the antinociceptive effect in a dose dependent manner compared to the respective controls in all assays. Inhibitions of the acetic acid-induced abdominal constrictions were observed at the doses of 12.5 (67.9%), 25 (85%) and 50 (99.5%) mg/kg compared to controls. Latex protein at the doses of 25 (39.8%; 42%) and 50 mg/kg (66.6%; 99.3%) reduced the nociception produced by formalin in the 1st and 2nd phases, respectively, and this effect was not reversed by pretreatment with naloxone (1 mg/kg). In the hot plate test, an increase of the reaction time was observed only at 60 min after the treatment with latex at the doses of 25 (79.5%) and 50 (76.9%) mg/kg, compared to controls and naloxone was ineffective to reverse the effect. It was concluded that the protein fraction derived from the whole latex of Calotropis procera possesses antinociceptive activity, which is independent of the opioid system.