Assessment of bioremediation potential of Calotropis procera and Nerium oleander for sustainable management of vehicular released metals in roadside soils.

Land transportation is a major source of heavy metal contamination along the roadside, posing significant risks to human health through inhalation, oral ingestion, and dermal contact. Therefore, this study has been designed to determine the concentrations of vehicular released heavy metals (Cd, Pb, Ni, and Cu) in roadside soil and leaves of two commonly growing native plant species (Calotropis procera and Nerium oleander).Two busy roads i.e., Lahore-Okara road (N-5) and Okara-Faisalabad roads (OFR) in Punjab, Pakistan, were selected for the study. The data were collected from five sites along each road during four seasons. Control samples were collected ~ 50 m away from road. The metal content i.e. lead (Pb), cadmium (Cd) nickel (Ni) and copper (Cu) were determined in the plant leaves and soil by using Atomic Absorption Spectrophotometer (AAS). Significantly high amount of all studied heavy metals were observed in soil and plant leaves along both roads in contrast to control ones. The mean concentration of metals in soil ranged as Cd (2.20-6.83 mg/kg), Pb (4.53-15.29 mg/kg), Ni (29.78-101.26 mg/kg), and Cu (61.68-138.46 mg/kg) and in plant leaves Cd (0.093-0.53 mg/kg), Pb (4.31-16.34 mg/kg), Ni (4.13-16.34 mg/kg) and Cu (2.98-32.74 mg/kg). Among roads, higher metal contamination was noted along N-5 road. Significant temporal variations were also noted in metal contamination along both roads. The order of metal contamination in soil and plant leaves in different seasons was summer > autumn > spring > winter. Furthermore, the metal accumulation potential of Calotropis procera was higher than that of Nerium oleander. Therefore, for sustainable management of metal contamination, the plantation of Calotropis procera is recommended along roadsides.

Combination of ethyl acetate fraction from Calotropis gigantea stem bark and sorafenib induces apoptosis in HepG2 cells.

The cytotoxicity of the ethyl acetate fraction of the Calotropis gigantea (L.) Dryand. (C. gigantea) stem bark extract (CGEtOAc) has been demonstrated in many types of cancers. This study examined the improved cancer therapeutic activity of sorafenib when combined with CGEtOAc in HepG2 cells. The cell viability and cell migration assays were applied in HepG2 cells treated with varying concentrations of CGEtOAc, sorafenib, and their combination. Flow cytometry was used to determine apoptosis, which corresponded with a decline in mitochondrial membrane potential and activation of DNA fragmentation. Reactive oxygen species (ROS) levels were assessed in combination with the expression of the phosphatidylinositol-3-kinase (PI3K)/ protein kinase B (Akt)/ mammalian target of rapamycin (mTOR) pathway, which was suggested for association with ROS-induced apoptosis. Combining CGEtOAc at 400 µg/mL with sorafenib at 4 µM, which were their respective half-IC50 concentrations, significantly inhibited HepG2 viability upon 24 h of exposure in comparison with the vehicle and each single treatment. Consequently, CGEtOAc when combined with sorafenib significantly diminished HepG2 migration and induced apoptosis through a mitochondrial-correlation mechanism. ROS production was speculated to be the primary mechanism of stimulating apoptosis in HepG2 cells after exposure to a combination of CGEtOAc and sorafenib, in association with PI3K/Akt/mTOR pathway suppression. Our results present valuable knowledge to support the development of anticancer regimens derived from the CGEtOAc with the chemotherapeutic agent sorafenib, both of which were administered at half-IC50, which may minimize the toxic implications of cancer treatments while improving the therapeutic effectiveness toward future medical applications.

Calotropis procera extract inhibits prostate cancer through regulation of autophagy.

Current treatment options available for prostate cancer (PCa) patients have many adverse side effects and hence, new alternative therapies need to be explored. Anticancer potential of various phytochemicals derived from Calotropis procera has been studied in many cancers but no study has investigated the effect of leaf extract of C. procera on PCa cells. Hence, we investigated the effect of C. procera leaf extract (CPE) on cellular properties of androgen-independent PC-3 and androgen-sensitive 22Rv1 cells. A hydroalcoholic extract of C. procera was prepared and MTT assay was performed to study the effect of CPE on viability of PCa cells. The effect of CPE on cell division ability, migration capability and reactive oxygen species (ROS) production was studied using colony formation assay, wound-healing assay and 2',7'-dichlorodihydrofluorescein diacetate assay, respectively. Caspase activity assay and LDH assay were performed to study the involvement of apoptosis and necrosis in CPE-mediated cell death. Protein levels of cell cycle, antioxidant, autophagy and apoptosis markers were measured by western blot. The composition of CPE was identified using untargeted LC-MS analysis. Results showed that CPE decreased the viability of both the PCa cells, PC-3 and 22Rv1, in a dose- and time-dependent manner. Also, CPE significantly inhibited the colony-forming ability, migration and endogenous ROS production in both the cell lines. Furthermore, CPE significantly decreased NF-κB protein levels and increased the protein levels of the cell cycle inhibitor p27. A significant increase in expression of autophagy markers was observed in CPE-treated PC-3 cells while autophagy markers were downregulated in 22Rv1 cells after CPE exposure. Hence, it can be concluded that CPE inhibits PCa cell viability possibly by regulating the autophagy pathway and/or altering the ROS levels. Thus, CPE can be explored as a possible alternative therapeutic agent for PCa.

Swarna Bhasma Induces Antigen-Presenting Abilities of Macrophages and Helps Antigen Experienced CD4+ T Cells to Acquire Th1 Phenotypes Against Leishmania donovani Antigens.

In leishmaniasis, the protective immunity is largely mediated by proinflammatory cytokine producing abilities of T cells and an efficient parasite killing by phagocytic cells. Notwithstanding a substantial progress that has been made during last decades, the mechanisms or factors involved in establishing protective immunity against Leishmania are not identified. In ancient Indian literature, metallic "bhasma," particularly that of "swarna" or gold (fine gold particles), is indicated as one of the most prominent metal-based therapeutic medicine, which is known to impart protective and curative properties in various health issues. In this work, we elucidated the potential of swarna bhasma (SB) on the effector properties of phagocytes and antigen-activated CD4+ T cells in augmenting the immunogenicity of L. donovani antigens. The characterization of SB revealing its shape, size, composition, and measurement of cytotoxicity established the physiochemical potential for its utilization as an immunomodulator. The activation of macrophages with SB enhanced their capacity to produce nitric oxide and proinflammatory cytokines, which eventually resulted in reduced uptake of parasites and their proliferation in infected cells. Further, in Leishmania-infected animals, SB administration reduced the generation of IL-10, an anti-inflammatory cytokine, and enhanced pro-inflammatory cytokine generation by antigen activated CD4+ T cells with increased frequency of double (IFNγ+/TNFα+) and triple (IFNγ+TNFα+IL-2+) positive cells and abrogated disease pathogeneses at the early days of infection. Our results also suggested that cow-ghee (A2) emulsified preparation of SB, either alone or with yashtimadhu, a known natural immune modulator which enhances the SB's potential in enhancing the immunogenicity of parasitic antigens. These findings suggested a definite potential of SB in enhancing the effector functions of phagocytes and CD4+ T cells against L. donovani antigens. Therefore, more studies are needed to elucidate the mechanistic details of SB and its potential in enhancing vaccine-induced immunity.

Assessment of Plant Growth-Promoting Parameters of Endophytes Isolated from Calotropis procera and Their Performance Under Irrigated and Non-irrigated Conditions.

In the present study, bacterial and fungal endophytes are isolated from Calotropis procera, a drought-resistant plant and studied for their role in plant growth promotion. Among bacterial sp. Enterobacter cloacae subsp. cloacae strain CPR5B and fungus, Penicillium citrinum strain CPL1F, were identified as potent endophytes as both strains were able to produce Indole Acetic Acid (IAA) and 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase and solubilize phosphate. Penicillium citrinum CPL1F also been shown to produce siderophore. The IAA production was observed to be 94.28 µg/mL and 17.1 µg/mL for bacterial and fungal sp., respectively. The phosphate solubilization was observed to be 76.41 µg/mL and 114.57 µg/mL, respectively. The in vitro plant treatment studies with bacterium and fungus under irrigated and non-irrigated conditions showed that both strains had promoted plant growth in both conditions with respect to their control. Both the strains showed significant changes in most of the growth parameters under endophyte-treated irrigated and non-irrigated conditions, suggesting their stress-dependent plant growth promotion. The present findings will contribute to exploring endophytes that enhance plant growth in adverse conditions and act as plant growth-promoting endophytes.

Identification of phytochemical, antioxidant, anticancer and antimicrobial potential of Calotropis procera leaf aqueous extract.

Since the dawn of civilization, people have turned to plants as a safe and efficient form of treatment for a variety of diseases. It has long been known that Calotropis procera has the potential to treat a number of diseases. In this study, the C. procera leaf aqueous extract was obtained using the maceration method, and p-coumaric was found to be the main compound. The extract was rich in phenols (174.82 mg gallic acid equivalent/g) and flavonoids (1781.7 µg quercetin equivalent/g). The extract had high antioxidant properties, as indicated by the IC50 values obtained for 2,2-diphenyl-1-picrylhydrazyl (DPPH) (366.33 µg/mL) and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) (169.04 µg/mL), as well as the ferric ions reducing antioxidant power (FRAP) (1.67 µg ascorbic acid equivalent/g of the extract). The cytotoxicity of the extract was evaluated against the survival of HT 29 cells, and the IC50 was found to be 236.87 µg/mL. The most resistant and sensitive strains to the extract were Escherichia coli and Staphylococcus aureus, respectively. The morphological changes of these strains were demonstrated through scanning electron microscopy and confocal laser scanning microscopy. The C. procera extract could be therefore used as an antioxidant, antimicrobial, and anticancer agent.

Comparative study of microscale and macroscale technique for encapsulation of Calotropis gigantea extract in metal-conjugated nanomatrices for invasive ductal carcinoma.

The encapsulation of plant extract in nanomatrices has limitations due to its adhesion to walls, size control, high cost and long durations that results in low yield. Macroscale and microscale level techniques for development of micro/nanoparticles may impact the encapsulation of plant extract. This study aimed to evaluate the relative efficiency of microscale and macroscale techniques for encapsulation of plant extract, which is not compared yet. Keeping this in view, encapsulation of Calotropis gigantea leaves extract (CaG) was attained in silver-conjugated poliglusam nanomatrices (POL/Ag) to induce apoptosis in invasive ductal carcinoma (IDC) cells. The ethanolic CaG extract was prepared using percolation method and characterized by chemical tests for its active phytochemical compounds. The droplet-based microfluidic system was utilized as microscale encapsulation technique for CaG in nanomatrices at two different aqueous to oil flow rate ratios 1.0:1.5, and 1.0:3.0. Moreover, conventional batch system was utilized as macroscale encapsulation technique consisted of hot plate magnetic stirrer. The prepared nanomatrices were analysed for antioxidant activity using DPPH test and for cytotoxicity analysis using MCF-7 cells. The characteristic peaks of UV-Vis, FTIR and XRD spectrum confirmed the synthesis of CaG(POL/Ag) by both the encapsulation methods. However, microfluidic system was found to be more expedient because of attaining small and uniform sized silver nanoparticles (92 ± 19 nm) at high flow rate and achieving high encapsulation efficiency (80.25%) as compared to the conventional batch method (52.5%). CaG(POL/Ag) nanomatrices found to have significant antioxidant activity (p = 0.0014) against DPPH radical scavenging activity. The CaG(POL/Ag) of the smallest sized formulated by the microfluidic system has also shown the highest cytotoxicity (90%) as compared to batch method (70%) at 80 µg/mL. Our results indicate that the microscale technique using microfluidic system is a more efficient method to formulate size-controlled CaG(POL/Ag) nanomatrices and achieve high encapsulation of plant extract. Additionally, CaG(Pol/Ag) was found to be an efficient new combination for inducing potent (p < 0.0001) apoptosis in IDC cells. Therefore, CaG(Pol/Ag) can be further tested as an anti-cancer agent for in-vivo experiments.

Calotropis procera (Aiton) Dryand (Apocynaceae): State of the art of its uses and Applications.

Calotropis procera (Aiton) Dryand (Apocynaceae), popularly known as milkweed, has been traditionally used to treat diseases particularly associated with gastric disorders, skin disease and inflammatory processes. The present study aimed to review the current scientific evidence regarding the pharmacological effects of C. procera extracted phytochemicals and possible research opportunities as complementary and alternative medicine. Scientific publications were searched in various electronic databases (PubMed, Scopus, Web of Science, Google Scholar, Springer, Wiley, and Mendeley) using the following search terms: Calotropis procera, medicinal plants, toxicity, phytochemical characterization, and biological effects. Collected data showed that cardenolides, steroid glycoside and flavonoids are the main classes of phytochemicals identified in C. procera latex and leaves. In addition, lignans, terpenes, coumarins, and phenolic acids have been reported. These metabolites have been correlated with their biological activities, including mainly antioxidant, anti-inflammatory, antitumoral, hypoglycemic, gastric protective, anti-microbial, insecticide, anti-fungal, anti-parasitic, among others. However, some of the studies were carried out with only a single dose or with a high dose not achievable under physiological conditions. Therefore, the validity of C. procera biological activity may be questionable. Not less important to highlight are the risks associated with its use and the possibility of accumulation of heavy metals that can be toxic. Furthermore, there are no clinical trials with C. procera to date. In conclusion, the need of bioassayguided isolation of bioactive compounds, bioavailability and efficacy, as well as pharmacological and toxicity studies, are needed using in vivo models and clinical trials in order to support the traditionally claimed health benefits.

In Vitro: The Effects of the Anticoccidial Activities of Calotropis procera Leaf Extracts on Eimeria stiedae Oocysts Isolated from Rabbits.

Hepatic coccidiosis is an infectious and mortal disease that causes global economic losses in rabbits. The research aimed to assess the efficacy of Calotropis procure leaf extracts on the inhibition of Eimeria stiedae oocysts and to determine the optimal dosage for suppressing the parasite's infective phase. In this experiment, oocyst samples per milliliter were tested, and 6-well plates (2 mL) of 2.5% potassium dichromate solution containing 102 non-sporulated oocysts on Calotropis procera leaf extracts were exposed after 24, 48, 72, and 96 h, and the treatments were as follows: a nontreated control, 25%, 50%, 100%, and 150% of C. procera for oocyst activities. In addition, amprolium was utilized as a reference drug. The Calotropis procera was analyzed by GC-Mass, and results showed that the botanical extract contained 9 chemical components that were able to inhibit the oocysts of E. stiedae at 100% and 150% concentrations by about 78% and 93%, respectively. In general, an increase in the incubation period and a greater dose resulted in a decrease in the inhibition rate. The results showed that C. procera has an effective ability, inhibitory potential, and protective effect on the coccidian oocyst sporulation of E. stiedae. It can be used in the disinfection and sterilization of poultry and rabbit houses to get rid of Eimeria oocysts.

Calotropis procera latex protein reduces inflammation and bone loss in ligature-induced period ontitis in male rats.

OBJECTIVE: Calotropis procera latex protein (CpLP) is a popular anti-inflammatory and therefore we aimed to study its effects on inflammatory bone loss. DESIGN: Male Wistar rats were subjected to a ligature of molars. Groups of rats received intraperitoneally CpLP (0.3 mg/kg, 1 mg/kg, or 3 mg/kg) or saline (0.9% NaCl) one hour before ligature and then daily up to 11 days, compared to naïve. Gingiva was evaluated by myeloperoxidase activity and interleukin-1 beta (IL-1ß) expression by ELISA. Bone resorption was evaluated in the region between the cement-enamel junction and the alveolar bone crest. The histology considered alveolar bone resorption and cementum integrity, leukocyte infiltration, and attachment level, followed by immunohistochemistry bone markers between 1st and 2nd molars. Systemically, the weight of the body and organs, and a leukogram were performed. RESULTS: The periodontitis significantly increased myeloperoxidase activity and the IL-1ß level. The increased bone resorption was histologically corroborated by periodontal destruction, leukocyte influx, and attachment loss, as well as the increasing receptor activator of the nuclear factor-kappa B ligand (RANKL)/osteoprotegerin (OPG) ratio, and Tartrate-resistant acid phosphatase (TRAP)+ cells when compared to naïve. CpLP significantly reduced myeloperoxidase activity, level of IL-1ß, alveolar bone resorption, periodontal destruction, leukocyte influx, and attachment loss. The CpLp also reduced the RANKL/OPG ratio and TRAP+ cells, when compared with the saline group, and did not affect the systemic parameters. CONCLUSIONS: CpLP exhibited a periodontal protective effect by reducing inflammation and restricting osteoclastic alveolar bone resorption in this rat model.

Anastomosing laticifer in the primary and secondary structures of Calotropis procera (Aiton) W.T.Aiton (Apocynaceae) stems.

An in-depth understanding of the development and distribution of laticifer (latex secretory structure) will be important for the production of both rubber and medicines and will support studies on plant adaptations to their environments. We characterize here and describe the ontogenesis of the laticifer sytem in Calotropis procera (Apocynaceae), an invasive subshrub species in arid landscapes. Anatomical and histochemical evaluations of the primary and secondary structures of the stem were carried out on a monthly basis during a full year, with ultrastructural evaluations of laticifer on the stem apex during the rainy season. In the primary structure, laticifer differentiate early from procambium and ground meristem cells of the cortex and medulla and become concentrated adjacent to the external and internal phloem of the bicollateral bundles. In the secondary structure, laticifer differentiates from fusiform derivative cells of the phloem close to the sieve-tube elements. The laticifer is of the articulated, anastomosing, branched type, and it originates from precursor cells that loose the transversal and longitudinal walls by dissolution. Latex is a mixture of terpenes, alkaloids, flavonoids, mucilage, and proteins. The apical meristem and vascular cambium where the laticifer system begins its development are active throughout the year, including during the dry season. The vascular cambium produces phloem with laticifer precursor cells during the rainy season, with high temperatures and long days. The ability of C. procera to grow under water deficit conditions and produce laticifer throughout the year contribute to its wide distribution in arid environments.

Use of Calotropis procera cysteine peptidases (CpCPs) immobilized on glyoxyl-agarose for cheesemaking.

Calotropis procera cysteine peptidases (CpCPs) have presented several potential biotechnological applications. Here, these enzymes were immobilized on glyoxyl-agarose (glyoxyl-CpCPs) with yields of 90-95 % and the recovered activities ranged from 10 % to 15 %, according to enzyme loadings (5, 10, 20, 40, and 50 mgBSAeq/g). Spectrophotometric assays and SDS-PAGE showed that the casein hydrolysis by glyoxyl-CpCPs was similar to soluble CpCPs. In addition, glyoxyl-CpCPs exhibited similar ratio of milk-clotting activity to proteolytic activity in comparison with soluble CpCPs and chymosin. Even after being stored for six months at 8 °C, the residual proteolytic activity of glyoxyl-CpCPs remained close to 100 %. Atomic force microscopy and dynamic light scattering techniques showed that the process of casein micelle aggregation after treatment with glyoxyl-CpCPs was very similar to its soluble form and chymosin. Glyoxyl-CpCPs performed well after five reaction cycles, producing cheeses with yield, moisture, protein, and fat similar to those produced with chymosin.

Phytochemical analysis, anti-inflammatory, antioxidant activity of Calotropis gigantea and its therapeutic applications.

ETHNOPHARMACOLOGICAL RELEVANCE: Herbal remedies can be used to treat a variety of chronic inflammatory illnesses, like rheumatoid arthritis and leprosy. The plant Calotropis gigantea (C. gigantea) belongs to the family Apocynaceae. To treat numerous contagious diseases, C. gigantea is utilized alone or combine with certain medicinal herbs. Traditional Asian and African practitioners employed C. gigantea to treat a variety of inflammatory conditions like boils, rheumatoid arthritis, gout, leprosy and other disorders. AIM OF THE STUDY: The goal of this study is to examine the anti-inflammatory and antioxidant activities of C. gigantea leaf extracts extracted using methanol, petroleum ether, and water. MATERIALS AND METHODS: The leaf extracts of C. gigantea were obtained using the Soxhlet extraction technique. The phytoconstituents present in all three C. gigantea leaf extracts were confirmed by qualitative analysis, and the amounts of the alkaloids, flavonoids, terpenoids and phenols found in the extracts were quantified. C. gigantea crude extracts were subjected to a nitric oxide scavenging experiment to assess their free radical scavenging activities. Protein denaturation and proteinase inhibition assays were used to investigate the effectiveness of extracts to restrict denaturation of protein and to inhibit key enzymes responsible for tissue damage. Further, the membrane stabilization efficacy of plant extracts were examined by the heat-induced hemolysis method. The DPPH and FRAP experiments were performed to determine the antioxidant effectiveness of phytoconstituents extracted using different solvents. The GC-MS study of plant C. gigantea methanolic, aqueous and petroleum ether extracts displayed a broad range of compounds that possess beneficial therapeutic effects. RESULTS: This study reveals that the methanolic extract of C. gigantea provides significantly more anti-inflammatory and antioxidant activity than other extracts. CONCLUSION: Compared to the aqueous and petroleum ether extracts, the methanolic leaf extract of C. gigantea demonstrated greater in vitro anti-inflammatory and antioxidant properties.

Polyphenols-Rich Extract of Calotropis procera Alone and in Combination with Trichoderma Culture Filtrate for Biocontrol of Cantaloupe Wilt and Root Rot Fungi.

Fungal diseases have always been a major problem for cantaloupe crops; however, synthetic fungicides are hazardous to humans and the environment. Consequently, a feasible alternative to fungicides without side effects could be by using bio agents and naturally occurring plants with antibacterial potential. This study has achieved a novel procedure for managing wilt and root rot diseases by potentially using Trichoderma sp. culture filtrates in consortium with plant extract of Calotropis procera, Rhizoctonia solani, Fusarium oxysporum, and Pythium ultimum, which were isolated from infected cantaloupe roots with identified root rot symptoms. The antagonistic activity of four Trichoderma isolates and analysis of antibiotics and filtrate enzymes of the most active Trichoderma isolate were determined as well as phytochemical analysis of C. procera plant extract using HPLC-UV. The obtained results showed that all Trichoderma isolates considerably lowered the radial growth of P. ultimum, R. solani, and F. oxysporum in varying degrees. The scanning electron micrographs illustrate the mycoparasitic nature of Trichoderma sp. on F. oxysporum. The phytochemical analysis of C. procera indicated that phenolic contents were the major compounds found in extracts, such as vanillin (46.79%), chlorogenic acid (30.24%), gallic acid (8.06%), and daidzein (3.45%) but including only a low amount of the flavonoid compounds rutin, naringenin, and hesperetin. The Pot experiment's findings showed that cantaloupe was best protected against wilting and root rot diseases when it was treated with both Trichoderma sp. culture filtrates (10%) and C. procera extract of (15 mg/mL), both alone and in combination. This study demonstrates that the application of bio agent Trichoderma spp. filtrate with C. procera phenol extract appears useful for controlling wilting and root rot disease in cantaloupe. This innovative approach could be used as an alternative to chemical fungicide for the control of wilting and rot root diseases.

Insecticidal potential of organic extracts of Calotropis procera to Spodoptera frugiperda

This study evaluated the toxic effects of organic extracts of Calotropis proceraleaves on the survival, development, and reproduction of Spodoptera frugiperda. Solutions of crude methanol extract and hexane and methanol fractions of C. proceraleaves were added at 1.15% and 2.14% concentrations to the artificial diet of S. frugiperda. The mortality and duration of larval and pupal phases, weights of female and male pupae, deformations of pupae and adults, the reduction of adults able to reproduce, pre-oviposition and oviposition periods, the number of postures per female, and the fecundity and fertility of S. frugiperdafemales were also evaluated. The extracts harmed the survival, development, and reproduction of S. frugiperda. The ingestion of extracts and fractions by caterpillars affected adults by decreasing the oviposition period, the number of postures, fecundity, and fertility. The crude MeOH extract at a2.14% concentration harmed the evaluated parameters of the insect, except for pupal mortality, female pupae weight, and pre-oviposition period. The MeOH fraction at 2.14% caused a 50.0% mortality of caterpillars and 16.0% deformation in pupae and 33.0% inadults, reducing by 72.0% the population able to reproduce. The MeOH fraction at the 2.14% concentration caused 25.0% and 38.0% of pupal mortality and deformation, respectively. Calotropis procerahas promising insecticidal properties for a biological insecticide, a convenient and sustainable strategy for protecting plants against S. frugiperda.

Ovicidal and miracicidal activities of Calotropis procera and its green-synthesized nanotized derivative: A quest for new antifasciola agents.

Fascioliasis is an important zoonotic disease but treatment with the mainstay drugs poses challenge of parasite resistance. The aim of the study was to determine the anthelmintic efficacy of ethanolic leaf extract of Calotropis procera (CP) and its synthesized silver nanoparticles (AgNPs) against the eggs and miracidia of Fasciola species. The ethanolic extract of C. procera was used to synthesise its corresponding green-synthesis derivative using silver nitrate (CP-AgNPs). The synthesized silver nanoparticles were characterized by Fourier Transform Infrared Spectroscopy (FTIR), X-ray Diffraction (XRD) and Scanning Electron Microscopy/Energy Dispersive X-ray (SEM/EDX). The ova and miracidia of Fasciola spp. were exposed to 1, 2, and 4 mg/ml CP ethanolic extracts and its corresponding AgNPs. FTIR showed that the formulation was capped with compounds present in the extract. The XRD showed the crystalline property of CP-AgNPs. The SEM image showed clusters of irregularly shape nanoparticles. The ovicidal activities were concentration dependent and showed highest activities 81.02±4.03% and 92.91±1.25% in 4 mg/ml CP and CP-AgNPs respectively (p < 0.05). The LC50 of CP (1.49 mg/ml) was more than 3 folds higher than that of CP-AgNPs (0.47 mg/ml). While CP did not cause miracidia death after 60 min exposure, however, 100% miracidia death were observed within 30 min exposure in all the tested concentration with CP-AgNPs. The positive control (ABZ) only showed 100% mortality after 60 min of exposure of miracidia. The study showed that green-synthesised C. procera nanoparticles showed superior ovicidal and miracicidal activities over C. procera leaf extracts and could be a source of potential antifasciola agent.

Purification, Partial Characterization, and Evaluation of the Antiulcer Activity of Calotropis procera Leaf Lectin.

BACKGROUND: Lectins are proteins with therapeutic and diagnostic potential that can be applied in battling various ailments. AIM AND OBJECTIVE: This study was designed to purify and characterize the hemagglutinating activity derived from the leaves of Calotropis procera and its possible role in protecting the stomach against ethanol-induced lesions. METHODS: The Calotropis procera leaf lectin (ProLec), was isolated by homogenization of the defatted leaf powder in Phosphate-Buffered Saline (PBS) and purified by affinity chromatography on Sephadex G-100. The lectin was eluted from the affinity column by 3% acetic acid and was physicochemically characterized. In a dose-dependent manner, ProLec was administered to rats with ethanol-induced ulcers, and biochemical, histopathological, and toxicological examinations were performed. RESULTS: ProLec is a heterodimer of 75 and 68 kDa. It agglutinated all human RBCs, whereas it showed weak interaction with animal erythrocytes. The protein was optimally active at 25 °C and was labile above this temperature. ProLec exhibited two pH optima and was a metalloprotein requiring Ca, Mn, and Ni. It contains 1.6% tryptophan residues of which about 1% is exposed and critical for lectin activity. The lectin exhibited a potent gastroprotective effect against ethanolinduced gastric lesions with no apparent toxicity to both kidneys and liver. Examination of the pH of the gastric juice of lectin-treated animals indicated a possible role of lectin in maintaining stomach acidity within the normal ranges compared to the gastric juice pH of animals that received ethanol only. CONCLUSION: These results may suggest that ProLec could conceivably be a good future drug for the treatment of gastric ulcers, however, extensive immunological and toxicological research remains to be done.

Green Synthesis and Antimicrobial Activities of Silver Nanoparticles Using Calotropis gigantea from Ie Seu-Um Geothermal Area, Aceh Province, Indonesia.

Herein, we report our success synthesizing silver nanoparticles (AgNPs) using aqueous extracts from the leaves and flowers of Calotropis gigantea growing in the geothermal manifestation Ie Seu-Um, Aceh Besar, Indonesia. C. gigantea aqueous extract can be used as a bio-reductant for Ag+→Ag0 conversion, obtained by 48h incubation of Ag+, and the extract mixture in a dark condition. UV-Vis characterization showed that the surface plasmon resonance (SPR) peaks of AgNPs-leaf C. gigantea (AgNPs-LCg) and AgNPs-flower C. gigantea (AgNPs-FCg) appeared in the wavelength range of 410-460 nm. Scanning electron microscopy energy-dispersive X-ray spectrometry (SEM-EDS) revealed the agglomeration and spherical shapes of AgNPs-LCg and AgNPs-FCg with diameters ranging from 87.85 to 256.7 nm. Zeta potentials were observed in the range of -41.8 to -25.1 mV. The Kirby-Bauer disc diffusion assay revealed AgNPs-FCg as the most potent antimicrobial agent with inhibition zones of 12.05 ± 0.58, 11.29 ± 0.45, and 9.02 ± 0.10 mm for Escherichia coli, Staphylococcus aureus, and Candida albicans, respectively. In conclusion, aqueous extract from the leaves or flowers of Calotropis gigantea may be used in the green synthesis of AgNPs with broad-spectrum antimicrobial activities.

First Report of Fruit Rot of Cherry and Its Control Using Fe2O3 Nanoparticles Synthesized in Calotropis procera.

Cherry is a fleshy drupe, and it is grown in temperate regions of the world. It is perishable, and several biotic and abiotic factors affect its yield. During April-May 2021, a severe fruit rot of cherry was observed in Swat and adjacent areas. Diseased fruit samples were collected, and the disease-causing pathogen was isolated on PDA. Subsequent morphological, microscopic, and molecular analyses identified the isolated pathogen as Aspergillus flavus. For the control of the fruit rot disease of cherry, iron oxide nanoparticles (Fe2O3 NPs) were synthesized in the leaf extract of Calotropis procera and characterized. Fourier transform infrared (FTIR) spectroscopy of synthesized Fe2O3 NPs showed the presence of capping and stabilizing agents such as alcohols, aldehydes, and halo compounds. X-ray diffraction (XRD) analysis verified the form and size (32 nm) of Fe2O3 NPs. Scanning electron microscopy (SEM) revealed the spinal-shaped morphology of synthesized Fe2O3 NPs while X-ray diffraction (EDX) analysis displayed the occurrence of main elements in the samples. After successful preparation and characterization of NPs, their antifungal activity against A. flavus was determined by poison technique. Based on in vitro and in vivo antifungal activity analyses, it was observed that 1.0 mg/mL concentration of Fe2O3 can effectively inhibit the growth of fungal mycelia and decrease the incidence of fruit rot of cherry. The results confirmed ecofriendly fungicidal role of Fe2O3 and suggested that their large-scale application in the field to replace toxic chemical fungicides.

Calotropis gigantea stem bark extracts inhibit liver cancer induced by diethylnitrosamine.

Several fractions of Calotropis gigantea extracts have been proposed to have potential anticancer activity in many cancer models. The present study evaluated the anticancer activity of C. gigantea stem bark extracts in liver cancer HepG2 cells and diethylnitrosamine (DEN)-induced primary liver cancer in rats. The carcinogenesis model induced by DEN administration has been widely used to study pathophysiological features and responses in rats that are comparable to those seen in cancer patients. The dichloromethane (CGDCM), ethyl acetate, and water fractions obtained from partitioning crude ethanolic extract were quantitatively analyzed for several groups of secondary metabolites and calactin contents. A combination of C. gigantea stem bark extracts with doxorubicin (DOX) was assessed in this study to demonstrate the enhanced cytotoxic effect to cancer compared to the single administration. The combination of DOX and CGDCM, which had the most potential cytotoxic effect in HepG2 cells when compared to the other three fractions, significantly increased cytotoxicity through the apoptotic effect with increased caspase-3 expression. This combination treatment also reduced ATP levels, implying a correlation between ATP and apoptosis induction. In a rat model of DEN-induced liver cancer, treatment with DOX, C. gigantea at low (CGDCM-L) and high (CGDCM-H) doses, and DOX + CGDCM-H for 4 weeks decreased the progression of liver cancer by lowering the liver weight/body weight ratio and the occurrence of liver hyperplastic nodules, fibrosis, and proliferative cells. The therapeutic applications lowered TNF-α, IL-6, TGF-ß, and α-SMA inflammatory cytokines in a similar way, implying that CGDCM had a curative effect against the inflammation-induced liver carcinogenesis produced by DEN exposure. Furthermore, CGDCM and DOX therapy decreased ATP and fatty acid synthesis in rat liver cancer, which was correlated with apoptosis inhibition. CGDCM reduced cleaved caspase-3 expression in liver cancer rats when used alone or in combination with DOX, implying that apoptosis-inducing hepatic carcinogenesis was suppressed. Our results also verified the low toxicity of CGDCM injection on the internal organs of rats. Thus, this research clearly demonstrated a promising, novel anticancer approach that could be applied in future clinical studies of CGDCM and combination therapy.