Development, validation, and application of a multimatrix UHPLC-MS/MS method for quantification of Datura-type alkaloids in food.

A quantitative ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the determination of tropane alkaloids (TAs), atropine and scopolamine, in a variety of food products. The sample preparation of cereal-based food, oilseeds, honey, and pulses consisted of a solid-liquid extraction with an acidified mixture of methanol and water, while an additional step of solid-phase extraction on a cation-exchange sorbent was introduced in the treatment of teas and herbal infusions, aromatic herbs, spices and food supplements. The limits of quantification of the method varied from 0.5 to 2.5 µg kg-1. Apparent recovery was in the range of 70-120%, and repeatability and intermediate precision were below 20%. The method was successfully applied in a proficiency testing exercise as well as in the analysis of various commercial foods. Only 26% of the analysed food samples contained one or both TAs. The mean concentrations for atropine and scopolamine amounted to 21.9 and 6.5 µg kg-1, respectively, while the maximum concentrations were 523.3 and 131.4 µg kg-1, respectively. Overall, the highest levels of TA sum were found in an herbal infusion of fennel and a spice mix containing fennel and anise seeds.

Withanolides from Datura ceratocaula and Datura discolor and their acetylcholinesterase inhibitory activity.

The investigation of the chemical constituents of Datura ceratocaula and D. discolor allowed to isolate three new withanolides, datucerolide A (1) from the first species, and datudiscolides A (8) and B (9) from the second. In addition, seven known withanolides and five ubiquitous compounds were isolated from these plants, along with 27-O-ß-d-glucopyranosyl dinnoxolide A (5), which was obtained as the tetraacetyl derivative 4. All the structures were elucidated by analyses of their spectroscopic and spectrometric data and that of dinnoxolide A (6) was confirmed by X-ray diffraction analysis. The structure 4 was assigned earlier to daturametelin G-Ac and that of 5 to datinolide B, therefore, it will be discussed whether these assignments are correct. On the other hand, the structure of datudiscolide A (8) was previously assigned to the aglycone of dinoxin B (14), however, a revision of its reported NMR data showed inconsistencies with the proposed structure. The inhibitory activity of withanolides 2, 3, 6-8, 12, and 13 against acetylcholinesterase enzyme (AChE) was evaluated. Compounds 6, 7, 12, and 13 exhibited the best activity with IC50 values ranging from 2.8 to 21.5 µM.

Datinolides E-I, five new withanolides with anti-inflammatory activity from the leaves of Datura inoxia Mill.

Five new withanolides, datinolides E-I (1-5), and three known withanolides (6-8) were separated from Datura inoxia Mill. leaves, and datinolide E (1) was the first withanolide with C-27 connected to a nitrogen-containing group. Their structures were clarified by comprehensive spectroscopic analysis and comparison with literature. The anti-inflammatory of isolated compounds against RAW264.7 cells was investigated by the CCK8 assay.

Four new withanolides with anti-inflammatory activity from Datura inoxia Mill. leaves.

Four previously undescribed withanolides, datinolides A-D (1-4) and eight known withanolides (5-12), were separated from the 70% ethyl alcohol extract of Datura inoxia Mill. leaves. All structures were clarified by comprehensive spectroscopic analysis. Furthermore, all withanolides were assessed for their anti-inflammatory activity and results showed that 1 exhibited a fairly good suppression against nitric oxide generation in lipopolysaccharide-stimulated RAW 264.7 cells (IC50 = 10.33 ± 1.53 µM).

Alkaloids of the Genus Datura: Review of a Rich Resource for Natural Product Discovery.

The genus Datura (Solanaceae) contains nine species of medicinal plants that have held both curative utility and cultural significance throughout history. This genus' particular bioactivity results from the enormous diversity of alkaloids it contains, making it a valuable study organism for many disciplines. Although Datura contains mostly tropane alkaloids (such as hyoscyamine and scopolamine), indole, beta-carboline, and pyrrolidine alkaloids have also been identified. The tools available to explore specialized metabolism in plants have undergone remarkable advances over the past couple of decades and provide renewed opportunities for discoveries of new compounds and the genetic basis for their biosynthesis. This review provides a comprehensive overview of studies on the alkaloids of Datura that focuses on three questions: How do we find and identify alkaloids? Where do alkaloids come from? What factors affect their presence and abundance? We also address pitfalls and relevant questions applicable to natural products and metabolomics researchers. With both careful perspectives and new advances in instrumentation, the pace of alkaloid discovery-from not just Datura-has the potential to accelerate dramatically in the near future.

Cytotoxic withanolides from Datura innoxia.

Chemical investigation of the aerial parts (except fruits) of the medicinal, hallucinogen and toxic plant Datura innoxia Mill. [Solanaceae] led to the isolation of the new withanolide, dinnoxolide A (1), along with the known compounds 21,27-dihydroxy-1-oxowitha-2,5,24-trienolide (2), daturamalakin B (3) and withametelin (4). Their structures were established by analysis of their spectroscopic data, including 1D and 2D NMR experiments and MS. Compounds 2 and 3 were isolated as natural products for the first time and the name dinnoxolide B was given to compound 2. The four withanolides showed in vitro cytotoxic activity against U251 (glioblastoma) and SK-LU-1 (lung adenocarcinoma) human cancer cell lines, with IC50 values ranging from 1.2 to 19.6 µM, and also against the noncancerous monkey kidney cell line (COS-7), with IC50 values ranging from 5.0 to 19.7 µM. Compound 4 was two times more active than the reference compound, etoposide, against lung adenocarcinoma cells.

Chemical and Fatty Acid Compositions of Crude and Purified Extracts Obtained from Datura innoxia Seeds Extracted with Different Solvents.

Oils play a key role as raw materials in a variety of industries. The aim of this study was to evaluate the potential of Datura innoxia seed oil cultivated in Saudi Arabia for industrial purpose and to study the effects of hexane, chloroform, and isopropanol as extraction solvents on the compositions of the extracts. The results showed that the hexane and chloroform extracts were mainly neutral oils which were rich in linoleic (≈46%) and oleic (≈31%) acids. However, the isopropanol extract contained large amount of neutral oil and organic acids. Neutral oil contained mainly palmitic acid (40.2%) and some important and valuable epoxy (15.4%) and cyclopropane (13.2%) fatty acids. Analysis of the sterol and tocopherol levels of the crude and purified oil extracted revealed that they were significantly affected by the extraction solvent used.

Determination of atropine and scopolamine in spinach-based products contaminated with genus Datura by UHPLC-MS/MS.

Datura species are well known because of their high concentration of tropane alkaloids, which has led to poisoning episodes when Datura is accidentally mixed with edible crops. Therefore, the European Union has set a maximum level in cereal-based infant food products of 1 µg kg-1 for atropine and scopolamine. However, the occurrence of these compounds in other commodities has become a global concern. Spinach and derived products can be contaminated with Datura innoxia leaves. In this study, we tested frozen spinachs and spinach-based infant food products. The determination was carried out by UHPLC-MS/MS after applying the QuEChERS method as sample treatment. The LOQs were below 0.016 µg kg-1, achieving satisfactory results in terms of precision, accuracy, and matrix effects. The obtained results (ranging between 0.02 and 8.19 µg kg-1) were close to the maximum level set by the European Union for 24% of the samples tested.

Datura quids at Pinwheel Cave, California, provide unambiguous confirmation of the ingestion of hallucinogens at a rock art site.

While debates have raged over the relationship between trance and rock art, unambiguous evidence of the consumption of hallucinogens has not been reported from any rock art site in the world. A painting possibly representing the flowers of Datura on the ceiling of a Californian rock art site called Pinwheel Cave was discovered alongside fibrous quids in the same ceiling. Even though Native Californians are historically documented to have used Datura to enter trance states, little evidence exists to associate it with rock art. A multianalytical approach to the rock art, the quids, and the archaeological context of this site was undertaken. Liquid chromatography-mass spectrometry (LC-MS) results found hallucinogenic alkaloids scopolamine and atropine in the quids, while scanning electron microscope analysis confirms most to be Datura wrightii Three-dimensional (3D) analyses of the quids indicate the quids were likely masticated and thus consumed in the cave under the paintings. Archaeological evidence and chronological dating shows the site was well utilized as a temporary residence for a range of activities from Late Prehistory through Colonial Periods. This indicates that Datura was ingested in the cave and that the rock painting represents the plant itself, serving to codify communal rituals involving this powerful entheogen. These results confirm the use of hallucinogens at a rock art site while calling into question previous assumptions concerning trance and rock art imagery.

Preclinical anticancer studies on the ethyl acetate leaf extracts of Datura stramonium and Datura inoxia.

BACKGROUND: Cancer is a horrific disease relentlessly affecting human population round the globe. Genus Datura encompasses numerous species with reported medicinal uses. However, its potential as a source of natural anticancer agents is yet to be determined. Datura stramonium (DS) and Datura inoxia (DI) are the two species chosen for this study. METHODS: Total phenolic and flavonoid content (TPC and TFC) as well as antioxidant activity were assessed through colorimetric method. Polyphenolic quantification was done by RP-HPLC. Following extract standardization ethyl acetate leaf extracts of both species (DSL-EA and DIL-EA) were chosen for anticancer studies. In vitro cytotoxicity using various models including cancer cell lines was monitored. Following toxicity studies, benzene (0.2 ml) was used to induce leukemia in Sprague-Dawley rats. Extracts were orally administered to preventive (100 and 200 mg/kg) and treatment (200 mg/kg only) groups. The antileukemic potential of extracts was assessed through haematological, biochemical, endogenous antioxidants and histological parameters. RESULTS: Significant TPC and TFC were estimated in DSL-EA and DIL-EA. RP-HPLC quantified (µg/mg extract) rutin (0.89 ± 0.03), gallic acid (0.35 ± 0.07), catechin (0.24 ± 0.02) and apigenin (0.29 ± 0.09) in DSL-EA while rutin (0.036 ± 0.004) and caffeic acid (0.27 ± 0.03) in DIL-EA. Both extracts exhibited significant brine shrimp cytotoxicity (LC50 < 12.5 µg/ml). DIL-EA exhibited greater cytotoxicity against PC-3, MDA-MB 231 and MCF-7 cell lines (IC50 < 3 µg/ml in each case) as well as higher protein kinase inhibitory action (MIC: 25 µg/disc) compared to DSL-EA. Leukemia induced in rats was affirmed by elevated serum levels of WBCs (7.78 ± 0.012 (× 103) /µl), bilirubin (7.56 ± 0.97 mg/dl), Thiobarbituric acid reactive substances (TBARs) (133.75 ± 2.61 nM/min/mg protein), decreased RBCs (4.33 ± 0.065 (× 106)/µl), platelets (344 ± 3.19 (× 103)/µl), total proteins (2.14 ± 0.11 g/dl), Glutathione S-transferases (GST) (81.01 ± 0.44 nM/min/ml), endogenous antioxidant enzymes levels and abnormal liver and kidney functionality in disease control rats. Both species revealed almost identical and significant (p < 0.05) alleviative effects in benzene induced leukemia. CONCLUSION: Comprehensive screening divulged the tremendous potential of selected species as potent source of natural anticancer agents in a variety of cancers particularly leukemia. Present study might provide useful finger prints in cancer research and mechanistic studies are prerequisite in logical hunt of this goal.

Probing the physio-chemical appraisal of green synthesized PbO nanoparticles in PbO-PVC nanocomposite polymer membranes.

Different plants can be used to prepare nanoparticles. This is termed as green technology. It is one of the best ecofriendly and low-cost method for the preparation of nanoparticles which has no harmful effects. PbO nanoparticles were prepared by green method using leaf extract of Datura Sternum plants. The preparation of Lead oxide was confirmed by color change from colorless to yellowish brown. UV-Visible peak obtained at 250 nm and XRD study clarified the formation of PbO NPs. These PbO nanoparticles were then applied for the preparation of Nano Composite Polymer Membranes (nCPMs). PbO-PVC nCPMs were prepared based on polyvinyl chloride (PVC) polymer and PbO filler with the help of solution casting method, using cyclohexanone as a solvent. Different percentage (5-35%) of filler was used. The physiochemical parameters studied were viscosity, water uptake (WU), perpendicular swelling (DT) in deionized water, density, porosity (ε), morphology, ion adsorption capacity (IAC) and electrical conductivity (σ). The values of all these parameters except viscosity and conductivity were increased on increasing filler percentage. Viscosity of the nCPMs solution was decreased from 171 to 46.21. The conductivity of nCPMs was first increased upto 25% filler and then decreased. The deformation in PVC structure was increased on enhancing PbO amount. The values of Density, porosity, water uptake, DT and IAC were found in range 1.15-5.02, 0.50-0.87, 72.01-141.30, 0.012-0.11, and 3.13 × 107-8.60 × 107 respectively.

Datura suaveolens and Verbena tenuisecta mediated silver nanoparticles, their photodynamic cytotoxic and antimicrobial evaluation.

Biogenic production of nanoparticles is eco-friendly, less expensive method with various medical and biological applications. Nanotechnology along with photodynamic therapy is gaining tremendous importance with enhanced efficacy. The present work was aimed to evaluate methanolic extracts and nanoparticles of two selected plants (Datura suavolens and Verbina tenuisecta) for cytotoxic photodynamic, antioxidant and antimicrobial study. Both extract and silver (5 mM) nanoparticles of Datura plant showed significant activities against bacterial strains. Maximum ZOI of 27.3 ± 1.6 mm was observed with nanoparticles of Datura branches with minimum inhibitory (MIC) value of 32 µg/ml. In case of antifungal and antioxidant assay samples were moderately active. Silver nanoparticles and extracts were effective against rhabdomyosarcoma cell line with lowest IC50 value of 42.5 ± 0.6 µg/ml and percent viability of 25.6 ± 1.3 of Verbena tenuisecta. However, nanoparticles of Datura leaves and branches were more potent with IC50 value of 2.4 ± 0.9 µg/ml and 7.8 ± 1.1 µg/ml respectively. The result of photodynamic study showed that efficacy of photosensitizer was enhanced and percent viability reduced when nanoparticles used as an adjunct. The color change and UV spectra (415‒425 nm) indicated the production of nanoparticles. Fourier transform infrared spectroscopy (FTIR) spectra showed presence of different functional groups e.g., hydroxyl, carbonyl and amino. Nanoparticles are sphenoid in morphology and size ranges between 20-150 nm. Current study showed these silver nanoparticles can be used as cytotoxic agent in photodynamic therapy and can play a critical role to establish medicinal potential of selected plants.

In vitro cytotoxicity of polyphenols from Datura innoxia aqueous leaf-extract on human leukemia K562 cells: DNA and nuclear proteins as targets.

Leukemia is a malignant hematological disease and chemotherapy remains the most important tool for its treatment. As chemotherapy has many side effects and could lead to resistance in cancer cells, plant-based medication is being considered as a new strategy in cancer treatment. Datura innoxia from the Solanaceae family is used in traditional medicine. The present study investigated the effect of an aqueous extract of D. innoxia aqueous leaf-extract on human chronic myeloid leukemia cells (K562 cell line) and human B lymphoblastoid cells (FS-2 cells) as the noncancerous cell line. The interaction of the D. innoxia extract with double-stranded DNA and histones was studied using multiple spectroscopic techniques. The total phenolic and flavonoid contents were determined through colorimetric analysis and the major polyphenols were quantified by HPLC-DAD analysis. The results demonstrated that the D. innoxia extract inhibited proliferation of the K562 cell line in a dose- and time-dependent manner (IC50 = 0.6 mg/ml), but had a slightly toxic effect on human B lymphoblastoid cells. The spectroscopy results suggest that the D. innoxia extract interacted with both DNA and histones in solution and that D. innoxia could be suggested as an anticancer drug.

Natural Bioactive New Withanolides.

Withanolides are a group of highly oxygenated steroids derived from a C28 ergostane skeleton, and have attracted significant scientific interest due to their complex structural features and multiple bioactivities. More than 170 new natural withanolides were isolated and identified in the last 5 years. This review provides a comprehensive summary of the structural, biological and pharmacological activities of these new compounds.

A novel phytosterol isolated from Datura inoxia, RinoxiaB is a potential cure colon cancer agent by targeting BAX/Bcl2 pathway.

Plant sterols have been widely used as chemotherapeutic agents for colorectal cancer for years together. In this study, a novel phytosterol was isolated and characterized from the leaf extract of a medicinal plant, Datura inoxia and was coined as RinoxiaB (RB). This phytosterol was observed to have antiproliferative activity against human colon adenocarcinoma cells, HCT 15. The cell viability assay revealed the IC50 value of the RB as 4 µM. Moreover, RB treated cells showed prominent morphological changes dose dependently and progressively increased the number of dead cells. Additionally, results of the comet, flow cytometry, and cell cycle analysis revealed that the majority of cells were arrested in their S and G2/M phase by blocking the mitotic spindle formation. The western blot analysis (Bcl-2, BAX, Cytochrome C, Caspases 9 & 3) clearly indicated that RB has the ability to induce apoptosis by significantly upregulating (P < 0.05) Bcl-2 and causing mitochondrial damage leading to Cytochrome C release and activation of caspases, which subsequently results in downregulation of BAX expression in the cytosol. Furthermore, the expression of tumor suppressors (p53 and p21) and cell cycle regulatory proteins (Cyclins D1 & B1) suggested that RB inhibit cell proliferation. Thus, the present finding concludes that RB can offer possible apoptotic effects by targeting BAX/Bcl2 pathway in HCT 15 cells, thus alleviating colon cancer.

Utilization of an Electrochemiluminescence Sensor for Atropine Determination in Complex Matrices.

A major challenge within forensic science is the development of accurate and robust methodologies that can be utilized on-site for detection at crime scenes and can be used for analyzing multiple sample types. The recent expansion of electrochemical sensors to tackle this hurdle requires sensors that can undergo analysis without any pretreatment. Given the vast array of samples that are submitted for forensic analysis, this can pose a major challenge for all electrochemical sensors, including electrochemiluminescent (ECL)-based sensors. Within this contribution, we demonstrate the capacity for an ECL-based sensor to address this challenge and it is potential to detect and quantify atropine from a wide range of samples directly from herbal material to spiked solutions. This portable platform demonstrates satisfactory analytical parameters with linearity across a concentration range of 0.75 to 100 µM, reproducibility of 3.0%, repeatability of 9.2%, and a detection limit of ∼0.75 µM. The sensor displays good selectivity toward alkaloid species and, in particular, the hallucinogenic tropane alkaloid functionality within complex matrices. This portable sensor provides rapid detection alongside low cost and operational simplicity, thus, providing a basis for the exploitation of ECL-based sensors within the forensic arena.

Antioxidant Potential, Phytochemicals Composition, and Metal Contents of Datura alba.

This study investigated the phytochemical characteristics and antioxidant activity in leaves, roots, stem, flower, and seed parts of Datura alba (D. alba). The study also assessed the heavy metal (Cr, Mn, Zn, and Cu) accumulation in each part of the plant. Among the phytochemicals, alkaloids were found only in leaves while tannins, flavonoids, and phenols were present in all parts of the plant. For antioxidant activity, free radical scavenging assay for 2,2-diphenyl-1-picrylhydrazyl (DPPH) was performed using ascorbic acid as the standard. Higher activity was shown by stem extract in methanol and leaf extract in n-hexane, ethyl acetate, and chloroform. Furthermore, all the target heavy metals were detected in all plant sections with the highest concentration of Zn in leaves and Cu in stem, root, flower, and seed. Due to stronger antioxidant potential and phytochemical composition, D. alba could prove as valuable prospect in pharmaceutical formulations by taking part in the antioxidant defense system against generation of free radicals.

Comprehensive Analysis of Phytochemical Constituents and Ethnopharmacological Investigation of Genus Datura.

The genus Datura comprises wild shrub plants that belong to the Solanaceae family. Naturally, they possess both medicinal and poisonous properties due to the presence of many biologically active phytochemical constituents. Traditionally, Datura had been used for mystic and religious purposes, as a natural drug to treat asthma, pain, gout, boils, abscesses, and wounds, and as psychoactive infusions and fumitories. Different Datura species exhibit diverse ethnopharmacological activities against different diseases, and many ancient and traditional cultures have used various forms of Datura to treat ailments and to prevent many diseases. In this article, we comprehensively summarize various phytochemical constituents isolated from Datura, their pharmacological properties against different diseases, parts of the plants used as traditional therapeutic agents, regions where they are located, and botanical descriptions of different Datura species. The ethnopharmacological properties of Datura may provide new insights for discovery and development of natural drugs. Further research is needed for the investigation of mechanisms of action and to develop safety profiles of the phytochemical constituents isolated from Datura species.

Valproic acid induces three novel cytotoxic secondary metabolites in Diaporthe sp., an endophytic fungus from Datura inoxia Mill.

Addition of the valproic acid (histone deacetylases inhibitor) to a culture of an endophytic fungus Diaporthe sp. harbored from Datura inoxia significantly altered its secondary metabolic profile and resulted in the isolation of three novel compounds, identified as xylarolide A (1), diportharine A (2) and xylarolide B (3) along with one known compound xylarolide (4). The structures of all the compounds (1-4) were determined by detailed analysis of 1D and 2D NMR spectroscopic data. The relative configurations of compounds 1-3 were determined with the help of NOESY data and comparison of optical rotations with similar compounds with established stereochemistry. All the isolated compounds were screened for antibacterial, antioxidant and cytotoxic activities. Xylarolide A (1) and xylarolide (4) displayed significant growth inhibition of MIAPaCa-2 with an IC50 of 20 and 32 µM respectively and against PC-3 with an IC50 of 14 and 18 µM respectively. Moreover, compound 1 displayed significant DPPH scavenging activity with EC50 of 10.3 µM using ascorbic acid as a positive control.

Identification of Datura Species Involved in a Food-Poisoning Case Using LC-MS/MS and DNA Barcording.

A food-poisoning case due to eating the roots of Datura occurred in Kawasaki City, Japan in 2014. The Datura plant was mistakenly collected instead of burdock in a domestic garden. The roots of these plants are quite similar to each other. We presumed that the specimen was the root of Datura, but it was difficult to classify it only from the morphology. Using LC-MS/MS, we detected atropine and scopolamine from the remaining plant specimen. Therefore, we applied the DNA barcoding method. The results showed that the specimen was classified into Solanaceae family, but not Asteraceae family. Thus, the specimen was confirmed to be Datura species based on both chemical and genetic analyses.