Isosteroidal Alkaloids Research Articles

Isosteroidal Alkaloids from Fritillaria karelinii

Three new isosteroidal alkaloids, karelinine (1), 5-epikarelinine (2), and 27-epiebeienine (3), were isolated from the bulbs of Fritillaria karelinii by column chromatography and semipreparative HPLC (high performance liquid chromatography), together with three known ones, ebeienine (4), persicanidine B (5), and heilonine (6). Their structures were identified by MS, 1D/2D NMR, and single-crystal X-ray diffraction. Compound 1 is a 5 beta-jervine isosteroidal alkaloid featuring a cis-fused A/B ring moiety, rarely found in the Fritillaria genus. Compounds 1 and 2 also represent the first jervine alkaloids with a 15 alpha-hydroxy group from this genus.

Isosteroidal alkaloids from the bulbs of Fritillaria tortifolia

Three new isosteroidal alkaloids, frititorines A-C (1–3), were isolated from the bulbs of Fritillaria tortifolia, together with ten known ones (4–13). Their structures were elucidated by extensive spectroscopic analyses, chemical methods, and single-crystal X-ray crystallographic analysis. Compound 1 is the first 5β-cevanine alkaloid with a cis A/B ring junction from the Fritillaria genus. Compound 2 is the first example of glycosylated isosteroidal alkaloid N-oxide. Compound 1 showed significant relaxant effect on Ach-induced tracheal contraction with pA2 and EC50 values equivalent to those of aminophylline.

Antitumor Activity of Isosteroidal Alkaloids from the Plants in the Genus Veratrum and Fritillaria.

Isosteroidal alkaloids are a category of promising bioactive compounds which mostly exist in plants of genus Veratrum and Fritillaria. The pharmacological activities of isosteroidal alkaloids include antihypertensive, antitussive, anti-inflammatory, antithrombosis, among others. Recently, some studies show that this kind of alkaloids exhibited significant antitumor activity. To the best of our knowledge, there is no review focusing on their antitumor activity and mechanism of their antitumor activity. To fill the gap, in this review, we summarized antitumor effects of the isosteroidal alkaloids from genus Veratrum and Fritillaria on different tumors and the mechanisms of their antitumor activity. In conclusion, this kind of alkaloids has extensive antitumor activity, and there are several main mechanisms of their antitumor activity, including the Hedgehog signaling pathway, caspase-3 dependent apoptosis, cell cycle, and autophagy.

A New Alkaloid with Tracheal Relaxant Effect from the Bulbs of Fritillaria pallidiflora

A new isosteroidal alkaloid, yibeinone E (1), along with five known ones, zhebeinone 3-O-β-D-glucoside (2), sinpeinine A (3), yubeinine (4), ebeinone (5), and delavinone (6), were isolated from the bulbs of Fritillaria pallidiflora. Their structures were confirmed on the basis of extensive spectroscopic analysis or comparison with the data in the literature (known compounds). Compound 3 showed an apparent relaxation effect on isolated tracheas.

Investigation of association of chemical profiles with the tracheobronchial relaxant activity of Chinese medicinal herb Beimu derived from various Fritillaria species

Ethnopharmacological relevanceFritillariae Bulbus (Beimu in Chinese) is derived from the bulbus of many Fritillaria species (family Liliaceae), which has been used as an antitussive herb in traditional Chinese medicine for more than 2000 years. Due to the complexity of plant origins and significant variations in chemical profiles, the characterization of the profile of the major bioactive constituents and its association with pharmacological activity are important for the quality control of Beimu herbs from different origins. Aim of the studyThis study aims to investigate the distribution of major bioactive isosteroidal alkaloids in Beimu herbs of different origins and its correlation with the tracheobronchial relaxant activity. MethodsQuantification of 7 main bioactive 5α-cevanine isosteroidal alkaloids, including ebeiedine, ebeiedinone, hupehenine, isoverticine, verticine, verticinone and imperialine, in 23 Fritillaria species was performed using gas chromatography. The relaxant effect of different extracts of 4 commonly used Beimu herbs, namely Zhe-Beimu (F. thunbergii Miq.), Chuan-Beimu (F. cirrhosa D. Don), Hubei-Beimu (F. hupehensis Hsiao et K. C. Hsia) and Yi-Beimu (F. pallidiflora Schrenk), was evaluated using rat isolated tracheal and bronchial preparations pre-contracted with carbachol, the well established in vitro antitussive model. ResultsAmongst 23 Fritillaria species detected, significant variations of the types and quantities of 7 major isosteroidal alkaloids were determined, which served as an important indicator for the classification of different Beimu herbs with distinct geographic distributions. Based on the type and quantity of these alkaloids, different origins of Beimu could be clearly clustered into several subgroups by principal component analysis. Furthermore, both crude alkaloid and water extracts of all 4 Beimu herbs showed a dose-dependent tracheobronchial relaxation with different potencies. The total content of alkaloids (weight adjusted based on the activity of individual alkaloids) in Beimu extracts significantly correlated with their tracheobronchial relaxation effects (r2 > 0.9, p < 0.001). ConclusionsThe results demonstrated that the differences in chemical profile of major bioactive isosteroidal alkaloids and pharmacological activity of Beimu could be incorporated into a simple and unified method for quality control and potential prediction of activity of Beimu herbs from different origins.

Isosteroidal alkaloids as potent dual-binding site inhibitors of both acetylcholinesterase and butyrylcholinesterase from the bulbs of Fritillaria walujewii

Five new isosteroidal alkaloids, walujewine A (1), walujewine B (4), walujewine C (5), walujewine D (6), walujewine E (10) were isolated from the bulbs of Fritillaria walujewii together with seven known isosteroidal alkaloids (2, 3, 7–9, 11, 12). Their structures were elucidated on the basis of IR, ESI-MS, HR-ESI-MS, 1D and 2D NMR spectroscopic data analyses and single-crystal X-ray diffraction. All the isolates were tested for ChE inhibiting activity by the Ellman's method. Compounds 3–5 and 8–10 were potent dual AChE−BChE inhibitors, and compound 1 showed highly selective AChE inhibition. The structure−activity relationship of compounds 1–12 was discussed in details. And kinetic analysis showed that compounds 1, 3–5, and 8–10 were mixed-type reversible inhibitors of AChE, simultaneously binding to the catalytic and peripheral anionic sites, which was verified by in silico docking studies. The docking simulation also showed that active compound 3 and 8 created many interactions with the CAS and PAS gorges of BChE, revealing their mixed-type inhibition. ADMET analysis further confirmed the therapeutic potential of some isosteroidal alkaloids based on their high BBB-penetration.

ChemInform Abstract: New Isosteroidal Alkaloids with Tracheal Relaxant Effect from the Bulbs of Fritillaria pallidiflora Schrenk.

AbstractFour new isosteroidal alkaloids, yibeinone A (I), yibeinone B (II), yibeinones C (III), and yibeinone D (IV) are isolated from the bulbs of Fritillaria pallidiflora together with four known ones.

A Non-Derivative Method For The Quantitative Analysis Of Isosteroidal Alkaloids From Fritillaria By High Performance Liquid Chromatography Combined With Charged Aerosol Detection

A Non-Derivative Method For The Quantitative Analysis Of Isosteroidal Alkaloids From Fritillaria By High Performance Liquid Chromatography Combined With Charged Aerosol Detection

New isosteroidal alkaloids with tracheal relaxant effect from the bulbs of Fritillaria pallidiflora Schrenk

Four new isosteroidal alkaloids, yibeinones A–D (1–4), along with four known ones, imperialine-3β-d-glucoside (5), imperialine (6), imperialine β-N-oxide (7), and dongbeinine (8), were isolated from the bulbs of Fritillaria pallidiflora. Their structures were confirmed on the basis of extensive spectroscopic analysis or comparison with the data in the literatures (known compounds). Compounds 2, 3, 4, and 6 showed an apparent concentration-dependant relaxation of isolated tracheal preparation, amongst 3 and 6 exhibited significant effects with pA2 values of 6.19±0.02 and 8.41±0.10, and EC50 values of 0.65±0.02μmolL−1 and 4.40±0.90nmolL−1, respectively.

A non-derivative method for the quantitative analysis of isosteroidal alkaloids from Fritillaria by high performance liquid chromatography combined with charged aerosol detection

A non-derivative method was developed for the qualitative and quantitative analysis of isosteroidal alkaloids from Fritillaria thunbergii. During method development the performance of two universal detectors, the charged aerosol detector (CAD) and evaporative light scattering detector (ELSD), were evaluated. The CAD was found to be 30 to 55 times more sensitive than ELSD enabling the measurement of low levels of reference compound impurities that could not be detected by ELSD. The peak area percent of the reference compound, peimisine, obtained by CAD was 50.10%, but 91.66% by ELSD showing that CAD is suitable to estimate the presence of impurities. The CAD showed good reproducibility with overall intra- and inter-day peak area RSD values of less than 1.8% and 2.7%, respectively and had a linear dynamic range of up to 4 orders of magnitude (0.06–44mg/L) for peimine and peiminine. The optimized method was used for the quantitative analysis of peimine and peiminine from F. thunbergii.

Characterization of the Isosteroidal Alkaloid Chuanbeinone from Bulbus of Fritillaria pallidiflora as Novel Antitumor Agent In Vitro and In Vivo.

Bulbus of Fritillaria pallidiflora, the dried bulb of F. pallidiflora, is widely used in Chinese folk medicine due to its powerful biological activities. The aim of this study was to investigate in vitro and in vivo antitumor activity of different fractions and isosteroidal alkaloids from bulbus of F. pallidiflora and clarify its putative mechanism of antitumor activity. Firstly, we assayed in vitro antitumor effects of different fractions from bulbus of F. pallidiflora and found that chloroform extracts and purified total alkaloids of bulbus of F. pallidiflora showed higher cytotoxic activity than other tested extracts. We further isolated four main alkaloids, chuanbeinone, imperialine-β-N-oxide, isoverticine and isoverticine-β-N-oxide, from the total alkaloids of bulbus of F. pallidiflora and found that they display significant cytotoxicity, whereby chuanbeinone showed the highest activity against Lewis lung carcinoma cells. Moreover, we found that chuanbeinone induced S phase arrest and further increased apoptosis of Lewis lung carcinoma cells. The results of Western blotting experiments showed that the expression of the antiapoptotic Bcl-2 was reduced by chuanbeinone treatment, while the proapoptotic protein Bax and caspase-3 were increased. Moreover, we investigated the in vivo antitumor activity of chuanbeinone and characterized its putative antitumor mechanism of action by the TUNEL assay and by histological and immunohistochemical analyses. Our results showed that chuanbeinone exhibited significant antitumor activity in vivo, while notably inhibiting tumor angiogenesis and inducing apoptosis characterized by an increased expression of caspase-3. Our findings show that chuanbeinone exhibits significant antitumor activity in vitro and in vivo, and points to possible therapeutic potential for this compound as well as for its natural source, bulbus of F. pallidiflora.

Phytochemical profiling of medicinal isosteroidal alkaloids of Iranian Fritillaria spp. (Liliaceae)

The genus Fritillaria (Liliaceae) embraces up to 165 species, including taxa of both medicinal and ornamental importance. In this study, 44 specimens of the genus representing 9 species were collected from their natural habitats located in 10 provinces of Iran. For the purpose of finding phytochemical diversity and a possible discriminant method among Fritillaria species, five major isosteroidal alkaloids (ISAs) of the genus (imperialine, verticine, verticinone, ebeiedine and ebeiedinone) were identified and characterized by gas chromatography (GC) and gas chromatography with mass spectrometry (GC–MS) methods. The estimated relative composition of ISAs was used to carry out a chemotaxonomic study on the species by means of hierarchical cluster analysis. Trimethyl silylimidazole (TMSI) was found to be an appropriate reagent for pre-column derivatization of the ISAs. The number of compounds detected in each specimen varied significantly, but imperialine, verticine, verticinone, ebeiedine and ebeiedinone were identified in nearly all samples. Verticine was the predominant alkaloid in most of the species. The specimens representing F. gibbosa, in which none of the examined ISAs were detected, showed a different phytochemical profile compared with the others, whereas the highest average of total contribution percentage was recorded in F. imperialis. Although, the qualitative and quantitative assessment of the five ISAs reveals a high level of phytochemical diversity within the genus, it is not sufficient to accurately distinguish the species and specimens within each species. Accordingly, evaluation of total ISAs profile of the species and/or specific alkaloids assigned to a certain species is believed to be required to find phytochemical affinities amongst Fritillaria species. This is the first study on phytochemistry of Fritillaria spp. in Iran.

Simultaneous Determination of Four Main Isosteroidal Alkaloids of Bulbus Fritillariae Cirrhosae in Rat Plasma by LC–MS–MS

A simple, sensitive and selective liquid chromatography–tandem mass spectrometry (LC–MS–MS) method was developed for simultaneous quantitation of four isosteroidal alkaloids (peimine, peiminine, peimisine, and delavine) in rat plasma. Carbamazepine was selected as internal standard (IS). The LC–MS–MS system coupled with an electrospray ionization (ESI) source was performed in the multiple reaction monitoring (MRM) mode. The transitions of the isosteroidal alkaloids executed as following: m/z 432.4 → 414.4 for peimine, m/z 430.5 → 412.4 for peiminine, m/z 428.4 → 114.2 for peimisine, m/z 416.5 → 105.2 for delavine, and m/z 237.2 → 194.2 for carbamazepine (IS). Sample preparation was performed with liquid–liquid extraction using ethyl acetate after carbamazepine (IS) spiked. The separation was performed on a Welch C18 column (3.5 μm, 2.1 × 100 mm), and a gradient elution of methanol and 5 mmol L−1 ammonium acetate in 0.1 % formic acid aqueous solution was used. The retention time was less than 8.0 min. Linearity was obtained over the concentration range of 0.2–200 ng mL−1 for peimine and peiminine, and 1.0–200 ng mL−1 for peimisine and delavine. The method was linear for all analytes with correlation coefficients >0.995. The intra-day and inter-day accuracy and precision of the assay were acceptable. This method has been successfully applied to the pharmacokinetic study of Bulbus Fritillariae Cirrhosae extract after oral administration to rats.

Phytochemical Variation in Fritillaria cirrhosa D. Don (Chuan Bei Mu) in Relation to Plant Reproductive Stage and Timing of Harvest1

Phytochemical Variation inFritillaria cirrhosaD. Don (Chuan Bei Mu) in Relation to Plant Reproductive Stage and Timing of Harvest. Economic development in southwest China and the increasing use of traditional Chinese medicine (TCM) worldwide have led to intensified collection of native medicinal plants. Fritillaria cirrhosa D. Don (Chuan Bei Mu), commonly used for the treatment of cough in TCM, is endemic to the Hengduan Mountain region of southwest China and is under increasing pressure from over-collection and decreasing suitable alpine habitat. The bioactive compounds in F. cirrhosa bulbs, isosteroidal alkaloids, are greatly influenced by environmental conditions and fluctuate in content and concentration with plant age and reproductive stage. Aiming at obtaining useful information for the sustainable management of wild F. cirrhosa populations, we evaluated how the phytochemical composition of F. cirrhosa bulbs varies at various stages of plant reproductive development. Using chemical methods and high-performance liquid chromatography, we extracted and analyzed two major bioactive alkaloids from F. cirrhosa bulb samples collected throughout the Hengduan Mountain region. Plant reproductive stage was found to affect the concentration of bioactive alkaloids in F. cirrhosa bulbs. Bulb alkaloid concentration was highest during the early stages of fruit development and decreased significantly with fruit maturation. These results lend biochemical support to the practice of harvesting F. cirrhosa during the early stages of plant senescence (i.e., early fruit development).

Antinociceptive Efficacy of Verticinone in Murine Models of Inflammatory Pain and Paclitaxel Induced Neuropathic Pain

Verticinone, an isosteroidal alkaloid separated from Bulbus Fritillaria (Chinese name "Bei-mu"), was evaluated for its analgesic activities in murine models of inflammatory and neuropathic pain. It was shown that oral administarion of verticinone could significantly inhibit acetic acid-induced writhing response in a dose-dependent way, and the writhing inhibition of 3 mg/kg verticinone was 66.2%, which was approximately higher than that of 200 mg/kg aspirin. In the formalin test, a high dose of (3 mg/kg) verticinone could inhibit the nociceptive response of both phases, but the lower dose (1.5 mg/kg) could only inhibit the second phase response, which suggested that verticinone might exert its analgesic effect through both central and peripheral mechanisms. In addition, in formalin and acetic acid tests, the spontaneous locomotive activities of the mice treated with verticinone were transiently greatly decreased when compared with the vehicle group. In the rat model of paclitaxel induced neuropathic pain, in contrast to the declined analgesic effect of morphine after repeated administration with the same dose, a relatively constant analgesic effect of verticinone was observed. These investigations suggested that verticinone could exert a good antinociceptive effect on inflammatory pain and cancer-related neuropathic pain probably through both peripheral and central mechanisms, and it might be partly involved with some sedation effects. Verticinone is expected to become a potentially novel sedative-analgesic agent without producing tolerance and dependence, but further studies are still urgently needed to elucidate the precise mechanisms and activities of it.

Microwave-Promoted Rearrangement of Hecogenin Derivatives into Cnor- D-homo Steroids

By means of microwave irradiation, the rearrangement from 12β-tosylate and 12β-alcohol of hecogenin derivatives into C-nor-D-homo-Δ17a(18)-olefin (2) and the Δ13(17a)-isomer (3) was achieved. With this method, the controlled solvolysis in anhydrous pyridine gave total conversion to desired exocyclic-olefin which is expected to be a building block of isosteroidal alkaloids. Keywords: Hecogenin derivatives, rearrangement, solvolysis, C-nor-D-homo, isosteroids, microwave irradiation

Differentiation-inducing effects of verticinone, an isosteroidal alkaloid isolated from the bulbus of Fritillaria ussuriensis, on human promyelocytic leukemia HL-60 cells.

The inducer of differentiation of human promyelocytic leukemia HL-60 cells is commonly accepted to have potential therapeutic importance. Verticinone, one of the major isosteroidal alkaloids from the bulbus of Fritillaria ussuriensis, was found to inhibit the growth of HL-60 cells by inducing these cells to differentiate toward granulocytes. Importantly, the combination of verticinone with all-trans retinoic acid (ATRA), a well-known inducer of HL-60 cells into granulocytic lineages, was more effective than either alone, suggesting its therapeutic use in minimizing the effective dose of ATRA.

Determination of the major isosteroidal alkaloids in bulbs of Fritillaria by high-performance liquid chromatography coupled with evaporative light scattering detection

A new direct HPLC analytical method using evaporative light scattering detection coupled with a low-temperature adapter for the simultaneous determination of the major biologically active isosteroidal alkaloids in Bulbus Fritillariae, a commonly used antitussive traditional Chinese medicinal (TCM) herb, has been developed. The simultaneous separation of eight Fritillaria alkaloids was achieved on a reversed-phase C 8 column with an isocratic mobile phase system consisting of acetonitrile–methanol–water (66.5:3.5:30, v/v) containing 0.006% triethylamine. This method provides good reproducibility and sensitivity for the quantification of six major isosteroidal alkaloids, namely peimissine, verticine, verticinone, imperialine, isoverticine and ebeiedine in different Fritillaria species with overall intra- and inter-day precision and accuracy of less than 11% and higher than 90%, respectively. The assay was successfully utilized to quantify the major biologically active alkaloids in five Fritillaria species. The results demonstrate that this method is simple, selective, and suitable for the quality control of this commonly used antitussive TCM herb, Bulbus Fritillariae.

Simultaneous determination of seven major isosteroidal alkaloids in bulbs of Fritillaria by gas chromatography

The present paper describes the development of a most simple, sensitive, and specific gas chromatographic method to date, for the direct determination of seven major bioactive isosteroidal alkaloids, namely ebeiedine, ebeiedinone, ebeienine, hupehenine, isoverticine, verticine, verticinone and imperialine, in Fritillaria species, a commonly used antitussive traditional Chinese medicinal (TCM) herb. In the present study, a commercially available Supelco SAC-5 capillary column (30 m×0.25 mm, 0.25 μm) specifically designed for the analysis of steroids was utilized for the direct determination of Fritillaria alkaloids. Calibration curves were obtained by spiking authentic compounds and the internal standard (solanidine) into herbal samples prior to extraction. Extraction was conducted simply by shaking the pre-alkalized diethyl ether solution (5.0 ml) containing dried herb (0.1 g) for 2 h. All calibration curves showed good linear regressions ( r 2>0.995) within test ranges. The assay was reproducible and accurate with the overall intra- and inter-day variation and accuracy of less than 10% and more than 90%, respectively. The developed GC method was successfully utilized to analyze seven major bioactive alkaloids in seven Fritillaria species, and the results demonstrate that this direct GC analytical method is suitable for the quality control of this commonly used antitussive TCM herb.