Diterpene Alkaloids Research Articles

Functional identification of the terpene synthase family involved in diterpenoid alkaloids biosynthesis in Aconitum carmichaelii

Aconitum carmichaelii is a high-value medicinal herb widely used across China, Japan, and other Asian countries. Aconitine-type diterpene alkaloids (DAs) are the characteristic compounds in Aconitum. Although six transcriptomes, based on short-read next generation sequencing technology, have been reported from the Aconitum species, the terpene synthase (TPS) corresponding to DAs biosynthesis remains unidentified. We apply a combination of Pacbio isoform sequencing and RNA sequencing to provide a comprehensive view of the A. carmichaelii transcriptome. Nineteen TPSs and five alternative splicing isoforms belonging to TPS-b, TPS-c, and TPS-e/f subfamilies were identified. In vitro enzyme reaction analysis functional identified two sesqui-TPSs and twelve diTPSs. Seven of the TPS-c subfamily genes reacted with GGPP to produce the intermediate ent-copalyl diphosphate. Five AcKSLs separately reacted with ent-CPP to produce ent-kaurene, ent-atiserene, and ent-13-epi-sandaracopimaradie: a new diterpene found in Aconitum. AcTPSs gene expression in conjunction DAs content analysis in different tissues validated that ent-CPP is the sole precursor to all DAs biosynthesis, with AcKSL1, AcKSL2s and AcKSL3-1 responsible for C20 atisine and napelline type DAs biosynthesis, respectively. These data clarified the molecular basis for the C20-DAs biosynthetic pathway in A. carmichaelii and pave the way for further exploration of C19-DAs biosynthesis in the Aconitum species.

Mechanism of action of a diterpene alkaloid hypaconitine on cytotoxicity and inhibitory effect of BAPTA-AM in HCN-2 neuronal cells.

Hypaconitine, a neuromuscular blocker, is a diterpene alkaloid found in the root of Aconitum carmichaelii. Although hypaconitine was shown to affect various physiological responses in neurological models, the effect of hypaconitine on cell viability and the mechanism of its action of Ca2+ handling is elusive in cortical neurons. This study examined whether hypaconitine altered viability and Ca2+ signalling in HCN-2 neuronal cell lines. Cell viability was measured by the cell proliferation reagent (WST-1). Cytosolic Ca2+ concentrations [Ca2+ ]i was measured by the Ca2+ -sensitive fluorescent dye fura-2. In HCN-2 cells, hypaconitine (10-50μmol/L) induced cytotoxicity and [Ca2+ ]i rises in a concentration-dependent manner. Removal of extracellular Ca2+ partially reduced the hypaconitine's effect on [Ca2+ ]i rises. Furthermore, chelation of cytosolic Ca2+ with BAPTA-AM reduced hypaconitine's cytotoxicity. In Ca2+ -containing medium, hypaconitine-induced Ca2+ entry was inhibited by modulators (2-APB and SKF96365) of store-operated Ca2+ channels and a protein kinase C (PKC) inhibitor (GF109203X). Hypaconitine induced Mn2+ influx indirectly suggesting that hypaconitine evoked Ca2+ entry. In Ca2+ -free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished hypaconitine-induced [Ca2+ ]i rises. Conversely, treatment with hypaconitine inhibited thapsigargin-induced [Ca2+ ]i rises. However, inhibition of phospholipase C (PLC) with U73122 did not inhibit hypaconitine-induced [Ca2+ ]i rises. Together, hypaconitine caused cytotoxicity that was linked to preceding [Ca2+ ]i rises by Ca2+ influx via store-operated Ca2+ entry involved PKC regulation and evoking PLC-independent Ca2+ release from the endoplasmic reticulum. Because BAPTA-AM loading only partially reversed hypaconitine-induced cell death, it suggests that hypaconitine induced a second Ca2+ -independent cytotoxicity in HCN-2 cells.

Anticancer Activity of Delphinium semibarbatum Alkaloid Fractions against LNCaP, and DU 145 Human Prostate Cancer Cells through the Intrinsic Apoptotic Pathway.

Prostate cancer is one of the common cancers with a high mortality rate in men. Therefore, there is always a necessity to discover new medications for treatment or alleviating its symptoms. In recent years, anticancer properties of a number of delphinium species were reported, but there is no study on the anticancer effects of Delphinium semibarbatum (D. semibarbatum) alkaloid contents. Therefore, this survey aimed to check the cytotoxicity and apoptotic properties of D. semibarbatum alkaloid fractions (DSAFs) against prostate cancer cells. Cytotoxicity was measured by MTT assay. We examined the apoptosis by detecting annexin V-FITC/PI staining, the mitochondrial membrane potential (ΔΨm) disruption, reactive oxygen species (ROS) generation, the activity of caspase-3, and expression of the Bax and Bcl-2 in cancer cells. DSAFs treatment inhibited the growth of LNCaP and DU‐145 cells by the increase of apoptotic (Q2+Q3) cells detected by annexin V/PI assay. We found over-generation of intracellular ROS and ΔΨm loss in mitochondrial membrane potential treated cell lines. Attenuation of anti-apoptotic Bcl-2 followed by the increase in pro-apoptotic Bax bands, as well as activation of the caspase-3 enzyme was shown in Western blot analysis. Phytochemical analysis suggested that hetisine type diterpene alkaloids were probably responsible for apoptotic activities. Conclusively, the present study demonstrated that D. semibarbatum alkaloid content exerted antiproliferative effects against prostate cancer cells by inducing the intrinsic pathway of apoptosis.

Hybrides of Alkaloid Lappaconitine with Pyrimidine Motif on the Anthranilic Acid Moiety: Design, Synthesis, and Investigation of Antinociceptive Potency.

Convenient and efficient routes to construct hybrid molecules containing diterpene alkaloid lappaconitine and pyrimidine fragments are reported. One route takes place via first converting of lappaconitine to 1-ethynyl-lappaconitine, followed by the Sonogashira cross-coupling-cyclocondensation sequences. The other involves the palladium-catalyzed carbonylative Sonogashira reaction of 5′-iodolappaconitine with aryl acetylene and Mo (CO)6 as the CO source in acetonitrile and subsequent cyclocondensation reaction of the generated alkynone with amidines. The reaction proceeded cleanly in the presence of the PdCl2-(1-Ad)2PBn∙HBr catalytic system. The protocol provides mild reaction conditions, high yields, and high atom and step-economy. Pharmacological screening of lappaconitine-pyrimidine hybrids for antinociceptive activity in vivo revealed that these compounds possessed high activity in experimental pain models, which was dependent on the nature of the substituent in the 2 and 6 positions of the pyrimidine nucleus. Docking studies were undertaken to gain insight into the possible binding mode of these compounds with the voltage-gated sodium channel 1.7. The moderate toxicity of the leading compound 12 (50% lethal dose (LD50) value was more than 600 mg/kg in vivo) and cytotoxicity to cancer cell lines in vitro encouraged the further design of therapeutically relevant analogues based on this novel type of lappaconitine–pyrimidine hybrids.

Chemical constituents in different parts of seven species of Aconitum based on UHPLC-Q-TOF/MS

Aconitum L., the main source of Aconitum medicinal materials, is rich in diterpenoid alkaloids. Several drugs derived from diterpenoid alkaloids are widely used to the current clinical treatment of pain, inflammation, and other symptoms. This paper aims to clarify the main metabolites and distribution of diterpenoid alkaloids in different parts of Aconitum plants. To that end, 7 species of Aconitum from three subgenera were analyzed by UHPLC-Q-TOF-MS under identical conditions. The fragmentation regularity of various types of diterpene alkaloids were determined and a total of 126 metabolites were identified by comparing the reference material and secondary mass spectrometry, with the literature. 67, 49, 17, 41, 14, 17 and 21 metabolites were identified from Aconitum carmichaeli, Aconitum stylosum, Aconitum sinomontanum, Aconitum vilmorinianum, Aconitum pendulum, Aconitum tanguticum and Aconitum gymnandrum, respectively. Meanwhile, the structure type of A. carmichaeli, A. stylosum, A. vilmorinianum, A. pendulum, A. gymnandrum were identified as C19 type, A. sinomontanum was C18 type, while A. tanguticum was C20 type. A high similarity of metabolites was found between A. stylosum and A. vilmorinianum. The quantitative analysis of 19 compounds and the relative peak area of all metabolites which obtained through internal standard berberine, highlighted compounds like karakoline, talatisamine and atisine as references for future study of metabolic pathways. Furthermore, results from metabolites distribution and relative peak area analysis suggest that the leaf of A. carmichaeli, the leaf and stem of A. stylosum and A. vilmorinianum, and the flower of A. pendulum have potential as medicinal resources and are worth further development. These results establish a foundation for the comprehensive utilization of Aconitum resources.

A UHPLC-QTOF-MS screening provides new insights into the phytochemical composition and biological properties of six Consolida species from Turkey

The genus Consolida is one of the most important within the Ranunculaceae family and several members of this genus contain important biologically active compounds, such as phenolics and alkaloids. The present study attempts for the first time to assess the biological properties and phytochemical constituents of six Consolida species (C. glandulosa (Boiss. & A. Huet) Bornm, C. hellospontica (Boiss.) Chater, C. raveyi (Boiss.) Schrödinger, C. regalis (Boiss.) Schrödinger, C. staminosa P.H. Davis & Sorger and C. stenocarpa (P.H. Davis & M. Hossain) P.H. Davis) growing in Turkey. A comprehensive phytochemical profiling of the different species was achieved by using an ultra-high-performance liquid chromatography-quadrupole time-of-flight (UHPLC-QTOF) mass spectrometry, targeting polyphenols and diterpene alkaloids. Also, the in vitro antioxidant assays (2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS), ferric reducing antioxidant power (FRAP), cupric reducing antioxidant capacity (CUPRAC), metal chelating and phosphomolybdenum) and enzyme inhibition (cholinesterases, tyrosinase, α-amylase, and α-glucosidase) potentials of the different Consolida methanolic extracts were evaluated. The UHPLC-QTOF profiling revealed 492 compounds, being 236 flavonoids, 93 phenolic acids, 78 tyrosol derivatives, 49 diterpene alkaloids, 29 lignans, and 7 stilbenes. The cumulative phenolic content ranged from 17.65 mg Eq./g for C. staminosa up to 43.04 mg Eq./g for C. glandulosa, whilst the total diterpene alkaloids content was exclusively found in C. glandulosa extracts (3.53 mg Eq./g). Consolida regalis (58. 61 mg trolox equivalent (TE)/g for DPPH and 65.38 mg TE/g for ABTS) and C. stenocarpa (46.81 mg TE/g for DPPH and 78.12 mg TE/g for ABTS) exhibited the highest anti-radical abilities, while the best reducing power values were recorded for C. glandulosa (129.46 mg TE/g for CUPRAC and 77.01 mg TE/g for FRAP) and C. raveyi (124.56 mg TE/g for CUPRAC and 78.36 mg TE/g for FRAP). Also, following the cholinesterases inhibition assays, C. hellospontica and C. glandulosa exhibited the highest inhibitory effects. Regarding tyrosinase inhibitory effects, C. raveyi showed the highest value, being 126.60 mg kojic acid equivalent (KAE)/g, followed by C. glandulosa (125.82 mg KAE/g) and C. stenocarpa (123.09 mg KAE/g). Besides, Consolida staminosa showed potential inhibition against α-amylase, but low glucosidase inhibitory properties. Therefore, the present findings could provide a starting point for further investigations to promote the industrial exploitation of these species with the aim of designing novel phyto-pharmaceuticals.

Therapeutic Potential of (-)-Agelamide D, a Diterpene Alkaloid from the Marine Sponge Agelas sp., as a Natural Radiosensitizer in Hepatocellular Carcinoma Models.

Radiation therapy (RT) is an effective local treatment for unresectable hepatocellular carcinoma (HCC), but there are currently no predictive biomarkers to guide treatment decision for RT or adjuvant systemic drugs to be combined with RT for HCC patients. Previously, we reported that extracts of the marine sponge Agelas sp. may contain a natural radiosensitizer for HCC treatment. In this study, we isolated (−)-agelamide D from Agelas extract and investigated the mechanism underlying its radiosensitization. (−)-Agelamide D enhanced radiation sensitivity of Hep3B cells with decreased clonogenic survival and increased apoptotic cell death. Furthermore, (−)-agelamide D increased the expression of protein kinase RNA-like endoplasmic reticulum kinase/inositol-requiring enzyme 1α/activating transcription factor 4 (PERK/eIF2α/ATF4), a key pathway of the unfolded protein response (UPR) in multiple HCC cell lines, and augmented radiation-induced UPR signaling. In vivo xenograft experiments confirmed that (−)-agelamide D enhanced tumor growth inhibition by radiation without systemic toxicity. Immunohistochemistry results showed that (−)-agelamide D further increased radiation-induced ATF4 expression and apoptotic cell death, which was consistent with our in vitro finding. Collectively, our results provide preclinical evidence that the use of UPR inducers such as (−)-agelamide D may enhance the efficacy of RT in HCC management.

Olive Oil Lignan (+)-Acetoxypinoresinol Peripheral Motor and Neuronal Protection against the Tremorgenic Mycotoxin Penitrem A Toxicity via STAT1 Pathway.

Penitrem A, PA, is an indole diterpene alkaloid produced by several fungal species. PA acts as a selective Ca2+-dependent K-channels (Maxi-K, BK) antagonist in brain, causing motor system dysfunctions including tremors and seizures. However, its molecular mechanism at the peripheral nervous system (PNS) is still ambiguous. The Mediterranean diet key ingredient extra-virgin olive oil (EVOO) provides a variety of minor bioactive phenolics. (+)-Pinoresinol (PN) and (+)-1-acetoxypinoresinol (AC) are naturally occurring lignans in EVOO with diverse biological activities. AC exclusively occurs in EVOO, unlike PN, which occurs in several plants. Results suggest that PA neurotoxicity molecular mechanism is mediated, in part, through distortion of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway. PA selectively activated the STAT1 pathway, independently of the interferon-γ (IFN-γ) pathway, in vitro in Schwann cells and in vivo in Swiss albino mice sciatic nerves. Preliminary in vitro screening of an EVOO phenolic compounds library for the ability to reverse PA toxicity on Schwann cells revealed PN and AC as potential hits. In a Swiss albino mouse model, AC significantly minimized the fatality after intraperitoneal administration of PA fatal doses and normalized most biochemical factors by modulating the STAT1 expression. The olive lignan AC is a novel lead that can prevent the neurotoxicity of food-contaminating tremorgenic indole alkaloid mycotoxins.

Antiarrhythmic agents based on diterpenoid alkaloid lappaconitine. Protonation of N-deacethyllappaconitine in methanol solutions

The data on antiarrhythmic drugs based on diterpene alkaloid lappaconitine are summarized. N-Deacetyllappaconitine is the main metabolite of allapinin and a potential antiarrhythmics for intravenous use. The basicity of two nitrogen atoms of this compound and the possibility of obtaining mono- and di-salts with the example of hydrochlorides and hydrobromides having good solubility in water have been studied. It was shown using NMR spectroscopy and potentiometric titration that in the methanolic solutions the atom N(20) is protonated at the first step (pKb1 = 6.77, 25 °C) in the pH range of 6—7. The nitrogen of the primary aromatic amino group is protonated in the pH range of 2—3 (pKb2 = 2.18, 25 °C).

The Cytotoxicity and Insecticidal Activity of Extracts from Delphinium formosum Boiss. & Huet

DOI : 10.26650/IstanbulJPharm.2019.19034 Delphinium species are well-known toxic plants with their diterpenoid alkaloid contents. There has been no previous investigation on the cytotoxicity of Delphinium formosum Boiss. & Huet. The extracts of the different parts of D. formosum, an endemic species in Turkey, were investigated for their cytotoxic activity against the human liver carcinoma cell line (HepG2) and primary human umbilical vein endothelial cells (HUVEC). The cytotoxic effects of twelve extracts and subfractions were determined against HepG2 cells using MTT assay. The only active extract was applied to the HUVEC as a model for healthy cells. Only one of the alkaloid-containing extracts from the aerial parts was toxic (IC50 = 244,9 μg/mL against HepG2 and 144,4 μg/mL against HUVEC), while the root extracts were inactive. The results were improbable although it’s predicted secondary metabolites, such as diterpene alkaloids (methyllycaconitine, browniine, lycoctonine, avardharidine, antranoyllycoctonine, delsemine A/B and lycoctonine). Based on previous studies in literature, the cytotoxic plants were also expected to exhibit insecticidal activity. Therefore, the cytotoxic extract of D. formosum was examined for its adulticidal and larvicidal activity against the yellow fever, dengue fevers and Zika virus vector Aedes aegypti L. You may cite this article as : Sen-Utsukarci B, Tabanca N, Estep AS, Akbal-Dagistan O, Kessler SM, Ozturk Z, Becnel J, Kiemer AK, Mat A (2019). The Cytotoxicity and Insecticidal Activity of Extracts from Delphinium formosum Boiss. & Huet. Istanbul J Pharm 10.26650/IstanbulJPharm.2019.19034.

Biodiversity of Convolvulaceous species that contain ergot alkaloids, indole diterpene alkaloids, and swainsonine

Convolvulaceous species have been reported to contain several bioactive principles thought to be toxic to livestock including the calystegines, swainsonine, ergot alkaloids, and indole diterpene alkaloids. Swainsonine, ergot alkaloids, and indole diterpene alkaloids are produced by seed transmitted fungal symbionts associated with their respective plant host, while the calystegines are produced by the plant. To date, Ipomoea asarifolia and Ipomoea muelleri represent the only Ipomoea species and members of the Convolvulaceae known to contain indole diterpene alkaloids, however several other Convolvulaceous species are reported to contain ergot alkaloids. To further explore the biodiversity of species that may contain indole diterpenes, we analyzed several Convolvulaceous species (n = 30) for indole diterpene alkaloids, representing four genera, Argyreia, Ipomoea, Stictocardia, and Turbina, that had been previously reported to contain ergot alkaloids. These species were also verified to contain ergot alkaloids and subsequently analyzed for swainsonine. Ergot alkaloids were detected in 18 species representing all four genera screened, indole diterpenes were detected in two Argyreia species and eight Ipomoea species of the 18 that contained ergot alkaloids, and swainsonine was detected in two Ipomoea species. The data suggest a strong association exists between the relationship of the Periglandula species associated with each host and the occurrence of the ergot alkaloids and/or the indole diterpenes reported here. Likewise there appears to be an association between the occurrence of the respective bioactive principle and the genetic relatedness of the respective host plant species.

Cytotoxicity of New Diterpene Alkaloids, Ceylonamides G-I, Isolated From Indonesian Marine Sponge of Spongia sp.

In the course of the search for cancer cell growth inhibitors, 3 new diterpene alkaloids, designated ceylonamides G-I (1-3), together with ceylonamide F (4) were isolated from an Indonesian marine sponge of Spongia sp. The chemical structures of compounds 1-3 were determined using spectroscopic analysis and compared with those of compound 4. Among the isolated compounds, 1 and 4 inhibited the growth of human prostate cancer DU145 cells in a two-dimensional monolayer culture, with an IC50 of 6.9 and 18.8 µM, respectively. Furthermore, these compounds are also effective on spheroid of three-dimensional cell culture model, which was prepared from DU145 cells. Based on the morphological changes in the spheroids, the minimum effective concentrations of compounds 1 and 4 were 10 and 25 µM, respectively.

Cardiotoxicity evaluation and comparison of diterpene alkaloids on zebrafish

Diterpene alkaloids (DAs) have a broad spectrum of pharmacological activities, but exhibiting extremely serious cardiotoxicity to induce arrhythmia, heart arrest, even death. This study aimed to evaluate the cardiotoxicity of three diester diterpene alkaloids (DDAs) including aconitine (AC), mesaconitine (MAC), hypaconitine (HAC) and three monoester diterpene alkaloids (MDAs) including 14-α-benzoylaconine (BAC), 14-α-benzoylmesaconine (BMAC), 14-α-benzoylhypaconine (BHAC) on zebrafish. Firstly, the zebrafish embryos after a 72-hour post fertilization were treated with different doses of AC, MAC, HAC, and BAC, BMAC and BHAC for 2, 10 and 24 h, respectively. The heart rates of the treated embryos were calculated and the morphological images of body, together with heart fluorescence were obtained. Results demonstrated that AC, MAC, and HAC at low doses (15.6 and 31.3 μM) decreased the heart rates and increased them at high doses (62.5, 125, and 250 μM), while BAC, BMAC, and BHAC decreased the heart rates in the dose range of 31.3–250 μM, but the highest dose (500 μM) of BAC and BMAC increased the heart rates. In addition, AC, MAC, and HAC exhibited serious organic and functional toxicities, while BAC, BMAC, and BHAC did not. It could be induced that DDAs expressed stronger cardiotoxicities than MDAs, which might be due to that they were known as the Na+ channel activators and K+ channel inhibitors, respectively. The β-acetate at C-8 position, along with the protonated nitrogen on ring A of their chemical structures contributed more for their different cardiotoxicities. This is the first study on cardiotoxicity comparison of DAs, providing references for the rational and safe application of these compounds and some plant species containing them to reduce side effects while retaining therapeutic efficacy.

Three new diterpenes and two new sesquiterpenoids from the endophytic fungus Trichoderma koningiopsis A729

Three new diterpenes koninginols A-C (1–3) and two new sesquiterpenoids 11-hydroxy-15-drimeneoic acid (4), koninginol D (5), together with twelve known compounds (6–17) were isolated from the endophytic fungus Trichoderma koningiopsis A729 derived from Morinda officinalis. Their structures were fully assigned with the aid of extensive spectroscopic analysis and reported data from the previous literatures. Koninginols A and B were disclosed as intriguing diterpene alkaloids, whereas the koninginol C was disclosed as one of the two examples of harziandione diterpene. All the isolates were evaluated for their antitumor activity, and compounds 1–2, 4–6, 11–13, 16–17 were evaluated for their antibacterial activity, wherein the new compounds 1 and 2 exhibited significant antibacterial activity against Bacillus subtilis with MIC values of 10 and 2 μg/mL, respectively.

Prenylated Indole Diterpene Alkaloids from a Mine-Soil-Derived Tolypocladium sp.

Ten new prenylated indole diterpene alkaloids, tolypocladin A-J (1-10), including four chlorinated metabolites, have been isolated from a culture of a mine-soil-derived fungus, Tolypocladium sp. XL115. The structures and absolute configurations of 1-10 were determined by spectroscopic analysis, ECD calculations, and comparison with known compounds. Compounds 1 and 8 displayed significant antimicrobial activities. In addition, compound 1 also showed weak cytotoxic activity against all tested human cancer cell lines and suppressed the growth and viability of the patient-derived HCC cells T1224.

SYNTHESIS OF HYBRID MOLECULES CONTAINING PYRIMIDINE AND DITERPENE ALKALOID LAPPACONITINE FRAGMENTS

The cross coupling of 5'-ethynyllappaconitine with benzoic acid chlorides under the conditions of the Sonogashira reaction in benzene leads to the corresponding 5-alkynones. By condensation of the obtained 5'-alkynones with amidines, compounds with hybrid structure containing fragments of a diterpene alkaloid and pyrimidine were synthesized. The yields of the target compounds were 67–90%. The possibility of carrying out the cross coupling – condensation reaction as a one-pot synthesis is demonstrated.

Cardiotoxicity of Consolida rugulosa, a poisonous weed in Western China

Poisonous weeds are a global problem since they not only hinder local economic development, but also cause ecological harm. Consolida rugulosa (family Ranunculaceae) is a weed that is widespread in Northwestern China and causes severe poisoning when ingested by livestock. In the present study, we purified the toxins in this plant and investigated their mechanism of action. Five natural diterpene alkaloids (compounds 1–5)—including two new compounds (1 and 2)—were isolated, and five semi-synthetic derivatives (6–10) were synthesised based on 4 or 5 for structure-activity analysis. The toxicity of the compounds was evaluated in vitro with lactate dehydrogenase (LDH) assay. All of the compounds—especially 1—stimulated LDH release in primary cultured rat myocardial cells, an effect that was blocked by the Na+ channel blocker lidocaine. Electrocardiography revealed that rats treated with 1 had severe arrhythmia, while heart Doppler echocardiography and analysis of serum biomarkers levels revealed that administration of 1 for 15 days induced changes in cardiac structure and myocardial enzyme levels. These effects were antagonised by lidocaine treatment. Thus, diterpene alkaloids are the main compounds responsible for the cardiotoxicity of C. rugulosa, which can be mitigated by co-administration of lidocaine.

Synthesis of hybrid molecules containing pyrimidine and diterpene alkaloid lappaconitine fragments

The cross coupling of 5'-ethynyllappaconitine with benzoic acid chlorides under the conditions of the Sonogashira reaction in benzene leads to the corresponding 5-alkynones. By condensation of the obtained 5'-alkynones with amidines, compounds with hybrid structure containing fragments of a diterpene alkaloid and pyrimidine were synthesized. The yields of the target compounds were 67–90%. The possibility of carrying out the cross coupling – condensation reaction as a one-pot synthesis is demonstrated.

Role of JAK/STAT3 Signaling in Functional Stimulation of Mesenchymal Progenitor Cells with Alkaloid Songorine.

JAK/STAT signaling pathway was examined comparatively during realization of growth potential of mesenchymal progenitor cells stimulated with diterpene alkaloid songorine or fibroblast growth factor. The stimulating role of JAKs and STAT3 on the mitotic activity and differentiation of progenitor cells cultured with songorine was revealed. Under these conditions, the study demonstrated suppression of fibroblast colony formation against the background of reduced number of actively proliferating CFU-fibroblasts and a drop of differentiation index of progenitor cells induced by pan-JAKs and STAT3 inhibitors. The observed changes were in almost complete agreement with the character of functional reactions of the progenitor elements in response to blockade of JAKs and STAT3 with fibroblast growth factor. In addition, blockade of JAKs with this factor enhanced the differentiation rate of the progenitor cells.

Targeted profiling and relative quantification of benzoyl diterpene alkaloids in Aconitum roots by using LC-MS/MS with precursor ion scan.

Benzoyl aconite alkaloids have myocardial protective effects at a low dose and produce toxic effects at high dose. Due to lack of enough reference compounds, most of the benzoyl alkaloids had few concerns, except the typical ones, i.e. aconitine, mesaconitine, and hypaconitine. To rapidly screen out and quantify benzoyl alkaloids, a high performance liquid chromatography combined with tandem mass spectrometry was proposed based on precursor ion scanning mode. First, a diagnostic ion at m/z 105 corresponding to benzoyl group was observed by using tandem mass spectrometry, which could be used for the rapid identification of benzoyl alkaloids. The targeted screening of these alkaloids was then conducted by using precursor ion scan of characteristic ion at m/z 105. Shengfuzi (the lateral root of A. carmichaelii) was taken as example, and 24 benzoyl-containing alkaloids were identified. The six major alkaloids including aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, and benzoylhypaconine were determined in the precursor ion scan mode by the standard curve method. Reliable linearity, sensitivity, precision, accuracy, and repeatability were obtained and validated. Then the relative response factors between these six analytes were calculated, which were not more than two times using any alkaloid as reference. Thus, the other 18 alkaloids lacking reference compounds were relatively quantified. This approach provides a useful tool for rapid identification and quantitative analysis of toxic benzoyl alkaloids, and also an efficient method for the safety assessment of Aconitum roots.