Croton Crassifolius Research Articles

Cyperenoic acid suppresses osteoclast differentiation and delays bone loss in a senile osteoporosis mouse model by inhibiting non-canonical NF-\u03baB pathway

Cyperenoic acid is a terpenoid isolated from the root of a medicinal plant Croton crassifolius with a wide range of biological activities. In this study, the effects of cyperenoic acid on osteoclast differentiation were investigated both in vitro and in vivo using receptor activator of nuclear factor-κB ligand (RANKL)-induced bone marrow-derived osteoclasts and senescence-accelerated mouse prone 6 (SAMP6). Cyperenoic acid significantly suppressed RANKL-induced osteoclast differentiation at the concentrations with no apparent cytotoxicity. The half maximum inhibitory concentration (IC50) for osteoclast differentiation was 36.69 μM ± 1.02. Cyperenoic acid treatment evidently reduced the expression of two key transcription factors in osteoclast differentiation, NFATc1 and c-Fos. Detailed signaling analysis revealed that cyperenoic acid did not affect MAPK pathways and canonical NF-κB pathway but impaired activation of p100/p52 in the non-canonical NF-κB pathway upon RANKL stimulation. Moreover, the expression of osteoclast-related genes, nfatc1, ctsk, irf8, acp5 and cfos were disrupted by cyperenoic acid treatment. The bone resorption activity by cyperenoic acid-treated osteoclasts were impaired. In a senile osteoporosis mouse model SAMP6, mice fed on diet supplemented with cyperenoic acid showed delay in bone loss, compared to the control. Taken together, plant-derived cyperenoic acid shows great potential as therapeutic agent for osteoporosis.

Selective Oxidations of Cyperenoic Acid by Slightly Reshaping the Binding Pocket of Cytochrome P450 BM3

AbstractThe selective hydroxylation of unactivated C−H bonds in complex natural products represents a formidable challenge in synthetic organic chemistry. Cyperenoic acid, a sesquiterpenoid isolated from Croton crassifolius, possesses an antiangiogenic‐promoting effect. Its C7‐ and C9α‐hydroxylated products can significantly inhibit the release of vascular endothelial growth factor (VEGF). To prepare these hydroxylated products, cytochrome P450 BM3 monooxygenase was chosen as a catalyst for the hydroxylation of cyperenoic acid. A simple and fast strategy, slightly reshaping the binding pocket of P450 BM3, was described to expedite the development of highly regio‐ and stereoselective P450 catalysts. P450 BM3 was evolved through one or two generations of mutations, and a highly enriched mutant library that contained fewer than 30 variants was then constructed. The obtained P450 BM3 variants achieved selective hydroxylation at positions C7 (94 % selectivity in the case of the F87A/A330W/F331L mutant) and C9 (90 % regioselectivity and 100 % stereoselectivity in the case of the L75V/F87A/T88F/A330W mutant) of cyperenoic acid and were also used to prepare the desired hydroxylated products at a preparative scale with high isolated yields.

Norcrassin A, a novel C16 tetranorditerpenoid, and bicrotonol A, an unusual dimeric labdane-type diterpenoid, from the roots of Croton crassifolius.

A novel C16 tetranorditerpenoid, norcrassin A (1), and an unusual dimeric labdane-type diterpenoid, bicrotonol A (2), were isolated from the roots of Croton crassifolius. Norcrassin A (1) featured a new carbon skeleton with an unprecedented 5/5/5/6 tetracyclic system. Bicrotonol A (2) possessed an unusual tetrahydroxypyran ring linkage connecting two labdane diterpenoid monomers. The structures of all compounds, including the absolute configuration, were elucidated by the interpretation of their NMR spectroscopic data, high resolution mass spectrometry, and single-crystal X-ray diffraction. A plausible biosynthetic pathway of 1 is proposed. The anti-Alzheimer's Disease (AD) activities of 1 and 2 are also evaluated using the AD pathological model.

Diterpenoids from Croton crassifolius include a novel skeleton possibly generated via an intramolecular [2+2]-photocycloaddition reaction

Five previously undescribed terpenoids (cracrosons D-H), including three clerodane diterpenoids, together with 16 known diterpenoids were isolated from Croton crassifolius (Euphorbiaceae). Cracroson D features a previously undescribed carbon skeleton with an unprecedented cyclobutane ring. Their structures, including their absolute configurations, were elucidated using spectroscopic and single-crystal X-ray diffraction analyses along with CD calculations. A plausible biogenetic pathway for cracroson D is also proposed, which was supported by the experimental results. Additionally, all of the compounds were evaluated in vitro for cytotoxicity against T24 and A549 cells using the CCK-8 method.

Biotransfomation of cyperenoic acid by Cunninghamella elegans AS 3.2028 and the potent anti-angiogenic activities of its metabolites

Cyperenoic acid (1) is one of the major sesquiterpenes isolated from Croton crassifolius, which exhibited potent anti-angiogenic activity. Traditional structural modification of 1 is difficult to perform by chemical technology due to the remarkable stability of the patchoulane skeleton. In order to overcome chemical synthesis difficulties and obtain structurally diverse derivations, microbial transformation of 1 by using Cunninghamella elegans AS 3.2028 was studied for the first time. Five new hydroxylated products 2–6 were obtained. Furthermore, cytotoxicity and anti-angiogenic activities of all the biotransformation products were evaluated by MTT assay and ELISA in HepG2 and MCF-7 cells. These results indicated that hydroxylated modification products 2–4 significantly inhibited VEGF release, which suggest the potential use of hydroxylated modification products for cancer therapy.

Crassins A-H, Diterpenoids from the Roots of Croton crassifolius.

Phytochemical investigation of the 70% acetone extract of Croton crassifolius roots afforded eight new diterpenoids, crassins A-H (1-8), and 19 known compounds. The structures of the new compounds were determined by spectroscopic methods, and their absolute configurations were determined by electronic circular dichroism, single-crystal X-ray diffraction analysis, comparison with literature data, and biogenetic considerations. Crassins A (1) and B (2) are new ring B-rearranged diterpenoids, whereas crassin C (3) is a new ring A-rearranged diterpenoid. Crassin H (8) exhibited selective cytotoxicity against A549 cells (IC50 5.2 μM) compared with HL-60 cells (IC50 11.8 μM). The known compound chettaphanin-II exhibited moderate activity against the A549 and HL-60 cell lines (IC50 8.4 and 10.5 μM, respectively).

Cytotoxic clerodane diterpenoids from Croton crassifolius

Hepatocellular carcinoma (HCC) is the most common type of liver cancer, and treatment options for HCC are limited. In addition, the discovery of new natural compounds with anti-hepatocarcinoma activity is attracting increasing attention. For this reason, phytochemical investigation of Croton crassifolius led to the isolation of 17 diterpenoids, including three new clerodane diterpenoids, named crassifolius A-C (1–3), along with 14 known ones (4–17). Their structures were established by 1D, 2D NMR, HR-ESI-MS, detailed calculated electronic circular dichroism (ECD) spectra and the assistance of quantum chemical predictions (QCP) of 13C NMR chemical shifts. The cytotoxicities of all these compounds against human liver cancer lines (HepG2 and Hep3B) were determined. Among them, compound 1 exhibited good cytotoxicity with IC50 value of 17.91μM against human liver tumor cells Hep3B. Following further studies of the anti-tumor mechanism of compound 1-induced cell growth inhibition, we found that compound 1 caused apoptotic cell death in Hep3B cells by detecting morphologic changes and Western blotting analysis.

Penduliflaworosin, a Diterpenoid from Croton crassifolius, Exerts Anti-Angiogenic Effect via VEGF Receptor-2 Signaling Pathway.

Anti-angiogenesis targeting vascular endothelial growth factor receptor-2 (VEGFR-2) has been considered as an important strategy for cancer therapy. Penduliflaworosin is a diterpenoid isolated from the plant Croton crassifolius. Our previous study showed that this diterpenoid possesses strong anti-angiogenic activity by inhibiting vessel formation in zebrafish. This study was conducted to further investigate the anti-angiogenic activity and mechanism of penduliflaworosin. Results revealed that penduliflaworosin significantly inhibited VEGF-induced angiogenesis processes including proliferation, invasion, migration, and tube formation of human umbilical vein endothelial cells (HUVECs). Moreover, it notably inhibited VEGF-induced sprout formation of aortic rings and blocked VEGF-induced vessel formation in mice. Western blotting studies showed that penduliflaworosin inhibited phosphorylation of the VEGF receptor-2 and its downstream signaling mediators in HUVECs, suggesting that the anti-angiogenic activity was due to an interference with the VEGF/VEGF receptor-2 pathway. In addition, molecular docking simulation indicated that penduliflaworosin could form hydrogen bonds within the ATP-binding region of the VEGF receptor-2 kinase unit. Finally, cytotoxicity assay showed that penduliflaworosin possessed little toxicity toward both cancer and normal cells. Taken together, our findings demonstrate that penduliflaworosin exerts its anti-angiogenic effect via the VEGF receptor-2 signaling pathway. The anti-angiogenic property and low cytotoxicity of penduliflaworosin suggest that it may be useful in cancer treatments.

A new patchoulane-type sesquiterpenoid glycoside from the roots of Croton crassifolius

A new patchoulane-type sesquiterpenoid glycoside (1), together with five known sesquiterpenoids (2–6), was isolated from the roots of Croton crassifolius. Their structures were elucidated on the basis of spectroscopic methods. This is the first report about the sesquiterpenoid glycoside from C. crassifolius. All the isolated compounds 1–6 were evaluated for their cytotoxic activities against human tumour cell lines HL-60 and A549, but none showed significant activity.

Pyran-2-one Derivatives from the Roots of Croton crassifolius

One new pyran-2-one derivative [crotonpyrone C (1)] and two known ones (2-3) were isolated from the supercritical fluid extract (SFE) of the roots of Croton crassifolius. Their structures were elucidated using spectroscopic (IR, UV, 1D and 2D NMR) and HRESIMS methods. The isolated compounds were evaluated for their anti-angiogenic activity using a zebrafish model, but showed little activity.

Diterpenoids from the roots of Croton crassifolius and their anti-angiogenic activity

Six diterpenoids [crassifolin J, K, L, M, N and O] along with eleven known ones were isolated from the supercritical fluid extract (SFE) of the roots of Croton crassifolius (Euphorbiaceae). Their structures were elucidated using spectroscopic methods (IR, UV, HRESIMS, 1D and 2D NMR). The structure and stereochemistry of crassifolin J was confirmed by single-crystal X-ray diffraction analysis, and the absolute configurations of crassifolin K–M were determined by CD spectra. Twenty-three diterpenoids from this plant were screened for their anti-angiogenic activity using a wild-type zebrafish in vivo model. Four of the known compounds were active, of which penduliflaworosin possessed the best activity relative to the positive control (SU5416). Further study demonstrated that penduliflaworosin could inhibit vessel formation on Tg(fli1a:EGFP)y1-type zebrafish embryos.

New clerodane diterpenoids from Croton crassifolius

Two new clerodane diterpenoids (1–2), one new clerodane diterpenoid alkaloid (3), as well as thirteen known compounds were isolated from Croton crassifolius. The structures of new compounds were established by a combination of spectroscopic methods, including HRMS, 1H NMR, 13C NMR, 1H 1H COSY, HSQC, HMBC, NOESY and X-ray crystallographic analysis. Compound 3 is firstly reported as the clerodane-type diterpenoid alkaloid in natural products. All of the compounds were evaluated for in vitro cytotoxic activities against CT26.WT cell using the MTT method.

A Novel Norclerodane Diterpenoid from the Roots of Croton crassifolius

A chemical investigation of Croton crassifolius afforded a novel norclerodane diterpenoid (1) with an unprecedented six-membered oxygen ring between C-1 and C-12, together with three known compounds. The structure of the new compound was elucidated based on spectroscopic (IR, 1D, and 2D NMR) and HR-ESI-MS techniques. This report describes the first example of a natural norclerodane with a 4H-chromene ring system.

Potent anti-angiogenic component in Croton crassifolius and its mechanism of action

Ethnopharmacological relevanceThe root of Croton crassifolius Geisel is traditionally used in China for the treatment of snake bites, stomach ache, sternalgia, joint pain, pharyngitis, jaundice and rheumatoid arthritis, while in Thailand, it has been used as an anticancer herbal medicine by the indigenous people. Yet, its pharmacological studies are still limited, especially towards its anticancer property. Anti-angiogenesis is a promising therapeutic strategy in the anti-cancer treatment. Previous studies have shown strong anti-angiogenic activity in the low polar fraction of the herb. Nevertheless, the potent compound which is responsible for the anti-angiogenesis, and its molecular mechanism have never been reported. Aim of the studyTo determine the potent anti-angiogenic component in C. crassifolius and its molecular mechanism of action. Materials and methodsC. crassifolius was extracted using supercritical fluid extraction and steam distillation. The anti-angiogenic activities of the two extracts were evaluated in the zebrafish model by quantitative endogenous alkaline phosphatase assay. The chemical compounds in the active extract were isolated using chromatographic methods, and their structures were elucidated using different spectroscopic techniques. The content/quantity of the active compounds in this extract was determined with HPLC analysis. The molecular mechanism of the most active compound was further studied using the real-time PCR assay. Besides, its cytotoxicity on various cancer and normal cell lines was evaluated using the cell-counting kit. ResultsSupercritical fluid extract (SFE) of C. crassifolius showed better anti-angiogenic activity than that of steam distillation extract (SDE). Three sesquiterpenes, namely, cyperenoic acid, 8-hydroxy-α-guaiene and (+)-guaia-l(10),ll-dien-9-one, were isolated and identified in the SFE. Among them, cyperenoic acid displayed the strongest anti-angiogenic activity by 51.7% of the control at 10μM, while the others showed little effect. HPLC results showed that cyperenoic acid was the major component in the SFE with 9.97% (w/w). Results of the real-time PCR assay suggested that the cyperenoic acid affected multiple molecular targets related to angiogenesis including vascular endothelial growth factor (Vegfa), angpiopoietin (Angpt), and their receptors. Cytotoxicity assay showed cyperenoic acid possessed little toxicity toward cancer and normal cells. ConclusionsCyperenoic acid is an important anti-angiogenic component present in C. crassifolius and serve as a potent inhibitor in the angiogenesis in the zebrafish embryo model. The anti-angiogenic property, but not the cytotoxicity, of C. crassifolius provides a scientific basis for its traditional use in cancer treatment.

ChemInform Abstract: Two New Clerodane Diterpenoids from Croton crassifolius.

AbstractThe two new clerodane diterpenoids crassifolin I (I) and crassifolin H (II) are isolated from the root of Croton crassifolius.

Cytotoxic and antibacterial pyran-2-one derivatives from Croton crassifolius

Two new pyran-2-one derivatives, crotonpyrones A (1) and B (2), were isolated from the roots of Croton crassifolius. Compounds 1 and 2 represent the first examples of the natural occurrence of the new pyran-2-one derivatives in the family Euphorbiaceae. Their structures were elucidated by extensive spectroscopic analysis (HRESIMS, 1D and 2D NMR), and the structure of compound 1 was further confirmed by single-crystal X-ray diffraction analyses. Compounds 1 and 2 showed good cytotoxicity against human cancer cell lines of HeLa and NCI-446. Compound 2 also exhibited antibacterial activity against Staphylococcus aureus.

A New Halimane Diterpenoid from Croton crassifolius

Agriculture and Animal Husbandry College, Qinghai University, Xining 810016, ChinaReceived December 9, 2013, Accepted January 13, 2014Key Words : Euphorbiaceae, Croton crassifolius, Diterpenoid, HalimaneCroton is a large genus of the Euphorbiaceae family,comprising approximately 1200 species of trees, shrubs andherbs distributed throughout tropical and subtropical regionsof the world.

Crocrassins A and B: two novel sesquiterpenoids with an unprecedented carbon skeleton from Croton crassifolius

Crocrassins A (1) and B (2), two novel sesquiterpenoids with a unique skeleton, were isolated from a Chinese medicinal plant Croton crassifolius.

Foliar trichomes of <i>Croton</i> L. (Euphorbiaceae: Crotonoideae) from China and its taxonomic implications

Foliar trichomes of 21 species of the genus Croton L. from China have been examined using stereomicroscopy and scanning electron microscopy. Five trichome types characterized by their morphology are identified, viz., stellate, lepidote, simple, dendritic and appressed-rosulate. Only stellate trichome is observed in most species, with only six species that are found to maintain two or three trichome types. Trichome types and density are useful for species identification and sectional classification for Chinese species. Based on the trichome types and other morphological characters, 21 Chinese species are proposed to be placed in five sections. Croton crassifolius belongs to sect. Andrichnia; C. cascarilloides belongs to sect. Monguia; C. mangelong, C. kongensis, C. laevigatus and C. laniflorus belong to sect. Argyrocroton; C. lauioides, C. howii and C. damayeshu belong to sect. Adenophylli. The remaining Chinese Croton species might be placed into sect. Croton. A key for Chinese Croton species based on trichome morphology is provided.DOI: http://dx.doi.org/10.3329/bjpt.v20i1.15468Bangladesh J. Plant Taxon. 20(1): 85-94, 2013 (June)

Clerodane Diterpenoids from Croton crassifolius

Seven new clerodane diterpenoids (1-7) were isolated from roots of Croton crassifolius, along with six known compounds. The structures were elucidated by extensive spectroscopic methods (IR, UV, HRESIMS, 1D NMR, and 2D NMR), and the structures of 1, 3, 4, and 7 were confirmed by single-crystal X-ray diffraction analyses. Compounds 1-13 were evaluated for in vitro antiviral activity against herpes simplex virus type 1 using the cytopathic effect reduction assay.