MOLECULES
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2020
·Vol. 25
·期 18
·4269
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IF 3.267
Ji-Heng Wu,Yue-Ting Cao,Hong-Ye Pan,Long-Hu Wang
(1) Background: Toad venom (Bufonis Venenum, known as ‘Chansu’ in Chinese), the secretion of the ear-side gland and skin gland of Bufo gargarizans cantor or Duttaphrynus melanostictus Schneider, has been utilized to treat several diseases in China for thousands of years. However, due to the chemical variability of the components, systematic chemical composition and the key pharmacophores in toad venom have not yet fully understood. Besides, it contains a variety of effective compounds with different physiological activity and chemotypes, mainly including alkaloids, bufogenins, bufotoxins, and so on. The recent pharmacological researches have demonstrated that several bufogenins have remarkable pharmacological effects, such as anti-inflammatory, analgesic effects, and anti-tumor effects. Aim of the study: To identify the bioactive compounds and pharmacophores originating from toad venom based on analyzing spectrum-effect relationship by chemometrics and to explore the anti-cancer mechanism primarily. (2) Materials and methods: Fingerprint of the 21 batches of samples was established using HPLC (High Performance Liquid Chromatography). The anti-tumor activity of extracts were determined by in-vitro assays. Chemometric analysis was used to establish the spectrum-effect model and screen for active ingredients. Pharmacodynamic tests for the screened active compound monomers were conducted with in-vitro assays. Further anti-tumor mechanisms were investigated using western blot and flow cytometry. (3) Results: The established spectrum-effect model has satisfactory fitting effect and predicting accuracy. The inhibitory effect of major screened compounds on lung carcinoma cells A549 were validated in vitro, demonstrating that arenobufagin, telocinobufogenin, and cinobufotalin had significant anti-tumor effects. Through further investigation of the mechanism by western blotting and flow cytometry, we elucidated that arenobufagin induces apoptosis in A549 cells with the enhanced expression of cleaved PARP (poly (ADP-ribose) polymerase). These results may provide valuable information for further structural modification of bufadienolides to treat lung cancer and a method for discovery of anti-tumor active compounds. Conclusions: Our research offers a more scientific method for screening the principal ingredients dominating the pharmacodynamic function. These screened compounds (arenobufagin, etc.) were proven to induce apoptosis by overactivation of the PARP-pathway, which may be utilized to make BRCA (breast cancer susceptibility gene) mutant cancer cells more vulnerable to DNA damaging agents and kill them.Keywords:toad venom;spectrum-effect relationship;chemometrics;apoptosis
研究领域:
药理学
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肿瘤学
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化学计量学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2021
·
IF 6.953
Yanan Zhang,Xue Li,Di Xu,Mengyun Wu,Yinghui Dai,Mingyu Xia,Dong Wang
Bufadienolides are the main active ingredients of Venenum Bufonis, which is a widely used traditional Chinese medicine secreted from parotoid gland and skin glands of Bufo bufo gargarizans . According to the transcriptome analysis, “cholesterol-bile acid-bufadienolidies pathway” was proposed as animal-derived bufadienolides biosynthesis pathway by us previously. In this pathway 3 β -hydroxysteroid dehydrogenase (3 β HSD) and steroid 5 β -reductase (SRD5 β ) might be the key enzymes to convert the A/B ring to cis -configuration. Therefore, as the second report of our group, here we report the cloning of the full length of SRD5 β cDNA of B. bufo gargarizans ( Bbg - SRD5β ) from the parotoid gland of B. bufo gargarizans for the first time, and site-directed mutagenesis was used to explored the character of Bbg -SRD5 β . Bbg -SRD5 β had an open reading frame of 981 bp and encoded 326 amino acids residues. The expression conditions of the recombinant Bbg -SRD5 β in E. coli BL21 (DE3) harbored with pCold- Bbg -SRD5 β was optimized as induction for 10 h at 15 °C with 0.1 mM IPTG. With NADPH as a cofactor, Bbg -SRD5 β can reduce the Δ 4,5 double bonds of progesterone to generate dihydroprogesterone õwithout substrate inhibition effect. The catalytic rate of mutant type Bbg -SRD5 β -Y132G was 1.8 times higher than that of wild type Bbg -SRD5 β . Although Bbg -SRD5 β was almost unable to reduce the progesterone to dihydroprogesterone after mutation of V309, the affinity of enzyme with NADPH changed significantly. Bbg -SRD5 β is the key enzymes to convert the A/B ring of steroid to cis -configuration, and V309 is a key site affecting the binding affinity of enzyme with NADPH, and the mutation of Y132 can adjust the catalytic rate of Bbg -SRD5 β .
研究领域:
生物化学
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分子生物学
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中医药学
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药理学
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DOI
PubMed
高影响因子
BIOMEDICAL CHROMATOGRAPHY
·
2019
·Vol. 33
·期 9
·e4555
·
IF 1.748
Ke-ke Cui,Yue-ting Cao,Jia-feng Shao,Zheng-yu Lu,Long-hu Wang
Bufadienolides are the main bioactive components of Venenum Bufonis (VB) and have been widely used to treat different types of human cancers for decades. The bufadienolide content in VB varies significantly in materials from different geographical origins. In this work, a new strategy for the quality assessment of VB was developed through quantitative analysis of multi-components by single marker (QAMS). Cinobufagin was selected as the internal reference substance; seven bufadienolides were separated and simultaneously determined based on relative correction factors. The correlation coefficient value ( r ≥ 0.9936) between QAMS and the normal external standard method proved the consistency of the two methods. According to the outcomes of 30 batches of VB samples, the contents of the seven bufadienolides were used for further chemometric analysis. All of the samples of VB from various geographical origins were divided into three categories based on hierarchical cluster analysis and radar plot, which indicated the crucial influence of geographical origins on VB. This study showed that QAMS combined with chemometristry could be used to comprehensively evaluate and effectively control the quality of VB from different geographical origins.
研究领域:
药理学
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分析化学
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中药学
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产品关联
DOI
PubMed
高影响因子
INTERNATIONAL IMMUNOPHARMACOLOGY
·
2024
·
IF 4.8
Li Guo,Sai Zhang,Cong Zhang,Shuang Ren,Zihan Zhou,Fengyuan Wang,Yuexuan Wang,Qiqi Chen,Yubing Wang,Wen-Hui Lee,Kui Zhu,Di Qin,Yuanyuan Gao,Tongyi Sun
Inflammatory pain is a chronic pain caused by peripheral tissue inflammation, seriously impacting the patient’s life quality. Cinobufacini injection, as a traditional Chinese medicine injection preparation, shows excellent efficacy in anti-inflammatory and analgesic treatment in patients with advanced tumors. In this study, a novel analgesic peptide CI5 with anti-inflammatory and analgesic bio-functions that naturally presents in Cinobufacini injection and its regulatory mechanism are reported. Our results showed that the administration of CI5 significantly relieved the pain of mice in the acetic acid twisting analgesic model and formalin inflammatory pain model. Furthermore, CI5 effectively reduced the inflammatory cytokines (IL-6, TNF-α and IL-1β) and inflammatory mediator (PGE2) expressions, and prevented the carrageenan-induced paw edema in mice. Further LC-MS/MS results showed the anti-inflammatory and analgesic bio-functions of CI5 depended on its interaction with the Rac-2 protein upstream of ERK1/2 and the inflammatory signaling pathway (ERK1/2/COX-2 axis). In summary, CI5, as a novel natural candidate identified from Cinobufacini injection, showed substantial clinical promise for inflammatory pain treatments.
研究领域:
药理学
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疼痛管理
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传统中药
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分子生物学
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产品关联
DOI
PubMed
高影响因子
MICROCHEMICAL JOURNAL
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2020
·
IF 3.594
Jinghui Zhang,Hongye Pan,Yang Chen,Zafar Iqbal Tanveer,Li Liu,Zongwei Cai,Yanjun Hong,Yongjiang Wu
Natural medicines are now gaining increasing acceptance around the global scope. Due to the complexity of composition , it is imperative to adopt a unique evaluation system, which is not confined to the content of finite marker compounds. This research presents the successful application of pharmacodynamics-based optimization to the ultrasound-assisted extraction of Chansu, a widely used natural medicine with complex bioactive ingredients. The effect of solvent concentration, solvent to solid ratio, extraction time and particle size, on two response indexes (extraction yield of total content of bufalin , cinobufagin and resibufogenin and anticancer activity) were evaluated by response surface methodology (RSM). Two models were built based on two response indexes, and both of them provided accurate predictions. Compared with extraction yield-based optimization, pharmacodynamics-based optimization aligns well with the integrity and complexity of natural medicines. Moreover, the pharmacodynamics-based optimization gives a more economical and efficient optimal condition, which is specifically embodied in the coarser requirement for particle size and the saving of extraction time and solvent. The recommended extraction conditions were methanol concentration of 83.54% (v/v), solvent to solid ratio of 26.23 mL/g, extraction time of 40.43 min and particle size of 61 mesh (0.212–0.25 mm). The present study elaborated the feasibility and implication of pharmacodynamics-based optimization, which provides useful information for further research on the extraction process of natural medicines.
研究领域:
药理学
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药物化学
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生物技术
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产品关联
DOI
高影响因子
危险品化学品经营许可证（带存储）